Central spherules in pulmonary corpora amylacea. Pulmonary corpora amylacea were found in 37 of 6,500 (0.6%) unselected autopsies. The patients had ranged in age from 48 to 87 years (average, 70). Among 1,000 corpora amylacea studied, there were 30 that contained centrally located, more or less intact, hollow spheres 15 to 20 mu in diameter. Morphologic considerations suggest that the spheres are derived from plant spores, and it is suggested that a possible origin is from inspired lycopodium spores, which are used as a dusting powder. Formation of pulmonary corpora amylacea as a reaction to lycopodium could account for the peculiar sporadic incidence of these bodies and their remarkable uniformity in size.


Recent advances in the use and indications of Ender's nailing. Ender's closed nailing was initially described for osteosynthesis of trochanteric fractures in elderly people. The authors performed about 500 cases, advanced their indications, and applied this technic to two particular types of lesions, segmental fractures of the femur including the neck and shaft, and metastatic fractures of upper femur. They discuss their indications, technical problems and results.


Keratopathy following retinal detachment surgery. Light microscopy, electron microscopy, and histochemical studies were carried out on the corneas of three patients in whom identical keratopathy developed following extensive retinal detachment surgery with silicone-oil injection. These studies revealed the presence of granular deposits in Bowman's layer. Histochemistry and electron-probe analysis showed these deposits to be calcium phosphate. No silicone was detected in the tissue.


Prevention of surface bacterial contamination of donor corneas. A simple method has been developed to reduce contamination in postmortem donor human eyes in anticipation of corneal transplantation. In vivo investigation of albino rabbits demonstrates that vigorous saline solution irrigation is extremely effective in decreasing the surface bacterial counts of the postmortem eye. In vitro and in vivo studies show that Neosporin kills bacteria at room temperature and further show that a tenfold increase in the thimerosal concentration of the Neosporin will kill fungus. Postmortem eyes contaminated by pathogenic organisms can be effectively cleaned by a combination of saline solution irrigation and the new Neosporin-thimerosal solution. No substantial damage of the donor tissue was noted by scanning electron microscopy. Human eyes cultured before this procedure were all contaminated, but after cleansing and immersion, no bacterial or fungal growth occurred.


"What's wrong with my child, doctor; do you think he is slow?". Intellectual handicap is the most frequent of the chronic handicapping conditions of childhood. It is important to make the diagnosis early without unnecessary delay, and also to refer on for special services as soon as possible. The family doctor's role in recognition and treatment of intellectual handicap, and parental reaction to this diagnosis in their children is discussed in this article.


Parathyroid identification by methylene blue infusion. A preoperative infusion of methylene blue was employed in 20 patients undergoing neck exploration for hyperparathyroidism. The dye was noted to stain adenomas and hyperplastic glands a deep purple-blue colour. Normal parathyroid tissue stained to a lesser extent or not at all. All unstained parathyroid tissue was normal histologically. Methylene blue infusion is a safe method of more rapidly identifying parathyroid tissue. Its preferential staining of abnormal parathyroid tissue can assist the surgeon in deciding the extent of his parathyroid excision.


Angiodysplasia of the small bowel: a method of intraoperative identification. Two cases of recurrent bleeding from the gastro-intestinal tract are described. The initial diagnosis and the identification of the lesions at laparotomy were difficult problems. In both cases the bleeding was due to a vascular lesion of the small bowel, referred to as angiodysplasia. The lesions were detected before operation by selective angiography and identified at laparotomy by a preoperative and an intraoperative injection of a vital stain through the catheter into the feeding vessels.


Long-term cultivation of functionally-active human foetal hepatocytes. Livers from human foetuses obtained from terminations of pregnancy by suction were used to establish long-term cultures of hepatocytes. Functional activities of the hepatocellular population in these cultures were monitored by immunoperoxidase staining during and up to the 90 days' life of the cultures. Intracellular localization of albumin and alpha-foetoprotien in cultures of the same age and from the same specimen indicate that syntheses of these two markers in foetal hepatocytes occur simultaneously. No qualitative differences were observed in the biosynthetic capacity of the cells maintained in tissue culture medium with and without serum. Fibroblast-like cells seen in the almost homogeneous population of hepatocytes did not stain for albumin and alpha-foetoprotein.


Morphological changes in the human conjunctival epithelium. I. In the normal elderly population. Forty-nine biopsies of bulbar conjunctiva from patients with no apparent conjunctival disease were studied by light and transmission electron microscopy. No significant morphological changes were recorded in those specimens from patients less than 79 years old. However, in the older age group the epithelium showed irregularities in thickness, a reduction in the goblet cell population, and in 25% the presence of 'hyaline bodies'. The epithelial irregularities consisted of mild superficial stratification, which was observed also in a few of the specimens from the younger age groups. The goblet cells appeared morphologically normal in all age groups. The significance of the hyaline bodies and their possible relationship to goblet cells is discussed.


Circular dichroism of beta turns in peptides and proteins. Circular dichroism (CD) spectra are reported for two groups of cyclic hexapeptides having beta turns whose geometry can be firmly established by X-ray crystallography and by NMR spectroscopy. One series contains the sequence L-Pro-D-Phe in the geometry of the classical type II beta turn, while the second group has the sequence D-Phe-L-Pro in the closely related geometry of the gramicidin S turn. CD data on the hydrogenated peptides show that in neither series do Cotton effects due to the aromatic phenylalanyl chromophore make a significant contribution to the spectra in the 195--240-nm region. In spite of the close geometric similarity of the beta turns of these two groups of peptides, their CD spectra are quite distinct. Furthermore, comparison of our data with the CD spectra of published models for beta-turn structures suggests that it may not be possible to characterize the contribution of all beta turns to the CD spectra of proteins by a single model curve. the CD spectra of model beta turns will be more useful in characterizing the folding of oligopeptides and sequence polypeptides, where a single type of turn is present.


The blockage of the electrical conductance in a pore-containing membrane by the n-alkanes. 1. In monooelein bilayers made highly conducting by the addition of a fixed amount of o-pyromellitylgramicidin, the membrane conductance has been shown to be strongly dependent on the chain length of the n-alkane with which the membrane is in equilibrium. Thus for n-hexadecane, the conductance is larger by approx. 10(4) times than it is for n-octane. This result is independent of whether the polypeptide is introduced via the aqueous or lipid phases. 2. The observed conductance variations have been accounted for in terms of a mechanism (outlined in earlier publications) which is based on the thickness and tension changes produced in bilayers by the adsorption of n-alkanes. Essentially quantitative agreement between theory and experiment is found.


Inhibition of activities of DNA polymerase alpha, beta, gamma, and reverse transcriptase of L1210 cells by phosphonoacetic acid. Phosphonoacetic acid has been shown to suppress replication of DNA tumor viruses by inhibiting the activity of virus-induced DNA polymerase and consequently viral DNA synthesis. We now have evidence to show that phosphonoacetic acid inhibits also the cellular DNA polymerases alpha, beta, and gamma of L1210 cells as well as reverse transcriptases of two type C viruses. Particularly, the DNA polymerase alpha is just as sensitive as the herpes virus induced DNA polymerase. The DNA polymerases beta and gamma required seven times more phosphonoacetic acid for a 50% inhibition of their activities. Phosphonoacetic acid inhibited the activities of the reverse transcriptase and terminal deoxyribonucleotidyltransferase only at higher concentrations. Kinetic analysis with the DNA polymerase alpha showed that the compound is a non-competitive inhibitor with respect to the substrates and uncompetitive inhibitor with the activated DNA template. Studies on time course of phosphonoacetic acid inhibition revealed that the compound is inhibitory even after the initiation of DNA synthesis. Phosphonoacetic acid also inhibited cell growth as well as the type C virus production; at concentrations above 50 microgram/ml, the inhibitory effect was more profound on the type C virus production than on cell growth.


Specificity of DNA base release by bleomycin. DNA was treated with bleomycin in the presence of Fe2+ and 2-mercaptoethanol under conditions where only a few percent of the bases were released. Release of all four bases was a linear function of bleomycin concentration, but the amount of thymine released was twice that of cytosine, 7 times that of adenine, and twelve times that of guanine. Unidentified minor products of thymine, of cytosine and of a purine were also released. Bromouracil did not sensitize DNA to bleomycin-induced breakage, and was released at the same rate as thymine.


Sequential passage of alpha2mu-globulin through the hepatic endoplasmic reticulum and Golgi apparatus during secretion. Studies on the synthesis and secretion of the sex-dependent urinary protein, alpha2mu-globulin, have been extended by establishing its sequential passage from the rough to the smooth endoplasmic reticulum and Golgi-rich fractions of the liver of adult male rats. After injection of 14C-labeled amino acids, the maximum radioactivity of alpha2mu occurred at 20 min in the rough, 25 min in the smooth microsomes and 30 or 35 min in the Golgi-rich fractions. Radioactive alpha2mu-globulin appeared in the bloodstream and kidneys after a lag of 20--25 min. Results indicate that alpha2mu-globulin follows a secretory pathway similar to that of serum albumin.


Bovine alpha-fetoprotein. Isolation and characterization. Bovine AFP was purified by ion exchange chromatograph on C.M. cellulose and DEAE Sephadex A-50, gel filtration and immunosorbent technique. AFP was homogeneous when studied by gel electrophoresis under non denaturing and denaturing conditions, by ultracentrifugation and by immunological methods. The following molecular data were obtained: 1. Sedimentation equilibrium indicated a molecular weight of 66,500 and sedimentation velocity gave s degrees 20, w = 4.71 S. A partial specific volume v = 0.737 ml g-1 was derived from density measurements. 2. From these data, a Stokes radius of 3.26 nm, a diffusion coefficient D20 w = 6.61 10(-7) cm2 sec-1 and a frictional ratio f/fo = 1.21 were calculated. 2. Sodium dodecylsulphate disc electrophoresis suggests a molecular weight of 67,000. 3. Gel filtration pointed to a molecular weight of 75,000. 4. Microheterogeneity of AFP was demonstrated by isoelectric focusing. The isoelectric point of the major component is 4.6. 5. The chemical composition was determined. AFP is a glycoprotein containing 7 per cent carbohydrate including 1.67 per cent hexoses, 2.38 per cent N-acetyl glucosamine and 1.8 per cent N-acetyl neuraminic acid.


