Bicyclic amidine inhibitors of nitric oxide synthase: discovery of perhydro-iminopyrindine and perhydro-iminoquinoline as potent, orally active inhibitors of inducible nitric oxide synthase

Syntheses and nitric oxide synthase inhibitory activity of cyclic amidines containing 5,6- 6,6- and 7,6-fused systems are described. X-ray structure determination facilitated the assignment of the stereochemistry of the most active compounds perhydro-2-iminoisoquinoline (8a) and perhydro-2-iminopyrindine (10a). Both 8a and 10a are very potent inhibitors of iNOS, with excellent selectivity over eNOS and they are orally active in rats with long duration suitable for once or twice a day dosing.     

Asterocheres crinoidicola n. sp., a copepod (Siphonostomatoida: Asterocheridae) parasitic on crinoids in Belize

A new species of siphonostomatoid copepod, Asterocheres crinoidicola, is parasitic on two closely related comasterid crinoids (Nemaster grandis and Davidaster rubiginosus) in Belize, Central America. An unusually long terminal prolongation of the third segment of the endopod of leg 1 distinguishes this species from all congeners. This is the first report of a copepod parasitic on a crinoid in the Caribbean.     

Structural requirements for the binding of prostaglandins

The binding of prostaglandins and analogs to the lipocyte PGE receptor was shown to exhibit a high degree of structural specificity. Small changes, particularly at the 9-keto or 15-hydroxy position, were found to drastically diminish interaction with the receptor. Studies of a rather substantial number of compounds revealed a close relationship between affinity for the lipocyte PGE receptor and the ability to stimulate cyclic AMP synthesis in the isolated mouse ovary. In general, activities in these two parameters follow the biological potencies generally recognized for these compounds.     

Apoptosis in C3H-10T1/2 cells: Roles of intracellular pH,protein kinase C,and the Na+/H+ antiporter

Changes in intracellular ion concentrations have been correlated with the activation of an endogenous endonuclease and thus internucleosomal DNA cleavage during apoptosis in many cell types. We investigated whether intracellular pH could play a significant role in apoptotic initiation and progression in C3H-10T1/2 cells, a cell strain that does not exhibit double-stranded DNA cleavage during apoptosis. Protein kinase C and the Na⁺/H⁺ antiporter, known regulators of intracellular pH, also were assessed for their involvement in apoptosis of C3H-10T1/2 cells. When a H⁺ ionophore was used to clamp intracellular pH to 6.0 or below, a significant level of apoptosis was induced in these cells within 6 h, whereas clamping at pH 6.75 did not induce significant amounts of apoptosis until 36 h after acidification. The acidified cells exhibited classic apoptotic morphology and chromatin condensation, similar to serum withdrawn cells, but failed to show internucleosomal DNA cleavage with electrophoresis of genomic DNA. Our results also suggest that the 12-O-tetradecanoylphorbol-13-acetate (TPA)-mediated inhibition of apoptosis in serum withdrawn C3H-10T1/2 cells functions through a sequential activation of protein kinase C and the Na⁺/H⁺ antiporter; thus, an alkalinization or an inhibition of acidification is involved in this apoptotic block. Serum withdrawal itself does not appear to act through a negative effect on either protein kinase C or the Na⁺/H⁺ antiporter. TPA was also capable of inhibiting the apoptosis induced by specific inhibitors of protein kinase C and the Na⁺/H⁺ antiporter, but the inhibition was successful only if the TPA was administered at least 20 min prior to the addition of the enzyme inhibitor. These results indicate that apoptosis in C3H-10T1/2 cells follows a pathway that involves intracellular acidification, but is independent of detectable endonuclease activity. J. Cell. Biochem. 67:231–240, 1997. © 1997 Wiley-Liss, Inc.     

Mitochondrial bioenergetics during the initiation of mercuric chloride-induced renal injury. I. Direct effects of in vitro mercuric chloride on renal mitochondrial function

