人端粒酶反转录酶基因诱导人永生化成骨细胞的生物学特性

目的:研究反转录病毒(hTERT)基因诱导的人永生化成骨细胞系Clone5的生物学特性,包括增殖能力、细胞周期和是否恶性变。方法:收集30～31代Clone5和6～7代人成骨细胞(hOB),绘制两种细胞的生长曲线并用流式细胞仪检测细胞周期特征,软琼脂集落实验和裸鼠致瘤实验检测Clone5是否恶性变。结果:由生长曲线计算出细胞倍增时间:hOB:190h;Clone5:62h;曲线显示Clone5的增殖能力明显增强;流式细胞仪检测发现,Clone5细胞周期中G1期细胞占53.46%,S期细胞占28.24%,凋亡率为0.76%,hOBG1期细胞占79.81%,S期细胞占19.76%,凋亡率为33.33%。和hOB相比,Clone5G1期细胞比例减少,S期比例增加,细胞的凋亡率显著降低;软琼脂集落实验未见细胞团形成,裸鼠接种不致瘤。结论:转导hTERT基因的Clone5增殖能力明显增强,凋亡率降低,且未发生恶性变,可用做组织工程的种子细胞和研究正常骨生理、生化的工具细胞。

家蚕微孢子虫反转录酶基因部分序列的克隆与分析

从家蚕微孢子虫 (Nosemabombycis)中通过RT PCR扩增出反转录酶 (reversetranscriptase)基因的部分序列(大小为 1 2kb) ,经 3′ RACE(RapidAmplificationofcDNAEnds)获得其 3′端序列 ,但经两次 5′ RACE后仍未发现起始密码子 (ATG)。目前得到的cDNA序列为 330 3bp ,推测其编码 110 1个氨基酸的多肽 ,经同源性比较分析 ,发现该序列与许多生物体的反转录酶基因有一定的同源性。

马传贫病毒白细胞弱毒疫苗毒株反转录酶基因克隆及序列分析比较

本实验从商品化马传贫弱毒疫苗培养物提纯马传贫疫苗毒病毒粒子并抽提病毒RNA后 ,采用RT_PCR方法扩增并首次克隆了马传贫驴强毒反转录酶 (RTase)基因。经核苷酸序列测定得出反转录酶基因全长 1 6 6 8bp ,编码 5 5 6个氨基酸。通过与已发表其它马传贫病毒反转录酶基因序列比较 ,发现在氨基酸和核苷酸水平差异率分别为 1 4 .0 %和 1 6 .6 %。变异氨基酸随机分布于整个基因上 ,无明显规律。在反转录病毒高度保守的酶基因上发现这样大的差异 ,展示出马传贫驴强毒作为马传贫弱毒疫苗亲本毒株的独特分子特性。

人端粒酶反转录酶基因修饰骨髓间充质干细胞调控肝细胞增殖和凋亡

背景:研究表明人端粒酶反转录酶基因的导入可以使骨髓间充质干细胞的生命周期得到显著延长,使其能够继续保持多向分化潜能。目的:探讨人端粒酶反转录酶基因修饰骨髓间充质干细胞对肝细胞增殖和凋亡的影响。方法:采取直接贴壁法,分离、培养大鼠骨髓间充质干细胞,利用脂质体转染法,将编码hT ERT基因的真核表达质粒pC Ineo-hT ERT导入骨髓间充质干细胞。将hT ERT基因转染骨髓间充质干细胞与肝细胞按1:1共培养(观察共培养组),同时设未转染骨髓间充质干细胞与肝细胞按1:1共培养组(对照共培养组)和肝细胞单独培养组,采用MTT比色法和免疫荧光染色法观察骨髓间充质干细胞对肝细胞增殖和凋亡的影响。结果与结论:观察共培养组的肝细胞增殖率明显高于对照共培养组和肝细胞单独培养组,差异有显著性意义(P<0.05);观察共培养组的肝细胞存活率明显高于肝细胞单独培养组,差异有显著性意义(P<0.05)。结果表明人端粒酶反转录酶基因修饰的骨髓间充质干细胞可以抑制肝细胞的凋亡,并促进其增殖,具有改善肝细胞功能的作用。

