Levels of the glutamic oxaloacetic transaminase of erythrocytes of pregnant women and of cord bloods of newborn infants.

The mean basal specific activity (S.A.) of the glutamic oxaloacetic transaminase of erythrocytes (EGOT) for a group of 64 pregnant women was lower (p less than 0.001) than the value for the cord bloods of newborn infants, and lower (p less than 0.001) than the value for adults who had a top limit of S.A. of EGOT. In establishing the top limit of the S.A., it is important that the mean basal S.A. of the cord bloods from 49 newborn infants was identical to the mean basal S.A. of adults who had an adequate supplement of pyridoxine. There were no differences in the mean basal S.A.'s of the cord bloods between asymptomatic mothers and mothers who had anemia, edema, hypertension, proteinuria and glucosuria. An infant may be born with a top limit of S.A. which is non-deficient in pyridoxal 5'-phosphate, but a mother can have a low level of the transaminase, and which is deficient in the coenzyme.


Hyperlipidemia in a Lebanese community: difficulties in definition, diagnosis and decision on when to treat.

Fasting serum cholesterol and triglyceride levels were measured in 131 randomly selected adult members of a Lebanese Community of Western New York. Mean cholesterol levels (males, 217 mg%; females 234 mg%) were higher than those reported from the Lebanon but similar to those reported in most other populations. Mean triglyceride levels (males, 153 mg%; females, 115 mg%) were higher than those reported in most other populations. Twenty-three subjects were hyperlipidemic on the basis of age and frequency distribution adjusted serum lipid levels above the 90th percentile. Clinical and family studies carried out on 13 of these 23 hyperlipidemic subjects suggested that 77% had monogenic hyperlipidemia and 23% primary non-monogenic hyperlipidemia. A high frequency of familial hypercholesterolemia (minimum estimate 0.7%) was found, in keeping with the high frequency of the disorder in Lebanon. The study serves to emphasize the difficulties in defining hyperlipidemia, in establishing a specific diagnosis in the individual patient, and in distinguishing between optimum and normal serum lipid levels.


Studies on platelet aggregation in vivo. VI. Effect of a pyrimido-pyrimidine derivative [RA233] on tumor cell metastasis.

The pyrimido-pyrimidine derivative RA 233 was found to increase circulation time and decrease metastatic settling of intravenously injected, radioisotope labeled ascites tumor cells in mice.


Ultrastructural study of the mitochondria in the skeletal muscle fibers in the diabetic Chinese hamster.

An ultrastructural study of skeletal muscle fiber biopsies from the tongue and soleus muscle of diabetic Chinese hamsters revealed the presence of lipid droplets within the muscle fibers. These droplets often were found in the cyptoplasm near mitochondira, sometimes inside or surrounded by one or more mitochondria. Glycogen granules were found in great quantities in the cytoplasm of the tongue and soleus muscle. Granules which appeared to be glycogen were also found within the mitochondria of the tongue muscle.


Experimental gingivitis in ODU plaque-susceptible rats. II. Biochemical nature of the rat plaque.

The characterization of the dental plaque from ODU plaque-susceptible rats is reported. The comparison between the rat dental plaque and a human dental plaque indicates that content of extractable hexose and inorganic salts and amino acid composition are different from each other. Electron microscopic study indicates that bacterial flora of the rat dental plaque seems to be little different from a human dental plaque. In spite spite of obvious differences in biochemical nature between the rat dental plaque and the human dental plaque, there are enough similarities to warrant intensive investigation regarding the relationship between the dental plaque and gingivitis.


The width of lingual mandibular attached gingiva.

One hundred and twenty individuals in good oral health were divided into six groups according to age. Measurements were made on the depth of the gingival sulcus, and the distance from the margin of the free gingiva to the mucogingival junction on the lingual mandibular teeth. The mean, extreme, standard deviation and analysis of variance were computed. A distinct pattern of the width of healthy lingual attached gingiva was found. The largest width was found lingual to the first and second molars with a mean height of 4.7 mm. This was followed by the third molar [3mm] second bicuspid [2.5 mm] first bicuspid [2 mm], and cuspid, lateral and central [1.9mm]. Lingual attached gingiva showed that there was a variation in width with each tooth and between individuals. There was no variation in width of lingual attached  gingiva between sexes. The extreme measuremtns ranged from  less than 1 mm to 8 mm. The data from the study gives the dentist some indication of the mean and extreme ranges of lingual attached gingiva. This data can serve as a guide in diagnoiss and treatment planning and in selecting procedures to reestablish these zones of attached gingiva.


Diabetes mellitus and periodontal disease.

A sample of 54 patients with diabetes mellitus were subjects to detailed assessment of periodontal disease levels using standard indices. In order to determine whether the severity of periodontal disease was related to the severity of diabets mellitus, a series of parameters of the diabetes mellitus population was simultaneously studied. There were no significant relationships between the levels of periodontal disease and the duration of diabetes, the type of treatment and the frequency of systemic complications. Periodontal disease in the diabetic appeared to the affected by the same etiologic factors [plaque, calculus, neglect] as would be expected in nondiabetic patients. Further studies with larger population samples would be appropriate.


The effect of chlorhexidine gluconate on the formation of experimental granulation tissue.

The aim of the study was to investigate the effect of chlorhexidine gluconate on the connective tissue formation. Granulation tissue was produced with a method in which sponges were inserted beneath the back skin of rats. Dry sterilized sponges were impregnated either with 0.45% NaCl solution (control) or or 0.2% chlorhexidine solution [test]. The sponges were removed after 3, 5, 7, 10, 14, 19 and 23 days. The content of hydroxyproline, hexosamines, DNA and RNA was estimated from the sponges. The acid glycosaminoglycans were further purified and analyzed. The results of the study indicate that chlorhexidine had a clear delaying effect on the formation of granulation tissue.


Effect of chlorhexidine gluconate on acute nonmicrobial inflammation reaction.

The effects of chlorhexidine on the appearance of leukocytes and some hydrolytic enzymes were studied in an experimental acute nonmicrobial inflammation reaction caused by implanation of viscose sponge beneath the backskin. The results showed that chlorhexidine gluconate decreased the number of leukocytes in the inflammatory exudate. The drug used had no measurable effect on the distribution of different types of leukocytes. Chlorhexidine has a clear inhibitory effect on the appearance of phosphatases, glycosidases and peptidases. On the other hand the effect on proteinases was small.


Histologic evaluation of the effects of intrasulcular toothbrushing on human sulcular epithelium.

Nineteen young adult Caucasian males had an average Gingival Index Score of 0.47. Each was asked to demonstrate his present method of toothbrushing: thirteen used the scrub method, and six combined the scrub and roll methods. Control gingival tissue specimens were taken from the mandibular left first bicuspid or first molar regions, and the area was allowed to heal for 40 days. Following complete healing, intrasulcular brushing was used in the mandibular left posterior segment, for 30 seconds daily for a period of 40 days. The brushing was done by the investigator on 28 days and by the subjects on 12 days. The average Gingival Index Score following intrasulcular brushing was 0.25. A gingival specimen was taken from the experimental area of the mandibular left second bicuspid region. Histological findings in control specimens from 13 subjects showed parakeratinization for varying distances in the sulcus epithelium apical to the gingival crest. In six specimens a complete absence of sulcus keratinization was observed. The average Inflammatory Index Score was 1.21. Following intrasculcular brushing, 18 specimens showed a parakeratin layer apical to the gingival crest; in 14, it extended the entire length of the coronal sulcular epithelium or apparent clinical sulcus, whereas in four more it extended well beyond the gingival crest into the sulcus. In the six subjects whose control specimens showed an absence of keratin apical to the gingival crest, parakeratinization of part or all of the clinical sulcus was found following intrasulcular brushing. The average Inflammatory Index Score was 1.015. A moderately intense acute inflammatory reaction was observed within the attachment epithelium in three subjects. Possible therapeutic benefits and hazards of intrasulcular brushing are discussed.


A histologic study of age changes in the adult human periodontal joint (ligament).

This study evaluated the histological age changes in 80 periodontal joints from 24 human cadavers ranging in age from 20 to 90 years. The limitations of such a descriptive study were reviewed to prevent overextrapolation of the findings to whole populations. In the young speciments, the bone surface of the tooth socket was smooth and regular with the insertion of Sharpey's fibers evenly distributed in the woven bone. Interstitial lamellae were found between secondary osteons in the alveolar bone. Copious periodontal fibers passed around small vascularized interstitial spaces to insert evenly into the cementum as Sharpey's fibers. Numerous cells, probably fibroblasts, were present within and between the fiber bundles. Like the bone surface, the cemental surface was smooth and regular. Both cellular and acellular cementum were present. Cemental splits were found both intracementally and at the cemento-dentinal junction. In the older specimens, the periodontal surface of the alveolar bone was jagged and uneven with Sharpey's fibers inserting irregularly into the woven bone. Subjectively an increased amount of internal remodeling was seen in the periodontal bone. A relative decrease in both suspensory fibers and cellular content was noted in the periodontal ligaments of the older specimens. Large vascularized interstitial spaces encroached upon areas formerly occupied by periodontal fibers and bone. Fat cells also were observed replacing the suspensory fibers. The cementum showed an irregular surface into which Sharpey's fibers were unevenly inserted. Both cellular and acellular cementum were present, and cemental splits were seen with the same frequency as in the young specimens. There was a tendency for greater cemental apposition, particularly in the apical region of the tooth. In many respects the aging periodontal joint showed atrophic and degenerative changes similar to those in cranio-facial sutures. However, these changes did not include either bony ankylosis or osteoporosis related to increasing age. In summary, the periodontal joints of the younger specimens were well organized while those of the older specimens showed a more irregular structure and a decrease in fiber and cellular content. Intermediate changes were seen in the intermediate age groups.


Primary hyperparathyroidism detected by gingival biopsy.

This case of primary hyperparathyroidism presented several unusual features: (1) The only presenting symptoms were two gingival giant cell lesions (brown tumors), (2) Bone changes occurred early in the disease, but apparently affected only the mandible and maxilla, (3) No renal disease could be detected. The diagnosis, treatment and postoperative course of this unusual case has been described.


Fractured genial tubercles.

It is apparent from these reports that fractures of genial tubercles occur periodically and that surgical treatment for most fractures is not necessary. This type of patient is quite likely to seek dental care because a denture may seem to be the obvious source of irritation. Review of all available information indicates that conservative management is the treatment of choice for fractures of genial tubercles.


Treatment prostheses in TMJ dysfunction-pain syndrome.

