The carboxylic acid groups of bovine luteinizing hormone. The effects of their modification on receptor site binding and subunit-subunit interaction. The modification of the carboxyl groups of the subunits of bovine luteinizing hormone to neutral derivatives by carbodiimide-mediated coupling with glycine methyl ester has been studied. The modified alpha subunit, which has 8 residues of glycine methyl ester incorporated, will no longer recombine with native beta (hormone-specific) subunit, but the modified beta subunit, with 6 to 7 glycine methyl esters incorporated, will recombine with native alpha to yield a partially active hormone. Derivatization of the intact hormone results in dissociation to subunits together with formation of a major side product which is covalently cross-linked. Significant cross-linked product was not obtained during modification of individual subunits, thus indicating an orientation between an activated carboxyl group(s) and a nucleophile(s) in the intact hormone which favors coupling. Separation of subunits from the derivatized, noncross-linked fraction by countercurrent distribution reveals a heterogeneous preparation of the modified alpha subunit which also will not recombine with either a native or modified beta subunit. The beta subunit from the modified intact hormone was indistinguishable from the modified isolated beta subunit in amino acid composition and in ability to recombine with native alpha subunit. The results are consonant with data from this and other laboratories in which various modifications of the alpha chain, the subunit common to the glycoproteins, more seriously affect recombination than similar modifications of the beta subunits. The number of carboxyl groups modified in each subunit is compatible with but not in total agreement with assignments of amides reported from sequence studies.


Site of synthesis of membrane and nonmembrane proteins of vesicular stomatitis virus. Upon infection of Chinese hamster ovary cells (CHO), vesicular stomatitis (VSV) virus synthesizes two membrane proteins (the VSV glycoprotein and the VSV matrix or membrane (M) protein) and three nonmembrane proteins (the VSV nucleocapsid, the viral transcriptase, and an NS protein). We have used the VSV-infected cell as a model system for the study of the site of synthesis of these membrane and nonmembrane proteins. We have isolated VSV mRNA from free polyribosomes, membrane-bound polyribosomes, and the postribosomal supernatant, and identified the individual species of VSV mRNA present in each fraction. The mRNA which encodes the VSV glycoprotein is found exclusively on membrane-bound polyribosomes, while the mRNAs which encode the VSV, M, N, and NS proteins are found in free polyribosomes, in the membrane fraction of the cell, and in the postribosomal supernatant. Our results suggest that the VSV glycoprotein is synthesized exclusively on membrane polyribosomes, while at least some of the M, N, and NS proteins are made on free polyribosomes.


Multiple thermotropic phase transitions in Escherichia coli membranes and membrane lipids. A comparison of results obtained by nitroxyl stearate paramagnetic resonance, pyrene excimer fluorescence, and enzyme activity measurements. At characteristic temperatures, membranes from Escherichia coli cells enriched in exogenous elaidic acid exhibit two abrupt changes in the slope of Arrhenius plots of two enzyme activities. For NADH oxidase, these changes occur at 27 degrees and 32 degrees, whereas for D-lactate oxidase, these changes occur at 31 degrees and 36 degrees. Pyrene excimer fluorescence and spin-labeled fatty acid paramagnetic resonance results indicate that the beginning, midpoint, and end of a single structural change(order leads to disorder transition) occurs at 25.5-29.0 degrees, 30.0-31.0 degrees, and 33.0-35.5 degrees, respectively. These data suggest that for NADH oxidase, the observed activity changes correspond to the beginning and midpoint of a single membrane lipid structural change, whereas for D-lactate, the activity changes correspond to the midpoint and end of that structural change. In addition to the membrane structural change spanning the range of 25.5-35.5 degrees, a second change (9.5-21.0 degrees) was also observed. This transition was detected by 5- and 16-2,2-dimethyloxazolidinyl-1-oxyl (doxyl) stearates, but not by 12-doxyl stearate or pyrene. Structural changes in the extracted lipids were observed in the temperature ranges 4.0-9.0 degrees, 14.0-20.0 degrees, and 25.0-35.5 degrees. The two higher ranges correlate well with the ranges for structural changes observed in the intact membrane. Observations of these multiple transitions in both intact membranes and extracted lipids strongly suggest that these lipids segregate into domains of different fluidity and composition.


Coordinate regulation of adenylate cyclase and carbohydrate permeases by the phosphoenolpyruvate:sugar phosphotransferase system in Salmonella typhimurium. Adenylate cyclase (EC 4.6.1.1) and several carbohydrate permeases are inhibited by D-glucose and other substrates of the phosphoenolpyruvate:sugar phosphotransferase system. These activities are coordinately altered by sugar substrates of the phosphotransferase system in a variety of bacterial strains which contain differing cellular levels of the protein components of the phosphotransferase system: Enzyme I, a small heat-stable protein, and Enzyme II. It is suggested that the activities of adenylate cyclase and the permease proteins are subject to allosteric regulation and that the allosteric effector is a regulatory protein which can be phosphorylated by the phosphotransferase system.


Arthritic disorders of the adult radiocarpal joint: anatomic considerations and an evaluation of fifty consecutive abnormal cases. The anatomy and pathology of the radiocarpal compartment of the adult wrist are described in a study of human cadavers and 50 consecutive patients with radiocarpal joint abnormalities. The most frequently encountered diseases were adult onset rheumatoid arthritis (42) and calcium pyrophosphate deposition disease (22%). Features allowing radiographic diagnosis included the degree of symmetry and the presence of demineralization, sclerosis, joint space narrowing, subchondral cysts and erosions. Evaluation of abnormalities in other compartments of the wrist and the ulnar styloid is mandatory.


[A functional flagella with a 6 + 0 pattern]. The male gamete of the Gregarine Lecudina tuzetae has been studied with transmission electron microscopy and microcinematography.  It is characterized by a flagellar axoneme of 6 + 0 pattern, a reduction of the chondriome, and the abundance of storage polysaccharide or lipid bodies.  The movements of the flagella are of the undulating type and they are performed in the three dimensions of space.  They are very slow, with a cycle time of about 2s.  The structure of the axoneme components are similar to those of flagella with a 9 + 2 pattern.  Each doublet has overall dimensions of 350 x 220 A; the space between the adjacent doublets is about 160 A.  The A subfiber bears arms like dynein arms.  The diameter of the axoneme is about 1,000 A.  The basal body consists of a cylinder of dense material 2,500 A long and 1,300-1,400 A in diameter; a microtubule 200 A in diameter is present in the axis.  This study shows that a 6 + 0 pattern can generate a flagellar movement.  The mechanism of the flagellar movement of the male gamete of L. tuzetae does not require the presence of central microtubules and it would include molecular interactions of the dynein-tubulin type between the adjacent peripheric doublets.  The slowness of the movements is discussed in terms of the axoneme's structure and its energy supply.  Finally, the phylogenetic significance of this flagella is examined on the basis of the morphopoietic potentialities of the centriolar structures.


The permeability of muscle capillaries to horseradish peroxidase. The main objective of this study was to determine the pathways by which horseradish peroxidase (HRP) can cross the endothelium of muscle capillaries.  Specimens of mouse diaphragm were fixed for cytochemical analysis at various intervals after intervenous injection of 0.5 mg HRP, at 4 min after intervenous injection of varied amounts of HRP, and at 4 min after intervenous injections in various volumes of isotonic NaCl.  Our findings indicate that endothelial junctions serve as a barrier which may allow passage of very limited amounts of HRP.  They also suggest that endothelial vesicles transfer HRP from the capillary lumen to the pericapillary interstitium as well as in the reverse direction.  Increasing the volume of solution injected to approximately 30% of total blood volume did not increase the amount of HRP that left the capillary lumen.  Our results with HRP do not provide clearcut evidence that endothelial junctions are the site of the small pore.


Characterization of an SV40-transformed 3T3 cell line expressing an unusual phenotype. A transformed variant derived as a clone from normal 3T3 cells infected with simian virus 40 (SV40) has been found to possess a phenotype intermediate between that of normal cells and that characteristic of the transformed state, yet cells of the variant still test positively for the SV40-specific nuclear T-antigen. The variant exercises growth control, although not as stringently as do normal cells. Its cell size more closely resembles that of normal cells than of transformed cells. The variant also exhibits levels of spontaneous agglutination that are in line with those characteristic of the normal cells from which it was derived, and far higher than corresponding values for cells exhibiting the fully transformed phenotype. Plasma membranes of variant cells more closely resemble those of transformed cells than of normal cells as estimated by polyacrylamide gel electrophoresis. Perhaps the most distinguishing characteristic of the transformed variant is its complete immunity to agglutination by concanavalin A (Con A), even at concentrations of the lectin as high as 500 mug/ml. Moreover, trypsinization does not render variant cells as agglutinable in the presence of Con A as are untreated fully transformed cells. By contrast the variant displays a low tolerance of Con A toxicity, as monitored by ability to grow after treatment with the lectin, and on this count resembles transformed cells. Moreover a survey of several normal cell lines has revealed that even they do not consistently show resistance to Con A toxicity. These observations indicate that Con A-mediated agglutination and inability to grow after treatment with Con A are quite independent and do not bear a cause and effect relationship.


The growth of the pollen tube wall in Oenothera organensis. The growth of the pollen tube wall of Oenothera is effected by the expulsion of fibrillar material from the cytoplasm into the developing wall. This material may also be seen in the cytoplasm, contained in membrane-bound vesicles. It is not clear how the content of the vesicles is discharged, but it appears not to involve the participation of microtubules. The source of the cytoplasmic fibrillar bodies depends upon the stage of development of the pollen tube. The earilest growth is derived from the inclusion into the wall of vesicles containing pre-formed materials present in the grain on pollination. During the next stage of growth the wall is derived from the content of double-membraned inclusions also present in the pollen. The content of the former vesicles is not so similar to the wall as the latter, but intermediates between the 2 types of vesicle may be seen in the cytoplasm, indicating that the former are formed from the latter. Most of the tube wall is derived from the products of dictyosomes in the pollen grain or tube. These dicytosomes are few in number and they must be exceedingly active. This, and the observation that dictyosome vesicles are frequently associated with banked complexes of mitochondria, indicates that some steps in the metabolism of the vesicular content, perhaps phosphorylation, take place distant from the dicytosomes. These different sources of fibrillar material presumably permit the rapid starting of tube growth, without any attendant metabolism. However, it would be impossible to include enough pre-formed wall material in the grain to enable the full growth of the tube, so once started, it seems that the tube then relies on the elaboration of simple reserves for the contruction of its wall. These reserves are likely to be held in the pollen, and may be the large numbers of starch grains characteristic of the pollen cytoplasm.


