MicroRNA-502-3p regulates GABAergic synapse function in hippocampal neurons

Gamma-aminobutyric acid(GABA)ergic neurons,the most abundant inhibitory neurons in the human brain,have been found to be reduced in many neurological disorders,including Alzheimer's disease and Alzheimer's disease-related dementia.Our previous study identified the upregulation of microRNA-502-3p(miR-502-3p)and downregulation of GABA type A receptor subunitα-1 in Alzheimer's disease synapses.This study investigated a new molecular relationship between miR-502-3p and GABAergic synapse function.In vitro studies were perfo rmed using the mouse hippocampal neuronal cell line HT22 and miR-502-3p agomiRs and antagomiRs.In silico analysis identified multiple binding sites of miR-502-3p at GABA type A receptor subunitα-1 mRNA.Luciferase assay confirmed that miR-502-3p targets the GABA type A receptor subunitα-1 gene and suppresses the luciferase activity.Furthermore,quantitative reve rse transcription-polymerase chain reaction,miRNA in situ hybridization,immunoblotting,and immunostaining analysis confirmed that overexpression of miR-502-3p reduced the GABA type A receptor subunitα-1 level,while suppression of miR-502-3p increased the level of GABA type A receptor subunitα-1 protein.Notably,as a result of the overexpression of miR-502-3p,cell viability was found to be reduced,and the population of necrotic cells was found to be increased.The whole cell patch-clamp analysis of human-GABA receptor A-α1/β3/γ2L human embryonic kidney(HEK)recombinant cell line also showed that overexpression of miR-502-3p reduced the GABA current and overall GABA function,suggesting a negative correlation between miR-502-3p levels and GABAergic synapse function.Additionally,the levels of proteins associated with Alzheimer s disease were high with miR-502-3p overexpression and reduced with miR-502-3p suppression.The present study provides insight into the molecular mechanism of regulation of GABAergic synapses by miR-502-3p.We propose that micro-RNA,in particular miR-502-3p,could be a potential therapeutic to rget to modulate GABAergic synapse function in neurological disorders,including Alzheimer's disease and Alzheimer's diseaserelated dementia.

Isolation and Characterization of Storage Protein Subunit-null-dwarf Mutants in Soybean(Glycine max(L.) Merr.)

Dwarfing is useful to reduce plant height,when breeding high-yielding and non-lodging crops.In this study,a set of natural storage protein subunit-null dwarf mutants of soybean was reported that showed strongly reduced plant stature and deficiency in various 7S and 11S subunits,designated as snd1 mutants.Under normal growth conditions,the snd1 mutants showed a severe dwarf phenotype,with plant height of about 25 cm.Compared with wild-type DN47,the mutant snd1 exhibited no obvious morphological differences at the early stage of development.All the snd1 mutants examined had fewer nodes and shorter than normal internodes;the leaves were similar in shape to normal parents,but were dark-green at the mature stage.The flower size was similar to DN47;however,the flowering period was shorter than in the wild-type.Significant variation was noted for protein content,oil content of the seeds and size of seeds(weight of 100 seeds)among 17 snd1 dwarf lines.Genetic analysis indicated that the dwarfism of snd1 was controlled by a single recessive gene.The snd1 dwarf mutant had markedly different dynamic levels of the endogenous hormones gibberellin(GA),brassinosteroid,indole-3-acetic acid and abscisic acid,at the seedling stage.Exogenous GA3 treatment led to recovery of the plant height phenotype of the snd1 mutant;GA3 at 0.1 mm had the largest effect on enhancing plant height.Using molecular markers,snd1 gene was approximately mapped in an interval of 603 kb between markers Satt166 and Satt561 on chromosome 19.Snd1 mutant provided valuable material for hypoallergenic soybean breeding and the snd1 gene might be a novel gene related to plant height in soybean.

Inhibition of DNA-dependent Protein Kinase Catalytic Subunit by Small Molecule Inhibitor NU7026 Sensitizes Human Leukemic K562 Cells to Benzene Metabolite-induced Apoptosis