Various types of acrylic resin therapeutic prostheses commonly used in the treatment of TMJ dysfunction-pain syndrome were described. Each design was related to recent data concerning optimum condylar positions in the fossae, the physiologic condylar suspension system, and individual treatment objectives for repositioning the mandibular condyles. For example, alteration of the vertical dimension of occlusion is a popular treatment procedure that is empirical in nature and is usually used without TMJ radiographs or a differential diagnosis. It can violate the physiologic requirements of the interocclusal distance or the speaking space and does not necessarily move the condyles forward as has been commonly thought. The dangers of empirical treatment procedures for a multicausal dysfunction syndrome have been pointed out. An example was cited where the mandible was moved forward for a long period of time with a repositioning prosthesis; this produced pathologic TMJ remodeling and continued pain. It was recommended that specific mandibular repositioning be based on the type of condylar displacement observed on the radiographs. Sometimes the condyles should be retruded, and other times they should be repositioned anteriorly or occasionally inferiorly on one side. Long-standing use of any acrylic resin repositioning prosthesis is contraindicated, particularly without close supervision. Acrylic resin anterior bite plates (with a minimum opening of 1 mm) were recommended for the relief of acute trismus or intractable pain. Usually the prosthesis is used in conjunction with heat and drug therapy. This type of prothesis can also be utilized to deprogram the muscles when a strong habit of eccentric occlusion develops as a result of missing teeth. (This should be confirmed by TMJ radiographs.) Occasionally atypical pain is present and a differential diagnosis can be established between TMJ dysfunction or neurologic etiology by the physiologic response to bite plate therapy. When it is necessary to reposition the mandibular condyles anteriorly the occlusion is adjusted to provide the planned anterior movement without increasing the vertical dimension of occlusion. A temporary acrylic resin prosthesis is used to retrain the muscle programming to the anterior therapeutic mandibular position. When the symptoms are relieved and the corrective condylar position is confirmed with TMJ radiographs, a more permanent repositioning prosthesis is made. The treatment of TMJ dysfunction-pain syndrome should be based on documented data including the pain history, TMJ radiographs, condylar position in the fossae, electromyographic evidence, and occlusal analysis. This information will help determine if the patient's centric relation is functional or dysfunctional and will indicate the recommended treatment procedures.


Creep and hardness testing of some denture base polymers.

A method of measuring creep, recovery, hardness, and the effect of cyclic loading on denture base polymers using a modified Wallace hardness tester is described. This apparatus is particularly sensitive and is capable of detecting very small variations in these properties of the materials. A representative group of denture base polymers that include heat-cured, pour-type, and other modified materials was tested. The results obtained are in good agreement with those shown by other workers.


Dehydroretronecine-induced skin tumors in mice.

Female Swiss mice susceptible to skin tumors received 6 sc injections and/or topical applications of dehydroretronecine, a metabolite of the pyrrolizidine alkaloid monocrotaline, and were observed for 15 months for tumor development. Of 92 animals examined, 63 had tumors at the site of application or injection; 47 of these had skin tumors, mainly basal cell and squamous cell carcinomas. These data indicate that dehydroretronecine is a proximate carcinogen capable of causing a high incidence of skin tumours in mice at the site of sc injection or topical application.


Immune reactivity of RBL-3, a nonproducer leukemia induced by Rauscher murine leukemia virus in C57BL/6 mice.

A cultured line of RBL-3 leukemia, induced by Rauscher murine leukemia virus (R-MuLV) in C57BL/6 mice, was found to be a virus nonproducer. An NB tropic virus indistinguishable from R-MuLV, however, was recovered after treatment with 5-iodo-2'-deoxyuridine (optimal concentration, 200 microgram/ml). Dexamethasone treatment slightly boosted the production of the activated virus but did not by itself activate the virus. RBL-3 cells were found to have Fc receptors but no surface immunoglobulin or Thy 1.2 antigen. RBL-3 cells exhibited the Friend-Moloney-Rauscher virus-specific transplantation antigen as shown by its immunosensitivity and immunogenicity in tumor rejection experiments. Crude membrane and crude soluble antigen preparations obtained from RBL-3 cell membranes retained the capacity to induce tumor rejection.


Quantitative kinetics of development of N-2-fluorenylacetamide-induced, altered (hyperplastic) hepatocellular foci resistant to iron accumulation and of their reversion or persistence following removal of carcinogen.

For study of the behavior of previously described carcinogen-induced foci of hepatocytes refractory to iron accumulation, iron deposition was induced in all hepatocytes of rat livers by feeding 8-hydroxyquinoline and ferrous gluconate to the animals for 3 months. The kinetics of development of the altered foci of hepatocytes free of stainable iron was then quantified for each lobe of liver for as long as 13 weeks of feeding of the carcinogen N-2-fluorenylacetamide and for 6 months following cessation of carcinogen exposure. The foci increased slowly in number and size for the first 12 weeks of carcinogen administration and then grew rapidly between weeks 12 and 13. At week 13, they corresponded to altered foci that have been observed in sections stained with hematoxylin and eosin, and they also displayed abnormal glycogen storage. After the carcinogen was discontinued, the foci disappeared by a reversion back to iron storage while temporarily retaining the nuclear abnormalities of altered cells. Over 95% of altered foci at this stage underwent reversion by 6 months following removal of the carcinogen. More advanced lesions developed in the lobes that displayed the greatest incidence and persistence of foci.


Induction of intestinal metaplasia in the stomachs of rats by N-methyl-N'-nitro-N-nitrosoguanidine.

N-Methyl-N'-nitro-N-nitrosoguanidine (MNNG) was administered orally to male Wistar rats at a concentration of 83 microgram/ml in the drinking water for 2, 4, 5, and 7 months; the rats were killed at about month 15. Intestinal metaplasia was found in the stomachs of 80-100% of the rats treated with MNNG for 4 or more months, of 37.5% treated with MNNG for 2 months, and of 10% of the controls. Metaplastic glands, composed of goblet cells and columnar cells with striated borders, were found in the pyloric region. Paneth's cells were found at the bottom of metaplastic glands in a rat treated with MNNG for 4 months. The incidence of well-differentiated adenocarcinomas of the stomach was 63-90% in rats treated with MNNG for 4 or more months and 25% in those treated with MNNG for 2 months.


Zinc deficiency and methylbenzylnitrosamine-induced esophageal cancer in rats.

Dietary zinc deficiency increases the incidence of and shortens the lag time for induction of esophageal tumors in rats by methylbenzylnitrosamine (MBN). Groups of control and zinc-deficient outbred Charles River CD rats were given 24, 17, 8, or 4 doses of MBN, administered twice weekly by intragastric intubation at doses of 2 mg/kg body weight. Between 58 and 93 days after the beginning of treatment, all rats were killed and examined. The frequency of esophageal tumors in the zinc-deficient groups was significantly higher than in the corresponding control groups. Following ip injection of [methyl-14C]MBN, DNA and RNA of esophagus and liver were more noticeably labeled than kidney, lung, and small intestine, which exhibited only modest labeling. In vitro incubation with [14C]MBN of tissue slices from esophagus, liver, kidney, and small intestine produced similar results. This pattern correlates well with the results of our studies of MBN-induced esophageal tumorigenesis.


Selective sex-related modification of diethylnitrosamine-induced carcinogenesis in BALB/c mice by concomitant administration of butylated hydroxytoluene.

Tumor induction in BALB/c mice by diethylnitrosamine (DENA) was shown to be sex related. Female mice killed 13.5 months after DENA exposure generally had higher incidences of forestomach and lung tumors than did males and also had higher numbers of lung tumors per tumor-bearing mouse. Although neither sex developed stomach tumors spontaneously, males had higher incidences of spontaneous lung tumors than did females (45 vs. 26%). When butylated hydroxytoluene (BHT) was given concomitantly with DENA treatment, females had an increased survival (68 vs. 43%) and a reduced incidence of forestomach squamous cell carcinomas (77 vs. 100%) compared with females given DENA alone. No such protective effect on survival or on the occurrence of stomach tumors was seen in males given BHT and DENA or on lung tumorigenesis in either sex. In males, BHT given alone increased survival of treated mice over that of untreated controls, but it had no such effect on the females. The results of this study suggested an as yet unexplained hormonal influence and/or dependence for this BHT modification of DENA tumorigenesis.


Effect of embryonic tissue immunization on chemically induced gastrointestinal tumors in rats. I. Can embryonic antigens act as rejection antigens?

Inbred WF rats were inoculated with crude suspensions prepared from liver and gut tissue of 12- to 15-day fetuses of the same strain. Rats previously unsensitized to syngeneic embryonic tissue were inoculated with fetal material sc three times during exposure to 1, 2-dimethylhydrazine dihydrochloride (DMH), a gastrointestinal (GI) carcinogen in rodents. Properly timed immunization inhibited the development, growth, and metastasis of primary GI tumors. This effects was observed in both sexes; however, it was more prounced in male rats. Nine WF rats with DMH-induced carcinoma of the GI tract were inoculated sc with syngeneic fetal tissue. Of 9 rats, 7 rejected the embryonal tissue implants, which thus demonstrated the presence of a concomitant immune response to embryonic antigen(s). Two rats in which fetal tissue grew out to palpable nodules had multiple GI tumors with metastasis and extra-GI tumors, i.e., a massive tumor load. Ten other rats with DMH-induced GI tumors were treated with unblocking serum. The unblocking serum was inoculated to counteract serum-blocking factors in vivo. These rats were inoculated intradermally with syngeneic fetal tissue. In all 10 rats, inflammation and necrosis were noted at the inoculation site after 24-72 hours, which thus demonstrated a delayed hypersensitivity reaction to embryonic antigens. Our experiments suggest that embryonic antigens common to fetal and tumor cells can induce immunity in an autochthonous host and can act as rejection antigens.


Radioimmunoassay for benzo[a]pyrene.