High speed liquid-solid chromatography of phenothiazines using columns packed with spherosil porous silica beads. Spherosil, a spherical porous silica, has been investigated for use in high speed liquid-solid chromatography and compared to Lichrosorb Si-60, using three phenothiazines as test solutes. Particle size distribution for four different size ranges with nominal mean diameters of 5, 10, 20 and 40 mu m is given. Distribution is very homogeneous. Columns from 15 to 100 cm in length and 1/4 or 1/8 in.o.d. have been prepared and efficiency measured by determination of HETP with the three phenothiazines: [methylamino-3-propyl]-10-chloro-3-phenothiazine [k' = 3]; [N-methyl-N-[dimethylamino-3-propyl]amino-3-propyl]-10-chloro-3-phenothiazine [k' = 6.5]; and [dimethylamino-3-propyl]-10-chloro-3-phenothiazine-N-oxide [k' = 15]. Influence of column length and incidence of bead diameter have been studied. 1/4 in.o.d. columns are easier to fill than 1/8 in.o.d. columns for Spherosil 5 mu m and have, therefore, a greater efficiency. HETP, H, varies according to flow rate as H = DVn with 0.4 less than n less than 0.6 and according to particle size ad H = A d beta p with 1.7 less than beta less than 1.8. The best figures for H are between 0.1 or 0.2 mm for a flow rate of 1800 ml hr-1  cm-2 [k' = 15]. The separation of a mixture of 6 phenothiazines with the mobile phase, anhydrous ethyl acetate 60 V, water saturated ethyl acetate 20 V, anhydrous methanol 20 V, 33% aqueous solution of ethylamine 0.25 V, is given. Its duration is 20 min. instead of 90 min. by thin-layer chromatography.


Urinary and plasma cyclic adenosine 3',5'-monophosphate in patients with idiopathic edema. Urinary excretion and plasma concentrations of adenosine 3',5'-monophosphate were determined in idiopathic edema patients both at rest and after assumption of the upright position. Patients and normal subjects responded similarly to upright posture with decreases in urinary volume and creatine excretion and their urinary excretion of cyclic AMP was not significantly different. The plasma concentration of cyclic AMP was high in patients, both in the recumbent and upright positions, and its renal clearance (unlike that of creatinine) was low. Plasma cyclic AMP increased in response to upright posture both in patients and normal subjects. Even if the significance of elevated plasma cyclic AMP is unknown, its presence without an increase in urinary cyclic AMP suggests that the renal handling of cyclic AMP is abnormal is idiopathic edema; this may be related to an excessive beta-adrenergic tone.


A radioreceptor assay for evaluation of the plasma glucocorticoid activity of natural and synthetic steroids in man. An assay for plasma glucocorticoid activity has been developed using specific glucocorticoid receptors. Unlike other assays for cortisol and certain synthetic corticosteroids, this radioreceptor assay measures the glucocorticoid activity of all natural an synthetic steroids. Steroids extracted from as little as 0.05 ml of plasma are incubated with 3H-dexamethasone and cytosol receptors from cultured rat hepatome cells. From 0.5 to 50 ng of cortisol are accurately detected. Glucocorticoid activities of adult determined by the assay correlate closely with corticoid levels obtained in the CBG-isotope and fluorometric assays. Other steroids are measured in proportion to both concentration and potency as glucocorticoids. Relative activities include: cortisol 100, dexamethasone 940, prednisolone 230, prednisone 3, estradiol 1 and androstenedione 1. A similar ranking of steroids was found using receptors from a human source (fetal lung). The assay has been useful in detecting glucocorticoid activity in unidentified medications and in measuring plasma glucocorticoid levels after administration of synthetic corticosteroids. A modification of the assay is described for also estimating the level of free (unbound) glucocorticoid activity in plasma. Assays are performed before and after removal by charcoal of steroids not bound to plasma transcortin. This consideration is shown to be particularly important with prednisolone where the unbound steroid level is much less than the total. Thus, the radioreceptor assay provides a relatively simple technique for measuring the total of free plasma of glucocorticoid activity in man due to any natural or synthetic steroid or combination of steroids.


Skeletal resistance to endogenous parathyroid hormone in pateints with early renal failure. A possible cause for secondary hyperparathyroidism. Studies were carried out to evaluate whether skeletal resistance to acute increments in endogenous parathyroid hormone exists in patients with mild to moderate renal insufficiency. Hypocalcemia was induced with the infusion of ethyl-enediamine-tetra-acetate (EDTA) in 10 normal subjects and 13 patients with mild renal failure. After the induction of hypocalcemia, the concentration of serum calcium increased gradually and reached preinfusion levels by 22 h in the normal subjects; in contrast, the levels of serum calcium in patients with mild renal insufficiency were significantly lower than the preinfusion values even at the end of 26 h following the EDTA infusion. This delayed recovery occurred despite significantly higher levels of serum immunoreactive parathyroid hormone (IPTH) in the patients than in the normal subjects. The increase in the levels of IPTH reflect elevations in the concentrations of biologically active hormone since urinary cyclic AMP increased significantly. Urinary calcium excretion following the EDTA infusion was not different in both groups and, therefore, could not account for the delayed recovery of serum calcium values in the patients with renal insufficiency. These results indicate that secondary hyperparathyroidism exists early in patients with renal failure and such patients have exaggerated parathyroid hormone secretion in response to acute hypocalcemia. The data are consistent with the concept that skeletal resistance to increments in endogenous parathyroid hormone is present in such patients. This abnormality is, at least partly, responsible for the hypocalcemia and secondary hyperparathyroidism of renal insufficiency.


Effects of progestins on estradiol receptor levels in human endometrium. Medroxyprogesterone acetate (Provera) was administered orally during 2-4 days to women undergoing endometrial curettage during the follicular phase of the menstrual cycle. Estradiol (E2) receptor levels were estimated from the amounts of 2H-E2 tightly bound in nuclei after incubation of endometrial tissue with an excess of the labeled hormone. An average value (9 subjects) of 1.5 pmole E2/mg DNA +/- 0.7 (mean +1- SD) was found. This value is significantly lower than the average E2 receptor levels in proliferative endometrium of untreated subjects (3.2 +/- 1.3, n = 33) and equals the level observed in the early secretory phase (1.5 +/- 0.5, n = 5). These results indicate that one of the progestin effects on human endometrium is the reduction of E2 receptor levels.


Evidence for partial 21-hydroxylase deficiency among heterozygote carriers of congenital adrenal hyperplasia. Concentrations of 17-hydroxyprogesterone are significantly greater in heterozygous carriers of CVAH than in controls 30 and 60 minutes after an infusion of 25 units of synthetic ACTH 1-24 and 2 hours after beginning a 4-hour infusion of 50 units ACTH. The majority of carriers were clearly above the control range at these collection times. Hence, heterozygous carriers have a partial enzyme deficiency although all cannot be diagnosed based on 17-hydroxyprogesterone levels after ACTH stimulation.


Pattern of plasma ACTH, hGH, and cortisol during menstrual cycle. The pattern of secretion of plasma ACTH, hGH, TSH, LH, FSH and cortisol was studied in 12 menstrual cycles, representing 5 normal volunteers. Results were plotted by taking the LH-FSH midcycle peak as day 0. The typical menstrual cyclic LH and FSH pattern was observed in each case. ACTH, cortisol and hGH varied significantly throughout the menstrual cycle. ACTH was characterized by a decrease during the follicular phase, a nadir at day -2, followed by a significant increase to a peak at day 0, then a subsequent decrease and constant levels during the luteal phase until day +8. Cortisol was lowest in the follicular phase until day -4, and highest from day -2 to day 0. During the luteal phase, cortisol remained constant but was significantly higher than in the follicular phase (days -7 to -4). hGH showed a significant increase during the periovulatory period (days -3 to +3). No significant changes of plasma TSH were observed. These results suggest that pituitary hormones other than gonadotropins may be involved in the ovulatory mechanism, and reveal a degree of stimulation of the pituitary-adrenal axis without establishing whether the effect is direct or indirect.


Radioreceptor assay of insulin: Comparison of plasma and pancreatic insulins and proinsulins. Porcine proinsulin, related intermediates and plasma immunoreactive insulin components have been studied by radioreceptor assay. Using the purified rat liver membrane or cultured human lymphocyte radioreceptor assay, porcine proinsulin is %5, split proinsulin 6% (54-55 split in connecting peptide) desdipeptide proinsulin 20% (deletion of amino acids 62 and 63 of connecting peptide) and desnonapeptide proinsulin 27% (deletion of amino acids 55-63 of connecting peptide) as active as porcine insulin in both assay systems; these values closely parallel the in vitro bioactivity of these preparations. In the lymphocyte radioreceptor assay the human plasma immunoreactive insulin-like component has the same potency as porcine insulin per immunoreactive unit, whereas the plasma immunoreactive proinsulin-like component is only 15% as active. Since both plasma immunoreactive components are somewhat less reactive than would be expected from puriified human insulin and proinsulin, the data suggest that both plasma components contain immunoreactive molecules that do not react in the radioreceptor assay.


Potentiation of adrenocortical response upon intermittent stimulation with corticotropin in normal subjects. Modifications of adrenocortical steroidogenic response to ACTH as a consequence of acute prior exposure to this hormone, were studied in 106 normal subjects divided in 15 experimental groups. Adrenocortical response was assessed by the changes in plasma cortisol level and in urinary excretion of cortisol, 17-ketogenic and 17-ketosteroids; in some experiments plasma 11-deoxycortisol, corticosterone, progesterone and 17-hydroxyprogesterone were determined as well, together with urinary excretion of the unconjugated form of 11-deoxycortisol and corticosterone. Slow (8-h) intravenous administration of ACTH in amounts producing maximal response, leaves the adrenal cortex in a hyperresponsive state in case of further stimulation for up to 3 days, while the adrenocortical secretion comes back to baseline in the meantime. This potentiation phenomenon seems to concern essentially cortisol secretion since, among the compounds measured only cortisol and 11-deoxycortisol secretions increased progressively in amplitude when ACTH was administered repeatedly. Futhermore the degree of ACTH-induced adrenocortical hyperresponsiveness was found to depend on the amount of ACTH injected and on the time during which the adrenal cells are exposed to high peptide hormone concentrations. Increased adrenocortical responsiveness to ACTH persists when endogenous corticotropin secretion was suppressed for a few days by dexamethasone in normal subjects. Thus the potentiation phenomenon is not critically dependent on continued exposure of adrenal cells to endogenous corticotropin.


