Association of β3 Adrenergic Receptor and Peroxisome Proliferator-activated Receptor Gamma 2 Polymorphisms With Insulin Sensitivity:A Twin Study

Objective To study the effect of β3 adrenergic receptor （β3AR） Trp64Arg and peroxisome proliferator activated receptor gamma 2 （PPAR72） Prol2Ala polymorphisms on insulin resistance. Methods One hundred and eight dizygotic twin pairs were enrolled in this study. Microsatellite polymorphism was used to diagnose zygosity of twins. Insulin sensitivity was estimated with logarithm transformed homeostasis model assessment （HOMA）. PCR-RFLP analysis was performed to detect the variants. As a supplement to the sib-pair method, identity by state （IBS） was used to analyze the association of polymorphisms with insulin sensitivity. Results The genotype frequencies of Trp64Trg, Trp64Arg, and Arg64Arg were 72.3%, 23.8%, and 3.9%, respectively, while the genotype frequencies of Pro12Pro, Pro12Ala, and Ala12Ala were 89.9%, 9.6%, and 0.5%, respectively. For β3AR Trp64Arg the interclass co-twin correlations of Waist-to-hip ratio （WHR）, blood glucose （GLU）, and insulin （INS）, homeostasis model assessment insulin resistance index （HOMA-IR） of the twin pairs sharing 2 alleles of IBS were greater than those sharing 0-1 allele of IBS, and HOMA4R had statistic significance. For PPAR3t2 Prol2Ala most traits of twin pairs sharing 2 alleles of IBS had greater correlations and statistic significance in body mass index （BMI）, WHR, percent of body fat （PBF） and GLU, but there were low correlations of either insulin or HOMA-IR of twin pairs sharing 1 or 2 alleles of IBS. The combined effects of the two variations showed less squared significant twin-pair differences of INS and HOMA-IR among twins sharing 4 alleles of IBS. Condusions β3AR Trp64Arg and PPAR）,2 Pro 12Ala polymorphisms might be associated with insulin resistance and obesity, and there might be slight synergistic effects between this two gene loci, and further studies are necessary to confirm this finding.

AduoLa Fuzhenglin Down-regulates Microwave-induced Expression of β_1-adrenergic Receptor and Muscarinic Type 2 Acetylcholine Receptor in Myocardial Cells of Rats

This paper is aimed to study the effect of ADL on expression of ~z-AR and Mz-AchR in myocardial cells of rats exposed to microwave radiation. Immunohistochemistry, Western blot and image analysis were used to detect the expression of ~I-AR and Mz-AchR in myocardial cells at 7 and 14 d after microwave exposure. The results show that the expression level was higher in microwave exposure group and 0.75 g/（kg.d） ADL group than in sham operation group and significantly lower in 1.5 and 3.0 g/（kg.d） ADL groups than in microwave group. So we have a conclusion that the expression of I^z-AR and Mz-AchR is down-regulated in myocardial cells of rats exposed to microwave radiation. ADL can protect rats against microwave-induced heart tissue injury.

Blocking beta 2-adrenergic receptor inhibits dendrite ramification in a mouse model of Alzheimer's disease

Dendrite ramification affects synaptic strength and plays a crucial role in memory. Previous studies revealed a correlation between beta 2-adrenergic receptor dysfunction and Alzheimer＇s disease （AD）, although the mechanism involved is still poorly understood. The current study investigated the potential effect of the selective β2-adrenergic receptor antagonist, ICI 118551 （ICI）, on Aβ deposits and AD-related cognitive impairment. Morris water maze test results demonstrated that the performance of AD-transgenic （TG） mice treated with ICI （AD-TG/ICI） was significantly poorer compared with NaCl-treated AD-TG mice （AD-TG/NaCl）, suggesting that β2-adrenergic receptor blockage by ICI might reduce the learning and memory abilities of mice. Golgi staining and immunohistochemical staining revealed that blockage of the β2-adrenergic receptor by ICI treatment decreased the number of dendritic branches, and ICI treatment in AD-TG mice decreased the expression of hippocampal synaptophysin and synapsin 1. Western blot assay results showed that the blockage of β2-adrener- gic receptor increased amyloid-β accumulation by downregulating hippocampal a-secretase activity and increasing the phosphorylation of amyloid precursor protein. These findings suggest that blocking the β2-adrenergic receptor inhibits dendrite ramification of hippocampal neurons in a mouse model of AD.