A benzo[a]pyrene (BP)-bovine serum albumin conjugate was synthesized and used to immunize 2.5- to 3.0-kg New Zealand White rabbits. The resulting antisera to BP bound trace amounts of [3H] (55 pg; 6,000 counts/min). A radioimmunoassay (RIA) to BP was developed by the antiserum first being titered with the [3H]BP and then a standard curve being constructed from the addition of unlabeled BP (0.4-15.8 pmoles; 0.1-4.0 ng/assay tube). The RIA could reliably detect 0.4 pmoles (0.1 ng) BP. The specificity with respect to structurally related polycyclic aromatic hydrocarbons was examined by means of competitive binding. Here concentrations of 4.0-400 pmoles of the compound were used, and the resulting competitive curves were compared for relative cross-reactivity at 50% B/B0 (counts per minute labeled BP bound to antiserum in the presence of corresponding concentrations of unlabeled BP/counts per minute labeled BP bound to antiserum in the absence of unlabeled BP). The antiserum B4-3 was specific to BP with the closest cross-reacting substance being 3-hydroxybenzo[a]pyrene (20% relative cross-reactivity). BP added to pooled human serum was measurable by the RIA at 1 ng/ml. A BP RIA may have potential use for the quantification of the absorbed dose of this carcinogen in man.


Carcinogenicity of subcutaneously injected N-nitrosoheptamethyleneimine in European hamsters.

Male and female European hamsters (45 of each sex) recieved sc injections once weekly for life of N-nitrosoheptamethyleneimine (NHMI) at one-fifth the median lethal dose (LD50) (females: 44 mg/kg body wt; males: 66 mg/kg body wt), one-tenth the LD50 (females: 22 mg/kg body wt; males: 33 mg/kg body wt), or one-twentieth the LD50 (females: 11 mg/kg body wt; males: 16.5 mg/kg body wt). Survival times for both males and females were dependent on the dose of NHMI. Pulmonary neoplasms were induced in almost all the treated animals. They were histologically diagnosed as adenocarcinomas, squamous cell carcinomas, and mixed cell carcinomas. In addition, nasal cavity tumors developed in all hamsters of all treatment groups; these were papillomas, squamous cell carcinomas, and a few adenocarcinomas. Only 1 tumor of the larynx and 1 tumor of the trachea were observed. Several papillomas and a few carcinomas were also detected in the forestomach. The results were discussed with reference to previous findings in rats and Syrian golden hamsters.


Enhancement of heterotransplanted human tumor graft survival in nude mice treated with antilymphocyte serum and in congenitally athymic-asplenic (Lasat) mice.

The latency period, success rate, and minimal cell inoculum size required for transplantation of continuously passaged human tumor lines into congenitally athymic (nude) mice, antilymphocyte serum (ALS)-treated congenitally athymic (nude) mice, and congenitally athymic-asplenic (lasat) mice were compared. The 11 tumor lines studied included examples of breast adenocarcinoma, transitional cell carcinoma, osteosarcoma, fibrosarcoma, Hodgkin's disease, malignant melanoma, and rhabdomyosarcoma. Of these 11 tumor lines, 3 were successfully transplanted into nude mice, compared to 5 of 10 tumor lines in ALS-treated nude mice and 9 of 11 lines in lasat mice. Moreover, the latency period was shorter and the minimal cell inoculum size was lower for lasat mice than for either nude or ALS-treated nude mice. Despite this enhancement of heterotransplantation into lasat mice and despite the growth of large local masses, no evidence of distant metastases was found.


A specific chromosome breakpoint associated with mouse plasmacytomas.

The chromosomes of uncultured cells of the near-diploid mouse plasmacytoma MOPC-31C were studied. The modal number of chromosomes was 44. The tumor lacked two marker chromosomes, reciprocal translocation [rcp t(12; 15)], that in previous studies were found to be common to 3 other uncultured myelomas and 1 cultured mouse myeloma. Through the formation of two markers, rcp t(6; 15), unique to this tumor, however, the tumor shared with other tumors and their specific markers a common breakpoint in chromosome "15 at band D3/E. This breakpoint has been found in all mouse plasmacytomas examined with banding thus far and is considered of possible importance in the development of this tumor.


Rate of cell division in atypias and invasive carcinoma of the uterine cervix of the mouse.

The rate of cell division during a 6-hour period was determined in cervical epithelia from 64 normal mice, 26 mice with cervical atypias, and 8 mice with invasive carcinomas. Cervical atypias and carcinomas were induced by local application of benzo[a]pyrene (BP), and mitosis was arrested with colchicine. Values for the ratio of dividing basal cells to total dividing cells and for the rates of basal cell division for the 6-hour period were both significantly greater in normal epithelia than in atypias, and these values for atypias were greater than those for invasive carcinomas. The differences were not estrous cycle-dependent. There was no evidence that the mechanism of host invasion by squamous tumor cells was due to an increased number of cells in mitoses. The occurrence of areas with mitotic activity suggested the existence of focally distributed and probably alternating, synchronous cell production in normal cervical epithelium. This property was apparently not altered by topical BP application or by carcinogenesis.


Decline of natural killer cell activity in sublethally irradiated mice.

Spleen cells of (C57BL/6 X C3H/He)F1 mice were assayed for natural killer (NK) cell activity against YAC-1 and FBL-3 lymphoma targets at several intervals after total-body exposure to a high sublethal dose of 137Cs or 60Co gamma-rays. NK cell activity did not decline for the first 12 days but decreased sharply thereafter and remained low until day 24. The recovery of splenic NK cell activity was delayed. Beginning on day 28, the activity was slowly increased, reaching near-normal levels (80% of controls) 41-59 days after irradiation. Suppressor cells detectable during the period of lowest NK cell activity, i.e., on days 17 and 19, may have been responsible for the delayed and slow recovery. These studies indicated that a) mature effectors of natural cytotoxicity are relatively radioresistant renewable cells with a lifespan of about 2 weeks whose progenitors are radiosensitive cells b) the kinetics of decline and especially of recovery of NK cell activity may be influenced by suppressor cells. Should NK cell activity confer resistance to autochthonous lymphomas in vivo, it may be a significant consideration for strategies of tumor therapy by cytotoxic agents that reconstitution of the NK cell pool is a slow process and that suppressor cell function must be overcome for full recovery.


Physicochemical properties and isoenzyme composition of hexokinase from normal and malignant human tissues.

The activity of hexokinase was studied in several normal and malignant human tissues. The enzyme activity in tumors was significantly higher. Isoenzyme studies on normal gastric mucosa and stomach cancer extracts showed that malignancy is accompanied by a "simplification" of the hexokinase isoenzyme pattern due to "deletion" of the slowest isoenzyme. Preparative polyacrylamide gel electrophoreis was used to isolate hexokinase isoenzymes from normal and malignant tissues. Tumor hexokinase isoenzymes displayed an increased affinity to glucose when compared to the corresponding normal prototypes (Km/glucose, 10(-6) M and 10(-5) M, respectively; Km = Michaelis constant). The molecular weights, subunit composition, and peptide patterns were identical for corresponding isoenzyme pairs from normal and tumor tissues.


Relationship between epidemiologic and endocrinologic aspects of cervical cancer.

In search of etiologic relevancy of the steroid deviations in urine, we compared the reproductive activities of patients having cervical cancer with those of normal controls. A premenopausal patient experienced the birth of her first child significantly earlier than did the urban control of corresponding age, despite their similarity of age at menarche. However, when the same patient was compared with the premenopausal rural control, this differences was not found. The same parameter also failed to differentiate a postmenopausal patient from the corresponding control of urban origin. The premenopausal patient who was indistinguishable from the rural control by age at first delivery was distinguished from the rural control by reduced excretions of adrenal steroids including 11-deoxy-17 ketosteroids and some corticosteroid metabolites. The spectrum and the degree of deviation of urinary steroids for a patient coincided with the spectrum and the degree of age dependency of the urinary steroids for a normal woman during and after adolescence. These findings indicated that the inability of the reproductive parameter to discriminate a patient was associated with growth retardation in a population, and a maturation deficiency of the adrenal gland (arrest of adrenarche) was implicated in the genesis of cervical cancer.


The multistage theory of carcinogenesis and the age distribution of cancer in man.

The multistage theory of carcinogenesis is briefly reviewed, and the incidence function predicted by the theory is expressed as an "infinite sum" (convergent power series). This expression for the incidence function has the Armitage-Doll approximation as the first non-zero term and explicitly exhibits the dependence of the incidence function on the transition rates. It is then clear that if the transition rates are not small enough (approximately 10(-4/cell/yr), the Armitage-Doll approximation is inappropriate. Retention of one more term (in the series) leads to a better approximation in these cases. This is illustrated by a hypothetical example. Other assumptions implicit in the theory are discussed.


Nuclear magnetic resonance and detection of human breast tumor.

Malignant, benign, and normal breast tissues corrected for fat appeared to have similar nuclear magnetic proton spin-lattice relaxation times, contrary to previously published reports. However, due to a lower fat content, benign and malignant tumors relaxed more slowly than the surrounding normal tissue. This provides a possibility for locating them (but not for discriminating between them) in a breast by the nuclear magnetic resonance imaging technique zeugmatography.


Solubilization and partial isolation of human melanoma tumor-associated antigens.

Human melanoma cell membrane tumor-associated antigens (TAA's) were solubilized in an active form by pronase digestion of either a fresh melanoma or cells from a melanoma cell line maintained in tissue culture. Upon elution from Sephadex G-200 column, TAA's solubilized from the melanoma cell line were found in four distinct peaks that had apparent molecular weights of approximately 48,000 (partition coefficient Kd, 0.426), 25,000 (Kd, 0.567)8 17,000 (Kd, 0.699), and 13,000 (Kd, 0.831) daltons, respectively. Fetal antigen activity was found in all but the 13,000-dalton peak. HLA antigen activity was detected in the 17,000-dalton material. TAA's prepared from the fresh tumor source eluted from Sephadex G-200 column with an apparent molecular weight of 14,000-25,000 (Kd, 0.786-0.572) daltons, as did HLA antigens. A partial resolution of the TAA's from the HLA antigens was achieved with the use of DEAE-cellulose chromatography. Results of antigenic stability assays suggested that the TAA structure is stable to prolonged exposure to low pH. Recovery of TAA activity from the strong denaturing agents 5 m urea, 0.5% (wt/vol) sodium dodecyl sulfate, and 4 m guanidine hydrochloride was partially successful. These properties of the TAA's may be useful for further isolation of the TAA's.


Selective killing of oncogenic human cells cocultivated with normal human fibroblasts.

Histidinol and theophylline reversibly arrested the growth of low-passage human foreskin fibroblasts (HFF) and protected these cells from an otherwise lethal combination of the S-phase drugs beta-cytosine arabinoside and hydroxyurea. In contrast, HeLa cells continued cell cycle transit in the presence of histidinol and theophylline. Consequently, these reagents did not after the vulnerability of HeLa cells to combined S-phase drugs. The differential S-phase drug sensitivity mediated by histidinol and theophylline persisted under cocultivation conditions, thereby allowing the selective and total eradiction of colonies of HeLa cells in the presence of subconfluent HFF cells. The HFF cells, spared S-phase drug toxicity by their unique response to histidinol and theophylline, showed total survival and viability.


