大鼠死后不同时间肝GAPDH mRNA的检测

目的 探讨大鼠死后不同时问肝甘油醛3-磷酸脱氢酶（GAPDH）mRNA的降解情况。方法 24只大鼠断颈处死后置于20℃温度控制系统内,利用荧光标记逆转录聚合酶链反应（RT—PCR）技术检测大鼠肝GAPDH mRNA在死后即刻至72h的相对表达量的变化情况。结果 在死后即刻至48h大鼠肝组织内均可检测到GAPDH mRNA,且其扩增产物呈下降趋势。结论 荧光RT—PCR技术检测大鼠肝组织GAPDH mRNA,在死后的降解呈现出一定的规律．可为死亡时间（PMI）推断提供新的指标。     

甲状腺功能减退大鼠脑海马组织T3核受体基因表达

目的了解甲状腺功能减退(简称甲减)大鼠海马组织T3核受体(T3NR)α、β亚型mRNA的特异性变化,探讨甲状腺激素对脑发育及功能调控的分子机制。方法采用丙基硫氧嘧啶(PTU)腹腔注射诱发甲减动物模型,运用RT-PCR检测技术测定实验性甲减大鼠脑海马组织T3NRα1mRNA、T3NRβ1mRNA的表达水平。结果甲减大鼠海马组织T3NRα1mRNA、T3NRβ1mRNA的表达水平明显下调,与正常对照组比较差异有统计学意义(P<0.01)。结论甲减时脑组织T3NRα、β亚型mRNA的表达水平下调,T3NR的合成减少,导致甲状腺激素的生物效应的降低,可能是甲减性脑损害发生的重要病理机制之一。     

Sex differences in androgen-regulated cytochrome P450 aromatase mRNA in the rat brain

The conversion of testosterone to estradiol by cytochrome P450 aromatase (P450_AROM) in the medial preoptic area is required for full expression of male sexual behavior in rats. Preoptic P450_AROM activity is stimulated by androgens through an androgen-receptor mediated mechanism that regulates P450_AROM gene expression. The mechanism of enzyme induction appears to be sexually dimorphic in several species leading to greater testosterone-stimulated P450_AROM activity in males than in females. The present study was designed to determine whether the sex difference in androgen-regulated P450_AROM activity is manifested at the levels of mRNA expression. We compared the concentrations of P450_AROM mRNA and enzyme activity between five different treatment groups: intact males, castrated males (CX), ovariectomized females (OVX), CX males treated with dihydrotestosterone (CX+DHT), and OVX females treated with DHT (OVX+DHT). We found that unstimulated levels of P450_AROM mRNA and enzyme activity in both the preoptic area and medial basal hypothalamus were similar in the CX and OVX groups. However, when treated with equivalent doses of DHT, the levels of P450_AROM mRNA and enzyme activity in both brain regions were significantly higher in males than in females (i.e., CX+DHT group >OVX+DHT group). These results demonstrate that sex differences in the regulation of P450_AROM in brain are exerted pretranslationally by androgen and suggest that gender differences in androgen responsiveness play an important role in regulating gene expression in the adult rat brain.     

局灶性缺血再灌注大鼠脑血管紧张素Ⅱ1型受体基因的表达

目的　研究大鼠局灶性脑缺血再灌注后血管紧张素Ⅱ 1型受体 (AT1 R)基因的表达及氯沙坦对其的影响。方法　 32只Wistar大鼠随机分为氯沙坦用药组、缺血再灌注组、假手术组和正常对照组 ,氯沙坦用药组给予氯沙坦(10mg·kg- 1 ·d- 1 )灌胃 ,2周后制作大鼠大脑中动脉栓塞再通模型 ,运用逆转录 聚合酶链反应法检测大脑中动脉供血区皮质AT1 RmRNA的表达 ;用放射免疫法测定血管紧张素Ⅱ (AngⅡ )的水平。结果　缺血再灌注组缺血区皮质AT1 RmRNA的表达和AngⅡ的水平均高于正常对照组 ,氯沙坦用药组缺血区皮质AT1 RmRNA的表达较缺血再灌注组降低。结论　局灶性脑缺血再灌注诱导AngⅡ水平和AT1 R基因的表达增加 ,氯沙坦可降低AT1 R基因的表达。     

