Control of DNA end resection by yeast Hmo1p affects efficiency of DNA end-joining

The primary pathways for DNA double strand break (DSB) repair are homologous recombination (HR) and non-homologous end–joining (NHEJ). The choice between HR and NHEJ is influenced by the extent of DNA end resection, as extensive resection is required for HR but repressive to NHEJ. Conversely, association of the DNA end-binding protein Ku, which is integral to classical NHEJ, inhibits resection. In absence of key NHEJ components, a third repair pathway is exposed; this alternative-end joining (A-EJ) is a highly error-prone process that uses micro-homologies at the breakpoints and is initiated by DNA end resection. In Saccharomyces cerevisiae, the high mobility group protein Hmo1p has been implicated in controlling DNA end resection, suggesting its potential role in repair pathway choice. Using a plasmid end-joining assay, we show here that absence of Hmo1p results in reduced repair efficiency and accuracy, indicating that Hmo1p promotes end-joining; this effect is only observed on DNA with protruding ends. Notably, inhibition of DNA end resection in an hmo1Δ strain restores repair efficiency to the levels observed in wild-type cells. In absence of Ku, HMO1 deletion also reduces repair efficiency further, while inhibition of resection restores repair efficiency to the levels observed in kuΔ. We suggest that Hmo1p functions to control DNA end resection, thereby preventing error-prone A-EJ repair and directing repairs towards classical NHEJ. The very low efficiency of DSB repair in kuΔhmo1Δ cells further suggests that excessive DNA resection is inhibitory for A-EJ.     

Attempts to label matrix synthesis of human root cementum in vitro

The present study describes the dynamic process of both acellular extrinsic (AEFC) and acellular/cellular intrinsic fiber cementum (AIFC/CIFC) matrix production on growing human teeth. Selected erupting maxillary and mandibular premolars with roots grown to about 70%–95% of their final length were placed in organ culture immediately following extraction. Twelve teeth for short-time labeling were pulse-incubated for 15 min in medium containing ³H-proline and chased for various times in order to follow the migration and secretion of the tracer. Eight teeth for long-time incubation were labeled continuously for 5 h before being chased for 1–8 days in order to label cementum matrix accumulation. After decalcification in ethylene diaminetetraacetic acid (EDTA), their roots were subdivided into about 20 slices each. Epon-embedded sections were prepared for light- and electron-microsopic as well as autoradiographic examination. During CIFC-formation, cementoblasts revealed high intracytoplasmic silver grain concentrations within the first hour after ³H-proline administration. The release of the tracer occurred between 60 to 120 min after administration. After 2 h, cementoblasts and the cementum matrix appeared to be labeled about equally. After 5 h, most of the labeled proteins appeared to be localized in the cementoid. Silver grains increased in number over the cementum matrix from 5–24 h. Very high intracellular grain concentrations within very large cementoblasts corresponded to regions of rapid cementum formation. Tracer-halos around entrapped cells lend support to a multipolar mode of matrix production during CIFC-initiation. The fate of the tracer during the development of early AEFC-matrix was less clear. However, fibroblasts revealed dense intracytoplasmic grain accumulations within the first hour after ³H-proline administration. Thereafter, the tracer localization was vague. This indistinct grain localization reflected the particular mode of AEFC-matrix production characterized by addition of new fibril segments to pre-existing fibers of a collagenous fringe.     

Changes in DNA double-strand break repair during aging correlate with an increase in genomic mutations

A double -strand break (DSB) is one of the most deleterious forms of DNA damage. In eukaryotic cells, two main repair pathways have evolved to repair DSBs, homologous recombination (HR) and non-homologous end-joining (NHEJ). HR is the predominant pathway of repair in the unicellular eukaryotic organism, S. cerevisiae. However, during replicative aging the relative use of HR and NHEJ shifts in favor of end-joining repair. By monitoring repair events in the HO-DSB system, we find that early in replicative aging there is a decrease in the association of long-range resection factors, Dna2-Sgs1 and Exo1 at the break site and a decrease in DNA resection. Subsequently, as aging progressed, the recovery of Ku70 at DSBs decreased and the break site associated with the nuclear pore complex at the nuclear periphery, which is the location where DSB repair occurs through alternative pathways that are more mutagenic. End-bridging remained intact as HR and NHEJ declined, but eventually it too became disrupted in cells at advanced replicative age. In all, our work provides insight into the molecular changes in DSB repair pathway during replicative aging. HR first declined, resulting in a transient increase in the NHEJ. However, with increased cellular divisions, Ku70 recovery at DSBs and NHEJ subsequently declined. In wild type cells of advanced replicative age, there was a high frequency of repair products with genomic deletions and microhomologies at the break junction, events not observed in young cells which repaired primarily by HR.     

