Tumor necrosis factor-dependent production of human immunodeficiency virus 1 in chronically infected HL-60 cells

Tumor necrosis factor (TNF) may play a central role in proviral activation and release from latency in cells infected with the human immunodeficiency virus (HIV). We studied viral production and its relation to TNF in a HL-60 cell line (J22-HL-60) infected with a monocytotropic strain of HIV-1JR-FL. Viral production was stimulated to similar levels by TNF, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), and 1,25-dihydroxyvitamin D3 (1,25[OH]2D3). Production of the virus was not suppressed by 3'-azido-3'-deoxythymidine (AZT), indicating that viral production was not caused by superinfection. Low concentrations of TNF (0.1 ng/mL) induced viral production with a short lag period of 8 hours, and this proviral activation was specifically suppressed by anti- TNF antibodies. However, induction of virus production by 1,25(OH)2D3 showed an extended lag period of 2 to 3 days. The effect of 1,25(OH)2D3 on virus production was also blocked by anti-TNF antibodies, which suggests the direct participation of TNF in this process. TNF accumulated in the culture supernatant of cells stimulated with 1,25(OH)2D3 with a kinetics consistent with its involvement in the action of 1,25(OH)2D3 on viral production. The J22-HL-60 cell line produced low levels of virus when cultured in the absence of an external stimulus; however, this basal viral production was suppressed greater than 80% in the presence of anti-TNF antibodies. Corresponding low levels of TNF were detected in the culture supernatants. Viral production decreased slowly with increasing passage of the cells, and no virus was detected in the supernatants of cells maintained in culture for several months. Concomitantly, TNF was no longer detected in the supernatant of these cells, which suggests that endogenous autocrine production of TNF drives viral production in the unstimulated cells. However, viral production was stimulated in these cells by low concentrations (0.1 ng/mL) of added TNF. These results argue for a central role for TNF in HIV proviral activation in chronically infected myeloid cells.     

Quantitation of MR relaxation effects of iron oxide particles in liver and spleen

The relaxation effects and organ distribution of superparamagnetic iron oxide particles for magnetic resonance imaging were measured in rats. T1 and T2 were measured for excised organs, and tissue iron levels were quantified with radiolabeling. Approximately 70% of the injected dose is present in the liver and 10% in the spleen 1 hour after injection. At 20 MHz, the doses required to reduce liver and spleen T2 to half the normal value, as measured with a Carr-Purcell-Meiboom-Gill sequence, were, respectively, 420 and 830 mumol iron injected per kilogram of rat. The transverse relaxation rates increase linearly with injected dose and showed no evidence of saturation. These results suggest that this material is less effective than previously suggested.     

Source of hydrogen peroxide and of chemiluminescence observed in activated human platelet preparations

Human platelet suspensions can be observed to produce small amounts of H2O2 (0.04 nmoles H2O2/min/2.5 X 10(5) cells/cu mm) and measurable chemiluminescence when exposed to target particles for phagocytosis, such as latex spherules. Both H2O2 production and chemiluminescence are characteristic of phagocytosing polymorphonuclear leukocytes (PMN) and analysis of the purified platelets indicates contamination by PMN at the level of 0.2%. The amount of H2O2 produced and the chemiluminescence observed can be duplicated by adding latex spheres to a preparation of PMN at a concentration equivalent to the contaminant in the platelet preparations. We conclude that the H2O2 produced and chemiluminescence observed from activated platelets is due to the presence of small amounts of contaminating PMN. These studies emphasize the importance of controlling for PMN contamination in studies of platelet biochemistry and physiology.     

Reductions in retinal gamma-aminobutyric acid (GABA) content and in [3H]flunitrazepam binding after postnatal monosodium glutamate injections in rats

Specific binding of [3H]flunitrazepam is found in the mammalian retina and its characteristics are similar in important respects to those in the cerebral cortex. Numerous reports have shown that monosodium glutamate (MSG) given neonatally to rats results in neuronal cell death with sparing of photoreceptor and glial cells. Sprague-Dawley rats were given MSG (3.2 mg/g i.p.) from day 2 to day 12 after birth; controls received equimolar injections of NaCl. At 8 to 9 weeks of age, the rats were killed and [3H]flunitrazepam binding was examined in the retinas are various brain regions. Histologic evidence showed the virtual absence of ganglion cells and a marked reduction of neurons in the inner nuclear layer of retinas from MSG-treated rats; photoreceptor and Muller cells appeared normal. In the retinas from MSG-treated rats, gamma-aminobutyric acid levels were decreased by 73% and the Bmax of [3H]flunitrazepam binding was decreased by 77%; there was no change in Kd. In the cerebellum, cerebral cortex and hypothalamus of MSG-treated rats, [3H]flunitrazepam binding was unchanged. These results strengthen the association of gamma-aminobutyric acid mechanisms with benzodiazepine binding and suggest a predominant neuronal localization of the binding sites.     

