Roles of protein phosphatase 4 in down-regulation of eNOS Ser633 phosphorylation induced by palmitic acid in human umbilical vein endotheli?al cells

AIMTo investigate the roles of protein phosphatase 4 (PP4) in down-regulation of endothelial nitric oxide synthase (eNOS) Ser633 phosphorylation induced by palmitic acid (PA). METHODSHuman umbilical vein endothelial cells (HUVECs) were treated with PA at 25 μmol/L, 50 μmol/L, 100 μmol/L and 200μmol/L for 36 h, or treated with PA at 100 μmol/L for 12 h, 24 h, 36 h and 48 h. Protein phosphatase 2A (PP2A) family inhibitor fostriecin (FST, 20 nmol/L) or okadaic acid (OA, 5 nmol/L) was selected to pretreat the HUVECs for 30 min. Protein phosphatase 4 catalytic subunit (PP4c) siRNA or protein phosphatase 2A catalytic subunit (PP2Ac) siRNA was transfected into the HUVECs. The protein expression levels of of eNOS, PP4c and PP2Ac, as well as the level of eNOS Ser633 phosphorylation, were detected by Western blot. The intracellular nitric oxide (NO) content was measured by DAF-FM DA. RESULTS(1) Compared with control group, the levels of eNOS Ser633 phosphorylation were decreased in PA groups in which the HUVECs were treated with 25 μmol/L, 50 μmol/L, 100 μmol/L and 200 μmol/L PA for 36 h (P<0.05) and 100 μmol/L PA for 24 h, 36 h and 48 h (P<0.05). No significant difference in the level of total eNOS protein expression among all the groups was observed. (2) Compared with control group, both FST and OA pretreatment reversed the reduction of eNOS Ser633 phosphorylation (P<0.05) and the decrease in intracellular NO content (P<0.05) induced by PA. No significant difference in the level of total eNOS protein expression among all the groups was observed. (3) Compared with si-Control group, the PP4c protein expression was significantly reduced (P<0.05), while the level of eNOS Ser633 phosphorylation was significantly increased in si-PP4c group (P<0.05). Although the levels of PP2Ac protein expression declined significantly (P<0.05), the level of eNOS Ser633 phosphorylation remained unchanged in si-PP2Ac group. No significant differencein the level of total eNOS protein expression among all the groups was found. CONCLUSION PA significantly reduces the level of eNOS Ser633 phosphorylation and the content of NO in the HUVECs, which may be due to PA inducing the activation of the PP2A family member PP4 rather than PP2A.     

Soluble Klotho protein suppresses THP-1-derived foam cell formation

AIMTo investigate the role of soluble Klotho protein in THP-1-derived foam cell formation. METHODSTHP-1 monocytes were induced into macrophages by treatment with 160 nmol/L phorbol myristate acetate for 48 h, and then were divided into 6 groups: negative control group (THP-1-derived macrophages), positive control group [THP-1-derived foam cells induced by oxidized low-density lipoprotein (ox-LDL) for 48 h], and 25, 50, 100 and 200 μg/L soluble Klotho protein groups (THP-1-derived macrophages pretreated with soluble Klotho protein at the indicat?ed concentraions for 2 h and then induced by ox-LDL for 48 h). Lipid droplets in cytoplasm were observed by oil red O staining. The cholesterol outflow rate was detected by scintillation counting technique. The content of intracellular total cholesterol, free cholesterol and cholesterol ester was detected by enzyme fluorescence analysis. The expression of acyl-coenzyme A:cholesterol acyltransferase 1 （ACAT1） and ATP-binding cassette transport?er A1 (ABCA1) at mRNA and protein levels was determined by RT-qPCR and Western blot, respectively. RESULTSOil red O staining and lipid mass quantification showed that THP-1-derived foam cell formation was dose-dependently suppressed by soluble Klotho protein. The cholesterol efflux rate of THP-1-derived foam cells was increased by soluble Klotho protein in a dose-dependent manner (P<0.05). In addition, soluble Klotho protein decreased the expression of ACAT1 and increased the expression of ABCA1 in a dose-dependent manner (P<0.05). CONCLUSION The soluble Klotho protein inhibits THP-1-derived foam cell formation in a dose-dependent manner by down-regulating the expression of ACAT1 and up-regulating the expression of ABCA1.     