荧光聚合酶链反应–毛细管电泳法与Sanger测序法检测胶质瘤中端粒酶反转录酶基因启动子突变状态对比分析

目的对比分析荧光聚合酶链反应-毛细管电泳(polymerase chain reaction-capillary electrophoresis,PCR-CE)法及Sanger测序法检测胶质瘤中端粒酶反转录酶(telomerase reverse transcriptase,TERT)基因启动子突变的敏感度、特异度及一致性,为临床检测提供方法学依据。方法收集2017年11月至2019年11月首都医科大学宣武医院265例胶质瘤标本及临床病理资料,分别采用Sanger测序法与荧光PCR-CE法检测TERT基因启动子突变。结果TERT基因启动子突变与年龄、组织学分型和WHO分级均显著相关(P<0.05)。Sanger测序法与荧光PCR-CE法的TERT基因启动子突变检出率分别为52.8%(140/265)及51.3%(136/265),敏感度和特异度分别为96.4%和99.2%,符合率为97.7%,具有较好一致性(Kappa=0.955)。Sanger测序法检出TERT C228T突变103例(38.9%),荧光PCR-CE法检出99例(37.4%),敏感度和特异度分别为95.2%和99.4%,符合率为97.7%,具有较好一致性(Kappa=0.952)。Sanger测序法和荧光PCR-CE法均检测出TERT C250T突变37例(14.0%),符合率为100.0%,具有较高一致性(Kappa=1.000)。结论荧光PCR-CE法检测TERT基因启动子突变率与Sanger测序法相当,敏感度、特异度及一致性均较高,且操作相对简便快速。

端粒酶反转录酶基因电转染人羊膜间充质干细胞移植治疗糖尿病

背景:羊膜是胎儿出生后的废弃物,羊膜间充质干细胞具有取材方便、增殖能力强、无伦理学争议、免疫原性低等优势。目的:通过端粒酶反转录酶(human telomerase reverse transcriptase,h TERT)基因电转染人羊膜间充质干细胞移植到大鼠糖尿病模型,探讨其对糖尿病大鼠的疗效。方法:分离培养人羊膜间充质干细胞,经h TERT基因电转染羊膜间充质干细胞,从50只SD大鼠中随机取10只作为对照组,剩余40只按45 mg/kg的剂量注射链脲霉素,建立糖尿病模型后,将建模成功的36只SD大鼠随机分为糖尿病组、人羊膜间充质干细胞移植组和h TERT-人羊膜间充质干细胞移植组,每组各12只;人羊膜间充质干细胞移植组和h TERT-人羊膜间充质干细胞移植组分别通过大鼠舌下静脉注射移植人羊膜间充质干细胞和经h TERT基因电转染羊膜间充质干细胞。移植后各组进行动态血糖水平监测,于移植后每周检测各组大鼠血浆胰岛素的浓度,胰腺切片苏木精-伊红染色观察病理变化。结果与结论:移植后4周,与糖尿病组比较,人羊膜间充质干细胞移植组和h TERT-人羊膜间充质干细胞移植组血糖水平明显下降(P<0.05),尤其是h TERT-人羊膜间充质干细胞移植组的空腹血糖水平接近于对照组水平(P>0.05),而糖尿病组空腹血糖维持在较高水平;移植后6周,与糖尿病组比较,人羊膜间充质干细胞移植组和h TERT-人羊膜间充质干细胞移植组的血浆胰岛素含量增加(P<0.05),胰腺病损程度减轻(P<0.05),h TERT-人羊膜间充质干细胞移植组更明显(P<0.05)。结果证实,提示h TERT转染的羊膜间充质干细胞移植能明显降低糖尿病大鼠血糖和减轻胰岛损伤,可以有效治疗大鼠糖尿病。

宫颈病变组织中端粒酶反转录酶基因表达的临床意义

目的探讨宫颈病变组织中端粒酶反转录酶(TERT)基因的表达和临床意义。方法对获取的208例宫颈脱落细胞标本进行细胞学诊断、组织学诊断,同时采用RT-PCR方法检测TERT基因的扩增情况。结果不明确意义鳞状上皮细胞(ASCUS)组、低度病变(LSIL)组、高度病变(HSIL)组、鳞状细胞癌(SCC)组与正常细胞学组比较,组间差异有统计学意义(χ2=16.441,P<0.0025);CIN1、2、3,SCC组与良性病变组TERT基因阳性率比较差异有统计学意义(P<0.001);TERT基因的表达率与细胞学诊断的病变程度呈正相关(R=0.342,P=0.0442<0.05),TERT基因的表达率与组织学诊断的病变程度呈正相关(R=0.527,P=0.0411<0.05)。结论TERT基因在宫颈癌和CIN病变中的表达有异常增加,可作为监测宫颈癌前病变病情进展的一个生物遗传学指标。

