Role of ghrelin in cell protection by up-regulating HSP70 through ERK1/2 signaling pathway in PC12 cells

AIM:To study the role of ghrelin in cell protection by up-regulating heat shock protein 70 (HSP70) and inhibiting apoptosis induced by oxidative stress through extracellular regulated protein kinases 1/2 (ERK1/2) signaling pathway in the PC12 cells. METHODS:Sodium nitoprusside (SNP) was used to induce oxidative stress injury in the PC12 cells. The cultured PC12 cells were divided into SNP-injured group (incubated with SNP at 0.5 mmol/L for 6, 12, 18 and 24 h), ghrelin pretreatment group (ghrelin at 100 nmol/L was given 30 min before adding SNP); HSP70 inhibitor group (quercetin at 10 μmol/L was added 60 min before ghrelin treatment), ERK inhibitor group (ERK 1/2 inhibitor PD98059 was added 60 min before ghrelin treatment) and control group (added same amount of culture medium only). The apoptotic rate was detected by flow cytometry. The protein expression was determined by Western blot and immunocytochemistry. RESULTS:Compared with control group, the apoptotic rate of PC12 cells in SNP-injured group was significantly increased (P<0.05). Compared with SNP-injured group, ghrelin (100 nmol/L) pretreatment significantly inhibited SNP-induced apoptosis of PC12 cells (P<0.05), and significantly up-regulated the protein expression of HSP70 (P<0.05). Time-effect analysis showed that ghrelin had the most significant effect at 18 h after SNP injury. Quercetin, an inhibitor of HSP 70, significantly reduced the anti-apoptotic effect of ghrelin (P<0.05). Ghrelin pretreatment promoted the phosphorylation of ERK1/2. ERK1/2 inhibitor PD98059 significantly inhibited the effects of ghrelin on up-regulation of HSP70 expression (P<0.05). CONCLUSION:Ghrelin upregulates the expression of HSP70 and inhibits the apoptosis in the PC12 cells induced by oxidative stress by promoting the phosphorylation of ERK1/2.     

三种融合标签对大熊猫轮状病毒结构蛋白VP6-VP7可溶性表达的影响

大熊猫轮状病毒(giant panda rotavirus,GPRV)是引起幼龄大熊猫腹泻的主要病原,对圈养大熊猫产生了较大的危害。轮状病毒结构蛋白VP6是一种载体蛋白,可介导黏膜免疫反应。VP7是轮状病毒结构蛋白中主要的中和抗原。因此,VP6-VP7的融合表达作为候选抗原对该病的防治具有重要的意义。传统大肠埃希菌原核表达存在表达量低、可溶性差以及纯度低等弊端。本研究使用醛缩酶(EDA)、谷胱甘肽S-转移酶(GST)、麦芽糖结合蛋白(MBP)3种融合标签,以实现获得表达量高和纯度高的GPRV-VP6-VP7重组表达蛋白。将扩增的VP6、VP7基因片段利用同源重组酶构建到含3种融合标签的表达载体pET21b上,将重组质粒转化至大肠埃希菌Rosetta(DE3)感受态细胞中进行低温诱导表达。用Ni-柱亲和层析法纯化目的蛋白,SDS-PAGE和Image J分析蛋白表达量和可溶性,Western blot分析得到表达的重组表达蛋白正确且具有蛋白活性。实验结果证明,EDA标签能显著促进VP6-VP7蛋白的原核可溶性表达,提高VP6-VP7蛋白表达量。     

茶树新品种--梦茗

"梦茗"系从安徽省岳西县来榜镇优良地方群体种中采用单株选育法育成的适制绿茶新品种。经安徽省内品比、推广示范和全国区试,发现"梦茗"物候期与福鼎大白茶相当,属早生种。品质优异,全国区试3个点感官审评总分均高于对照;内含物含量高,氨基酸含量4.9%、茶多酚总量26.9%、水浸出物总量40%。产量高,品比试验高于福鼎大白茶26.45%,全国区试产量与对照相当。抗旱能力强,抗寒能力与对照相当。     

“金花散茶”及“金花菌粉”对被动吸烟小鼠肺组织JAK2/STAT3炎性及磷酸化蛋白表达的影响

为探究“金花散茶”及其“金花菌粉”对被动吸烟(Cigarette smoking environment,CSE)小鼠肺组织受损的预防及修复机制,建立C57BL/6小鼠CSE模型,以600 mg∙kg^-1剂量的金花散茶茶汤(Eurotium cristatum tea extract,ECTE)及金花菌粉浸提液(Eurotium cristatum powder extract,ECPE)进行灌喂处理。与CSE模型组相比,小鼠灌喂ECPE和ECTE后,肺组织病理学切片显示其可保护小鼠肺组织形态结构完整;酶联免疫分析显示,灌喂ECPE和ECTE可显著抑制小鼠血清IL-6、IL-8、IL-1β、IFN-γ和TNF-α表达量上调;Western blot结果表明,灌喂ECPE和ECTE对小鼠肺组织p-JAK2、p-STAT3、p-JAK2/JAK2、p-STAT3/STAT3高表达起到抑制作用。以上研究结果表明,灌喂ECPE、ECTE对CSE肺受损小鼠具有明显保护作用,总体趋势为ECPE组优于ECTE组、预防组优于治疗组。     