Effect of tanshinone ⅡA on liver lipid deposition and ferroptosis-related protein expression in ApoE-/- mice

AIM To observe the effect of tanshinone ⅡA on liver lipid deposition and ferroptosis-related protein expression in ApoE^-/- mice. METHODS Thirty-two ApoE^-/- mice were randomly divided into model group, high-dose (60 mg/kg) tanshinone ⅡA group, low-dose (30 mg/kg) tanshinone ⅡA group and simvastatin group, and C57BL/6J mice (n=8) were used as normal control group. The mice in normal control group were given the basic feeding, while the others were given high-fat diet. The mice in tanshinone ⅡA groups and simvastatin group were given corresponding drugs. The mice in normal control group and model group were intraperitoneally injected with equal volume of saline. Eight weeks later, the serum levels of triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were tested by automatic biochemistry analyzer. The liver tissues were stained with HE and oil red O. The contents of reactive oxygen species (ROS) and glutathione (GSH) in liver tissues of the mice were measured by commercially available kits. The liver glutathione peroxidase 4 (GPX4) and p53 were detected by immunohistochemical method. The protein and mRNA expression levels of ferroptosis-related factors GPX4, xCT/SLC7A11, p53 and ferritin heavy chain 1 (FTH1) were determined by Wes automatic Western blot quantitative analysis system and RT-qPCR. RESULTS Compared with normal control group, the serum levels of TC, TG and LDL-C in model group were increased significantly (P<0.05 or P<0.01), and HDL-C did not change significantly. The fat vacuoles were clearly visible in liver tissue. The content of ROS in liver tissue was increased significantly,and GSH was decreased significantly (P<0.01). The mRNA and protein expression levels of p53 were increased significantly, and GPX4, xCT/SLC7A11 and FTH1 were decreased significantly (P<0.05 or P<0.01). Compared with model group, tanshinone ⅡA significantly decreased the serum levels of TC, TG and LDL-C (P<0.05 or P<0.01), and HDL-C did not change significantly. High-dose and low-dose tanshinoneⅡA also significantly decreased the degree of steatosis, and the size of lipid droplets. The content of ROS in liver tissues was decreased significantly, and GSH was increased significantly (P<0.01). The mRNA and protein expression levels of GPX4, xCT/SLC7A11 and FTH1 were increased significantly, and p53 were decreased significantly (P<0.05 or P<0.01). CONCLUSION Tanshinone ⅡA reduces liver lipid deposition and lipid peroxidation damage in ApoE^-/- mice, which may be related to the intervention of ferroptosis-related proteins in the liver cells.     

Effect of forsythiaside A on immune function in rats with ulcerative colitis

AIM To investigate the effect of forsythiaside A (FA) on immune function in rats with ulcerative colitis and its related mechanism. METHODS Healthy SD rats were randomly divided into 5 groups: control group (no treatment, normal feeding), model group (establishment of rat ulcerative colitis model), and low, medium and high doses of FA groups (treatment of the model rats with FA at 5 mg/kg, 20 mg/kg and 80 mg/kg, respectively). The malondialdehyde (MDA) content and superoxide dismutase (SOD) activity in rat colon tissues were measured by colorimetry, and the serum levels of tumor necrosis factor-α (TNF-α), interleukin-2 (IL-2) and IL-4 were detected by ELISA. The spleen index and thymus index, the percentages of CD3⁺, CD4⁺ and CD8⁺ T-lymphocytes in peripheral blood mononuclear cells （PBMC）, the serum IgA and IgG levels, and the serum complement C3 and C4 levels were also determined. RESULTS The colon tissues of the rats in model group showed obvious inflammation and ulceration, indicating that the animal model was successfully established. Compared with model group, the colonic inflammation and ulceration were significantly attenuated in FA groups, among which the high dose had the best effect. Compared with control group, the spleen index and thymus index of the rars in model group were decreased (P<0.05), MDA content in colon tissues was increased (P<0.05), and SOD activity in colon tissues was decreased (P<0.05). The levels of CD3⁺, CD4⁺, CD8⁺ and CD4⁺/CD8⁺ T-lymphocytes in PBMC, and the serum levels of C3, C4 and IL-4 were decreased (P<0.05), while the serum levels of IgA, IgG, TNF-α, and IL-2 were increased in model group as compared with control group. Furthermore, the spleen index and thymus index of the rats in FA groups were increased (P<0.05), the MDA content in the colon tissues was decreased (P<0.05), and the SOD activity in the colon tissues was increased (P<0.05). The levels of CD3⁺, CD4⁺, CD8⁺ and CD4⁺/CD8⁺ T-lymphocytes in PBMC, and the serum levels of C3, C4 and IL-4 were increased (P<0.05), while serum IgA, IgG, TNF-α and IL-2 levels were decreased in FA groups as compared with model group (P<0.05). CONCLUSION Forsythiaside A effectively attenuates the colonic lesions in rats with ulcerative colitis, and its mechanism may be related to reinforcement of oxygen free radical scavenging power, alleviation of inflammatory response, and enhancement of immune function.     