Serial measurement of beta2 microglobulin, acute phase reactant proteins and the ESR in non-Hodgkin's lymphomas and chronic lymphocytic leukaemia. Long-term observation of non-Hodgkin's lymphomas indicates they can be sub-divided into two groups with respect to changes in the plasma proteins. The first group has acute phase reactant proteins raised during active disease and sometimes a raised B2m, whilst in remission the protein profile is normal. The second group is typified by a chronic elevation of B2m and ESR but has normal C-RP levels. Chronic lymphocytic leukaemia usually has a raised B2 m level and normal acute phase proteins, a subset with low B2 m is described.


[Comparative study of the myoid cells of human embryonal and adult thymus by the indirect immunofluorescence method]. The indirect immunofluorescence method showed that decreased (in comparison with myoid cells of adult human thymus) content of muscle antigens in the myoid elements of the embryonic organ caused a greater secretory activity of these elements at the early embryogenesis. Due to increased secretory activity of the myoid cells internal medium of the embryonic thymus contained more antigens common to the muscle tissue than the adult human thymus. The fact that during the ontogenesis the functional activity of the myoid cells correlated with the rate of lymphoid tissue formation favours a suggestion that heteroorganic antigens provide the thymus lymphocytes with information concerning the autoantigen structure necessary to induce natural immunological tolerance.


Amniotic fluid cell morphology in early antenatal prediction of abortion and low birth weight. The morphology of rapidly adherent (RA) amniotic fluid cells was examined in 201 pregnant women referred for amniocentesis because of two sequential high serum alpha-fetoprotein (AFP) concentrations. Out of 43 amniotic fluid samples containing increased amounts of AFP, 42 had neural or peritoneal cells predominating among the RA cells, the outcome being an infant with a neural-tube defect or exomphalos. In the other case with a raised amniotic fluid AFP concentration but only anterior placental cells the infant was normal. In 25 amniotic fluid samples containing normal amounts of AFP distinctive new patterns of RA cells were observed, termed fetal distress cells. These pregnancies resulted in five spontaneous abortions and 20 infants with birth weights under 2500 g. Fetal distress cells were not detected in any of the remaining 133 samples. One pregnancy was terminated because of a chromosomal abnormality, and there were seven twin pairs not recognised on ultrasonography before amniocentesis. The remaining 125 pregnancies went to term, resulting in infants with birth weights exceeding 2500 g. The results suggest that RA-cell morphology will prove to be of value in the early antenatal prediction of spontaneous abortion and low birth weight.


Changes in synaptic function induced by blockage of axonal transport in the rabbit optic pathway. This study was undertaken to elucidate the physiological significance of material involved in the rapid axonal transport. The effects of colchicine-induced inhibition of axonal transport in the retinal ganglion cells on the electrophysiological properties of the retrobulbar visual pathways were investigated in Albino rabbits. An impaired signal transmission to the contralateral visual cortex, superior colliculus and lateral geniculate body following flash light stimulation as well as direct optic nerve stimulation appeared 4--6 days after an intravitreous injection of 10--25 microgram colchicine. It was concluded that inhibition of the fast axonal transport within the retinal ganglion cells interferes with transsynaptic signal transmission from optic nerve terminals in the subcortical nuclei. This indicates a functional relationship between material supplied via the rapid phase of axonal transport and an unimpaired transsynaptic signal transmission, previously not revealed in the central nervous system of mammals.


5-azacytidine in acute leukemia.

5-azacytidine in acute leukemia. 101 patients with acute leukemia in relapse were treated with 5-azacytidine according to three schedules: Regimen A--300 mg/m2(day divided intravenously at 8 hour intervals for 5 days; Regimen B--750 mg/m2 as a single iv pulse dose administered at 2 to 3 weeks intervals; and Regimen C--300 mg/m2/day by continuous infusion daily for 5 days. Twelve patients achieved a complete remission (CR) and six achieved a partial remission (PR) for an overall 18% response rate. Of 78 patients receiving an adequate trial the response rate was 23%. An average of 1.5 courses and a median of 5 weeks were necessary to achieve a response. The median duration of CR patients was 21 weeks and for PR patients it was 5 weeks. Response rates were 24% for Regimen A, 0 for Regimen B, and 1 of 8 for Regimen C. The CR rate for AML and AMML was 13%. Two of eight AMoL patients achieved a CR. Only 2 of 23 ALL patients responded, one of whom achieved a CR. Toxicity included moderate to severe nausea and vomiting, diarrhea, stomatitis, skin rash, and prolonged myelosuppression. 5-azacytidine has significant activity in the acute nonlymphoblastic leukemias.


Germ cell tumors (II): VAB II in metastatic testicular cancer. Between June 1974 and January 1976, 50 patients with metastatic non-seminometous testicular carcinoma were treated with the VAB II protocol. The induction phase consisted of vinblastine (0.4 mg/kg) and actinomycin D (0.02 mg/kg) on day 1. Bleomycin (0.5 mg/kg) was given by continuous infusion for 7 days, and cis-diammine-dichloroplatinum (II) (DDP) (1 mg/kg) was given on day 8. A weekly maintenance of vinblastine and bleomycin, with actinomycin D and DDP on a rotating schedule was given followed by vinblastine, actinomycin D and chlorambucil every 3--4 weeks. Therapy was discontinued after 30--36 months of treatment in the face of continued remission. The response rate was 50% CR, 34% PR with 60% CR and 36% PR in previously untreated patients. Second-look surgery and excision of residual lesions was performed in selected cases. Alopecia, mucositis, nausea, and vomiting were universal. One patient died in the postsurgical period of toxicity from the combination of bleomycin and high concentrations of oxygen. There were seven instances of allergic reactions to DDP. Eleven of 25 complete responders and 4 partial or minor responders who underwent excision of stable disease remain alive from 19 to 35 months following start of therapy, 12 of them without evidence of disease.


Serum alpha-fetoprotein and human chorionic gonadotropin as markers for the effect of postoperative radiation therapy and/or chemotherapy in testicular cancer. Human alpha1-fetoprotein (AFP) and chroiongonadotropin (hCG) have been studied in 67 patients with malignant germ cell neoplasia of the testis admitted to the Radium Centre, Aarhus Municipal Hospital within an 18 month period. In 34 patients with nonseminoma, AFP was elevated in 35% of cases and hCG in 38% of cases. Nonseminoma patients who presented with or developed distant metastases showed a significantly higher incidence of elevated tumor markers than patients with local or regional disease. In patients with distant metastases monitoring of AFP and/or hCG was a most sensitive and reliable indicator of disease activity. In the management of such patients these tests are as important as clinical and x-ray examinations.


Dietary lipotropes, hepatic microsomal mixed-function oxidase activities, and in vivo covalent binding of aflatoxin B1 in rats. Weanling male Sprague-Dawley rats were fed either a nutritionally complete synthetic diet (Diet 1) or a diet marginally deficient in choline and methionine, and lacking folacin (lipotrope deficient, Diet 2) to determine the role of hepatic mixed-function oxidase metabolism of aflatoxin B1 (AFB1) in the Diet 2-induced enhancement of AFB1 hepatocarcinogenesis previously reported. Hepatic microsomal mixed-function oxidase activities, as assayed by ethylmorphine N-demethylation, ethoxycoumarin O-dealkylation, cytochrome c reduction, AFB1 metabolism, and cytochrome P-450 content, were all depressed by Diet 2. Furthermore, the proportion of an i.p. dose of AFB (1 mg/kg) that became covalently bonded to DNA and RNA was similarly reduced when measured 6 hr after administration. The formation of AFB1-protein adducts was not influenced by dietary treatment. The depression of DNA and RNA adduct formation in the Diet 2 animals was probably related to the lower mixed-function oxidase activities and not to an alteration of glutathione levels, which remained unchanged by dietary treatment. These results suggest that the marginally lipotrope-deficient diet does not enhance tumor formation through an increased microsomal activation of AFB1. Alternative hypotheses without data are suggested.


Isolation and partial characterization of radioiodinated myeloblastic leukemia-associated cell surface glycoprotein antigen. Peripheral blood myeloblasts from five patients with acute myeloblastic leukemia and peripheral remission leukocytes from two of these patients were radiolabeled by the lactoperoxidase-catalyzed surface radioiodination technique and incubated in a nutrient medium at 37 degrees. Radioactive materials shed from viable cells into the supernatant at 24 hr were purified by gel filtration and by DEAE-cellulose chromatography. The radiolabeled leukemic cells shed relatively few molecular species into the culture medium. The DEAE-cellulose eluate usually contained one major peak in which radioactivity and protein levels were coincident; the molecular weight of this compound was 350,000 to 400,000, and it contained carbohydrate as well as protein. Glycoprotein shed from leukemic cells was specifically reactive in a coprecipitation assay with defined antimyeloblast alloantisera obtained from leukemic patients receiving immunotherapy. No reaction was seen with antisera directed against HLA or B-cell antigens. Material shed from remission cells did not coprecipitate with antileukemic antisera. The isolation of radioactively labeled antigen derived from myeloblasts may ultimately allow the monitoring of human antigen levels in leukemic blood by radioimmunoassay.


Differential control of synergistic effect with polyene macrolide antibiotics upon Chinese hamster cells in vitro. An amphotericin B-resistant cell (AMBR-1), which was isolated from aneuploid Chinese hamster cells (V79), was found to show much higher resistance than the parent V79 cells to other polyene antibiotics, such as pentamycin and filipin. To obtain the 50 to 60% inhibition of the control protein synthesis activity by a synergistic combination of fusidic acid and amphotericin B, 50 microgram fusidic acid per ml were combined with 10 microgram amphotericin B in V79 cells, whereas in AMBR cells 50 microgram fusidic acid per ml were combined with 100 microgram polyene antibiotic per ml. Bleomycin (10 microgram/ml), which alone did not affect cellular DNA synthesis, inhibited DNA synthesis of V79 cells by more than 90% of the control activity when combined with only 1 microgram pentamycin per ml, whereas a similar extent of inhibition in AMBR cells was observed by combination with more than 5 microgram pentamycin per ml.


Computerized nuclear morphometry as an objective method for characterizing human cancer cell populations. A new method for measuring differences in nuclear detail in chrome alum gallocyanin-stained nuclei of cells from human breast cancers was compared with conventional subjective grading and classification systems. The new method, termed computerized nuclear morphometry (CNM), gives a multivariate numerical score that correlates well with nuclear atypia and gives a higher reproducibility of classification than do subjective observations with conventional histological preparations. When 100 individual nuclei from each of 137 breast cancers were examined by CNM, there was a broad CNM score variation between patients but a good reproducibility for each tumor. When different parts of the same tumor were sampled, there was good reproducibility between samples, indicating that some breast cancers at least are "geometrically monoclonal." When these cancers were compared by the grading systems of WHO and Black, correlations of 0.43 and 0.48, respectively, were found. There was a poor correlation between CNM and classifications of tumor type, but in general there were high values for CNM in medullary tumors and low values in mucous tumors. Correlations between CNM and tumor progression and prognosis await future study of patients participating in the study.


