Differential requirement for p56lck in fetal and adult thymopoiesis

The protein tyrosine kinase p56lck is critical for the generation of mature thymocytes in adult mice. However its requirement during the maturation of thymocytes from the fetal to the adult stage has not been clearly defined. We analyzed prenatal and postnatal thymocyte maturation in mice deficient for p56lck (lck[-/-]). Before birth, lck appears to play a crucial role in the expansion and proliferation of CD4+CD8+ double positive thymocytes, whereas proliferation and absolute numbers of CD4-CD8- double negative thymocyte precursors remained within the normal range until the end of the second week postnatal. Three weeks after birth, the total numbers of double negative and immature single positive thymocytes underwent a dramatic reduction that correlated with a decrease in the double positive population. This ontogenic defect was associated with a significant decrease in the proliferation rates of thymocyte precursors. Our data suggest that signaling via p56lck kinase is differentially required within a given phenotypically defined thymocyte subpopulation, depending on its stage of thymocyte maturation.     

Compartmental pharmacokinetics and tissue drug distribution of the pradimicin derivative BMS 181184 in rabbits

The pharmacokinetics of the antifungal pradimicin derivative BMS 181184 in plasma of normal, catheterized rabbits were characterized after single and multiple daily intravenous administrations of dosages of 10, 25, 50, or 150 mg/kg of body weight, and drug levels in tissues were assessed after multiple dosing. Concentrations of BMS 181184 were determined by a validated high-performance liquid chromatography method, and plasma data were modeled into a two-compartment open model. Across the investigated dosage range, BMS 181184 demonstrated nonlinear, dose-dependent kinetics with enhanced clearance, reciprocal shortening of elimination half-life, and an apparently expanding volume of distribution with increasing dosage. After single-dose administration, the mean peak plasma BMS 181184 concentration (Cmax) ranged from 120 microg/ml at 10 mg/kg to 648 microg/ml at 150 mg/kg; the area under the concentration-time curve from 0 to 24 h (AUC0-24) ranged from 726 to 2,130 microg . h/ml, the volume of distribution ranged from 0.397 to 0.799 liter/kg, and the terminal half-life ranged from 4.99 to 2.31 h, respectively (P < 0.005 to P < 0.001). No drug accumulation in plasma occurred after multiple daily dosing at 10, 25, or 50 mg/kg over 15 days, although mean elimination half-lives were slightly longer. Multiple daily dosing at 150 mg/kg was associated with enhanced total clearance and a significant decrease in AUC0-24 below the values obtained at 50 mg/kg (P < 0.01) and after single-dose administration of the same dosage (P < 0.05). Assessment of tissue BMS 181184 concentrations after multiple dosing over 16 days revealed substantial uptake in the lungs, liver, and spleen and, most notably, dose-dependent accumulation of the drug within the kidneys. These findings are indicative of dose- and time-dependent elimination of BMS 181184 from plasma and renal accumulation of the compound after multiple dosing.     

Treatment of thyroid disease in pregnancy

With appropriate therapy, complications related to thyroid disease in pregnancy can be minimized. Although the diagnosis of thyroid endocrinopathy may be difficult in pregnancy, few therapies are contra-indicated. Because medications may cross the placenta, however, clinicians need always to be mindful of potential fetal effects and should work to use the minimal dose necessary to achieve maternal euthyroidism. Thyroid function tests, in particular free T4 and TSH, remain good measures of thyroid function and therapy in pregnancy.     

Comparison of nociceptive effects produced by intrathecal administration of mGluR agonists

The present study examined the mGluR subtypes involved in (1S, 3R)-ACPD-induced spontaneous nociceptive behaviours (SNB) by administering the following selective agonists by the intrathecal (i.t.) route: (RS)-DHPG, trans-ADA (Group I; mGluR1/5 and mGluR5, respectively), (1S, 3S)-ACPD, (2R, 4R)-APDC (Group II), and L-AP4 (Group III). (RS)-DHPG administration induced SNB that were of significantly greater intensity and longer duration than those induced by an equal dose of (1S, 3R)-ACPD. No other agonists produced SNB, except (1S, 3S)-ACPD, which may be attributable to a nonselective action at mGluR1. Intrathecal treatment with the mGluR antagonist (+)-MCPG or the NMDA antagonist D-AP5 prior to (RS)-DHPG administration dose-dependently reduced SNB. It is suggested that a possible interaction between NMDA and mGluR1 is a critical event in the maintenance of persistent nociception.     

