658 nm低能量激光照射对人牙周膜细胞生物学效应的影响

目的:探讨658 nm低能量激光照射对人牙周膜细胞增殖、碱性磷酸酶活性及纤维连接蛋白合成的影响.方法:改良组织块法体外培养人牙周膜细胞.通过658 nm激光照射人牙周膜细胞,观察能量密度为1.86 J/cm2和3.72 J/cm2激光照射后不同时间点细胞增殖效应、碱性磷酸酶活性和纤维连接蛋白的变化.结果:1.86 J/cm2和3.72 J/cm2能量密度的激光照射人牙周膜细胞,可显著促进细胞增殖效应.3.72 J/cm2能量密度的激光照射可提高人牙周膜细胞碱性磷酸酶活性;能量密度为1.86 J/cm2的激光照射人牙周膜细胞72 h后,细胞中纤维连接蛋白分泌量增加.结论:658 nm低剂量激光照射可促进人牙周膜细胞增殖;适量的低剂量激光照射人牙周膜细胞可促进其碱性磷酸酶活性及纤维连接蛋白的分泌.     

Two new secondary metabolites isolated from Avena sativa L. (Oat) seedlings and their effects on osteoblast differentiation

Seedlings of natural crops are valuable sources of pharmacologically active phytochemicals. In this study, we aimed to identify new active secondary metabolites in Avena sativa L. (oat) seedlings. Two new compounds, avenafuranol (1) and diosgenoside (2), along with eight known compounds (3–10) were isolated from the A. sativa L. seedlings. Their chemical structures were elucidated via 1D and 2D NMR spectroscopy, high-resolution ESIMS, IR spectroscopy, optical rotation analysis, and comparisons with the reported literature. The effect of each isolated compound on alkaline phosphatase (ALP) activity for osteoblast differentiation induced by bone morphogenetic protein-2 (BMP-2) was investigated using the C2C12 immortal mouse myoblast cell line. Compounds 1, 4, 6, 8, and 9 induced dose-dependent increases in ALP expression relative to ALP expression in cells treated with only BMP-2, and no cytotoxicity was observed. These results suggest that A. sativa L. seedlings are a natural source of compounds that may be useful for preventing bone disorders.     

镉、铅对蟾蜍精巢毒作用的酶学研究

为研究镉、铅对精巢的生化毒作用机理 ,应用氯化镉、硝酸铅溶液对成年雄性蟾蜍进行腹腔染毒 (按镉计 0 .1、0 .2、0 .4、0 .8mg/ kg体重 ;按铅计 1、2、4、8mg/ kg体重 ) ,连续染毒 7d后活体解剖 ,测定分析精巢中各种酶的活性。结果显示 ,在镉染毒下 ,精巢乳酸脱氢酶 (L DH)和酸性磷酸酶 (ACP)的活性随镉染毒剂量的增加而降低 ,而碱性磷酸酶 (AL P)未发现明显变化 ;在铅染毒下 ,乳酸脱氢酶 (L DH)和碱性磷酸酶 (AL P)的活性随铅染毒剂量的增加而降低 ,而酸性磷酸酶 (ACP)未发现明显变化 ;乳酸脱氢酶 (L DH)同工酶在镉、铅染毒下则主要表现为酶带 L DH1 、L DH2 的抑制或缺失和 L DH5活性的增强。因此 ,镉、铅对蟾蜍的雄性生殖毒性机理可能与酶的活性存在着一定的关系 ,L DH可以考虑作为反映镉、铅中毒对精巢功能影响程度的一种有价值的生化指标     

Toxicity of Cry1A toxins from Bacillus thuringiensis to CF1 cells does not involve activation of adenylate cyclase/PKA signaling pathway

