Daunomycin, cytosine arabinoside and 6-thioguanine (DAT) vs vincristine, cytosine arabinoside and 6-thioguanine (VAT) in the induction treatment of acute nonlymphocyte leukemia: a randomized collaborative study.

One hundred patients were entered in a cooperative study comparing the efficacy of two different regimens in the induction treatment of acute nonlymphocytic leukemia (ANLL). Patients were randomly allocated to receive either the DAT or VAT combination; half of the patients were also randomized to receive CNS prophylaxis including intrathecal methotrexate + prednisone and cranial irradiation. Consolidation and maintenance therapy were uniform in responding patients. Out of 82 evaluable patients 41 (50%) attained complete remission (CR) with no significant difference between the two regimens. Median remission duration was slightly longer in the DAT group (32.5 vs 22 weeks); median survival was 34 weeks for all evaluable patients with no difference between the two schedules. Meningeal relapse occurred only in two patients after 19 and 99 weeks of continuous remission. Fourteen patients are still alive after 61 to greater than or equal to 155 weeks, of whom seven are in their initial remission (six in the DAT and one in the VAT group). We conclude that 1) DAT and VAT are equally effective in inducing CR in a high proportion of ANLL patients; 2) until marrow remission can be prolonged significantly, preventing CNS leukemia will not have any significant impact of the course of ANLL.


[Results of polychemotherapy in the advanced stages of cervico-facial tumors].

An association of cyclophosphamide, fluorouracil and methotrexate already employed with success against solid tumours in other sites was used in the treatment of 62 patients with advanced tumours of the head and neck. Complete (40.32%) and partial (43.55%) regressions were obtained relatively quickly. A satisfactory period of remission occurred in patients for whom no further treatment had seemed possible, either on account of the extent of the disease, or because surgery and/or radiotherapy had already been performed.


[Cryotherapy, polychemotherapy and radiotherapy in the treatment of oral carcinoma. Preliminary note].

Following a short introduction to outline the therapeutic approach to primary carcinoma of the oral cavity, technical modalities, and the techniques of using this new type of cryo-polychemo-radiotherapeutic association are reviewed. The extremely interesting and flattering results obtained in 30 cases of patients suffering from this form of cancer must, however, be considered fairly modest if allowance is made for the time limits (insignificant). Results will be reviewed when a longer period has gone by.


[Prosthetic rehabilitation by means of removable prostheses with distribution of force by means of wires, in cases of subtotal edentia].

Dental arches, where almost all the teeth are missing, have been rehabilitated by prostheses leaving one or at most three contiguous teeth. Cases like this have been resolved with the use of a removable prosthesis with stress distribution by wires so as to obtain axial load on the abutment tooth and uniform distribution on the remaining tooth or teeth and on the alveolar mucosa. Examination 7 cases followed up for a period varying from 2 tp 13 years has shown that such prostheses are extremely functional.


[Importance of form in dental implantation].

Thirteen implantations have been made in the mandibular arches of 6 dogs. They consisted of commerical titanium blades and screws and of replicas of extracted dental roots variously shaped and made of titanium. All replicas and some of the commerical implantations were covered with pure aluminium using the plasmajet tecnique. Clinico-radiological, histopathological and microradiographical observation showed that only some of the replicas of dental roots give promising results.


[Effects of the combination of magnesium fluosilicate and manganese on experimental carries in white rats].

The pattern of experimental caries was examined in rats treated with magnesium fluosilicate associated with manganese and a cariogenic diet. A 33.34% fall in the incidence of caries was observed with respect to a control series, whereas Mg fluosilicate and manganese administered alone caused 30% and 20% reductions respectively. It was also noted that a slight toxic action, expressed as a 3 g fall in body weight, was found with the Mg fluosilicate dose employed; this effect was attenuated by simultaneous administration of manganese.


Chemoimmunotherapy of acute leukemia in children.

Comparative results of chemo- and chemoimmunotherapy with Soviet strain of BCG vaccine in children with acute leukemia are presented. The immunotherapeutical regimen includes 20 intracutaneous injections of 0.1 mg BCG vaccine given weekly during 5--6 months with subsequent 5--6 months intervals simultaneously with maintenance chemotherapy. In the control group children were only given conventional chemotherapy. The median duration of remission and survival in patients who had received BCG chemoimmunotherapy was 25.2 +/- 1.5 and 32.3 +/- 1.2 months respectively. In the group of patients with chemotherapy alone--13.2 +/- 0.9 and 21.8 +/- 1.1 months respectively (p less than 0.01). The best results were obtained in children who had received before BCG immunotherapy neuroleukemia prophylactic treatment.


The Milton survey: Part 1, General methods, height, weight and 24-hour excretion of sodium, potassium, calcium, magnesium and creatinine.

This paper describes the methods and general aims of a survey carried out on just over 1200 subjects (about 82% of the adult population) in Milton, a small town south of Dunedin. Data were collected on personal and family history, present medication if any, 24-hour dietary recall, psychometric scores, height, weight, skin-fold thickness, anthropometry, blood pressure, heart rate, electrocardiogram, mass miniature X-ray of chest, forced expiratory volume (FEV1), fasting plasma cholesterol and triglycerides, lipoprotein pattern and fasting and post-prandial blood sugar in a selected subgroup, and 24-hour urine volume and excretion of sodium, potassium, calcium, magnesium and creatinine. Mean values are given in the paper for each 10-year age group for height, weight, Quetelet's index, 24-hour urine volume, creatinine, sodium, potassium, calcium and magnesium. These and other results will be discussed in detail in subsequent papers.


Recurrent respiratory disease in Polynesian children.

Studies were performed on 13 Polynesian children who suffered from recurrent respiratory disease characterised by cough, wheeze, tachypnoea and radiological opacities. Only one was substantially underweight. Most had iron deficiency anaemia. Serum IgG levels were high with a less definite trend to raised IgA and IgM levels. Precipitating antibodies to cow's milk protein were found in 6 out of 10 children tested who were currently on cow's milk feeds. The proportion of rosette-forming lymphocytes was reduced in 8 of the 12 tested. The Mantoux test using using 10 TU of PPD was negative in 7 out of 10 children who had had BCG in the newborn period. The likelihood of cow's milk contributing to the respiratory disease in these children is discussed.


Carcinoma of the urachus: report of four cases and review of the literature.

Four cases of urachal carcinoma are presented. The features of this uncommon disease are outlined and the literature is reviewed briefly.


Epidemiology of cancers: I. Cancerous diseases of the large bowel.

A pilot study of cancerous diseases of the large bowel in Canterbury is described. The experience of 346 patients derived from the Cancer Registry for 1970-72 is presented. The feasibility of the method is demonstrated with the identification of guidelines for future work. The preliminary results suggest some special features in New Zealand. While demographic characteristics do not appear to help in defining persons at high risk from cancerous diseases of the large bowel, nevertheless other features of the natural history of this disorder indicate that it might be feasible to define high risk profiles.


Scalp infection after fetal monitoring in labour.

Neonatal scalp infection was reviewed over a four year period. The occurrence of such infection was found to be 0.14% of those infants monitored in labour using a spiral scalp electrode and 0.16% of those infants undergoing instrumental delivery. The duration of labour and the membrane-rupture delivery interval did not differ significantly from the total population. Scalp infection after monitoring occurred significantly later (day seven) than after instrumental delivery (day three) and was less often associated with positive bacterial cultures. Scalp damage is probably important in the aetiology of scalp infection, and the course of infection related to the degree of trauma.


The management of Hodgkin's disease in a regional centre over an eight-year period.

The staging and management of Hodgkin's disease has been discussed in relationship to experience obtained over an eight year period in a regional centre. Eighty cases were shown to be evaluable and the data pertaining to them has been discussed. A complete remission has been achieved by either wide-field radiotherapy or combnation chemotherapy in 78 percent of these cases. Sixty-six percent of the evaluable patients were alive and well five years after their diagnosis. The importance of interdisciplinary co-operation in management and the referral of such patients to a regional centre has been stressed.


Urinary tract obstruction and renal failure due to uterine prolapse.

Three women are described in whom obstructive uropathy was found secondary to uterine prolapse. Two of these patients had severe renal failure. It is important to exclude this condition in any woman presenting with renal functional impairment. Potentially damaging urinary tract obstruction should be considered in every patient with a uterine prolapse.


Doctor-patient communication in patients with arthritis.

Fifty patients with a chronic form of arthritis were given a standard interview concerned with establishing what knowledge they had of their disease and the drugs they were receiving for its treatment. Eighty-six percent of patients overall knew the name of their disease and 74 percent, the names of the drugs which they were receiving. Seventy-three percent of patients receiving non-steroidal anti-inflammatory drugs were unaware that these drugs were likely to have gastro-intestinal side effects even though about half of these patients were receiving either soluble aspirin or indomethacin. Younger patients were more likely to be aware of potential gastro-intestinal side effects (46 percent) than older patients (19 percent). Thirteen patients with rheumatoid arthritis were receiving myocrisin or penicillamine which are more likely to be associated with severe side effects than non-steroidal anti-inflammatory drugs. Four of five younger patients had some knowledge of the likely side effects of these drugs but none of the eight older patients knew this information. Written instructions should be given to older patients receiving these potentially toxic drugs.


Blood alcohol levels. How accurately can they be guessed?

Volunteers were asked to guess their blood alcohol level immediately before having their venous level measured by gas chromatography. Thirty-one percent were able to guess to within 20 mg per 100ml of the measured amount. There was a tendency for people to over-estimate if the level was low and to under-estimate the higher values. Blood alcohol concentrations ranged from nil to 295mg per 100ml with both high and low being recorded throughout the evening.


Lingual thyroid. Review of the literature and report of a case.

A case of lingual thyroid nodule was presented which demonstrates the need for oral surgeons to be aware of this entity in order to avoid the indiscriminate excision of lesions of the posterior dorsal surface of the tongue without proper medical examination of all systems of the body.


Postinjury myofascial pain dysfunction syndrome: its etiology and prognosis.

Clinical data are producted which appear to show that the response to therapy of myofascial pain dysfunction syndrome (MPDS) is much less favorable when the patient has been involved in a road traffic or similar accident which precipiated the condition. Reasons for this difference and the differences between postinjury MPDS and nonpostinjury MPDS patients may be a consequence of litigation and, in addition, may be due to the personality of the patient. Evidence to support this hypothesis is seen when the condition is compared with such disorders as low back pain. Further research is needed to explore the etiology and treatment implications of these differences.


Aphthous ulcer--a treatment complication. Report of a case.

