Blood thymidine level and iododeoxyuridine incorporation and reutilization in DNA in mice given long-acting thymidine pellets.
A long-acting thymidine pellet consisting of 190 mg of cholesterol and 60 mg of thymidine has been developed for the study of thymidine metabolism and reutilization in vivo. Implantation of such a pellet s.c. in adult mice will maintain the blood plasma concentration of thymidine at levels between 40 and 8 X 10(-6) M, which are from 36 to 7 times those of normal mice, for periods up to 48 hr. During this period, in vivo uptake and reutilization of [125I]iododeoxyuridine, a thymidine analog, into intestinal and tumor DNA were almost completely suppressed. While iododeoxyuridine reutilization is not large in normal proliferative tissue even in the absence of pellet implants, reutilization of over 30% was measured in large, rapidly growing ascites tumors. The inhibition of iododeoxyuridine incorporation by elevated thymidine blood levels is directly proportional to serum concentration. This appears to be due to a thymidine pool in rapid equilibrium with blood thymidine. This pool is at least 10 times larger than the 4-nmole pool of extracellular thymidine. (+info)
Tissue pharmacokinetics, inhibition of DNA synthesis and tumor cell kill after high-dose methotrexate in murine tumor models.
In Sarcoma 180 and L1210 ascites tumor models, the initial rate of methotrexate accumulation in tumor cells in the peritoneal cavity and in small intestine (intracellularly) after s.c. doses up to 800 mg/kg, showed saturation kinetics. These results and the fact that initial uptake in these tissues within this dosage range was inhibited to the expected relative extent by the simultaneous administration of leucovorin suggest that carrier mediation and not passive diffusion is the major route of drug entry at these extremely high doses. Maximum accumulation of intracellular drug occurred within 2 hr and reached much higher levels in small intestine than in tumor cells at the higher dosages. At a 3-mg/kg dose of methotrexate s.c., intracellular exchangeable drug levels persisted more than four times longer in L1210 cells than in small intestine, but differences in persistence (L1210 cell versus gut) diminished markedly with increasing dosage. At 96 mg/kg, the difference in persistence was less than 2-fold. In small intestine and L1210 cells, theduration of inhibition of DNA synthesis at different dosages correlated with the extent to which exchangeable drug was retained. Toxic deaths occurred when inhibition in small intestine lasted longer than 25 to 30 hr. Recovery of synthesis in small intestine and L1210 cells occurred synchronously and only below dosages of 400 mg/kg. Within 24 hr after dosages of greater than 24 mg/kg, the rate of tumor cell loss increased to a point characterized by a single exponential (t1/2=8.5 hr). The total cell loss, but not the rate of cell loss, was dose dependent. (+info)
Lung weight parallels disease severity in experimental coccidioidomycosis.
Evidence provided by histopathological study of lesions is a valuable adjunct for evaluating chemotherapeutic efficacy in experimental animal models, In addition, this should be correlated with a measure of disease severity in the same animal. The latter could be obtained by homogenization of infected organs and quantitative enumeration of viable cells of the etiological agent, but this would preclude histopathological studies in the same animal. Progression of disease in pulmonary infection is associated with replacement of air space by fluid, cells, and cellular debris. Therefore, an increase in lung weight should reflect severity of disease. Results with the murine model of coccidioidomycosis demonstrate that increasing lung weight parallels the increasing census of fungus cells in the lungs of both treated and nontreated infected mice. This was supported with evidence obtained from microscopic studies of lesions indicating that specific chemotherapy limited spread of the infection and inhibited multiplication of the fungus in the lung. Therefore, lung weight can be used as a measure of disease severity in the murine model of coccidioidomycosis. (+info)
Prevention of collagen-induced arthritis by gene delivery of soluble p75 tumour necrosis factor receptor.