Increasing data suggest that mitochondrial dysfunction may be an important early component of nephrotoxin-induced changes in renal cell function and viability. This study was designed to obtain more detailed information about the effects on several basic bioenergetic parameters of the direct interaction of Hg2+ with renal cortical mitochondria in vitro as a necessary prelude to studies of mitochondrial functional changes after treatment with mercuric chloride in vivo. Beginning at a threshold level of 2 nmol of Hg2+/mg of mitochondrial protein, Hg2+ induced marked stimulation of State 4 respiration, mild inhibition of State 3 respiration, and 2,4-dinitrophenol uncoupled respiration, a striking increase in atractyloside-insensitive ADP uptake and stimulation of both basal- and Mg2+-activated oligomycin-sensitive mitochondrial ATPase activity. These effects of Hg2+ could be prevented and reversed by the sulfhydryl reagent dithioerythritol and by albumin but were not affected by Mg2+. Detailed studies on the addition of HgCl2 to the preparation at different stages of the mitochondrial isolation procedure demonstrated that the presence of other proteins decreased mitochondrial Hg2+ binding, that the Hg2+ was not readily washed off the mitochondria by nonprotein-containing solutions, and that prolonged exposure of mitochondria to Hg2+ during the isolation procedure did not markedly alter its functional effects or their reversibility as assessed on the final mitochondrial preparation. These data provide an important basis for critically assessing the changes in function of mitochondria isolated after in vivo treatment with mercuric chloride.     

Adaptation of subtype a human immunodeficiency virus type 1 envelope to pig-tailed macaque cells

The relevance of simian/human immunodeficiency virus (SHIV) infection of macaques to HIV-1 infection in humans depends on how closely SHIVs mimic HIV-1 transmission, pathogenesis, and diversity. Circulating HIV-1 strains are predominantly subtypes C and A and overwhelmingly require CCR5 for entry, yet most SHIVs incorporate CXCR4-using subtype B envelopes (Envs). While pathogenic subtype C-based SHIVs have been constructed, the subtype A-based SHIVs (SHIV-As) constructed to date have been unable to replicate in macaque cells. To understand the barriers to SHIV-A replication in macaque cells, HIVA(Q23)/SIV(vif) was constructed by engineering a CCR5-tropic subtype A provirus to express SIV vif, which counters the macaque APOBEC3G restriction. HIVA(Q23)/SIV(vif) replicated poorly in pig-tailed macaque (Ptm) lymphocytes, but viruses were adapted to Ptm lymphocytes. Two independent mutations in gp120, G312V (V3 loop) and A204E (C2 region), were identified that increased peak virus levels by >100-fold. Introduction of G312V and A204E to multiple subtype A Envs and substitution of G312 and A204 with other residues increased entry into Ptm cells by 10- to 100-fold. G312V and A204E Env variants continued to require CCR5 for entry but were up to 50- and 200-fold more sensitive to neutralization by IgG1b12 and soluble CD4 and had a 5- to 50-fold increase in their ability to utilize Ptm CD4 compared to their wild-type counterparts. These findings identify the inefficient use of Ptm CD4 as an unappreciated restriction to subtype A HIV-1 replication in Ptm cells and reveal amino acid changes to gp120 that can overcome this barrier.     

Two common nonsynonymous paraoxonase 1 (<Emphasis Type="Italic">PON1</Emphasis>) gene polymorphisms and brain astrocytoma and meningioma

Background  

Human serum paraoxonase 1 (PON1) plays a major role in the metabolism of several organophosphorus compounds. The enzyme is encoded by the polymorphic gene PON1, located on chromosome 7q21.3. Aiming to identify genetic variations related to the risk of developing brain tumors, we investigated the putative association between common nonsynonymous PON1 polymorphisms and the risk of developing astrocytoma and meningioma.     

Evaluation of pyrrolidin-2-imines and 1,3-thiazolidin-2-imines as inhibitors of nitric oxide synthase

Syntheses and evaluation of pyrrolidin-2-imines and 1,3-thiazolidin-2-imines as inhibitors of nitric oxide synthase (NOS) are discussed. An extensive SAR was established for pyrrolidin-2-imines class of compounds. The amidines came out as the most potent inhibitors in addition to displaying selectivity.     

Antigen-antibody complexes stimulate the synthesis and release of prostaglandins by mouse peritoneal macrophages

Antigen-antibody complexes (Ag/Ab) formed at equivalence stimulate the release of arachidonic acid and synthesis of prostaglandin E₂ and 6-keto-prostaglandin F_1α by resident mouse peritoneal macrophages. Prostaglandin synthesis and secretion is stimulated by submicrogram quantities of Ag/Ab which increases in a dose-dependent manner. This release is time-dependent and occurs in the absence of any loss of cell viability as indicated by increased cellular levels of lactate dehydrogenase without concomitant loss of this activity to the media and the continued secretion of a constitutive cellular product, lysozyme. The stimulated synthesis of prostaglandins by Ag/Ab is inhibited by indomethacin and physiological levels of antiinflammatory glucocorticoids.