水稻抽穗期基因OsDof6功能的初步研究

【目的】以前期通过穗发育芯片筛选到一个水稻Dof家族转录因子OsDof6为研究对象,进一步探究OsDof6的表达模式与生物学功能。【方法】对OsDof6的基因、蛋白及启动子序列进行生物信息学分析,利用CRISPR/Cas9技术对该基因进行定点编辑,通过实时定量PCR和亚细胞定位技术分析该基因的表达模式。【结果】对该基因的启动子序列分析表明,该基因启动子中存在大量与光响应、激素响应及胁迫响应相关的顺式调控元件。利用CRISPR/Cas9技术获得了2种不同突变类型的功能缺失突变体9522^Dof6-3和9522^Dof6-4。对突变体T₁株系进行表型观察发现,相较于对照,两种突变体在营养生长阶段分蘖数明显降低,转入生殖生长阶段后,两种突变体的抽穗时间推迟约3 d。实时定量PCR结果显示OsDof6在水稻根、茎、叶和穗中均有不同程度的表达,在穗发育后期相对表达量明显提高;通过亚细胞定位分析发现OsDof6定位于细胞核。【结论】初步判断OsDof6基因会影响水稻分蘖数与抽穗期。     

麦/油-稻轮作下秸秆还田与氮肥管理对直播杂交稻氮素利用特征的影响

【目的】探明秸秆还田和氮肥管理对麦/油后直播杂交稻氮素积累、转运、氮肥利用效率及籽粒产量的影响。【方法】选用优质三系杂交稻宜香优2115,采用二因素裂区设计,麦、油茬田同步开展试验,处理完全一致。主区为麦/油秸秆全量翻埋还田(M₁)和秸秆不还田(对照,M₀),副区设4个氮肥管理,即不施氮对照(N₀)、m_基肥∶m_分蘖肥∶m_促花肥∶m_保花肥比例分别为1∶0∶0∶0(N₁)、3∶3∶2∶2(N₂)、2∶2∶3∶3(N₃),测定了直播杂交稻主要生育时期各器官的氮素积累量及籽粒产量。【结果】结果表明,两种轮作方式下,氮肥管理对直播杂交稻主要生育时期的氮素积累、齐穗后茎鞘、叶片的氮素转运及稻株氮素利用效率均存在显著或极显著的调控效应。秸秆还田显著提高麦/油茬杂交稻中后期的氮素积累量、茎鞘和叶片的氮素转运量以及氮肥利用效率,其中,氮肥农学利用率、氮肥偏生产力和氮肥表观利用率较秸秆不还田分别提高了34.96%/28.76%、2.52%/2.61%和31.91%/22.30%。同时,油菜秸秆还田下直播杂交稻各生育时期氮素积累和产量较麦秆还田表现更好,籽粒产量提高481 kg/hm²(5.22%)。M₁N₂处理、M₀N₃处理下,直播杂交稻各阶段的氮素积累速率明显加大,促进结实期茎鞘和叶片的氮素向穗部转运,成熟期稻株氮素积累量优势明显且有较高的氮素利用效率(麦/油茬稻氮肥农学利用率、偏生产力和表观利用率分别达17.87 kg∙kg^-1/17.85 kg∙kg^-1、67.27 kg∙kg^-1/71.28 kg∙kg^-1、74.93%/75.05%),最终实现高产。【结论】在麦/油-稻轮作下秸秆全量还田,配合N₂氮肥管理,可有效提高直播杂交稻氮素吸收、利用效率,增加籽粒产量,尤以油菜秸杆还田的效果更好。     

秸秆还田对东北半干旱区玉米‖花生轮作系统土壤水热特性及作物产量的影响

以东北半干旱区玉米‖花生轮作系统为研究对象,试验设传统玉米单作、花生单作、不同秸秆还田量下(30%、60%和100%)玉米‖花生轮作5个处理。通过连续3年田间定位试验,研究玉米秸秆不同还田量对农田土壤水热特性、产量及土地当量比(LER)的影响。结果表明,与传统单作相比,秸秆还田处理玉米区播前0～40 cm土壤平均含水量提高了1.9%～3.9%,花生区提高了11.0%～13.9%;成熟期,秸秆还田花生区0:00～8:00平均地温比传统单作提高了0.2℃～1.1℃,10:00～22:00平均地温较传统单作降低了0.5℃～1.0℃。2016年和2017年玉米‖花生轮作系统LER为1.05～1.16,2018年各处理LER均小于1。吉林半干旱区玉米‖花生轮作系统进行秸秆还田改善了作物播前的土壤水分状况,但对于作物产量的影响在不同年份有所差异。     