Radioreceptor-inactive growth hormone associated with stimulated secretion in normal subjects. We have previously reported systematic discrepancies between radioreceptor (RRA) and radioimmunoassay (RIA) measurements of growth hormone (hGH) in acromegalic patients. Due to limitations in RRA sensitivity, such comparisons could not be made in normal subjects. RRA methodology has now been adapted to allow detection of hGH at normal circulating levels. Since variations in Na+, K+, Ca++, and Mg++, incubation at 37 C and 4 C, and delayed tracer addition failed to improve assay sensitivity, specimen size was increased to 300 mul and incubation volume to 1.5 ml, while holding the quantity of added receptor constant. Best assay sensitivity, in room temperature incubations in 25 mM Tris for 16 h at pH 7.6 and 10 mM Ca++, was 0.66 +/- 0.30 ng hGH per ml serum. Under these conditions, 200 mug hepatic receptor protein bount 15.8 +/- 0.83% of added 125I-hGH, and 8.72 +/- 0.85% of bound tracer was displaced by 0.25 ng added unlabeled hGH. Nonspecific depression of binding by serum did not impair assay sensitivity with most receptor preparations. The basal hGH measured by RIA (antiserum 68-416) in a group of normal short children was 1.97 ng/ml, similar to the RRA result, 1.89 ng/ml (P = NS). Comparative measurements were also made in selected samples of sufficient volume during the 1 1/2 h following administration of hGH secretagogues (insulin, arginine, L-dopa). In these samples, the RIA value was 9.34 +/- 0.68 and the RRA value 6.29 +/- 0.62 ng/ml (P less than 0.01); the RIA/RRA was 1.77 +/- 0.18. Thus, no significant measurement discrepancy was found in basal samples from normal subjects, in contrast to previous findings in acromegalics. The appearance of such a discrepancy within 90 min after stimulation of hGH might be due to RIA/RRA discordance in secreted molecular subspecies, or might arise from peripheral hGH metabolism.


Circulating NSILA-s in man: Preliminary studies of stimuli in vivo and of binding to plasma components. The insulin-like peptide NSILA-s (non-suppressible insulin-like activity soluble in acid ethanol) was measured in plasma using gel filtration in acid and a new radioreceptor assay. Total plasma NSILA-s levels, which are elevated in some patients with extrapancreatic tumors and hypoglycemia, were also found to be elevated in pregnancy. Patients with hypopituitarism had subnormal concentrations of total plasma NSILA-s which rose promptly and reached a peak 1 h after intravenous hGH. In contrast ot this acute stimulus, chronic elevation of hGH resulted in an elevated level of NSILA-s in only 1 of 10 acromegalics. In normal subjects, oral glucose loading resulted in a rise in NSILA-s (mean peak equals 151% of basal) while insulin induced hypoglycemia produced a small and variable fall. Among the other conditions studied, anorexia nervosa was associated with low levels of NSILA-s. Using gel filtration we found that at physiologic pH about 93% of endogenous NSILA-s is reversibly bound to larger molecules. We show that in normal adults the insulin-like bioactivity in plasma due to free NSILA-s in the basal state is equal to that exerted by insulin itself, and that the free NSILA-s in the circulation of patients with extra-pancreatic tumors approximates the elevated level of insulin in patients with insulin-secreting pancreatic tumors.


The effect of iodized oil on the TSH response to TRH in endemic goiter patients. The TSH and T3 response to synthetic TRH was evaluated in 4 groups of patients: normal controls and goitrous subjects from the urban area of Sao Paulo (urinary iodine excretion: 172.2 +/- 48.3 mug I/g creatinine) and nongoitrous and goitrous subjects from the endemic areas of Sao Bento (urinary iodine excretion: 53.8 +/- 17.1 mug I/g). Plasma T4 and T3 were within our normal range in all groups of patients. The mean plasma TSH was significantly higher (5.2 +/- 3.3 muU/ml) in goitrous subjects living in Sao Bento as compared to normal control groups both in urban or endemic areas, and after TRH these patients had an exaggerated and sustained TSH response with a significantly higher peak level (21.1 +/- 7.9 muU/ml). T3 concentration rose in all subjects following TRH and all patients from the Sao Bento endemic areas had a significantly higher proportionate increase in plasma T3 at 120 min. After an injection of iodized oil basal plasma TSH returned to the normal range in the goitrous subjects from Sao Bento. The mean peak TSH response to TRH was 9.1 +/- 3.8 muU/ml at 3 months after the iodized oil injection, and only at 6 months after the iodized oil TSH response was significantly reduced (peak level: 6.1 +/- 2.4 muU/ml). It is confirmed that plasma TSH levels are increased in endemic goitrous patients but not in normal controls living in the same endemic area and it is suggested that the pituitary threshold for inhibition of secretion of TSH by T4 and T3 has been reset in these goitrous subjects to achieve a persistently higher secretion rate of TSH.


Effect of the diphosphonate EHDP on bone mineral metabolism during prolonged bed rest. The effect of disodium ethane-1-hydroxy-1, 1-diphosphonate (EHDPTM) on bone mineral metabolism was tested in 4 healthy young men during 20 weeks of continuous bed rest. Two subjects received an oral dose of 5 mg/kg/day and the other 2 20 mg/kg/day. The low dose had two minor effects: the increase in bone accretion rate which usually occurs during bed rest was prevented, and there was an accentuation of the bed rest induced increase in hydroxyproline excretion. Skeletal mineral loss, assessed by calcium balance measurements and gamma ray absorptiometry of the calcaneus, occurred at the rate previously noted in untreated control subjects. Two types of drug effect were apparent at the higher dosage: one was immediate and sustained--a rise in serum phosphorus concentration and a fall in serum alkaline phosphatase activity. The other was delayed and progressive--a decline in urinary hydroxyproline excretion and in the rates of bone accretion and resorption. Skeletal mineral loss may have been affected; the usual negative mineral balance developed during the first half of the study, then disappeared during the last few weeks. However, gamma ray absorptiometry revealed no attenuation of the calcaneal mineral losses.


Increase in numbers of gonadotropin receptors on granulosa cells during follicle maturation. The binding of both human chorionic gonadotropin (hCG) and human luteinizing hormone (hLH) to a homogeneous population of isolated intact granulosa cells from increasingly mature porcine ovaan follicles has been studied. The number of receptor sites per granulosa cell increases 35-fold as the follicle enlarges, although cell size remains constant. This may explain the increased biologic responsiveness to gonadotropin of mature cells from large follicles. The affinity for both hormones, as determined by equilibrium dissociation constants, is high, and does not appear to change significantly as the cells mature. Comparison of dissociation constants, numbers of binding sites, and competitive inhibition between hCH and hLH, indicates that these two hormones probably interact with the same receptor on the granulosa cells.


A radioreceptor assay for follicle-stimulating hormone. A highly specific and sensitive radioreceptor assay for FSH has been developed, using partially purified plasma membranes from bovine testes. Highly purifed hFSH was radioiodinated by the lactoperoxidase method. The limit of detection of purified hFSH was 1 ng/ml on the basis of 2 times standard deviation of the zero value. The assay was applicable for measurements of FSH in serum; however, the sensitivity decreased slightly to 2.5 ng/ml. The precision of the assay was less than +/- 10% within-assays and +/- 15% between-assays as expressed by the coefficient of variation. The assay was highly specific for FSH of various species. Slight inhibition of uptake of 125I-hFSH was observed with LH and TSH; but no competition was seen with 10,000 ng/ml of insulin, prolactin, hGH, hCG, and subunits of LH. Purified hFSH (LER-1575C) was measured to be 200 times the potency of the reference FSH/LH (LER-907); and purified ovine FSH (LER-1491) and rat FSH (FSH-I-1) were estimated to be 35 times and 71 times that of oFSH-S-1 (NIH), respectively. The content of FSH in human pituitary was estimated to be 226.8 +/- 118.8 mIU/mg, and the index of discrimination (radioimmunoassay/radioreceptor assay) for pituitary FSH was demonstrated to be 1.71 +/- 0.49. For measurements of serum hFSH, the index of discrimination (radioimmunoassay/radioreceptor assay) was demonstrated to be 1.08 +/- 0.20.


ACTH and prostaglandin receptors in human adrenocortical tumors. Apparent modification of a specific component of the ACTH-binding site. The failure of certain adrenal tumors to respond to ACTH was investigated in vivo be administration of corticotropin-(1-24)-tetracosapeptide (ACTH1-24) and dexamethasone and in vitro by studying the binding properties of ACTH1-24 and prostaglandin E1 (PGE1) and their effect on adenylate cyclase activity of the tumors' crude membranes; in addition, in five cases the stimulation of cortisol production in isolated adrenal cells by both hormones and dibuttyryl cyclic adenosine 3',5'-monophosphate (cAMP) was also studied. The results obtained in 13 hormone-producing tumors of the human adrenal cortex, i.e. 10 carcinomas and 3 adenomas, were compared with those found in normal human adrenal glands. According to the adenylate cyclase responses to ACTH1-24 and PGE1, the tumors fall into different categories. In the first group are six rumors in which the adenylate cyclase was stimulated by both ACTH1-24 and PGE; in addition specific binding could be demonstrated for the two hormones in all six. The binding affinity for 125I-ACTH1-24 was found to be about 10 times higher than that for 125I-ACTH11-24. In the one tumor in which the experiment was performed, bound 125I-ACTH1-24 was displaced by ACTH1-10. These results are similar to the ones found in normal human adrenal preparations. For two rumors of the group in which ACTH did not increase steroidogenesis in vivo, the biochemical abnormality might be located beyond cAMP formation. A second group encompasses six tumors in which the steroidogenesis in vivo and the adenylate cyclase activity were insensitive to ACTH1-24 but in which the enzyme was stimulated by PGE1 and NaF. However, these preparations bound 125I-ACTH1-24 and 125I-ACTH11-24, the binding affinity being similar for both peptides but 10 times lower than the one found in normal adrenal cortex for 125I-ACTH1-24. In the only case of this group where it was tested, ACTH1-10 did not displace bound 125I-ACTH1-24. This result strongly suggests the possibility of a modification or a loss of the receptor site that binds the N-terminal sequency (1-10) of ACTH, the biologically active part of the molecule. In the last tumor, both PGE1 and ACTH were unable to stimulate adenylate cyclase activity and steroid production in a preparation of isolated adrenal cells, although steroidogenesis was stimulated by dibutyryl though steroidogenesis was stimulated by dibutyryl cAMP. No specific binding for PGE1 could be demonstrated. However, 125I-ACTH1-24 and 125I-ACTH11-24 were found to be bound to the tumor with the same affinity.


The role of superoxide anion and hydrogen peroxide in phagocytosis-associated oxidative metabolic reactions. The contribution of hydrogen peroxide (H2O2) and one of its unstable intermediates, superoxide anion (O2), to the oxidative reactions that occur in phagocytizing leukocytes was explored by depleting these cells of O2. This was accomplished by allowing them to phagocytize latex particles coated with superoxide dismutase (SOD), which catalyzes the generation of H2O2 from O2. Although the rate and extent of phagocytosis of latex coated with bovine serum albumin was similar to latex coated with SOD, the rate of oxygen consumption, [14C]formate oxidation, [1-14C]glucose oxidation, and iodination of zymosan particles was significantly enhanced by SOD. In contrast, the rate and extent of reduction of nitroblue tetrazolium (NBT) was diminished by 60%. These studies indicate that the majority of NBT reduction by leukocytes is due to O2, whereas stimulation of the hexose monophosphate shunt and iodination of ingested particles requires H2O2 generated from the increased reduction of oxygen by phagocytizing leukocytes.