Central nervous system lesions in childhood leukemia.

Central nervous system (CNS) lesions were found in 85 of 91 children with acute leukemia. Cerebral atrophy was the most common lesion, occurring in 65 percent. Atrophy was most severe in children treated with intrathecal methotrexate alone or in combination with radiation therapy, in children youngest at time of onset of leukemia, and in children in whom duration of leukemia was shortest. Longer duration was the determining factor among children who were older at onset. Lesions previously reported by other investigators, such as leptomeningeal infiltration of leukemic cells, infection, hemorrhage, degenerative grey and white matter disease and leukoencephalopathy were also seen. Central pontine myelinolysis occurred in four children.


Psychiatric care in a general practice.

The morbidity and patient management in a small general practice in a suburb of Auckland was analysed over a period of three months, comparing medical and psychiatric cases. Psychiatric patients tend to be more often frmale, older (though not in the geriatric age group), make slightly greater demands on practice time and have somewhat more chronic illnesses, but these differences from medical patients are small and psychiatric patients do not form a major problem in practice in the community. The findings, while showing a slightly higher frequency of psychiatric patients, are otherwise comparable with those from other New Zealand studies.


Flucloxacillin (floxapen) in the treatment of skin and upper respiratory tract infections.

An appraisal of flucloxacillin (floxapen) for the treatment of infection of the skin and upper respiratiory tract was carried out in general practice by 34 doctors. The success rate in the treatment of skin infections was assessed as 92 percent and in upper respiratory tract infections as 84 percent. The incidence of side effects overall was 9.5 percent and their nature and severity are described. Also the results of the present study are compared with previous appraisals of flucloxacillin in general practic and the difference in dose regimes used in these studies is discussed.


The functioning of a short-stay emergency gynaecological unit.

Admissions to a small constantly staffed emergency gynaecological unit over a two month period are reported. Of 408 patients seen, 230 were in hospital for less than 36 hours and of these 128 were not admitted to a hospital ward. Of a yearly total of 2000 emergency admissions about 1000 did not need admission to a hospital ward. Full staffing of such a unit for 24 hours each day would thus allow a considerable number of beds to be released for elective work. The referring diagnosis of incomplete abortion proved to be incorrect in 100 of 203 patients. Clinical features and management in these patients are discussed. Pain is an unreliable guide, but opening of the cervix is not often seen unassociated with incomplete abortion.


MCQ assessment of introductory clinical course for fourth year medical students.

A 60-item MCQ as administered to 46 fourth year medical students at the beginning, half-way, and at the end of a five week introductory clinical course. The stated goals were achieved: (1) a baseline measure was provided, indicating abilities brought by newly-arriving students; (2) data gave opportunity for "mid-course" corrections by students in their study and by tutors in their teaching; (3) overall student change was assessed, offering a measure of the educational effectiveness of the course, and (4) tutors were made aware of individual students requiring particular attention. Such assessment techniques provide an effective monitoring and motivating instrument at this level  of medical education.


The measurement of complement as an aid in diagnosis: 1. Nephritis and collagen disease.

A selection of patients with nephritis and collagen diseases were studied for complement abnormalities. This communication briefly reviews the current comcepts of complement physiology, details standard assay used in this laboratory and describes clinical examples where these tests have provided  significant clinical information. Quantitative measurements of individual components, the presence of circulating immune complexes, and the demonstration of in vivo C3 cleavage products were assessed simultaneously. The importance of measuring cleavage products and immune complexes is stressed. Clinical examples are given where misleading or erroneous interpretation of results might have occurred if these tests had not been performed.


Establishing a health planning and research unit: the first two years of operation.

The Christchurch Health Planning and Research Unit was established in April 1975 to develop an approach to regional health services planning based upon researched community need, making the best use of available resources and involving agencies, health professionals and the community. This paper reports on the background to its establishment, the development of a planning strategy and progress and problems to date, especially in the fields of geriatrics, mental health and primary care.


Child health of Great Barrier Island.

The health of 69 children on Great Barrier Island was studied. This is an isolated community without a permanent doctor, and most children were healthy. Ear disease was the commonest problem, especially in Maori children. Problems of impetigo, scabies, rheumatic fever and dental caries were not present. Comparisons are made with the results of a recent Auckland study.


The state of school health: a Bay of Plenty study.

A health survey was carried out on 131 primary school children in the Bay of Plenty. Ninety children had no health defect. Inadequate hearing was the commonest defect and was more prevalent in Maoris. In all 55 health defects were found of which 18 were newly detected. There was a total absence of iron deficiency anaemia.


Social patterns of adolescent cigarette smoking in a rural community.

A survey of a secondary school serving a small town and rural district in New Zealand revealed that 46 percent of the students, aged 12 to 18, were current cigarette smokers. The smoking habits of elder siblings and peers were closely related to subjects' smoking. Parental opposition to smoking, beliefs that smoking was bad for health and a degree of self-esteem appeared to have protective value. The key to prevention of cigarette smoking may lie in parent and early childhood education aimed at increasing awareness of the dangers of smoking, fostering a soical climate of opposition to smoking, and increasing the child's self-confidence.


Pathology medical manpower planning in New Zealand results of 1977 survey.

As part of Medical Manpower Planning in New Zealand, a survey conducted by the New Zealand Society of Pathologists in association with the Royal College of Pathologists of Australia shows that at present there are 116 pathologists practising in New Zealand, a ratio of one to 27 000 population. This is far short of the requirement of 1:20 000, for which an additional 40 pathologists is required now, with an average annual requirement of 11 additional pathologists over the next nine years.


Acute selenium poisoning: case report.

A case of self-poisoning with sodium selenate sheep drench, along with blood and urine levels of selenium, is reported. Treatment included gastric lavage, diuresis, vitamin C, and dimercaprol, and the patient recovered without sequelae.


Oral scleral heterografts. A pilot study.

This study showed that scleral heterografts in the oral cavity work. Such grafts demonstrate the following characteristics: (1) attach firmly to submucosal, periosteal, and bony tissues; (2) remain firm and fibrotic in nature, indicating the possibility of a stressbearing tissue; (3) show little or no scarring or contracture; (4) show minimal, or no- host antigenic response based on clinical acceptance; (5) appear to become vascularized and cellular, blending with adjacent collagen. As projected by this study, scleral heterografts might well be used to obliterate bony undercuts and perhaps to obtund cystic cavities and other major bony defects. Further studies of layering of the sclera are essential before any definitive result can be determined in large bony defects. In those areas in which no bone regeneration would be feasible, such as in the mylohyoid region of the mandible, this application does present a possibility for removing undercuts as opposed to removing overlying bone. The tenacity of adherence  to submucosal tissues, periosteum, and bone makes scleral heterografting even more intriguing in the possible obliteration of oroantral fistulas. We might theorize that the sclera would not only attach to the buccal and palatal flaps but also show a rather firm and tenacious adherence to the bony walls of the antral defect. The availability of human bank sclera for use as presently made by periodontists and as pioneered by Klingsberg is somewhat limited. From our study, an extrapolation would indicate that, because of the comparably low and antigenicity of bovine sclera and human sclera, an investigation in this area should be completed in order to determine whether scleral heterografts can replace scleral homografts in human periodontal surgery. Sclera is easily preserved in anhydrous glycerine (as preserved by King and associates19) and can certainly be stored in banks. In our opinion, the final application of scleral heterografts, human or bovine, the role of antigenicity, and the ability of osteoblasts to invade either a single piece or layers of sclera warrant further study.


Osteosarcoma of the mandible. Report of a case.

A case of osteosarcoma of the mandible with recurrence is presented, and the pertinent literature is reviewed. There was dramatic response to chemotherapy and return of full sensation to the left inferior alveolar nerve distribution.


Effects of systemic cortisone on the healing of tooth sockets in rats. A histologic study.

Young male rats were divided randomly into experimental and control groups of twenty-seven each. Animals in the experimental group received daily subcutaneous injections of cortisone, in dosages sufficient to allow only slight gains in weight during the 38--day study period, whereas animals in the control group received a similar volume of saline solution. All injections were started 3 days before the upper right first molars were extracted. Histologic evaluation of the sockets of animals killed at intervals during healing showed no significant differences between the experimental and control groups.


Osteoradionecrosis of maxillae.

A case of osteoradionecrosis of the maxilla has been presented. It is hoped that this review and discussion of osteomyelitis and osteoradionecrosis will point out the need for proper management of the dentition of cancer patients who are subjected to radiotherapy.


Persistent sialadenitis in an accessory parotid gland.

The patient who was followed had a persistent painful condition of the side of her face. Following a total parotidectomy, the symptoms presevered and this was ultimately diagnosed as being due to sialadenitis in the accessory parotide gland as well.


Diphenylhydantoinate-induced hyperplasia of the masticatory mucosa in an edentulous epileptic patient.

Gingival hyperplasia caused by phenytoin sodium is rarely encountered in edentulous persons. A case of generalized palatal hyperplasia is reported in which retained roots and teeth were suspected of having perpetuated a preextraction lesion, whereas a subsequently placed complete denture initiated a midpalatal hyperplasia. The lesion remained in spite of nonuse of the denture for several years.


Epidemiologic study of potential adverse drug reactions in dentistry.

In an effort to prevent and document adverse drug reactions, the University of Southern California Dental School conducted a 2--year study in providing clinical pharmacy services to its patients. The study comprised prescription screening of 2,653 drugs dispensed to 2,418 patients, with 110 reactions being uncovered. These consisted of fifty reports of drug interaction, twenty-nine reports of contraindication, and thirty-one actual cases of side effects. All but the latter group of reactions were avoided by careful review of patients' medical data. The study also discussed the relationship of drug reactions to specific medication prescribed by determining epidemiologic categories of risk.


Dental management of mild hemophilia with polycythemia vera.

Classic hemophilia (hemophilia A. Factor VIII deficiency, antihemophilic factor deficiency) and polycythemia vera were diagnosed in a patient who presented for management of his dental disease. This case report summarizes the history, laboratory findings, and clinical course and discusses the various diagnostic considerations in the management of two rare disorders. A review of current concepts of hemophilia and polycythemia vera is presented.


Oral findings in the autoimmune polyendocrinopathy-candidosis syndrome (APECS) and other forms of hypoparathyroidism.

