Estimation of Type-Specific Neutralizing Antibody to Herpes Simplex Virus Type 2 in Uterine Cervical Cancer Patients by a New Absorption Method

A simple method of estimating type-specific neutralizing antibody to type 2 herpes simplex virus (HSV-2) was devised with the use of the microneutralization system. Serially diluted serum was mixed in the well with a constant amount of type 1 virus (HSV-1), and after 3 days' incubation at 37 C, the plate was irradiated with ultraviolet light. The absorbing HSV-1 consisted of culture fluid plus an extract of infected Vero cells not especially concentrated. The well then received indicator HSV-1 or HSV-2, and after being left at 37 C for 1 hr a suspension of dispersed Vero cells was dropped into the wells, following our standard neutralization procedure. Preliminary tests with rabbit antisera showed that even a low level of HSV-2 antibody was detected by this method, unless an exceptionally high titer of HSV-1 antibody originally coexisted with the HSV-2 antibody. Sera from acutely infected persons testified to the specificity of the antibody so detected. It was revealed by means of the new technique that the rate of HSV-2 antibody was significantly higher in uterine cervical cancer patients than in control women. There was no correlation between the clinical stage of cervical cancer and the presence of HSV-2 antibody.     

Stimulation of anaphylaxis in the mouse footpad by dietary fish oil fatty acids

The effect of fish oil-derived omega-3 (omega-3) fatty acids on anaphylaxis, Arthus and delayed type hypersensitivity reactions in mice has been investigated. Mice on a normal chow diet were fed eicosapentaenoic acid and docosahexaenoic acid at a dose of 500 and 333 mg/kg/day, respectively, by a gastric tube over a period of 61 days. Control groups were given water, safflower oil or oleic acid. Anaphylactic and Arthus type reactions were induced in the mouse footpad using bovine serum albumin as an antigen. Carrageenin was utilized to produce a delayed type hypersensitivity reaction. The animals fed omega-3 fatty acids induced a more anaphylactic foodpad reaction. There was no significant effect of the diet on Arthus and delayed type hypersensitivity responses. There was no effect of the fish oil-supplemented diet on production of antibodies to bovine serum albumin. Synthesis of prostaglandin E2 by peritoneal macrophages was significantly inhibited in the animals fed omega-3 fatty acid-enriched fish oil, while leukotriene B4 production was not affected. These results suggest that a diet enriched in omega-3 fatty acids modulates production of arachidonic acid metabolites and this may influence anaphylaxis, but not Arthus and cellular mediated hypersensitivity responses.     

Effect of forskolin on collagen production in clonal osteoblastic MC3T3-E1 cells

The effect of forskolin on collagen production in osteoblasts was investigated by using clonal osteoblastic MC3T3-E1 cells cultured in a-minimum essential medium containing 0.1% bovine serum albumin. Forskolin increased the adenylate cyclase activity in membranes pelleted from homogenates of the cell line in a dose-dependent manner. The drug caused a 13-fold stimulation at 10(-4) M, indicating that the compound directly acts on adenylate cyclase, leading to an increase in the intracellular cAMP content of the cells. Collagen accumulation in the cultures was elevated by one-day treatment with 5 X 10(-5) M forskolin to about twice that in the controls. The stimulation was mainly due to an elevation in collagen synthesis but not to an inhibition of intracellular collagen degradation because forskolin dose-dependently increased collagen synthesis; it also significantly increased the amount of low-molecular-weight hydroxyproline found in the cultures. Cells treated with forskolin produced mainly type I collagen, as found in bone matrix in situ, with only small amounts of other types of collagen. Furthermore, forskolin time-dependently inhibited DNA synthesis in the cells, indicating that the increase in type I collagen synthesis by forskolin was not due to stimulated cell proliferation. These results suggest that cAMP is closely linked to the differentiation of osteoblasts in vitro.     

A method for measuring the amount of retrogradely transported HRP in the rat masseteric motoneuron using Mesulam's HRP histochemical protocol and an image processing system. An investigation of the effect of dopamine receptor antagonists on the retrograde transport of HRP

We developed a method for a determination of the amount of retrogradely transported HRP in the rat masseteric motoneuron using a modification of Mesulam's HRP histochemical protocol and an image processing system combined with a light microscope and a television camera. To test the validity and reproducibility of the new method, a quantitative comparison of the amount of dark blue granules of HRP-product in the cell body of masseteric motor neurons was performed between the right and left trigeminal motor nuclei of 70 rats, which resulted in no significant difference. An additional study used the method was made of the effects of administration of five dopamine receptor antagonists with different biochemical and pharmacologic properties on retrograde transport of HRP in the rat masseteric motoneuron. As a result, chlorpromazine, haloperidol, SCH 23390, and sulpiride significantly enhanced retrograde transport of HRP as against domperidone which showed no significant change in the transport. A possible regulatory system for retrograde transport of HRP in the masseteric motoneuron was discussed in relation to the action of the dopamine receptor.     