Characterization of remnants produced during the metabolism of triglyceride-rich lipoproteins of blood plasma and intestinal lymph in the rat. The metabolism of intravenously injected large and small chylomicrons from intestinal lymph and of very low density lipoproteins from blood plasma was studied in functionally eviscerated "supradiaphragmetic" rats. For studies with lymph lipoproteins, recipient animals were injected with 4-amino-pyrazolopyrimidine 18 h before injection of lipoprotein to prevent secretion of very low density lipoproteins into their blood plasma. In all cases, most of the triglycerides (labeled with 14C) were rapidly metabolized, whereas cholesteryl esters (labeled with 3H) persisted in the blood. Most of the cholesteryl esters remained in smaller "remnant" lipoproteins, less dense that 1.006, which retained an apparently spherical shape, as determined by electron microscopy of negatively stained preparations. Whereas the diameters and chemical compositions of large chylomicrons were substantially different from those of small chylomicrons and very low density lipoproteins, all remnants were similar in these respects. Average remnant diameters were 400-600 A and remnants were enriched in cholesteryl esters and in protein insoluble in tetramethylurea. In addition to triglycerides, remnants were depleted of phospholiarticle size, the composition of remnants, like that of their precursors, was consistent with the "pseudomicellar" model of lipoproteins, in which a core of nonpolar lipids is covered by a monolayer of polar lipids and protein. These results domonstrate the fundamental similarity of the initial step in the metabolism of triglyceride-rich lipoproteins from intestinal mucosa and liver and show that loss of triglycerides from the core of the particles is accompanied by removal of polar components from the surface.


Cryoimmunoglobulinemia in rheumatoid arthritis. Significance in serum of patients with rheumatoid vasculitis. Cryogloculins were examined in a standardized manner in an unselected group of 35 patients with rheumatoid arthritis (RA) and 8 patients with RA complicated by cutaneous vasuclitis and neuropathy. Optimum conditions for detection and characterization of cryoglobulins were established; the proportion of resolubilized to total precipitable protein remained constant in an individual patient under these conditions. All 8 vascultis patients and 9 of 35 other patients with RA exhibited cryoglobulins; total protein and immunoglobin content were significantly higher in the cryoglobulins of patients with vasculitis. Immunoglobulins G and M constituted two-thirds and three-quarters of the total protein in the cryoglobulins from uncomplicated rheumatoid and vasculitis patients, respectively. Serum antiglobulin titers were higher, and serum C3 levels were lower, in vasculitis patients compared to rheumatoid patients without vasclitis. Anti-gamma globulin activity was detected in all cryoglobulins from vasculitis patients. Cryoglobulin IgG and IgM were polyclonal. Density gradient analyses demonstrated the majority of the cryoglobulin activity to reside in the 19S IgM fraction. There was no evidence of a light weight (8S) IgM. A monoclonal rheumatoid factor did not detect 7S-ANTI-7S complexes in the cryoprecipitates, but acid eluates from some cryoglobulins absorbed with insoluble IgG revealed an antiglobulin of the IgG class. Serial studies performed on vasculitis patients treated with cyclophosphamide disclosed a relationship between clinical evidence of vasculitis and the presence of cryoglobulins. The antigen (IgG) and antibody (largely IgM rheumatoid factor) nature of these cryglobulins is presented as evidence that the widespread vascular complications of RA are mediated, at least in part, by circulating immune complexes.


Insulin receptor deficiency in genetic and acquired obesity. We have previously shown that in the insulin-resistant obese hyperglycemic mouse (ob/ob) there is a deficiency in the number of insulin receptor sites on hepatocytes, adipocytes, and thymic lymphocytes. We now find that concentration of insulin receptors on liver plasma membranes is decreased in the db/db mouse, another form of inherited obesity, and in normal mice that became obese after treatment with gold thioglucose, while thin mice, heterozygous for the ob mutation (ob/+), have normal insulin binding. With acute and chronic food restriction of the ob/ob and gold thioglucose obese mice, there is reduction in hyperinsulinemia and an associated increase in the insulin receptor concentration toward normal. In contrast, when fasting ob/ob mice were given exogenous insulin to maintain the hyperinsulinemia, insulin receptors failed to increase. Thus, in all cases, there was a consistent relationship between the degree of hyperinsulinemia and of insulin receptor loss. These findings suggest that decreased insulin binding is a characteristic feature of the insulin resistance of obesity, and that sustained hyperinsulinemia is a major factor in the control of the concentration of insulin receptors on target cells.


Effect of Klebsiella pneumoniae enterotoxin on intestinal transport in the rat. The effects on intestinal transport of either a semipurified preparation of enterotoxin elaborated by Klebsiella pneumoniae or similaryly prepared control material were tested by marker perfusion studies in the small intestine of rats. At a concentration of 2 mg/ml, the enterotoxin produced net secretion of water, Na, and Cl in both jejunal and ileal segments; HCO3 transport was not affected. Net secretion was evident within 30 min after intorduction of the toxin and was maximal after 90 min. The addition of 56 mM glucose to the enterotoxin-containing perfusion fluid resulted in reversal of water and Na transport to net absorption in both intestinal segments. The enterotoxin also produced a significant depression of xylose absorption in both the jejunum and ileum but did not affect the absorption of either glucose or L-leucine. Intestinal structure was not altered after perfusion of the toxin but insillation of approximately one-quarter of the total perfusion dose into a ligated jejunal loop for 18 h produced fluid secretion and structural abnormalities. These observations confirm the fact that other species of coliform bacteria in addition to tescherichia coli are capable of elaborating an enterotoxin. Such species commonly contaminate the small intestine of persons with tropical sprue and it is suggested that chronic exposure of the intestinal mucosa to the enterotoxin elaborated by these bacteria may be a factor in the pathogenesis of intestinal abnormalities in thid disorder.


Thrombin-induced protein phosphorylation in human platelets. Intact human platelets loaded with 32PO4 contain multiple phosphorylated proteins. Thrombin treatment of intact 32PO4-loaded platelets results in a 2-6-fold increase in phosphorylation of a platelet protein (designated "peak 7" protein) of approximately 40,000 mol wt as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis and by gel filtration on Sephadex G-150. A similar increase in phosphorylation was observed in a platelet protein (designated "peak 9" protein) of approximately 20,000 mol wt. The time for half-maximal phosphorylation of peak 7 and peak 9 protein was 10-14 s. The concentration of thrombin at half-maximal phosphorylation was 0.25 U/ml for both proteins. Prior incubation of platelets with dibutyryl cyclic adenosine 3',5'-monophosphate or prostaglandin E1 inhibited thrombin-induced peak 7 and peak 9 protein phosphorylation. The erythroagglutinating phytohemagglutinin of Phaseolus vulgaris, a non-proteolytic release-inducing agent, induced peak 7 and peak 9 protein phosphorylation. Thus, the characteristics of peak 7 and peak 9 protein phosphorylation are similar to those of the platelet release reaction, suggesting that the phosphorylation of these proteins may play a role in the platelet release reaction. When platelet sonicates or the supernatant fraction from platelet sonicates were incubated with [gamma-32P]ATP there was phosphorylation of both peak 7 and peak 9 proteins. This phosphorylation was unaffected by either added thrombin or adenosine 3',5'-cyclic monophosphate (cAMP) despite the presence of the phosphodiesterase inhibitor 1-methyl-3-isobutylxanthine. Thus, the thrombin-dependent phosphorylation depends upon intact platelets. When the supernatant fraction from platelet sonicates was fractionated by histone-Sepharose affinity chromatography, two distinct protein kinase enzymes were resolved, one a cAMP-dependent holoenzyme and the other a cAMP-independent enzyme. The isolated cAMP-dependent enzyme fraction catalyzed the cAMP-(but not thrombin-) stimulated phosphorylation of a protein that co-electrophoresed with peak 7 protein.


The significance of inhibitor-resistant alkaline phosphatase in the cytochemical demonstration of transport adenosine triphosphatase. The hydrolysis of disodium p-nitrophenyl phosphate at pH 9.0 by slices of formaldehydee-fixed rat renal cortex was investigated by colorimetric estimation of the nitrophenol liberated. It was found that three types of activity could be identified on the basis of their responses to inhibitors and cations: (a) alkaline phosphatase sensitive to inhibition by L-tetramisole; (b) potassium-dependent phosphatase, probably identifiable with the phosphatase component of sodium-potassium-dependent transport adenosine triphosphatase (?Na-K-ATPase); and (c) alkaline phosphatase insensitive to L-tetramisole. It was found that in the presence of strontium ions, as used in Na-K-ATPase cytochemistry, the activities of the second and third types of enzyme were approximately equal. The implications of these findings for the cytochemical demonstration of Na-K-ATPase are discussed.


Use of the unlabeled antibody immunohistochemical technique for the detection of human antibody. Two methods have been developed which permit use of the unlabeled antibody immunohistochemical technique for detection of human antibody, without the need for immunization of humans with peroxidase. Human antibody to herpes simplex virus (HSV) reacted with human cell cultures infected with HSV was the experimental system. In the first method an attempt was made to employ rabbit peroxidase-antiperoxidase (PAP) soluble complexes in connectin with human antibody. This was done by sequential addition to the HSV-infected cells of (a) human anti-HSV, (b) rabbit antihuman globulin, (c) guinea pig antirabbit globulin (the bridging reagent) and (d) rabbit PAP. Strong specific staining of HSV-infected cells was obtained; however, difficulties were encountered with nonspecific reactions on uninfected cells. In the second method PAP soluble complexes prepared with baboon antiperoxidase were bridged to the human anti-HSV antibody by rabbit antihuman globulin. Because of the phylogenetic relatedness of human and baboon globulins this resulted in firm binding which gave strong specific staining of HSV-infected cells without significant reaction in uninfected cells.


Epidermal activity of NAD-dependent isocitrate dehydrogenase in psoriasis during treatment with dithranol. The activity of ICDH(NAD) was measured in subcorneal and basal epidermal layers in 8 patients with psoriasis and in 7 healthy controls treated once a day with 0.15% dithranol in white petrolatum for 2 weeks. Skin biopsies were taken before and on days 2, 6, and 14 of the treatment. Lowry's microtechniques were used in conjunction with a bioluminescent system (bacterial luciferase) for enzymatic assays. The enzymic activity could be related to the type of keratinization present in the stratum corneum overlying the epidermal areas under study. In orthokeratotic areas from the controls, in noninvolved, and in treated involved skin the activity was low. In parakeratotic areas, as found in treated noninvolved and in involved psoriatic skin, the enzymic activity was increased to a level at least twice that found in orthokeratosis. Since ICDH(NAD) activity reflects an aspect of mitochondrial function, the results suggest that mitochondrial activity may be important in control of keratinization.