Collagen type II-induced arthritis (CIA) in DBA/1 mice can be passively transferred to SCID mice with spleen B- and T-lymphocytes. In the present study, we show that infection ex vivo of splenocytes from arthritic DBA/1 mice with a retroviral vector, containing cDNA for the soluble form of human p75 receptor of tumour necrosis factor (TNF-R) before transfer, prevents the development of arthritis, bone erosion and joint inflammation in the SCID recipients. Assessment of IgG subclass levels and studies of synovial histology suggest that down-regulating the effector functions of T helper-type 1 (Th1) cells may, at least in part, explain the inhibition of arthritis in the SCID recipients. In contrast, the transfer of splenocytes infected with mouse TNF-alpha gene construct resulted in exacerbated arthritis and enhancement of IgG2a antibody levels. Intriguingly, infection of splenocytes from arthritic DBA/1 mice with a construct for mouse IL-10 had no modulating effect on the transfer of arthritis. The data suggest that manipulation of the immune system with cytokines, or cytokine inhibitors using gene transfer protocols can be an effective approach to ameliorate arthritis. (+info)
Tissue-specific knockout of the insulin receptor in pancreatic beta cells creates an insulin secretory defect similar to that in type 2 diabetes.
Dysfunction of the pancreatic beta cell is an important defect in the pathogenesis of type 2 diabetes, although its exact relationship to the insulin resistance is unclear. To determine whether insulin signaling has a functional role in the beta cell we have used the Cre-loxP system to specifically inactivate the insulin receptor gene in the beta cells. The resultant mice exhibit a selective loss of insulin secretion in response to glucose and a progressive impairment of glucose tolerance. These data indicate an important functional role for the insulin receptor in glucose sensing by the pancreatic beta cell and suggest that defects in insulin signaling at the level of the beta cell may contribute to the observed alterations in insulin secretion in type 2 diabetes. (+info)
The cerebral metabolic consequences of nitric oxide synthase deficiency: glucose utilization in endothelial and neuronal nitric oxide synthase null mice.
Nitric oxide has multiple physiologic roles in the CNS. Inhibiting nitric oxide synthesis might therefore alter functional activity within the brain. We used [14C]-2-deoxyglucose in vivo autoradiography to measure local CMRglc in "knockout" mice lacking the genes for either the endothelial (eNOS) or neuronal (nNOS) isoforms of nitric oxide synthase, and in the progenitor strains (SV129, C57B1/6). Glucose utilization levels did not significantly differ between nNOS and eNOS knockout mice and C57B1/6 mice in any of the 48 brain regions examined, but were relatively lower in some subcortical regions in SV129 mice. (+info)
T cell subsets in experimental lupus nephritis: modulation by bacterial superantigen.
Chronic graft-vs-host disease (GvH), induced by injection of DBA/2 lymphocytes into (C57BL/6 x DBA/2)F1 hybrids, is a murine model for lupus nephritis, associated with a Th2-dependent polyclonal B cell activation. The development of glomerulosclerosis in this model is preceded by a glomerular influx of LFA-1+ T cells. We investigated whether exposure to bacterial superantigen would modulate the course of this autoimmune syndrome. Injection of the bacterial superantigen staphylococcal enterotoxin B (SEB) in mice has been shown to induce the activation of TcRVbeta8+ T cells. Within 2 weeks after GvH induction, mice were injected twice with 20 microg of SEB and the following parameters were examined: cytokine and Ig profile, proteinuria and renal pathology. The second SEB injection induced in GvH mice an increased release of both interferon-gamma (IFN-gamma) and interleukin-10 (IL-10) as compared with control F1 mice. No differences were observed in IL-2 production. SEB-treated GvH mice demonstrated a delayed onset of proteinuria. Histological analysis of the kidney showed that SEB-challenged GvH mice displayed significantly more interstitial inflammation and mesangial proliferation together with more IgG2a deposits in glomeruli than non-injected GvH mice. From these results, we conclude that GvH mice are more responsive to SEB in terms of cytokine production and that bacterial infection can modulate the course of this renal disease from a membranous to a more proliferative type of nephropathy. (+info)
Development and function of autospecific dual TCR+ T lymphocytes.
Recent studies have challenged the long held concept that each T lymphocyte expresses on its surface only a single, unique alphabetaTCR. Dual TCR+ T cells have been recognized, however, their origin and potential to escape screening for self-reactivity remain obscure. We now report the thymic generation of dual alphabetaTCR+ T cells in the H-2Db/H-Y-specific TCR transgenic (Tg) mouse. Dual TCR+ thymocytes were positively selected less efficiently than single TCR+ thymocytes, although a subset attained maturity. Importantly, when TCR Tg mice were bred onto a negatively selecting background, auto-specific cells survived central deletion and matured as CD4+ dual TCR+ cells. These cells were autoreactive when CD8 expression was restored. The existence of autospecific, dual TCR+ T cells may have implications for the maintenance of self tolerance. (+info)