Boric acid inhibits LPS-induced TNF-α formation through a thiol-dependent mechanism in THP-1 cells

Oxidative stress plays an important role during inflammatory diseases and antioxidant administration to diminish oxidative stress may arrest inflammatory processes. Boron has been implicated to modulate certain inflammatory mediators and regulate inflammatory processes. Here we investigated the role of the tripeptide glutathione (GSH) in modulating the effects of boric acid (BA) on lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNF-alpha) formation in THP-1 monocytes. Interestingly, we found that BA had no significant effects on both TNF-alpha production and intracellular GSH contents, whereas it could inhibit LPS-induced TNF-alpha formation and ameliorated the d,l-buthionine-S,R-sulfoximine (BSO)-induced GSH depletion. Twenty-four hour incubation with BSO induced a decrease of the intracellular GSH and an increase of TNF-alpha. Treatment with N-acetyl-l-cysteine (NAC) did not significantly increase intracellular content of GSH but significantly reduced the secretion of TNF-alpha. BSO-pretreatment for 24h enhanced the LPS-induced secretion and mRNA expression of TNF-alpha further. BA inhibited LPS-stimulated TNF-alpha formation was also seen after GSH depletion by BSO. These results indicate that BA may have anti-inflammatory effect in the LPS-stimulated inflammation and the effect of BA on TNF-alpha secretion may be induced via a thiol-dependent mechanism.     

Multiple signals regulate phospholipase CBeta3 in human myometrial cells

Phospholipase CB3 (PLCB3) serine(1105) (S(1105)), a substrate for multiple protein kinases, represents a potential point of convergence of several signaling pathways in the myometrium. To explore this hypothesis, the regulation of PLCB3-S(1105) phosphorylation (P-S(1105)) was studied in immortalized and primary human myometrial cells. 8-[4-chlorophenylthio] (CPT)-cAMP and calcitonin gene-related peptide (CALCA) transiently increased P-S(1105). Relaxin also stimulated P-S(1105); this effect was partially blocked by the protein kinase A (PRKA) inhibitor, Rp-8-CPT-cAMPS. Oxytocin, which stimulates Galphaq-mediated pathways, also rapidly increased P-S(1105), as did prostaglandin F2alpha and ATP. Oxytocin-stimulated phosphorylation was blocked by protein kinase C (PRKC) inhibitor Go6976 and by pretreatment overnight with a phorbol ester. Cypermethrin, a PP2B phosphatase inhibitor, but not okadaic acid, a PP1/PP2A inhibitor, prolonged the effect of CALCA on P-S(1105), whereas the reverse was the case for the oxytocin-stimulated increase in P-S(1105). PLCB3 was the predominant PLC isoform expressed in the myometrial cells and PLCB3 short hairpin RNA constructs significantly attenuated oxytocin-stimulated increases in intracellular calcium. oxytocin-induced phosphatidylinositol (PI) turnover was inhibited by CPT-cAMP and okadaic acid, but was enhanced by pretreatment with Go6976. CPT-cAMP inhibited oxytocin-stimulated PI turnover in the presence of overexpressed PLCB3, but not overexpressed PLCB3-S(1105)A. These data demonstrate that both negative crosstalk from the cAMP/PRKA pathway and a negative feedback loop in the oxytocin/G protein/PLCB pathway involving PRKC operate in myometrial cells and suggest that different protein phosphatases predominate in mediating P-S(1105) dephosphorylation in these pathways. The integration of multiple signal components at the level of PLCB3 may be important to its function in the myometrium.     

Protopanaxadiol 6-hydroxylase and its role in regulating the ginsenoside heterogeneity in Panax notoginseng cells