Relationship between catecholamine level and gene polymorphism of β1 adrenergic receptor G1165C in children with EV71 infection in hand foot and mouth disease

Objective:To investigate the relationship between the levels of plasma adrenaline and norepinephrine and gene polymorphism of β1 adrenergic receptor G1165 C in children with enterovirus 71(EV71) infection in hand foot and mouth disease(HFMD). Methods:The polymerase chain reaction(PCR) was used to detect the expression of gene polymorphism of β1 adrenergic receptor G1165 C in vitro. The levels of plasma adrenaline and norepinephrine were measured by enzyme-linked immunosorbent assay(ELISA). Results:The plasma norepinephrine level of severe group was significantly higher than the mild group in children with EV71 infection in HFMD(P<0.05); however,the levels of plasma adrenalinein in two groups had no statistical differences(P>0.05); There was no significant difference in the distribution of β1 adrenergic receptor G1165 C genotype and allele between EV71 infection group and healthy control group(P> 0.05). Further analysis of EV71 infection group by dividing it into mild and severe groups showed that there was no significant difference in the distribution of genotype and allele between these two groups as well(P> 0.05). There was no significant difference in the levels of epinephrine and norepinephrine in different genotypes of EV71 infection group(P> 0.05),and in the levels of plasma epinephrine and norepinephrine in the mild and severe groups(P> 0.05). Conclusions:As the disease gets worse,the plasma norepinephrine level has a rising trend in children with EV71 infection in HFMD,which is an important indicator to evaluate the progress of the disease. However,the gene polymorphism of eptor G1165 C have no significant correlation,not only with the susceptibility and severit β1 adrenergic recy of EV71 infection in hand,foot and mouth disease,but also with the levels of catecholamine.

Association between essential hypertension and polymorphisms of beta 1 adrenergic receptor gene G1165C (Gly389Arg) in Chinese Mongolian population

BACKGROUND： The prevalences of hypertension, cerebrovascular diseases, etc. are higher in Mongolian population because of the influence of various factors including genetics, geography, diet, etc. Therefore, it is helpful to develop researches on the genetics of various diseases including hypertension in Mongolian population. OBJECTIVE： To analyze the association between the polymorphism of beta1 adrenergic receptor （β1-AR） gene G1165C （Arg389Gly）, an important candidate gene for various diseases of cardiovascular system, and essential hypertension in Mongolian population. DESIGN ： A cross-sectional study SETTINGS： Department of Neurology, the First Affiliated Hospital of Inner Mongolia Medical College; Wulate Houqi Red Cross Society. PARTICIPANTS： The survey was carried out from February 2003 to March 2005. Totally 239 Mongolian residents, whose blood relations of 3 generations were all Mongolians, were selected from Wulate Houqi, Inner Mongolia, and they were all informed with the survey and detected items. Based on the diagnostic standard of hypertension set by WHO in 1999, the subjects were divided into two groups according to the level blood pressure： ① Normal blood pressure group （n=117）： systolic blood pressure （SBP） 〈 140 mm Hg （1 mm Hg =0.133 kPa）, diastolic blood pressure （DBP） 〈 90 mm Hg, and those having histories of cerebrovascular disease, heart disease, diseases of liver, kidney and tiroides, and diabetes mellitus were excluded. ② Essential hypertension group （n=122）： including 51 patients with simple high SBP. All the enrolled subjects had no blood relationship with each other, and had no history of miscegenation. METHODS ： The body height, body mass, waist circumference and blood lipids were measured routinely, and their habits of smoking and drinking were also investigated. Penpheral venous blood （5 mL） was drawn, the genome DNA was extracted, and the polymorphisms of the β1-AR Gl165C （Gly389Arg） genotype were detected with the Sequenom system. Polymerase chain reaction （PCR） experiment and SNP detection were performed in Huada Gene Laboratory of Bejing, then the univariate analysis of variance was applied in the sample comparison among groups, and the chi-square test was used to compare the genotypes and allele frequencies. The odd ratio （OR） and 95% confidence interval （CO were calculated. MAIN OUTCOME MEASURES： The distributions of β1-AR Gl165C （Gly389Arg） genotypes and alleles were observed. RESULTS： A11 the 239 subjects were involved in the analysis of results, and no one missed, ①Comparison of β1-AR G1165C （Gly389Arg） genotypes and allele distnbutions： In Mongolian population, the frequencies of CC and GG＋GC genotypes at β1-AR G1165C （Gly389Arg） site in the essential hypertension group （72%, 28%） were not significantly different from those in the normal blood pressure group （67%, 33%） （xz=0.841, P=-0.359; OR 0.773, 95%Cl： 0.445-1.342）; The frequencies of C and G alleles also had no significant differences between the essential hypertension group （85%, 15%） and the normal blood pressure group （82%, 18%） （x^2=1.136, P=-0.287; OR： 0.769, 95%Cl： 0.747-1.248）. ②The frequencies of CC and GG＋GC genotypes at β1-AR G1165C （Gly389Arg） site had no significant differences between the patients with simple high SBP （71%, 29%） and the normal blood pressure group （x^2=0.250, P=-0.617; OR： 0.833, 95%C/： 0.408-1.703）; The frequencies of C and G alleles were not significantly different between the patients with simple high SBP （86%, 14%） and the normal blood pressure group （x^2=0.670, P=-0.413; OR 0.766, 95%Cl： 0.404-1.453）. CONCLUSION： In Mongolian population, the distributions of the genotypes and alleles of β1-AR Gl165C （Gly389Arg） have no obvious differences between the subjects with normal blood pressure and the patients with essential hypertension （including simple SBP increase）, which suggests that G1165C （Glu389Asp） site of β1-AR gene may be not a genetic mark of essential hypertension and simple high SBP in Mongolian population.

