B lymphocyte colony-forming cells in the SJL/J mouse.

Mercatoethanol-induced B lymphocyte cloning in semi-solid agar has been used to study lymphocyte colony formation by cells from the SJL/J mouse thymus. From the 3rd month of life, the SJL/J mouse thymus. From the 3rd month of life, the SJL thymus develops an increasing frequency of cells forming B lymphocyte colonies in agar. The peak frequency in 6- to 12-month-old mice was one colony per 1000 to 2000 cultured thymus cells. In contrast, 10 to 100 times lower frequencies were found in the thymus of five other inbred mouse strains. The rise in B lymphocyte colony-forming cells correlated well with the age-related rise in Ig-positive cells and approximately 50% of the colony cells reacted with anti-micron-serum indicating the B lymphocyte nature of the colony cells. Colony-forming cells from the thymus showed higher sensitivity than colony-forming spleen cells to cortisol and irradiation. Cell transfer experiments and thymus grafting suggested that the increased frequency of colony-forming cells in the thymus is caused by development of special thymus-seeking B lymphocytes in ageing SJL/J mice. Finally, B lymphocyte colony-forming cells were found to be more frequent in the thymus, spleen, and lymph nodes from healthy aged mice than in lymphoid organs from mice with spontaneous reticulum cell tumors.     

gamma-Vinyl GABA (4-amino-hex-5-enoic acid), a new selective irreversible inhibitor of GABA-T: effects on brain GABA metabolism in mice

Abstract— γ-Vinyl GABA (4-amino-hex-5-enoic acid, RMI 71754) is a catalytic inhibitor of GABA-T in vitro. When given by a peripheral route to mice, it crosses the blood-brain barrier and induces a long-lasting, dose-dependent, irreversible inhibition of brain GABA transaminase (GABA-T). Glutamate decarboxylase (GAD) is only slightly affected even at the highest doses used. γ -Vinyl GABA has little or no effect on brain succinate semialdehyde dehydrogenase, aspartate transaminase and alanine transaminase activities. GABA-T inhibition is accompanied by a sustained dose-dependent increase of brain GABA concentration. From the rate of accumulation of GABA it was estimated that GABA turnover in brain was at least 6.5 μmol/g/h. Based on recovery of enzyme activity the half-life of GABA-T was found to be 3.4 days, that of GAD was estimated to be about 2.4 days. γ -Vinyl GABA should be valuable for manipulations of brain GABA metabolism.     

Bioaccumulation and depuration of enteroviruses by the soft-shelled clam, Mya arenaria.

Low levels of feces-associated natural virus, simulating virus numbers estimated to exist in moderately polluted shellfish-growing waters, were used to evaluate the effectiveness of depuration as a virus depletion procedure in soft-shell clams. Depuration effectiveness depended upon the numbers of virus bioaccumulated and whether virus was solids associated. Virus uptake was greatest when viruses were solids associated and pollution levels were equivalent or greater than those likely to be found in grossly polluted growing waters. Virtually all bioaccumulated feces-associated natural virus was deposited within either the hepatopancreas or siphon tissues. Viruses usually were eliminated within a 24- to 48-h depuration period. Dependence upon depuration of clams to elimate health hazards of virus etiology involved a risk factor not measureable in the study. The greatest reduction of health risks would come from the routine depuration of clams harvested from growing waters of good sanitary quality.     

Defect in macrophage effector function confers Salmonella typhimurium susceptibility on C3H/HeJ mice

The defective allele of the endotoxin response locus (Lps^d) renders mice (e.g., C3H/HeJ strain) both endotoxin hyporesponsive and susceptible to Salmonella typhimurium. In this study, the mechanism of Lps^d-regulated susceptibility to murine typhoid was examined. C3H/ HeJ mice became significantly more resistant to S. typhimurium by reconstitution with bone marrow from syngeneic C3H/HeN mice (Lpsⁿ, salmonella resistant). Thus, the Lps^d resistance defect appeared to reside in a radiosensitive bone marrow-derived cell(s). At least one of the abnormal cell types appeared to be a macrophage because C3H/HeJ mice preinfected with Mycobacterium bovis (BCG) were, in contrast to controls, able to restrict early salmonella replication in their spleens and displayed a signficant increase in mean time to death. In contrast, no deficiency in uptake of salmonellae by C3H/HeJ macrophages was observed. These results indicate that the early deaths of C3H/HeJ mice following S. typhimurium challenge reflect a failure of their macrophages to limit the growth of these gram-negative bacteria.     

