Synthesis of two nitro analogs of tranylcypromine: Relations of aromatic substitution of nitro groups to MAO-Inhibitory activity

Two new nitro analogs of tranylcypromine, (E)-2-(p-nitrophenyl)cyclopropylamine ((E)-p-NTCP) and (E)-2-(m-nitrophenyl)cyclopropylamine ((E)-m-NTCP) were synthesized in order to examine the effect of aromatic nitro substitution on the MAO-inhibitory activity of 2-phenylcyclopropylamines. The compounds were obtained by treatingt-butyl (E)-2-(p-nitrophenyl) cyclopropanecarbamate andt-butyl (E)-2-(m-nitrophenyl)cyclopropanecarbamate withp-toluenesulfonic acid in CH₃CN. Inhibitions of rat brain mitochondrial MAO-A and B by the compounds were examined using serotonin and benzylamine as the substrate at bothin vitro andex vivo levels. It was found fromin vitro measurements that(E)-p-NTCP at 6.0×10^?5M elicited merely 22.5% inhibition against MAO-B without any effect on MAO-A. In contrast,(E)-m-NTCP showed fair degrees of inhibitions of MAO-A and B with IC₅₀ values, 2.5×10^?7M and 1.4×10^?6M, respectively. It was also noted from(E)-m-NTCP thatm-nitro substitution caused a shift of selectivity of the inhibition toward MAO-A. According toex vivo measurements at 1.5, 3, 6, and 12 hr following the administration of a dose of 0.015 mmol/kg, i.p. to the rats, the inhibition percents of MAO-A by(E)-m-NTCP were 58.6, 63.7 63.6, and 46.6%, slightly lower than those observed by tranylcypromine. Whereas,(E)-p-NTCP at the same dose level did not show significant inhibitions against both MAO-A and MAO-B. Possible reasons for the difference in potencies between(E)-m-NTCP and(E)-p-NTCP were sought in relation to differing electron withdrawing effects ofm-andp-substituents which will influence electron density of the side chain amino functions and the partitions.     

Structures of Flavamine and Flavadine from Aconitum flavum

Two new diterpenoid alkaloids, flavamine ( 1) and flavadine ( 2), were isolated from the roots of ACONITUM FLAVUM Hand-Mazz. The structures were established on the bases of spectral analyses and chemical correlations with napelline ( 3) and lucidusculine ( 4), respectively.     

Interaction of 2-halogenated dATP analogs (F, Cl, and Br) with human DNA polymerases, DNA primase, and ribonucleotide reductase

Recently, 2-halogenated deoxyadenosine analogs (F, Cl, and Br) have been shown to have antitumor activity. These analogs are phosphorylated by cells and are believed to exert their cytotoxic action at the nucleoside triphosphate level. In this work the interaction of these nucleoside triphosphate analogs with potential targets, such as DNA polymerase alpha, beta, and gamma, DNA primase, and ribonucleotide reductase was examined in detail. All of these compounds competitively inhibited the incorporation of dAMP into DNA by DNA polymerase alpha, beta, or gamma. F-dATP was able to completely substitute for dATP using DNA polymerase alpha and gamma, but not with DNA polymerase beta. Cl-dATP and Br-dATP substituted poorly for dATP using DNA polymerase alpha and beta. Extension of a 32P-labeled primer by DNA polymerase alpha, beta, or gamma on a single-stranded M13 template showed that these compounds were incorporated into the 3' end of the growing DNA chain and that elongation beyond the incorporated analogs was significantly retarded for Cl-dATP and Br-dATP using either DNA polymerase alpha or beta. DNA primase using poly(dC) as template was inhibited by these compounds at a concentration 4 to 5 times greater than that required for 2-F-araATP. The 2-halogenated dATP analogs were potent inhibitors of ADP reduction by ribonucleotide reductase. In conclusion, the cytotoxic action of 2-Cl-deoxyadenosine and 2-Br-deoxyadenosine may partially be mediated through the mechanism of "self-potentiation," by depression of the deoxynucleoside triphosphate pools due to inhibition of ribonucleotide reductase, which would facilitate their incorporation into DNA and result in the inhibition of DNA synthesis.     