Of twenty-eight patients aged 4 to 28 years with the autoimmune polyendocrinopathy-candidosis syndrome (APECS), twenty-seven had chronic oral candidosis, twenty-five had HPT, twenty-three had enamel hypoplasia, and nineteen had hypoadrenocorticism. Of three patients with other forms of hypoparathyroidism, none had any associated disease or oral abnormality. In nineteen cases candidosis was the first manifestation of APECS. Its occurrence was independent of the other components of the syndrome. The occurrence of enamel hypoplasia also was independent of HPT. Possibly, the enamel organ was affected by an autoimmune episode. No delayed maturation, resorption, or root hypoplasia was observed. Tooth impaction occurred in three patients.


Superficial melanomas of oral mucous membranes.

In accordance with microscopic and clinical criteria established for superficial melanomas of the skin (superficial spreading melanoma, lentigo maligna melanoma, acral-lentiginous melanoma), three oral lesions have been evaluated. The literature on oral melanomas has also been reviewed, with special attention given to those cases that had pre-existing melanosis. One patient with a diagnosis of superficial spreading melanoma eventually died of his untreated lesion 11 years after its first appearance. Two patients had lesions diagnosed as acral-lentiginous melanoma (a group which also includes volar and subungual melanomas) that exhibited aggressive, recurrent behavior. These lesions had microsocpic features similar to lentigo maligna melanoma but did not behave in a manner consistent with that diagnosis. Electron microscopic study of one acral-lentiginous melanoma demonstrated malenosomes and premelanosomes that were like those seen in normal melanocytes and nevus cells. The superficial or radial growth phase of many oral melanomas has apparently gone unrecognized. Melanosis has been reported to be a common feature of invasive oral melanomas but has not generally been related to the natural history of these lesions. Oral lesions with a prolonged intra-epithelial or radial growth phase would be expected to have a better prognosis than nodular melanomas, but meaningful survival data are not available because of the infrequency with which oral melanomas have been subclassified.


Metastatic retinoblastoma of the mandible.

Two cases of oral metastasis of retinoblastoma are reported. The first case was seen in Iraq and the second case in England. Review of the literautre revealed that when oral metastasis occurs, the prognosis must be considered to be grave; this was confirmed by Case 2. In each case there was metastasis in the symphyseal region, which followed enucleation of the affected orbit. Although the prognosis is poor, recent studies indicate that immunologic techniques may be useful to monitor the response to treatment of the primary lesion.


Intraoral blue (Jadassohn-Tieche) nevus.

Blue nevus of the oral mucosa is a distinctly uncommon clincial entity. Careful review of the literature yielded thirty-one previously reported cases. The present article reports the occurrence of a blue nevus of the hard palate in a 58-year-old man. It is of interest since it is the smallest (1 by 1 mm.) intraoral blue nevus to be reported. A clinicopathologic study of the previous thirty-one cases and of our case suggests that this lesion has no age or sex predilection. The most common site of occurrence was the hard palate. There appears to be no tendency toward recurrence. A brief review of the historical background, clinical features, theories of possible origin, and differential diagnosis is presented. Excisional biopsy of localized areas of oral pibmentation, together with histopathologic study, is indicated to rule out melanoma.


Verruciform xanthoma. First case reported in a black person.

The twenty-sixth case of verruciform xanthoma is reported. The clinical and light microscopic findings in this case are similar to those previously described of a shaggy, parakeratin-covered, pebbly, or verrucous surface and elongated rete pegs which extend a uniform depth into the underlying conncetive tissue. Large "foam cells" or xanthoma cells filled only the connective tissue papillae between the epithelial pegs. The 1.5 by 4 cm. lesion in our patient is the largest verruciform xanthoma reported to date and is the first to be reported in a black person.


Induced reattachment in periodontic-endodontic lesions by root demineralization in situ.

A new method of root preparation to enhance reattachment of gingival tissues to root dentin has been developed through extensive animal research. The improved technique may aid periodontists and endodontists in the management of deep combined lesions. This article reviews essentials of experimental animal data and presents case reports of three different clinical approaches to combined lesion closure attempts utilizing surgical and chemical root preparation.


Blood flow and oxygen consumption in steroid-treated dental pulps.

This study examined pulpal blood flow and oxygen consumption by the dental pulps of dogs. Blood flow was measured by the radioactive microsphere method and oxygen uptake was determined by the indirect Warburg approach. These variables were assessed 2 days after the canine teeth had been exposed and treated with either a cotton pellet impregnated with 1 per cent prednisolone or normal saline. The analysis indicated that the pulpal blood flow as well as the oxygen consumption did not differ between the steriod-treated tooth and the corresponding saline-treated tooth in the same jaw.


Submergence of roots for alveolar bone preservation. I. Endodontically treated roots.

Mandibular premolars in four dogs were endodontically treated and then totally submerged. Histologic and radiographic findings showed that this procedure should be considered as an alternative to extraction of key teeth in an effort to preserve alveolar bone. 1. Periodic follow-up on this procedure should be done to rule out cystic formation. 2. Beveling the coronal portion to the buccal is advocated to compensate for the thin buccal plate. 3. The procedure should be done in two steps to allow the root canal sealer to set, or an amalgam should be placed over the pulp canal. 4. New cementum and connective tissue will form over the coronal surface separating the dentin from new bone.


Granulopoiesis in chronic myeloproliferative disorders in children.

Four children with chronic myeloproliferative disorders (three with Philadelphia [Ph1] chromosome-positive chronic myelogenous leukemia [CML] were studied with soft agar culture at diagnosis (before therapy) in an attempt to define abnormalities in granulopoiesis. The three patients with CML had elevated peripheral blood golony-forming cells (CFCs) and/or normal or decreased bone marrow CFCs (in those studied). Colony-stimulating activity (CSA) was markedly decreased or absent at diagnosis in all three. Maturation of myeloid cells eithin the colonies in agar was normal, indicating that no block in myeloid maturation was present. These findings are in general agreement with results previously reported in untreated adults with Ph1 chromosome-positive CML and further define the similarity with the adult form of the disease. One Ph1 chromosome-negative patient with a clinically similar chronic myeloproliferative disorder was studied and had similarly elevated peripheral blood CFCs. She had normal CSA with a similarly high WBC count. This finding was unexpected and suggests that, unlike the patients with CML, her monocytes were capable of elaboration CSA. This difference might prove helpful in the classification of this type of disorder in cases where the Ph1 chromosome abnormality is not present.


Congenital hypoplastic anemia terminating in acute promyelocytic leukemia.

Acute promyelocytic leukemia developed in a patient with congenital hypoplastic anemia (CHA). The patient had Turner's phenotype, but no chromosomal abnormality. The literature on CHA is briefly reviewed, and the relationship between leukemia and cytotoxic therapy is considered.


[Induction of jaw bone formation by tooth autotransplantation].

A case-report of segmental underdevelopment of the alveolar process in the upper jaw of an eleven-year old girl is presented. At the site where the alveolar ridge was underdeveloped, two impacted malformed incisors were found. The malformed teeth had almost normal crowns, but the root-development had terminated 1-2 mm beyond the cementoenamel junction. The patient was treated by removal of the two malformed impacted teeth, followed by autotransplantation of the developing second premolar with the intention of inducing bone formation. The result was promising. After 3 years the alveolar process had developed to almost normal vertical hight. Some considerations are made regarding the possible origin of the bone induction factors in tooth buds. The fact that this patient demonstrated an underdevelopment of the alveolar process in spite of the formation of almost normal crowns, is taken as evidence that the bone stimulating principle in tooth buds are not to be found in the enamel organ proper, but must be related to Hertwig's epithelial sheath, root development and tooth eruption.


[Ocular localization appearing in a child with acute lymphoblastosis after hematological remission for 18 months].

Case report of a 4-year-old girl in whom a unilateral ocular localization appeared after 1 1/2 year of maintained remission of acute lymphoblastic leukaemia. The evolution of this localization can be divided in three phases: 1. During the first 17 months, an apparent simple relapsing conjunctivitis of the left eye. 2. During the following 5 months, repeated episodes of the left iritis with mydriasis and discolouration of the iris. 3. Finally, left uveitis involving the anterior chamber, with hypopion, hyphaema and glaucoma; the clinical presentation at this stage was suggestive of ocular lymphoblastosis. Radiotherapy to the lesion (total dose, 1 200 rads) was quickly followed by resolution without sequelae. No relapse was observed during the following 30 months, till death. The ocular disease remained the only manifestation of leukaemia during its evolution. A haematological relapse occurred 11 months after cure of the iritis, followed 1 year later by a second, treatment-sensitive relapse, and finally by a third, fatal relapse. The patient survived 5 years 9 months.


Morphology of osteosarcoma: new qualitative and quantitative investigations.

Results on seven cases of osteosarcoma are reported, based on new morphologic methods and quantitative procedures. Tumor tissue was embedded without prior decalcification in plastic and sectioned. Imprint cytology preparations were produced from fresh tumor tissue, and cell nuclei were measured with an electronic image analysing computer system. The loss of differentiation seen in osteosarcomas differs among osteoblasts, osteocytes, and osteoclasts. The differentiation of osteoclasts, namely their resorptive characteristics, disappears relatively early. Tumor osteocytes show loss of differentiation in their osteocyte processes. The new formation of tumor bone tissue remains in the near normal range of volume density when nuclear polymorphy is limited. The formation of ground substance and mineralization are apparently closely couplet to one another, since in our cases mostly ordered osteoid seams were observed. The capacity for mineralization of bone tissue is lost with marked polymorphy. The significance of these results for diagnostic statements and therapeutic consequences will be further discussed in long term studies.


Ion flow through a membrane: effect of chemical reaction on time dependence.

The membrane model previously described [Hays, T.R., Buckwalter, C.Q., Lin S.H. & Eyring, H. (1978) Proc. Natl. Acad. Sci. USA 75, 1612-1615] for ion flow through a membrane is expanded to include the effect of binding of the mobile ion at the occupiable sites in the membrane. Two different effects were investigated: alteration of the association-dissociation rates at constant equilibrium constant and alteration of the equilibrium constant at constant dissociation constant. Increasing the rates of association and dissociation initially causes an increased slowing of the relaxation to the final steady state, though ultimately the curves for the faster rates cross those for the slower states. Increasing the equilibrium constant causes a greater delay in the relaxation curve, with the curves for different equilibrium constants not crossing. Overall, the effect of binding is not very great unless the equilibrium constant for binding is quite large.


Energies of polypeptides: theoretical conformational study of polyglycine using quantum mechanical partitioning.