Multiple theories of the cause and treatment of aphthous ulcers are described in the medical and dental literature. A case is reported in which there were posttreatment sequelae from the application of a sillver-nitrate stick (pencil) to an aphthous ulcer on the tongue. Compications subsequent to the use of silver nitrate may contraindicate the use of this agent in its highly concentrated form on the mucous membranes of the mouth.


The effect of oral rinses on organoleptic mouth odor ratings and levels of volatile sulfur compounds.

A test mouthwash, saline rinse, and no treatment were evaluated for their effects on organoleptic mouth odor ratings (OR) and corresponding concentrations of volatile sulfur compounds (VSC) in sixty-two subjects. The mouthwash was significantly superior to the saline rinse and to no treatment in reducing OR and VSC for 3 hours, which was the duration of the study.


The concept of hamartomas: its relevance to the pathogenesis of odontogenic lesions.

The term hamartoma should be applied only to tumorlike lesions which arise during development of the organ or tissue in question and which are apparently not neoplastic; it should not be used instead of benign tumor. A hamartoma is composed on an abnormal mixture of tissues native to the part, with an excess of one or more of these tissues. The relevance of the concept of hamartomas in the pathogenesis of odontogenic lesions is discussed briefly.


Ultrastructure of odontogenic keratocysts.

The ultrastructural features of an odontogenic keratocyst are described. The results of the investigation suggest that the basal lamina complex in the odontogenic keratocyst appears to be morphologically normal. The frequently observed splitting of the epithelium from the capsular tissue is marked by the separation of the epithelium and the basal lamina complex from the connective tissue. The morphologic features of the parakeratinized surface zone of the epithelium suggest that the odontogenic keratocyst may be characterized by "incomplete" parakertinization.


Focal epithelial hyperplasia in Abu Dhabi.

Focal epithelial hyperplasia of the oral mucosa has not been described in the United Arab Emirates, and its incidence in the Middle East is sporadic. A typical case with a familial background was seen in Abu Dhabi in an 11-year-old girl.


Bilateral dens invaginatus in the mandibular incisor region.

A case of bilateral dens invaginatus in the mandibular incisor regions has been reported. Radiographs demonstrated invaginations in both of the incisors in the mandibular incisor region and also demonstrated a periapical radiolucency. Clinical examination revealed two anomalous incisors and apparently there was congenital absence of two mandibular incisors. Dens invaginatus in the mandibular incisors would appear to be a most unusual occurrence.


Alveolar bone loss associated with vertical root fractures. Report of six cases.

Six cases of vertical root fractures accompanied by vertical bone loss are presented. Possible causes for these fractures are suggested. Four vertical fractures probably resulted from the use of excessive pressure at the time of obturation of the canals. One vertical root fracture may have been caused by the cementation of a post, and another may have been caused by the cementation of an inlay in an endodontically treated tooth. All of the fractures resulted in alveolar bone loss to the apical extent of the fracture lines. The definitive treatment in each case was extraction of the tooth or root amputation. In one case successful treatment was accomplished by apically positioning the flap.


Pulpal response to the application of phosphoric acid to dentin.

Experimental cavities were prepared in forty-six clinically intact premolars of young individuals to evaluate the biologic effects of 50 per cent phosphoric acid etchant containing 7 percent zinc oxide by weight. The experimental periods ranged from 30 minutes to 150 days. The remaining dentin thickness varied from 1.8 to 3.5 mm. The teeth treated with the acid showed no more than a moderate pulpal response at the varying time periods. All of the acid-treated teeth demonstrated pulpal imflammation in the absence of clinical pain. Acid etchants should not be used on exposed or unprotected dentin surfaces.


Brain abcess following dental infection.

A 48-year-old woman underwent root canal treatment of the upper left lateral incisor and lower right second premolar. Close to the conclusion of the endodontic treatment she complained about headaches. Later on, because of aggravation of her condition, with headaches, fever, malaise, weakness, and numbness of the right limbs, she was admitted to the hospital. The disease progressed to an epileptic state, with appearance of a right hemiparesis. A brain scan and carotid arteriogram revealed the presence of a mass occupying the left parietal space. Craniotomy disclosed an abscess containing yellow pus from which Streptococcus viridans was cultured. After thorough surgical cleansing of the area, removal of the bone for decompression, and treatment with ampicillin the patient improved gradually and slowly regained the mobility of her right side.


Comparison of xeroradiographs with occlusal and screen films.

Since the advent of xeroradiography, many questions have been asked about its image quality and speed relative to that of periapical and screen films. The experiment reported here measures the "resolution" of occlusal film, screen film, and xeroradiographs at different k Vp and mAs settings. The occlusal film was used instead of the periapical film because it has the same speed as a periapical film and is sufficiently large for the placement of the measuring instrument (a test pattern).


Dental x-ray spectrometry with an Si(Li) semiconductor.

Spectra from a dental x-ray apparatus with varying tube potentials and various added filters were measured with the use of an Si(Li) semiconductor. The mean energies of the spectra were calculated to estimate radiation qualities under various conditions. A phantom lip made of Arcylite was used to compare the attenuated quantities of radiation incident on the lip, according to the radiation quality, resulting from various tube potentials and added filtration.


Periapical actinomycosis. An unusual case report.

An unusual case of actinomycosis in the oral cavity is reported. The strict anaerobic qualities of Actinomyces israelii make laboratory culturing and growing of this organism difficult. Accordingly, the diagnosis was based on the histopathologic report. Because of the atypical clinical presentation, i.e., lack of any discharging sinus tract on the skin surface, and the histopathologic reports, it was postulated that the actinomycosis was superimposed on an existing granulomatous tissue.


Sealing quality of a temporary filling material.

Endodontic access cavities were prepared in forty extracted human teeth. The access cavities were obturated with Cavit and tested for leakage with methylene blue. The data suggested that at least a 3.5 mm. thickness of Cavit should be used in order to prevent leakage. Examination under the scanning electron microscope showed areas in which the constituents of Cavit were improperly mixed, which may lead to increased penetration.


Beam-guiding instruments for simplified dental radiography, with a training device.

Needs for exposure procedures in dental radiography with improved reliability and radiation hygiene, which can be applied with a minimum expenditure of time, have been documented. This article presents dental instruments designed for use in recently published general purpose x-ray procedures which may meet such requirements. They can provide accurate radiographs in endodontic, periodontic, and general practice and are adaptable to more time-consuming special-purpose procedures, including "serial identical" exposures and exposures of handicapped or anesthetized patients.


Xeroradiography of dental structures. II. Image analysis.

Xeroradiography provides an attractive, convenient, low-radiation-dose alternative to conventional intraoral film radiography. In this study the image properties of the two techniques were compared quantitatively and qualitatively. Over all, image quality was very similar, but xeroradiography delivered radiation doses seven to fourteen times less than conventional film radiography.


The syndrome of oral complaints: etiology and therapy.

Many patients who were examined at the oral medicine clinic have had almost no findings, despite their severe complaints, and we propose to diagnose their condition as SOC (syndrome of oral complaints). It is more frequent in middle-aged postmenopausal women who thereby tend to express all three determinants of illness--the biologic, the psychological, and the social. Such a triple assessment of the determinants, together with a thorough physical examination, brings the SOC into the diagnostic category of hypochondria. Therefore, it should be treated by the family dentist or by the general internist, with reassurance, local measures, and minor tranquilizers.


Clinical observations on osteoradionecrosis of the mandible. Report of a case with sequestration.

A case of osteoradionecrosis of the right mandible is presented. One and one half years after the onset of osteoradionecrosis, sequestration came about in a relatively large region, and the changes were observed on the radiographs over this period of time. A favorable course of the case is described in detail and discussed with reference to the literature.


Multiple fibroepithelial hyperplasias of the oral mucosa.

Reported is a rare case of multiple fibroepithelial hyperplasias of the oral cavity, and the suggestion is made that several previous authors may have confused this lesion with focal epithelial hyperplasia (Heck's disease). The differential diagnosis of multiple fibroepithelial hyperplasias is discussed in order to elucidate some of the confusions in the diagnosis of cases of multiple papules and nodules of the oral mucosa.


Wegener's granulomatosis: a case with primary mucocutaneous lesions.

A case of Wegener's granulomatosis is recorded in which the first signs were skin ulceration, segmental gingivitis, and oro-antral fistula. Diagnosis, management, and the significance of the microscopic appearances are discussed.


Focal epithelial hyperplasia in Israeli families of Libyan origin.

The occurrence of focal epithelial hyperplasia (Heck's disease) in seven Israeli families is reported. All the cases involved children and young adults whose parents had immigrated to Israel from Libya. The familial occurrence and possible etiologic factors of the condition are discussed. A genetic predisposition for the disease is suggested.


Reversible cavernous sinus involvement due to minor dental infection. Report of a case.

Described is a case of a cavernous sinus involvement due to minor dental infection. The early dental diagnosis and treatment reversed the course of cavernous sinus thrombosis.


Intravascular angiomatosis.

For 55 years much has been written about a peculiar benign intravascular process which can be misdiagnosed as angiosarcoma unless careful attention is paid by the pathologist to its benign cellular morphology and growth pattern. The cases reported in the literature demonstrate an increased incidence in the extremities and the head and neck, with a propensity for the subcutaneous tissues of the perioral region. Four cases are presented to further point out the perioral incidence and emphasize the differential diagnostic criteria for benignancy, including intravascular proliferation without atypical mitotic activity and necrosis in areas devoid of vascular differentiation.


Perineural invasion by squamous-cell carcinoma of the lower lip. Review of the literature and report of a case.

Although perineural metastases have been studied extensively for various regions of the body, there is a relative paucity of data on cancers of the head and neck region. A case of perineural metastasis from a squamous-cell carcinoma of the lower lip is presented, with symptoms mimicking other neurologic diseases.


The effects of partially filled polyethylene tube intraosseous implants in rats.

Polyethylene tubes obturated flush at one end and 1 mm. short at the opposite end with gutta percha and Grossman's cement as the cementing media were implanted in rat tibias. The gutta-percha, the set Grossman's cement, and the polyethylene implant were well tolerated by the rat intraosseous tissues. There was no significant inflammatory response at either the flush end or the short end of the polyethylene implant.


Osmotic fragility of leukaemic and normal lymphocytes.

The osmotic fragility of cord blood lymphocytes (CBLs), normal adult lymphocytes (NALs) and leukaemic cells was studied using hypotonic saline. Leukaemic cells were more resistant to hypotonic treatment than cord blood or adult lymphocytes. At low hypotonic saline concentrations (less than 0.2% NaCl), almost all of the NALs and CBLs were lysed after thirty minutes at 4 degrees C, whilst a large proportion of chronic lymphatic leukaemia cells remained intact. This phenomenon could conceivably be used to separate tumour cells from normal lymphoid cells.