The influence of experimentally produced oligohydramnios on lung growth and pulmonary surfactant content in fetal rabbits.

To study the effect of oligohydramnios on lung growth and biochemical lung development in fetal rabbits, amniotic fluid was drained through a tube inserted into the maternal peritoneal cavity on the 23 day of gestation. Littermate fetuses without an amniotic shunt were used as controls. The fetuses were delivered abdominally on the 28 day of gestation. In a total of 8 pregnant does, 17 fetuses underwent amniotic shunting and 22 fetuses were used as controls. The amniotic shunt produced a significant reduction in the amniotic fluid volume. There were no differences in the wet weights of the fetal body, liver or brain between the two groups. However, the amniotic shunt significantly decreased the wet weight of the fetal lung, fetal lung wet weight/body weight ratio, and protein concentration per lung as compared to the control fetuses. In the fetal liver and brain tissues, no changes were found in the concentrations of total phospholipids, phosphatidylcholine (PC) or disaturated phosphatidylcholine (DSPC, the main component of lung surfactant) per g of wet tissue and per mg of protein. However, the lungs of the fetuses with amniotic shunts contained significantly more PC and DSPC, and the L/S ratio was higher than in the control fetuses. These results suggest that the oligohydramnios produced by an amniotic shunt causes pulmonary hypoplasia, but raises the pulmonary surfactant content of fetal rabbit lung.     

Cytologic diagnosis of laryngeal and hypopharyngeal squamous cell carcinoma in sputum

A prospective study of the value of sputum cytology in the diagnosis of squamous cell carcinoma of the larynx and hypopharynx is reported. Sputum cytology established the diagnosis in 63.5% of the patients with laryngeal lesions and in 77.4% of the patients with hypopharyngeal lesions. In laryngeal cancer, a positive diagnosis by sputum cytology was related to clinical T factors (according to the TNM classification): while only 29.4% of T1 lesions were positively detected by sputum cytology, 63.3% of T2 lesions, 69.7% of T3 lesions and 79.2% of T4 lesions were so detected. In hypopharyngeal cancer, there was no discernible relationship between sputum cytodiagnosis and clinical T factors. Generally, there was only a small number of cancer cells present in the sputum in these cases. Some of the squamous cancer cells were not very conspicuous and would require careful screening of the sputum specimens to be detected.     

A kinetic study of the inhibition of yeast AMP deaminase by polyphosphate

Inorganic pyrophosphate and polyphosphates have acted as potent inhibitors of purified AMP deaminase (EC 3.5.4.6) from yeast: the activity fell to a definite limit with the increase in the concentration of the inhibitor. The effect of polyphosphate was largely on the maximal velocity of the enzyme with some decrease in affinity. The cooperative effect of AMP, analyzed in terms of a Hill coefficient, remained at 2 in the absence and presence of polyphosphate. Binding of polyphosphate to the enzyme showed no cooperativity. The inhibition of AMP deaminase by polyphosphate can be qualitatively and quantitatively accounted for by the partial mixed-type inhibition mechanism. Both the Ki value for the inhibitor and the breakdown rate of the enzyme-substrate-inhibitor complex are dependent on the chain length of polyphosphate, suggesting that the breakdown rate of the enzyme-substrate-inhibitor complex is regulated by binding of polyphosphate to a specific inhibitory site.     