右美托咪定联合依托咪酯对老年直肠癌根治术患者术后炎症反应、胃肠功能恢复和认知功能的影响

摘要 目的：探讨右美托咪定联合依托咪酯对老年直肠癌根治术患者术后炎症反应、胃肠功能恢复和认知功能的影响。方法：选择2019年3月~2021年5月南京鼓楼医院收治的180例老年直肠癌患者,均接受腹腔镜下直肠癌根治术治疗。根据随机数字表法将患者分为对照组和研究组,各为90例。对照组患者麻醉选用依托咪酯,研究组患者麻醉选用右美托咪定联合依托咪酯,对比两组术中血流动力学、术后炎症反应、胃肠功能恢复和认知功能,同时记录两组围麻醉期不良反应发生情况。结果：插管即刻（T2）~拔管即刻（T4）时间点,两组心率（HR）、平均动脉压（MAP）先下降后升高,且研究组的波动幅度小于对照组（P<0.05）。两组进食时间组间对比无统计学差异（P>0.05）,研究组的肠鸣音恢复时间、首次排气时间短于对照组（P<0.05）。两组气管拔管时间、呼吸恢复时间、麻醉苏醒时间对比无统计学差异（P>0.05）。两组术后3 d白介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）和C反应蛋白（CRP）水平均升高,且研究组的变化幅度小于对照组（P<0.05）。两组术后1 d、术后3 d简明智能状态量表（MMSE）评分先下降后升高,且研究组的波动幅度小于对照组（P<0.05）。两组不良反应发生率组间对比无差异（P>0.05）。结论：老年直肠癌根治术患者麻醉方案选用右美托咪定联合依托咪酯,可减轻机体炎性应激,稳定机体血流动力学,有利于胃肠功能恢复,同时还可减轻对机体认知功能的损害。     

术前AGR、NLR、FOXQ1联合检测对低位直肠癌根治性切除手术患者术后复发的预测价值

摘要 目的：探讨术前白蛋白-球蛋白比值（AGR）、中性粒细胞-淋巴细胞比值（NLR）、叉头框蛋白Q1（FOXQ1）联合检测对低位直肠癌根治性切除手术患者术后复发的预测价值。方法：选取2017年3月至2019年4月重庆市第九人民医院就诊的拟行低位直肠癌根治性切除手术患者110例,术前均检测血清AGR、NLR、FOXQ1水平。随访3~48个月,中位随访时间为25.5个月,失访9例,101例根据术后复发情况将患者分为复发组（n=16）和未复发组（n=85）。比较两组患者术前血清AGR、NLR、FOXQ1水平,收集患者的临床资料,以Logistic回归分析探讨低位直肠癌患者术后复发的危险因素,绘制受试者工作曲线（ROC）判定术前血清AGR、NLR、FOXQ1水平对低位直肠癌患者术后复发的预测价值。结果：复发组术前血清AGR水平低于未复发组,术前血清NLR、FOXQ1水平高于未复发组（P<0.05）;复发组患者肿瘤细胞分化程度为低分化、TNM分期为Ⅲ期、糖链抗原19-9（CA19-9）阳性占比高于未复发组（P<0.05）,复发组术后化疗占比低于未复发组（P<0.05）;Logistic回归分析结果显示,TNM分期为Ⅲ期、细胞分化程度为低分化、术前血清AGR降低、NLR升高、FOXQ1升高均为低位直肠癌患者术后复发的危险因素（P＜0.05）;ROC曲线结果显示,术前血清AGR、NLR、FOXQ1水平及三者联合对低位直肠癌患者术后复发的预测的曲线下面积（AUC）值分别为0.738、0.747、0.731、0.842。结论：术前检测血清AGR、NLR、FOXQ1水平对低位直肠癌根治性切除术患者术后复发预测具有一定的价值,且联合的预测价值更高。     