Benzene is an established leukotoxin and leukemogen in humans. We have previously re- ported that exposure of workers to benzene and to benzene metabolite hydroquinone in cultured cells induced DNA-dependent protein kinase catalytic subunit （DNA-PKcs） to mediate the cellular response to DNA double strand break （DSB） caused by DNA-damaging metabolites. In this study, we used a new, small molecule, a selective inhibitor of DNA-PKcs, 2-（morpholin-4-yl）-benzo[h]chomen-4-one （NU7026）, as a probe to analyze the molecular events and pathways in hydroquinone-induced DNA DSB repair and apoptosis. Inhibition of DNA-PKcs by NU7026 markedly potentiated the apoptotic and growth inhibitory effects of hydroquinone in proerythroid leukemic K562 cells in a dose-dependent manner. Treatment with NU7026 did not alter the production of reactive oxygen species and oxidative stress by hydroquinone but repressed the protein level of DNA-PKcs and blocked the induction of the kinase mRNA and protein expression by hydroquinone. Moreover, hydroquinone increased the phos- phorylation of Akt to activate Akt, whereas co-treatment with NU7026 prevented the activation of Akt by hydroquinone. Lastly, hydroquinone and NU7026 exhibited synergistic effects on promoting apop- tosis by increasing the protein levels of pro-apoptotic proteins Bax and caspase-3 but decreasing the protein expression of anti-apoptotic protein Bcl-2. Taken together, the findings reveal a central role of DNA-PKcs in hydroquinone-induced hematotoxicity in which it coordinates DNA DSB repair, cell cycle progression, and apoptosis to regulate the response to hydroquinone-induced DNA damage.

大豆7S球蛋白α-亚基缺失型种质创新

采用常规杂交育种方法,人工去雄、授粉,以10份综合农艺性状优良的黑龙江省主栽品种(或优良品系)为母本、亚基组成为(α'+α+11S酸性亚基)-缺失型育种材料日B为父本进行有性杂交。将F1杂交种南繁,单粒点播、单株收获得到F2代分离群体。聚丙烯酰胺凝胶电泳(SDS-PAGE)法分析表明,F1代杂交种子7S球蛋白α'-与α-亚基的表现型全部为正常型。在杂交F2代种子中获得了具有中国大豆遗传背景的α-缺失、(α+A1aA1bA2)-缺失、A3-缺失、(α'+A4)-缺失和(α'+α)-缺失的贮藏蛋白亚基组成新类型种质,为我国大豆蛋白组分育种选择提供了重要的中间材料。

SSR标记辅助回交转育大豆7S球蛋白α-亚基致敏蛋白缺失新品系

为快速培育7S球蛋白致敏蛋白α-亚基缺失的优质大豆新品系,以(α'+α)-亚基双缺失型材料日B为供体亲本,东农47为受体亲本,以杂交-回交转育方法为基础,结合SSR标记辅助遗传背景选择与主要农艺性状鉴定及品质分析,在BC2F4选育到7S球蛋白α-亚基缺失新品系Cb80。Cb80综合农艺性状优良,遗传背景回复率大于90%,其蛋白质及氨基酸总量为44.1%、41.95%,分别比轮回亲本东农47提高3.5和3.76个百分点。

Losartan Protects Podocytes against High Glucose-induced Injury by Inhibiting B7-1 Expression

The role of B7-1 in podocyte injury has received increasing attention.The aim of this study was to investigate whether losartan protects podocytes of patients with diabetic kidney disease(DKD)by regulating B7-1 and the underlying mechanisms.Rats with streptozotocin-induced DKD were treated with losartan for 8 weeks.Biochemical changes in blood and urine were analyzed.Kidneys were isolated for electron microscopy,immunofluorescence,real-time quantitative PCR(RT-PCR),and Western blot analysis.Immortalized mouse podocyte cells were cultured in normal or high glucose medium in the presence or absence of losartan for 48 h,and then the cells were collected for immunofluorescence,PCR,Western blotting and monolayer permeability detection.The phosphatidylinositol 3-kinase(PI3K)110a subunit and angiotensin II type 1 receptor(AT1R)plasmids were transfected into podocytes,respectively,and then Western blotting was performed to assess the expression of B7-1 protein.The results showed that losartan ameliorated podocyte structure and function in the rat model of DKD,and reduced the expression of B7-1 protein.Overexpression of PI3K 110a subunit in podocytes attenuated the inhibitory effect of losartan on B7-1 expression in high glucose-stimulated podocytes.The expression of B7-1 was significantly increased by overexpression of ATI R and significantly reduced by blocking PI3K 110a subunit.We conclude that losartan protects podocytes against high glucose-induced injury by inhibiting AT1R-mediated B7-1 expression.This effect is dependent on the AT1R-PI3K 110a subunit pathway.