Clearance and fate of leukemia-inhibitory factor (LIF) after injection into mice

Leukemia-inhibitory factor (LIF) elicits effects on a broad range of cell types, including cells of the monocytic and megakaryocytic series, embryonal stem cells, hepatocytes, adipocytes, and osteoblasts. Native and recombinant LIF, injected intravenously into adult mice, had an initial half-life of 6-8 min and a more prolonged second clearance phase. Clearance of 125I-LIF from the circulation was paralleled by a rapid accumulation in the kidneys, liver, lungs, and spleen and a more gradual accumulation in the thyroid gland. Labeling of the renal glomerular tufts, parenchymal hepatocytes, splenic red pulp, alveolar pneumocytes, and thyroid follicular cells as well as of megakaryocytes and osteoblasts in the bone cavities, placental trophoblasts, and cells of the choroid plexus was demonstrable autoradiographically. The appearance of a large amount of nonprecipitable 125I in the urine suggested that the kidneys were the major route of LIF clearance from the body.     

Crystallization and initial crystallographic results for pepstatin A inhibited bovine cathepsin D.

Cathepsin D was purified from bovine liver by a method using two pepstatin A affinity columns. The eluted protein was combined with pepstatin A and the complex crystallized from 15% polyethylene glycol 8000 at pH 5.9. The crystals diffract to a resolution of 3.0 A and have space group P2(1)2(1)2(1) with unit cell dimensions a = 74.8 A, b = 76.0 A, c = 157.7 A. There are two molecules in the asymmetric unit. The structure was solved by molecular replacement using a pepsin search model and both molecules showed clearly interpretable density in the position expected for pepstatin A in a preliminary difference map. The refined model has r.m.s. deviations from ideal bond lengths and angles of 0.014 A and 3.2 degrees, respectively, and a crystallographic R factor of 17%.     

Concentration and detection of hepatitis A virus and rotavirus from shellfish by hybridization tests.

A modified polyethylene glycol precipitation method for concentration of virus followed by a new method to recover nucleic acid was used to detect hepatitis A virus (HAV) and rotavirus (SA11) in shellfish (oysters and hard-shell clams) by hybridization tests. Infectious virus, seeded into relatively large quantities of shellfish, was recovered consistently, with greater than 90% efficiency as measured by either in situ hybridization (HAV) or plaque assay (rotavirus SA11). Viral nucleic acid for dot blot hybridization assays was extracted and purified from virus-containing polyethylene glycol concentrates. Separation of shellfish polysaccharides from nucleic acid was necessary before viral RNA could be detected by dot blot hybridization. Removal of shellfish polysaccharides was accomplished by using the cationic detergent cetyltrimethylammonium bromide (CTAB). Use of CTAB reduced background interference with hybridization signals, which resulted in increased hybridization test sensitivity. After polysaccharide removal, dot blot hybridization assays could detect approximately 10(6) physical particles (corresponding to approximately 10(3) infectious particles) of HAV and 10(4) PFU of SA11 rotavirus present in 20-g samples of oyster and clam meats. These studies show continuing promise for the development of uniform methods to directly detect human viral pathogens in different types of shellfish. However, practical applications of such methods to detect noncultivatable human viral pathogens of public health interest will require additional improvements in test sensitivity.     

Chlorine resistance of poliovirus isolants recovered from drinking water.

Poliovirus 1 isolants were recovered from finished drinking water produced by a modern, well-operated water treatment plant. These waters contained free chlorine residuals in excess of 1 mg/liter. The chlorine inactivation of purified high-titer preparations of two such isolants was compared with the inactivation behavior of two stock strains of poliovirus 1, LSc and Mahoney. The surviving fraction of virus derived from the two natural isolants was shown to be orders of magnitude greater than that of the standard strains. These results raise the question whether indirect drinking water standards based on free chlorine residuals are adequate public health measures, or whether direct standards based on virus determinations might be necessary.     

Bioaccumulation and depuration of enteroviruses by the soft-shelled clam, Mya arenaria.

Low levels of feces-associated natural virus, simulating virus numbers estimated to exist in moderately polluted shellfish-growing waters, were used to evaluate the effectiveness of depuration as a virus depletion procedure in soft-shell clams. Depuration effectiveness depended upon the numbers of virus bioaccumulated and whether virus was solids associated. Virus uptake was greatest when viruses were solids associated and pollution levels were equivalent or greater than those likely to be found in grossly polluted growing waters. Virtually all bioaccumulated feces-associated natural virus was deposited within either the hepatopancreas or siphon tissues. Viruses usually were eliminated within a 24- to 48-h depuration period. Dependence upon depuration of clams to elimate health hazards of virus etiology involved a risk factor not measureable in the study. The greatest reduction of health risks would come from the routine depuration of clams harvested from growing waters of good sanitary quality.