经尿道等离子前列腺剜除术与经尿道前列腺电切术治疗良性前列腺增生症的临床疗效对比研究

摘要 目的：对比经尿道等离子前列腺剜除术（TUKEP）与经尿道前列腺电切术（TURP）治疗良性前列腺增生症（BPH）的临床疗效。方法：回顾性分析我院2018年1月1日至2021年3月17日期间收治的200例BPH患者的临床资料。根据手术方式的不同将患者分为A组（n=83）和B组（n=117）,A组手术方式为TURP,B组手术方式为TUKEP,比较两组围术期指标,随访6个月,对比两组性功能、尿流动力学变化及并发症发生情况。结果：B组手术时间、术中出血量、尿管留置时间、住院时间、术后冲洗时间短于A组（P<0.05）。术后6个月,两组患者的最大尿流速（Qmax）、膀胱顺应性（BC）升高,剩余尿量（PVR）降低（P<0.05）,且B组患者的Qmax、BC高于A组,PVR低于A组（P<0.05）。术后6个月,两组患者的勃起功能评分表（IIEF-5）、射精功能评分表（CIPE-5）评分降低（P<0.05）,但B组、A组IIEF-5、CIPE-5评分组间对比无明显统计学差异（P>0.05）。B组的并发症发生率小于A组（P<0.05）。结论：TUKEP、TURP治疗BPH,疗效相当,TUKEP在缩短手术时间、尿管留置时间、术后冲洗时间、住院时间,降低术中出血量,减少并发症发生率,改善尿流动力学方面更有优势。     

右美托咪定联合七氟醚对老年腹腔镜胃肠肿瘤切除术患者应激反应和脑氧代谢的影响

摘要 目的：观察老年腹腔镜胃肠肿瘤切除术患者使用七氟醚联合右美托咪定复合麻醉后,机体应激反应和脑氧代谢的变化情况。方法：将我院2020年1月至2020年12月期间收治的100例老年腹腔镜胃肠肿瘤切除术患者根据随机数字表法分为对照组（n=50,七氟醚麻醉）和研究组（n=50,右美托咪定联合七氟醚麻醉）。观察两组血流动力学、应激反应、脑氧代谢、认知功能和不良反应。结果：两组手术60 min（T1）~手术后30 min（T3）时间点较麻醉诱导前（T0）时间点心率（HR）先下降后升高,平均动脉压（MAP）先升高后下降（P<0.05）。研究组T1~T3时间点HR及MAP低于对照组（P<0.05）。两组术后3 d多巴胺（DA）、肾上腺素（AD）、去甲肾上腺素（NE）水平升高,但研究组低于对照组（P<0.05）。两组术后3 d血氧含量差（DajvO₂）和脑氧摄取率（CERO₂）下降,且研究组低于对照组（P<0.05）。研究组术后6 h、术后1 d、术后3 d简易精神状态检查量表（MMSE）评分高于对照组（P<0.05）。两组不良反应发生率,对比无显著性差异（P>0.05）。结论：七氟醚联合右美托咪定应用于老年腹腔镜胃肠肿瘤切除术患者,可有效减轻机体应激反应,减轻机体认知功能损伤,维持血流动力学稳定和脑氧代谢。     