人端粒酶反转录酶重组绿色荧光表达载体转染椎间盘正常髓核细胞

背景:椎间盘髓核细胞分离培养困难,老化较快,迫切需要一种标准细胞株用于实验研究。目的:探讨人端粒酶反转录酶重组绿色荧光表达载体的构建及其转染正常髓核细胞构建永生化细胞的可行性研究。方法:通过目的基因克隆、真核表达质粒中目的基因序列测定、目的基因真核表达质粒的构建、转染人端粒酶反转录酶表达检测等步骤进行实验。结果与结论:构建出人端粒酶反转录酶重组绿色荧光表达载体,成功转染正常髓核细胞并在细胞中稳定表达。结果表明运用人端粒酶反转录酶转染椎间盘髓核细胞构建永生化细胞是一种可行的方法。

联合表达人端粒酶反转录酶hTERT及VEGF与肿瘤复发的关系

目的应用肿瘤发生、发展不同阶段的联合瘤标对膀胱移行细胞癌(BTCC)检测,比较其肿瘤复发诊断的特异性。方法对67例BTCC和作为对照的10例正常膀胱上皮石蜡标本分别用免疫组化法和原位杂交法,检测其血管内皮生长因子(VEGF)和人端粒酶反转录酶(hTERT)的表达。结果两者在正常膀胱上皮中不表达,在BTCC中均不同程度表达,67例BTCC组织中,hTERT强阳性表达28例,其中有肿瘤复发的18例(64.3%),VEGF表达为强阳性共51例,其中有肿瘤复发的30例(58.8%),联合瘤标均强阳性例数为21,有肿瘤复发的18例(85.7%),其特异性与单一瘤标均有显著差异(P<0.05)。结论把针对肿瘤生物学行为不同阶段的瘤标联合,能有效提高诊断的准确度,有希望作为预测BTCC预后复发的有效手段。

转导hTERT基因致人牙髓干细胞永生化的研究

目的 :建立永生化人牙髓干细胞系(human dental pulp stem cell,h DPSC),供口腔医学临床和基础研究使用。方法:原代培养人牙髓细胞并筛选牙髓干细胞,以慢病毒为载体,导入端粒酶反转录酶基因(h TERT)全长c DNA,经筛选得到阳性克隆。体外连续传代,应用反转录PCR(reverse transcription PCR,RT-PCR)和蛋白质印迹技术(Western blot)检测h TERT的整合和表达情况。结果 :成功地将h TERT基因转入人牙髓干细胞,得到的转化细胞端粒酶表达阳性,增殖旺盛,至今已传至第35代。结论:转导外源性h TERT基因可以导致人牙髓干细胞永生化。

膀胱癌中端粒酶RNA的原位表达与肿瘤预后

观察人端粒酶mRNA(hTR)在各级膀胱移行细胞癌 (TCC)中的表达 ,分析其与肿瘤分级及预后的关系。作者回顾性分析了 6 7例膀胱移行细胞癌及对照组的正常膀胱上皮、膀胱良性病变各 10例 ,对石蜡标本用地高辛标记的端粒酶反转录酶基因 (hTRET)寡核苷酸作为RNA探针 ,原位检测端粒酶mRNA在膀胱移行细胞癌细胞中的表达。结果显示 ,标本的端粒酶mRNA表达强度与膀胱TCC分级、预后显著相关 (P <0 0 1)。