Protein synthesis by rat transplantable yolk sac tumor and its relation to the cytosol levels of translatable messenger RNA's. alpha-Fetoprotein (AFP) was shown to be the major secretory protein produced in vitro by normal rat yolk sacs. While not so active, AFP production was also detected in the transplantable tumors derived from normal yolk sacs. The major secretory protein synthesized by the tumor cells had a molecular weight of 40,000 and was reactive with an anti-rat albumin antibody. The functional messenger RNA's coding for these proteins were quantitated by translation in a cell-free system derived from wheat germ followed by specific immunoprecipitation of the newly synthesized peptides. The overall template activity of the RNA prepared from the normal yolk sacs and yolk sac tumor cells was virtually identical. The cytosol RNA prepared from the normal yolk sacs was approximately 12 times more active than that from the tumor cells in directing the synthesis of AFP. The presence of the cytosol RNA prepared from the tumor cells was required for the synthesis of proteins immunoprecipitable with the antialbumin antibody. These results suggest that the changes in AFP and albumin synthesis can be accounted for by a corresponding change in the levels of functional messenger RNA's coding for these proteins.


Quantitative description of cell cycle kinetics under chemotherapy utilizing flow cytometry. A discrete time cell cycle kinetics model is developed to account for the effects of cytotoxic chemotherapy, particularly including the existence of cells destined to die. A model structure is determined from related experiments, leaving key parameter values undetermined. These values are found by determining the best least squares fit of the predicted to the observed DNA distribution data at a series of time intervals. The numerical methods include separable least squares, linear inequality constrained least squares and the Gauss--Newton method. This approach is applied to an experiment in which the Ehrlich ascites tumour was given a single dose of bleomycin. The results include several different parameters, including the age response function and a time series of cell age and DNA distributions, which can be used as a basis for further treatment.


The hypothalamic neurosecretory systems of the Japanese quail as revealed by retrograde transport of horseradish peroxidase. Within 24-48 h after injection of horseradish peroxidase (HRP) into the neural lobe or into the median eminence of adult Japanese quail dense accumulations of its reaction product (HRP-RP) can be demonstrated in axons of the hypothalamo-hypophysial tract and in the magnocellular neurosecretory perikarya of the supraoptic and paraventricular nuclei as well as in scattered neurons of the accessory hypothalamic neurosecretory nuclei. The HRP-RP-containing nerve fibers, which are beaded in appearance, occur prominently in the internal zone of the median eminence. They turn dorsally at its anterior border to become widely distributed in the retrochiasmatic region and extended to the paraventricular, supraoptic areas. These observations confirm more directly conclusions drawn earlier from Gomori-type preparations and from immunologic demonstration of arginine vasotocin, mesotocin and neurophysin. HRP-RP was also found in perikarya of parvocellular secretory neurons in the infundibular nucleus 48 h after injection of HRP into the median eminence but not after injection into the pars nervosa. This provides direct evidence that a conspicuous component of the tubero-infundibular tract is formed by axons of tuberal neurons that originate from the infundibular nucleus and pass directly into the median eminence.


Effects of benzo(a)pyrene adducts of DNA synthesis in vitro. Two diol epoxides of benzo(a)pyrene (BP), and benzo(a)pyrene 4,5-oxide, have been used to make adducts in the homopolymers polyribocytidylic acid, (rC); polyriboadenylic acid (rA), polydeoxycytidylic acid (dC) and polydeoxyadenylic acid (dA). With appropriate oligomers as primers these modified and unmodified polynucleotides were used as templates for DNA synthesis with avian myeloblastosis virus DNA polymerase (AMV) or E. coli Pol I DNA polymerase. We have found that: (1) the size of the DNA product is not markedly decreased by the presence of these these polycyclic aromatic hydrocarbon adducts in the templates; (2) the presence of adducts does not lead to increased incorporation of erroneous bases. These results, supported by kinetic data, suggest that these polymerases can bypass a site containing an adduct on the template without leaving a gap or causing misincorporation of a base and they imply that mutagenesis by BP may not be attributable to either of these mechanisms.


[Abortive effect of autologous anti-alpha-fetoprotein antibodies in the rat]. Female Commentry Rats 80--85 days old, were immunized by 5 intramuscular injections of Mouse alphafetoprotein (AFP) and then mated. After fertilization a supplementary injections was administered. The animals were bled at different times and killed immediately after the last bleeding on the 19 or 20th day of gestation. Titers of AFP and of autologous anti-AFP antibodies in the maternal blood were determined as well as the AFP concentration in the pooled amniotic fluids from live embryos of each litter. Compared to non-immunized control series, the total of live and dead embryos per litter in animals immunized with Mouse AFP showed no difference. However, the number of live embryos was on the average 50% lower than that in the control series. The serum titers of AFP and of antibodies to autologous AFP in immunized pregnant Rats bearing dead embryos decreased concomitantly with the number of live embryos. The results reported herein demonstrate that the presence of anti-autologous AFP antibodies in pregnant Rats correlates with the interruption of development in a significant proportions of embryos. This suggests that certain spontaneous abortions in the Rat and perhaps in other mammals can be explained by the rupture of immunological tolerance to autologous AFP.


Polystyrene tube immunoradiometric assay for human alpha1-fetoprotein, and its use for mass screening. We describe a two-site immunoradiometric assay for human alpha1-fetoprotein, with use of antibody-coated polystyrene tubes as solid phase. The sensitivity, precision, and simplicity of this system make it eminently suitable for mass-screening purposes. We currently use it for neonatal detection of hereditary tyrosinemia in the Province of Quebec; measurements are made on blood spotted and dried on paper. This system could be well suited for other mass surveys, such as prenatal screenings for fetal abnormalities.


Serum elastase inhibitors in cardio-vascular diseases. Serum elastase inhibiting capacity was measured in three groups: 150 control subjects, 38 hospitalized children without cardiovacular diseases and 202 hospitalized patients suffering from cardiovascular diseases. The values obtained were 53% in control adult subjects and 79% (range 45--90%) in the hospitalized patient groups. The highest levels were recorded at the acute phase of myocardial infarction. The levels of alpha 1-antitrypsin (alpha1-AT) and alpha 2-macroglobulin (alpha2-M) were determined by radial immunodiffusion technique for various levels of inhibitory power. No correlation was found between the inhibitory power levels and the alpha1-AT and alpha2-M levels. This study suggests that other proteins may intervene in the inhibition process of elastolysis.


Human alpha2-macroglobulin and its antitrypsic and antithrombin activities in serum and plasma. alpha2-Macroglobulin level, trypsin protein esterase and progressive antithrombin activities were measured in normal and nephrotic sera and plasma. Trypsin protein esterase activity was proportional to the alpha2-macroglobulin concentration in serum and plasma from both normal and nephrotic patients. The results were different, however, with progressive antithrombin activity: in normal plasma, antithrombin III is the main thrombin inhibitor, then alpha2-macroglobulin and alpha1-antitrypsin, whereas in nephrotic syndrome patients, alpha2-macroglobulin is the main thrombin inhibitor.


A rapid and inexpensive laser nephelometric assay for plasma pregnancy specific beta1-glycoprotein levels. A laser neophelometric method for the measurement of maternal plasma pregnancy specific beta1-glycoprotein levels is described. Fifty plasma specimens can be analysed within four hours at a low reagent cost. The assay has been used to determine the range of plasma pregnancy specific beta1-glycoprotein concentrations in 212 normal pregnancies of from 31 to 41 weeks gestation.


The fatty acid composition of skin and plasma lipids in Refsum's disease. The fatty acid composition of the skin and plasma lipids is described in a patient severely affected by Refsum's disease whose plasma phytanic acid concentration was very high (3.1 mg/ml). In the epidermal lipids, especially in the phospholipid fraction, phytanic acid tended to replace linoleic acid and to some extent arachidonic acid. In some respects the changes in the skin in Refsum's disease resemble those of essential fatty acid deficiency.


Inhibition of viral reverse transcriptase and human sperm DNA polymerase by anti-sperm antibodies. The IgG fraction of serum from a rabbit immunized with detergent-prepared human sperm nuclei inhibited the DNA polymerase activities in human sperm and seminal fluid as well as the partially purified reverse transcriptase of the baboon endogenous type-C retrovirus (BEV). The analogous enzymes from lysates of oncogenic type-C viruses was unaffected. IgG from the serum of individual partners from infertile marriages similarly inhibited both purified BEV reverse transcriptase and human sperm DNA polymerase, but not a DNA polymerase isolated from human prostatic fluid. The data suggest that BEV reverse transcriptase and the human sperm DNA polymerase are antigenically related. Furthermore, the sperm appears to be auto-antigenic and the antibodies thus formed may be capable of interfering with reproductive success.


Electron microscopic detection of phleomycin during different waves of replication in regenerating skeletal muscle of mice following staining with mercuric chloride. Phleomycin has devastating effects on regeneration of skeletal muscle when applied during an early wave of replication in the pre-myotube (new fibre) period of the reaction. The effect cannot be explained from the contribution of dividing cells to myotubes. Phleomycin's effects on regeneration are much less severe when the challenge coincides with a later wave of pre-myotube proliferation, effects that can be explained from the contribution of such cells to new muscle. An attempt has been made, by means of electron microscopy, to explain how phleomycin distributes early versus late wave cells, using a mercury substitution stain to detect the antibiotic. Cells of the two periods showed conspicuous differences in the staining characteristics of their chromatin. Positive staining reactions outside the nucleus were confined mainly to ribosomes. Exceptions included materials in transit across nuclear and cytoplasmic membranes.


[Suggestions on the control and stimulation of cellular immunity in cancer]. Seventy patients suffering from cancer have been treated since 1969 with intracutaneous injections of autogenous carcinoma cells. This should stimulate cellular immunity through the skin. A stimulation and reactivation of cellular immunity seems to be possible. The cellular reaction at the injection site supplied useful hints for the evaluation of the state of immunity. The possibility of improving cellular resistance through immune gamma globulin in pigs treated with carcinoma cells is discussed.


The very low birthweight infant: impact on parents during the pre-school years. This paper presents follow-up data on 75 families who had children born between 1970 and 1971 at birthweights under 1501 g (very low birthweight: VLBW) and a comparison group of 55 families who had children born at the same time but at birthweights over 2500 g. An interview done when the children were 3--5 yr old revealed that parents with a VLBW child gave a less favourable evaluation of their child in 2 of 7 areas considered: development and play. These differences of evaluation were caused by a subgroup of 22 VLBW children with neurological and/or intellectual defects. No evidence came forth to indicate that the experience of having a VLBW child had significant or continuing impact on the family. An absence of serious social problems such as child abuse or the child's failure to thrive were noted.