A method is developed for evaluating the total energy of polypeptides based on a combination of quantum mechanical and empirical potentials. Adjacent and nonadjacent peptide units are allowed to interact through these respective means. Our hybrid procedure is applied to a study of polyglycine and compared to the results obtained by the method of Scheraga and coworkers. We find the alpha helical conformation of a single strand of polyglycine to be most stable in vacuo. Other less-stable configurations include the 3(10) helix, the 2(7) ribbon structure, and the fully extended conformation.


General characteristics of protein degradation in diabetes and starvation.

The enhanced protein degradation associated with diabetes and starvation is fundamentally different from normal protein catabolism. In normal eukaryotic cells large molecular weight proteins tend to be degraded more rapidly than small proteins, acidic proteins tend to be degraded more rapidly than neutral or basic proteins, and glycoproteins tend to be degraded more rapidly than nonglycoproteins. All three of these general correlations are absent or markedly reduced in liver and muscle of diabetic and starved rats. In contrast, the correlations between proteins size and half-life, between protein net charge and half-life, and between protein carbohydrate content and half-life are not affected in brain of diabetic or starved animals. These results suggest that diabetes and starvation alter the general characteristics of intracellular protein degradation in target tissues of insulin. Degradation of serum proteins is also affected in diabetes and starvation. In normal animals a general correlation exists between isoelectric points of serum proteins and their degradative rates. This relationship is abolished in diabetes and starvation, as it is among liver and muscle proteins. The implications of our findings are discussed with regard to possible mechanisms of the enhanced protein breakdown.


Micrococcus luteus DNA gyrase: active components and a model for its supercoiling of DNA.

Two active components alpha and beta of micrococcus luteus DNA gyrase, of peptide weights of 115,000 and 97,000, respectively, have been purified. Each individual component exhibits little DNA gyrase activity; the ATP-dependent negative supercoiling of a covalently closed circular DNA duplex is catalyzed by a combination of the two. Covalent closure by Escherichia coli ligase of a circular DNA containing single-chain scissions, when carried out in the presence of a combination of the DNA gyrase components alpha and beta, gives a positively supercoiled DNA upon removal of the bound protein molecules. ATP was not present during the ligase treatment; therefore the positive supercoiling of DNA observed is a result of the binding of gyrase molecules, presumably as multi-subunit oligomers, during the ligation step. This is in contrast to the negative supercoiling of DNA catalyzed by gyrase in the presence of ATP. A model in which negative supercoiling of DNA is achieved by ATP-modulated repetitive wrapping of the DNA around gyrase is described. The model also suggests a plausible mode of action by which translocation of a DNA along its helix axis can be actively driven by an ATPase.


Kinetics of hemoglobin and transition state theory.

Experimental evidence indicates that the way cooperativity in hemoglobin is manifested kinetically depends on the nature of the ligand. It is shown that the qualitative differences in the kinetics of NO, O2, CO, and bulky isocyanides can be understood in structural terms in a framework based on transition state theory.


Assembly of new nucleosomal histones and new DNA into chromatin.

The assembly of chromatin from newly synthesized nucleosomal histones (labeled with [3H]arginine) and new DNA (density-labeled with [125I]iododeoxyuridine)was studied in growing cultured mouse cells. The nucleosomal histones were specifically examined by dissociating histone H1 and nonhistone proteins from unsheared chromatin either by incubation in 0.6 M NaCl or by digestion with micrococcal nuclease to release nucleosomes. In both cases, the four nucleosomal histones (H2A, H2B, H3, and H4) are essentially the only proteins that remain bound to DNA and that are labeled by [3H]arginine. After formaldehyde fixation, H1-depleted chromatin containing dense DNA can be completely resolved in CsCl buoyant density gradients from that containing unreplicated DNA; separation of nucleosomes is satisfactory although less complete. New DNA and new histones are already assembled into chromatin possessing characteristic nucleosomal structure after 3 min of synthesis (the shortest time studied), as shown by the kinetics of digestion of new DNA by micrococcal nuclease, by the distribution of new DNA and new histones in nucleosomes. However, after 3-30 min of synthesis most new nucleosomal histones are associated with unreplicated DNA rather than with new DNA. It is concluded that new nucleosomes are assembled on DNA at some distance from DNA replication sites, with concomitant migration of preexisting nucleosomes onto new DNA.


Adenovirus DNA-directed transcription of 5.5S RNA in vitro.

A cell-free system developed from human KB cells was used to transcribe 5.5S RNA from deproteinized adenovirus DNA in vitro. The cell-free RNA synthesis is dependent upon exogenous templates, ribonucleoside triphosphates, and cell-free postmitochondrial supernatant of human KB cells. The synthesis of 5.5S RNA is inhibited only by high levels of alpha-amanitin; therefore it is carried out by RNA polymerase III. The rate of synthesis was linear for at least 2 hr, indicating reinitiation. The 5.5S RNA synthesized in vitro is similar to the corresponding in vivo RNA in size, sequence, and coding region on adenovirus type 2 DNA. In this report is demonstrated in vitro synthesis of a facsimile of an in vivo transcript directed by deproteinized DNA in a mammalian cell-free postmitochondrial supernatant system.


A general method to assess similarity of protein structures, with applications to T4 bacteriophage lysozyme.

A method is proposed that permits the structural similarity between any pair of proteins to be analyzed in a completely general manner. In the proposed procedure, all possible structural segments of a given length from one protein are compared with all possible segments from the other protein. This set of comparisons reveals any structural similarities between the two proteins being compared, and also provides a basis for estimating the probability that a particular degree of structural homology could have occurred by chance. Application of the method to the comparison of T4 bacteriophage lysozyme and carp calcium-binding protein suggests that the previously reported structural similarity between parts of these two proteins [Tufty, R. M.& Kretsinger, R. H. (1975) Science 187, 167-169] is no better than would be expected by chance. On the other hand, the structural correspondence between phage lysozyme and hen egg-white lysozyme [Rossman, M.G. & Argos, P. (1976) J. Mol. Biol. 105, 75-96] does appear to be significant.


Construction of a novel plasmid-phage hybrid: use of the hybrid to demonstrate ColE1 DNA replication in vivo in the absence of a ColE1-specified protein.

A hybrid bacteriophage, P420, was constructed in vitro; it contains part of bacteriophage P4 and a 3.6-kilobase derivative of plasmid ColE1. In Escherichia coli, the plasmid-phage hybrid can exist as a stable plasmid or can be packaged into infective bacteriophage particles. Replication of P420, directed by the ColE1 replicon, was found to occur after P420 phage infection of E. coli cells that had been incubated in the presence of chloramphenicol. Replication began without a lag period and resulted in the synthesis of covalently closed circles of P420 DNA. Like ColE1 DNA replication but unlike that of P4, replication was dependent on DNA polymerase I and was sensitive to rifampicin. The presence of a resident ColE1 plasmid in the infected cells resulted in an inhibition of the replication of the incoming P420 DNA. These results indicate that ColE1 does not require a plasmid-coded protein to replicate its DNA in vivo and demonstrate the utility of P4 bacteriophage for coupling bacteriophage properties to a plasmid replicon.


The ovalbumin gene: structural sequences in native chicken DNA are not contiguous.

The sequence organization of the structural ovalbumin gene and flanking sequences in native chicken DNA was studied by restriction mapping and filter hybridization using a nick-translated probe generated from pOV230, a recombinant plasmid that contains a full-length ovalbumin DNA synthesized from ovalbumin mRNA. The structural sequences of the ovalbumin gene in native chicken DNA were found to be noncontiguous because at least two restriction endonucleases that do not cut the structural sequence do cleave the natural gene into multiple fragments by cleaving within nonstructural sequences interspersed between the structural sequences. The observation that all ovalbumin DNA-containing sequences were contained within a single DNA fragment generated by BamHI digestion of total chicken DNA has allowed us to construct an inclusive restriction map of the natural ovalbumin gene which contains at least two "insert regions." These regions may be further subdivided into alternating structural and insert sequences. Both insert regions were located within the peptide-coding regions of the gene and the sizes of these insert regions were estimated to be approximately 1.0 and 1.5 kilobase pairs, respectively.


Hepatic receptor that specifically binds oligosaccharides containing fucosyl alpha1 leads to 3 N-acetylglucosamine linkages.

Evidence is presented suggesting that hepatocytes contain a receptor that binds glycoproteins specifically through fucose in alpha1-->3 linkage to N-acetylglucosamine. Human lactoferrin, which contains this type of linkage, is rapidly cleared from the circulation of mice after intravenous injection, and greater than 90% of the injected material is found in hepatocytes. Binding of lactoferrin is mediated through its carbohydrate groups, since its clearance is prolonged after periodate oxidation or after its oligosaccharide groups are extensively degraded with glycosidases. In addition, glycopeptides from lactoferrin inhibit lactoferrin clearance. That lactoferrin clearance is mediated through binding to its fucosyl groups is suggested for several reasons. First, transferrin and asialotransferrin, whose oligosaccharide groups are essentially structurally identical to those of lactoferrin but devoid of fucose, are not cleared on intravenous injection. Second, when fucose is incorporated into asialotransferrin by alpha1-->3 N-acetylglucosamine fucosyl transferase, the resulting fucosylated derivative is cleared rapidly. Neither mannan nor derivatives of orosomucoid that are cleared by binding to receptors for galactose, N-acetylglucosamine, or mannose, inhibit clearance of lactoferrin although clearance is inhibited by fucoidin. Finally, glycoproteins containing fucose in alpha1 --> 2 linkage to galactose or alpha1 --> 6 linkage to N-acetylglucosamine do not inhibit lactoferrin clerance by the liver. Since clearance of other glycoproteins, such as human lactoperoxidase, also appears to be mediated through binding to the same hepatocyte receptor as lactoferrin, it is concluded that the fucose-specific receptor studied here may fulfill other functions than binding lactoferrin. Preliminary studies with liver homogenates and detergent extracts of liver show binding in vitro.


Identification of the N gene protein of bacteriophage lambda.

