Aging effect on myeloperoxidase in rat kidney and its modulation by calorie restriction

Myeloperoxidase (MPO), a heme protein existing in neutrophil and monocyte, is implicated in various stages of inflammatory conditions with the production of a variety of potent oxidants. To investigate the extent of the involvement of MPO in aging, we measured MPO activities in kidney of rats at different ages maintained with an ad libitum (AL) or a calorie restriction (CR) dietary regimen. Results showed that the MPO activities increased during aging in AL rats, but were significantly attenuated by CR. This result was consistent with altered protein level of MPO during aging. In addition, we were able to detect dityrosine that is a stable end MPO-oxidation product. The amount of dityrosine increased in old AL, but not in old CR rats. To examine the source responsible for increased MPO activity during aging for leukocyte recruitment and infiltration, the levels of vascular cell adhesion molecule (VCAM-1) protein were measured. The level of VCAM-1 showed age-dependent increase in AL rats, which was correlated with higher activity of MPO in old AL rats. Furthermore, we have found that LPS-induced inflammation increased the activity and protein levels of MPO, and VCAM-1 expression in young rat kidneys. These findings suggest that increased MPO activity with aging may related to increased recruitment of inflammatory cells, contributing to protein oxidation accumulation in the aging process. We propose that age-related alterations of MPO, dityrosine, and VCAM were modulated by CR through its anti-inflammatory action.     

狂犬病毒鼠脑复壮后的典型形态恢复及感染细胞内的形态发生学

[目的]观察比较鼠脑复壮前后狂犬病毒的形态变化,并观察病毒感染BHK-21细胞后不同时间的形态发生情况.[方法]以保存时间较长的SRV9毒株为原始材料,经乳鼠脑传代复壮后接种BHK-21细胞,浓缩、纯化后观察.[结果](1)未经复壮的病毒中DI粒子占较高比例,典型粒子只占少数,而复壮后典型粒子所占比例升高到病毒粒子总数的90%.(2)感染24h后在细胞浆内可以观察到典型病毒粒子,其数量随着培养时间的延长而增加.带毒传代之后的培养过程中细胞内病毒数量增加不明显.(3)病毒可以在细胞内的空泡膜表面以多种方式成堆出芽.[结论](1)鼠脑复壮可恢复狂犬病毒中典型粒子所占比例.(2)带毒传代1～2次时为狂犬病毒收获的最佳时机.(3)本研究为狂犬病毒的装配机制补充了数据.     

检测心肌细胞钾离子通道蛋白活化水平方法的优化

介绍了一种如何合理的利用蛋白质免疫沉淀和蛋白质免疫印迹相结合的方法检测大鼠心肌细胞钾离子通道蛋白Kv1.2和Kv1.5的表达与活化水平。实验结果表明, 与单独利用免疫印迹的方法相比较, 本实验是对钾离子通道蛋白及其它亚家族的钾通道蛋白磷酸化表达水平检测方法的一种优化, 从而获得一套可行、简单、合理的实验方案, 同时也提高了检测的准确性, 敏感性及特异性。     

Effects of sap velocity on the daytime increase of stem CO2 efflux from stems of Schima superba trees

Stem CO₂ efflux (E _s) has been estimated from a temperature-related equation, but sap flux often affects measurements of E _s, which leads to misunderstanding real stem respiration. In order to observe the relationship between E _s and stem temperature and to analyze the effect of sap velocity on E _s, stem temperature, E _s and sap flux were measured from a subtropical Schima superba plantation in South China on three trees for consecutive 3 days in July and October 2009. Stem temperature, E _s and sap velocity were significantly higher in July than in October. Stem temperature could explain 17–41 and 54–75% variations of E _s in July and October, respectively. A negative relationship between E _s and stem temperature was found during 1800–2300 hours in July. The daytime E _s was 9.2, 4.3 and 2.4% higher than the predicted for three trees in July, and this occurred only on Tree 1 in October. Sap velocity was positively correlated with E _s for three trees in July, and the increase of E _s with the increase of sap velocity was only observed on Tree 1 in October. These results demonstrated that the occurrence of sap flux could account for the increase of daytime E _s, and the effect of sap velocity on E _s varied with the seasons from the S. superba stem.     

LncRNA TUG1 alleviates cardiac hypertrophy by targeting miR-34a/DKK1/Wnt-β-catenin signalling

The current study was designed to explore the role and underlying mechanism of lncRNA taurine up-regulated gene 1 (TUG1) in cardiac hypertrophy. Mice were treated by transverse aortic constriction (TAC) surgery to induce cardiac hypertrophy, and cardiomyocytes were treated by phenylephrine (PE) to induce hypertrophic phenotype. Haematoxylin-eosin (HE), wheat germ agglutinin (WGA) and immunofluorescence (IF) were used to examine morphological alterations. Real-time PCR, Western blots and IF staining were used to detect the expression of RNAs and proteins. Luciferase assay and RNA pull-down assay were used to verify the interaction. It is revealed that TUG1 was up-regulated in the hearts of mice treated by TAC surgery and in PE-induced cardiomyocytes. Functionally, overexpression of TUG1 alleviated cardiac hypertrophy both in vivo and in vitro. Mechanically, TUG1 sponged and sequestered miR-34a to increase the Dickkopf 1 (DKK1) level, which eventually inhibited the activation of Wnt/β-catenin signalling. In conclusion, the current study reported the protective role and regulatory mechanism of TUG1 in cardiac hypertrophy and suggested that TUG1 may serve as a novel molecular target for treating cardiac hypertrophy.     