Differential TNF secretion by wound fibroblasts compared to normal fibroblasts in response to LPS

Fibroblasts cultured from wound sites have been shown to have an altered phenotype compared to normal dermal fibroblasts and are generally regarded as target cells of the cytokine response at sites of injury. This study was undertaken to determine whether wound fibroblasts can contribute to proinflammatory cytokine production in wounds and, in particular, whether they are capable of secreting TNF. Wound fibroblasts were cultured from polyvinyl alcohol sponges implanted subcutaneously for 2 weeks in Balb/c mice. Fibroblasts harvested from the skin and subcutaneous tissue of untreated mice served as a control population of cells. All cells were passaged at least twice and then stimulated with a dose range of LPS. Supernatants were harvested 8 hr following stimulation and TNF was assayed using a standard L929 cell-killing assay. There was a significant TNF response to LPS by wound fibroblasts, evident as early as 4 hr following exposure to LPS and associated with an upregulation of TNF mRNA. Normal dermal fibroblasts did not secrete any measurable amounts of TNF in response to LPS. The results indicate that wound fibroblasts generate a brisk TNF response to stimulation with LPS, in contrast to normal subcutaneous fibroblasts. These data reveal an additional unique property of wound-harvested fibroblasts and suggest a possible contributing mechanism to disordered wound healing in the face of infection or conditions characterized by excessive fibrosis.     

Nitric oxide-related vasoconstriction in lungs perfused with red cell lysate

The present study in isolated rat lungs demonstrates that nitric oxide gas (.NO, 70 nM) added to the perfusate containing a small amount of hemolysate [175 microliters of lysed red blood cells (RBC) per 50 ml of Earle's balanced salt solution (EBSS)] triggered profound and sustained vasoconstriction. Vasoconstriction was not observed when .NO was added to lungs perfused with washed intact rat or human RBC or with oxyhemoglobin (Hgb 20 microM). The presence of hemolysate in the perfusate also caused vasoconstriction in response to n-acetylcysteine (50 microM), glutathione (10(-4) M), or ascorbic acid (10(-4) M) and potentiated greatly the vasoconstrictor response to 5 mM KCl. Not only .NO, but also nitroprusside (SNP) or L-arginine and paradoxically three .NO synthesis inhibitors, including N-monomethyl L-arginine, L-NAME, and nitroblue tetrazolium, which have different mechanisms of action, each caused in the presence of hemolysate large vasoconstrictive responses. Hemolysate itself enhanced O2 consumption by slices of lung; no effects of this dose of .NO on lung slice respiration were seen in the absence of hemolysate. Both Hgb and hemolysate lowered perfusate cGMP levels to the same degree suggesting that the vasoconstrictive response was not due to unique effects of hemolysate on guanylyl cyclase. Addition of superoxide dismutase (SOD) and catalase (CAT) to the hemolysate containing perfusate, or addition of a cyclooxygenase or 5-lipoxygenase inhibitor, virtually abolished the .NO induced vasoconstriction. The latter data are consistent with the concept that exposure of the vasculature to hemolysate may result in the formation of peroxynitrite. However, SOD and CAT did not abolish the pulmonary vasoconstriction induced by L-arginine or by NAC. Our data indicate that hemolysate has profound effects on lung vessel tone regulation and on lung tissue mitochondrial function, yet the precise molecular mechanisms responsible for the action of hemolysate are likely to be very complex.     

Multideterminant role of calcium in hippocampal synaptic plasticity

Hippocampal CA1 cells possess several varieties of long-lasting synaptic plasticity: two different forms of long-term potentiation (LTP) and at least one form of long-term depression (LTD). All forms of synaptic plasticity are induced by afferent activation, all involve Ca2+ influx, all can be blocked by Ca2+ chelators, and all activate Ca(2+)-dependent mechanisms. The question arises as how different physiological responses can be initiated by activation of the same second messenger. We consider two hypotheses which could account for these phenomena: voltage-dependent differences in cytosolic Ca2+ concentration acting upon Ca2+ substrates of differing Ca2+ affinities and compartmentalization of the Ca2+ and its substrates.