B2 adrenergic receptors and morphological changes of the enteric nervous system in colorectal adenocarcinoma

AIM To study the morphology of the enteric nervous system and the expression of beta-2 adrenergic(B2A) receptors in primary colorectal cancer.METHODS In this study, we included forty-eight patients with primary colorectal cancer and nine patients for control tissue from the excision of a colonic segment for benign conditions. We determined the clinicopathological features and evaluated the immunohistochemical expression pattern of B2 A receptors as well as the morphological changes of the enteric nervous system(ENS). In order to assess statistical differences, we used the student t-test for comparing the means of two groups and one-way analysis of variance with Bonferroni's post hoc analysis for comparing the means of more than two groups. Correlations were assessed using the Pearson's correlation coefficient.RESULTS B2 A receptors were significantly associated with tumor grading, tumor size, tumor invasion, lymph node metastasis(P < 0.05), while there were no statistically significant associations with gender, CRC location and gross appearance(P > 0.05). We observed, on one hand, a decrease of the relative area for both Auerbach and Meissner plexuses with the increase of the tumor grading, and on the other hand, an increase of the relative area of other nervous elements not in the Meissner plexus or in the Auerbach plexus with the tumor grading. For G1 tumors we found that epithelial B2 A area showed an inverse correlation with the Auerbach plexus areas [r(14) =-0.531, P < 0.05], while for G2 tumors, epithelial B2 A areas showed an indirect variation with both the Auerbach plexus areas [r(14) =-0.453, P < 0.05] and the Meissner areas [r(14) =-0.825, P < 0.01]. For G3 tumors, the inverse dependence increased for both Auerbach [r(14) =-0.587, P < 0.05] and Meissner [r(14) =-0.934, P < 0.05] plexuses.CONCLUSION B2 A receptors play an important role in colorectal carcinogenesis and can be utilized as prognostic factors. Furthermore, study of the ENS in colorectal cancer may lead to targeted molecular therapies.

肺腺癌组织中前列腺素内过氧化物合酶2表达水平与表皮生长因子受体基因突变的相关性分析

目的 分析肺腺癌组织中前列腺素内过氧化物合酶2(PTGS2)表达水平与表皮生长因子受体(EGFR)基因突变的相关性。方法 选取57例肺腺癌患者为研究对象,收集肺腺癌组织及其相应癌旁组织。采用实时荧光定量PCR(RT-PCR)法检测癌组织及癌旁组织中PTGS2 mRNA表达水平;采用免疫组织化学法分析PTGS2蛋白表达;采用RT-PCR法检测癌组织及癌旁组织EGFR基因突变情况。分析肺腺癌患者临床病理参数与PTGS2 mRNA水平、EGFR基因突变情况的关系。采用Spearman秩相关分析探讨肺腺癌患者EGFR基因突变情况与PTGS2 mRNA水平的相关性。结果 57例肺腺癌患者中,5例癌旁组织EGFR基因突变型患者,其对应癌组织也均发生突变,且为同一突变类型。26例(45.61%)癌组织EGFR基因突变型患者,未发现双重突变,其中19外显子突变17例(29.82%),均为缺失突变;21外显子突变9例(15.79%),均为L858R点突变。癌组织中PTGS2mRNA表达水平、PTGS2蛋白阳性率及EGFR基因突变型比例高于癌旁组织,EGFR基因野生型比例低于癌旁组织(P<0.01)。肺腺癌患者性别、TNM分期、吸烟与PTGS2 mRNA表达水平、EGFR基因突变情况有关(P<0.05或0.01)。EGFR基因突变型PTGS2 mRNA表达水平高于EGFR基因野生型(P<0.01)。肺腺癌患者EGFR基因突变与PTGS2 mRNA表达水平呈正相关(r=0.512,P<0.01)。结论 肺腺癌患者癌组织中PTGS2mRNA表达水平及PTGS2蛋白阳性率升高,且与EGFR基因突变关系密切,二者可能共同影响疾病进程。