Examination of Subunit Composition of Bombyx Mori Silk Fibroin

Main subunits of the silk fibroin were separated by GFC(gel filtration chromatography)technigue.The nativesilk fibroin and α,c subunits were measured by gel elec-trophoresis.The aminoacid compositions of the nativesilk fibroin and α,c subunits were analyzed by means ofaminoacid measurement.Some properties of silk was inter-preted in view of subunit composition of silk fibroin.

CD4^(+)T细胞亚型及其相关炎性因子在NE-FE-COPD患者中的表达及意义

目的探讨中性粒细胞优势型频繁急性加重型的慢性阻塞性肺疾病(NE-FE-COPD)患者CD4^(+)T细胞亚型及其相关炎性因子的表达及意义。方法选取2019年3月至2021年3月在该院治疗的COPD患者为研究对象,按照不同表型分为非频繁急性加重型COPD组(IE-COPD组,n=11)、嗜酸性粒细胞优势型频繁急性加重型COPD组(Eos-FE-COPD组,n=13)、中性粒细胞优势型频繁急性加重型COPD组(NE-FE-COPD组,n=15),肺功能正常及吸烟史>10包/年者为对照组(CTRL组,n=9)。采集各组支气管肺泡灌洗液(BALF),流式细胞术检测CD4^(+)T细胞亚型表达,ELISA测定炎性因子水平,分析其与肺功能和急性加重频率的相关性。CD4^(+)CD28^(null)T细胞和CD4^(+)CD28^(+)T细胞与人气道上皮细胞(hAECs)进行共培养后分为co-culture组和Control组,免疫荧光染色观察hAECs紧密连接损伤情况,RT-qPCR和Western blot检测ZO-1和occludin mRNA及蛋白表达水平。结果NE-FE-COPD组BALF中CD4^(+)CD28^(null)T细胞占比、白细胞介素(IL)-1β水平高于CTRL组、IE-COPD组、Eos-FE-COPD组,差异有统计学意义(P<0.05)。CD4^(+)CD28^(null)T细胞占比、IL-1β水平与肺功能呈负相关(P<0.05),与急性加重频率呈正相关(P<0.05)。与Control组比较,co-culture组hAECs紧密连接受损,ZO-1和occludin mRNA及蛋白表达水平降低,差异有统计学意义(P<0.05)。结论CD4^(+)CD28^(null)T细胞和IL-1β可能参与NE-FE-COPD的发生和发展。

大豆7S、11S球蛋白亚基缺失突变体豆腐加工特性的鉴定

以4份7S、11S球蛋白不同亚基组成大豆种质为试材,采用小样本大豆加工方法制作豆腐,测定其豆腐产量、豆腐干重(DTW)、豆腐蛋白含量(PC)、豆腐硬度和豆浆蛋白率(RPA)等豆腐品质指标,并分析和鉴定其豆腐加工特性来明晰以上四种大豆种质在豆腐品质加工上的利用潜力。结果表明,大豆7S球蛋白亚基表现分别为:α'-亚基缺失,[(α'+α)+11S(Ⅱb+Ⅱa)]-亚基缺失和β-亚基极少型变异种质日A-5、日B-1和越南褐脐做成的豆腐在豆腐产量、豆腐干重、豆腐蛋白质含量、豆腐硬度和豆浆蛋白率等品质性状上存在显著的差异,四个大豆材料间的湿、干豆腐产量的遗传变异系数较大。α'-亚基缺失型日A-5是适用于以硫酸钙作为凝固剂加工豆腐的大豆种质资源。低大豆7S球蛋白品系日B-1因同时伴随有11S酸性亚基缺失,所制豆腐硬度低、成型差。越南引进的越南褐脐(β-少型)可在优质豆腐用品种亲本选配中加以利用。

大豆脂氧酶与7S球蛋白亚基双缺失株系的主要品质性状分析

本研究以脂氧酶完全缺失品系SI0162为母本,以α'-、α-、β-亚基完全缺失品系1003-44为父本,从后代中成功地获得了大豆脂氧酶及7S球蛋白亚基双缺失材料。对双缺失材料的蛋白质、脂肪含量及组分等进行了测定,并与黑龙江省主栽品种黑河38进行比较,以探究脂氧酶与7S球蛋白亚基双缺失材料的相关品质性状。研究结果表明:双缺失材料蛋白质平均含量为39.89%,与黑河38基本持平;脂肪平均含量为18.73%,比黑河38低1.79%;含硫氨基酸含量为1.45%,比黑河38高25.0%;脂肪酸中油酸含量比黑河38高13.3%。观察表明双缺失的大豆种质能够正常生长发育,同时在含硫氨基酸及油酸含量等有益人类健康的成分上的变化符合育种目标,为培育食品加工专用大豆品种提供了基础材料。

