动物模型中人骨形态发生蛋白2重组腺病毒活性观测

目的:利用基因工程技术构建人骨形态发生蛋白2重组腺病毒,并进行体内表达,测定活性;为深入开展的基因治疗研究创造条件。方法:实验于2000-05/2002-06在西安交通大学第一附属医院骨科实验室完成。应用PCR技术扩增出人骨形态发生蛋白2全长基因,体外同源重组构建重组腺病毒后,随机将30只SD大鼠分为治疗组和对照组,每组15只,将纯化后的重组腺病毒50μL注入SD大鼠股部肌肉陷窝病损处,通过组织学染色、X射线片对动物模型的组织变化进行观测。结果:6周后治疗组可见明显骨诱导形成,2周时治疗组碱性磷酸酶活性明显高于对照组,重组腺病毒治疗组有明显的诱导成骨活性。结论:应用于动物实验中目的基因表达,并具有生物学活性。本研究是在骨缺损基因治疗的一次尝试,且人骨形态发生蛋白2重组腺病毒的构建成功,也为国内骨科疾病基因治疗的进一步研究提供了基础。     

五例恙虫病东方体感染基因型及其基因变异

目的旨在了解浙闽地区人群恙虫病东方体感染的基因型,并分析其基因变异。 方法从浙闽地区5例散发恙虫病发热期患者血中分离病原体并进行细胞培养;提取感染细胞DNA,巢式PCR扩增完整恙虫病东方体56-kD型特异性抗原（TSA）基因和热休克蛋白基因（groESL）并测序;采用MEGA 7.0软件,进行序列比对和系统发育分析。 结果病原学确认5例恙虫病患者,并分离到恙虫病东方体菌株。序列比对表明,5菌株中有2株的56-kD TSA基因和groESL基因100%一致,另2菌株的此二个基因序列一致性亦为100%,分别将两组菌暂时命名为浙江-1型和浙江-2型。56-kDa TSA基因比对和系统发育分析表明,浙江-1型和浙江-2型分别与台湾-A基因型和Gilliam-C基因型亲缘较近（98.45%和98.50%）,但有明显变异;另1株菌Wuj/2014与台湾-A基因型高度相似（99.94%）。56-kDa TSA基因各基因型支系的时间树分析表明,台湾-A基因型、浙江-1型和浙江-2型3支系与祖先的分歧时间相对其他原型株晚,尤其是浙江-2型,说明这些基因型或亚型在恙虫病东方体的进化过程中出现较晚。 结论本研究病例所感染恙虫病东方体基因型不同,可能为未被认识的新的基因亚型,迫切需要进行全基因组测序确认,并探讨其基因变异与人恙虫病严重程度间的关系。     