Various structure-similar plant secondary metabolites like ginseng saponins (ginsenosides) possess different or even totally opposite biological activities. Intentional manipulation of the ginsenoside heterogeneity in cellular biosynthesis is of great interest and significance [Zhong and Yue (2005); Adv Biochem Eng Biotechnol 100:53-88]. In this work, CO-binding spectra of microsomes prepared from the suspended cells of Panax notoginseng showed increases in absorption at 450 nm compared with the control without CO sparging, and protopanaxadiol 6-hydroxylase (P6H), a new enzyme catalyzing the conversion of ginsenoside aglycone protopanaxadiol into protopanaxatriol, was found. P6H was dependent on NADPH and molecular oxygen. The enzymatic reaction was inhibited by carbon monoxide and partially reversible upon illumination with blue light, and sensitive to cytochrome P450 inhibitors. The results supported the contention that P6H was a cytochrome P450-dependent hydroxylase, whose catalytic product was confirmed to be protopanaxatriol by HPLC-MS. Induction of P6H activity by phenobarbital, a cytochrome P450 inducer, was observed. A maximal activity of P6H was obtained with addition of 0.5 mM phenobarbital on day 4 of shake-flask cultivation. The maximum content of protopanaxatriol-type ginsenosides (Rg(1) and Re, Rg group) and the maximum ratio of the content of protopanaxatriol: protopanaxadiol reached 6.88 +/- 0.21 mg g(-1) dry weight and 7.0, respectively, which was about 1.4 and 2.0-fold that of respective controls (without addition of phenobarbital). Oxidative burst was also observed in the cell cultures with addition of phenobarbital. P6H was concluded as a key enzyme in regulating Rg-group ginsenoside biosynthesis in P. notoginseng cells.     

Thrombin peptide TP508 prevents nitric oxide mediated apoptosis in chondrocytes in the endochondral developmental pathway

TP508 is a 23-amino acid peptide derived from human prothrombin that increases cartilage matrix production and reduces alkaline phosphatase activity without changing chondrocyte proliferation. This study tested the hypothesis that TP508 acts by blocking the onset of apoptosis associated with hypertrophy. Rat costochondral resting zone chondrocytes and human auricular chondrocytes were cultured in DMEM containing 50microM ascorbic acid and 10% FBS. Apoptosis was induced by treatment of confluent cultures with chelerythrine, tamoxifen, or inorganic phosphate (Pi) for 24h. One half of the cultures received TP508 (0, 0.7, or 7microg/ml). Apoptosis was assessed as a function of DNA fragmentation ([3H]-thymidine labeled DNA fragments), TUNEL staining, and cell viability using the MTT assay, as well as by assessing the Bcl-2/Bax mRNA and protein ratios and caspase-3 activity. The universal NO synthase inhibitor l-NMMA was used to assess the effect of NO production on chondrocyte apoptosis and specific NO synthase subspecies were identified using iNOS inhibitor 1400W and nNOS inhibitor vinyl-l-NIO, as well as l-NAME, which inhibits both iNOS and eNOS. Finally, we assessed if TP508 would block NO production induced by the apoptogens. Chelerythrine, tamoxifen and Pi-induced apoptosis and this was reversed by TP508. All apoptogens increased NO production and this was reduced by TP508. TP508 reduced NO levels to the same extent as 1400W but not to the same extent as l-NAME, suggesting that its effects are mediated primarily by iNOS. In addition, TP508 reduced the effect of chelerythrine to the same extent as 1400W and l-NAME, again indicating that it acts via inhibition of an iNOS pathway. TP508 also regulated Bcl-2/Bax mRNA in a time and dose-dependent manner. The Bcl-2/Bax mRNA ratio was 0.11 in the absence of TP508 at 1h and 4.95 at 7microg/ml TP508; by 3h the ratio was approximately 1 in both groups. The Bcl-2/Bax protein ratio also increased by 63% at 1h. TP508 did not affect caspase-3 activity. TP508 also caused a dose-dependent increase in protein kinase C (PKC) activity within 9min that was maximal at 270min. These results show that TP508 prevents apoptosis in growth plate chondrocytes via inhibition of iNOS-dependent NO and suggest a possible role for PKC in the mechanism.     

首例通过人工授精技术孕育的麋鹿在北京南海子麋鹿苑诞生

2008年4月11日,首例通过人工授精技术而孕育的麋鹿在北京南海子麋鹿苑诞生,它的降生具有十分重要的科研和经济价值.     

A test of rats' tolerance for 3beta-acetoxyandrosta-1,5-dien-17-one ethylene ketal (ADEK), a new anti-androgen

