The role of inducible nitric oxide synthase in the host response to Coxsackievirus myocarditis

The host response to Coxsackievirus infection is complex, including T lymphocytes, B lymphocytes, natural killer cells, and macrophages. Although Coxsackievirus infection induces expression of inducible nitric oxide synthase (NOS2; EC 1.14.13.39) in macrophages, the precise role of NOS2 in the host response to Coxsackievirus myocarditis has been unclear. We show, by using mice homozygous for a disrupted NOS2 allele, that Coxsackievirus replicates to higher titers in NOS2(-/-) mice, that the host lacking NOS2 clears virus more slowly than the wild-type host, and that myocarditis is much more severe in infected NOS2(-/-) mice. These data show that NOS2 is crucial for the host response to Coxsackievirus in the mouse.     

Niacin decreases removal of high-density lipoprotein apolipoprotein A-I but not cholesterol ester by Hep G2 cells. Implication for reverse cholesterol transport

Increasing the expression of c-FMS (colony-stimulating factor 1 receptor) by introduction of a transgene reduced the concentration of retinoic acid or 1,25-dihydroxy vitamin D3 needed to cause myeloid or monocytic cell differentiation and hypophosphorylation of the retinoblastoma tumor suppressor protein (RB) typically associated with cell cycle G0 arrest and differentiation of HL-60 human myelo-monoblastic precursor cells. The data are consistent with a model in which signals originating with retinoic acid and c-FMS integrate to cause differentiation, RB hypophosphorylation, and G0 arrest. Furthermore, these two signals can compensate for each other. Three HL-60 sublines described previously (A. Yen et al., Exp. Cell Res., 229: 111-125, 1996) expressing low (wild-type HL-60), intermediate, and high cell surface c-FMS were treated with various concentrations of retinoic acid. The lowest concentration tested, 10(-8) M, induced significant differentiation of only the high c-FMS-expressing cells, with no accompanying hypophosphorylated RB or G0 arrest. The low and intermediate c-FMS expressing cells showed no induced differentiation, hypophosphorylation of RB, or G0 arrest. A 10-fold higher retinoic acid concentration, 10(-1) M, induced significant differentiation of both intermediate and high c-FMS-expressing cells. It induced RB hypophosphorylation only in high c-FMS-expressing cells but with no accompanying G0 arrest in any of the cells. The highest retinoic acid concentration, 10(-6) M, elicited differentiation, hypophosphorylation of RB, and G0 arrest in low, intermediate, and high c-FMS-expressing cells. As the concentration of retinoic acid increased, cell differentiation, hypophosphorylation of RB, and G0 arrest were progressively elicited within this ensemble of cells with different c-FMS expression levels. Thus, for example, at the lowest concentration of retinoic acid, expression of high enough c-FMS still allowed differentiation. At higher concentrations, progressively less c-FMS was needed for differentiation. The apparent threshold for the sum of the retinoic acid plus c-FMS originated signals to elicit differentiation, hypophosphorylation of RB, and G0 arrest increased, in that order. Thus retinoic acid-induced cell differentiation, RB hypophosphorylation, and G0 arrest have different signal threshold requirements. 1,25-Dihydroxy vitamin D3, also a ligand for a member of the steroid thyroid hormone receptor superfamily, caused monocytic differentiation with a similar c-FMS dependency, indicating that these effects characterize both myeloid and monocytic differentiation.     

A randomised dose escalation study of subcutaneous interleukin 2 with and without levamisole in patients with metastatic renal cell carcinoma or malignant melanoma

We have examined the efficacy, toxicity and host immunological response of two different dose schedules of interleukin 2 (IL-2) given subcutaneously, daily for 3 months in patients with renal cell carcinoma (RCC) or metastatic melanoma (MM). We also examined the effect of adding the immune modulator levamisole to the two different schedules of IL-2. Thirty-nine patients were entered into two sequential phase I/II studies. Eighteen patients entered study 1 and were randomised to receive IL-2, 3 x 10(6) IU m-2 day-1, subcutaneously for 3 months with or without levamisole 50 mg t.d.s. p.o. on days 1-3 on alternate weeks. Twenty-one patients entered study 2 and were randomised to receive 5.4 x 10(6) IU m-2 day-1 subcutaneously for 3 months with or without levamisole 50 mg t.d.s. p.o. on days 1-3 on alternate weeks. Blood was taken for peripheral blood lymphocyte (PBL) phenotype analysis, and measurement of IL-2, soluble IL-2 receptor (sIL-2R) and neopterin concentration. Two patients with metastatic melanoma, one in each study, responded (11.8%); both received IL-2 alone. Observations of immunological parameters showed that treatment with subcutaneous IL-2 resulted in a significant rise in the percentage of PBLs bearing CD25, CD3/HLA-DR, CD56 and levels of IL-2 receptor and neopterin. The total white blood cell count (WBC) and total lymphocyte count rose significantly on day 18 compared with pretreatment levels. The addition of levamisole to either IL-2 schedule resulted in no significant changes in any immunological parameters. This study illustrates that prolonged subcutaneous IL-2 can be given safely in the outpatient setting. There was no evidence that levamisole acts as an immunomodulator in this study.     