Behavioral ototoxicology.

Behavioral ototoxicology. Methods for the evaluation in experimental animals of toxic substances that produce hearing impairment are described. In the experiments reported here, animals were trained by positive reinforcement operant conditioning procedures so that their hearing could be examined by behavioral means. When normal hearing was established, aminoglycosidic antibiotics (kanamycin and dihydrostreptomycin) were given daily and hearing tests administered in order that the course of hearing loss could be closely followed. Initial loss of sensitivity to the high frequencies always progressed in time to impairment at the low frequencies, and these changes in hearing were correlated with a loss of receptor cells in the inner ear which started in the basal region of the cochlea and advanced toward the apex. Although such behavioral procedures are moderately expensive to instrument and relatively time-consuming to apply, they are shown to yield valid quantitative measures of hearing. Further, they provide for reliable early detection of the toxic process and a measure of behavioral impairment that can be precisely related to the histopathological changes that occur simultaneously in the inner ear and auditory nerve.


Experimental allergic encephalitis: study of cellular immunity during disease suppression. Administration in complete Freund's adjuvant of encephalitogenic protein (EP), derived from central nervous tissue to guinea pigs, regularly results in the development of experimental allergic encephalitis (EAE) which leads to the death of the animals. Administration of EP in incomplete Freund's adjuvant at an appropriate time will completely suppress the clinical development of disease. Results reported herein show that animals receiving suppressive injections of EP for 7 days show depression of lymphocyte DNA synthesis and macrophage migration inhibition, but not of skin reactivity, in response to EP immediately following the injections, and subsequently show recovery of lymphocyte reactivity but do not develop clinical manifestations of EAE. Humoral or other factors may prevent the development of disease in these animals. Guinea pigs receiving injections of EP for 14 days show profound and prolonged depression of lymphocyte reactivity to EP and macrophage migration inhibition. Possible mechanisms for these results include a diminished number or function of reactive cells or activity of a population of cells with the capacity to suppress cellular immune responses. Nonspecific suppression of reactivity to an unrelated antigen during the suppressive injections was not observed.


The in vitro immune response to a T-independent antigen. I. The effect of macrophages and 2-mercaptoethanol. The in vitro immune response of murine spleen cells to the T cell-independent antigen, 2,4-dinitrophenyl-beta-alanyl-glycyl-glycyl-N(2-aminoethyl)carbamyl-methylated Ficoll (DAGG-Ficoll), requires the presence of macrophages, or 24-hr culture supernates from peritoneal cells. Complete reconstitution of the response by supernates, however, is dependent on the addition of 2-mercapto-ethanol. Antigen is required to fully express the in vitro immune response, but is not required in cultures of peritoneal cells in order to obtain active supernates. Active supernates can be obtained from allogeneic or syngeneic peritoneal cell cultures. The enhancing effects of 2-mercaptoethanol and peritoneal cell supernates appear to be synergistic nonspecific events directed toward lymphocyte activation and differentiation.


Specificities in natural cell-mediated cytotoxicity against lymphoblastoid cell lines. I. Selective inhibition by cross-competition. The apparent nonselective effects of natural cell-mediated cytotoxicity tested directly on different lymphoblastoid cell targets were found to be quite specific in the cross-competition assay. The specificity was detected through inhibition of cytotoxicity by competitor cells sharing common specificities with the target cells. Cross-competition tests were performed employing eight lymphoblastoid lines including four T and four B cells. Selective inhibition observed between lymphoblastoid cell lines indicated that T and B cell lines were more antigenically similar within each group than between them. The target antigens that distinguish T cell from B cell lines have been tentatively called TA-T and TA-B.


Effects of biotin deficiency on serum proteins and plasma amino acids. In biotin-deficient rats, a decrease of total proteins, attributable to a decrease of albumin and alpha1-globulin fractions, a decrease of the pre-beta-lipoproteins and an increase of the alpha-lipoproteins, was observed, together with a rise of total amino acids. Such a situation  may be related to the influence of biotin on the synthesis of RNA and proteins.


Effect of tannic acid on the Ehrlich ascites tumor cells. Tannic acid was used to fix and stain Ehrlich ascites tumor cells in suspension. An increase in surface microvilli and cytoplasmic blebs in the tumor cells was observed. The mechanism of tannic acid induced surface morphological changes in tumor cells and the formation of a precipitate of protein-tannic acid-heavy metal complex are discussed.


Light microscopy as an aid in predicting ureaplasma infection in human semen. The potential role of Ureaplasma urealyticum in human infertility make it desirable to screen barren couples for this infection. Semen specimens from 96 consecutive patients were evaluated in our clinic. Microbiologic results were correlated with the percentage of coiled and fuzzy tails in seminal cytology. These features were then used in a double-blind study to predict Ureaplasma infection in another group of 100 randomly selected patients. It was possible to predict the presence or absence of Ureaplasma prior to laboratory culture in 70% of specimens. False-positive diagnoses were made in 19%. The diagnosis was false-negative in 11%.


Stimulation of cyclic adenosine 3':5'-monophosphate accumulation in human testes in vitro by luteinizing hormone, follicle-stimulating hormone, and prostaglandins. The gonadotropins luteinizing hormone (LH) and follicle-stimulating hormone (FSH) stimulate cyclic adenosine 3':5'-monophosphate (cyclic AMP) accumulation in human testicular incubates. The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine acts synergistically with LH and FSH to enhance cyclic AMP accumulation. These studies indicate that cyclic AMP accumulation may be involved in the mechanism of action of LH and FSH in the human testes, as has been proposed for rat testes. The prostaglandins (PG PGE1, PGE2, PGA1, and PGF2alpha stimulate cyclic AMP levels at 10(-4) M in human testes. The E type prostaglandins are the most potent. They induce half-maximal stimulation of cyclic AMP at 7 x 10(-7) M.


[Effect of iodobenzoteph and cytostasan on several immune responses in animals]. Tests staged on line-bred mice immunized with sheep erythrocytes brought evidence that cystostasan and iodobenzoteph cut down the number of antibody-forming cells both with the primary and secondary immune response. The maximal effect becomes manifest following introduction of the drugs on the 1--2nd day after immunization of the animals. Iodobenzoteph brings down the number of hemopoietic stem cells colonies while cytostosan leaves them unchanged.


Regulation of allotype expression in heterozygous rabbits. I. Concomitant modulation of cell surface allotypes on peripheral blood lymphocytes from b4b6 rabbits. Treatment of b4b6 rabbit peripheral blood lymphocytes with b5b5 anti-b4 antibodies at 4 degrees resulted in the modulation (disappearance) ob b4 and b6 cell surface allotype after subsequent incubation in serum-free medium for 1 h at 37 degrees. A clear dose dependence on the sensitizing anti-b4 antibody was observed. Similarly, b5b5 anti-b6 treatment demonstrated a dose dependence for b6 modulation and a threshold dose effect for b4 comodulation. Cells which formed rosettes with anti-b4-coupled SRBC (anti-b4 direct antiglobulin (DAG) rosettes) also demonstrated concomitant modulation of b4 and b6 allotype when incubated at 37 degrees. When cells formed anti-b6 DAG rosettes, subsequent b6 modulation could also be demonstrated, but no b4 comodulation occurred. Concomitant modulation did not occur when cells were incubated with anti-allotype antibodies at 37 degrees. Blocking studies disclosed that the two allotypes are not contiguous in the membrane since uptake of one antiallotype antibody did not block the uptake of another at 4 degrees. We therefore propose that concomitant modulation might occur during a process similar to patch formation.


Regulation of allotype expression in heterozygous rabbits. II. Concomitant suppression of b4 and b6 allotypes in the same cell. Cells from heterozygous b4b6 rabbits were treated at 4 degrees with anti-b4 or anti-b6 antibodies and then warmed at 37 degrees. A disappearance of both b4 and b6 allotypes (concomitant modulation) ensued. When cells which had undergone extensive comodulation were cultured overnight we noted that those cells were unable to re-express either allotype at pre-modulation levels. This suppression was likely linked to the initial events which culminated in comodulation. Those cells were not further suppressible when suppressive antibodies were added to the cultures whereas cell cultures which had undergone little or no previous modulation or comodulation were readily suppressed for both allotypes after anti-allotype antibodies had been added to the cultures overnight (concomitant suppression). This indicated that in vitro suppression of allotype may depend on cell surface allotype being present at a sufficiently high density. We present data which show that events at the cell surface may play a role in the regulation of cell surface allotype expression and propose that concomitant suppression may have bearing on cellular mechanisms which control allotype expression and also allotype suppression.


Effects of proteolytic enzymes and neuraminidase on the I and i erythrocyte antigen sites. Quantitative and thermodynamic studies. The effects of neuraminidase and proteolytic enzymes on I and i reactivities was studied with I and i adult red cells, using radioimmunological methods. An enhanced reactivity after enzyme treatment is not exclusively due to a membrane charge reduction. The increase in site numbers and association constants bring about the gain of the cold agglutinin fixation. The release of N-acetylneuraminic acid residues gradually increases the I antigen site density of I red cells and the i site density of i red cells. Similar behaviour was observed after proteolytic enzyme treatment with papain, bromelin or ficin. The proteolytic treatment of I erythrocyte reveals underlying i receptors on these cells. Following membrane glycoprotein chain removal, anti-i antibodies are specifically fixed on I erythrocytes. The accessibility to antibodies of the determinants responsible for I and i erythrocyte activities was influenced significantly by steric hindrance factors. While N-acetylneuraminic acid release increased antibody affinities for the antigenic receptor, the removal of glycopeptide chains greatly diminished steric hindrance and brought about higher affinity constants. After enzyme treatment, the antigenic structures become more homogeneous in their reaction with antibodies. The heterogeneity of binding constants observed with antigenic determinants of non-treated erythrocytes is probably due to the wide range of spatial distribution of these receptors within the membrane.