Intensive organic vegetable production increases soil organic carbon but with a lower carbon conversion efficiency than integrated management

Intensive vegetable production in greenhouses has rapidly expanded in China since the 1990s and increased to 1.3 million ha of farmland by 2016, which is the highest in the world. We conducted an 11‐year greenhouse vegetable production experiment from 2002 to 2013 to observe soil organic carbon (SOC) dynamics under three management systems, i.e., conventional (CON), integrated (ING), and intensive organic (ORG) farming. Soil samples (0–20 and 20–40 cm depth) were collected in 2002 and 2013 and separated into four particle‐size fractions, i.e., coarse sand (> 250 µm), fine sand (250–53 µm), silt (53–2 µm), and clay (< 2 µm). The SOC contents and δ¹³C values of the whole soil and the four particle‐size fractions were analyzed. After 11 years of vegetable farming, ORG and ING significantly increased SOC stocks (0–20 cm) by 4008 ± 36.6 and 2880 ± 365 kg C ha^?1 y^?1, respectively, 8.1‐ and 5.8‐times that of CON (494 ± 42.6 kg C ha^?1 y^?1). The SOC stock increase in ORG at 20–40 cm depth was 245 ± 66.4 kg C ha^?1 y^?1, significantly higher than in ING (66 ± 13.4 kg C ha^?1 y^?1) and CON (109 ± 44.8 kg C ha^?1 y^?1). Analyses of ¹³C revealed a significant increase in newly produced SOC in both soil layers in ORG. However, the carbon conversion efficiency (CE: increased organic carbon in soil divided by organic carbon input) was lower in ORG (14.4%–21.7%) than in ING (18.2%–27.4%). Among the four particle‐sizes in the 0–20 cm layer, the silt fraction exhibited the largest proportion of increase in SOC content (57.8% and 55.4% of the SOC increase in ORG and ING, respectively). A similar trend was detected in the 20–40 cm soil layer. Over all, intensive organic (ORG) vegetable production increases soil organic carbon but with a lower carbon conversion efficiency than integrated (ING) management.     

基于GIS与RS的北方防沙屏障带生态系统格局演变

研究北方防沙屏障带生态系统格局演变特征,对加强屏障带建设、改善生态环境具有重要意义。以北方防沙屏障带为研究区,根据2005、2010、2015年MODIS遥感影像,将北方防沙屏障带生态系统类型划分为森林、草地、湿地、农田、城镇、荒漠和裸地7类。采用空间统计、转移矩阵、景观指数分析法、PNTIL模型等方法,对北方防沙屏障带2005—2015年生态系统类型时空演变特征进行分析。结果表明,2005—2015年北方防沙屏障带生态系统整体呈稳定状态;从分屏障带看,内蒙古防沙屏障带荒漠面积明显减少,塔里木防沙屏障带森林面积迅速增加,而河西走廊防沙屏障带草地面积明显增加;从正/逆转换方向来看,屏障带内农田、城镇、荒漠及裸地生态系统正向转换率高于逆向转换率。10年间,在景观水平上,北方防沙屏障带斑块数量、斑块密度呈减少趋势;在类型水平上荒漠景观整体呈现破碎化萎缩的趋势特征,森林、草地向形状简单化的方向发展。研究表明,2005—2015年,北方防沙屏障带生态总体向好,治沙效果明显,但局部仍面临较大压力。     

Effect of CHOP expression knock-down on apoptosis of renal tubular epithelial cells

AIM:To study the effect of C/EBP homologous protein (CHOP) on the apoptosis of renal tubular epithelial HK2 cells. METHODS:The serum mRNA levels of CHOP in the patients with acute kidney injury and healthy controls were detected by qPCR. In vitro, renal tubular epithelial HK2 cells were divided into control group, negative group (transfected with negative control siRNA), si-CHOP group (transfected with CHOP siRNA), and induced by transforming growth factor-β1 (TGF-β1). The viability of the cells was measured by MTT assay, and the apoptotic rate was analyzed by flow cytometry. The protein levels of nuclear antigen Ki-67, proliferating cell nuclear antigen (PCNA), caspase-3 and cleaved caspase-3 were determined by Western blot. RESULTS:Compared with the healthy controls, the serum mRNA levels of CHOP in the patients with acute kidney injury were increased significantly (P<0.05). Transfection with CHOP siRNA significantly decreased the expression of CHOP in the renal tubular epithelial HK2 cells (P<0.05). Knock-down of CHOP expression by siRNA significantly increased the viability of renal tubular epithelial HK2 cells (P<0.05), decreased the apoptotic rate (P<0.05), increased the expression of Ki-67 and PCNA (P<0.05), and down-regulated the protein level of cleaved caspase-3 (P<0.05). CONCLUSION:The serum mRNA levels of CHOP were increased in the patients with acute kidney injury. Knock-down of CHOP expression inhibits the apoptosis of renal tubular epithelial cells by regulating the expression of proliferation-and apoptosis-related proteins.