Expression and function of circular RNA_0000231 in non-small-cell lung cancer

AIM: To investigate the expression and function of circular RNA_0000231 (circ_0000231) in non-small-cell lung cancer (NSCLC). METHODS: RT-qPCR was used to detect the expression of circ_0000231 in the NSCLC tissues and cell lines. circ_0000231 small interfering RNA (si-circ_0000231) or negative control siRNA of circ_0000231 (NC) was transfected into the NSCLC cells. The proliferation and apoptosis of the NSCLC cells were detected by CCK-8 assay, colony formation assay and flow cytometry, respectively. The expression of cyclin D1 (CCND1) and anti-apoptotic protein Bcl-2 were determined by RT-qPCR and Western blot. RESULTS: The expression of circ_0000231 in the NSCLC tissues and cell lines was significantly up-regulated compared with precancerous tissues and lung epithelial cells BEAS-2B (P<0.05). After transfection of NSCLC cells with si-circ_0000231, the cell viability, colony formation numbers were significantly decreased, and the apoptotic rate in si-circ_0000231 group was significantly increased as compared with NC group (P<0.01). In addition, the results of RT-qPCR and Western blot showed that transfection of si-circ_0000231 inhibited the expression of CCND1 and Bcl-2 (P<0.01). CONCLUSION: The expression of circ_0000231 is significantly increased in the NSCLC tissues and cells. Knock-down of circ_0000231 expression significantly inhibits the proliferation of NSCLC cells.     

菌剂与钾肥配施对连作魔芋生长、生理代谢和发病率的影响

以安康当地花魔芋为材料进行盆栽试验,研究娄彻氏链霉菌（Streptomyces rochei）菌剂D74与钾肥配施对连作魔芋生物学性状、块茎产量、叶绿素含量、活性氧代谢、渗透调节物质含量及软腐病的影响,为魔芋连作障碍微生物-化学双重修复提供理论依据。结果表明：菌剂与钾肥配施对连作魔芋具有一定的促生防病作用,使魔芋单株块茎鲜质量和生长系数分别较对照显著增加86.7%和85.9%,软腐病病情指数降低 33.0%。配施处理中,钾肥和菌剂分别在促进魔芋块茎膨大和抑制软腐病方面发挥主导作用。菌剂与钾肥配施促进魔芋叶片叶绿素的合成,其叶绿素a及叶绿素(a+b)含量分别较对照显著增加51.0%及34.6%,同时叶绿素a/b及叶绿素(a+b)/类胡萝卜素比值分别提高41.8%及22.2%。配施的菌剂效应对叶绿素合成的促进作用占主导地位。菌剂与钾肥配施显著提高魔芋的诱导抗性,其叶片过氧化物酶活性、可溶性蛋白和可溶性糖含量分别为对照的3.1倍、2.9倍和2.2倍。采用主成分分析法对魔芋相关19个生理生化指标分析得出特征值、贡献率、累积贡献率和主成分载荷矩阵,提取3个主成分（叶绿素b、叶幅和相对防效）综合评价菌剂与钾肥配施对魔芋连作障碍的修复效果。不同施肥处理综合得分高低顺序为：菌剂与钾肥配施>单施钾肥>单施菌剂>不施肥对照。综上所述,娄彻氏链霉菌菌剂D74与钾肥配施能提高连作魔芋产量并降低植株发病率,有效缓解连作障碍。     

奶牛乳脂性状候选基因EEF1D突变位点功能分析

提高乳脂含量、提升奶质已成为当前奶牛业研究的重点方向之一。前期经GWAS方法研究发现,EEF1D基因5'非翻译区(5'-UTR)的G/A突变与乳脂率极显著相关,为了进一步对候选基因EEF1D的突变位点进行功能分析,首先通过人工合成获得了EEF1D基因突变型与野生型片段,并在细胞水平上对其开展了功能验证实验。研究发现,EEF1D基因5'-UTR的G/A突变引起Sp3转录起始位点后移33 bp,并且突变型的活性大约是野生型的3倍。研究结果表明,该突变可改变转录因子的转录起始位点,从而使EEF1D的表达上调。     