Non-ST elevation myocardial infarction (NSTEMI) in three hospital settings in South Africa: does geography influence management and outcome? A retrospective cohort study

Background

Guidelines advise early angiography in non-ST elevation myocardial infarction (NSTEMI) to ensure an optimal outcome. Resource limitations in secondary hospitals in the Western Cape dictate a local guideline to treat NSTEMIs medically with out-patient assessment for angiography, unless mandatory indications for early angiography occur.

Methods

A retrospective cohort study assessed NSTEMIs at Tygerberg Hospital (TBH), Karl Bremer Hospital (KBH) and Worcester Hospital (WH) over one year. Two cohorts were analysed, secondary hospitals (KBH and WH; SH) and secondary service within a tertiary hospital (TBH). Where differences were found, sub-analysis compared WH and KBH.

Results

TBH and SH were similar at baseline and in clinical presentation. Cases at TBH were more likely to receive in-patient angiography (94 vs 51%, p < 0.0001), and had a lower in-patient mortality rate (6 vs 23%, p = 0.0326). There was no difference between KBH and WH in sub-analysis.

Conclusion

This study confirmed that the management and mortality of NSTEMIs in the public health sector in the Western Cape, South Africa is not influenced by geography, but rather by the level of service available in the hospital of first presentation.     

Generation of natural killer cells from Thy 1.1+ bone marrow precursor cells in the rat

We have recently described a long-term bone marrow culture (LTBMC) system in the rat for the generation of natural killer (NK) cells from bone marrow (BM) precursors in the presence of interleukin-2 (IL-2). We found that the LTBMC-conditioned medium was essential to render the NK precursor cells responsive to IL-2. In this report, we isolated by flow cytometric cell sorting Thy 1.1+ BM cells, which have been shown to contain pluripotent stem cells and early precursor cells of various hematopoietic lineages. These Thy 1.1+ immature BM precursors did not generate any detectable NK activity when cultured for 7 days with IL-2 alone. However, when the cells were cultured with IL-2 in the presence of LTBMC-conditioned medium, NK cells were generated as demonstrated by cytolytic activity against NK-sensitive tumor targets, large granular lymphocyte (LGL) morphology, and the acquisition of NK cell-associated phenotype (72% of the cells were 3.2.3+/OX41-/CD5-). This study demonstrates the existence of an IL-2 unresponsive Thy 1.1+ NK precursor in the BM of the rat, that can differentiate to a mature NK cell in the presence of LTBMC-conditioned medium and IL-2.     

The relationship between agonist states of the M1 muscarinic receptor and the hydrolysis of inositol lipids in transfected murine fibroblast cells (B82) expressing different receptor densities

We studied the relationship between the M1 muscarinic receptor density and the receptor-mediated hydrolysis of inositol lipids in cloned murine fibroblast B82 cells which were transfected with the m1 muscarinic receptor gene. Of the seven clones examined, the M1 muscarinic receptor densities in these cells characterized by (-)[3H]methyl-3-quinuclidinyl benzilate ([-)-[3H]MQNB binding ranged from 12 fmol/10(6) cells in LK3-1 cells to 260 fmol/10(6) cells in the LK3-8 cells. Carbachol/(-)[3H]MQNB competition curves for the LK3-1 cells (with low receptor density) had a Hill coefficient close to unity. The competition curves for carbachol in the clones with higher receptor densities had Hill coefficients less than 1 and were best fitted by a computerized nonlinear least-squares regression program for the two-site model. The percentage of the M1 muscarinic receptors which had high affinity for carbachol decreased as the receptor density increased, suggesting that the presence of endogenous factors in these cells may be important for the agonist affinity state of the receptor. Concentration-response curves for carbachol-stimulated [3H]inositol monophosphate [( 3H]IP1) accumulation were also obtained. A significant correlation was observed between the density of M1 muscarinic receptor with high affinity for carbachol and the maximum [3H]IP1 accumulation in these cells. There is no significant difference among the EC50 values and the dissociation constant of high-affinity state values of the carbachol/(-)[3H] MQNB competition curves. These results suggest that the high-affinity state for carbachol may be the functional state of the M1 muscarinic receptors in these transfected B82 cells.(ABSTRACT TRUNCATED AT 250 WORDS)