Bacillus thuringiensis (Bt) bacteria produce Cry toxins that are able to kill insect pests. Different models explaining the mode of action of these toxins have been proposed. The pore formation model proposes that the toxin creates pores in the membrane of the larval midgut cells after interaction with different receptors such as cadherin, aminopeptidase N and alkaline phosphatase and that this pore formation activity is responsible for the toxicity of these proteins. The alternative model proposes that interaction with cadherin receptor triggers an intracellular cascade response involving protein G, adenylate cyclase (AC) and protein kinase A (PKA). In addition, it was shown that Cry toxins induce a defense response in the larvae involving the activation of mitogen-activated kinases such as MAPK p38 in different insect orders. Here we analyzed the mechanism of action of Cry1Ab and Cry1Ac toxins and a collection of mutants from these toxins in the insect cell line CF1 from Choristoneura fumiferana, that is naturally sensitive to these toxins. Our results show that both toxins induced permeability of K⁺ ions into the cells. The initial response after intoxication with Cry1Ab and Cry1Ac toxins involves the activation of a defense response that involves the phosphorylation of MAPK p38. Analysis of activation of PKA and AC activities indicated that the signal transduction involving PKA, AC and cAMP was not activated during Cry1Ab or Cry1Ac intoxication. In contrast we show that Cry1Ab and Cry1Ac activate apoptosis. These data indicate that Cry toxins can induce an apoptotic death response not related with AC/PKA activation. Since Cry1Ab and Cry1Ac toxins affected K⁺ ion permeability into the cells, and that mutant toxins affected in pore formation are not toxic to CF1, we propose that pore formation activity of the toxins is responsible of triggering cell death response in CF1cells.     

Potential chemoprevention of N-nitrosodiethylamine-induced hepatocarcinogenesis by polyphenolics from Acacia nilotica bark

Chemopreventive potential of Acacia nilotica bark extract (ANBE) against single intraperitoneal injection of N-nitrosodiethylamine (NDEA, 200 mg/kg) followed by weekly subcutaneous injections of carbon tetrachloride (CCl₄, 3 ml/kg) for 6 weeks induced hepatocellular carcinoma (HCC) in rats was studied. At 45 day after administration of NDEA, 100 and 200 mg/kg of ANBE were administered orally once daily for 10 weeks. The levels of liver injury and liver cancer markers such as alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), γ-glutamyl transferase (γ-GT), total bilirubin level (TBL), α-feto protein (AFP) and carcinoembryonic antigen (CEA) were substantially increased following NDEA treatment. However, ANBE treatment reduced liver injury and restored liver cancer markers. ANBE also significantly prevented hepatic malondialdehyde (MDA) formation and reduced glutathione (GSH) in NDEA-treated rats which was dose dependent. Additionally, ANBE also increased the activities of antioxidant enzymes viz., catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione-S-transferase (GST) in the liver of NDEA-administered rats. Eventually, ANBE also significantly improved body weight and prevented increase of relative liver weight due to NDEA treatment. Histological observations of liver tissues too correlated with the biochemical observations. HPLC analysis of ANBE showed the presence of gallic, protocatechuic, caffeic and ellagic acids, and also quercetin in ANBE. The results strongly support that A. nilotica bark prevents lipid peroxidation (LPO) and promote the enzymatic and non-enzymatic antioxidant defense system during NDEA-induced hepatocarcinogenesis which might be due to activities like scavenging of oxy radicals by the phytomolecules in ANBE.     

Erv26p-Dependent Export of Alkaline Phosphatase from the ER Requires Lumenal Domain Recognition

Active sorting at the endoplasmic reticulum (ER) drives efficient export of fully folded secretory proteins into coat protein complex II (COPII) vesicles, whereas ER-resident and misfolded proteins are retained and/or degraded. A number of secretory proteins depend upon polytopic cargo receptors for linkage to the COPII coat and ER export. However, the mechanism by which cargo receptors recognize transport-competent cargo is poorly understood. Here we examine the sorting determinants required for export of yeast alkaline phosphatase (ALP) by its cargo receptor Erv26p. Analyses of ALP chimeras and mutants indicated that Erv26p recognizes sorting information in the lumenal domain of ALP. This lumenal domain sorting signal must be positioned near the inner leaflet of the ER membrane for Erv26p-dependent export. Moreover, only assembled ALP dimers were efficiently recognized by Erv26p while an ALP mutant blocked in dimer assembly failed to exit the ER and was subjected to ER-associated degradation. These results further refine sorting information for ER export of ALP and show that recognition of folded cargo by export receptors contributes to strict ER quality control.     

Sphingolipid synthesis is involved in autophagy in Saccharomyces cerevisiae

In eukaryotes, autophagy is a conserved protein degradation system that degrades cytoplasmic components by encompassing them with double-membrane structures, called autophagosomes, and delivering them to the lytic compartments of vacuoles/lysosomes. Certain Atg proteins are known to be involved in autophagy, yet the identity and function of lipid molecules involved remain largely unknown. We investigated the involvement of sphingolipids in autophagy using Saccharomyces cerevisiae. Inhibiting synthesis of the simplest complex sphingolipid, inositol phosphorylceramide (IPC), resulted in reduced autophagic activities. Similar results were obtained using myriocin, an inhibitor of the first step in sphingolipid synthesis. Our results indicate that sphingolipids, especially IPC, are required for autophagy. Inhibition of sphingolipid synthesis had no effect on formation of Atg12-Atg5 or Atg8-phosphatidylethanolamine conjugates, on maturation of vacuolar proteases, or on formation of the pre-autophagosomal structure (PAS). These results suggest that sphingolipids are not involved in the cellular signaling that leads to formation of the PAS, but may be involved in the process of autophagosome formation.     