Ontogeny of B lymphocytes in mice. Quantification of surface membrane immunoglobulins by immunoperoxidase assay. Surface membrane immunoglobulins (SmIg) of splenic lymphocytes were investigated by immunoperoxidase assay in newborn Swiss mice, their mothers and adult controls. The mean number of SmIg + cells in adult mice was 45-8% and the mean number of antigenic sites per positive cells was 108,500. In newborn mice during the first 7 days of life, values for both parameters were low (24-25--28-5% and 38,000-45,773 respectively) and began to rise after the 10th day to reach adult levels between the ages of 2 and 3 weeks. A peak above adult levels for the mean number of sites per cell was observed on day 21 with a subsequent drop to adult levels on day 28. Post-partum females were found to have consistently fewer sites per positive cell (69,000-86,600) during the 14 days following delivery than their adult control counterparts, though the difference was not statistically significant.


Primary in vitro sensitization of virus specific cytotoxic T lymphocytes. CBA mouse-derived splenic lymphocytes treated with either beta-priopionlactone-inactivated or u.v. light-inactivated parainfluenza (sendai) virus stimulated in vitro unprimed syngeneic T-lymphocytes to differentiate into cytotoxic T-lymphocytes (CTL). The CTL generated were virus specific and H-2 restricted. For optimal CTL responses to be induced (i) a critical treatment of stimulator cells, (ii) an optimal ratio of responder to stimulator cells and (iii) an in vitro incubation period of 5 days was required. The in vitro system for the induction of primary virus-specific CTL responses may be useful to analyse the sensitization phase of H-2 restricted virus-specific CTL.


[Effect of various plasma expanders on blood group serological study results]. Plasma expanders can disturb in serological blood typing tests to varying amounts. In order to avoid possibly fatal errors in index reading, it is, therefore, necessary when demanding this type of examination to inform about any infusion of plasma expanders carried through before the blood test and to take it into consideration.


Developmental changes in carbohydrate moiety of human alpha-fetoprotein. Human AFP purified from fetal serum  and amniotic fluid was separated into three different variants by chromatography on concanavalin A insolubilized on Sepharose (Con A--Sepharose). The three variants were indistinguishable in immunodiffusion and radioimmunoassay. Sera from patients with yolk-sac tumor and amniotic fluid from early pregnancy were found to contain a high proportion (15-45%) of AFP which does not bind to Con A, while AFP in fetal and newborn sera, and in amniotic fluid from late pregnancy, contained less (2-6%) of this variant. The use of a large excess of Con A--Sepharose and the fact that the non-bound AFP consistently eluted as non-bound in rechromatography showed that this AFP is non-reactive with Con A. Fractionation of radiolabelled AFP from cord serum in a mixture with amniotic fluid verified the difference in the amount of the Con-A nonreactive variant in AFP from these two sources. These results suggest that AFP synthesized by the yolk-sac tissue and by the liver are glycosylated differently. The variant may prove to be diagnostically useful.


Longitudinal study of Epstein-Barr virus antibody titers and excretion in pediatric patients with Hodgkin's disease. Nineteen pediatric patients with Hodgkin's disease (HD) who had experienced primary Epstein-Barr virus (EBV) before, or in one case after, diagnosis, were studied longitudinally for changes in the titers and spectra of EBV-related antibodies, excretion of EBV into the oropharynx, the number of EBV-carrying lymphoid cells in the peripheral blood, and clinical signs and symptoms suggestive of reactivation of the latent virus. The incidence and geometric mean titers of IgG antibodies to viral capsid antigen (VCA) in the HD patients at the time of diagnosis and in the controls were similar. The anti-VCA titers of the patients rose above control levels during and after therapy and remained elevated for up to 7 years of observation. At no time were heterophil or VCA-specific IgM antibodies detected. Antibodies to EBV-induced early antigens were more common in patients (ultimately 80%) than in controls (9%). In contrast, antibody levels to EBV-associated nuclear antigen were disproportionally low in the patients. Excretion of EBV was noted at increased frequency in the patients but the number of circulating, EBV-carrying lymphoid cells was the same as in controls. No discrete clinical syndrome was associated with rising antibody titers or viral excretion. While these results are best explained by a presumed reactivation of the persistent EBV infection by immunosuppressive effects of HD or its therapy, they have not provided direct evidence for this suggestion.


Vascular permeability increasing action of the saline extract of Dictyocaulus filaria--a preliminary report. The cutaneous hypersensitivity reaction in sheep infected with Dictyocaulus filaria was investigated. The saline extract of the worms increased the cutaneous capillary permeability in both the infected and uninfected control animals. Similar increase in the permeability was also observed in uninfected healthy rabbits. The results suggest that the vascular permeability increasing action of the worm extract is, partly, due to histamine release.


Study of Epstein-Barr virus-determined nuclear antigen (EBNA) by chromatography on fixed cell nuclei. Low ionic strength (50 to 100 mM NaCl) and pH 6.0 were found to be optimal conditions for in vitro conversion of Epstein-Barr virus (EBV)-determined nuclear antigen (EBNA)-negative nuclei to EBNA-positive nuclei by addition of the complement-fixing (CF) antigen extracted from Raji cells. In vitro conversion of nuclei to EBNA-positively was sensitive to DNase but not to RNase treatment. This suggests that nuclear DNA is a specific target substance to which EBV-CF antigen binds. If nuclei were fixed with methanol/acetic acid and subsequently treated with 0.6 M NaCl, EBNA could be eluted from in vitro-converted Ramos nuclei with 0.3 and 0.4 M NaCl. The same conditions were also found to be optimal for the adsorption and elution of EBV-CF antigen in DNA-cellulose chromatography. This indicates that the DNA-binding properties of EBNA antigen can be studied by "chromatography" on fixed nuclei followed by the ACIF test. The obvious advantages of this method over chromatography on DNA-cellulose are its simplicity, the possibility of testing many samples in one experiment and, especially, the use of minimal amounts of material. Significant differences in elution patterns for EBNA were found when nuclei derived from different cell lines (Ramos, Raji, and P3HR-1) were converted in vitro to EBNA-positivity. EBNA is eluted from in vitro-converted nuclei of EBV genome-positive P3HR-1 cells at an almost 0.1 M higher concentration of NaCl than is necesssary for a similar degree of elution from nuclei of EBV genome-negative Ramos cells.


Frequent elevation of tissue polypeptide antigen in the sera of workers exposed to bladder carcinogens. A group of 108 male workers previously exposed to bladder carcinogens and a group of 26 patients with carcinoma of the bladder differed significantly from a control group comprising 63 healthy male donors with respect to their levels of tissue polypeptide antigen (TPA). The risk group exhibited 4-6 times more frequent elevations of TPA than did the group of healthy donors.


Surface membrane changes of T cells induced by syngeneic tumour cells. I. Formation and uptake of complexes of Ig and tumour antigens by T cells. A large dose of tumour cells (10(7)) from two different tumours produced a significant increase of Ig-bearing spleen cells 6 h after intraperitoneal administration in syngeneic mice, like conventional antigens previously tested. Incubation of normal spleen cells in serum taken 6 h after administration of tumour cells reproduced in vitro the changes observed in the spleen cell population of the serum donors, demonstrating the presence of a cytophilic Ig, which was shown to be taken up by T cells. Serum collected 6 h after tumour cell inoculation contains also a 4S factor which can generate in vitro cytophilic Ig for T cells in the presence of a foreign soluble protein or tumour antigen. Extracts obtained from tumours cells, which were shown to contain tumour antigens, were labelled with 125I and mixed with the 4S factor and normal 7S Ig. Upon fractionation of the mixture, high molecular weight material containing radioactivity and 7S Ig were eluted in the void volume, well ahead of the position of both the tumour cell preparation and the 7S Ig. This material contained Ig cytophilic for splenic T cells and it is likely that it represents cytophilic complexes of Ig and tumour antigens. It is postulated that the cytophilic Ig detected in the serum of animals 6 h after injection of a large number of tumour cells represents cytophilic complexes of Ig and tumour antigens formed through the mediation of a soluble factor.


Biochemical studies on RNA tumor virus information and its transmission in B16 murine melanoma. The mouse melanoma B16 contains particles encapsulating a high molecular weight RNA of 60--70S size associated with a reverse transcriptase. The particles possess a density of 1.14--1.18 g/cm3. The RNA shares sequences with the 70S RNAs of several mammalian C-type RNA tumor viruses. The nuclear DNA of the mouse melanoma B16 possesses particle-related sequences not present in the genome of normal C57BL mice.


Expression of human mammary epithelial (HME) antigens in primary cultures of normal and abnormal breast tissue. The expression of human mammary epithelial (HME) antigens was studied in primary cell cultures of normal and abnormal breast tissue and of tissues from other organs. By indirect immunofluorescence it was shown that heterologous anti-HME bound to the cell surface of both suspension and monolayer cultures of human breast epithelial cells from normal breast tissue, normal breast fluid, non-malignant atypical lesions of the female breast, gynecomastia in the male breast, breast carcinomas, and peritoneal and pleural effusions of patients with breast carcinomas. The fibroblast-like cells cultured from either normal, non-malignant atypical, or malignant breast tissue never bound anti-HME, nor did histiobyte-like cells from breast fluid or a sarcoid granuloma of the breast. From a Paget tumor of the breast, two distinguishable epithelial-like colonies appeared in the cultures, only one of which expressed HME-antigens. Anti-HME did not bind to cells cultured from tissues of other organs, whether epithelial-like or fibroblast-like; these organs included thyroid, ovary, endometrium, and skin.


Somatic cell hybrids between human lymphoma cell lines. V. IdUrd inducibility and P3HR-1 superinfectability of Daudi/HeLa (DAD) and Daudi/P3HR-1 (DIP-1) cell lines. We have studied two types of somatic cell hybrid with regard to expression of the Epstein-Barr virus (EBV) cycle and its regulation. The first, DIP-1, a hybrid formed between two human lymphoma EBV producers (Daudi and P3HR-1), contained EBV DNA, expressed the virus-determined nuclear antigen (EBNA), andwas a producer of the EBV-associated antigens EA (early antigen) and VCA (viral capsid antigen). The second, DAD, a hybrid series of clones formed between Daudi and a HeLa cell derivative (D98), differed with regard to the expression of EBNA, EA, VCA and the content of EBV DNA. EA was regularly induced in the EBV DNA-containing hybrids following treatment with iododeoxyuridine (IdUrd). This induction was greater in lines spontaneously expressing EA. In two hybrids, DIP-1 and DAD10, VCA and virus DNA synthesis were also induced in the presence of IdUrd, the latter being detected by in situ hybridization with P3HR-1 EBV complementary RNA. Finally, while DIP-1 was superinfectable by the P3HR-1 EBV strain, the DAD series of hybrids were refractory to P3HR-1 superinfection and lacked EBV receptors.


