‘Default’ generated neonatal regulatory T cells are hypomethylated at conserved non‐coding sequence 2 and promote long‐term cardiac allograft survival

Regulatory T (Treg) cells play an important role in the maintenance of immune self-tolerance and homeostasis. We previously reported that neonatal CD4⁺ T cells have an intrinsic ‘default’ mechanism to become Treg (neoTreg) cells in response to T-cell receptor (TCR) stimulation. However, the underlying mechanisms are unclear and the effects of neoTreg cells on regulating immune responses remain unknown. Due to their involvement in Foxp3 regulation, we examined the role of DNA methyltransferase 1 (DNMT1) and DNMT3b during the induction of neoTreg cells in the Foxp3^gfp mice. The function of neoTreg cells was assessed in an acute allograft rejection model established in RAG2^−/− mice with allograft cardiac transplantation and transferred with syngeneic CD4⁺ effector T cells. Following ex vivo TCR stimulation, the DNMT activity was increased threefold in adult CD4⁺ T cells, but not significantly increased in neonatal cells. However, adoptively transferred neoTreg cells significantly prolonged cardiac allograft survival (mean survival time 47 days, P < 0·001) and maintained Foxp3 expression similar to natural Treg cells. The neoTreg cells were hypomethylated at the conserved non-coding DNA sequence 2 locus of Foxp3 compared with adult Treg cells. The DNMT antagonist 5-aza-2′-deoxycytidine (5-Aza) induced increased Foxp3 expression in mature CD4⁺ T cells. 5-Aza-inducible Treg cells combined with continuous 5-Aza treatment prolonged graft survival. These results indicate that the ‘default’ pathway of neoTreg cell differentiation is associated with reduced DNMT1 and DNMT3b response to TCR stimulus. The neoTreg cells may be a strategy to alleviate acute allograft rejection.     

肌醇多磷酸5-磷酸酶(PIPP)在人下咽癌组织的表达及临床意义

目的研究肌醇多磷酸5-磷酸酶(PIPP)在人下咽癌中的表达,探讨其临床意义。方法对15例人下咽癌组织学标本和15例正常的癌旁组织进行免疫荧光和免疫印迹检测。结果 PIPP的表达量及荧光强度在下咽癌组织中明显低于正常组织。结论 PIPP在下咽癌组织中低表达可能为临床提供新的诊断指标。     

三维经食管超声心动图导航左心室室壁瘤手术中的作用分析简

目的室壁瘤切除术中应用三维经食管超声心动图（3D-TEE）导航,定量分析手术前后室壁瘤患者左心室功能,探讨对患者远期愈后的作用。方法64例行左心室室壁瘤切除手术患者,分2组,每组32例。行3D-TEE为试验组,其中男性29例.女性3例：年龄46～76岁,平均年龄59.9岁。对照组术中未行3D-TEE检查,其中男性29例,女性3例;年龄38。79岁,平均年龄60.0岁。在患者手术前3d内、术后1年分别行3D-TTE检查,对照分析同期行室壁瘤切除手术患者心功能,并记录分析两组患者手术时间、术后辅助治疗、术后不良事件发生及治疗前后左心室结构与功能改善的情况。结果术后试验组左心室射血分数为（45.88±6.60）％,每搏量（49.95±7.18）mL,心指数（2.17±0.44）IJ（min·m^2）;对照组左心室射血分数为（37.88±6.82）％,每搏量（40.54±10.12）mL,心指数（1.68±0.38）I／（min·m^2）;试验组明显高于对照组,二者差异有统计学意义（P〈0.05）。两组左心室舒张末容积差异无统计学意义（P〉0.05）.但试验组左心室收缩末容积明显低于对照组.二者差异有统计学意义（P〈0.05）。两组患者手术时间相差无统计学意义。结论术中应用3D-TEE检查能够安全、便捷地评价室壁瘤形成患者的左心室。在术中应用3D-TEE为导航.更利于外科医生的判断和处理.对患者的远期效果应该更好。     

Highly sensitive and specific screening of EGFR mutation using a PNA microarray-based fluorometric assay based on rolling circle amplification and graphene oxide

Screening epidermal growth factor receptor (EGFR) mutations, especially deletions, is essential for diagnosis of non-small cell lung cancer (NSCLC) and also critical to inform treatment decisions for NSCLC patients. Here, we demonstrated a facile peptide nucleic acid (PNA) microarray-based fluorometric method for sensitive and specific detection of EGFR mutation, using rolling circle amplification (RCA), graphene oxide (GO), and a fluorescently-labeled detection probe (F-DP). First, the EGFR gene sequence was efficiently captured by the label-free PNA probe which was attached on the surface of a 96-well plate. And then, the EGFR mutation sequence was specifically amplified by RCA using the circular DNA, which was formed by the ligation of the padlock probe when hybridizing with the EGFR mutation, as a template. The single-stranded RCA product (RCAP) was then sensitively detected with the F-DP and GO system. This method has a detection limit of 0.3 pM for EGFR mutation and a high discrimination capability to target EGFR mutation against EGFR wildtype. The use of a PNA microarray and a fluorescence quenching platform make this system quite suitable for high-throughput analysis of EGFR mutations in resource-limited settings without the need of costly and cumbersome equipment. Furthermore, this detection system provides a novel way for the diagnosis of other diseases that are caused by gene deletion mutations.

A facile peptide nucleic acid microarray-based fluorometric method was developed for sensitive and specific detection of EGFR mutation by using rolling circle amplification, graphene oxide, and a fluorescently-labeled detection probe.     