宫颈粘液过氧化物酶在月经周期中的变化规律

本文对29例月经周期正常妇女的宫颈粘液过氧化物酶进行了30个周期的研究。在月经周期不同时间测定宫颈粘液过氧化物酶(CMPx)活性及血清促黄体生成素(LH)、雌二醇(E_2)和孕酮(P)。结果表明:在排卵前三天酶活性明显下降,至排卵后一天开始上升。卵泡期,酶活性与E_2呈负相关(r=-0.67);黄体期,酶活性与P呈正相关(r=0.79)。本研究提示:1.CMPx在排卵周期具有特定的变化规律,其变化受体内激素水平影响,可作为预告排卵的指标。2.如简化测定方法,可为自然避孕提供新途径。     

GALC transduction leads to morphological improvement of the twitcher oligodendrocytes in vivo

Globoid cell leukodystrophy (GLD, Krabbe disease) is a severe demyelinating disease caused by a genetic defect of beta-galactocerebrosidase (GALC). To date treatment to GLD is limited to hematopoietic stem cell transplantation. Experimental approaches by means of gene therapy in twitcher mouse, an authentic murine model of human GLD, showed significant but only marginal improvements of the disease. To clarify whether the introduction of GALC could provide beneficial effects on the oligodendrocytes in GLD, we transduced twitcher oligodendrocytes by stereotactically injecting recombinant retrovirus encoding GALC-myc-tag fusion gene into the forebrain subventricular zone of neonatal twitcher mouse. In vivo effects of exogenous GALC on twitcher oligodendrocytes were studied histologically by combined immunostaining for the myc-epitope and the oligodendroglial specific marker, pi form of glutathione-S-transferase, at around 40 days of age. We show here that GALC transduction led to dramatic morphological improvement of the twitcher oligodendrocytes comparing with those in untreated twitcher controls. This study provided direct in vivo evidence that GALC transduction could prevent or correct aberrant morphology of oligodendrocytes in GLD which may be closely related to the dysfunction and/or degeneration of oligodendrocytes and the demyelination in this disease.     

Identification and characterization of clonal NK-like cells from channel catfish (Ictalurus punctatus)

TcR alpha, beta, and gamma chain negative cytotoxic NK-like cells were cloned from alloantigen-stimulated PBL obtained from nai;ve channel catfish. Stimulation with allogeneic cells and growth promoting factors are required for their continued in vitro proliferation and cytotoxic activity. These granular cells kill not only the stimulating allogeneic cells, but also unrelated allogeneic targets by a perforin/granzyme-mediated apoptosis pathway. In addition, they are negative for markers that define neutrophils, monocytes/macrophages, and non-specific cytotoxic cells. Although these NK-like clones kill a number of different allogeneic targets, they display interclonal variation in cytotoxicity toward a panel of allogeneic targets, i.e. some clones have no apparent target specificity, while others display a target preference. In addition, flow cytometric analyses revealed that expression of a putative FcmuR, an LFA-1-like molecule, and a putative thymocyte/T cell antigen varies among the different clones, with no clear correlation between surface antigen expression and cytotoxic activity. Although not all clones express a putative FcmuR, it was noted that they all expressed an ITAM containing FcepsilonR gamma chain homolog. This finding suggests that the catfish FcepsilonR gamma chain may potentially be used as an accessory molecule for not only FcmuRs, but also for other unknown activation receptors. These results support the hypothesis that catfish NK-like cells are heterogeneous in terms of target specificities and cell surface phenotype.     