Thyrotropin-releasing hormone and insulin in chemically induced pancreatic islet cell tumors in rats

Thyrotropin-releasing hormone (TRH) and insulin were measured by radioimmunoassay in acetic-acid extracts of 19 pancreatic islet cell tumors induced by streptozotocin and nicotinamide in rats. In addition, gel filtration properties of TRH-immunoreactivity and immunoreactive insulin (IRI) were examined in 5 and 14 tumors, respectively. TRH was demonstrated in 10 of 19 tumors, with a mean of 166 +/- 47 (SEM) pg/mg wet weight, whereas the concentration was less than 3 pg/mg wet weight in the other tumors. In contrast, all tumors contained IRI, with a mean of 11.0 +/- 1.6 micrograms/mg wet weight. Ten tumors in which TRH was demonstrated contained more IRI than those in which TRH was not detected (13.1 +/- 1.8 vs 6.5 +/- 1.7 micrograms/mg wet weight, P less than 0.02). After gel filtration, all TRH immunoreactivity was eluted at the same place as synthetic TRH in the 5 tumors. In addition, gel filtration elutes showed essentially the same pattern of IRI in the 14 tumors, with 3 peaks. The predominant IRI peak comigrated with marker insulin (95.7 +/- 0.8%), another prominent peak occurred coincident with proinsulin standard (3.3 +/- 0.5%), a third peak was present in the void volume (0.28 +/- 0.04%). These distributions of IRI were similar to those in extracts of normal pancreases. The present studies demonstrate TRH immunoreactivity in pancreatic islet cell tumors induced by streptozotocin and nicotinamide in rats. Chemically induced insulinomas can serve as a model for insulin storage which is analogous to islet B cells.     

Surface antigens of Biomphalaria glabrata (Gastropoda) hemocytes: functional heterogeneity in cell subpopulations recognized by a monoclonal antibody

A hemocyte surface membrane marker (BGH₁) has been identified using hemocyte-specific monoclonal antibodies (mABs) generated by somatic cell fusion methods. The BGH₁ epitope was expressed on a subpopulation of circulating, glass-adherent blood cells from two strains of the snail, Biomphalaria glabrata. Approximately 40% of the circulating hemocytes from the PR albino (M-line) B. glabrata strain were BGH₁^?, compared to a prevalence of 10% BGH₁⁺ cells in the 10-R2 snail strain. When hemocytes were firmly attached and spread on a glass surface, BGH₁⁺ cells were morphologically distinguishable from BGH₁^? cells by their ovoid shape and the presence of short, thin filopodial projections along the ectoplasmic border. In contrast, BGH₁^? hemocytes were more pleomorphic and possessed long, spike-like filopodia. Moreover, the BGH₁ epitope was trypsin-resistant and retained its antigenic reactivity with probe mABs following fixation with paraformaldehyde or paraformaldehyde/MeOH. Fixation with glutaraldehyde, however, significantly reduced mAB binding to the BGH₁ surface epitope. There was no apparent age-dependent expression of the BGH₁ determinant since circulating hemocyte populations in very young (1–2 mm) to adult (10–12 mm) snails were composed of both BGH₁⁺ and BGH₁^? subpopulations. Quantitative shifts in the prevalence of epitope-bearing hemocytes between the smallest snail size class (1–2 mm) and the larger snails (3–4 and 10–12 mm) are believed to be due to a differential production and/or release of BGH₁^? hemocytes within the blood circulation rather than a gradual age-related change in the expression of surface antigens on individual cells. Experiments designed to assess the in vitro phagocytic capability and lysosomal acid phosphatase (APase) activity of mAB-reactive hemocytes revealed that BGH₁⁺ cells, when compared to those lacking the surface marker, were significantly reduced in both their phagocytic and APase-producing activities. Since the PR albino strain of B. glabrata possesses a higher proportion of BGH₁^? hemocytes and a lower total concentration of circulating cells than do snails of the 10-R2 strain, PR albino snails are thus potentially reduced in their natural capacity to mount cellular reactions against foreign materials.     

Differential expression of 2′∶3′-cyclic nucleotide 3′-phosphodiesterase in cultured central,peripheral, and extraneural cells

The relative levels of the central nervous system myelin marker enzyme 2:3-cyclic nucleotide 3-phosphodiesterase (EC 3.1.4.37, CNPase) were determined in neuroblastoma, astrocyte, oligodendrocyte and Schwann cell cultures and in freshly isolated human lymphocytes and platelets. The highest specific activities were associated with the cells that elaborate myelin membrane in the central and peripheral nervous system, oligodendrocytes and Schwann cells, respectively. Antiserum to bovine CNPase recognized both CNP1 and CNP2 in CNS myelin and human oligodendroglioma. In addition, a 53,000 dalton protein was evident on autoradiographs of immunoblotted PNS myelin and human oligodendroglioma proteins. Cultured rat oligodendrocyte, C₆ and mouse NA neuroblastoma CNPase appear to share common determinants with the corresponding normal rat CNS enzyme.