On the mechanism of an increase of muscle performance and of vasodilation during emotional stress in man.

The mechanism of the increase of muscle performance and of vasodilation during emotional stress was studied. The "emotional" increment of voluntary performance does not depend on the level of blood supply to the working muscles, and the effect is maintained under conditions of cessation of arterial inflow. Augmentation of muscle performance is also observed during emotional stress when isometric contraction is evoked by electrical tetanic stimulation of the nerve, when the number of muscle fibres participating in the evoked response does not increase during emotional stress. The "emotional" vasodilation is greatly reduced in patients suffering from McArdle's syndrome, in whom the normal course of glycolysis in muscles is disrupted. It is suggested that acetylcholine liberated from sympathetic fibres causes the activation of glycolysis in muscles, which in its turn induces vasodilation in resting skeletal muscles and increase of muscle performance during emotional stress.


Cardiovascular effects of epidural analgesia. I. Thoracic epidural analgesia. An experimental study in sheep of the effects on central circulation, regional perfusion and myocardial performance during normoxia, hypoxia and isoproterenol administration.

Some circulatory effects of thoracic epidural analgesia (TEA) were investigated in splenectomized, open-chest sheep during normoxia, hypoxia and isoproterenol administration. During normoxia, TEA caused comparatively marked reductions in systemic arterial blood pressure, total peripheral resistance and cardiac output. A fall in heart rate was not compensated for by any rise in stroke volume. Myocardial contractility (LV dd/dt/IP) was not affected by TEA. The proportion of cardiac output diverted to the blocked area was markedly increased. Compensatory vasoconstriction was not observed within the unblocked area in six out of nine animals. Myocardial blood flow showed a pronounced reduction in accordance with the calculated changes of heart work, so that myocardial oxygen extraction remained unchanged. Studies under hypoxia revealed that cardiac responses to hypoxia in the sheep are mediated chiefly by neurogenic factors. TEA abolished the hypoxia-induced rise in heart rate but did not affect the increase in pulmonary vascular resistance caused by hypoxia. The administration of isoproterenol during TEA increased systemic arterial blood pressure, but due to further fall in total peripheral resistance it was not fully normalized. Cardiac output and heart rate increased markedly. Myocardial oxygen consumption and blood flow increased but did not reach control levels.


Nanosecond relaxation processes of phospholipid bilayers in the transition zone.

Ultrasonic relaxation spectra of dipalmitoyl lecithin vesicles have been recorded as a function of temperature over the frequency range 14-265 MHz. A relaxation process is observed with a time constant of about 10(-8) sec. At the mid-point of the crystalline-liquid crystalline transition (about 41.3 degrees), the relaxation amplitude is maximal. This suggests that the relaxation process is intimately associated with the order-disorder transition. Further support for this conclusion comes from the finding that the volume change of the reaction, as calculated from the relaxation amplitude at the transition midpoint, agrees with that determined independently by equilibrium dilatometry measurements of the deltaV of the transition. The results show that a major step in the transition occurs on a far shorter time scale than previously recognized. Similar fast processes have also been detected in dimyristoyl and distearoyl lecithin vesicles. From a consideration of various lines of evidence, it appears that the relaxation monitors the elementary step associated with the isomerization of lipid chains, such as kink formation through internal bond rotations, as the bilayer transforms between ordered and disordered phases.


Interacting enzyme systems at steady state: location of the phase transition in approximations of the mean field type.

We consider a phase transition "loop," obtained from a mean field type of approximate treatment of a closed steady-state Ising system. Where is the cut (stable path) across the loop located? The general procedure, in answering this question, is to pass to an open version of the same system and use the cut that appears automatically in this case (no loop is possible in an open system). This is equivalent to finding the point at which the two phases have equal total probability in the open system. It is shown here that this procedure, when applied to a system of two-state enzyme molecules, is formally equivalent to well-known thermodynamic methods (Maxwell's theorem, etc.). These can be applied directly to the closed system without considering the open system explicitly. However, for enzyme molecules with more than two states, the "thermodynamic" method generally fails and one must fall back on the open system procedure mentioned above. Practical implementation of this procedure is not easy.


Structure-function relationships of heparin species.

We have fractionated porcine heparin species of low molecular weight, with an average specific anticoagulant activity of 96 units/mg by affinity chromotography. Highly active and relatively inactive preparations of similar size were obtained with specific anticoagulant activities of 360 and 4 units/mg, respectively. The highly active heparin fraction possesses 1.1 additional residues of glucuronic acid and 1.5 fewer residues of N-sulfated glucosamine per molecule compared to the relatively inactive species. This decrease in N-sulfated glucosamine appears to be secondary to a corresponding increase in N-acetylated glucosamine. This form also contains a tetrasaccharide sequence with a N-sulfated glucosamine at its reducing end as well as equivalent amounts of glucuronic acid and iduronic acid. Furthermore, the internal glucosamine residue of this sequence appears to be N-acetylated. Sufficient amounts of this tetrasaccharide sequence are present within the highly active preparation such that each molecule may be endowed with this structure. The relatively inactive product contains a significantly decreased quantity of this tetrasaccharide sequence such that only [unk]20% of these molecules may possess this structure. The mean distance between nonsulfated uronic acid residues of the highly active species is smaller than that separating similar residues of the relatively inactive product. In addition, a larger number of the nonsulfated uronic acid residues of the highly active material appears either to be present in a restricted region of the molecule separated only by glucosamine residues or to be located at penultimate positions within the polysaccharide chain.


Insulin receptor: interaction with nonreceptor glycoprotein from liver cell membranes.

In crude receptor preparations (either particulate or soluble) of rat liver membranes, the insulin receptor exhibits complicated binding kinetics (two binding plateaus, half-saturated at approximately 60 pM and 700 pM insulin) and an apparent chromatographic heterogeneity, suggested by the presence of two detectable, soluble insulin-binding components with apparent Stokes radii of 72 A and 38 A. In contrast, the insulin receptor isolated by affinity chromatography exhibits a simple binding isotherm (half-maximal saturation of binding at 700 pM insulin) without evidence for negative cooperativity and behaves as a single component (apparent Stokes radius of 38 A) upon chromatography on Sepharose 6B. The apparent discrepancies between the properties of the unpurified insulin receptor and the affinity-purified receptor can be attributed to the presence in crude preparations of a nonreceptor constituent(s) having properties consistent with those of a membrane glycoprotein. A glycoprotein fraction from such crude soluble membrane preparations, freed from insulin receptor and subsequently partially purified using concanavalin-A-agarose, when combined with affinity-purified insulin receptor, causes both a reappearance of the complicated binding kinetics and an increase in the receptor's apparent Stokes radius from 38 A to 72 A. Similar results are observed for a glycoprotein fraction obtained from rat adipocyte membranes but are not observed for an identical fraction isolated from human erythrocyte membranes. We conclude that the insulin receptor in rat liver membranes can interact with another nonreceptor membrane glycoprotein that may represent either a nonrecognition moiety of the receptor oligomer or an effector molecule to the biological action of insulin.


Synthetic peptide derivatives that bind to fibrinogen and prevent the polymerization of fibrin monomers.

A series of small peptides corresponding to the amino termini of the fibrin alpha- and beta-chains has been synthesized. The peptides glycyl-L-prolyl-L-arginyl-L-proline and glycyl-L-prolyl-L-arginylsarcosine are potent inhibitors of fibrin polymerization. Moreover, these peptides have a natural stability stemming from their inherent resistance to proteolysis because of the involvement of amino acids in each of their peptide bonds. The peptide glycyl-L-prolyl-L-arginyl-L-proline binds to fibrinogen and to fragment D, in both cases with an association constant of approximately 5 x 10(4); it does not bind to fragment E. The number of binding sites is two for fibrinogen and one for fragment D. The tripeptide glycyl-L-prolyl-L-arginine binds less tightly and is less than half as effective in preventing polymerization. The peptide glycyl-L-histidyl-L-arginyl-L-proline, which corresponds exactly to the amino terminus of the fibrin beta-chain, does not inhibit the aggregation of fibrin monomers under the conditions used. It does bind weakly to fibrinogen, however, suggesting the involvement of sites other than those binding the alpha-chain analogues. Various other peptides were found not to inhibit polymerization; these included glycine-L-proline, L-prolyl-L-arginine and glycyl-L-prolyl-L-seryl-L-proline. The last-named corresponds to the serine/arginine amino acid replacement previously reported for a defective human fibrinogen.


Site-specific initiation of a DNA fragment: nucleotide sequence of the bacteriophage G4 negative-strand initiation site.

The synthesis of the bacteriophage G4 negative strand is an example of the de novo initiation of a polynucleotide chain. This initiation is performed by the Escherichia coli replication protein dna G which selects a unique site on 5400-base positive-strand template. In this paper we present the nucleotide sequence of the G4 negative-strand initiation site. This is the template element recognized by the dna G priming protein. In conjunction with the sequence of the nascent negative strand, obtained by Bouché, Rowen, and Kornberg [Bouché, J.-P., Rowen, L. & Kornberg, A. (1978) J. Biol. Chem. 253, 765-769], the present data provide a description of a dna G-dependent origin of replication in which one knows the place at which polymerization starts at the nucleotide level.


Transposition of plasmid DNA segments specifying hydrocarbon degradation and their expression in various microorganisms.

The conjugative TOL plasmid (75 Mdal), specifying biodegradation of xylenes, toluene, and trimethylbenzene derivatives, undergoes dissociation in Pseudomonas aeruginosa PAO to a nonconjugative TOL(*) plasmid (28 Mdal) and a transfer plasmid termed TOLDelta (48 Mdal). The TOL(*) plasmid is rendered transmissible through introduction of a number of conjugative plasmids such as factor K, CAM, and TOLDelta but not by the FP2 derivative pR0271. Transfer of TOL(*) via factor K or TOLDelta is mediated by the formation of plasmid cointegrates; no recombination is observed with CAM. A recombinant RP4-TOL plasmid (76 Mdal), which has lost resistance to tetracycline, has been isolated. The TOL(*) segment can be transposed from this RP4-TOL recombinant plasmid to other antibiotic resistance plasmids such as R702. A segment of DNA, specifying salicylate degradation from SAL plasmid, was transposed onto pAC10, the TOL(*-) derivative of RP4-TOL recombinant plasmid, which has lost resistance to tetracycline but retains the transfer genes of RP4. Transposition of the salicylate degradative genes onto pAC10 results in the loss of kanamycin resistance. It has been possible to isolate SAL(+) segregants from pAC10[unk]SAL transposition derivatives that have lost the pAC10 plasmid. Such segregants harbor the salicylate degradative genes in the form of a nonconjugative plasmid (SAL(*)). Transfer of RP4[unk]TOL(*) or pAC10[unk]SAL(*) transposition derivatives to Escherichia coli, Salmonella typhimurium, Agrobacterium tumefaciens, or Azotobacter vinelandii results in the functional expression of the antibiotic resistance genes but not of the hydrocarbon degradative genes. Such genes, however, are fully expressed on being transferred back to Pseudomonas.