miR-10b介导NKG2D调节脑胶质瘤细胞免疫效应的实验研究

目的:观察微小核糖核酸-10b(miR-10b)对脑胶质瘤细胞免疫效应的调节作用并探讨其作用机制。方法:取人脑胶质瘤细胞U251进行培养和传代,获得处于对数生长期的细胞。按照1.0×105个/ml浓度制备细胞悬液,并设置对照组、过表达组、低表达组、空白组,每组6个复孔。对照组、过表达组、低表达组分别采用脂质体转染法转染阴性对照、miR-10b模拟物、miR-10b抑制剂,空白组予以等量无菌生理盐水。分离和培养1例健康志愿者外周血自然杀伤(NK)细胞。MTT法检测不同效靶比时NK细胞的杀伤活性;流式细胞仪检测各组NK细胞表面NK细胞激活受体(NKG2D)表达,并检测各组人脑胶质瘤细胞U251表面主要组织相容性复合物Ⅰ链相关基因A(MICA)、UL16结合蛋白2(ULBP2)、UL16结合蛋白3(ULBP3)表达。结果:对照组、过表达组、低表达组转染效率分别为(93.55±2.05)%、(95.67±3.14)%、(94.18±3.26)%;与对照组和空白组相比,过表达组miR-10b表达升高,低表达组miR-10b表达降低,差异均有统计学意义(P<0.05),且对照组和空白组miR-10b表达差异无统计学意义(P>0.05);与对照组和空白组相比,过表达组NK细胞不同效靶比杀伤活性均降低、NKG2D表达降低,低表达组NK细胞不同效靶比杀伤活性均增高、NKG2D表达增高,差异均有统计学意义(P<0.05),各组NK细胞杀伤活性均随效靶比增加而增高,差异均有统计学意义(P<0.05),且对照组与空白组相比,相同效靶比NK细胞杀伤活性、NKG2D表达差异均无统计学意义(P>0.05);与对照组和空白组相比,过表达组人脑胶质瘤细胞U251表面MICA、ULBP2、ULBP3表达均降低,低表达组人脑胶质瘤细胞U251表面MICA、ULBP2、ULBP3表达均增高,差异均有统计学意义(P<0.05),且对照组与空白组人脑胶质瘤细胞U251表面MICA、ULBP2、ULBP3表达差异均无统计学意义(P>0.05)。结论:抑制miR-10b表达能够增加NK细胞表面NKG2D和人脑胶质瘤细胞U251表面MICA、ULBP2、ULBP3表达,增强NK细胞对人脑胶质瘤细胞U251的杀伤活性。

哮喘治疗中β_(2)-AR激动药诱发受体脱敏的发生机制及预防进展

β_(2)-肾上腺素受体(β_(2)-AR)激动药作为治疗支气管哮喘(以下简称“哮喘”)的一线药物,在临床中广泛应用,然而长期使用可导致β_(2)-AR脱敏,降低其临床疗效,致使部分哮喘患者症状控制欠佳。β_(2)-AR激动药引起β_(2)-AR脱敏的机制主要包括慢速脱敏(与气道黏膜β_(2)-AR密度减小有关)和快速脱敏(与刺激性G蛋白脱偶联机制有关)。环磷酸腺苷(cAMP)-蛋白激酶A和cAMPcAMP激活的交换蛋白信号通路与β_(2)-AR脱敏过程关系密切。糖皮质激素、过氧化物酶体增殖物激活受体γ激动药、ASM-024、中药单药及成方等与β_(2)-AR激动药联合使用时能改善β_(2)-AR的敏感性,从而更好地控制哮喘症状。

β_(2)-肾上腺素能受体与神经丝在山羊脾中的分布和共定位研究

为探讨肾上腺素能神经与脾功能联系,用免疫组织化学SP法和免疫荧光双标记法观察β_(2)-肾上腺素能受体(β_(2)-AR)和羊脾神经丝(NFs)在山羊脾中的表达分布特点。结果显示,β_(2)-AR免疫反应阳性产物在山羊脾中分布广泛,白髓、红髓、中央动脉、被膜、小梁、边缘区均有不同程度的着色,白髓中免疫组化染色阳性相对表达量极显著高于红髓(P<0.01),极极显著高于边缘区和被膜(P<0.001);红髓中免疫组化染色阳性相对表达量极显著高于边缘区和被膜(P<0.01);NFs免疫反应阳性产物在山羊脾中红髓、被膜、小梁、小血管、白髓和边缘区均有不同程度的着色,红髓中免疫组化染色阳性相对表达量显著高于白髓(P<0.05),极显著高于边缘区(P<0.01);被膜中免疫组化染色阳性相对表达量极显著高于边缘区和白髓(P<0.01)。共定位研究表明β_(2)-AR和NFs在3个区域存在共定位关系,在平滑肌细胞构成的被膜和小梁中存在,在富含平滑肌细胞的小血管(如中央动脉)中存在,在T、B淋巴细胞丰富区域(如白髓、红髓和边缘区)存在。表明交感-肾上腺素能神经系统对机体免疫具有调控作用。β_(2)-AR和NFs共定位结构证明脾受到交感-肾上腺素能神经的调控,交感神经纤维通过释放神经递质-去甲肾上腺素,作用于脾上的肾上腺素能受体实现功能调控。