Expression of gamma-aminobutyric acid A receptor subunits α1，β1，γ2 mRNA in rats with hepatic encephalopathy

AIM: To investigate the mRNA expression of gammaaminobutyric acid A (GABAA) receptor subunitsα1,β1,γ2 in different parts of the brain of rats with hepatic encephalopathy.METHODS: Twelve adult male Sprague-Dawley rats were randomly divided into two groups: (1) hepatic encephalopabhy model group (n = 6), which was induced by intraperitoneal injection of thioacetamide (TAA, 350 mg/kg) for three consecutive days; (2) control group (n = 6), in which the rats were treated with same dose of normal saline solution. After the freeze slice of cerebrum was made, in situ hybridization was used to detect the mRNA of GABAA receptor subunits α1,β1,, and γ2 in rat cerebral cortex,basal nuclei, substantia nigra and hippocampi. Image data were collected and analyzed quantitatively by QWin550CW model image signal gather and analysis system.RESULTS: In rats with hepatic encephalopathy, mRNA expression levels of GABAA receptor subunits α1,β1 increased significantly in basal nuclei, substantia nigra pars compacta, substantia nigra pars reticularis and hippocampi (144.7&#177;15.67/184.14&#177;4.41, 60.61&#177;33.66/113.07&#177;32.44,87.71&#177; 21.25/128.40&#177;18.85, 122.34&#177;5.56/161.60&#177;4.56,123.29&#177;5.21/140.65&#177;4.15, 123.40&#177;4.42/140.09&#177;4.52,124.76&#177;4.18/140.09&#177;4.12, 141.62&#177;15.09/182.80&#177;5.20,69.13&#177;30.74/134.21&#177;43.76, 87.87&#177;25.16/151.01&#177;19.49,122.14&#177;6.30/162.33&#177;3.92, 122.81&#177;5.09/137.19&#177;7.12,123.00&#177;4.63/138.11&#177;5.92, 125.75 &#177;2.43/138.81&#177;6.10,P&lt;0.01), but did not change in the cerebral cortex compared to the control group. Similar changes were found in the mRNA expression levels of GABAA receptor subunit γ2,which increased significantly in basal nuclei, substantia nigra pars compacta, substantia nigra pars reticularis (136.81&#177;26.41/167.97&#177;16.23, 51.00&#177;36.14/113.18&#177;36.52,86.35&#177;20.30/126.90&#177;19.74, P&lt;0.01), CA1 of hippocampal(162.15:1:9.05/178.62:1:6.45, P&lt;0.05), and no changes were found in the cerebral cortex and CA2, CA3, CA4 of hippocampi.CONCLUSION: In rats with hepatic encephalopathy,mRNA expression levels of GABAA receptor subunits α1,β1,γ2 increase significantly in basal nuclei, substantia nigra and hippocampi, suggesting that the changes of mRNA expression levels in GABAA receptor subunits may contribute to the pathogenesis of hepatic encephalopathy.     

垂体肿瘤转化基因在大肠癌组织中的表达及临床意义

目的探讨垂体肿瘤转化基因（PTTG）在大肠癌发生、发展中的作用。方法采用RT-PCR及免疫组化法检测80份大肠癌组织、20份正常大肠组织中PTTG mRNA、PTTG蛋白表达,分析其与大肠癌临床病理参数的关系。结果大肠癌组织中PTTG mRNA及蛋白表达量明显高于正常大肠组织,P均〈0.01;PTTG mRNA及PTTG蛋白表达量与大肠癌浸润深度、淋巴结转移、Duke＇s分期有明显相关性。结论 PTTG高表达在大肠癌的发生发展过程中起促进作用;PTTG表达可作为大肠癌转移及预后的预测指标。     

RT-PCR/Southern杂交方法检测肝癌患者外周血AFP mRNA

目的寻找一种敏感的方法以检外外周血中的肝癌细胞.方法分离原发性肝癌患者以及肝硬化、急慢性肝炎、肝转移癌、肝脏良性肿瘤患者和健康志愿者的外周血单个核细胞,提取单个核细胞的总RAN,进行RT-PCR/Southern杂交检测AFPmRNA.结果原发性肝癌患者外周血AFP mRAN的检出率为41.4%,其检出率与肿瘤大小以及TNM分期有关,对照组均为阴性.结论肝癌患者外周血中的AFPmRNA可以作为肝癌细胞的血液微小转移的标志物.     