Macrophage activation and Fcgamma receptor-mediated signaling do not require expression of the SLP-76 and SLP-65 adaptors

The Src-homology 2 domain-containing, leukocyte-specific phosphoprotein of 76 kDa (SLP-76) is a hematopoietic adaptor that plays a central role during immunoreceptor-mediated activation of T lymphocytes and mast cells and collagen receptor-induced activation of platelets. Despite similar levels of expression in macrophages, SLP-76 is not required for Fc receptor for immunoglobulin G (IgG; FcgammaR)-mediated activation. We hypothesized that the related adaptor SLP-65, which is also expressed in macrophages, may compensate for the loss of SLP-76 during FcgammaR-mediated signaling and functional events. To address this hypothesis, we examined bone marrow-derived macrophages (BMM) from wild-type (WT) mice or mice lacking both of these adaptors. Contrary to our expectations, SLP-76(-/-) SLP-65(-/-) BMM demonstrated normal FcgammaR-mediated activation, including internalization of Ig-coated sheep red blood cells and production of reactive oxygen intermediates. FcgammaR-induced biochemical events were normal in SLP-76(-/-) SLP-65(-/-) BMM, including phosphorylation of phospholipase C and the extracellular signaling-regulated kinases 1 and 2. To determine whether macrophages functioned normally in vivo, we infected WT and SLP-76(-/-) SLP-65(-/-) mice with sublethal doses of Listeria monocytogenes (LM), a bacterium against which the initial host defense is provided by activated macrophages. WT and SLP-76(-/-) SLP-65(-/-) mice survived acute, low-dose infection and showed no difference in the number of liver or spleen LM colony-forming units, a measure of the total body burden of this organism. Taken together, these data suggest that neither SLP-76 nor SLP-65 is required during FcgammaR-dependent signaling and functional events in macrophages.     

Receptors for IgA on phagocytic cells

IgA receptors have been detected on monocytes, polymorphonuclear neutrophils (PMNs) and eosinophils, and on phagocytic cells at mucosal sites. These receptors bind both secretory and serum forms of immunoglobulin A (IgA) and require the Ca2 region of the IgA molecule for ligand recognition. Monocytes and PMNs modulate their expression of the IgA receptor upon treatment with cytokines, such as granulocyto-macrophage colony-stimulating factor, and lipopolysaccharide. Purified IgA receptors appear as heavily glycosylated molecules with an average molecular weight of 60 kD, dropping to 32 and 36 kD upon treatment with N-glycanase. The cDNA sequence encoding the IgA receptor has been determined by expression cloning, and predicts that the receptor consists of two Ig-like extracellular domaines, a transmembrane region and a cytoplasmic tail of 41 residues. Ligation of IgA receptors on phagocytic cells by multivalent IgA complexes induces a variety of responses, including superoxide generation, release of inflammatory mediators, phagocytosis, and killing of various pathogenic microorganisms. Thus the apparent role of these receptors is to amplify the protective effects of the IgA antibody, a function of potential importance to mucosal defense.     

Preparation and characterization of porous beta-tricalcium phosphate/collagen composites with an integrated structure

Porous beta-tricalcium phosphate (TCP)/collagen composites with different beta-TCP/collagen weight ratio were prepared. The influences of the preparation conditions on the microstructure of porous composite and the joint status of beta-TCP particles with collagen fibrils were characterized by X-ray diffractometer, scanning electron microscopy and transmission electron microscopy. The results showed: (1) an acid treatment could effectively disassemble collagen fibrils; (2) in the resulting porous composites, beta-TCP particles homogenously existed on the skeleton of the collagen fibril network and bonded tightly to both the fibrils and themselves. The tight bonding formation could be due to the reaction between Ca ions in the particles and carboxyl groups in collagen polypeptide chains and due to the reprecipitation of partially dissolved beta-TCP during synthesis. The tight bonding between beta-TCP particles and collagen fibrils in the composites demonstrated an integrated structure, which was reproducible when beta-TCP/collagen ratio ranged from 2 to 4. Such integrated structure would make significant contributions in reliably tailoring properties of the porous composites by varying beta-TCP content. In addition, the porous composites had large porosity (approximately 95%) and appropriate pore size (approximately 100 microm), showed no negative impact in cytotoxicity assay and complete bone tissue regeneration after 12 weeks in animal test.