精确循证促进中药注射剂合理合法应用的策略与路径

中药注射剂临床应用主要存在两大问题：说明书对临床用药指导不足和临床应用环节不合理。问题的核心是缺少精确循证研究的推进。针对当前问题,应采取由政府药政部门主导,逐步形成医疗机构-研究者-企业多方协作平台共同开展精确、循证研究的策略。精确临床定位、完善药品不良反应监测体系、评估用药风险,医疗机构充分实践精确循证研究产出的证据可为具体实现路径。中药注射剂做到临床定位清晰、具体,同时机制清晰,靶点明确。优胜劣汰,从而增强中药注射剂优势品种的核心竞争力,在保护人类生命健康事业中做贡献。     

套式聚合酶链反应及限制酶分析检测新生儿肝炎巨细胞病毒感染

在人巨细胞病毒（HCMV）AD169株直接早期蛋白EcoRIJDNA片段内，自行设计二对引物，建立套式PCR检测HCMVDNA。经实验证明有高度的特异性与敏感性，对其它疙疹类病毒和正常人基因组DNA无交叉反应。一次性PCR检测HCMVAD169株基因组的最小检出量为100fg，而套式PCR可达10fg，经PStI酶切后，得到的片段与设计相符。对23例新生儿肝炎临床标本进行病毒分离，一次性PCR，套式PCR检测，结果表明套式PCR能进一步提高一次性PCR的特异性与敏感性，适用于新生儿肝炎HCMV感染的临测。     

重组质粒pRNAT-U6.1/CFB siRNA的构建、鉴定及其对人脐静脉内皮细胞增生的影响

背景 脉络膜新生血管(CNV)是多种眼部疾病致盲的原因之一,研究发现补体系统在CNV的发病机制中起重要作用.目的 构建针对补体因子B(CFB)的小干扰RNA(siRNA)重组质粒,体外观察其对人脐静脉内皮细胞ECV-304增生的影响.方法 根据人CFB的基因序列设计引物,经PCR扩增后与质粒pRNAT-U6.1连接,得到重组质粒pRNAT-U6.1/CFB siRNA,并进行测序鉴定和PCR鉴定.ECV-304细胞株进行常规培养,用电转染技术将重组质粒或空质粒分别转染人ECV-304细胞株,分为CFB-siRNA转染组和空质粒转染组,未转染的细胞为空白对照组.细胞转染后继续培养48 h,于倒置荧光显微镜下观察绿色荧光蛋白(GFP)的表达并计算转染效率;采用半定量逆转录PCR(RT-PCR)法测定各组细胞中CFB mRNA的相对表达量;用MTT法检测各组细胞转染24、48和72 h时细胞的增生值(A)并计算生长抑制率;利用流式细胞技术检测各组细胞的生长周期变化.结果 PCR扩增的目的片段序列与CFB基因序列完全相符,ECV-304细胞转染后,倒置荧光显微镜下可见CFB-siRNA转染组和空质粒转染组的细胞中GFP呈绿色荧光.半定量RT-PCR结果显示,CFB-siRNA转染组、空质粒转染组和空白对照组的CFB mRNA相对表达量分别为0.07±0.04、0.14±0.02和0.14 ±0.03,总体比较差异有统计学意义(F=233.05,P=0.00);其中CFB-siRNA转染组CFB mRNA相对表达量明显低于空质粒转染组和空白对照组,差异均有统计学意义(均P＜0.05).MTT法检测结果显示,各组不同时间细胞增生抑制率总体比较差异有统计学意义(F分组=212.99,P=0.00);CFB-siRNA转染组细胞转染24、48、72 h后细胞增生的抑制率分别为(23.45±0.01)％、(33.48±0.02)％和(45.49±0.01)％,明显高于同时间点空质粒转染组和空白对照组,差异均有统计学意义(均P＜0.05).流式细胞仪检测结果显示,CFB-siRNA转染组、空质粒转染组和空白对照组G1期的细胞数占总细胞数的(44.4±0.5)％、(25.8±0.4)％和(27.9±0.6)％,总体比较差异有统计学意义(F=58.98,P=0.00);CFB-siRNA转染组G1期和G2期细胞所占百分比显著高于空白对照组和空质粒转染组,差异均有统计学意义(均P＜0.05).结论 重组质粒pRNAT-U6.1/CFB siRNA可通过将细胞阻滞在G1期而有效抑制人脐静脉内皮细胞的增生.     