Transcriptome sequencing and experiments reveal the effect of formyl peptide receptor 2 on liver homeostasis

BACKGROUND Formyl peptide receptor 2(Fpr2)is an important receptor in host resistance to bacterial infections.In previous studies,we found that the liver of Fpr2-/-mice is the most severely damaged target organ in bloodstream infections,although the reason for this is unclear.AIM To investigate the role of Fpr2 in liver homeostasis and host resistance to bacterial infections.METHODS Transcriptome sequencing was performed on the livers of Fpr2-/-and wild-type(WT)mice.Differentially expressed genes(DEGs)were identified in the Fpr2-/-and WT mice,and the biological functions of DEGs were analyzed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis.Quantitative real time-polymerase chain reaction(qRT-PCR)and western blot(WB)analyses were used to further validate the expression levels of differential genes.Cell counting kit-8 assay was employed to investigate cell survival.The cell cycle detection kit was used to measure the distribution of cell cycles.The Luminex assay was used to analyze cytokine levels in the liver.The serum biochemical indices and the number of neutrophils in the liver were measured,and hepatic histopathological analysis was performed.RESULTS Compared with the WT group,445 DEGs,including 325 upregulated genes and 120 downregulated genes,were identified in the liver of Fpr2-/-mice.The enrichment analysis using GO and KEGG showed that these DEGs were mainly related to cell cycle.The qRT-PCR analysis confirmed that several key genes(CycA,CycB1,Cdc20,Cdc25c,and Cdk1)involved in the cell cycle had significant changes.The WB analysis confirmed a decrease in the expression of CDK1 protein.WRW4(an antagonist of Fpr2)could inhibit the proliferation of HepG2 cells in a concentration dependent manner,with an increase in the number of cells in the G0/G1 phase,and a decrease in the number of cells in the S phase.Serum alanine aminotransferase levels increased in Fpr2-/-mice.The Luminex assay measurements showed that interleukin(IL)-10 and chemokine(C-X-C motif)ligand(CXCL)-1 levels were significantly reduced in the liver of Fpr2-/-mice.There was no difference in the number of neutrophils,serum C-reactive protein levels,and liver pathology between WT and Fpr2-/-mice.CONCLUSION Fpr2 participates in the regulation of cell cycle and cell proliferation,and affects the expression of IL-10 and CXCL-1,thus playing an important protective role in maintaining liver homeostasis.

β_(2)肾上腺素受体在erastin诱导的前列腺细胞铁死亡和自噬中的作用及其机制

目的探讨β2肾上腺素受体(ADRB2)在铁死亡激活剂erastin(Era)诱导的前列腺癌细胞铁死亡、自噬中的作用及可能的分子机制。方法取慢病毒或对照感染的PC-3细胞,设置sh-NC组(正常培养)、sh-NC+Era组(10μmol/L Era处理24 h)、sh-ADRB2组(正常培养)与sh-ADRB2+Era组(10μmol/L Era处理24 h)。采用CCK-8法检测铁死亡激活剂Era和铁死亡抑制剂ferrostatin-1(Fer-1)处理后的细胞存活率,透射电镜观察细胞形态变化,丙二醛(MDA)检测试剂盒检测MDA含量,铁离子比色法检测铁离子含量。Western blotting检测胱氨酸谷氨酸转运体(XCT)、铁蛋白重链1(FTH1)、谷胱甘肽过氧化物酶(GPX4)、p62、LC3、c-Jun N-末端激酶(JNK)、c-Jun、p-c-Jun蛋白表达水平。注射ADRB2敲低的PC-3稳转株,构建裸鼠成瘤模型并用Era处理,分为sh-NC组、sh-NC+Era组、sh-ADRB2组与sh-ADRB2+Era组,每组4只。隔天记录瘤体体积及最终瘤体重量,并采用免疫组化检测ADRB2、JNK、c-Jun、p-c-Jun的表达情况。结果Era处理后PC-3细胞存活率下降(P<0.01),加用Fer-1处理后PC-3细胞存活率恢复(P<0.01)。透射电镜下可见,Era处理后PC-3细胞发生铁死亡和自噬形态学改变,MDA和铁离子含量增高(P<0.05或P<0.01);敲低ADRB2并用Era处理后PC-3细胞存活率进一步下降(P<0.05),MDA和铁离子含量进一步升高(P<0.01),FTH1、XCT、GPX4和LC3蛋白表达水平降低(P<0.05或P<0.01),p62蛋白和JNK通路相关基因JNK、c-Jun、p-c-Jun蛋白表达水平升高(P<0.01);JNK抑制剂处理后,FTH1、XCT和LC3表达水平升高,p62表达水平降低(P<0.01)。PC-3裸鼠模型中,sh-ADRB2+Era组瘤体体积明显小于sh-NC+Era组和sh-ADRB2组(P<0.05或P<0.01);免疫组化检测结果显示,与sh-NC组比较,sh-ADRB2组ADRB2蛋白表达水平降低(P<0.05),JNK、c-Jun和p-c-Jun蛋白表达水平升高(P<0.01);与sh-NC+Era组比较,sh-ADRB2+Era组ADRB2蛋白表达水平降低,JNK、c-Jun和p-c-Jun蛋白表达水平升高(P<0.05)。结论ADRB2可能通过JNK/c-Jun通路调控前列腺癌细胞中Era诱导的铁死亡和自噬。