Antigenic characterization of a new gibbon ape leukemia virus isolate: seroepidemiologic assessment of an outbreak of gibbon leukemia. A type-C virus recently isolated from a leukemic gibbon in a colony located on Hall's Island, Bermuda, was characterized with respect to the antigenic properties of its gag and env gene-coded proteins. This virus, designated GaLV-H, was found to be closely related immunologically to type-C viruses previously isolated from gibbons (GaLV-SF, GaLV-SEATO, GaLV-Br) and from woolly monkey (SSAV). However, GaLV-H was readily differentiated from these isolates in a radioimmunoassay for its env gene product, gp70. Seroepidemiology established that GaLV-H was horizontally transmitted among gibbons within the colony. There was no evidence of exposure leading to an immune response to the virus or viral antigenemia in humans working in association with these animals.


Studies on Epstein-Barr virus-related antigens. III. Purification of the virus-determined nuclear antigen (EBNA) from non-producer Raji cells. The purification of Epstein-Barr virus-determined nuclear antigen (EBNA) was attempted on the basis of its biochemical and physicochemical properties and its immunological specificity, assayed by the indirect single radial immunodiffusion test and anti-complement immunofluorescence absorption test. When non-producer Raji cell extract was subjected to 20--60% saturated ammonium sulfate fractionation, followed by DNA-cellulose chromatography and immunoadsorbent chromatography, EBNA was purified more than 3,000 times with a 8% yield. Such purified protein was composed of three polypeptides with molecular weights of 100,000 +/- 5,000, 70,000 +/- 5,000 and 50,000 daltons, respectively, on SDS-polyacrylamide gel electrophoresis. Similar purification was achieved by heating the extract at 70 degrees C for 10 min, instead of ammonium sulfate fractionation, followed by DNA-cellulose chromatography and immunoadsorbent chromatography. This final preparation consisted almost exclusively of 100,000 +/- 5,000 daltons polypeptide, 50,000 polypeptide, and the 70,000 +/- 5,000 polypeptide passing through the adsorbent column. These findings suggest that EBNA is probably a molecular complex of three smaller subunits of heat-stable 100,000 +/- 5,000 and 50,000 daltons and heat-labile 70,000 +/- 5,000 daltons polypeptides, respectively.


Unresponsiveness to exogenous TSH in obesity. (1). Assessment of thyroidal and other indices in 275 instances of obesity with body weight excesses up to 200 percent or more of the ideal revealed absent thyroidal I131 uptake responses to TSH in about one out of five patients. Moreover, basal thyroidal I131 uptake of 10 percent or less, prolongation of ankle reflex time, or high levels of serum cholesterol were present in a minority. Also, occasional instances of unduly elevated serum TSH titers were found. Some of the indices deviated from normal more often with the greater excesses of body weight or with increased age. (2). These findings are consonant with a hypothesis that routine thyroidal or related indices are sporadically abnormal in massive obesity almost always without overt hypothyroidism or myxedema, that total unresponsiveness to exogenous TSH is surprisingly frequent, and that such unresponsiveness represents an unexplained endocrine anomaly in association with gross overweight. (3). Our data suggest that some obese persons are not able to respond to exogenous TSH, nor, presumably, to increases of endogenous TSH. This could result in an economy of caloric expenditure and play a contributory role in the genesis or the perpetuation of the obesity.


Thyroid and obesity: survey of some function tests in a large obese population. Tests of thyroid function in 493 obese patients were compared with thyroid function in 3076 non-obese patients. No differences in 131I uptake by the thyroid were observed at 6 h or 24 h; the frequency distribution was gaussian or normal in both populations; the frequencies of normal, high or low T3 and T4 values in the two populations were likewise comparable. Further, no correlation was found in the obese subjects between 131I uptake (6 h and 24 h), T3, T4 and overweight; on the other hand, there was a significant negative correlation between 131I uptake (6 h and 24 h) and age. It would appear that thyroid function is normal in obesity.


[Cyclic adenosine 3-5 monophosphate (c-AMP) and allergy]. Cyclic adenosin-3-5-monophosphate (c-AMP) plays a central role in regulating immune responses. Increased intracellular c-AMP has an inhibitory effect on release of histamine and other mediators from basophile leukocytes or mast cells as well as lysosomal enzymes from polymorphs. It depresses cell-mediated immune reactions and antibody formation. The hormonal factors which increase intracellular c-AMP are either of neuroendocrine origin (e.g. epinephrin) or inflammatory products (e.g. prostaglandins and histamine). This seems to be an endogenous mechanism of regulation which is able to stop immune or allergic reactions. Patients suffering from atopic diseases (asthma and/or atopic dermatitis) show a decreased reactivity to betaadrenergic stimuli. The defect is demonstrable in vitro in peripheral leukocytes which show a significantly smaller increase in intracellular c-AMP after betaadrenergic stimulation than normal leukocytes.


The possible involvement of a plasmid(s) in actinomycin synthesis by Streptomyces parvulus and Streptomyces antibioticus. The treatment of Streptomyces parvulus and Streptomyces antibioticus with acriflavine or novobiocin resulted in the loss of ability to produce actinomycin. The concomitant loss of ability to form aerial mycelium and the incidence of auxotrophic progeny (S. parvulus) were both low relative to the loss of the antibiotic-producing property. Protoplast fusion induced by polyethylene glycol 4000, using suitable auxotrophic strains of S. parvulus, resulted in high recombination frequencies to prototrophy (9.6 approximately 15%). When fusion was carried out between auxotrophic act+ and act- strains, respectively, there was a high frequency (84 approximately 95%) of the actinomycin synthesizing character among the prototrophic recombinants examined. No actinomycin-producing recombinant was detected in similar experiments between auxotrophic non-producing strains.


Human leucocyte neutral proteases, with special reference to collagen metabolism. Three different types of neutral proteases related to collagen metabolism have been found in the granule fraction of human leucocytes from normal adults, using collagen, gelatin, and synthetic peptides as substrates. These are collagenase, an enzyme showing a potent hydrolytic activity against gelatin but little against native collagen, and one splitting the cross-links region of collagen. Their molecular weights were estimated to be about 75,000 150,000, and 25,000, respectively, by gel chromatography. The former two enzymes were inhibited by a alpha2-macroglobulin and ethylenediaminetetraacetate, but not by alpha1-proteinase inhibitor (alpha1-antitrypsin) or phenylmethylsulfonylfluoride, while the latter enzyme, associated in behavior with an enzyme hydrolyzing succinyl-(l-alanyl)3-p-nitroanilide, was inhibited by alpha1-proteinase inhibitor, alpha2-macroglobulin, and phenylmethylsulfonylfluoride, but not by ethylenediaminetetraacetate. A possible cooperative function of these enzymes in collagen catabolism is discussed.


Differential inhibition of DNA polymerases of calf thymus by 9-beta-D-arabinofuranosyladenine-5'-triphosphate. The effect of 9-beta-D-arabinofuranosyladenine-5'-triphosphate (araATP) on the reactions of DNA polymerases alpha and beta [E.C. 2.7.7.7] purified from calf thymus was examined. The reaction of DNA polymerase alpha was shown to be more sensitive to the inhibition than that of DNA polymerase beta. The K1 value of DNA polymerase beta for araATP was 45 micrometer; 15 times higher than that of DNA polymerase alpha (3 micrometer). The mode of inhibition by araATP was essentially competitive to deoxyadenosine triphosphate (dATP) in the reactions catalyzed by both DNA polymerase alpha and beta using activated DNA as a template-primer. However, in the reactions of the alpha-enzyme, araATP also inhibited the incorporation of deoxyribonucleotides othan than dATP non-competitively.


The use of gel filtration to follow conformational changes in proteins. Conformational flexibility of bovine myelin basic protein. The hydrodynamic behavior of bovine myelin basic protein was studied by gel filtration through Sephadex G-100 under conditions which included variations in pH from 2 to 12, variations in ionic strength from 0.01 to 1.5 M at pH 2 and from 0.1 to 2 M at pH 7, and variations in guanidinium chloride concentration from 0 to 6 M. A number of well characterized compact globular proteins were subjected to the same conditions for comparison. Compact globular proteins showed major conformational transitions due to acid, alkali, and guanidinium chloride denaturation and, possibly, minor transitions as well. Myelin basic protein behaved like a flexible linear polyelectrolyte, expanding continuously between pH 11 and pH 2 to 3 at ionic strength 0.1 M and contracting continuously with increase in ionic strength at pH 2 and at pH 7 to the point of salting-out. Relatively low concentrations of guanidinium chloride (approximately 0.5 M) were sufficient to cause the basic protein to expand. With increasing concentration of the denaturant the molecule continued to expand, but in a noncooperative manner. These results demonstrated the lack of significant intramolecular stabilization in the protein.


Movies and dental perceptual-motor skills. The burgeoning popularity of motion pictures in dental education, despite the absence of supporting data, presents the possibility that the medium is being used uncritically. Assumptions underlying the choice and production of dental films are enumerated and critiqued. The assumption that the typical dental procedure film provides a useful model of dental motions is seriously questioned. Potentially superior alternatives are proposed.


Assessment of the influence of irrelevant IgE on allergic sensitivity to two independent allergens. Serum samples from 31 patients sensitive to both ragweed and rye grass were quantitated for IgE specific for ragweed antigen E (AgE) and rye grass group I (rye I) antigens employing the previously described radioallergosorbent test (RAST) elution technique. IgE anti-AgE ranged from 0.4 to 320 ng/ml and comprised 0.4% to 14.2% of total serum IgE. The same sera contained IgE anti-rye I ranging from 7.9 to 1,168 ng/ml, comprising 1.6% to 29.6% of total serum IgE. A new theoretical parameter (RF), representing the percent of total IgE-Fc receptors of target cells occupied by allergen-specific IgE, was calculated for each IgE specificity by using the determinations of allergen-specific IgE antibody, total serum IgE, and assuming an equilibrium constant for binding of IgE molecules to basophils of 10(9)M-1. This "% occupancy" parameter correlated with skin test titration, basophil cell sensitivity, and hay fever symptom scores as well as, but not better than, the absolute values of allergen-specific IgE. This finding suggests that in most atopic patients, the quantity of irrelevant IgE plays a relatively minor role in determining allergic sensitivity to a given allergen. The implications of this finding are discussed.


Evaluation of a patient with cold and cholinergic urticaria. A-20-year-old male Army paratrooper presented with a history of inducible urticaria associated with exercise as well as cold exposure. Upon evaluation, he not only had a positive ice cube test, but also had a positive mecholyl skin test with numberous satellite lesions and generalized punctate urticaria following exercise challenge. Thus, he appeared to have combined cold and cholinergic urticaria. When mediator release was examined during cold and exercise challenge, histamine release was observed in each instance; a rapid rise and fall of plasma histamine was seen after cold challenge, while a lag phase followed by sustained elevation of plasma histamine was associated with exercise challenge. This represents the fourth reported case of combined cold and cholinergic urticaria and is the first in whom mediator release was assessed. The time-course of histamine release was characteristic of each disorder.