Effect of buthionine sulfoximine, a synthesis inhibitor of the antioxidant glutathione, on the murine nigrostriatal neurons

This study analyzed the effects of acute systemic treatment with buthionine sulfoximine (BSO), a synthesis inhibitor of the antioxidant reduced glutathione (GSH), on dopaminergic neurons of the murine nigrostriatal pathway. Part 1 of the study established a dose-response curve and the temporal pattern of GSH loss and recovery in the substantia nigra and striatum following acute BSO treatment. Part 2 of the study determined the effect of acute BSO treatment on the morphology and biochemistry of nigrostriatal neurons. We found that decreases in GSH levels had profound morphological effects, including decreased catecholamine fluorescence per cell, increased levels of lipid peroxidation and lipofuscin accumulation, and increased numbers of dystrophic axons in dopaminergic neurons of the nigrostriatal pathway. However, no measurable effects were observed in biochemical levels of either dopamine or its metabolites. These changes mimic those that have been reported to occur in the nigrostriatal system of rodents with advancing age. Our data suggest that reduction of GSH via BSO treatment results in the same types of nigrostriatal degenerative effects that occur during the aging process and consequently is a good model system for examining the role of GSH in protecting this area of the brain against the harmful effects of age-related oxidative stress.     

Transfer of leptophos in hen eggs and tissues of embryonic rats

This study reports the relations among normal aging, intelligence, and frontal lobe functioning. Intelligence tasks and frontal lobe functioning tasks were administered to 107 adults from two age groups (25 to 46 years and 70 to 99 years). Intelligence measures were assessed with two crystallized tests (WAIS Vocabulary and Information subtests), one fluid intelligence test (Cattell's Matrices), and one mixed, crystallized and fluid test (WAIS Similarities subtest). Frontal functioning was assessed using the Wisconsin Card Sorting Test (WCST) and two tests of verbal fluency. Significant age differences in favor of the young were found on the two intelligence tests with a fluid component and on all measures of frontal lobe functioning. Correlational analyses examining the relationship of intelligence measures to frontal variables indicated that these last measures were significantly correlated with only fluid intelligence tests in the elderly group. The implications for the relations among aging, fluid intelligence, and frontal lobe functioning are discussed.     

Growth hormone-binding protein in normal and pathologic gestation: correlations with maternal diabetes and fetal growth

To date, measurements of GH-binding protein (GHBP) during human pregnancy have been carried out using assays susceptible to interference by the elevated levels of human placental GH typical of late gestation. We recruited a large cohort of pregnant women (n = 140) for serial measurements of GHBP and used the ligand immunofunctional assay for GHBP. For normal gravidas, GHBP levels fell throughout gestation. Mean levels were 1.07 nmol/L (SE = 0.18) in the first trimester, 0.90 nmol/L (SE = 0.08) at 18-20 weeks, 0.73 nmol/L (SE = 0.05) at 28-30 weeks, and 0.62 nmol/L (SE = 0.06) at 36-38 weeks. GHBP levels in the first trimester correlated significantly with maternal body mass index (r = 0.58; P < 0.01). GHBP levels in pregnancies complicated by noninsulin-dependent diabetes mellitus (NIDDM) were substantially elevated at all gestational ages. The mean value in the first quarter (2.29 nmol/L) was more than double the normal mean (P < 0.01). In contrast, patients with insulin-dependent diabetes mellitus (IDDM) showed reduced GHBP concentrations at 36-38 weeks. The correlation between body mass index and GHBP is consistent with a metabolic role for GHBP during pregnancy, as is the dramatic elevation in GHBP observed in cases of NIDDM. At 36 weeks gestation, GHBP was significantly elevated (P < 0.01) in those women whose neonates had low birth weight (< 10th percentile). In early gestation (< 14 weeks), GHBP tended to be higher in women whose fetuses were designated to be at risk of intrauterine growth retardation (1.39 nmol/L; n = 4; compared with 1.07 nmol/L in normals), but this did not reach statistical significance. Although both NIDDM and IDDM pregnancies are at risk of fetal macrosomia, their GHBP concentrations are markedly divergent. This paradox and the roles of glucose and insulin in the regulation of GHBP during gestation warrant further investigation.     