Variable expression of lineage regulators in differentiated stromal cells indicates distinct mechanisms of differentiation towards common cell fate

Mesenchymal stem cells (MSCs) possess a multi-lineage differentiation capacity that makes them important players in the field of regenerative medicine. MSC populations derived from different tissues or donors have been shown to exhibit variable gene expression patterns. Further, it is widely acknowledged that MSC isolates are heterogeneous mixtures of cells at different developmental stages. However, the heterogeneity of expression of lineage regulators has not been linked to differentiation potential of different MSC populations towards mesenchymal lineages. Here, we analyzed variation of expression of differentiation markers across whole population and between single differentiating cells of multipotent stromal cell populations derived from adipose tissue (AdMSCs) and skin (FBs) of seven donors. The results of the analyses show that all cell populations exhibit similar differentiation potential towards adipocyte, osteoblast and chondrocyte lineages despite tissue type- and donor-specific variations of expression of differentiation-associated genes. Further, we detected variable expression of lineage regulators in individual differentiating cells. Together, our data indicate that single cells of stromal cell populations could use distinct molecular mechanisms to reach a common cell fate.     

EGFR trans-activation mediates pleiotrophin-induced activation of Akt and Erk in cultured osteoblasts

Pleiotrophin (Ptn) plays an important role in bone growth through regulating osteoblasts’ functions. The underlying signaling mechanisms are not fully understood. In the current study, we found that Ptn induced heparin-binding epidermal growth factor (HB-EGF) release to trans-activate EGF-receptor (EGFR) in both primary osteoblasts and osteoblast-like MC3T3-E1 cells. Meanwhile, Ptn activated Akt and Erk signalings in cultured osteoblasts. The EGFR inhibitor AG1478 as well as the monoclonal antibody against HB-EGF (anti-HB-EGF) significantly inhibited Ptn-induced EGFR activation and Akt and Erk phosphorylations in MC3T3-E1 cells and primary osteoblasts. Further, EGFR siRNA depletion or dominant negative mutation suppressed also Akt and Erk activation in MC3T3-E1 cells. Finally, we observed that Ptn increased alkaline phosphatase (ALP) activity and inhibited dexamethasone (Dex)-induced cell death in both MC3T3-E1 cells and primary osteoblasts, such effects were alleviated by AG1478 or anti-HB-EGF. Together, these results suggest that Ptn-induced Akt/Erk activation and some of its pleiotropic functions are mediated by EGFR trans-activation in cultured osteoblasts.     

Comparative evaluation of hepatotoxic and nephrotoxic effects of aroclors 1221 and 1254 in female rats

Polychlorinated biphenyls (PCBs) are persistent environmental pollutants. This study compared effects of two PCB mixtures, Aroclors 1221 (A1221) and 1254 (A1254) on serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), urea, creatinine and uric acid in female rats. Histopathological changes in the liver and kidney were also examined. A group of adult Wistar rats served as controls. Groups II and III were subcutaneously injected with A1221 and A1254 at 10 mg/kg every other day for 6 weeks. At the end of this period, all animals were decapitated and blood samples were collected. Serum urea, creatinine, uric acid, ALT, AST and ALP levels were determined. Liver and kidney were collected for histopathological examination. They were fixed in formaldehyde and processed for light microscopy. Both A1221 and 1254 significantly elevated serum ALT (p < 0.05) and AST (p < 0.01) levels compared to the control group. Serum ALP values were significantly increased by A1221 (p < 0.05), but they were unaffected in the A1254 group. Treatment with both A1221 and A1254 significantly increased serum levels of urea (p < 0.05), creatinine (p < 0.01) and uric acid (except in the A1221 group; p < 0.005). Distinct histopathological changes including renal corpuscular atrophy, peritubular vascular congestion and dilated cortical tubules, sinusoidal dilatation, congestion and mononuclear cell infiltration were observed. These findings suggest that PCBs may cause nephrotoxicity and hepatotoxicity in female rats.