Varicella-Zoster Immunoglobulins during Varicella, Latency, and Zoster. Antibodies to membrane antigens of cells infected by varicella-zoster virus were detected by immunofluorescence. Rises in varicella-zoster IgA, IgG, and tigM were detected after both varicella and zoster. Prompt antibody responses were observed in patients with generalized zoster as well as in those with localized zoster. A delayed response was found, moreover, in one patient who did not develop disseminated disease. Antibody to varicella-zoster virus was detected in the sera of all three patients tested prior to and in all but one of 63 patients tested soon after onset of zoster. The level of varicella-zoster IgA and IgG rose in two of three immune patients after household exposure to infection with varicella-zoster virus. Varicella-zoster IgG was detected in the cord blood of infants whose mothers were immune to varicella.


Spread of Streptococcus pneumoniae in families. II. Relation of transfer of S. pneumoniae to incidence of colds and serum antibody. Factors that affect the spread of Streptococcus pneumoniae and the antibody responses associated with colonization were studied in 64 families for periods of eight to 52 weeks. Surveillance included daily recording of respiratory symptoms and bimonthly pharyngeal cultures for identification of the pneumococcal carrier state. Rhinovirus cultures were included for a portion of the study period. Intrafamilial carriage of a single type of S. pneumoniae and simultaneous spread to more than one family member were commonmspread often occurred in association with an upper respiratory tract infection; simultaneous transmission of S. pneumoniae and a rhinovirus was documented. Preexisting, type-specific serum antibody did not prevent acquisition of homotypic S. pneumoniae but did appear to shorten the duration of pharyngeal carriage. Sera of all 11 adults had greater than 150 ng of antibody nitrogen/ml of homotypic serum antibody (measured by a radioimmunoassay) before colonization. In contrast, only one of 13 preschool children had homotypic antibody concentrations of this magnitude before colonization. A threefold or greater rise in the concentration of homotypic antibody occurred in 13 of 24 children (54%) after acquisition of S. pneumoniae; the increase in antibody concentration was associated with illness in six of the children. On the other hand, acquisition of S. pneumoniae in adults was not associated with an increase in concentration of homotypic serum antibody.


Experimental infection with herpes simplex virus type 2 in newborn rats: Effects of treatment with iododeoxyuridine and cytosine arabinoside. Sprague-Dawley rats were inoculated, within 36 hr of birth, with either of two strains of herpes simplex virus type 2. The effects of treatment for five days with either 5-iododeoxyuridine or cytosine arabinoside in near toxic doses were studied. Evaluation of treatment based on rate of survival, incidence of lesions, and isolation of virus from the central nervous system showed no appreciable difference between treated and untreated littermates. Drug-related effects of retardation of growth and defective development of the cerebellum and retina were found. The validity of therapy with either of these drugs in generalized herpetic infections is questioned because activity was minimal against the infecting agent even when the dosage of the drug was sufficient to produce serious defects in certain developing tissues.


Antiviral activity of adenine arabinoside and iododeoxyuridine in human fetal intestinal and tracheal organ cultures. In vitro antiviral activities of two potentially clinically useful antiviral compounds, adenine arabinoside and iododeoxyuridine, were examined in human fetal intestinal and tracheal organ cultures infected with Herpesvirus hominis (types 1 and 2) or vaccinia virus. The two compounds were similarly active against the viruses in organ culture, and minimal inhibitory concentrations could be determined by titration of organ culture fluid harvests into tissue culture or directly in the organ cultures themselves. Minimal inhibitory concentrations were consistently lower in tracheal than in  intestinal organ cultures and were consistently higher for H. hominis type 2 than for H. hominis type 1. Thus the organ cultures are promising systems in which to evaluate antiviral activity against those agents that replicate in vitro only in organ culture, and they may have particular application to the study of herpetic tracheitis and esophagitis.


Immunofluorescent staining for the measurement of antibodies to Herpesvirus hominis. Frozen sections of Herpesvirus hominis-inoculated mouse brain provide excellent antigens for indirect immunofluorescent staining of antibodies to H. hominis in serum. Indirect immunofluorescent staining is a rapid, sensitive, and reproducible method for the measurement of antibodies to H. hominis. The method can be applied to the clinical and epidemiological studies of H. hominis infections.


Variations in hybridization of RNA from different mouse tissues and embryos to endogenous C-type virus DNA transcripts. Several adult tissues, newborns, and embryos of uninfected BALB/c mice were analysed for RNA complementary to [3H]-DNA transcripts synthesized from an endogenous type-C virus of BALB/c 3T3. The technique of RNA:DNA hybridization was used and the extent of hybridization was measured by the use of a single-strand-specific nuclease (S-I), purified from Aspergillus oryzae. Virus-specific RNA was detected in all adult and embryonic tissues tested. However, the RNA extracted from tissues having higher proliferative activity, such as spleen, small intestine, uterus and embryos, hybridize the [3H]-DNA probe to a greater extent than the RNA from tissues with low proliferative activity, such as kidney and liver. These observations add further support to the view that the repression of the virus genome in normal cells is not complete, and suggest the existence of a correlation between a qualitative or quantitative change in the endogenous C-type virus genome transcription pattern and cell proliferation.


Genetic susceptibility of chicken times quail hybrid embryos to avian RNA tumour viruses. An attempt was made to hybridize the chicken (Gallus domesticus) male with Japanese quail (Coturnix coturnix japonica) female in order to study the genetic susceptibility of hybrid embryos to avian RNA tumour viruses of subgroups, A, B, D and E. In the hybrids the results supported the prevailing concept that susceptibility is dominant over resistance regardless of the dominant trait contributed by either parent. It was also observed that the Ie gene of the chicken was unable to suppress the 'quail-coded' susceptibility to subgroup E virus in the hybrid system, suggesting the lack of penetrance of the Ie gene. Despite the fact that some hybrids were resistant to viruses of subgroups B and D, they were susceptible to subgroup E virus, which was not expected on the basis of the concept that subgroup B-resistant cells cannot be E-susceptible. Also, the hybrids were susceptible to E virus regardless of gs antigen expression and presence of the Ie gene in the genome. This indicates that our earlier suggestion that the Ie gene is another expression of the gs antigen-determining gene is inconsistent.


Virus-specific proteins associated with the replication complex of poliovirus RNA. The poliovirus replication complex was isolated and purified from infected HeLa S3 cells. Preparations with RNA-dependent RNA polymerase activity were concentrated 200- and 1000-fold with respect to the original virus and total protein content. The enzyme activity was found to be associated with the proteins NCVPI, 2, 3, 4, (5), 6 and VPl/NCVPx. The structural proteins VP2, 3 and 4 were not present. Addition of cycloheximide to infected cells resulted in a decrease in the in vitro polymerase activity and a loss in NCVPI content. Treatment of the infected cells with toloylsulphonyl-phenylalanine chloromethyl ketone (TPCK) and iodoacetamide (IAA) led to an inhibition of in vivo RNA synthesis. The 750 g supernatant fluids obtained from extracts of these cells were able to block RNA synthesis in vitro. Electrophoretic profiles of the respective protein compositions indicate that large virus precursor proteins are responsible for the inhibition of poliovirus RNA synthesis in vivo and in vitro.


Alcoholic neuropathy. An electron-microscopic study. The sural nerves of 6 patients with different signs of alcoholic neuropathy were studied qualitatively and quantitatively by electron microscopy. Myelinated and unmyelinated fibres showed degenerative changes of the Wallerian type. Concomitant involvement of the myelin appears to be secondary to axonal lesions. Regenerative processes, although frequently observed, did not balance the destruction of fibres in the degenerative phase. Quantitative studies indicated a reduced number of myelinated fibres and decreased percentage of the area covered in cross-section by myelinated and unmyelinated axons. The histograms of myelinated fibres showed a shift to the left of the peak of large and small fibres and an increased number of small-sized fibres. Similarly the histograms of unmyelinated fibres showed a shift to the left and a bimodal distribution with an increased number of small-sized fibres. Imbalance in degenerative and regenerative processes seems to be the basis of the chronic partial denervation observed in the nerves of alcoholic patients in this study.


Ultrastructural studies in human polymyositis. Biopsy of muscle from 13 cases of human polymyositis permitted an ultrastructural study in which attention was directed to the cellular population of the inflammatory reaction. The constant feature was the presence of small and large lymphocytes, and cells of the latter type were frequently in intimate contact with the surface of the muscle fibres. Macrophages were seen both within and outside degenerating muscle fibres. Plasma cells were seen in 2 cases. Filaments bearing some resemblance to virus inclusions were seen in one case. The implications of the cellular reaction in human polymyositis are discussed and the literature of ultrastructural observations on the muscle in this disease is briefly reviewed.


Prognosis in adult polymyositis.

Prognosis in adult polymyositis. The effects of long-term corticosteroid treatment have been assessed in 20 patients with polymyositis. Cases were only accepted if a muscle biopsy showed the characteristic features of inflammation and muscle fibre degeneration. If the muscle disorder was only a minor feature of generalised collagen disease or there was evidence of coexisting neuropathy, the patients were excluded. Over a mean follow-up period of 5 years, 8 patients died and only 4 improved. These results have been compared with those in other series. An attempt has been made to correlate muscle biopsy findings with prognosis, and the reasons for the poor outcome are discussed.


Demonstration of enzymes related to myelinogenesis in established human brain cell cultures. CNP, CGalT and PAPS-CST, enzymes involved in myelinogenesis were demonstrated in cultures derived from human brain. Little or no CNP activity could be found in cultures derived from non-neurogenic tissue. Transformation of brain cultures by PML-SV40 virus resulted in a 3 to 10-fold increase in the specific activity of CNP.


Analysis of liver scanning in a general hospital. The specificity and sensitivity of liver scanning in a general referral population of 125 patients was studied. All patients had a liver biopsy prior to the scan or not more than 10 days later. The results of this study were compared with one earlier and one ongoing study in the same laboratory. The differences in accuracy are mostly attributable to the difference in the populations studied. It is apparent that the multiple-view scintillation camera technique is not superior to the rectilinear two-view scans for studying the liver.


Evaluation of soft-tissue calcifications in dermatomyositis with 99mTc-phosphate compounds: case report. A whole-body scan with 99mTc-pyrophosphate and 85Sr-nitrate demonstrates extension of calcinosis in one case of dermatomyositis with cutaneous, subcutaneous, and muscular calcinosis. The authors suggest the potential use of 99mTc-phosphate compounds as an auxiliary instrument in the evaluation of dermatomyositis-polymyositis syndrome.


Gallium uptake in benign tumor of liver: case report. A case of positive tracer localization in a benign tumor of the liver on a 67Ga-citrate scan is reported. The authors were unable to find any previous reports of positive localization of gallium in this type of liver tumor.