Regression of cardiovascular remodeling in hypertension:Novel relevant mechanisms

Asymptomatic organ damage due to progressive kidney damage, cardiac hypertrophy and remodeling put hypertensive patients at high risk for developing heart and renal failure, myocardial infarction and stroke. Current antihypertensive treatment normalizes high blood pressure, partially reverses organ damage, and reduces the incidence of heart and renal failure. Activation of the renin-angiotensin system(RAS) is a primary mechanism of progressive organ damage and, specifically, a major cause of both renal and cardiovascular fibrosis. Currently, inhibition of the RAS system [mainly with angiotensin I converting enzyme inhibitors or angiotensin II(Ang II) receptor antagonists] is the most effective antihypertensive strategy for normalizing blood pressure and preventing target organ damage. However, residual organ damage and consequently high risk for cardiovascular events and renal failure still persist. Accordingly, in hypertension, it is relevant to develop new therapeutic perspectives, beyond reducing blood pressure to further prevent/reduce target organ damage by acting on pathways that trigger and maintain cardiovascular and renal remodeling. We review here relevant novel mechanisms of target organ damage in hypertension, their role and evidence in prevention/regression of cardiovascular remodeling and their possible clinical impact as well. Specifically, we focus on the signaling pathway Rho A/Rho kinase, on the impact of the vasodilatory peptides from the RAS and some insights on the role of estrogens and myocardial chymase in cardiovascular hypertensive remodeling.

Gabapentinoid Insensitivity after Repeated Administration is Associated with Down-Regulation of theα2δ-1 Subunit in Rats with Central Post-Stroke Pain Hypersensitivity

The α2δ-1 subunit of the voltage-gated Ca2＋ channel （VGCC） is a molecular target of gabapentin （GBP）, which has been used as a first-line drug for the relief of neuropathic pain. GBP exerts its anti-nociceptive effects by disrupting trafficking of the α2δ-1 subunit to the presynaptic membrane, resulting in decreased neurotrans- mitter release. We previously showed that GBP has an anti- allodynic effect in the first two weeks; but this is followed by insensitivity in the later stage after repeated adminis- tration in a rat model of central post-stroke pain （CPSP） hypersensitivity induced by intra-thalamic hemorrhage. To explore the mechanisms underlying GBP insensitivity, the cellular localization and time-course of expression of the α2δ-1 subunit in both the thalamus and spinal dorsal horn were studied in the same model. We found that the α2δ-1 subunit was mostly localized in neurons, but not astrocytes and microglia. The level of α2δ-1 protein increased in the first two weeks after injury but then decreased in the third week, when GBP insensitivity occurred. Furthermore, the c^2g-1 down-regulation was likely caused by later neuronal loss in the injured thalamus through a mechanism other than apoptosis. In summary, the present results suggest that the GBP receptor ~2^-1 is mainly expressed in thalamic neurons in which it is up-regulated in the early stage of CPSP but this is followed by dramatic down-regulation, which is likely associated with GBP insensitivity after long-term use.

域外传真

Ottakringer Helles;UO - Urban Ottakringer;Null Komma Josef.

High temperature increases centromeremediated genome elimination frequency and enhances haploid induction in Arabidopsis

Double haploid production is the most effective way to create true-breeding lines in a single generation.In Arabidopsis,haploid induction via mutation of the centromere-specific histone H3(cenH3)has been shown when the mutant is outcrossed to the wild-type,and the wild-type genome remains in the haploid progeny.However,factors that affect haploid induction are still poorly understood.Here,we report that a mutant of the cenH3 assembly factor Kinetochore Null2(KNL2)can be used as a haploid inducer when pollinated by the wild-type.We discovered that short-term temperature stress of the knl2 mutant increased the efficiency of haploid induction 10-fold.We also demonstrated that a point mutation in the CENPC-k motif of KNL2 is sufficient to generate haploid-inducing lines,suggesting that haploidinducing lines in crops can be identified in a naturally occurring or chemically induced mutant population,avoiding the generic modification(GM)approach at any stage.Furthermore,a cenh3-4 mutant functioned as a haploid inducer in response to short-term heat stress,even though it did not induce haploids under standard conditions.Thus,we identified KNL2 as a new target gene for the generation of haploid-inducer lines and showed that exposure of centromeric protein mutants to high temperature strongly increases their haploid induction efficiency.