2001-2017年天山中段积雪时空变异研究

【目的】为干旱区提供冰雪资源的合理利用及灾害防治的科学理论不服务信息。【斱法】采取趋势分析法、相关性分析法,开展了2001-2017年积雪年内年际变化特征以及分布变化原因斱面的研究。【结果】①天山中段SCP(积雪覆盖率)年际变化呈"单峰"型曲线,6月到最小值,12月到次年1月达到最大值,从2月开始逐渐下降到最小值;从7月初开始SCP开始逐渐上升到最大值。②天山中段年际SCP呈减少趋势、春季的SCP变化呈略微减少趋势、夏季不秋季SCP变化均呈增加趋势、冬季SCP变化趋势跟全年年际SCP变化趋势基本相似,且均呈减少趋势,最高值为98.6%,最小值出现在2009年为83.2%。③天山中段54.08%区域的SCD(积雪日数)呈减少趋势,其中4.36%区域呈显著减少趋势;45.92%区域的SCD呈增加趋势,其中3.03%区域呈显著增加趋势。④天山中段积雪不LST(地表温度)年际相关性呈负相关性比较明显,负相关占总面积的55.89%,主要分布在高海拔的天山山脉,博格达尔;34.98%区域呈极显著负相关,主要分布在伊犁河谷、天山南北坡地区不阿克苏、库尔勒、博斯腾湖流域周边。【结论】天山中段积雪时空变化丌仅是LST变化的影响,还受人类活劢和海拔高度差异等因素的共同作用。     

重组GSTA3蛋白对福美双诱导的肉鸡TD中抗凋亡基因BAG-3表达的影响

【目的】肉鸡胫骨软骨发育不良（TD）是肉鸡常见的一种骨骼性疾病,研究重组GSTA3蛋白对福美双诱导的TD肉鸡软骨细胞中抗凋亡基因BAG-3表达的影响,为治疗TD提供新的思路和方法。【方法】将120羽1周龄肉雏鸡随机分为6组（编号为A、B、C、D、E、F组）。A、B、C组为基础日粮对照组,D、E、F组为添加福美双日粮诱导TD组。试验饲喂福美双2 d诱发TD,在添加福美双第1、3、5、7天,腿部肌肉注射重组鸡GSTA3蛋白和磷酸盐缓冲液,A组与D组注射（100 μg·kg ^-1）磷酸盐缓冲液;B组与E组注射低剂量（100 μg·kg ^-1）GSTA3;C组与F组注射高剂量（200 μg·kg ^-1）GSTA3。试验历时23 d。添加福美双后1、2、4、6、10和15 d采集胫骨生长板。通过Real-time qPCR检测BAG-3基因的mRNA水平,利用免疫组化来检测BAG-3蛋白表达水平。【结果】Real-time qPCR结果显示,TD损伤修复期内,相比较于基础日粮对照组,福美双对照组肉鸡胫骨生长板中BAG-3 mRNA的表达水平基本都显著上调（P<0.05）;相比较于福美双对照组,E和F组在第2、4、10、15天都有显著差异,且在第10和15天显著低于福美双对照组（P<0.05）,表明与D组相比恢复较快。免疫组化结果表明BAG-3蛋白在肉鸡胫骨软骨细胞的增殖区和前肥大区无表达,只在肥大区细胞质中表达;福美双组与空白对照组相比,BAG-3蛋白表达增加;福美双高低剂量组与未注射蛋白的福美双组相比,重组GSTA3增加了肥大区的蛋白表达水平（第10和15天）。【结论】在福美双诱导肉鸡发生TD的过程中,GSTA3重组蛋白能够通过调控BAG-3表达参与凋亡途径,抑制细胞凋亡。在TD损伤修复期,注射GSTA3后使抗凋亡基因BAG-3蛋白表达增强,从而可参与细胞凋亡来缓解TD损伤,使得肉鸡TD生长板功能较快地恢复正常。     

美国农学教习眼中的张之洞与湖北农务学堂

自1889年张之洞出任湖广总督后的十余年间,张之洞在荆楚大地开创了著名的区域近代化模式——“湖北新政”,使其谋求早期工业化、教育改革以及军队近代化的尝试在湖北获得了一个巨大的实验场。张之洞曾为改良农业,创办新式农务学堂而首先从美国聘请了农学教习布里尔。现存于美国的布里尔相关档案,以其亲历者和旁观者的视角,对于张之洞“湖北新政”期间的举措,尤其是教育方面的改革尝试进行了一些记录和述评。这些新近发掘的史料,为深化相关研究提供了新的依据和视角。