The 5-HT2c receptor gene Cys23Ser polymorphism influences the intravaginal ejaculation latency time in Dutch Caucasian men with lifelong premature ejaculation

It has been postulated that the persistent short intravaginal ejaculation latency time （IELT） of men with lifelong premature ejaculation （LPE） is related to 5-hydroxytryptamine （HT）2c receptor functioning. The aim of this study was to investigate the relationship of Cys23Ser 5-HT2c receptor gene polymorphism and the duration of IELT in men with LPE. Therefore, a prospective study was conducted in 64 Dutch Caucasian men with LPE. Baseline IELT during coitus was assessed by stopwatch over a 1-month period. All men were genotyped for Cys23Ser 5-HT2c receptor gene polymorphism. Allele frequencies and genotypes of Cys and Ser variants of 5-HT2c receptor gene polymorphism were determined. Association between Cys/Cys and Ser/Ser genotypes and the natural logarithm of the IELT in men with LPE were.investigated. As a result, the geometric mean, median and natural mean IELT were 25.2, 27.0, 33.9s, respectively. Of all men, 20.0%, 10.8%, 23.1% and 41.5% ejaculated within 10, 10-20, 20-30 and 30-60s after vaginal penetration. Of the 64 men, the Cys/Cys and Ser/Ser genotype frequency for the Cys23Ser polymorphism of the 5-HT2c receptor gene was 81% and 19%, respectively. The geometric mean IELT of the wildtypes （Cys/Cys） is significantly lower （22.6s; 95% CI 18.3-27.8s） than in male homozygous mutants （Ser/Ser） （40.4s; 95% CI 20.3-80.4s） （P = 0.03）. It is concluded that Cys23Ser 5-HT2c receptor gene polymorphism is associated with the IELT in men with LPE. Men with Cys/Cys genotype have shorter IELTs than men with Ser/Ser genotypes.

Effect of Dai-Saiko-To (Da-Chai-Hu-Tang) on LDL-Receptor Gene Expression in Human Hepatoma Cell Line (HepG2)

We previously reported that Dai-saiko-to (Da-Chai-Hu-Tang), a traditional Japanese kampo medicine, increased LDL receptor mRNA expression in the liver of the hypercholesterolemic rabbits. In this study, we focused on LDL receptor gene expression in a human hepatoma cell line (HepG2) treated with Dai-saiko-to extract and the extracts of eight herbs presented in Dai-saiko-to. Dai-saiko-to extract significantly increased LDL receptor gene and SREBP2 gene expression compared with the control. The extracts of four herbs, Bupleurum root, Pinellia tuber, Scutellaria root and Peony root significantly increased the LDL receptor gene expression. Whereas, Jujube, Immature orange, Ginger and Rhubarb extracts did not change the gene expression. These results suggest that Dai-saiko-to increased the expression of the cholesterol transport gene (LDL receptor) regulated by SREBP2 gene in the human hepatoma cell line. The pharmacological activity of Dai-saiko-to against hypercholesterolemia and atheromatous lesions related for these four herbal components.