Effects of dietary protein and calcium levels on renal glucose-6-phosphatase activity of intact and thyroparathyroidectomized rats. In order to determine the relationship of parathyroid hormone and levels of dietary protein and calcium with the activity of renal glucose-6-phosphatase (G6Pase), effects of two levels of dietary protein, namely, 25 and 75%, on the enzyme activity were compared at three levels of dietary calcium, namely, 0.06, 0.63, and 1.83%, with the use of intact and thyroparathyroidectomized (TPTX) rats. In intact rats, 0.06% dietary calcium caused an increase in renal G6Pase activity in rats fed the high carbohydrate diet, and dietary calcium in excess (1.83%) caused the enzyme activity to decrease. Similar responses in the activity of renal G6Pase to the variation of dietary calcium levels were seen in rats fed the high protein diet, but significant differences were not obtained. In TPTX rats fed the high carbohydrate diet, the activity of renal G6Pase was significantly decreased compared with that of intact rats. When TPTX rats were fed the high protein diet, however, no significant decrease in the enzyme activity was observed. Free access to aqueous 0.1% CaC1(2) solution by TPTX rats tended to restore the activity of renal G6Pase and serum calcium concentrations depressed by thyroparathyroidectomy. In addition, a significant correlation was observed between the total activity of renal G6Pase and serum calcium concentrations. Hypothyroidism produced by oral administration of propylthiouracil (0.05% of diets) did not affect the enzyme activity in the kidneys of rats fed the high carbohydrate and the high protein diets. The results suggest that the activity of renal G6Pase of rats fed the high protein diet might be less susceptible both to dietary calcium levels and to parathyroid function than that of rats fed the high carbohydrate diet.


Radiographic survey of perlite workers.

Radiographic survey of perlite workers. Chest roentgenograms of 240 perlite workers employed for 1 to 23 years in the industry, showed no evidence of pneumoconiosis associated with perlite exposures. One individual, found to have simple pneumoconiosis, and one found to have complicated pneumoconiosis, had formerly been diatomaceous earth workers. Since only 28 of the men had been in the industry over 15 years and only seven for 20 years or more, continued surveillance is essential to make sure that there are no effects with more prolonged exposures. Studies of pulmonary function of the individuals who have had relatively long exposures are needed to supplement radiographic evidence.


Pulmonary sarcoidosis: A clinico-pathological study. The light and electron microscopic changes in biopsy tissue from the lung of a 30-year-old housewife severeley incapacitated by diffuse pulmonary sarcoidosis with pulmonary hypertension are presented. The lung tissue was distorted by numerous granulomas in the interstitial tissues and within alveoli. Many pulmonary blood vessels including arteries were damaged by the granulomas. The ultrastructural features of the epithelioid cells were found to be distinctive and probably specific. The giant cells which accompanied the epithelioid cells contained two types of inclusion body: one appeared to be related to the Schaumann body but the nature and origin of the second type was not clear. Many of the granulomas were surrounded by avascular fibrous tissue which contained, in addition to mature fibroblasts, myofibroblasts and a primitive form of cell that appeared to be a fibroblast precursor. It was conjectured that the myofibroblasts, through their contractile powers, might increase the distortion of the lung architecture and thereby the patient's disability. The alveolar walls were thickened by a diffuse infiltrate of macrophages and epithelioid cells but there was no excess of collagen and elastic fibres. The evidence suggested that the epithelioid cells developed from macrophages. From the cellular nature of the diffuse infiltration of the alveolar walls and the absence of fibrosis it seemed that the disease was still at an early and active stage, a conclusion strengthened by the fact that treatment with corticosteroids led to marked and sustained clinical improvement.


Malignant chondroid syringoma of the thigh. Report of a case with electron microscopy of the tumour. A case of malignant chondroid syringoma occurring in a 52-year-old female is reported. The tumour presented as a large, cystic lesion of the thigh. Local recurrence and secondary deposits in the inguinal region occurred at an early stage. The rarity of these malignant tumours and metastasising sweat gland tumours in general is emphasised.


Hepatotoxicity of chemotherapy following nephrectomy and radiation therapy for right-sided Wilms tumor. Two children developed hepatotoxicity during treatment for right-sided Wilms Tumor. Treatment consisted of nephrectomy, irradiation, and chemotherapy with actinomycin D and vincristine. Hepatic enlargement, thrombocytopenia, and abnormalities in liver function and seen on the liver scan occurred at the time of the course of chemotherapy administered 20 days after completion of irradiation. These abnormalities disappeared when treatment was temporarily suspended. Vincristine and actinomycin D were subsequently reintroduced without evidence of hepatotoxicity. Actinomycin D after irradiation for right-sided Wilms tumor may produce severe liver toxicity.


Adriamycin cardiotoxicity in a child with Wilms tumor. Report of a case and review of the literature. A case of adriamycin-induced carciomyopathy in a 5 3/4-year-old girl with metastatic Wilms tumor is reported. Potentiation of the adriamycin cardiotoxicity by irradiation to the heart is proposed; the clinical and pathologic findings are discussed.


Interactions of morphine, adenosine, adenosine triphosphate and phosphodiesterase inhibitors on the field-stimulated guinea-pig ileum. Inhibition of the electrically induced contractions of the guinea-pig ileum has been shown to be a reliable index to the relative potency of various narcotic analgesics. This property suggests that this preparation might be used as a model in attempts to elucidate the mechanism(s) by which morphine induces analgesia in the central nervous system. Since it has been demonstrated that some adenosine derivative may function as an endogenous inhibitory transmitter in the gut, the effects of adenosine, adenosine triphosphate (ATP) and morphine on the ileum were further characterized and compared. Morphine, adenosine and ATP produce a substantial inhibition of the isometric contractions induced by transmural field stimulation. The inhibition produced by each is antagonized by 2.5 times 10(-7) M tolazoline whereas that produced by ATP is potentiated by 4 times 10(-7) M 5-hydroxytryptamine. The inhibitory effects of morphine and ATP can also be markedly potentiated by two of the several phosphodiesterase inhibitors tested, Ro 20-1724 and dipyridamole. In addition, pretreatment of the ileum with either adenosine, ATP or morphine can produce a significant potentiation of the inhibitory effects of norepinephrine. The above suggests that cyclic adenosine 3',5'-monophosphate may play a role in mediating some of the inhibitory effects produced by exogenous adenosine, ATP and morphine. In addition, the similarities between the effects produced by these substances indicates that the biochemical pathways responsible for mediating the effects of each may share some common elements.


Effect of chlorpropamide on renal response to parathyroid hormone in normal subjects and in patients with hypoparathyroidism and pseudohypoparathyroidism. Chlorpropamide inhibited by 35 to 65% the increase in the urinary excretion of adenosine 3',5'-monophosphate (cyclic AMP) following a large dose (250 U) of parathyroid hormone (PTH) in normal subjects and patients with hypoparathyroidism and pseudohypoparathyroidism. By contrast, in normal subjects, the response of urinary cyclic AMP excretion to smaller amounts of PTH (15 and 30 U) was not decreased by chlorpropamide. Chlorpropamide did not decrease the phosphaturic response to any dose of PTH. The probable explanation for the discrepant effects of chlorpropamide on urinary cyclic AMP and phosphate excretions is that phosphaturia results from a minimal elevation of cyclic AMP and that chlorpropamide does not decrease cyclic AMP production to such a low level. Chlorpropamide decreased the accumulation of renal cyclic AMP in response to PTH in the parathyroidectomized rat, suggesting that this may be the mechanism of action for this drug in decreasing the urinary excretion of cyclic AMP in response to PTH in man. Tolazamide, another sulfonylurea, did not inhibit the elevation of urinary cyclic AMP excretion after PTH. Therefore, the sulfonylurea part of the molecule is probably not involved in the inhibition produced by chlorpropamide.


Protein kinases activated by cAMP in the genital tract of spayed mice treated with oestradiol-17beta. The cAMP-dependent protein kinase (ATP:protein phosphotransferase, EC 2.7.1.37), has been studied in the vaginal epithelium, vaginal stroma, endometrium, and whole uterus of spayed mice treated with oestradiol-17 beta, and in the vaginal epithelium and uterus of spayed mice. Two protein kinase isoenzymes (PK I and PK II) were found in whole uterus, endometrium, and vaginal stroma. Vaginal epithelium contained only one isoenzyme (PK II). Oestradiol treatment increased PK I relative to PK II in the uterus. The isoenzyme pattern in the vaginal epithelium was unaltered after such treatment. The total protein kinase activity was 70% higher in uterine extracts (cytosol) than in extracts from vaginal epithelium. Oestradiol treatment did not influence the total protein kinase activity in either tissue.


Regulation of seminiferous tubular function by FSH and androgen. Seminiferous tubules contain a cytoplasmic androgen receptor similar to the receptors in the epididymis and ventral prostate. The presence of a cytoplasmic receptor indicates that androgens maintain spermatogenesis by a direct action on certain types of cells within the seminiferous tubule. The Sertoli cell appears to be one of the cell types containing androgen receptors and the receptor might also be present in spermatogonia, primary spermatocytes, or peritubular cells. The Sertoli cell is stimulated by FSH to produce an androgen-binding protein which may serve to increase the accumulation of androgen in the seminiferous epithelium and make it available for binding by intracellular androgen receptors. This may be a way in which FSH enhances the action of androgen on spermatogenesis. Androgens act on the Sertoli cell to increase its response to FSH. This action of androgens on the Sertoli cell results in increased production of androgen-binding protein and may enhance the production of other substances which exert trophic effects on spermatogenesis.


Synthesis and cell culture studies on the antiviral activity of 5-mercaptomethyl-2'-deoxyuridine. Treatment of 5-mercaptomethyluracil (I) with trimethylsilyl chloride in the presence of triethylamine gave 2,4,5-tris-(trimethylsilyl)-5-mercaptomethyluracil (II) which, upon coupling with 2-deoxy-3,5-di-O-(p-toluoyl)-D-erythro-pentofuranosyl chloride, furnished as anomeric mixture of fully substituted 2'-deoxy ribonucleosides. The nucleoside with beta configuration (III) was predominantly formed and was isolated as a crystalline solid. The free nucleoside IV was obtained by removal of blocking groups by sodium methoxide catalyzed deacylation, deionization under reducing atmosphere, and chromatography on neutral alumina. IV is oxidized to the corresponding disulfide V in solution in the absence of thiols. IV was found to be markedly inhibitory against the herpes virus of infectious bovine rhinotracheitis (IBR). Against this virus, IV was found to be as potent as 5-iododeoxyuridine and cytosine arabinoside when added 18 hr before virus infection.