不同剂量右美托咪定辅助麻醉对老年结肠癌根治术患者麻醉效果和术后谵妄的影响及谵妄的相关因素分析

摘要 目的：探讨不同剂量右美托咪定辅助麻醉对老年结肠癌根治术患者麻醉效果,并分析术后谵妄的影响因素。方法：选取2019年4月~2021年1月期间我院收治的160例老年结肠癌根治术患者,根据随机数字表法将患者分为A组（53例）、B组（53例）和C组（54例）。A组和B组均于麻醉诱导前给予0.5 μg/kg右美托咪定,以0.2 μg/kg?h速率静脉输注至手术结束前30 min,A组术后镇痛时再给予右美托咪定0.05 μg/kg?h。C组给予等容量和等速率的生理盐水。观察三组的应激反应、麻醉效果、不良反应发生率、谵妄发生率。根据是否发生谵妄分为谵妄组（n=28）和无谵妄组（n=132）。采用Logistic回归分析术后谵妄的影响因素。结果：三组术后第3 d C反应蛋白（CRP）、白介素-6（IL-6）、多巴胺、肾上腺素均较手术结束升高（P<0.05）。A组、B组术后第3 d CRP、IL-6、多巴胺、肾上腺素低于C组（P<0.05）。A组、B组术后第3dCRP、IL-6、多巴胺、肾上腺素组间对比,无明显差异（P>0.05）。A组、B组、C组的不良反应总发生率组间的对比无明显差异（P>0.05）。B组的谵妄发生率明显低于A组、C组（P<0.05）。A组、C组谵妄发生率组间对比无明显差异（P>0.05）。单因素分析结果显示,术后谵妄的发生与年龄、术前抑郁、术前合并基础疾病数量、术中低氧血症、气腹后PaCO₂、白蛋白有关（P<0.05）。多因素Logistic回归分析结果显示：术前抑郁、年龄≥70岁、术前合并基础疾病数量≥3、术中低氧血症、气腹后PaCO₂偏高、白蛋白偏低是导致老年结肠癌根治术患者术后谵妄的危险因素（P<0.05）。结论：小剂量右美托咪定辅助麻醉可提高老年结肠癌根治术患者的麻醉效果,减少谵妄的发生率,同时术前抑郁、年龄≥70岁、术前合并基础疾病数量≥3、术中低氧血症、气腹后PaCO₂偏高、白蛋白偏低是引起谵妄发生的危险因素。     

术前中性粒细胞绝对值/淋巴细胞比值联合血清瘦素、睾酮对前列腺癌根治术后生化复发的评估价值

摘要 目的：探讨术前中性粒细胞绝对值/淋巴细胞比值（NLR）联合血清瘦素、睾酮对前列腺癌（PCa）根治术后生化复发的评估价值。方法：选取2018年1月～2020年1月于我院接受根治性切除术治疗的82例PCa患者。术前均检测NLR、血清瘦素、睾酮水平。术后对所有患者均进行2年随访观察,按照是否发生生化复发分为复发组（n=34）以及无复发组（n=48）。比较两组NLR、血清瘦素、睾酮水平差异。收集患者临床资料,采用多因素Logistic回归分析PCa根治术后生化复发的影响因素。采用受试者工作特征（ROC）曲线分析NLR以及血清瘦素、睾酮预测PCa根治术后生化复发的评估价值。结果：复发组术前NLR以及瘦素水平均高于无复发组（P＜0.05）,而睾酮水平低于无复发组（P＜0.05）。复发组术前前列腺特异抗原（PSA）≥10 ng/mL、TNM分期T2期人数占比以及Gleason评分均高于无复发组（P＜0.05）。多因素Logistic回归分析显示,术前PSA≥10 ng/mL、TNM分期T2期、Gleason评分较高、术前NLR较高、瘦素水平较高、睾酮水平较低是PCa根治术后生化复发的危险因素（P＜0.05）。ROC曲线分析显示,联合检测术前NLR、血清瘦素、睾酮水平预测PCa根治术后生化复发的ROC曲线下面积为0.897,高于三项指标单独检测的0.678、0.712、0.733。结论：PCa根治术后生化复发受术前PSA、NLR、血清瘦素、睾酮水平、TNM分期、Gleason评分等因素影响,术前NLR联合血清瘦素、睾酮对PCa根治术后生化复发的预测价值较高。     

Sumoylation regulates EXO1 stability and processing of DNA damage

DNA double-strand break repair by the error-free pathway of homologous recombination (HR) requires the concerted action of several factors. Among these, EXO1 and DNA2/BLM are responsible for the extensive resection of DNA ends to produce 3′-overhangs, which are essential intermediates for downstream steps of HR. Here we show that EXO1 is a SUMO target and that sumoylation affects EXO1 ubiquitylation and protein stability. We identify an UBC9-PIAS1/PIAS4-dependent mechanism controlling human EXO1 sumoylation in vivo and demonstrate conservation of this mechanism in yeast by the Ubc9-Siz1/Siz2 using an in vitro reconstituted system. Furthermore, we show physical interaction between EXO1 and the de-sumoylating enzyme SENP6 both in vitro and in vivo, promoting EXO1 stability. Finally, we identify the major sites of sumoylation in EXO1 and show that ectopic expression of a sumoylation-deficient form of EXO1 rescues the DNA damage-induced chromosomal aberrations observed upon wt-EXO1 expression. Thus, our study identifies a novel layer of regulation of EXO1, making the pathways that regulate its function an ideal target for therapeutic intervention.