The N gene protein, pN, of bacteriophage lambda stimulates early gene transcription by allowing mRNA chain elongation to proceed into genes distal to transcription termination sites normally recognized by the Escherichia coli transcription termination protein rho. pN has previously eluded detection on sodium dodecyl sulfate/polyacrylamide gels because of its small size, its instability, and the difficulty of distinguishing pN itself both from host proteins and from other early lambda proteins whose synthesis depends on pN action. These problems have now been overcome and we find that the major form of pN present in crude cell extracts of infected cells has an apparent molecular weight of 13,500. lambdabio256, a deletion-substitution mutant terminating in N, codes for a shorter pN of molecular weight 12,500. A nonsense fragment of 10,500 molecular weight coded by lambdaN(am7) has also been identified. These conclusions are based on examination of the electrophoretic profiles of the proteins synthesized after infection of UV-irradiated E. coli by various lambdaN(-) temperature-sensitive, nonsense, and deletion-substitution mutants. It has also been possible to distinguish pN itself from other early lambda polypeptides by infecting ron(-) cells with either lambdaN(mar) phage allowing pN synthesis but not pN action or lambdaN(am) phage defective in pN synthesis and pN action. Our results together with previous data are discussed with respect to the possible existence of multiple molecular weight forms of pN and the location of the coding sequences in the N gene region.


Butyrate suppression of histone deacetylation leads to accumulation of multiacetylated forms of histones H3 and H4 and increased DNase I sensitivity of the associated DNA sequences.

Exposure of HeLa cells to Na butyrate leads to an accumulation of multiacetylated forms of histones H3 and H4. Our studies of histone acetylation in HeLa S-3 cells show that 7 mM butyrate suppresses the deacetylation of histones without influencing the rate of radioactive acetate incorporation. An alteration in nucleosome structure in highly acetylated chromatin is indicated by an increased rate of DNA degradation by DNase I. A close association of acetylated histones with the DNase I-sensitive sequences is confirmed by the finding that histones remaining after limited DNase I digestion are depleted in the multiacetylated forms of histones H3 and H4. DNase I treatment has also been found to selectively release [3H]acetyl-labeled H3 and H4 from avian erythrocyte nuclei under conditions previously shown to preferentially degrade the globlin genes in erthyrocyte chromatin. Our results are consistent with the view that histone acetylation provides a key to the mechanism for altering chromatin structure at the nucleosomal level, and that this may explain the selective DNase I sensitivity of transcriptionally active DNA sequences in different cell types.


Novel enzyme from rat liver that cleaves an ADP-ribosyl histone linkage.

A novel enzyme that splits a bond between ADP-ribose and histone was discovered and partially purified from rat liver cytosol. The 105,000 X g supernatant of rat liver homogenate was precipitated by 45% saturated ammonium sulfate and then chromatographed on a DEAE-cellulose column. The enzyme activity was eluted in a single peak at about 0.2 M NaCl and clearly separated from poly(ADP-ribose) glycohydrolase which came out at 0.13 M NaCl. In contrast to the latter enzyme, this new enzyme catalyzed the spliting of a linkage between ADP-ribose and a protein portion in mono ADP-ribosylated histone H2B but little, if any, of the glycosidic ribosyl(1"-2') ribose bonds within poly(ADP-ribose). Analysis of the reaction product by paper chromatography and Dowex 1 column chromatography indicated that the split product contained the ADP-ribose moiety but was not exactly identical with ADP-ribose. Available evidence suggested that it was either an altered ADP-ribose molecule produced by a structural rearrangement or ADP-ribose itself linked to an unidentified compound. The enzyme had a pH optimum of about 6.0 and was inhibited by 80-90% in the presence of 5 mM ADP-ribose.


Nature of the gel to liquid crystal transition of synthetic phosphatidylcholines.

High sensitivity differential scanning calorimetry is employed in the study of the gel to liquid crystal phase transition of a highly purified sample of dipalmitoylphosphatidylcholine in multilamellar bilayer suspension. It is concluded from the calorimetric data that the purity of the sample is better than 99.94 mol % and that the transition closely approximates an isothermal first-order transition.


Effect of calcium on structure and function of a hyaluronic acid matrix: carbon-13 nuclear magnetic resonance analysis and the diffusional behavior of small solutes.

Natural-abundance (13)C NMR at 25.16 MHz has been used to study a 2.5% matrix of hyaluronic acid at various degrees of polymerization and at various ionic strengths. Peak assignment is facilitated by comparing proton-decoupled and off-resonance-decoupled spectra of a hyaluronidase-depolymerized matrix with spectra from relevant monosaccharides. In contrast to the spectrum following depolymerization, the spectrum for intact matrix has considerable broadening, particularly for peaks assigned to the N-acetylglucosamine moiety. This is most dramatic for the hydroxymethylene carbon. With the addition of Ca(2+) above 5 mM these broadened peaks narrow and approach the sharpness observed for the hyaluronidase digest. There is no shift in resonance peak positions. These changes are quantitatively less impressive if Na(+) is substituted for Ca(2+). The data suggest the existence of a considerable degree of order in regions of the matrix at physiological concentrations of Ca(2+). Within such a matrix the translational movement of lysine and glucose is enhanced relative to that in a matrix of agarose. Further addition of Ca(2+) abrogates not only matrix order, but the enhanced diffusivity as well.


Genetic recombination and complementation between bacteriophage T7 and cloned fragments of T7 DNA.

Frafments of phage T7 DNA have been cloned in Escherichia coli by using the plasmid pMB9. Such cloned fragments are able to recombine with infecting phages, thus providing a means to integrate the physical and genetic maps of T7 DNA. Approximately 65% of the T7 DNA molecule has been found in clones so far, and analysis of these clones has mapped genes 12-17 with an accuracy of about 1% the total length of T7 DNA. At least some cloned segments can supply T7 functions to infecting phages.


Synapse turnover: a mechanism for acquiring synaptic specificity.

Neurons are generated in chick retina that are able to form synapses with striated muscle cells for only a brief period during embryonic development. The ability to form synapses is lost with a half-life of 21 hr. Retina neuron-myotube synapses form rapidly but soon are terminated. Chick embryo spinal cord neurons also form synapses with muscle cells for only a limited time during development, but these synapses are long lived. These results show that different classes of synapses turn over at different rates and suggest that part of the specificity of synaptic circuits may be acquired during development by a process of selection based on synapse termination rates.


Changes in the three-dimensional structure of concanavalin A upon demetallization.

When the Mn(2+) and Ca(2+) ions normally present in concanavalin A are removed, the protein becomes incapable of binding saccharides. To explore the structural differences between the native and demetallized forms and their effects on the saccharide-binding properties of the protein, we have refined and compared the crystal structures of both forms. Refinement, carried out by automated difference Fourier methods, has revealed a number of differences between the two structures as well as minor differences between the two crystallographically independent monomers in the demetallized structure. Significant differences between the holo- and apoproteins are confined to the region where the metals are bound. These differences include a reorganization and disordering of the loop, consisting of residues 7-25, that contains all of the direct metal ligands of the protein. In some molecules, the side chain of arginine-228 appears to move into the metal-binding region, possibly compensating in part for the absence of the metal's positive charge. The cis peptide observed in the native protein at alanine-207 is apparently not present in the demetallized protein. The conformational differences affect many of the residues currently thought to be involved in the specific binding of saccharides.


Fe-O2 bonding and oxyheme structure in myoglobin.

In the polarized electronic absorption spectrum of oxymyoglobin in single crystals, charge-transfer states involving orbitals of the iron and dioxygen ligand are defined as probes of oxyheme orbital structure and coordination geometry. The spectrum of sperm whale oxymyoglobin is diagnostic of a bent (formula: see text) oxheme coordination geometry with totally spin-paired, ground-state electronic configurations of the iron and of the dioxygen ligand. In contrast, Aplysia myoglobin is distinguishably different in oxyheme structure, indicating that the geometry of Fe-O2 bonding in heme proteins can be altered by the protein environment.


Decreased microviscosity of membrane lipids in leukemic cells: two possible mechanisms.

Steady-state fluorescence polarization studies with the fluorescent lipid probe 1,6-diphenyl 1,3,5-hexatriene were done to determine the degree of microviscosity of cellular membrane lipids and serum lipoproteins in human normal donors and leukemic patients. The results show a marked decrease in microviscosity of cellular membrane lipids in both intact lymphocytes and isolated cellular plasma membranes obtained from leukemic patients in clinical relapse as compared to intact lymphocytes and isolated cellular plasma membranes obtained from normal donors and leukemic patients in complete clinical remission. Concomitant to these dynamic changes in cellular membrane lipids, the degree of microviscosity of lipids in the blood serum of leukemic patients in clinical relapse is markedly reduced as compared to serum obtained from normal donors and leukemic patients in complete clinical remission. Moreover, an in vitro incubation of leukemic lymphocytes with normal low density lipoproteins results in an increased microviscosity of cellular membrane lipids. In addition to the interrelation between cellular membrane lipids and serum lipoproteins, plasma membrane vesicles with a high degree of lipid microviscosity were isolated from the blood serum and pleural effusion of leukemic patients in clinical relapse. Such membrane vesicles could not be detected in normal serum. Therefore, we suggest that the two major mechanisms associated with the decreased microviscosity of membrane lipids in human leukemic cells are an abnormal exchange in lipids between the leukemic cell surface membrane and leukemic serum lipoproteins and an exfoliation of plasma membrane vesicles with a high degree of microviscosity from the cell surface of leukemic cells.


Interactions between adenovirus, a tumor promoter, and chemical carcinogens in transformation of rat embryo cell cultures.

The tumor-promoting agent 12-O-tetradecanoyl-phorbol-13-acetate (TPA) caused a 2- to 3-fold enhancement of transformation of secondary rat embryo cells that had been injected with a temperature-sensitive mutant of adenovirus type 5(H5ts 125). In addition, transformed foci appeared earlier and were larger in cultures grown in the presence of TPA. The addition of TPA could be delayed until up to 7 days after viral injection and still enhancement was observed. Exposure of the cells to 7,12-dimethylbenz[a]pyrene prior to H5ts125 infection also resulted in a 2- to 4-fold enhancement of transformation, and this enhancement was further augmented 2- to 3-fold when cells were grown in TPA after virus infection. Whereas TPA did not enhance the cloning efficiency of normal rat embryo cells, it did enhance the cloning efficiency of isolated colonies of adenovirus-transformed cells when these were grown alone or cocultured with a 100-fold excess of normal rat embryo cells. The enhancement of adenovirus transformation by TPA appears to be due to its ability to facilitate expression of the transformed state rather than an effect on virus uptake or integration.


Carbohydrate structure and cell differentitation: unique properties of fucosyl-glycopeptides isolated from embryonal carcinoma cells.