Flavonoids. 8. Synthesis and antifertility and estrogen receptor binding activities of coumarins and delta3-isoflavenes. A series of coumarins and delta3-isoflavenes was prepared. Although antifertility activity was shown by all of these compounds, the required dosage in mice varied from 13.5 mug/kg/day to 50 mg/kg/day. The most potent compounds were the 2-methyl-4-ethylisoflavenes, two of which (2a and 2b) were about equipotent with DES on a molar basis. They were followed by the 2,2-dimethylisoflavenes, the 2-unsubstituted isoflavene, and the coumarins. The most active compounds possessed an acetoxy group at C-7 and an oxygen function at C-4'. Presence of fluorine at C-4' or diethylaminoethoxy at C-M decreased the antifertility activity. The uterotropic activity followed the same trends as the antifertility activity with some evidence for the separation of the two effects in the 2,2-dimethylisoflavene series. Based on a limited study it appears that two phenolic hydroxyl groups are required for the presence of good estrogen receptor binding activity. An apparent lack of correlation between the estrogen binding activity and uterotropic or antifertility effects is probably explained by in vivo metabolism.


Uptake in vitro of nucleic acid precursors and nucleic acids by Zajdela ascitic hepatoma cells. Zajdela ascitic hepatoma cells are shown to take up pyrimidine bases at much lower rates than obtained in slices from normal rat liver. The rates of uptake of adenine and uridine by the Zajdela cells are, however, as high as in the slices. Like the slices, again, the Zajdela cells take up E. coli RNA and DNA at very low rates but, unlike the slices, thses cells degrade rapidly the RNA taken up. The Zajdela cells resemble parenchymal cell suspensions derived from normal rat liver in regard to the uptake of pyrimidine bases and the ability to degrade heterologous RNA.


Quantum chemical study of relative reactivities of a series of amines and nitriles: relevance to prebiotic chemistry. Using the Iterative Extended Hucken Theory (IEHT) calculations of the elctron distribution and orbital energies of a series of thirteen amines, nitriles and amino-nitriles relevant to prebiotic and cosmo-chemistry have been carried out. Ground state properties such as the energy and nature of the highest occupied (HOMO) and lowest empty (LEMO) molecular orbitals, net atomic charges and number of non-bonding electrons have been identified as criteria for correlating the relative nucleophilicity of amine and nitrile nitrogens and the electrophilicity of nitrile and other unsaturated carbon atoms. The results of such correlations can be partially verified by known chemical behavior of these compounds and are used to predict and understand their role in prebiotic organic synthesis.


Study of double and triple simultaneous substitutions of amino acids in cytochromes C. A statistical analysis aimed at obtaining some informations on a possible correlation between simultaneous amino acid substitutions is proposed.  This method is applied to a set of cytochrome C, at the level of tandem and triple substitutions separated along the peptide chain by 1 to 15 peptide bonds.  Monte-Carlo simulations are performed and the results are compared.  We find a significant occurrence of three adjacent amino acid substitutions in which the first replacement requires a two nucleotide substitution. A possible explanation of this fact is proposed on  the basis of covarions.


Evaluation of antitumor activity of Bordetella pertussis in two murine tumor models. The antitumor activity of three preparations of killed Bordetella pertussis (Bp) (Eli Lilly crude and fluid pertussis vaccines and Parke-Davis pertussis vaccine) was studied in the B16 melanoma and CaD2 mammary adenocarcinoma models. In these tumor systems; Bp had weak and variable tumor inhibitory activity and did not augment tumor rejection immunity. The intratumor injection of Bp did not affect the growth of the B16 tumor but significantly inhibited the growth of the CaD2 tumor. However, the established tumor did not regress. Admixture of Bp with B16 cells before inoculation inhibited tumor growth and prolonged survival of inoculated mice. Admixture of Bp with CaD2 cells completely suppressed tumor cell growth in 60% of inoculated mice. Intratumor injection of CaD2 with Bp combined with surgery provided no protection against subsequent development of metastases.


Tumor induction by carcinogenic agents in aquarium fish. The effects of nine carcinogens on 1,220 guppies [Poecilea reticulata (Lebistes reticulatus)] and 40 zebra fish (Danio rerio) were studied. Multiple exposure techniques included skin application, im and ip injections, feeding, implantation in pellets, and dissolving of the compound in the aquarium water. 7-12-Dimethylbenz[a]anthracene, 3-methylcholanthrene, and benzidine produced no tumors in the fish. N-2-Fluorenylacetamide, omicron-aminoazotoluene, 4-dimethylaminoazobenzene, diethylnitrosamine, and dimethylnitrosamine induced tumors in the livers of some of the fish. These neoplasms included cholangiomas, hepatoadenomas, cholangiocarcinomas, and hepatocellular cancers. Nitrosomorpholine caused not only hepatic tumors in guppies and zebra fish but also intestinal adenocarcinomas and poorly differentiated connective-tissue lesions in the abdominal cavities of zebra fish. Experimental induction of tumors in aquarium fish offers wide possibilities for comparative cancer research. Fish are a suitable model for the testing of compounds for carcinogenic activity and for the screening of environmental carcinogens.


Response of microbodies in Morris hepatoma 9618A to clofibrate. Regulation of the formation of microbodies in Morris hepatoma 9618A was studied by examination of the response of the organelles to clofibrate. The fine structures of microbodies in the hepatoma cells closely resembled those in hepatocytes of normal adult rats. In clofibrate-treated rats, the tumor cells showed a slight increase in the size of microbodies and in catalase activity; however, the tumor microbodies did not increase in number. In contrast, in adult clofibrate-treated rats and rats on the day of birth whose mothers received clofibrate during the gestation period, the hepatocytes showed microbodies that were greater in both number and size, and the catalase activity in the liver was definitely elevated.


Zinc intake, neoplastic DNA synthesis, and chemical carcinogenesis in rats and mice. DNA synthesis in a transplanted hepatoma induced by 3'-methyl-4-dimethylaminoazobenzene was significantly reduced (P less than 0.01) in rats maintained on diets low (0.4 mug/g) or high (greater than or equal to 500 mug/g) in zinc when compared with control animals given 60 mug zinc/g. 3-Methylcholanthrene-induced carcinogenesis was considerably lowered in mice receiving the same low or high zinc diets during the induction periods.


C-Type virus particles in a cell line from a lymphosarcoma of a nude mouse. C-type virus particles were observed by electron microscopy in cultures of a cell line (NML-1) derived from a lymphosarcoma that arose spontaneously in a nude NIH outbred mouse. The presence of such particles indicated that human tumors heterotransplanted in nude mice might become contaminated with murine oncornaviruses.


Oncogenesis by Marek's disease herpesvirus in chickens lacking expression of endogenous (gs, chick helper factor, Rous-associated virus-O) and exogenous avian RNA tumor viruses. Chickens free of exogenous avian leukosis virus (ALV) infection, replicating endogenous ALV (Rous-associated virus-O), gs antigen, and chick helper factor were fully susceptible to induction of Marek's disease (MD) by ALV-free MD viruses. Dual infection with Rous-associated virus-2 and MD virus did not significantly alter the character of the MD lesions. Thus exogenous ALV infection was not requisite for MD virus-induced oncogenesis. Although participation of endogenous RNA tumor virus genes in MD lesion induction could not be excluded, expression of such genes in MD tumors as gs antigen was not established.


Antibodies to Herpes simplex virus in Jewish women with cervical cancer and in healthy Jewish women of Israel. Herpes simplex virus (HSV) antibody titers were examined in sera from 39 Jewish women with squamous cell carcinoma of the uterine cervix (CaCx) and in sera from controls matched by age and country of origin. Highly significant differences were found between the cases and controls for both HSV type 1 (HSV-1) and HSV type 2 (HSV-2). Compared to findings in other demographic areas, the geometric mean titer (GMT) of HSV-1 among the CaCx cases were considerably higher, whereas the GMT for HSV-2 was in the same range. The percentage of HSV-2-positive patients, as defined by the HSV-2/HSV-1 antibody titer ratio was low compared to that found in other demographic areas; this was presumably due to the high incidence of HSV-1 infection in the population. The HSV-1 and HSV-2 infection rate in the Israeli Jewish female population was estimated by antibody titers for 94 healthy subjects. The GMT of HSV-1 was considerably higher, whereas the GMT of HSV-2 was lower, than those reported elsewhere. The association found previously between HSV-2 and CaCx remained true for Jewish women. Their low incidence of CaCx, which did not seem to result from lower susceptibility, might be explained by the low incidence of HSV-2 infection.


Aging changes in CD-1 HaM/ICR mice reared under standard laboratory conditions. Three hundred CD-1 HaM/ICR mice were observed for 2 years, and useful necropsies were done on 99 males and 102 females. Mortality was 50% at 16 months in the males and 18 months in the females. Among mice that came to autopsy, total tumor incidence was 54% for males and 75% for females, with most neoplasms occurring after 18 months. Adenomas or adenocarcinomas of the lung were the most frequent, followed by lymphoreticular tumors, vascular tumors, hepatomas, subcutaneous fibrosarcomas, and adenocarcinomas of the mammary glands. Some degree of amyloidosis was seen in half the mice of both sexes, beginning at 8 months in males and 12 months in females. Variability in tumor incidence among small groups if mice emphasized the need for adequate samples.


Extraction of tumor-specific antigen from cells and plasma membranes of line-10 hepatoma. Tumor-specific antigen was extracted with 3 M KCl from line-10 guinea pig hepatoma cells. The yield of antigenic activity, estimated by production of delayed cutaneous hypersensitivity reactions in line-10 immune guinea pigs, was 10-30% of the antigen present in intact cells. By ultracentrifugation criteria, the extracted antigen was soluble. Gel filtration, ion exchange chromatography, and salting-out studies showed that the antigen was heterogeneous in size and net charge. The possibility that 3 M KCl extracted a homogeneous population of molecules associating into polymers of various sizes at low ionic strength was ruled out by heterogeneity on Sephadex G-200 chromatography at high ionic strength. After osmotic lysis of sucrose-loaded line-10 cells, whole plasma membranes or large membrane fragments were obtained in a yield of about 20%. The isolation procedure did not cause detectable loss of membrane antigenic activity. The membranes had 33 skin test U/mg membrane protein, compared to the intact cell value of 1.7 skin test U/mg cell protein. Extracts of plasma membranes had 10-20% of the antigenic activity of the starting membrane material. In contrast to the wide variety of proteins liberated from intact cells, much of the protein extracted from the membranes was in the molecular weight range above 250,000.