Reconstitution of cholera toxin-activated adenylate cyclase.

Reconstitution of adenylate cyclase activity responsive to stimulation by guanylyl-5'imidodiphosphate or NaF may be achieved by mixing dilute Lubrol 12A9-solubilized extracts of wild-type S49 membranes with membranes of an adenylate cyclase-deficient variant. Experiments using N-ethylmaleimide to inactivate components of the adenylate cyclase system indicate that distinct components from both wild-type detergent extracts and adenylate cyclase-deficient membranes are essential for reconstitution. These results and conclusions confirm those of E. M. Ross and A. G. Gilman [J. Biol. Chem. (1977) 252, 6966-6969]. Detergent extracts of cholera toxin-treated wild-type membranes yield a reconstituted adenylate cyclase as responsive to GTP as to guanylyl-5'-imidodiphosphate whereas, in the absence of cholera toxin treatment, GTP has little or no effect. Cholera toxin-treated adenylate cyclase-deficient membranes and Lubrol 12A9 extracts from them, however, fail to yield a reconstituted adenylate cyclase that responds to GTP with an increase in cyclase activity. Because treatment of the adenylate cyclase-deficient variants with cholera toxin is without effect on the reconstituted cyclase, we propose that the cholera toxin substrate is absent or altered in the adenylate cyclase-deficient phenotype.


Evidence that two forms of bovine erythrocyte cytochrome b5 are identical to segments of microsomal cytochrome b5.

Homogeneous preparations of two forms of soluble cytochrome b5 have been obtained from bovine erythrocytes by successive chromatography on DEAE-cellulose, Bio-Gel P-60, and DEAE-Sephadex. Although the two forms could be separated on disc gel electrophoresis, they appeared to have similar molecular weights of approximately 12,000 and identical visible absorbance spectra. The tryptic hemepeptides derived from the two forms of bovine erythrocyte cytochrome b5 are electrophoretically indistinguishable from each other and from the tryptic core hemepeptide derived from liver microsomal cytochrome b5. The bovine erythrocyte tryptic hemepeptide was purified to homogeneity; its amino acid composition was shown to be identical to that of tryptic hemepeptide from liver microsomal cytochrome b5. The amino acid compositions of the two isolatable forms of erythrocyte cytochrome b5 correspond well to the compositions of the 97- and 95-residue segments of native liver microsomal cytochrome b5 that begin at the NH2 terminus. These results agree with the hypothesis that soluble erythrocyte cytochrome b5 is derived from microsomal protein by proteolysis during erythroid maturation.


Regio- and stereoselectivity of various forms of purified cytochrome P-450 in the metabolism of benzo[a]pyrene and (-) trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene as shown by product formation and binding to DNA.

Highly purified cytochromes P-450(LM2) and P-450(LM4) and partially purified P-450(LM1), P-450(LM3b), and P-450(LM7) from rabbit liver microsomes exhibit different catalytic activities in the metabolism of benzo[a]pyrene (BzP) and (-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene [(-)trans-7,8-diol] in a reconstituted enzyme system. The two highly purified cytochromes also exhibit differences in the activation of BzP and (-)trans-7,8-diol to intermediates that bind to DNA, as well as in the stereoselective conversion of (-)trans-7,8-diol to the highly mutagenic and carcinogenic diol-epoxides r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10- tetrahydrobenzo[a]pyrene (diol-epoxide I) and r - 7,t - 8 - dihydroxy - c - 9,10 - oxy - 7,8,9,10 - tetrahydrobenzo[a]pyrene (diol-epoxide II). P-450(LM2) is more active than P-450(LM4) in the metabolism of BzP and in its conversion to products that bind to DNA. In contrast, P-450(LM4) is more active than P-450(LM2) in the metabolism of (-)trans-7,8-diol and in its conversion to products that bind to DNA. The ratio of activity (percent substrate metabolized) with BzP relative to that with (-)trans-7,8-diol is 21 for P-450(LM2) and 0.3 for P-450(LM4); P-450(LM1), P-450(LM3b), and P-450(LM7) gave intermediate ratios. Marked stereoselectivity in the oxygenation of the (-)trans-7,8-diol to the highly mutagenic and putatively carcinogenic diol-epoxides I and II was observed with P-450(LM4), whereas the other preparations showed less selectivity. The ratio of diolepoxide I to diol-epoxide II ranges from 0.3 for P-450(LM7) to 11 for P-450(LM4). The substrate specificity and regio- and stereo-selectivity of the different forms of cytochrome P-450 may regulate the balance between activation and detoxification pathways of BzP and therefore determine the susceptibility of individual tissues, strains, and species to the carcinogenic action of BzP.


Soluble biospecific macromolecule for purification of estrogen receptor.

An alternative to affinity chromatography purification of proteins based on the use of a water-soluble biospecific polymer has been applied to the estrogen receptor from calf uterus. The receptor (4S-trypsin form) was bound to a dextran-estradiol conjugate (molecular weight approximately 500,000) and the complex was isolated by gel filtration. Highly purified receptor was subsequently released from the dextran-estradiol conjugate by exchange with [3H]estradiol.


In vitro biosynthesis and chemical characterization of beta-lipotropin, gamma-lipotropin, and beta-endorphin in rat pars intermedia.

Three 3-hr incubations of pars intermedia cells from 40 rat pituitaries with [35S]methionine, [3H]lysine, and [3H]leucine sufficed for the identification and chemical characterization of biosynthesized beta-lipotropin, gamma-lipotropin, and beta-endorphin. From the molecular weight, migration on polyacrylamide gels, and sequence Met5, Lys9, Leu14,17, rat beta-endorphin was shown to be identical to its sheep homologue and no trace of Leu5 beta-endorphin could be detected. Rat beta-lipotropin differs from that of sheep in its elution properties on CM-cellulose, and its sequence shows Leu2,10,14, Lys20. Rat gamma-lipotropin shows the same NH2-terminal sequence as beta-lipotropin and is again different from its sheep homologue. The identification of rat beta-lipotropin was confirmed by its selective cleavage into beta-endorphin after trypsin digestion of the citraconylated peptide, a property not observed with rat gamma-lipotropin.


Cycloheximide resistance can be mediated through either ribosomal subunit.

Two cycloheximide-resistant mutants of Tetrahymena thermophila were analyzed to determine the site of their cycloheximide resistance. The mutations in both strains had been previously shown to be genetically dominant and located at separate loci (denoted Chx-A and Chx-B). Strains carrying these mutations were readily distinguished by the extent to which they were resistant to the drug. The homozygous double mutant was more resistant than either single mutant. Cell-free extracts of wild type and of the three mutant strains, assayed for protein synthetic activity by both runoff of natural mRNA and poly(U)-dependent phenylalanine polymerization, demonstrated that in vitro the mutants were all more resistant than the wild type. Further fractionation of the cell-free systems into ribosomes and supernates localized cycloheximide resistance to the ribosome for both Chx-A and Chx-B homozygotes. Ribosome dissociation and pairwise subunit mixing in the in vitro system indicated that ribosome resistance was conferred by the 60S subunit from one strain whereas resistance in the other strain was mediated through the 40S subunit. This was further confirmed by reconstruction of all four cycloheximide-resistance "phenotypes" by mixing ribosomal subunits from appropriate strains. This finding suggests that the mechanisms by which these mutations confer resistance to cycloheximide are different.


Role of nucleotides in tubulin polymerization: effect of guanosine 5'-methylene diphosphonate.

Incubation of purified rat brain tubulin with guanosine 5'-methylene diphosphonate [GMP(CH2)P] (1 mM), a GDP analog resistant to hydrolysis, results in the polymerization of 20-30% of the total tubulin present. Analogous incubations with GDP (1 mM) do not result in tubulin polymerization. Polymerization with GMP(CH2)P occurs in the presence of alkaline phosphatase (EC 3.1.3.1) under conditions that completely hydrolyze the likely phosphate donors (GTP, GDP, and GMP) as well as the potential product [GMP(CH2)PP] of the transphosphorylase activity present in purified tubulin preparations. Tubulin polymerization in vitro thus can occur in the absence of gamma-phosphate and phosphate bond hydrolysis at the exchangeable nucleotide-binding site of tubulin. Polymerization of tubulin by GMP(CH2)P is neither prevented nor reversed by concentrations of calcium (2 mM) that prevent microtubule assembly and disrupt already formed microtubules induced by GTP. However, tubulin polymerized with GMP(CH2)P is readily depolymerized by cold (4 degrees, 30 min). The possible involvement of GTP alpha-beta bond hydrolysis must be considered seriously as playing a role in the process of microtubule depolymerization.


Hydrogen peroxide production, chemiluminescence, and the respiratory burst of fertilization: interrelated events in early sea urchin development.

After fertilization of the sea urchin, Strongyl-ocentrotus purpuratus, a crosslinked fertilization membrane is formed; the crosslinks (dityrosine residues) are synthesized in a reaction catalyzed by an ovoperoxidase that is released from the cortical granules during fertilization. The substrate for ovoperoxidase activity, hydrogen peroxide, is generated by the egg coincident with the "respiratory burst" that follows parthenogenetic activation by the divalent ionophore A23187 or fertilization. This burst of oxygen consumption may be almost quantitatively accounted for by hydrogen peroxide evolution, as measured by the peroxidase-catalyzed quenching of scopoletin fluorescence. Neither the burst of oxygen consumption nor hydrogen peroxide production occurs when the inhibitor of cortical granule discharge, procaine, is present at fertilization. Fertilization or parthenogenetic activation with A23187 also is associated with a burst of light emission. This chemiluminescence is inhibited in vivo by inhibitors of the ovoperoxidase, such as 3-amino-1,2,4-triazole, phenylhydrazine, sulfite, or azide. A crude ovoperoxidase preparation catalyzes hydrogen peroxide-dependent chemiluminescence that is similarly inhibited. Thus, the bursts of oxygen uptake, peroxide production, and chemiluminescence appear to be several manifestations of the peroxidative system released at fertilization. This system may additionally be responsible for spermicidal activity and thus may act as a component of the block to polyspermy.