Effects ofβ_2-Adrenergic Antagonist on Cytosolic Ca^(2+) in Ventricular Myocytes from Infarcted Rat Heart

Objectives To investigate the effects of β2-adrenergic antagonist on cytosolic Ca^2 ＋ （[Ca^2＋ ]i） in ventricular myocytes from infarcted rat heart. Methods A ligature was placed around left anterior descending coronary artery of rat hearts. Rats in the control group were sham-operated. Cardiomyocytes were dissociated at two, four, eight weeks after myocardial infarction （MI） and [Ca^2＋]i was measured via fura-2 fluorescence. The response of cardiomyocytes to isoproterenol in presence or absence of betal-adrenergic antagonist atenolol, beta2-adrenergic antagonist ICI118, 551 or non-selective β1, 2- adrenergic antagonists propranolol was examined. Results The followings were found that ICI 118, 551 had no significant effects on the rise of [Ca^2＋]i induced by isoproterenol in normal ventricular myocytes （P 〉 0.05）, ICI118, 551 only significantly attenuated the rise of [Ca^2＋]i induced by isoproterenol at four weeks and eight weeks after MI （24.5%±5.7% vs 57.8% ± 13.2%, P〈 0.01; 12.2%±7.9% vs 44.6%±11.3%, P〈 0.01）. Atenolol had suppressive effects only in the control group and the post-MI group of two weeks （P 〈 0.05）, and propranolol had suppressive effects in the control and all the three post-MI groups （P 〈 0.01）. Conclusions Beta2-adrenergic antagonist ICI118, 551 may exert negative effects on Ca^2＋ overload initiated by sympathetic stimulation after MI.

α_(1A)-肾上腺素能受体与增强型绿色荧光蛋白标记的活化T细胞核因子2稳定共表达细胞的构建

目的建立α_(1A)-肾上腺素能受体(α_(1A)-AR)与增强型绿色荧光蛋白(EGFP)标记的活化T细胞核因子2(NFAT2)稳定共表达细胞。方法①将pcDNA3.1-α_(1A)-AR-3×FLAG重组质粒转染至U2OS-EGFPNFAT2细胞,经潮霉素B(Hygro-B)200 mg·L^(-1)压力筛选后加入α_(1A)-AR激动剂去甲肾上腺素(NE,10μmol·L^(-1))孵育30 min,通过高内涵筛选系统检测细胞核内绿色荧光强度,验证EGFP-NFAT2核转位,筛选得到稳定表达α_(1A)-AR的U2OS-EGFP-NFAT2-α_(1A)-AR细胞。②采用实时荧光定量PCR(RT-qPCR)和Western印迹法检测该细胞和对照细胞U2OS-EGFP-NFAT2中α_(1A)-AR mRNA和蛋白的表达水平。③将U2OS-EGFP-NFAT2-α_(1A)-AR细胞接种于96孔板,分别加入NE(10^(-8)~10^(-5) mol·L^(-1))或α2-AR激动剂右美托咪定(DMED,10^^(-8.8)~10^(-5) mol·L^(-1))孵育30 min,通过高内涵筛选系统检测EGFP-NFAT2核转位。④将U2OS-EGFP-NFAT2-α_(1A)-AR细胞分为溶剂对照组、α1-AR拮抗剂萘派地尔(1μmol·L^(-1))组、NE(1μmol·L^(-1))组、萘派地尔+NE(各1μmol·L^(-1)共孵育)组、α2-AR拮抗剂阿替美唑(0.1μmol·L^(-1))组、DMED(0.1μmol·L^(-1))组、阿替美唑+DMED(各0.1μmol·L^(-1)共孵育)组和萘派地尔+DMED(萘派地尔1μmol·L^(-1)与DMED 0.1μmol·L^(-1)共孵育)组,药物孵育时间均为30 min,通过高内涵筛选系统检测EGFP-NFAT2核转位,验证该细胞α_(1A)-AR功能的特异性。结果①Hygro-B压力筛选得到58株U2OS-EGFP-NFAT2-α_(1A)-AR细胞,NE 10μmol·L^(-1)孵育后,其中50号细胞核内绿色荧光强度最强,故选定其为稳定共表达α_(1A)-AR和EGFPNFAT2的U2OS-EGFP-NFAT2-α_(1A)-AR细胞。②Western印迹法结果显示,U2OS-EGFP-NFAT2-α_(1A)-AR细胞可明显表达α_(1A)-AR蛋白,而对照细胞U2OS-EGFP-NFAT2中未见α_(1A)-AR蛋白表达。RT-qPCR结果显示,该细胞在传代5~20代内α_(1A)-AR mRNA均稳定表达,其表达水平为对照细胞的500~800倍。③NE或DMED使U2OS-EGFP-NFAT2-α_(1A)-AR细胞中EGFP-NFAT2核转位明显增加,半数有效浓度(EC50)分别为5.94×10^(-7)和6.15×10^(-8) mol·L^(-1)。④与溶剂对照组和萘派地尔组比较,NE组U2OS-EGFP-NFAT2-α_(1A)-AR细胞EGFP-NFAT2核转位明显增强(P<0.01),而萘派地尔+NE组EGFP-NFAT2核转位较NE组明显减弱(P<0.01)。与溶剂对照组和阿替美唑组比较,DMED组EGFP-NFAT2核转位明显增强(P<0.01),阿替美唑+DMED组EGFP-NFAT2核转位与DMED组比较无明显差别,而萘派地尔+DMED组EGFP-NFAT2核转位较DMED组明显减弱(P<0.01)。结论成功构建稳定共表达α_(1A)-AR和EGFP-NFAT2的U2OS-EGFPNFAT2-α_(1A)-AR细胞,可用于靶向α_(1A)-AR化合物筛选和受体分子机制研究。