输尿管软镜碎石术治疗上尿路结石的疗效和安全性

目的探讨输尿管软镜碎石术(retrograde intrarenal stone surgery,RIRS)治疗上尿路结石的疗效和安全性。方法回顾性分析2016年4月至2019年1月武汉大学人民医院行RIRS的640例上尿路结石患者的临床资料。男424例,女216例。年龄(46.2±12.8)岁。结石最大径(1.4±0.7)cm。肾下盏结石126例,非肾下盏结石514例。单侧输尿管上段结石196例,单侧输尿管上段结石合并肾结石118例,单侧肾结石236例,双肾结石90例。104例术前留置双J管。马蹄肾8例,孤立肾合并肾功能不全30例,盆腔异位肾合并旋转不良4例,先天性输尿管畸形6例,海绵肾2例。术前血红蛋白(133.2±5.6)g/L,血清肌酐(84.4±12.2)μmol/L。手术均采用全麻,患者取截石位。采用输尿管软镜联合钬激光碎石,软镜顺利进入肾盂后首先观察肾盂及各肾盏并寻及结石。使用200μm光纤碎石,钬激光功率为12~45 W(0.5~1.5 J/10~30 Hz),根据实际情况辅助取石网篮套取结石。术中检查各肾盂、肾盏,确保结石已完全粉末化(结石最大径<0.3 cm),留置双J管和尿管。手术均由同等资历的术者完成。结果所有手术均顺利完成,手术时间(45.6±14.6)min。术后第1天复查血清肌酐(76.0±10.6)μmol/L,与术前比较差异有统计学意义(t=64.76,P<0.05);血红蛋白(126.4±9.6)g/L,与术前比较差异无统计学意义(t=2.02,P=0.064)。术后住院时间(4.8±1.5)d。术后3例(0.9%)发生严重并发症,分别为2例脓毒血症,1例包膜下血肿。术后3个月596例获得随访,其中552例达到结石清除标准,结石清除率(stone-free rate,SFR)为92.6%;余44例未达到结石清除标准者采用体外冲击波碎石、再次输尿管软镜手术或拔除双J管后观察。结石大小(χ^2=29.569,P<0.05)和位置(χ^2=44.949,P<0.05)是SFR的影响因素。多因素回归分析结果显示结石大小不是影响SFR的独立危险因素(P=0.639),结石位置是影响SFR的独立危险因素(P=0.013)。结论RIRS对于上尿路中小结石患者是一种可靠的治疗方式,疗效确切,并发症少,安全性高。但对于大结石及肾下盏结石的治疗还存在一定的局限性。肾下盏结石是影响RIRS疗效的独立危险因素。     

碘伏治疗鼻咽癌放疗后口腔溃疡的护理

放射治疗(简称放疗)是目前治疗鼻咽癌的首选治疗方法,而大剂量的放疗对口腔黏膜有直接损伤作用,特别是同时接受化学疗法的病人,极易发生口腔溃疡,影响进食。加之病人免疫力降低,易发生严重感染,从而影响病人的正常治疗,导致治疗中断乃至失败。自2001年7月—2005年7月作者采用碘     

Radiological studies on the best entry point and trajectory of anterior cervical pedicle screw in the lower cervical spine

Objective

To explore the best entry point and trajectory of anterior cervical transpedicular screws in the lower cervical spine by radiological studies, and provide reference for clinical application.

Methods

Fifty patients were scanned by computed tomography and confirmed no obvious defect of the cervical spine. On horizontal axis, camber angle (α) and axial length (AL) were measured from C₃ to C₇. On sagittal view, the cranial or caudal angle (β) and sagittal length (SL) were also measured from C₃ to C₇. On the sagittal and horizontal planes vertebrae were respectively divided into four areas, ordered 1–4, on the anterior side of the pedicle. The areas and angles of pedicle intersect into the vertebral body were recorded. We inserted six anterior pedicle screws into the lower cervical spine of three patients by this technique.

Results

On transverse plane, camber angle (α) of C₃–C₅ increased gradually, while it decreased from C₅ to C₇. On sagittal view, C₃ and C₄ pedicles showed cranial tilting, while C₅ to C₇ were caudally tilted. AL and SL values increased gradually from C₃ to C₇. The number of the intersections of C₃–C₇ in each area was also different. Six pedicle screws of three cases were inserted into the lower cervical spine with proper placement and no complications.

Conclusion

Anterior transpedicular screw (ATPS) is a theoretically feasible option for internal fixation. The technique described in this paper was subsequently used in three patients without complication. Future improvement of ATPS insertion remains necessary for this technically demanding procedure.     

Geminivirus–Host Interactions: Action and Reaction in Receptor-Mediated Antiviral Immunity

In plant−virus interactions, the plant immune system and virulence strategies are under constant pressure for dominance, and the balance of these opposing selection pressures can result in disease or resistance. The naturally evolving plant antiviral immune defense consists of a multilayered perception system represented by pattern recognition receptors (PRR) and resistance (R) proteins similarly to the nonviral pathogen innate defenses. Another layer of antiviral immunity, signaling via a cell surface receptor-like kinase to inhibit host and viral mRNA translation, has been identified as a virulence target of the geminivirus nuclear shuttle protein. The Geminiviridae family comprises broad-host range viruses that cause devastating plant diseases in a large variety of relevant crops and vegetables and hence have evolved a repertoire of immune-suppressing functions. In this review, we discuss the primary layers of the receptor-mediated antiviral immune system, focusing on the mechanisms developed by geminiviruses to overcome plant immunity.     