Embryonic neural retina collagen: in vitro synthesis of high molecular weight forms of type II plus a new genetic type.

Chicken neural retina cells from 6- to 7-day embryos were labeled with [(3)H]proline for 24 and 72 hr and the collagenous proteins of the medium were analyzed. Ninety percent of the collagenous proteins eluted from DEAE-cellulose columns as a peak near the middle of the gradient. Upon sodium dodecyl sulfate/polyacrylamide gel electrophoresis, this material consisted predominantly of a slowly migrating procollagen, a smaller amount of an intermediate migrating form, and some alpha-chain material. Limited enzymatic digestion of this DEAE peak material plus mapping of its CNBr peptides identified this material as a type II precursor. The remaining 10% of the collagenous proteins eluted from DEAE-cellulose at the end of the gradient. Electrophoresis of this fraction showed four major bands. Two migrated near the type II precursors, a third migrated somewhat more slowly, and the fourth was near beta-chain dimers. In addition, there were two minor bands. Limited pepsin digestion of this DEAE peak material produced two bands: one migrated slightly more slowly than alpha1, and the second, considerably more slowly. The CNBr peptide pattern of this material appears to be different from any previously described. Thus, neural retina cells in culture synthesize at least two genetically distinct classes of collagenous proteins. One represents precursors to type II. The second is composed of multiple, very high molecular weight forms which may represent precursors (procollagens) of a new genetic type(s) of collagen.


Disulfide-linked high molecular weight protein associated with human cataract.

A major high molecular weight disulfide-linked protein has been isolated from cataractous lenses. It is only present in the water-insoluble protein fractions. This species has not been found in normal lenses of comparable age. Upon reduction of this fraction, polypeptides having molecular weights of approximately 60,000, 43,000, and 20,000 as well as a noncharacterized heterogeneous species are released. Similar sized polypeptides have been found in various noncovalently linked aggregates in both normal and cataractous lenses. Examination of the disulfide-linked protein fraction indicates that approximately 70% of the sulfhydryl groups are in the oxidized state. Although little change in the sizes of the other major polypeptides in the water-insoluble fraction is observed upon reduction, these components were also found to contain an appreciable disulfide content. Such results indicate that the only major lens fraction containing disulfide-linked polypeptide is the high molecular weight species and that the disulfides present in the remaining fractions are either intrachain disulfides or link polypeptides to small peptides.


Isolation and characterization of mitochondrial DNA from Chlamydomonas reinhardtii.

Mitochondrial DNA (mtDNA) has been isolated from a mitochondrial pellet of Chlamydomonas reinhardtii. The mtDNA has a buoyant density of 1.706 g/ml in CsCl, a melting temperature of 87.9 degrees in standard saline citrate, and a nucleoside composition of 47.5% deoxyguanidine plus deoxycytidine with no odd nucleosides. Thermal denaturation and renaturation studies have shown that (i) mtDNA contains no extensive intramolecular heterogeneity nor significant base bias between the complementary polynucleotide chains and (ii) mtDNA renatures as a single homogeneous class with a kinetic complexity of 9.78 X 10(6) daltons. Although rare (less than or equal to 1%), both open and supercoiled circular mtDNA molecules have been observed in the electron microscope. Contour lengths of linear and open and closed circular molecules are all within the range of 4.0-5.4 micron with a mean of 4.67 +/- 0.30 micron. This size is similar to that of animal mtDNA but approximately 1/8 that of the higher plant mtDNAs. The magnitude of mtDNA reiteration in C. reinhardtii is estimated to be of the same order as that of chloroplast DNA.


Identification, localization, and role of fibronectin in cultured bovine endothelial cells.

We have examined bovine aortic endothelial cell cultures for the presence of fibronectin, a high molecular weight cell-surface glycoprotein. Sparse cultures contain fibronectin only on dorsal cell surfaces at regions of cell-cell contact, as detected by immunofluorescence. In contrast, when the endothelial cells reached confluence as a highly contact-inhibited monolayer, fibronectin was detected in an extracellular matrix underneath the cell monolayer but not on top of the monolayer. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis of isolated extracellular matrix revealed that a predominant component of the matrix is a protein of approximately 2.3 X 10(5) molecular weight, which has been identified as fibronectin.


Changes in genome composition of the Friend virus complex in erythroleukemia cells during the course of differentiation induced by dimethyl sulfoxide.

The Friend spleen focus-forming virus (SFFV) complex released by Friend virus-transformed erythroid cells has been analyzed with respect to changes in the genome composition that may occur during induction of erythropoiesis with dimethyl sulfoxide. It is shown that: (a) There are three types of virus particles, one with buoyant density 1.20 g/ml, one with density 1.17 g/ml (the density of the cloned lymphatic leukemia virus helper component of the complex), and a major fraction that has a density of 1.14 g/ml. (b) Three RNA subunits-35S, 32S, and 30S-have previously been shown to be detectable in the Friend virus complex. The 1.20-g/ml particles contain only 30S RNA, whilst the 1.14- to 1.17-g/ml particles contain a mixture consisting of predominantly 30S and 32S RNA and about 5-10% 35S RNA. (c) Induction of differentiation results in an increase in the 1.14-g/ml particles and 32S RNA. The amount of 30S RNA does not change. (d) Hybridization of the different genomic viral RNAs with full-length virus cDNA shows that the 30S RNA (of induced and uninduced Friend virus) is more closely related to the 32S RNA of the induced Friend virus than to the 32S RNA of the constitutively released Friend virus. (e) The 30S RNA contains SFFV-specific sequences. (f) A hypothesis is presented in which the induction of the new 32S RNA species is related to the increase of SFFV activity and to a specific function of the SFFV during induction of erythropoiesis.


Association of Thy-1 differentiation alloantigen with synaptic complexes isolated from mouse brain.

Conventional fractionation procedures were used in an effort to define the subcellular distribution of the Thy-1 alloantigen in whole mouse brain. After discontinuous sucrose density gradient centrifugation of isotonic postnuclear particulate fractions, the bulk of Thy-1 was recovered in regions of the gradients containing synaptosomes. The synaptosome fraction that banded at 1.2 M sucrose yielded a specific activity for Thy-1 significantly greater than the synaptosomes separating at 1.4 M sucrose. Osmotic lysis of both synaptosome fractions resulted in further enrichment in Thy-1 activity, with no concomitant decrease in yields. The synaptosomal membranes obtained in this way were subsequently treated with Triton X-100 and subjected to further density gradient centrifugation. Although the detergent treatment resulted in some loss of antigenic activity, the gradient fractions that contained Thy-1 also were found by electron microscopy to be richest in synaptic junctional complexes. These findings suggest that Thy-1 is associated with synaptosomes and synaptic junctional complexes and therefore may be involved in the formation and/or maintenance of synaptic connections.


Metabolic cooperation between mouse embryonal carcinoma cells and their differentiated derivatives.

Metabolic cooperation has been used as an index of intercellular communication between mouse embryonal carcinoma (EC) and other cell types. EC cells do not cooperate with differentiated cells of various properties or origins. In contrast they cooperate with cells of all other EC lines tested, including a human teratocarcinoma line, but with different efficiencies. This might reflect differences either in cell-cell interactions or in the formation of gap junctions. During  in vitro differentiation, EC cells appear to remain isolated from differentiated cell types.


Late S phase cells (Chinese hamster ovary) induce early S phase DNA labeling patterns in G1 phase nuclei.

Cells (Chinese hamster ovary) in G1 phase were fused with cells in late S phase to determine if a cell in late S phase can induce DNA synthesis in the nucleus of a G1 cell and, if so, to determine if the DNA synthesis so induced in a G1 phase nucleus has an autoradiographic pattern characteristic of early or of late S phase synthesis. The results indicate (i) that 89% of the G1 nuclei in late-S/G1 binucleates synthesized DNA, while only 2% of the control unfused G1 cells synthesized DNA, and (ii) that in all late-S/G1 binucleates the G1 nucleus was induced to synthesize early S phase DNA. These results are compatible with the idea that a cytoplasmically transmissible factor initiates DNA synthesis but that an intranuclear mechanism defines the temporal order of replication.


Free ribosomal RNA genes in Paramecium are tandemly repeated.

The genes coding for 17S and 25S rRNA in Paramecium tetraurealia were isolated. The macronuclear ribosomal DNA (rDNA) exists as relatively small, extrachromosomal molecules with both linear and circular forms. Electron microscopy and restriction endonuclease analysis revealed that the rDNA is arranged as tandem repeats with an average repeat size of 5.5 X 10(6) daltons. Some heterogeneity of repeat lengths was found both by electron microscopy and by restriction enzyme analysis. The rDNA does not snap back after denaturation. This study provides additional evidence that extrachromosomal rDNA may be a common feature among lower eukaryotes. However, in contrast to several other cases, the rDNA of Paramecium is not palindromic, but occurs as tandem repeats as in higher eukaryotes.


Structural characterization of H-2 antigens purified from mouse liver.

Papain solubilized H-2a histocompatibility antigens (H-2Kk plus H-2Dd) have been purified by a large-scale procedure that can routinely provide 2-3 mg of heavy chain from 1 kg of mouse liver. The heavy chains were homogeneous by sodium dodecyl sulfate electrophoresis. Disc gel electrophoresis resolved two protein bands that were identified as H-2Dd and H-2Kd by immune complex formation and autoradiography. Comparative amino acid composition and NH2-terminal sequence analyses of unfractionated H-2a, H-2Kk, and HLA suggested close structural relationships. However, the following observations suggest that papain cleaves these membrane bound antigens at different positions with respect to the COOH terminus: the molecular weight of the peptide portion of papain solubilized HLA is smaller than that of H-2 (30,000 versus 33,700); the COOH-terminal sequences are different; and, finally, papain-solubilized H-2 contains a free cysteine residue in addition to the two disulfide bridges that are present in both H-2 and HLA.


Identification of stage-specific polypeptides synthesized during murine preimplantation development.

[35S]Methionine-labeled extracts of mouse ova and preimplantation embryos were analyzed by two dimensional polyacrylamide gel electrophoresis. Of the 400-600 molecular species that have been resolved as distinct spots on autoradiograms of gels for every stage of development from unfertilized eggs to early blastocysts, particular attention has been paid to the identification of 36 of these proteins, each of which is expressed only for a portion of the period under investigation. These molecules are referred to as stage-specific polypeptides and they are biochemical markers of early embryonic development and differentiation.