An atlas of subgross pathology of the human breast with special reference to possible precancerous lesions. One hundred ninety-six whole human breasts were examined by a subgross sampling technique with histologic confirmation. The method permitted the enumeration and identification of essentially all the focal dysplastic, metaplastic, hyperplastic, anaplastic, and neoplastic lesions. Of the 196, 119 were suitable for complete quantitative morphologic analysis of the focal lesions by type. They consisted of 67 breasts obtained by autopsy, 29 cancerous breasts obtained by mastectomy, and 23 contralateral to those with cancer. All lesions, photographed subgrossly, were subsequently confirmed and correlated histologically. Morphologic evidence supported the hypothesis that most lesions traditionally grouped as mammary dysplasia or fibrocystic disease, including apocrine cysts, sclerosing adenosis, fibroadenomas, various forms of lobules (sclerotic, dilated, hypersecretory, hyperplastic, atypical, or anaplastic), ductal carcinoma in situ (DCIS), and lobular carcinoma in situ (LCIS), arose in terminal ductal-lobular units (TDLU) or in the lobules themselves. A probable exception was papilloma of ducts larger than terminal ones. Isolated foci of DCIS within the TDLU were seen in 40% of cancerous breasts, which indicated that the disease often was multifocal. Of the contralateral breasts, the 60% with clinical cancer contained such lesions, and data were in accord with the clinically known fact that women with previous breast cancer have a high rate of the disease in the remaining one. An atypical lobule (AL) of type A (ALA) had the following characteristics: a) It was more common in cancerous breasts or in those contralateral to cancer than in breasts not so identified; b) it had lobular morphology and was a terminal structure on the mammary tree; c) it tended to persist after the menopause, whereas normal lobules usually atrophied; d) it variable degrees of anaplasia forming an arbitrary continuum from normal lobules to ductal carcinoma in situ; and e) as ALA progressed to DCIS, the unfolded lobule resembled a duct which gave the false impression that DCIS was a ductal lesion. The morphologic evidence supported that hypothesis that the lesions herein called AL were derived from TDLU and were precancerous.


Aerosolized BCG (Tice strain) treatment of bronchogenic carcinoma: phase I study. Twenty patients with pathologically proved non-resectable bronchogenic carcinoma were treated with 100 aerosolized BCG (Tice strain) doses in addition to conventional treatment. The procedure is based on findings that, generally BCG must be closely associated with neoplastic cells to be effective as an immunotherapeutic agent. Bronchogenic malignancy, usually of mucosal origin, is logically treated in this manner. We report here the findings and developments of 10 patients who were treated at least five times each (for a total of 81 treatments) and pertinent experience relating to these and another 10 patients treated a total of 19 times. Local and systemic reactions were frequent and consisted of fever, cough, dyspnea, nausea, vomiting, anorexia, and malaise. Four of the 20 patients (20%) had reactions with the first treatment; by the fourth treatment 6 of 6 (100%) were affected. Prednisone given prophylactically reduced the intensity and the frequency of reactions. There were no severe side effects, obvious BCG infections, or significant changes in pulmonary or liver functions or hematologic values. No patient acquired purified protein derivative sensitivity, although 3 persons converted other skin tests to positive. There was no improvement in actuarial survival time.


Malignant tumors in American black and Nigerian children: a comparative study. Results of a study on the relative frequencies of tumors in American black and Nigerian children were compared with data from the Childhood Cancer Registries in Manchester, United Kingdom, and Kampala, Uganda. The American black child living in Washington, D.C. and the Caucasian child living in Manchester had similar high frequencies for leukemia and glioma, whereas the incidence of lymphoma and retinoblastoma was low. African children living in Nigeria or Uganda had the opposite frequency patterns. These differences in frequencies of tumors between two ethnologically related population groups, American black and Nigerian, suggested the influence of environmental factors in the etiology of these tumors, even though exposure to environmental carcinogens was short. The rarity of Ewing's sarcoma and testicular tumors in American black and Nigerian children suggested a genetic influence.


Hemolytic anemia induced by murine erythroblastosis virus: possible mechanisms of hemolysis and effects of an interferon inducer. Murine erythroblastosis virus (MuEV), also called murine leukemia virus-Kirsten, is a member of the murine type-C-RNA leukemia-sarcoma group of oncogenic viruses. Like other members of this group, MuEV can elicit both a hemolytic disorder and an oncogenic response. Neonatal rats infected with MuEV succumb to this hemolytic disorder unless they are treated with the synthetic double-stranded polyribonucleotide, polyinosinic-polycytidylic acid (poly I-poly C). Animals receiving poly I-poly C had markedly reduced levels of virus reproduction as measured by bioassay and electron microscopy. The proliferation of erythroblasts after MuEV infection in animals not receiving poly I-poly C appeared to be an erythropoietin-dependent compensatory response to hemolysis. The hemolysis itself seemed to require virus reproduction in the cell types affected. Administration of poly I-poly C to MuEV-infected rats inhibited virus reproduction and thus may circumvent the hemolytic disease syndrome. The ultrastructure of the virus and of the virus reproduction was also studied.


Inhibitory effect of adult bovine serum on release of infectious Epstein-Barr virus from a virus-carrier cell line. When certain bovine sera (BS) were used to culture P3HR-1 cells, the amount of infectious Epstein-Barr virus (EBV) released into the culture fluid was significantly less than that obtained when calf serum (CS) or fetal calf serum (FCS) was used. A direct dose-response relationship was noted between the concentration of BS and the degree of inhibition of virus production. Cell growth was not inhibited nor was virus infectivity neutralized by BS. Similarly, EBV infectivity was not reduced by spent medium containing BS. The inhibition of virus production by BS could be reversed by replacement of the serum supplement with CS. The formation of viral capsid antigen and of infectious virus within the cells was not inhibited by BS, whereas the amount of virus released was significantly decreased in comparison with culture media containing FCS or CS. These findings suggested the presence of a factor(s) in the BS which inhibited the release of EBV from the cell.


Establishment of two canine sarcoma cell lines: productive infection with feline leukemia virus. Two canine sarcoma cell lines (11028, 11031) were established in vitro and have been transferred 213 and 306 times, respectively, since 1970. These cell lines had a chromosome pattern consistent with their canine origin. Both cultures were infected with feline leukemia virus (FeLV), which caused a morphologic and karyotypic changes. The cells became rounded after infection with FeLV and both cultures showed the presence of a single, large, acrocentric chromosome considered to be a marker chromosome. All tumors were transplanted into newborn beagle pups, but only the 11028-FeLV formed metastatic tumors. No helper or focus-forming activity or virus particles were found in the uninfected cultures. Helper activity and virus were demonstrated in both sarcoma lines after infection with FeLV, though no focus-forming activity was noted. Helper activity of progeny virus could be assayed on either cat or dog embryo cells.


Effect of BCG on dimethylhydrazine induction of colon tumors in rats. The effect of BCG injection into the colon wall on the induction of colon tumors by 1,2-dimethylhydrazine dihydrochloride (DMH) was assessed in male rats. Persistent, generalized BCG infection followed the intracolonic injection of 6.7 times 10(6) organisms in otherwise untreated rats. In rats given DMH (30 mg/kg/wk) intragastrically for 5 weeks and then infected with BCG, colon tumors developed at the same rate and in the same incidence as in uninfected rats, but more tumors were mucinous adenocarcinomas and metastasized to the abdominal lymph nodes. In a few rats with large, established colon tumors, the injection of BCG into tumors induced no apparent change in them when examined at autopsy 1-22 weeks later.


Lymphocytes and leukemia viruses: tropism and transtropism of murine leukemia virus. The tropism of naturally occurring murine leukemia virus (MuLV) was investigated in short-term lymphocyte cultures. The tropism of MuLV was readily defined in fibroblast cultures, but not in lymphocyte cultures. Lymphocytes free of infectious MuLV could be infected across the tropism barrier by partially purified MuLv or by in vitro contact with MuLV-producing lymphocytes. Stimulation of lymphocytes was not required for this cross-infection and replication of MuLV. When cross-infected lymphocytes and was specifically associated with lymphocytes were stimulated in vitro by allogeneic cells, they facilitated MuLV infection of ordinarily non-permissive fibroblasts. This phenomenon (transtropism) required antigenically stimulated lymphocytes and was specifically associated with infection of the lymphocyte by MuLV across the tropism barrier. Thus in contrast with the resting lymphocyte, the transformed lymphocyte acquired the ability to disseminate infectious MuLV to nonpermissive cells. These findings suggest a novel relationship between lymphocytes and leukemia viruses. They indicate one mechanism whereby antigenic stimulation may enhance the development of virus-induced lymphoid neoplasms.


Feline oncornavirus-associated cell membrane antigen. IV. Antibody titers in cats with naturally occurring leukemia, lymphoma, and other diseases. Cats with naturally occurring leukemia and lymphoma had low or negative humoral antibody titers to the feline oncornavirus-associated cell membrane antigen (FOCMA). Geographic differences were seen in the relative frequencies of various forms of lymphoproliferative neoplasms. Lymphatic leukemia and thymic lymphoma were most common in Boston, whereas alimentary lymphoma was most frequent in Glasgow. No significant differences were found in geometric mean FOCMA antibody titers for the various forms of leukemia-lymphoma or for feline leukemia virus (FeLV)-positive as compared to FeLV-negative cats. Approximately 70% of 76 Boston cats with nonregenerative anemias were FeLV gs antigen (gsa) positive; this was similar to the percentage with leukemia-lymphoma from the same population that was positive. Fifty-five to 62% of the Boston cats with other infectious diseases, such as peritonitis and septicemia, were gsa positive. We postulate that this is due to a predisposition to infectious diseases by the immunosuppressive action of FeLV. Young cats from the Boston population that developed lymphoma, infectious peritonitis, and certain other diseases were more likely to be FeLV gsa positive than older cats with the same diseases.


Antitumor effects of antibody-diphtheria toxin conjugates. II. Immunotherapy with conjugates directed against tumor antigens induced by simian virus 40. Hamster and rabbit antibodies against antigens on the surface of simian virus 40 (SV40)-transformed hamster sarcoma or lymphoma cells were purified by immunoadsorption on SV40-transformed cells and conjugated to diphtheria toxin with glutaraldehyde. Treatment with a single dose of antibody-toxin conjugate partially protected hamsters concurrently challenged with 10(3) SV40-transformed sarcoma cells, as evidenced by a reduction in tumor incidence, an increase in tumor latency, and a prolongation of the life-spans of hamsters that developed tumors. Treatment of established sarcomas was ineffective, but repeated treatment with the conjugates induced complete regressions in 20-56% of hamsters bearing established transplants in the SV40-induced lymphoma.


Effect of host age, virus dose, and route of inoculation on tumor incidence, latency, and morphology in Syrian hamsters inoculated intravenously with oncogenic DNA simian virus 40. Three-week-old to 12-month-old male Syrian hamsters were inoculated iv with 10(8.5) median tissue culture infective dose of simian virus 40 (SV40). Three-week-old hamsters were similarly inoculated with aliquots of SV40 of progressively decreasing titers. The tumor incidence and, to a lesser extent, the tumor latency were directly dependent on the age of the animals at the time of virus exposure and on the dose of the virus. However, this age-dose dependence was not of the magnitude usually observed in hamsters inoculated with SV40 sc or im. Moreover, the wide morphologic spectrum of neoplasms induced, i.e., lymphosarcoma, reticulum cell sarcoma, and osteogenic sarcoma, by iv route of inoculation, contrasted sharply with the anaplastic and spindle-cell sarcomas which were the only types of malignant tumors resulting when other routes were used.