The human δ2 glutamate receptor gene is not mutated in patients with spinocerebellar ataxia

The human glutamate receptor delta 2 gene （GRID2） shares 90%homology with the orthologous mouse gene. The mouse Grid2 gene is involved with functions of the cerebellum and sponta-neous mutation of Grid2 leads to a spinocerebellar ataxia-like phenotype. To investigate whether such mutations occur in humans, we screened for mutations in the coding sequence of GRID2 in 24 patients with familial or sporadic spinocerebellar ataxia and in 52 normal controls. We de-tected no point mutations or insertion/deletion mutations in the 16 exons of GRID2. However, a polymorphic 4 nucleotide deletion （IVS5-121_-118 GAGT） and two single nucleotide polymor-phisms （c.1251G〉T and IVS14-63C〉G） were identiifed. The frequency of these polymorphisms was similar between spinocerebellar ataxia patients and normal controls. These data indicate that spontaneous mutations do not occur in GRID2 and that the incidence of spinocerebellar ataxia in humans is not associated with GRID2 mutation or polymorphisms.

Congenital nephrogenic diabetes insipidus arginine vasopressin receptor 2 gene mutation at new site:A case report

BACKGROUND Congenital nephrogenic diabetes insipidus(CNDI)is a rare hereditary disorder.It is associated with mutations in the arginine vasopressin receptor 2(AVPR2)gene and aquaporin 2(AQP2)gene,and approximately 270 different mutation sites have been reported for AVPR2.Therefore,new mutations and new manifestations are crucial to complement the clinical deficiencies in the diagnosis of this disease.We report a case of a novel AVPR2 gene mutation locus and a new clinical manifestation.CASE SUMMARY We describe the case of a 48-d-old boy who presented with recurrent fever and diarrhea 5 d after birth.Laboratory tests showed electrolyte disturbances and low urine specific gravity,and imaging tests showed no abnormalities.Genetic testing revealed a novel X-linked recessive missense mutation,c.283(exon 2)C>T(p.P95S).This mutation results in the substitution of a proline residue with a serine residue in the AVPR2 protein sequence.The diagnosis of CNDI was confirmed based on the AVPR2 gene mutation.The treatment strategy for this patient was divided into two stages,including physical cooling supplemented with appropriate amounts of water in the early stage and oral hydrochlorothiazide(1-2 mg/kg)after a clear diagnosis.After follow-up of one and a half years,the patient gradually improved.CONCLUSION AVPR2 gene mutations in new loci and new clinical symptoms help clinicians understand this disease and shorten the diagnosis cycle.

肾上腺素β受体基因、G蛋白基因和CYP2D6基因多态性对美托洛尔导致脂代谢异常的影响

目的:研究肾上腺素β受体基因、G蛋白基因和CYP2D6基因多态性对美托洛尔导致脂代谢异常的影响。方法:选取2019年10月至2022年10月该院收治的初治高血压患者120例,检测β_(1)受体基因Ser49Gly、Gly389Arg多态;β_(2)受体基因Arg16Gly、Gln27Glu多态;β3受体基因Trp64Arg多态;代谢酶基因CYP2D6*10的P34S多态;G蛋白基因GNB3 C825T多态。比较美托洛尔单药治疗3个月后不同表型总胆固醇(TC)、三酰甘油(TG)、低密度脂蛋白(LDL)和高密度脂蛋白(HDL)水平变化。结果:各基因位点不同基因型患者的年龄、性别、体重指数、基线TC和基线TG等指标比较,差异均无统计学意义(P>0.05)。治疗3个月后,β_(2)肾上腺素受体基因Arg16Gly患者的TG水平升高,其中Gly/Gly型患者升高幅度最高,Arg/Gly型次之,二者均高于Arg/Arg型,差异有统计学意义(P<0.05)。结论:β_(2)肾上腺素受体Arg16Gly多态与美托洛尔引起的血浆TG水平升高相关,携带2个G等位基因会导致服药后血脂水平明显升高。