Isolation and characterization of mutants of human diploid fibroblasts resistant to diphtheria toxin.

Stable mutants highly resistant to the protein synthesis inhibitor diphtheria toxin (dipr) have been selected in human diploid fibroblast cells at a frequency of 1-8 X 10(-6). Treatment of cells with mutagens, (e.g., ethylmethanesulfonate, nitrosoguanidine, and ICR-170), increased the frequencies of dipr mutants by 50- to 500-fold in different experiments, and the optimal expression time for dipr mutation was about 5 days. All mutants examined thus far have bred true, and no effects of cell density or cross feeding have been observed on the selection. Fluctuation analysis showed that the dipr mutation occurs in these fibroblasts at the rate of 5-6 X 10(-7) mutations per cell per generation. Protein synthesis in mutant extracts was resistant to diphtheria toxin, indicating that the dipr lesion in such mutants lies in the protein synthesis machinery. The characteristics of the dipr marker should make this system particularly useful for studies of quantitative mutagenesis in human diploid cells.


Biochemical evidence that "new" influenza virus strains in nature may arise by recombination (reassortment).

Oligonucleotide analysis of two avian influenza A viruses (Hav6N2 and Hav6Nav4) isolated in nature showed identical or almost identical patterns for the corresponding M and HA genes; 24 of 25 and 13 of 13 large oligonucleotides were indistinguishable by two-dimensional gel analysis. On the other hand, remarkable differences in the oligonucleotide patterns of the remaining genes were observed. Only 2 of 11 oligonucleotide spots of the NS gene, 10 of 27 spots of the NA/NP genes, and 22 of 49 spots of the P genes were indistinguishable between the two strains. On the basis of this observation that at least two genes of these viruses are virtually identical whereas others show easily detectable differences, we conclude that the two avian strains are related to each other by a recombinational event. In addition, it was found that animals in nature can be doubly infected with influenza viruses. Both lines of evidence strongly suggest that recombination is at least one mechanism by which "new" influenza virus strains emerge in nature.


Co-transfer of human X-linked markers into murine somatic cells via isolated metaphase chromosomes.

Transformation frequencies of 4 x 10(-5) were obtained in chromosome-mediated gene transfer experiments using human cell line HeLa S3 as donor and mouse cell line A9 as recipient. This high frequency of interspecific transformation was achieved by treating the recipient cells with dimethylsulfoxide in addition to other facilitators. The high frequency of transformation correlated positively with transgenome size on the basis of both co-transfer of linked markers and chromosome analysis. The syntenic human markers glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate:NADP(+) 1-oxidoreductase, EC 1.1.1.49) and phosphoglycerate kinase (ATP:3-phospho-D-glycerate 1-phosphotransferase, EC 2.7.2.3) were sometimes transferred together with the selected X-linked prototrophic marker hypoxanthine phosphoribosyltransferase (IMP: pyrophosphate phosphoribosyltransferase, EC 2.4.2.8) into murine somatic cells. Donor human chromosome material could be demonstrated cytologically in some of the transformed cell lines. Transformants exhibited various rates of loss of the human hypoxanthine phosphoribosyltransferase marker when grown under nonselective conditions. These results reveal a broader range of possible interspecific transgenome sizes than has been recognized in the past. The largest transgenomes consist of cytologically detectable donor fragments and contain syntenic markers that are not closely linked to the selected marker.


Mass isolation and fertility testing of temperature-sensitive mutants in Tetrahymena.

A set of 239 heat-sensitive (38 degrees) and 16 cold-sensitive (18 degrees) conditional mutants of Tetrahymena has been generated by combining techniques to manipulate large numbers of clones with a method for the selection of self-fertilized cells after mutagen treatment. A simple technique is presented for determining the fertility of individual clones; 179 of the clones in this set (71%) are fertile. The fertile conditional mutants have already been shown to have lesions in a number of diverse functions, including nucleic acid metabolism, mobility, cell cycle, and cortical pattern.


Subunit molecular weight and genetic variability of proteins in natural populations.

The relationship between subunit molecular weight and heterozygosity was studied in six different groups of organisms, i.e., 9 species of primates, 32 species of rodents, 56 species of reptiles, 12 species of salamanders, 64 species of teleost fishes, and 29 species of Drosophila. The correlation coefficient between them was positive in all groups, and the magnitude of correlation was roughly in agreement with the theoretical expectation under the mutation-drift hypothesis when the incomplete correlation between molecular weight and mutation rate was taken into account. Furthermore, the correlation was higher when the average heterozygosity was high than when this was low, as theoretically expected.


Base ratio, DNA content, and quinacrine-brightness of human chromosomes.

Human chromosomes were labeled with base-specific radioactive DNA precursors and examined autoradiographically to measure their DNA content and base ratio (percentage A-T base pairs). The requirement that incorporation of labeled bases be uniform during DNA synthesis was met by the use of inhibitors of de novo synthesis of DNA precursors. The genome was subdivided into 75 segments based on quinacrine banding, and the base ratio of each was calculated by a method that corrects for bias due to the scatter of grains about their source. Estimates of base ratio are shown to be sufficiently precise to detect variability among chromosomes and among segments within a chromosome. Analysis of these data and of measurements of the quinacrine fluorescence intensity of segments leads to the following conclusions. Base ratio is positively correlated with brightness, as predicted from independent in vitro studies. Larger chromosomes tend to have higher base ratios and to be brighter than smaller ones. The best prediction of the brightness of a segment must take into account not only its base ratio but also its DNA content. To explain these results, we suggest an evolutionary model in which chromosomes containing repeated sequences of A-T-rich DNA tend to grow by means of unequal sister chromatid and meiotic exchanges.


Milk precipitins, circulating immune complexes, and IgA deficiency.

Twenty-two patients with selective IgA deficiency were studied for the presence of serum precipitins to bovine milk, bovine and fetal calf serum, and circulating immune complexes. Fifty-nine percent had circulating immune complexes, 50% had milk precipitins, 23% had precipitins to bovine serum, and 13% had precipitins to fetal calf serum. All patients with precipitating antibodies against milk or against bovine or fetal calf serum had circulating immune complexes and the precipitin titers correlated with the amount of circulating immune complexes. After one IgA-deficient patient had drunk 100 ml of milk, studies of sequential serum samples showed the presence of casein in the circulation at 60 min and the appearance of increasing amounts of immune complexes for 120 min. These findings are interpreted to indicated that in human beings the IgA system may provide a major barrier to absorption of immunogens from the gastrointestinal tract.


Primary structure of murine major histocompatibility complex alloantigens: amino acid sequence studies of the cyanogen bromide fragments of the H-2Kb glycoprotein.

Radiochemical microtechniques have been used in the amino acid sequence analysis of five major CNBr fragments of the glycoprotein specified by the murine major histocompatibility complex gene H-2k(b). These fragments have been tentatively aligned and represent the NH(2)-terminal 80% of the intact molecule. All amino acids except Asp, Asn, and Gln have been assigned in 128 out of 149 possible positions in the NH(2)-terminal portions of each of these fragments. These assignments, which represent approximately 50% of the total sequence from these fragments, are listed below in the order of their alignment in the intact H-2K(b) molecule: IIIn, -PHSLRYFVTAVSRP(G)L(G)(E)PRYM; IIIa, EVGYV--TEFVRF-S-AE(A)PRYEPR(A)--M; Ib, E-EGPEYWERET-KAK(G)-E-SFR--LRTLL(G)YY--TK; Ia, AALITK-KWE-AGEAERLRAYLEGTC-E-L; Ic, ELVETRPAG-GTF-KWAS-VVPLGKE-YY(T). The unassigned positions represented by dashes in the above sequences may be tentatively assigned as Asp, Asn, or Gln. The NH(2)-terminal sequence obtained for the H-2K(b) molecule was compared to the limited sequence information available for other major histocompatibility complex gene products. An 84% homology (16 of 19 residues) to the H-2K(q) and H-2K(k) molecules, which are identical to one another in the positions compared, was observed. A similar comparison with 28 of the 31 NH(2)-terminal residues of HLA-B7 indicated 68% homology. Furthermore, significant homology was observed between H-K(b) and HLA-B7 in a region of glycosylation, which occurs between positions 85 and 100 in the two molecules.


Tuftsin (an Ig-associated tetrapeptide) triggers the immunogenic function of macrophages: implications for activation of programmed cells.

The immunoglobulin heavy-chain-associated tetrapeptide, tuftsin (Thr-Lys-Pro-Arg), known for its phagocytosis-stimulating activity, was found to augment the antigen-specific, macrophage-dependent education of T lymphocytes. The investigation of stereospecific characteristics of the tetrapeptide, by use of structural analogs with different modifications, revealed strict structural requirements for eliciting the immunogenic activity of macrophages. It was found that the most important moiety for its activity is the dipeptide Pro-Arg. This finding is of interest in view of the appearance of this particular dipeptide in other bioregulatory peptides, including many of the peptide hormones. The significance of the appearance of a common structure in such molecules, which may act through specific receptors on different target cells, is discussed.


Biochemical evidence for a deficiency of vitamin B6 in the carpal tunnel syndrome based on a crossover clinical study.

In a patient with severe carpal tunnel syndrome and a significant deficiency of vitamin B(6), the evidence for the deficiency was an extraordinarily low basal specific activity of the glutamic-oxaloacetic transminase of the erythrocytes (EGOT). This enzyme was also deficient in pyridoxal phosphate. The patient was treated with the recommended dietary allowance of pyridoxine, 2 mg/day, for 11 weeks, then 100 mg/day for 12 weeks, a placebo for 9 weeks, and again pyridoxine at 100 mg/day for 11 weeks. Sixty-one monitorial assays of EGOT over 48 weeks supported the following interpretations. (i) His diet permitted the development of a debilitating carpal tunnel syndrome. (ii) Treatment with pyridoxine at 2 mg/day reduced the deficiency of EGOT activity from about 70% to 50%, maintained a deficiency of pyridoxal phosphate, and relieved but allowed a marginal syndrome. (iii) Treatment at 100 mg/day for 12 weeks nearly achieved a "ceiling" level of EGOT and eliminated the deficiency of pyridoxal phosphate. (iv) After placebo for 7 weeks, the deficiencies of EGOT activity and pyridoxal phosphate reappeared, and clinical symptoms become worse. (v) Retreatment at 100 mg/day reestablished a "ceiling" EGOT, with no deficiency of pyridoxal phosphate, and the patient was asymptomatic. These data also support the concept that a deficiency of vitamin B(6) is significant in the etiology of the carpal tunnel syndrome. Mechanistically, a state of deficiency of the coenzyme seems to lower the level of the apoenzyme; a state of no deficiency of the coenzyme regulates a ceiling level of the transaminase. The latter state is presumably desired for health.