Activation of latent collagenase of human leukocytes and gingival fluid by bacterial plaque. Collagenase activity was studied in human leukocytes, gingival crevicular fluid and bacterial plaque, with soluble radioactive collagen as substrate. Inflamed gingiva liberated vertebrate type collagenase into the crevicular fluid in active form. Healthy gingiva, in contrast, released collagenase in a latent form that could be activated by trypsin or plaque. Plaque also stimulated leukocytes to release collagenase, and activated the latent enzyme.


Endogenous thyrotropin-releasing hormone in the anterior pituitary: sites of activity as identified by immunocytochemical staining. Thyrotropin-releasing hormone (TRH) immunoreactivity was localized in the rat anterior pituitary with rabbit anti-TRH sera and the unlabeled antibody peroxidase-antiperoxidase complex (PAP) technique. Stain was present in secretory granules of cells possessing morphological characteristics of thyrotropes, gonadotropes and lactotropes. Antibody absorption studies with anti-TRH sera absorbed with TRH, 3 diastereoisomeric analogues of TRH, gonadotropin-releasing hormone (GnRH), bovine serum albumin, thyrotropin, prolactin, adrenocorticotropin, luteinizing hormone, follicle stimulating hormone were performed to determine the specificity of the staining reaction. Only absorption with TRH resulted in a significant reduction in staining intensity. In vitro experiments were then begun with hemipituitaries to ascertain if intrapituitary TRH might originate by sequestration of exogenous, plasma membrane bound TRH or by de novo synthesis. The results suggest that anterior pituitary TRH is of endogenous origin.


Immunoperoxidase stains cortisol in adrenal and pituitary. The unlabeled peroxidase-anti-peroxidase (PAP) method of Sternberger was used to localize cortisol within paraffin embedded sections of cat adrenal and pituitary tissue. Incubation of the cortisol antiserum used in this method with increasing concentrations of cortisol led to progressive extinction of cortisol staining of the adrenal fasciculata cells, (as measured with a scanning integrating microdensitometer). This result suggests strongly that the staining achieved with this method was specific for cortisol. Cortisol staining was demonstrated not only within cells that synthesize cortisol (the adrenal fasciculata) but also in cells of the adrenal medulla and of the anterior pituitary, two target sites for cortisol action.


Masking of protein antigen by modification of amino groups with carbobenzoxychloride (benzyl chloroformate) and demasking by treatment with nonspecific protease. Cryostat sections of various substrates were treated with carbobenzoxychloride in acetone to modify antigens. By applying specific fluorescent antibodies, it could be shown that the antigenic determinants of rabbit gamma-globulin and bovine insulin were totally masked. The antigenicity of ACTH was markedly reduced, whereas the polysaccharide antigens of Salmonella typhimurium were only partially masked. After masking, antigenicity could be restored by treatment with nonspecific protease. The reversible protection of amino groups by carbobenzoxychloride may be a way to preserve protein antigens during embedding in plastics, as such materials also bind to amino groups, blocking the antigenicity of proteins.


Rapid hypotonic method for flow cytofluorometry of monolayer cell cultures. Some pitfalls in staining and data analysis. The rapid hypotonic staining procedure developed by Krishan for DNA determinations by flow cytofluorometry has been proven accurate for in vivo cell samples and for cell lines growing in suspension culture. We show that the unmodified procedure may produce distorted DNA histograms when used for staining cells growing in monolayer cultures, however. To eliminate these distortions, it was necessary to avoid the use of trypsin by staining the attached cells directly, using a hypotonic fluorochrome solution to which nonionic detergent was added. Two sublines of HeLa S3 cells are shown to exhibit major differences in their staining characteristics. By using our revised staining procedure, the two sublines appear to produce very satisfactory DNA histograms. However, in only one subline does the S phase fraction calculated from the histograms agree with the autoradiographical labeling index. Mitotic cells remain intact under these staining conditions, and the principal observed effect of nonionic detergents in this case is to decrease the coefficient of variation of fluorescence intensity.


Studies on the immunosuppressive effect of organochlorine and organophosphoric pesticides in subacute experiments. Subacute experiments were carried out on rabbits by daily oral administration of Lindane, Malathion and Dichlorophos. A dose-dependent significant decrease in Salmonella typhi induced immune response was found in sera of animals treated with these pesticides compared to untreated controls. Inihbition of cholinesterase activity confirmed the immunological findings.


Failure to detect mouse species-specific antigens in Toxoplasmin Sevac. Toxoplasmin--a highly purified extract from the protozoa Toxoplasma gondii propagated in mice--was tested for the presence of the mouse species-specific antigens by immunodiffusion in agar gel and by PCA test. Neither test gave positive reaction. It was concluded therefore that Toxoplasmin Sevac used for the detection of dermal hypersensitivity to toxoplasma antigens in humans does not contain any detectable contamination by mouse species-specific antigens within the limits of sensitivity of both methods used.


Twelve serum protein profiles in children with acute nonbacterial gastroenterocolitis. In thirty children hospitalized with acute benign, short-duration gastroenterocolitis, no obligate pathogens were isolated from stools. Five bleedings were established from each patient in order to obtain the protein profiles of albumin, orosomucoid, haptoglobin, alpha2-macroglobulin, ceruloplasmin, transferrin, C3-component, C-reactive protein, immunglobulins IgG, IgA, IgM and IgD. The proteins were quantitated by the single radial immunodiffusion method. The initial drop in some of the proteins followed may be related to general protein loss, negative nitrogen balance or hemodilution. The absence of a significant increase in all the investigated immunoglobulin classes contrasted with remarkable increase in haptoglobin and orosomucoid, both reaching normal levels in late convalescence. C-reactive protein could be demonstrated in half of the children showing early normalization with disappearance of clinical symptoms. In contrast to ceruloplasmin and C3- component, alpha2-macroglobulin was not involved in the acute phase protein reaction.


Regulation of the immune response to tumor antigen. IV. Tumor antigen-specific suppressor factor(s) bear I-J determinants and induce suppressor T cells in vivo. The nature and function of suppressor factor(s) elaborated by suppressor T cells in response to certain chemically induced tumors have been further defined. Thus, suppressor factor(s) specific for the S1509a methylchol-anthrene-induced fibrosarcoma have been shown to bear determinants encoded by the I-J subregion of the murine MHC since suppressive activity is removed by passage of the factor through an immunoadsorbent composed of anti-I-Jk coupled to Sepharose. No loss of activity was observed after passage of factor through control columns composed of normal mouse globulin. Furthermore, activity could be recovered from the relevant immunoadsorbent by elution with high salt. The administration of crude suppressor factor(s) to normal animals for 4 days resulted in the development of a population of suppressor cells that act in a manner analogous to the suppressor cell population used for production of factor. These factor-induced suppressor cells are T cells and exhibit an antigen specificity similar to that displayed by the tumor-induced suppressor cells. Thus, tumor-specific suppressor factor(s) bear I-J determinants and are capable of inducing the appearance of suppressor T cells in the nontumor-bearing host, which may then act in a specific manner to limit host responsiveness to tumor antigen.


Properties of H-2L locus products in allogeneic and H-2 restricted, trinitrophenyl-specific cytotoxic responses. The role of the recently defined L antigen (a second D region product) in allogeneic and TNP-specific syngeneic primary CML responses has been investigated. The lysis by anti-L specific cytotoxic effector cells was not inhibited when the target cells were pretreated with an antiserum directed against K and D, whereas an antiserum against L completely abrogated this response. Therefore, H-2L products are recognized on the target cell independently of H-2K and H-2D locus products. Both A.SW cells as well as B10 cells were found to respond to Ld alloantigens, in addition to Dd alloantigens when stimulated by cells differing only in the D region. The results of cold target blocking and antiserum inhibition experiments failed to detect cytotoxic cells with specificity of L antigens in association with TNP, under conditions in which TNP-specific effectors to K and D antigens were demonstrable. These findings suggest that there is a more limited involvement of H-2L locus products than the H-2K or H-2D  locus products in the induction and specificity of these responses.


Antigen-induced murine B cell lymphomas. II. Exploitation of the surface idiotype as tumor specific antigen. In the accompanying report, we have described the characterization of two unusual murine B cell lymphomas, CH1 and CH2. A heterologous antiserum, which we refer to as "anti-idiotype" serum, has been raised to the detergent-solubilized surface immunoglobulin of CH1. The following criteria have established that this antiserum is specific for the CH1 tumor and that it reacts with V region determinants of the tumor surface IgM: 1) the antiserum reacts with CH1 tumor cells, but not normal mouse lymphoid cells or CH2 tumor cells, in indirect immunofluorescence and C-dependent cytotoxicity testing, 2) capping with the anti-idiotype serum removes all or most of the tumor surface Ig, 3) the antiserum forms a single band of precipitation against serum from CH1 tumor-bearing mice, when tested by double diffusion precipitin analysis, and 4) a single band of precipitation is formed in the electrophoretic migration position of IgM when the anti-idiotype antiserum is tested against serum from CH1 tumor-bearing mice in immunoelectrophoresis. Furthermore, we have demonstrated that this antiserum is useful in monitoring tumor growth and is a potent immunotherapeutic agent. Specifically, 50% of mice injected with a lethal tumor inoculum and given a small dose of anti-idiotype serum 2 days later remain tumor free, whereas all tumor-challenged control mice died within 30 days.


Regulation of the immune response to tumor antigen. VI. Differential specificities of suppressor T cells or their products and effector T cells. Suppressor T cells arising during the development of certain murine methylcholanthrene-induced fibrosarcomas have previously been shown capable of limiting only those effector responses generated against the homologous tumor. Thus, S1509a-induced suppressor T cells inhibit immune reactivity only to the S1509a tumor in S1509a immune mice and have no effect on the rejection of SAI tumors in SAI-immune animals. In contrast to this is the cross-reactivity of effector cells in this system, whereby animals rendered immune to either the S1509a or SAI sarcoma are equally capable of rejecting a challenge of the opposite tumor. The specificity of suppression has been further defined in the present study, which demonstrates that S1509a-induced suppressor cells can inhibit responsiveness only to the S1509a sarcoma, even in the simultaneous presence of both the S1509a and SAI tumors. Furthermore, the suppressor factor that is obtainable from suppressor T cells demonstrates a similar precise specificity in its ability to limit selectively reactivity only against the inducing tumor, regardless of the simultaneous expression of antigens on other tumors recognized by cross-reactive effector cells. These results suggest that the antigenic determinants recognized by effector and suppressor T cells are different, and may provide a model for further dissection of suppressor cell function in vivo.