市售炮制辅料醋中5-羟甲基糠醛含量的比较

目的 比较不同品种市售炮制辅料醋中5-羟甲基糠醛的含量。方法 采用高效液相色谱法测定5-羟甲基糠醛的含量,色谱条件为流动相甲醇-0.05%磷酸（8:92）,流速1.0 mL·min^-1,检测波长284 nm,进样量20 mL。结果 3批白醋中未检测到5-羟甲基糠醛,以黑糯米、小米等优质粮食为原料酿造的米醋中5-羟甲基糠醛含量相对较高,天立老醋、水塔老陈醋和恒顺镇江陈醋等有陈酿工艺的米醋中5-羟甲基糠醛含量高。结论 5-羟甲基糠醛含量的高低反映了米醋质量的优劣,5-羟甲基糠醛可作为米醋的质量控制指标。     

根痛平胶囊和根痛平片中川续断皂苷Ⅵ的含量测定方法研究及续断的投料情况分析

目的 建立根痛平胶囊和根痛平片中川续断皂苷Ⅵ的含量测定方法,监测生产企业续断投料情况。方法 采用高效液相色谱(HPLC)法,Agilent Zorbax SB-C₁₈色谱柱(4.6 mm×250 mm,5 μm),以乙腈-水（27:73）为流动相等度洗脱,流速1.0 mL·min-1,检测波长为212 nm。结果 川续断皂苷Ⅵ在30~280 μg·mL^-1的范围内呈良好的线性关系,回归方程为Y = 2 029.357 0 X + 3.688 5 ,R² = 0.999 7,平均回收率为96.08%,RSD为1.01%。结论 该法简单准确,可用于测定根痛平胶囊和根痛平片中川续断皂苷Ⅵ的含量,为根痛平系列制剂的质量标准提高提供理论依据;制定了川续断皂苷Ⅵ含量参考限度值,考察企业续断的投料情况,提示个别企业未投料或投料用续断质量差。     

两种方法治疗垂直不稳定型股骨颈骨折的近期疗效对比

目的评价单纯空心螺钉与3枚空心钉加内侧支撑钢板两种方式处理股骨颈Pauwels Ⅲ型骨折的近期治疗效果。方法选取锦州地区55例股骨颈Pauwels Ⅲ型骨折患者,随机分为空心加压螺钉组30例（A组）,3枚空心螺钉+支撑钢板组25例（B组）。记录术后髋关节Harris评分,对比各项手术指标、愈合时间、复位质量、股骨头坏死与骨不连等并发症的发生情况。结果A组手术时间（68.73±4.91）min,低于B组（110.76±5.85）min;A组术中出血量（69.47±5.15）mL,低于B组（110.92±10.10）mL;A组的骨折愈合时间（5.03±0.47）个月,高于B组（4.37±0.56）个月;差异均有统计学意义（P<0.05）。两组内固定失效和复位情况的对比,B组优于A组（P<0.05）;在股骨头坏死和骨折不愈合方面的比较,A组高于B组（P>0.05）。术后第1、3个月的Harris分值,A组为（70.67±1.88）分、（74.83±3.69）分,低于B组（71.60±1.78）分、（76.60±3.10）分,差异无统计学意义（P>0.05）;术后第6、12个月的Harris分值,A组为（78.97±1.88）分、（88.17±3.36）分,低于B组（80.08±1.93）分、（92.04±3.72）分,差异有统计学意义（P<0.05）。结论采用空心钉+支撑钢板治疗骨折线较垂直的股骨颈Pauwels Ⅲ型骨折的效果优于单纯3枚空心钉,拥有骨折复位质量好、更加牢靠的固定、有效抵抗垂直剪切力、缩短愈合时间,患髋功能恢复好等优点。     

结核感染T细胞斑点试验联合胸腔积液Xpert MTB/RIF检测诊断结核性胸腔积液的价值

目的评估结核感染T细胞斑点试验（T-SPOT.TB）联合胸腔积液结核分枝杆菌/利福平耐药实时荧光定量核酸扩增检测技术(Xpert MTB/RIF) 在结核性胸腔积液的诊断价值。方法收集胸腔积液患者112例,其中确诊或临床诊断为结核性胸腔积液76例,非结核性胸腔积液36例。所纳入患者治疗前均行胸腔积液T-SPOT.TB、血T-SPOT.TB、胸腔积液Xpert MTB/RIF、胸腔积液腺苷脱氨酶（ADA）检测,并对其检测结果进行分析。结果胸腔积液T-SPOT.TB诊断结核性胸腔积液的敏感度（97.37%,74/76),高于外周血T-SPOT.TB （76.32%,58/76）、胸腔积液Xpert MTB/RIF（65.79%,50/76）和胸腔积液ADA（28.95%,22/76）,差异均有统计学意义（χ2=14.74、25.22、76.45,P<0.01）。胸腔积液Xpert MTB/RIF检测的特异度（100%,36/36）均高于胸腔积液T-SPOT.TB（77.78%,28/36）、外周血T-SPOT.TB（55.56%,20/36）和胸腔积液ADA（50.00%, 18/36）,差异均有统计学意义（χ2=6.89、20.57、24.00,P均＜0.01）。胸腔积液T-SPOT.TB和胸腔积液Xpert MTB/RIF联合检测的敏感度（64.47%, 49/76）低于胸腔积液T-SPOT.TB单独检测（97.37%,74/76）,差异有统计学意义（χ2=26.63,P<0.01）,但特异度由77.78%(28/36)提高至100%(36/36),差异有统计学意义（χ2=45.82,P<0.01）。结论胸腔积液T-SPOT.TB检测结核性胸腔积液具有较高的敏感度和特异度,胸腔积液T-SPOT.TB和胸腔积液Xpert Mtb/ＲIF联合检测可明显提高诊断的特异度。