Anti-interferon globulin inhibits the development of glomerulonephritis in mice infected at birth with lymphocytic choriomeningitis virus.

Swiss mice infected at birth with lymphocytic choriomeningitis virus develop glomerulonephritis. Injection of potent anti-mouse interferon globulin at the time of viral infection inhibited the development of these renal lesions. We conclude that the production of endogenous interferon by this virus in the first few days of life plays an important role in the pathogenesis of this glomerulonephritis.


Direct determination of volume- and time-dependence of alveolar surface tension in excised lungs.

We measured alveolar surface tension directly by observing the spreading properties of fluid droplets placed by micropipette within individual alveoli. Alveolar surface tension in excised rat lungs at total lung capacity and 37 degrees was 29.7 +/- 1.4 mN-m-1. In lungs held at functional residual capacity, surface tension changed slowly, rising to 9 mN-m-1 at 30 min with a subsequent approximately linear rise to 16 mN-m-1 at 115 min. Thereafter it rose more rapidly (20 mN-m-1 at 140 min), suggesting that it was not asymptotically approaching an equilibrium value.


Effect of myasthenic patients' immunoglobulin on acetylcholine receptor turnover: selectivity of degradation process.

Antibodies in the sera of patients with myasthenia gravis are believed to play an important role in the pathogenesis of the disorder. They have recently been shown to accelerate the degradation of acetylcholine receptors in cultured mammalian skeletal muscle and at intact neuromuscular junctions. To elucidate the mechanism of the antibody-accelerated degradation process, we have prepared cultures in which one set of acetylcholine receptors was exposed to myasthenic immunoglobulin while a second set of acetylcholine receptors, newly incorporated after exposure to the immunoglobulins, was not. The set of acetylcholine receptors with bound myasthenic immunoglobulin was degraded at 2 to 3 times the normal rate, while the second set of acetylcholine receptors without bound immunoglobulin was degraded at the control rate. This suggest that the binding of antibody from myasthenic patients alters the acetylcholine receptors in some way that causes them to be selected for preferential degradation by the muscle cells. New synthesis and incorporation of the acetyl-choline receptors into the surface membrane of cultured skeletal muscle was unaffected by exposure to myasthenic immunoglobulin.


Proteolytic activity in erythrocyte precursors.

Thalassemia is characterized by unequal rates of synthesis of the alpha and beta globin chains that are part of the hemoglobin tetramer. In the type of thalassemia due to a defect in beta-chain synthesis (beta-thalassemia), this imbalance results in a relative exoess of alpha-chains. We have studied the susceptibility of excess free alpha-chains to proteolysis. Incubation of isotopically labeled peripheral blood lysates from individuals with beta-thalassemia trait in the presence of bone marrow or normoblast lysates from thalassemic or hematologically normal individuals resulted in a decrease in the alpha/beta ratio and a loss of free alpha-chain radioactivity. Neither contamination with leukocytes nor higher ATP contents in young erythrocytes appeared to be responsible for this activity in normoblasts and bone marrow. We propose that erythroid precursor cells possess proteolytic activity that is markedly diminished in mature cells. This activity serves an important control function in the regulation of hemoglobin synthesis. It accounts at least in part for the more balanced synthesis of alpha- and beta-chains observed in bone marrow than in peripheral blood in heterozygous beta-thalassemia. It also plays a fine-tuning role in maintaining balanced synthesis in non-thalassemic erythrocytes.


Growth hormone modulation of murine erythroleukemia cell growth in vitro.

There are few studies showing a biological effect of growth hormone (somatotropin) on cell proliferation in vitro at physiological concentrations. We report here that Friend virus-infected erythroleukemia cells are responsive to growth hormone in vitro. Using a serum-free clonogenic assay we found as little as 0.1 ng of human growth hormone per ml caused a prominent stimulation of cell proliferation. Peak activity of human growth hormone occurred at 200 ng/ml, resulting in a 2-fold increase in cloning. Human chorionic somatomammotropin and the Cys(Cam)53-hGH(1-134) fragment of human growth hormone were also active, but a biologically inert oxidized human growth hormone had no growth-promoting effect in vitro. Cell proliferation was stimulated by insulin with peak potentiation occurring at 1 ng/ml, and prolactin had a demonstrable stimulatory effect between 50 and 100 ng/ml. These observations indicate that growth hormone and related polypeptides have a direct effect on the in vitro proliferation of erythroleukemia cells in the absence of serum. The results confirm a direct action of growth hormone on mammalian cells and suggest that pituitary hormones may affect leukemic cell growth.


Secretion of lipoprotein-X by perfused livers of rats with cholestasis.

The major abnormal plasma lipoprotein of cholestasis (LP-X) was isolated from blood plasma and from perfusates of isolated livers of rats with biliary obstruction. In both cases LP-X was composed mainly of about equimolar parts of phospholipids and unesterified cholesterol; the small protein component was primarily the arginine-rich apolipoprotein. By electron microscopy, LP-X appeared as a unilamellar liposome (690 A mean diameter, range 400-1000 A) with the trilaminar staining image typical of phospholipid bilayers. Extensive block staining of cholestatic livers for 48 hr with warmed uranyl acetate (37 degrees) permitted the visualization of vesicles indistinguishable from LP-X within hepatic parenchyma. These trilaminar-staining vesicles occurred predominantly within bile canaliculi. They also were seen in nearby cytoplasmic vacuoles or invaginations between hepatocytes and in the space of Disse. Similar vesicles were not seen in the endoplasmic reticulum or Golgi cisternae. These observations raise the possibility that the vesicles are formed within bile canaliculi and are transported from the canaliculi to the space of Disse within pinocytotic vacuoles.


Insulin resistance due to a defect distal to the insulin receptor: demonstration in a patient with leprechaunism.

We have studied a 2-year-old girl with acanthosis nigricans, glucose intolerance, marked hyperinsulinemia, and somatic features characteristic of the leprechaunism syndrome. Circulating plasma insulin levels were increased up to 50-fold and the patient showed a blunted hypoglycemic response to an injection of exogenous insulin (0.2 units/kg), indicating the presence of severe insulin resistance. Insulin purified from the patient's plasma was normal on the basis of chromatographic, electrophoretic, and immunologic criteria. Furthermore, the purified insulin competed effectively with (125)I-labeled insulin for binding to insulin receptors on cultured IM-9 lymphocytes and rat fat cells and also exhibited normal biological potency when tested on rat fat cells. Anti-insulin receptor and anti-insulin antibodies were not detected in the patient's plasma, and plasma levels of glucagon, growth hormone, and cortisol were normal. Insulin binding to the patient's circulating monuclear leukocytes was only slightly depressed into the low normal range and could not account for the severe insulin resistance. Studies on the patient's fibroblasts revealed normal levels of insulin receptors but a total absence of insulin's ability to accelerate glucose transport. Because rates of glucose transport and metabolism were normal in the basal state in the absence of insulin, we conclude that this patient's insulin resistance is due to an inherited cellular defect in the coupling mechanism between occupied insulin receptors and the plasma membrane glucose transport system.


Separation of younger red cells with improved survival in vivo: an approach to chronic transfusion therapy.

Transfusion of donor blood containing predominantly younger red cells with prolonged survival in vivo could significantly reduce the iron overload of patients requiring chronic transfusion. Age-dependent separation of red cells can be obtained by buoyant density centrifugation on isotonic solutions of arabino-galactane. By this technique, rabbit red cells were separated on a single layer of arabino-galactane and the appropriate fraction, after being labeled with (51)Cr, was reinfused into the donor. The survival in vivo was calculated by a mathematical model which corrects for both (51)Cr elution and random loss. There was a significant difference in survival in vivo between the light young red cells and the heavy old red cells. The potential survival in vivo of the 50% lightest red cells was 56 days, compared to 28 days for the heaviest red cells. Arabino-galactane appeared to be devoid of acute toxicity and of strong antigenicity and it did not appear to adhere to the red cell stroma. These data extrapolated to humans indicate that it may be feasible and advantageous to use red cells fractionated by this technique for transfusion. The 50% lightest human red cells can be expected to have a mean survival of 88 days, compared with 60 days for unfractionated blood. Transfusion of young red cells could significantly reduce the iron overload for patients requiring chronic transfusion, by avoiding infusion of the oldest red cells, which contribute equally to iron overload yet offer only transient survival in vivo.


"Conjugal" transfer of plasmid DNA among oral streptococci.

The beta plasmid from Streptococcus faecalis strain DS5, which codes for resistance to erythromycin and lincomycin, was introduced into a Lancefield group F streptococcus, strain DR1501, by transformation. This strain, DR1501 (beta), was found to be an excellent donor of the beta plasmid and readily transferred the resistance markers to various lactic acid bacteria, including certain strains of S. mutans, S. sanguis, and S. salivarius. Evidence is presented indicating that the transfer of the beta plasmid is mediated by a mechanism that requires cell-to-cell contact. The transfer of plasmid DNA during conjugation has been confirmed by the isolation of beta plasmid from transconjugant clones and by their ability to then serve as donors of the erythromycin and lincomycin resistance markers.


Regulation of acetylcholine receptor levels by a cholinergic agonist in mouse muscle cell cultures.

The effects of continuous exposure to carbamylcholine (CbCho) on regulation and stabilization of acetylcholine receptors (AcChoR) were studied in cell cultures of G8, a continuous mouse muscle cell line. Exposure of cultures to 10-100 muM CbCho for 24-48 hr produced a 30-50% reduction in (125)I-labeled alpha-bungarotoxin binding. CbCho was not found to alter cell morphology, protein metabolism, or amino acid incorporation. Electrophysiological experiments demonstrated a 75% reduction in the maximum sensitivity of the myotubes to iontophoretic application of acetylcholine (AcCho). The reduction in AcCho sensitivity appeared to represent a true loss of functional receptors because there were no changes in the passive electrical properties of the cells or in the AcCho reversal potential and because receptor desensitization appeared not to be involved. Tetrodotoxin had no effect on receptor levels, either alone or in combination with CbCho. Receptor degradation in control cells could be described kinetically as a first-order process with a half-time of 19.2 hr; turnover rate in receptors remaining after prolonged exposure to CbCho was indistinguishable from that in control cells. We conclude that a receptor-active ligand can exert negative control over AcChoR levels and that prolonged exposure to an AcCho analog is not sufficient to induce a stable population of receptors in these cells.