From embryonal carcinoma cells labeled with fucose, two main classes of glycopeptide products of Pronase digestion can be distinguished by Sephadex G-50 column chromatography: one eluted near the excluded volume and a smaller one. The large fucosyl-glycopeptides are scarcely present in differentiated cells derived from embryonal carcinoma cells (i.e., fibroblastlike cells, myoblasts, and parietal yolk-sac carcinoma). During in vitro differentiation of embryonal carcinoma cells, these large glycopeptides disappear almost completely. The small glycopeptides were analyzed by paper electrophoresis, concanavalin A-Sepharose affinity chromatography, and digestion with an endoglycosidase. The major components of these glycopeptides from embryonal carcinoma cells appear to be different from complex glycopeptides known to occur in adult cells. The glycopeptide pattern of mouse preimplantation embryos resembles that of embryonal carcinoma cells. These results suggest that the carbohydrate profile changes fundamentally during early stages of mammalian development.


Expression of a high molecular weight cell surface glycoprotein (LETS protein) by preimplantation mouse embryos and teratocarcinoma stem cells.

The expression of a high molecular weight cell surface glycoprotein (LETS, fibronectin) by preimplantation mouse embryos as well as cultured teratocarcinoma stem cells was detected by using indirect immunofluorescent staining. When each stage of preimplantation embryonic development was tested for the presence of LETS protein, none was observed on two-cell, four-cell, or eight-cell embryos, or on the morula or the outer cell layer (trophectoderm) of the early or late blastocyst. However, when the inner cell mass was isolated by immunosurgery, positive staining was observed. The intensity of the staining was significantly greater on the inner cell mass isolated from the expanded (day 4) blastocyst than on that from the early (day 3) blastocyst. Certain established cell lines of teratocarcinoma stem cells (embryonal carcinoma cells) also express cell surface LETS protein. "Nullipotent" (Nulli-SCC-1) as well as pluripotent (PSA 1) embryonal carcinoma cell lines have deposits of LETS protein concentrated in areas of cell-cell contact. In addition, a teratocarcinoma-derived endodermal cell line (PYS) was found to be capable of depositing LETS onto the substratum in a fibrillar network.Taken together, our results indicate that LETS protein is synthesized at a specific stage of preimplantation mouse embryonic development. In particular, they suggest that LETS protein is a product of the embryonic ectoderm, and that some types of embryonic endoderm are also capable of synthesizing this protein.


Isolation, preliminary characterization, and interferon antagonistic effect of a mammalian lectin-like substance.

We have isolated from mouse costal cartilage a tissue antagonist of interferon (TAI) which accelerates the decay of an established antiviral state. The effect is reminiscent of substances previously isolated from the basement membrane of human amnion. Since we have recently shown that phytohemagglutinin can mimic the biological effect of TAI, we have explored the possibility that TAI could be an animal lectin-like material. First, TAI agglutinates mouse cells; second, this cell agglutination is inhibited by some sugars. Preliminary characterization indicates that the active molecule is a protein. After Sephadex G-100 gel filtration, TAI is found in a peak of 150,000 molecular weight. When purified by isoelectric focusing, this peak shows maximal activities corresponding to an isoelectric point of pH 8.8 TAI binds to polysaccharide residues of the cell membrane which could be its primary site of action, comparable to phytohemagglutinin.


Coupled cell-free synthesis, segregation, and core glycosylation of a secretory protein.

mRNA from rat mammary glands 13-15 days post partum was translated in a wheat germ cell-free system either in the absence or in the presence of ribosome-denuded membranes prepared from isolated rough microsomes of dog pancreas. Newly synthesized alpha-lactalbumin was identified by immunoprecipitation with a monospecific rabbit antiserum against rat alpha-lactalbumin and was characterized by partial amino-terminal sequence determination and by lectin affinity chromatography. In the absence of membranes a presumably unglycosylated form of alpha-lactalbumin was synthesized that bound neither to concanavalin A-Sepharose nor to Ricinus communis lectin-agarose and that contained an amino-terminal signal peptide region comprising 19 amino acid residues. In the presence of membranes a processed form was synthesized that lacked the signal peptide portion and that had an amino-terminal sequence identical to that of mature alpha-lactalbumin. Furthermore, this processed form was found to be segregated, presumably within the microsomal vesicles, because it was resistant to post-translational proteolysis. It was also found to be glycosylated, and because it bound to concanavalin A-Sepharose, from which it could be eluted specifically by alpha-methyl mannoside, but not to R. communis lectin-agarose, it was presumably core-glycosylated. Processing, segregation, and core glycosylation were observed to proceed only when membranes were present during translation and not when they were added after translation.


Enzymatic synthesis and rapid translocation of phosphatidylcholine by two methyltransferases in erythrocyte membranes.

The synthesis of phosphatidylcholine from phosphatidylethanolamine is carried out by two methyltransferases in erythrocyte membranes. The first enzyme uses phosphatidylethanolamine as a substrate, requires Mg2+, and has a high affinity for methyl donor, S-adenosyl-L-methionine. The second enzyme methylates phosphatidyl-N-monomethylethanolamine to phosphatidylcholine and has a low affinity for S-adenosyl-L-methionine. The first enzyme is localized on the cytoplasmic side of the membrane and the second enzyme faces the external surface. This asymmetric arrangement of the two enzymes across the membrane makes possible the stepwide methylation of phosphatidylethanolamine localized on the cytoplasmic side and facilitates the rapid transmembrane transfer of the final product, phosphatidylcholine, to the external surface of the membrane. A mechanism for an enzyme-mediated flip-flop of phospholipids from the cytoplasmic to the outer surface of erythrocyte membranes is described.


Induction of maturation in Xenopus laevis oocytes by a steroid linked to a polymer.

A progesterone analog has been covalently linked via an amide bond to polyethylene oxide (molecular weight, 20,000). This macromolecular steroid molecule displays the biological activity of progesterone in inducing meiotic maturation when incubated with Xenopus laevis oocytes (stage VI) in vitro. Its efficiency (half-maximum effective concentration, 30 muM) is approximately 10 times lower than that of its low molecular weight homolog (3 muM). Control experiments with polyethylene oxide and an estradiol derivative (up to 1 mM) assessed the specificity of the progesterone macromolecular analog. Uptake experiments using radioactive derivatives revealed a small (if not negligible) intake of the macromolecular progesterone analog by the oocytes compared to that of free steroids, and no parallelism was found between radioactivity incorporation and effect. The possibility of cleavage of the macromolecular derivative during the incubation was ruled out. Furthermore, injection of the polymer-linked progesterone into the oocytes did not induce maturation. These observations suggest that the macromolecular progesterone analog itself is responsible for the biological effect and that the presence of this compound inside the cell is neither necessary nor sufficient for triggering reinitiation of meiosis. These conclusions are in agreement with the proposal that interaction with the plasma membrane of the oocyte is necessary for progesterone action in this particular system, in contrast to the case of somatic cells which have intracellular steroid receptors.


Tumorigenicity and its suppression in cybrids of mouse and Chinese hamster cell lines.

The effect of cytoplasm upon the expression of tumorigenicity was examined with a pair of mouse and a pair of Chinese hamster cell lines, in intraspecies cybrids formed by reciprocal fusions between either tumorigenic or nontumorigenic cells and cytoplasms derived from them. With the mouse cells, 3T3 and the simian virus 40-transformed line SVT2, the cybrid clones were tumorigenic when SVT2 cells were fused with 3T3 cytoplasts, but not in the reciprocal fusion. With the hamster cells, CHEF/18 and the spontaneous transformant CHEF/16, however, tumorigenicity was partially suppressed in cybrid clones formed by fusion of tumorigenic CHEF/16 cells with CHEF/18 cytoplasts; cybrids were nontumorigenic in the reciprocal fusion. Thus, cybrid analysis has shown that tumorigenicity is not cytoplasmically transmitted in these two cell pairs, but suppression of tumor-forming ability may be cytoplasmically transmitted in the hamster cybrids.


Complement C3 convertase: cell surface restriction of beta1H control and generation of restriction on neuraminidase-treated cells.

The alternative or properdin pathway of complement is primarily controlled by the endopeptidase C3b inactivator (C3bINA) and the nonproteolytic glycoprotein beta1H. The molecular mechanisms of control were investigated by performing binding studies of radiolabeled complement proteins to C3b bearing sheep erythrocytes (E(S)C3b). C3b was found to have distinct binding sites for beta1H, C3bINA, Factor B, and properdin. beta1H binding increased C3bINA binding 30-fold, while Factor B binding prevented C3bINA action on C3b and was competitive with beta1H binding. Properdin binding, which facilitates Factor B interaction with C3b, had no effect on the beta1H and C3bINA sites. Activators such as rabbit erythrocytes (E(R)) have previously been shown to interfere with the effectiveness of the control by C3bINA and beta1H, thereby allowing unrestricted formation of C3 convertase. Such restriction of control does not occur on the surface of E(S), a nonactivator of the alternative pathway. On the basis of comparative binding studies, restriction of control is explained entirely by reduced binding of beta1H to E(R)C3b relative to E(S)C3b. Access of properdin, Factor B, C3bINA, and the Fab fragment of anti-C3 to the two cell types was unrestricted. Restriction of beta1H control could be generated on the surface of E(S) by removal of cell-surface sialic acid with neuraminidase (acylneuraminyl hydrolase; EC 3.2.1.18). This enzymatic treatment converted E(S) from a nonactivator to an activator of the alternative pathway.


Studies on the murine Ss protein: demonstration that the Ss protein is functionally the fourth component of complement.

This report describes a simple and reproducible assay for the fourth component of murine complement (C4). Utilizing this assay, experiments are described which conclusively demonstrate that the murine Ss protein (serum substance) is functional C4 and that ascitic fluid may be used as a source for isolating Ss. The evidence that Ss is functional C4 is: (i) pretreatment of mouse plasma with F(ab')2 anti-Ss completely abrogated the C4 hemolytic titer; and (ii) in the isolation of the Ss protein from ascitic fluid, Ss immunochemical activity and C4 functional activity coeluted in each step of a four-step purification procedure.


Model for the incidence of embryonal cancers: application to retinoblastoma.

The two-mutation theory of cancer initiation hypothesizes that some cancers originate after two successive mutations, of which the second mutation is always somatic and the first mutation may be germinal (hereditary cases) or somatic (nonhereditary cases). A quantitative model using the Poisson distribution is developed for ages at diagnosis for hereditary and nonhereditary cases. This model relates age-specific incidence data explicitly to the number of divisions of embryonal cells and to rates of somatic mutations per cell division. A good fit is obtained when the model is applied to data on ages at diagnosis for one such embryonal tumor, retinoblastoma.