人端粒酶反转录酶催化酶亚单位基因对肝母细胞瘤细胞株体外生物学特性的影响

目的 研究人端粒酶反转录酶催化酶亚单位 (hTERT)基因对肝母细胞瘤细胞株体外生物学特性的影响。方法 体外将已构建好的正、反义hTERT真核表达载体转染HepG2 肝母细胞瘤细胞株 ,经G4 18及聚合酶链反应 (PCR)筛选鉴定获得分别转入了正、反义hTERT载体的抗性克隆细胞HepG2 s和HepG2 as。运用逆转录 PCR(RT PCR)技术及TRAP 银染法对各组细胞内源性hTERTmRNA的表达及端粒酶的活性进行了检测 ;同时还采用MTT法、双层软琼脂克隆形成试验、流式细胞术观察和分析了反义hTERT对肝母细胞瘤细胞体外生长增殖活力的影响及是否能诱导瘤细胞的凋亡。结果 与空白对照组、正义hTERT组相比 ,反义hTERT能显著降低HepG2 细胞内源性hTERTmRNA的表达 (n =10 ,t =7 6 1,P <0 0 1)和下调端粒酶活性。当各组细胞传至第 2 0代后 ,与HepG2 、HepG2 s比较 ,HepG2 as细胞的生长速度和集落形成能力明显地减慢和降低 (n =10 ,t=4 5 4 ,P <0 0 1和n =10 ,t=3 96 ,P <0 0 1) ,同时伴有凋亡率的显著增加 (n =10 ,t=9 2 4 ,P <0 0 1和n =10 ,t=8 37,P <0 0 1)。结论 反义hTERT体外可抑制肝母细胞瘤细胞的生长增殖能力 。

siRNA targeting of Cdx2 inhibits growth of human gastric cancer MGC-803 cells

AIM:To investigate the effects of small interference RNA(siRNA) targeting of Cdx2 on human gastric cancer MGC-803 cells in vitro and in vivo.METHODS:The recombinant pSilencer 4.1-Cdx2 siRNA plasmids were constructed and transfected into gastric cancer MGC-803 cells in vitro.The stable transfectants were selected.The effects of Cdx2 siRNA on growth,proliferation,cell cycle,apoptosis,migration and invasiveness of human gastric cancer MGC-803 cells were evaluated and the expression of phosphatase and tensin homolog(PTEN),caspase-9 and caspase-3 was observed in vitro by reverse transcription polymerase chain reaction(RT-PCR) and Western blotting analysis.We also investigated the effect of Cdx2 siRNA on growth of MGC-803 cells in nude mice in vivo.RESULTS:Cdx2 siRNA led to inhibition of endogenous Cdx2 mRNA and protein expression as determined by RT-PCR and Western blotting analysis.Cdx2 siRNA significantly inhibited cell growth and proliferation,blocked entry into the S-phase of the cell cycle,induced cell apoptosis,and reduced the motility and invasion of MGC-803 cells.Cdx2 siRNA also increased PTEN expression,and activated caspase-9 and caspase-3 in MGC-803 cells in vitro.In addition,siRNA targeting of Cdx2 inhibited the growth of MGC-803 cells and promoted tumor cell apoptosis in vivo in nude mice tumor models.CONCLUSION:Cdx2 was involved in regulating progression of human gastric cancer cells MGC-803.Manipulation of Cdx2 expression may be a potential therapeutic strategy for gastric cancer.

Establishment of an orthotopic transplantation tumor model of hepatocellular carcinoma in mice

AIM:To improve the outcome of orthotopic transplantation in a mouse model,we used an absorbable gelatin sponge(AGS) in nude mice to establish an orthotopic implantation tumor model.METHODS:MHCC-97L hepatocellular carcinoma(HCC)cells stably expressing the luciferase gene were injected into the subcutaneous region of nude mice.One week later,the ectopic tumors were harvested and transplanted into the left liver lobe of nude mice.The AGS was used to establish the nude mouse orthotopic implantation tumor model.The tumor suppressor gene,paired box gene 5(PAX5),which is a tumor suppressor in HCC,was transfected into HCC cells to validate the model.Tumor growth was measured by bioluminescence imaging technology.Semi-quantitative reverse transcription polymerase chain reaction(RT-PCR) and histopathology were used to confirm the tumorigenicity of the implanted tumor from the MHCC-97L cell line.RESULTS:We successfully developed an orthotopic transplantation tumor model in nude mice with the use of an AGS.The success rate of tumor transplantation was improved from 60% in the control group to 100% in the experimental group using AGS.The detection of fluorescent signals showed that tumors grew in all live nude mice.The mice were divided into 3 groups:AGS-,AGS+/PAX5-and AGS+/PAX5 +.Tumor size was significantly smaller in PAX5 transfected nude mice compared to control mice(P < 0.0001).These fluorescent signal results were consistent with observations made during surgery.Pathologic examination further confirmed that the tissues from the ectopic tumor were HCC.Results from RT-PCR proved that the HCC originated from MHCC-97L cells.CONCLUSION:Using an AGS is a convenient and efficient way of establishing an indirect orthotopic liver transplantation tumor model with a high success rate.