Structure and physiological activity of the motoneurons of the nematode Ascaris.

The nervous system of the nematode worm Ascaris contains about 250 nerve cells; of these, the motoneurons consist of five segmental sets, each containing 11 cells. Morphologically, the motoneurons can be divided into seven different types. Their geometry is simple: some are unbranched, others have one branch point, and the most complex have two. There is no neuropil in the nerve cords; synapses are made by axo-axonal contact or onto short spines. These features enable us to study the anatomy and physiology of the system with a degree of completeness that would be difficult in other systems. The physiological activity of five of the motoneurons has been investigated, three being excitatory and two inhibitory. The excitatory motoneurons receive input from intersegmental interneurons. The inhibitory motoneurons do not receive input from the interneurons; instead they receive their input from the excitatory motoneurons in a circuit that can mediate reciprocal inhibition between the dorsal and the ventral musculature.


In vitro release of [5-methionine]enkephalin and [5-leucine]-enkephalin from the rat globus pallidus.

Endogenous [5-methionine]enkephalin (Metenkephalin) and [5-leucine]enkephalin (Leu-enkephalin) are released from perfused slices of rat globus pallidus by increased K(+) in a Ca(2+)-dependent manner. Tissue perfused for 40 min contained only 26% of the Met-enkephalin and 44% of the Leu-enkephalin found in the freshly dissected tissue. After perfusion, the mean (+/-SEM) ratio (wt/wt) of Met-enkephalin to Leu-enkephalin was 3.4 +/- 0.2 compared with 5.8 +/- 0.2 in the fresh tissue. The degradation of trace amounts of synthetic [(3)H]enkephalins in the perfusing medium during stimulated release seems to reflect the accelerated degradation of enkephalin released from the tissue: 63% of the Met-enkephalin and 23% of the Leu-enkephalin were degraded in a medium containing bacitracin (30 mug/ml). The mean ratio (wt/wt) of the Met-enkephalin to the Leu-enkephalin recovered after release by exposure of slices to 50 mM K(+) was 2.7 +/- 0.3. When perfusates were corrected for degradation, this ratio increased to about 5.5 which is higher than that found in the perfused tissue. The differences in release, tissue loss, and catabolism of the two enkephalins may be reflecting differences in the metabolic systems operating on the pentapeptides, but this interpretation will have to be validated by in vivo release experiments. In any event these observations strongly suggest that both enkephalins can be considered candidate neurotransmitters in the rat globus pallidus.


Responses to light of solitary rod photoreceptors isolated from tiger salamander retina.

Single, isolated rod photoreceptors were obtained by enzymatic dissociation of the tiger salamander (Ambystoma tigrinum) retina. These solitary cells retained the morphological features of rods of the intact retina and could be maintained in culture for several days. When impaled with micropipettes for electrophysiological recording, dark-adapted solitary rods had during darkness a resting potential of approximately -45 mV and a steady-state slope resistance of 500 Momega at rest. The current-voltage relationship showed both inward- and outward-going rectification. The responses to light of solitary rods were similar to those recorded from rods in the intact retina stimulated with large-diameter spots of light. The reversal potential of the light response of solitary rods was near 0 mV when measured in either the inner or outer segment.


Evidence for existence of a yearly clock in surgically and self-blinded chipmunks.

By use of simplified technique and constant environmental conditions, I have demonstrated the existence in the chipmunk of a yearly clock. In a blinded chipmunk the clock manifested itself by remarkably consistent changes in running activity, food and water intake, and body weight over 6 1/2 yr. Studies on freshly trapped chipmunks kept in the same laboratory environment but with alternating light and darkness (12 hr each), showed that, when their eyes were covered for much of the light period, they reduced their exposure to light to preserve the activity of the yearly clock. Laboratory-adapted chipmunks that do not shield their eyes from light do not show the clock. The yearly clock has all the characteristics of the 24-hr clock, including sharply defined active and inactive phases, and must likewise play an important part in the animal's survival. Light would appear to be the chief or only cue for the clock. The period lengths did not change with age during the 6 1/2 yr.


[Indirect lymphography of sub-mental and sub-maxillary lymph nodes (author's transl)].

This report presents our observations in 81 cases; and we have used indirect lymphography only, we injected the contrast medium, lipiodole ultra fluide, intramuscularly to both sides of the tongue. The injection sites are first anaesthetized. The amount of contrast medium should not exceed 5 cc. It is injected in about 10 mn. The first radiological examination is performed 4, 8 and 12 hours after the injection. X-rays are taken using the occipitomental and also lower intraoral occlusal views. The following radiological studies are made after 24 and 48 hours. If necessary further x-rays can be taken as well.


[Variations in salivary immunoglobulins in Sjögren's syndrome (author's transl)].

A study of salivary immunoglobulins (IgA, IgG, IgM) was made in 74 subjects with or without Sjögren's syndrome. In the normal subjects, only IgA could be detected by classical immunodiffusion techniques. A correlation was sought between the presence of IgG and/or IgM in the saliva, and various clinical or objective clinical examinations, as well as with accessory salivary gland biopsy. This study indicates that in Sjögren's syndrome the detection of IgG and/or IgM is strongly correlated with sialography which is the surest examination used clinically. Thus the detection and estimation of salivary immunoglobulins is thus a simple and specific complementary examination in Sjögren's syndrome.


[Stomatitis and antimitotics (author's transl)].

The authors report a case of stomatitis, in a patient receiving antimitotic therapy, which was found to be due to an acute leukosis. They feel that patients on antimitotics who develop stomatitis should have a white cell count to exclude leukosis, which is not a very rare finding today.


[The greatest horizontal contour-line of the face. Meeting of anthropology and maxillofacial surgery (author's transl)].

The greatest horizontal contour-line of the face is convex, passes through the malar bone and the external side of the zygomatic arch, and can be defined as the geometric connection of the facial points most distant from the median plane. The study of this facial line of greatest contour is particularly important in the case of fractures of the malar and zygomatic arch. The roentgenography of this line is done with an x-ray plate held parallel to the Reid line (orbital meatus) and the principal ascending radius passing through the middle of the line which connects the two gonions. This technique leads to extreme accuracy in the reduction of zygomato-malar fractures. The method can be used in the operating room.


[Reflections concerning reconstruction of the floor of the orbit using a metal grill (author's transl)].

The author presents four cases of patients with collapse of the floor of the orbit, total in three cases and partial in one. The material used was cut from the perforated plates used classically in maxillo-facial surgery. In one cases, the approach was mixed--sinusal and sub-orbital--whilst in the other three a classical sub-orbital-palpebral approach was used. There was one complication of infection 15 days after insertion in the case in which a mixed approach was used, this having no sequelae--one and a half year having elapsed. In all cases, the Lancaster test returned to normal, proving the perfect mobility of the eyeball over the plate. Despite the small number of cases, and a follow-up of no more than one and a half years, the author feels that the ease of use of these plates and the good results obtained here offer a possible future for this method.


[Inflammatory granuloma of the face with a malignant course. Atypical Hodgkin's disease (author's transl)].

A case of Hodgkin's disease beginning in the skin of the face with local lymph node involvement is reported. This was a lymphogranulomatous form without characteristic Reed-Sternberg cells in skin biopsy specimens, rendering the diagnosis difficult. There was a combination of granulomatous inflammation, plaques of necrosis and lesions of fibrinoid necrosis in the vessel walls. This form entered the classification of clinical Stage IV, with its associated serious prognosis.


On the pharmacokinetics of cephalosporin antibiotics.

A review is given on the pharmacokinetic characteristics of some cephalosporin antibiotics. The use of the pharmacokinetic parameters serum concentration, serum protein binding, serum half life, and apparent volume of distribution as a basis for the selection of the clinically most effective cephalosporin derivative is discussed. It is concluded that these parameters must be used in conjunction with data on tissue distribution to specialized sites, and with information on antibacterial activity and toxicity.


Experimental models for studies on transportation of antibiotics to extravasal compartments.

A short review is presented illustrating different methods employed to determine the antibiotic concentration in different tissues in man and the limitation to perform thorough pharmacokinetic studies. The most often used experimental models (different tissue cage models the fibrin clot, skin blisters, skin windows, skin chambers) applied in animal and man for studies of antibiotics are presented as well as a discussion concerning their relevance to the clinical situation.


Studies on the pharmacokinetics of cefoxitin, cefuroxime, cephaloridine and cephalothin using subcutaneous tissue cages.

The present study was undertaken to compare the ability of three cephalosporins--cephalothin, cephaloridine and cefuroxime--and a cephamycin--cefoxitin--to penetrate into extravasal compartments. The study was performed using female adult rabbits with subcutaneously implanted steel net cages. All antibiotics were given in a dose of 50 mg/kg b.w. parenterally as i.v. injections. The concentrations of the different antibiotics were determined simultaneously in serum and in fluid obtained from the cages. The concentrations were followed after single doses as well as after repeated doses using an agar well technique for the assays. To control the experimental conditions electrolyte and protein contents of the fluid in the different cages were followed and were found to be stable during the experimental period. All antibiotics were rapidly eliminated from serum. The concentrations in the cages reached its maximum 1.5-3 hours after administration. The highest concentrations in the extravasal compartments were obtained with cephaloridine and the lowest with cephalothin with the values of the other antibiotics ranging in between.


Implanted tissue-cages: a critical evaluation of their relevance in measuring tissue concentrations of antibiotics.

In several species of laboratory animals, the concurrent serum and tissue-cage fluid levels of several antibiotics after a single parenteral dose have markedly different profiles. New work has shown that these differences are mainly due to the relatively impermeable granulomatous sheath which develops around the implanted tissue chambers. The concentrations of an antibiotic in tissue-cage fluid are therefore not representative of its concentration in "interstitial fluid" or extravascular fluid in general. Tissue-cage levels are probably more representative of antibiotic levels to be found inside abscesses, in chronic disease situations, or relatively avascular sites in the body.


The correlation of in-vitro susceptibility tests with in-vivo results of antibiotic treatment.

The reason why the results of the clinical response to therapy do not always correspond to antibiotic laboratory testing include errors in performance or interpretation of test results, imprecise definition of resistance and host factors which influence the outcome of treatment regardless of the antibiotic being used. These factors are reviewed. Cephalosporins and penicillins pose problems because the prime effects of enzymes in influencing the outcome of treatment are not always clear in standard tests for susceptibility. Inter-laboratory differences in interpretation of tests makes comparisons of tests between centres difficult and the use of reference strains for validation of testing is proposed.