Following the demonstration that the androgen activity of androsta-5-ene-3beta,17beta-diol (Adiol) is not inhibited by the anti-androgens currently used to treat prostate cancer, we sought agents that would inhibit the androgenic function of Adiol as well as of dihydrotestosterone. The steroid 3beta-acetoxyandrosta-1,5-dien-17-one ethylene ketal (ADEK) met this criterion. Its tolerance was assessed in rats by oral and by subcutaneous administration for four weeks. Neither route of ADEK administration resulted in any behavioral changes. There was no effect on weight gain during the 28 days of steroid intake and no effect on the weight of the kidneys, heart, liver, testes, adrenals or the ventral lobe of the prostate glands. The seminal vesicles of the treated rats were 23-29% and the weights of the anterior prostates of the respective groups were 17-26% smaller than the controls. In contrast, the dorsolateral prostates were increased 26-55% as compared with the controls. There were no detectable changes in the histology of the kidneys, hearts, livers, testes and adrenals of any of the rats, but both groups of ADEK-treated rats had mild atrophic changes in their seminal vesicles and in the ventral lobe of their prostate glands. Both ADEK-treated groups showed focal glandular epithelial hyperplasia in the dorsolateral lobes in comparison with the control group. Orally administered ADEK was rapidly converted to several metabolites, which were nearly completely cleared from the blood within 4h.     

Optimization of cold-active protease production by the psychrophilic bacterium Colwellia sp. NJ341 with response surface methodology

Culture conditions were optimized for an extracellular cold-active protease production by the psychrophilic bacterium Colwellia sp. NJ341. Response surface methodology was applied for the most significant fermentation parameters (casein, citrate sodium, temperature and Tween-80) identified earlier by one-factor-at-a-time approach. A 2(4) full factorial central composite design was employed to determine the maximum protease production. Using this methodology, the quadratic regression model of producing cold-active protease was built and the optimal combinations of media constituents for maximum protease production (183.21 U/mL) were determined as casein 5.18 g/L, citrate sodium 3.84 g/L, temperature 7.96 degrees C, Tween-80 0.23 g/L. Protease production obtained experimentally coincident with the predicted value and the model was proven to be adequate.     

北疆高产棉花密度与打顶时序调控的生态位适宜度研究

以北疆棉区高产 (2 0 0 0kg·hm-2 )棉花 (G .hirsutumL .)为研究对象 ,引进生态位适宜度理论对受密度和打顶时序调控的高产棉花生态位适宜度进行了研究。经果表明 ,在密度梯度上 ,随密度增加棉花生态位适宜度值增加。 1 5 0× 1 0 3 株·hm-2 密度下 ,分次打顶降低了生态位适宜度 ,导致减产 ;在高密度种植条件下 ,分期打顶能提高生态位适宜度 ,有利于提高产量。在 2 2 5× 1 0 3 和 30 0× 1 0 3 株·hm-2 密度条件下 ,与同密度下 1次打顶 (7/5 )相比 ,2次打顶增产 4 5 %和 1 1 3%,3次打顶增产 7 5 %和 5 9%。 1 5 0× 1 0 3 株·hm-2 密度下 ,棉田苗蕾期LAI较小、漏光损失大 ,光合势低 ,是实现超高产的主要限制因子 ,花铃期的分期打顶导致减产。在 2 2 5× 1 0 3 和 30 0× 1 0 3 株·hm-2 密度条件下 ,棉花苗蕾期LAI增加 ,分期打顶保证了花铃期较适宜的LAI ,提高了棉花生态位适宜度 ,有利于提高产量 ;1次打顶处理 (7月 5日 )比本区生产上的打顶时间 (7月 1 0日左右 )提早了 5d左右 ,使高密度棉花群体LAI的发展受到了一定程度地限制 ,避免了因密度高群体透光条件的恶化而降低了其生态位适宜度值 ,造成减产损失。     

Genistein对腺样囊性癌cyclinB1蛋白表达和细胞增殖周期的影响

目的研究酪氨酸蛋白激酶抑制剂genistein抑制人涎腺腺样囊性癌(SACC-83)细胞生长与cyclin B1蛋白表达和细胞增殖周期的关系.方法 MTT法测定genistein对SACC-83细胞体外生长的作用;流式细胞仪测定细胞增殖周期;Western Blot技术检测cyclin B1和Cdk1蛋白,并利用电泳凝胶成像分析软件对其结果进行量化分析;采用SPSS11.5统计软件对结果进行统计学分析.结果 Genistein对SACC-83细胞生长有抑制作用,且当其作用到一定时间达到一定的浓度后,该作用与剂量及时间呈依赖关系;Genistein作用的细胞与对照细胞比较,G0/G1期细胞数减少, G2/M期细胞数增多,cyclin B1和Cdk1蛋白水平降低.结论 Genistein对SACC-83细胞生长的抑制作用与其调节cyclin B1和Cdk1蛋白表达和细胞增殖周期有关.