Triterpenoids from <i>Uncaria rhynchophylla</i> and Their PTP1B Inhibitory Activity

Uncaria rhynchophylla (Gouteng) is a famous traditional Chinese medicine used for psychiatric and hypotensive purposes in China. In this study, the ethyl acetate (EtOAc) part of U. rhynchophylla was revealed with protein tyrosine phosphatase 1B (PTP1B) inhibitory activity. Subsequent investigation on the EtOAc part yielded one new triterpenoid, 3β-formyloxy-6β,19α-dihydroxyurs-12-en-28-oic acid (1) and four known ones, 3β,6β,19α-trihydroxyurs-12-en-28-oic acid (2), 2-oxopomolic acid (3), 3β,19α-dihydroxy-6-oxo-olean-12-en-28-oic acid (4) and sumaresinolic acid (5). The structure of compound 1 was determined by extensive HRESIMS, IR, 1D and 2D NMR spectroscopic analyses. Two ursane-type triterpenoids (2 and 3) showed selective inhibition on PTP1B with IC₅₀ values of 48.2 and 178.7 μM. The enzyme kinetic study suggested that compounds 2 and 3 were mix-type inhibitors on PTP1B with K_i values of 15.6 and 132.5 μM. This investigation manifests the antidiabetic potency of U. rhynchophylla with triterpenoids as the active constituents.     

Distribution and characterization of simple sequence repeats in Gossypium raimondii genome

Simple sequence repeats (SSRs) can be derived from the complete genome sequence. These markers are important for gene mapping as well as marker-assisted selection (MAS). To develop SSRs for cotton gene mapping, we selected the complete genome sequence of Gossypium raimondii, which consisted of 4447 non-redundant scaffolds. Out of 775.2 Mb sequence examined, a total of 136,345 microsatellites were identified with a density of 5.69 kb per SSR in the G. raimondii genome leading to development of 112,177 primer pairs. The distributions of SSRs in the genome were non-random. Among the different motifs ranging from 1 to 6 bp, penta-nucleotide repeats were most abundant (30.5%), followed by tetra-nucleotide repeats (18.2%) and di-nucleotide repeats (16.9%). Among all identified 457 motif types, the most frequently occurring repeat motifs were poly-AT/TA, which accounted for 79.8% of the total di-nt SSRs, followed by AAAT/TTTA with 51.5% of the total tetra-nucleotede. Further, 18,834 microsatellites were detected from the protein-coding genes, and the frequency of gene containing SSRs was 46.0% in 40,976 genes of G. raimondii. These genome-based SSRs developed in the present study will lay the groundwork for developing large numbers of SSR markers for genetic mapping, gene discovery, genetic diversity analysis, and MAS breeding in cotton.     

Up-regulation of BMP-2 antagonizes TGF-β1/ROCK-enhanced cardiac fibrotic signalling through activation of Smurf1/Smad6 complex

Rho‐associated kinase (ROCK) plays a critical role in pressure overload‐induced left ventricular remodelling. However, the underlying mechanism remains unclear. Here, we reported that TGF‐β1‐induced ROCK elevation suppressed BMP‐2 level and strengthened fibrotic response. Exogenous BMP‐2 supply effectively attenuated TGF‐β1 signalling pathway through Smad6‐Smurf‐1 complex activation. In vitro cultured cardiomyocytes, mechanical stretch up‐regulated cardiac TGF‐β1, TGF‐β1‐dependent ROCK and down‐regulated BMP‐2, but BMP‐2 level could be reversed through blocking TGF‐β1 receptor by SB‐431542 or inhibition of ROCK by Y‐27632. TGF‐β1 could also activate ROCK and suppress endogenous BMP‐2 level in a dose‐dependent manner. Knock‐down BMP‐2 enhanced TGF‐β1‐mediated PKC‐δ and Smad3 signalling cascades. In contrast, treatment with Y‐27632 or SB‐431542, respectively suppressed ROCK‐dependent PKC‐δ and Smad3 activation, but BMP‐2 was only up‐regulated by Y‐27632. In addition, BMP‐2 silencing abolished the effect of Y‐27632, but not SB‐431542 on suppression of TGF‐β1 pathway. Further experiments showed that Smad6 Smurf1 interaction were required for BMP‐2‐evoked antagonizing effects. Smad6 overexpression attenuated TGF‐β1‐induced activation of PKC‐δ and Smad3, promoted TGF‐β RI degradation in BMP‐2 knock‐down cardiomyocytes, and could be abolished after knocking‐down Smurf‐1, in which Smad6/Smurf1 complex formation was critically involved. In vivo data showed that pressure overload‐induced collagen deposition was attenuated, cardiac function was improved and TGF‐β1‐dependent activation of PKC‐δ and Smad3 was reduced after 2 weeks treatment with rhBMP‐2(0.5 mg/kg) or Y‐27632 (10 mg/kg) in mice that underwent surgical transverse aortic constriction. In conclusion, we propose that BMP‐2, as a novel fibrosis antagonizing cytokine, may have potential beneficial effect in attenuating pressure overload‐induced cardiac fibrosis.