Identification and validation of an autophagy-related long non-coding RNA signature as a prognostic biomarker for patients with lung adenocarcinoma

BackgroundLung adenocarcinoma (LUAD) is the most predominant pathological subtype of lung cancer, accounting for 40–70% of all lung cancer cases. Although significant improvements have been made in the screening, diagnosis, and precise management in recent years, the prognosis of LUAD remains bleak. This study aimed to investigate the prognostic significance of autophagy-related long non-coding RNAs (lncRNAs) and construct an autophagy-related lncRNA prognostic model in LUAD.MethodsThe gene expression data of LUAD patients were obtained from The Cancer Genome Atlas (TCGA) database. All autophagy-related genes were downloaded from the Human Autophagy Database (HADb). Spearman’s correlation test was exploited to identify potential autophagy-related lncRNAs. The multivariate Cox regression analysis was used to construct the prognostic signature, which divided LUAD patients into high-risk and low-risk groups. Subsequently, the receiver operating characteristic (ROC) curves were generated to assess the predictive ability of this prognostic model for overall survival (OS) in these individuals. Then, the Gene set enrichment analysis (GSEA) was conducted to execute pathway enrichment analysis. Finally, a multidimensional validation was exploited to verify our findings.ResultsA total of 1,144 autophagy-related lncRNAs were identified to construct the co-expression network via Spearman’s correlation test (|R²| >0.4 and P≤0.001). Ultimately, a 16 autophagy-related lncRNAs prognostic model was constructed, and the area under the ROC curve (AUC) was 0.775. The results of GSEA enrichment analysis showed that the genes in the high-risk group were mainly enriched in cell cycle and p53 signaling pathways. The results of the multidimensional database validation indicated that the expression level of BIRC5 was significantly correlated with the expression level of TMPO-AS1. Furthermore, both TMPO-AS1 and BIRC5 had a higher expression level in LUAD samples. LUAD patients with high expression levels of TMPO-AS1 and BIRC5 were correlated with advanced disease stage and poor OS.ConclusionsIn summary, our results suggested that the prognostic signature of the 16 autophagy-related lncRNAs has significant prognostic value for LUAD patients. Furthermore, TMPO-AS1 and BIRC5 are potential predictors and therapeutic targets in these individuals.     

Synthesis and Evaluation of a CBZ‐AAN‐Dox Prodrug and its in vitro Effects on SiHa Cervical Cancer Cells Under Hypoxic Conditions

Although doxorubicin (Dox) is widely used in clinical treatment for solid tumors, it causes many side‐effects such as heart and kidney damage, bone marrow suppression, and drug resistance. Legumain is a lysosomal protease that is elevated and associated with an invasive and metastatic phenotype in a number of solid tumors. In this study, we designed and synthesized a Dox prodrug, N‐benzyloxycarbonyl‐Ala‐Ala‐Asn‐Doxorubicin (CBZ‐AAN‐Dox), with 94% purity. Single substrate kinetic assays demonstrated hLegumain‐specific enzymatic cleavage and activation of the prodrug in vitro, and this enzymatic cleavage of the prodrug substrate was more sensitive in acidic conditions, releasing more than 70% of Dox after 24 h. Treatment of tumor cells with our prodrug demonstrated a much higher IC₅₀ value, significantly enhanced uptake of the prodrug, and considerably less cellular toxicity compared to Dox treatment alone. Our study presents a novel prodrug, CBZ‐AAN‐Dox, to potentially increase both the safety and efficacy of clinical treatment of tumors by exploiting the tumor's innate expression of legumain.