Mechanisms of innate resistance to Toxoplasma gondii infection

The interaction of protozoan parasites with innate host defences is critical in determining the character of the subsequent infection. The initial steps in the encounter of Toxoplasma gondii with the vertebrate immune system provide a striking example of this important aspect of the host-parasite relationship. In immuno-competent individuals this intracellular protozoan produces an asymptomatic chronic infection as part of its strategy for transmission. Nevertheless, T. gondii is inherently a highly virulent pathogen. The rapid induction by the parasite of a potent cell-mediated immune response that both limits its growth and drives conversion to a dormant cyst stage explains this apparent paradox. Studies with gene-deficient mice have demonstrated the interleukin-12 (IL-12)-dependent production of interferon gamma (IFN-gamma) to be of paramount importance in controlling early parasite growth. However, this seems to be independent of nitric oxide production as mice deficient in inducible nitric oxide synthase (iNOS) and tumour necrosis factor receptor were able to control early growth of T. gondii, although, they later succumbed to infection. Nitric oxide does, however, seem to be important in controlling persistent infection; treating chronic infection with iNOS metabolic inhibitors results in disease reactivation. Preliminary evidence implicates neutrophils in effector pathways against this parasite distinct from that described for macrophages. Once initiated, IL-12-dependent IFN-gamma production in synergy with other proinflammatory cytokines can positively feed back on itself to induce 'cytokine shock'. Regulatory cytokines, particularly IL-10, are essential to down-regulate inflammation and limit host pathology.     

Nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mouse as a model system to study the engraftment and mobilization of human peripheral blood stem cells

Mobilized CD34(+) cells from human peripheral blood (PB) are increasingly used for hematopoietic stem-cell transplantation. However, the mechanisms involved in the mobilization of human hematopoietic stem and progenitor cells are largely unknown. To study the mobilization of human progenitor cells in an experimental animal model in response to different treatment regimens, we injected intravenously a total of 92 immunodeficient nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice with various numbers of granulocyte colony-stimulating factor (G-CSF) -mobilized CD34(+) PB cells (ranging from 2 to 50 x 10(6) cells per animal). Engraftment of human cells was detectable for up to 6.5 months after transplantation and, depending on the number of cells injected, reached as high as 96% in the bone marrow (BM), displaying an organ-specific maturation pattern of T- and B-lymphoid and myeloid cells. Among the different mobilization regimens tested, human clonogenic cells could be mobilized from the BM into the PB (P = .019) with a high or low dose of human G-CSF, alone or in combination with human stem-cell factor (SCF), with an average increase of 4.6-fold over control. Therefore, xenotransplantation of human cells in NOD/SCID mice will provide a basis to further study the mechanisms of mobilization and the biology of the mobilized primitive human hematopoietic cell.     

Improvement of arterial oxygenation by selective infusion of prostaglandin E1 to ventilated lung during one-lung ventilation

Stride length, stance width and gait symmetry were measured as parameters of gait analysis in young (5 months) and old (28 months) Sprague-Dawley rats following 3 months of treadmill running as well as in sedentary controls. Stride length decreased significantly with age and increased significantly following training in both young and old rats. Means of stance width increased significantly with age, but were unaffected by training. Gait symmetry declined with age; however, treadmill running counteracted the decline of gait symmetry. It was concluded that treadmill running compensates for or prevents age-related changes in the neuromuscular function of rats, which lead to gait deterioration.     

Sequential hemodynamic changes for large volume paracentesis in post-hepatitic cirrhotic patients with massive ascites

Large volume paracentesis (4.8 to 15.5 liters) was performed in 42 patients with post-hepatitic cirrhosis and massive ascites, not only to derive parameters capable of predicting the development of severe clinical hypotension after large volume paracentesis, but also to determine the optimal time to introduce preventive volume expanders. Systemic hemodynamics were sequentially measured for 72 hours in thirty-two patients. Severe clinical hypotension occurred in 13 (31.0%) patients 4-62 hours from the start of paracentesis. Univariate analysis, with the Mantel-Cox test used to compare Kaplan-Meier curves, and the subsequent multivariate analysis by stepwise Cox regression procedure were utilized to identify two variables, withdrawn ascitic fluid greater than 7.5 liters (p = 0.0121) and the absence of peripheral edema (p = 0.0148), reaching statistical significance to predict the occurrence of severe clinical hypotension. Compared to the baseline value, the cardiac output of patients not developing severe clinical hypotension increased (6.26 +/- 0.66 vs. 6.65 +/- 0.69 liter/min, p < 0.01) one hour from the start of paracentesis and right atrial pressure decreased (11.2 +/- 2.4 vs. 8.7 +/- 2.3 mmHg, p < 0.05). The cardiac output returned to the baseline value at the 9th hour. Based on the results presented herein, we can conclude that severe clinical hypotension occurs in a high percentage of patients with post-hepatitic cirrhosis and massive ascites within 72 hours from the start of large volume paracentesis. At potential risk of this occurring are those patients without peripheral edema and withdrawn ascitic fluid greater than 7.5 liters. Volume expanders should be introduced before 4th hour from the start of large volume paracentesis.