Expression of hypothalamic GnRH-I mRNA in the female turkey at different reproductive states and following photostimulation

In birds, changes in hypothalamic gonadotropin-releasing hormone-I (GnRH-I) content and release are correlated with reproductive stages. This study examined the distribution and expression level of GnRH-I mRNA in anatomically discrete hypothalamic nuclei throughout the turkey reproductive cycle and following photostimulation. GnRH-I mRNA expression was determined using in situ hybridization in non-photostimulated (NPS), egg-laying (LAY), incubating (INC) and photorefractory (REF) hens. Overall, GnRH-I mRNA expression was greatest in the nucleus commissurae pallii (nCPa) and around the organum vasculosum lamina terminalis (OVLT), with less expression observed in the nucleus septalis lateralis (SL), cortico-habenula cortico-septum area, and within the nucleus preopticus medialis. GnRH-I mRNA expression was significantly increased in nCPa, OVLT, and SL after NPS hens (6L:18D) were exposed to a 30 or 90 min pulse of light beginning 14 h after first light (dawn). GnRH-I mRNA abundance within nCPa, OVLT and SL was greater in LAY than in NPS and INC hens, while mRNA expression was least in REF hens. These results indicate that GnRH-I mRNA expression in birds is sensitive to light stimulation during the photosensitive period and can be used to more precisely characterize their different reproductive stages.     

实时荧光定量RT-PCR检测结直肠癌中ERCC1 mRNA的表达及其临床意义

目的传统的影像学检查及肿瘤标志物检测对结直肠癌微转移的诊断价值有限。本研究旨在探讨ERCC1 mRNA在结直肠癌患者外周血中的表达情况及其临床意义。方法用实时定量荧光逆转录聚合酶链反应(RT-PCR)方法,对53例结直肠癌患者和21名健康人的外周血标本进行ERCC1 mRNA检测,分析其表达与肿瘤临床病理特征的关系并应用接受者操作特征曲线(ROC曲线)分析ERCC1 mRNA检测对结直肠癌的诊断价值。结果结直肠癌组外周血ERCC1 mRNA表达水平显著高于正常对照组(P<0.05),其表达与肿瘤淋巴结转移、血行转移和TNM分期相关(P<0.05)。ERCC1 mRNA诊断结直肠癌的ROC曲线下面积0.89,诊断界值为Ct≤32。结论实时定量荧光RT-PCR检测ERCC1 mRNA可以更好地评估结直肠癌血道微转移,有利于判断预后。     

An analysis of T cell antigen receptor variable beta genes during the clinical course of patients with chronic hepatitis B

BACKGROUND: The degree of hepatocyte injury in patients with chronic hepatitis B appears consistent with the number of T cells that respond to hepatitis B virus-related antigens. METHODS: By using a polymerase chain reaction (PCR)-based approach, we monitored a ratio of the T cell antigen receptor (TcR) variable (V) beta gene families against a total TcR V beta gene expression in the peripheral T cells obtained from five patients and four healthy controls. RESULTS: In the healthy controls, there was no significant change in the ratios at an interval of four or eight weeks. In contrast, several TcR V beta families showed the significant changes in the ratios of their gene expression during the follow-up period in all patients. No common highly fluctuated TcR V beta, however, was found among the patients. Furthermore, there was no correlation between their changes and serum levels of alanine aminotransferase. CONCLUSIONS: These findings suggest that the skewing of the TcR family with multiclone is the result of T-cell responses to viral antigens in peripheral blood.     

外周血中白蛋白mRNA作为肝癌微小转移标志物的进一步研究

目的　检测不同肝脏疾病患者的外周血中白蛋mRNA(ALBmRNA) ,探讨外周血中ALBmRNA作为肝癌微小转移标志物的特异性。方法　分离原发性肝癌患者以及肝硬化、急慢性肝炎、肝转移癌、肝脏良性肿瘤患者和健康志愿者的外周血单个核细胞 ,提取单个核细胞的总RNA ,进行RT PCR ,将RT PCR产物再进行Southern杂交。结果　原发性肝癌患者组外周血中ALBmRNA的检出率为 44 .8% ,对照组检出率为 2 9.2 % ,两者之间的差异无统计学意义 (P≥ 0 .0 5 )。结论　作为肝癌微小转移的检测标志物ALBmRNA的特异性不高