Effect of realgar on telomerase activity and hTERT-mRNA expression in NB4 cells

Objective: To evaluate whether realgar could down-regulate human telomerase reverse transcriptase (hTERT) gene expression and telomerase activity in acute promyelocytic leukemia cell line-NB4 cells. Methods: The expression of hTERT-mRNA was analyzed by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). Telomerase activity was determined by polymerase chain reaction enzyme-linked immunoassay (PCR-ELISA). Flow cytometry using PI staining was applied to analyze the cell cycle and apoptosis. Results: Treatment of NB4 cells with 155, 300, 600 μg/L realgar reduced telomerase activity significantly accompanying with decrease of hTERT-mRNA and increasing cell apoptosis. G2/M phase arrest appeared when treated with realgar in 300, 600 μg/L. Conclusion: It is suggested that telomerase activity of NB4 cells can be specifically inhibited by realgar through the down-regulation of hTERT gene expression. G2/M phase arrest and apoptosis by realgar in NB4 cells might be related to the reduction of telomerase activity and hTERT-mRNA expression.

CXCR4/SDF-1 axis is involved in lymph node metastasis of gastric carcinoma

AIM:To investigate the role of CXC chemokine receptor-4 (CXCR4) and stromal cell-derived factor-1 (SDF-1) in lymph node metastasis of gastric carcinoma.METHODS:In 40 cases of gastric cancer,expression of CXCR4 mRNA in cancer and normal mucous membrane and SDF-1 mRNA in lymph nodes around the stomach was detected using quantitative polymerase chain reaction (PCR) (TaqMan) and immunohistochemistric assay.SGC-7901 and MGC80-3 cancer cells were used to investigate the effect of SDF-1 on cell proliferation and migration.RESULTS:Quantitative reverse transcription PCR and immunohistochemistry revealed that the expression level of CXCR4 in gastric cancer was significantly higher than that in normal mucous membrane (1.6244 ± 1.3801 vs 1.0715 ± 0.5243,P < 0.05).The expression level of CXCR4 mRNA in gastric cancer with lymph node metastasis was also significantly higher than that without lymph node metastasis (0.823 ± 0.551 vs 0.392 ± 0.338,P < 0.05).CXCR4 expression was significantly related to poorly differentiated,high tumor stage and lymph node metastasis.Significant differences in the expression level of SDF-1 mRNA were found between lymph nodes in metastatic gastric cancer and normal nodes (0.5432 ± 0.4907 vs 0.2640 ± 0.2601,P < 0.05).The positive expression of SDF-1 mRNA in lymph nodes of metastatic gastric cancer was consistent with the positive expression of CXCR4 mRNA in gastric cancer (r=0.776,P < 0.01).Additionally,human gastric cancer cell lines expressed CXCR4 and showed vigorous proliferation and migratory responses to SDF-1.AMD3100 (a specific CXCR4 antagonist) was also found to effectively reduce the migration of gastric cancer cells.CONCLUSION:The CXCR4/SDF-1 axis is involved in the lymph node metastasis of gastric cancer.CXCR4 is considered as a potential therapeutic target in the treatment of gastric cancer.

Magnetic iron oxide nanoparticles carrying PTEN gene to reverse cisplatin-resistance of A549/CDDP cell lines

To evaluate the feasibility of using magnetic iron oxide nanoparticle as wild PTEN gene carrier for transfection in vitro to reverse cisplatin-resistance of A549/CDDP cells, A549/CDDP cells were transfected with the wild PTEN gene expression plasmid (pGFP-PTEN) by magnetic iron nanoparticle and lipo2000. The transfection efficiency was detected by fluorescence microscope and flow cytometer. The expression levels of PTEN mRNA and protein were detected by reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry analysis. The effect of PTEN transfection on cell cycle enhances the sensitivity of A549/CDDP to cisplatin and nanoparticle-mediated transfection has a higher efficiency than that of the liposome-mediated group. The apoptosis level was up-regulated in PTEN transfection group. The magnetic iron oxide nanoparticle could be used as one of the ideal gene carriers for PTEN gene delivery in vitro. PTEN can be an effective target for reversing cisplatin-resistance in lung cancer.