Expression of I-A and I-E,C region-coded Ia antigens on functional B cell subpopulations. Ia antigens from specific subregions have been examined on functional B cell populations. Expression of both I-A and I-E,C region antigens was demonstrated on cells required for both lipopolysaccharide mitogenesis and polyclonal activation. Similar I-A and I-E,C subregion expression was found on cells required for response to the T-independent antigen, polyvinylpyrrolidone. TNP-specific IgM and hen egg lysozyme-specific IgG plaque-forming cells also express I-A and I-E,C region antigens. No evidence was found for an Ia- population responsive in the systems tested. Further, no evidence of preferential expression of I-A or I-E,C region antigens was observed in any system examined. Therefore, it appears that B cells express both I-A and I-E,C region-coded Ia antigens.


Enterically induced immunologic tolerance. I. Induction of suppressor T lymphoyctes by intragastric administration of soluble proteins. Specific immune unresponsiveness was induced in inbred mice (BDF1) by the administration of soluble ovalbumin (OVA) by gastric intubation. Anti-hapten (DNP) responses likewise were specifically diminished when animals were fed autologous carrier (OVA or keyhole limpet hemocyanin). Adoptive transfer of spleen cells demonstrated that the tolerant state could be maintained in irradiated recipient mice, and specific anergy could be transferred to normal recipient animals. Adoptive suppression was mediated by T lymphocytes, as demonstrated by nylon wool fractionation and susceptibility of the cells to anti-Thy 1.2 and complement. Transferred B cells had neither suppressive nor augmentative effects. Enteric administration of OVA also specifically diminished antigen-induced DNA synthesis of primed lymph node T cells, although suppressor cells were not identified in the lymph nodes per se.


Genetic control of the immune response to the H-2Dk private specificity, H-2.32. II. Genetic linkage analysis of a backcross generation. B10.AKM mice (H-2M) when immunized with H-2k cells showed very low cytotoxic antibody responses to the H-2Dk private specificity H-2.32, whereas AKR.M and (AKR.M X B10.AKM)F1 mice that possess the same H-2m haplotype mounted reasonable anti-H-2.32 antibody responses. The genetic nature of the non-H-2 linked gene(s) controlling the anti-H-2.32 response was analyzed on the backcross progeny raised between (AKR.M X B10.AKM)F1 and B10.AKM mice. The anti-H-2.32 antibody response was found to be predominantly controlled by a single locus. This locus segregated independently of the Ig heavy chain locus, the Ly2 locus, and the Mls locus. Despite the observed difference in antibody production, no significant differences between AKR.M and B10.AKM mice were detected in induction of H-2Dk-specific killer T cells. Thus, the defect in the response of B10.AKM mice to H-2.32 can be detected at the level of B cell function and is controlled by a single non-H-2-linked genetic locus, but is not attributable to genes linked to the major immunoglobulin structural genes nor to the Mls locus.


Adherent cell function in murine T lymphocyte antigen recognition. II. Definition of genetically restricted and nonrestricted macrophage functions in T cell proliferation. The mechanisms by which adherent cells, presumably of mononuclear phagocytic lineage, influence in vitro antigen-specific activation of murine T lymphocytes was examined. Two distinct functions for macrophages could be discerned. One macrophage function is dependent on a soluble factor produced by cultured adherent cells and is most easily studied with complex multideterminant antigens. This factor is neither antigen-specific nor MHC-restricted in its action in that PEC, regardless of haplotype, produce factor in the absence of antigen. A second function, antigen-specific T cell activation, is seen when antigens of more restricted heterogeneity are used, such as those under the control of Ir genes. This latter activity demands identity or partial identity between the antigen-presenting cell and the primed T cell, thus suggesting an additional specific, genetically restricted function for macrophages in in vitro antigen recognition. Whether these adherent cell functions are mediated by all or distinct subsets of cells was not established.


Purification of alpha-2 macroglobulin by immunoadsorbent chromatography. A method for preparation of alpha-2 macroglobulin (alpha2M) by immunoadsorbent chromatography utilizing rabbit--anti-human alpha2M-conjugated Sepharose 4B is described. This procedure offers a rapid, simple and inexpensive method for purification of alpha2M from as little as 2.0 ml of human serum. By multiple applications to the absorbent column, as much as 10 mg of purified alpha2M may be easily prepared in one day. This technique is especially useful for isolation of alpha2M from individual sera and for studies of cultured cell supernatants containing human alpha2M.


Plasmacytoma spleen colonization: a sensitive, quantitative in vivo assay for idiotype-specific immune suppression of MOPC-315. A spleen colony-forming assay for the measurement of idiotype-specific transplantation resistance to MOPC-315 is described. The assay is highly quantitative, sensitive, reproducible, less time consuming and distinctly superior to conventional in vivo assays which measure tumor incidence, tumor size, and/or host survival time after tumor challenge. The assay directly measures those clonogenic cells of the MOPC-315 myeloma which have a sufficient proliferative capacity to form macroscopic splenic foci within 14 days after intravenous challenge.


Electrophoresis of immunoglobulin G. Facilitated migration of minute amounts in agarose. Migration of very small amounts of immunoglobulin (20 ng) is restricted in agarose electrophoresis. Incorporation of a stable protein matrix (rabbit gamma globulin 1 mg/ml) in the agarose permits unrestricted migration so that immunoelectrophoresis of this quantity of radiolabelled antibody is possible. Very small amounts of radiolabelled and non-radiolabelled antibody were subjected to successful crossed immunoelectrophoresis through barriers of antigen under conditions which provide favorable ratios of antibody to antigen. These methods should be useful for studies of antibody eluted from tissue in acquired and autoimmune diseases associated with tissue bound immunoglobulin.


A rapid, novel method for the solid-phase derivatization of IgG antibodies for immune-affinity chromatography. We show that a solid-phase immune adsorbent can be prepared from rabbit antiserum by isolating the IgG fraction with the A protein of Staphylococcus aureus associated covalently with a Sepharose matrix. The IgG is then coupled to the matrix using the cross-linking agent dimethylsuberimidate. IgG antibody bound in this fashion is in the proper orientation for combination with antigen because association with protein A occurs via the Fc portion of the IgG molecule, thus leaving the combining site of the molecule free to interact with antigen.


Induction of haemolytic plaque-forming cells with sheep lymphoid cells in vitro. The in vitro induction of specific primary and secondary immune responses in sheep lymph node cell suspensions is described and suitable culture conditions determined. The induction of primary immune responses required supplementation of the culture medium with antigen-absorbed homologous serum or lymph, whereas the requirements for the induction of a secondary response were less stringent. The addition of 2-mercaptoethanol to the medium was required. The amounts of heterologous erythrocytes used for immunization were critical and optimal responses were obtained when 50 microleters of a 1% suspension were added to 1 ml cultures. Lymphocyte densities of about 5 X 10(6)/ml were found optimal in primary immune responses in vitro. Less than 2 X 10(6) cells/ml rarely gave rise to plaque-forming cell (PFC) generation, whereas densities of 10 x 10(6) and above reduced the number of PFC obtained per number of cultured cells. Lymphocytes obtained from the efferent lymph draining lymph nodes previously immunized with heterologous erythrocytes were found to generate PFC in vitro when specific antigen was added to the cultures, but attempts to generate PFC in vitro with cells from efferent lymph draining non-immunized nodes failed.


Partial purification of pemphigus-related epidermal antigens. Epidermal antigens partially purified by either isoelectric focusing (the pH 5.2 peak) or concanavalin A (Con A) affinity chromatography react with Con A in tube precipitation reactions. Bands of identity between crude skin antigens, the Con A affinity antigens eluted with alpha-methyl glucoside and the pH 5.2 peak are formed in Ouchterlony gel with rabbit antisera (Rab) to the pH 5.2 antigen. Absorption of Rab or pemphigus antibodies (Pab) with A+ erythrocytes does not affect complement fixation reactions of Rab with the skin antigen nor abolish the ability of Pab to interact with the intercellular cement. The pH 5.2 epidermal antigens react weakly with Pab in tube precipitation reactions and only weakly, if at all, to inhibit Pab reactions in the region of the intercellular cement. High concentrations of Con A inhibit the Pab, peroxidase-anti-IgG tissue reaction whereas the converse inhibition does not occur. Simultaneous use of both Pab and Con A-perodixase reactions at Con A concentrations which do not inhibit Pab, causes enhanced tissue peroxidase reactions in the region of the intercellular cement. These preliminary data indicate that the Pab and Con A-reacting sites are localized on different molecules or antigenic determinants in the intercellular cement. They exclude the possibility that A-blood substances are involved in either site.


Delayed neurotoxicity induced by organophosphorus compounds in the wild mallard duckling: effect of leptophos. Feeding of 260 ppm of leptophos to mallard ducklings caused delayed neurotoxicity similar to that reported for hens. Thus leptophos caused ataxia, with subsequent paralysis, loss of appetite, and slow-down in the growth rate of the treated birds. Spinal lesions were identical in morphology and distribution to those seen in hens following leptophos administration. The severity of histologic changes correlated both with the clinical condition and the duration of intoxication.


The presence of F9 antigen on the surface of mouse embryonic cells until day 8 of embryogenesis. The fate of F9, a surface antigen common to embryonal carcinoma cells and cleavage embryos, as well as to male germ line cells has been studied in post-implantation mouse embryos. The antigen is readily detected on the surface of 7-day and 8-day, but not on 9-day embryonic cells, using anti-F9 serum absorption experiments and indirect immunofluorescence staining. In addition, it is shown that an anti-F9 serum absorbed with 8-day (but not with 9-day) embryonic cells does not react with cleavage embryos (morulae). It is concluded that the antigenic determinants, which are recognized on the surface of morulae by the anti-F9 serum, persist until day 8 of embryogenesis, but are not detected on day 9, either due to their absence, masking or quantitative reduction.


Kepone: a literature summary.

Kepone: a literature summary. This overview provides a narrative survey of the reference literature on Kepone. The circumstances surrounding the "Hopewell incident" have increased the awareness and vigilance of state and federal agencies regarding human and environmental exposure to toxic chemicals. Highlights of this overview include history, nomenclature, uses, toxicology, environmental aspects, epidemiology, and conclusions. A chronologic history of Kepone is detailed in the Appendix.