QUANTIFICATION OF P4HA2 mRNA OF FIBROBLASTS WITH SYBR GREEN BASED RT-PCR FOR CORRECTING CMV INACTIVATION EFFICIENCY IN DONOR BLOOD

Objective To quantify proline 4-hydroxylase, alpha polypeptide ii （ P4HA2 ） mRNA of human embryo lung fibroblast （HELF） with SYBR green based reversed transcript PCR （RT-PCR） for correcting cytomegalovirus （CMV） inactivation or clearance efficiency in donor blood. Methods A pair of specific primers of exon 12a of P4HA2 was designed, and the related PCR-reaction system and condition were optimized. Then the recombinant plasmid containing the target fragment was constructed for making standard curve with SYBR green based real-time RT-PCR. Finally, the sensitivity, reproducibility, and specificity of this method were fully estimated. Results The sensitivity of the method was 1.5E ＋ 04 copies/mL of P4HA2 mRNA, corresponding to 10^3 fibroblasts. In addition, existence of 8. 67E ＋ 06 leukocytes could not interfere with the accurate quantification of HELF in the large dynamic range. The intra-assay variability and inter-assay variability both varied in different concentrations, being higher in low concentrations and lower in high concentrations. But all of them were below 13. 76% in variation, which showed acceptable stability of this method. Conclusion SYBR green and specific primer based real-time RT-PCR show up a good quality for quantifying HELF P4HA2 mRNA with good specificity, stability, and high sensitivity. Approximate 10 copies of P4HA2 mRNA per cell in average can be detected by the method. Therefore, this method can be used to deduct fibroblast-associated CMV for correcting CMV inactivation efficiency in leukocytes.

The molecular mechanisms of Yang Wei Kang Liu powder on anticancer and reducing chemotherapy side-effect in combination with chemotherapy

Objective:We studied the molecular mechanisms of Yang Wei Kang Liu Power(YWKL,traditional Chinese medicine for nourishing stomach and anticancer) on anticancer and reducing chemotherapy side-effect in combination with chemotherapy.Methods:615 pre-cancer mouse model of YWKL for 10 days and CTX 1 time,semi-quantitative reverse transcription-polymerase chain reaction(RT-PCR) to detect bone marrow granulocyte-macrophage colony-stimulating factor(GM-CSF) gene and cancer proto-oncogene Bcl-2,c-myc expression.Results:YWKL in combination with chemotherapy could obviously promoted the expression of GM-CSF gene and inhibited the expression of Bcl-2 and c-myc oncogenes of FC 615 mice.Conclusion:The molecular mechanisms of anticancer and reducing chemotherapy side-effect of YWKL in combination with chemotherapy are to promote the expression of GM-CSF gene and inhibit the expression of Bcl-2 and c-myc oncogenes.

贵州省HIV抗病毒治疗病人群耐药基因突变研究

目的:研究贵州省经抗反转录病毒治疗的艾滋病毒感染者/艾滋病患者(HIV/AIDS)的人免疫缺陷病毒(Human Immunodeficiency virus,HIV)的反转录酶区和蛋白酶区耐药基因变异情况。方法:采集贵州省抗病毒治疗的感染者/艾滋病患者(HIV/AIDS)56例的血样,进行RNA核酸提取、采用套式反转录-聚合酶链反应(Reverse Transcription-Polymerase Chain Reaction,RT-PCR)扩增HIV-1 pol基因基因片段及基因测序,对所得结果登录美国斯坦福大学HIV耐药数据库进行在线分析确定耐药突变情况。结果:分析获得的44例目的基因片段,有24例发生基因突变。亚型分布为:17例AE亚型、15例BC亚型、10例C亚型及2例B亚型。逆转录酶抑制剂(reverse transcriptase inhibitors,RTIs)突变率和蛋白酶抑制剂(protease inhibitors,PIs)突变率分别为45.5%(20/44)和13.6%(6/44),其中核苷酸类逆转录酶抑制剂(nucleotide reverse transcriptase inhibitors,NRTIs)耐药突变发生率为34.1%(15/44),非核苷类逆转录酶抑制剂(non-Nucleoside reverse transcriptase inhibitors,NNRTIs)耐药突变发生率38.6%(17/44)。结论:贵州已经出现了针对NRTIs与NNRTIs高度耐药毒株,有必要对未进行抗病毒治疗的感染者进行耐药监测。