腹腔巨噬细胞核因子转位在小鼠内毒素诱导葡萄膜炎发病中的作用

目的观察野生型C3H/HeN小鼠和Toll样受体4(TLR4)基因缺陷型C3H/HeJ小鼠腹腔巨噬细胞在脂多糖(LPS)刺激下激活状态的差异,从体外途径探讨TLR4传导信号在前葡萄膜炎发病中的可能作用机制。设计实验研究。研究对象健康成年C3H/HeN和C3H/HeJ小鼠。方法常规提取两种小鼠腹腔巨噬细胞体外培养,按处理因素不同随机分六组:C3H/HeN小鼠LPS组、正常对照组、抗TLR4单克隆抗体加LPS组、抗TLR4单克隆抗体组;C3H/HeJ小鼠LPS组、正常对照组。各组在不同处理后1h、3h、6h、12h、24h五个时间点通过免疫荧光组织化学染色进行观察。主要指标TLR4传导途径中关键分子TLR4、髓样分化因子88(MyD88)、核因子-κB(NF-κB)免疫荧光强度及表达位置差异。结果所有组TLR4均表达于细胞膜,MyD88表达于细胞质和细胞核。C3H/HeN小鼠LPS组和C3H/HeJ小鼠LPS组TLR4荧光强度比较,差异有统计学意义;余各组TLR4荧光强度两两比较,差异无统计学意义。C3H/HeN小鼠腹腔巨噬细胞LPS组随LPS刺激时间的延长,各观察时间点MyD88荧光强度逐渐减弱,总体差异有统计学意义;余各组各时间点MyD88荧光强度无明显变化。C3H/HeN小鼠正常对照组、抗TLR4单克隆抗体组,C3H/HeJ小鼠正常对照组腹腔巨噬细胞各时间点NF-κB均表达于胞质,C3H/HeN小鼠腹腔巨噬细胞LPS刺激1h后NF-κB表达于胞核,3h依旧表达于胞核,此后无法检测到NF-κB的表达。C3H/HeN小鼠抗TLR4单克隆抗体加LPS组、C3H/HeJ小鼠LPS组巨噬细胞经LPS刺激1h后NF-κB转移至胞膜,直至24h一直表达于胞膜。结论腹腔巨噬细胞TLR4传导途径中核因子的转位可能与前葡萄膜炎的发病有关,特异性阻断该途径或许为前葡萄膜炎的治疗提供新思路。     

全喉切除发音重建术后喉咽部颈段食管影像学研究

目的:研究全喉切除发音重建术后患者咽部、食管在发音时的作用机制、影像学表现。方法:采用全喉切除发音重建术后1年以上随访病人,利用频闪动态喉镜、纤维喉镜、食道钡餐透视下观察发音时分流口及食管粘膜振动情况并照像记录。结果:发音时喉刚部及食管粘膜均振动,参与发音,发不同音时食管振动及扩张方式不一,但声源位于我们设计的分流口。结论:本发音方法设计合理,发音流畅,进食时无明显病人不能耐受之呛咳。     

Prior statin and short‐term outcomes of primary intracerebral hemorrhage: From a large‐scale nationwide longitudinal registry

IntroductionThe relationship between statins and intracerebral hemorrhage outcomes is unclear.AimWe aimed to compare the in‐hospital mortality and evacuation of intracranial hematoma rates in patients with primary intracerebral hemorrhage between prior statin users and nonusers.ResultsThe final study population included 66,263 patients. Multivariable logistics analyses showed that prior statin use was not associated with in‐hospital mortality for primary intracerebral hemorrhage (adjusted odd ratio 0.78, 95% CI 0.61–1.01), but reduced the proportion of patients undergoing evacuation of intracranial hematoma (adjusted odd ratio 0.70, 95% CI 0.61–0.82). Propensity score matching analyses yielded similar results.ConclusionPrior statin use was not associated with in‐hospital mortality but did reduce evacuation of intracranial hematoma rates.     

Melatonin inhibits EMT and PD‐L1 expression through the ERK1/2/FOSL1 pathway and regulates anti‐tumor immunity in HNSCC

Melatonin is an endogenous hormone with various biological functions and possesses anti‐tumor properties in multiple malignancies. Immune evasion is one of the most important hallmarks of head and neck squamous cell carcinoma (HNSCC) and is closely related to tumor progression. However, as an immune modulator under physiological conditions, the roles of melatonin in tumor immunity in HNSCC remains unclear. In this study, we found that the endogenous melatonin levels in patients with HNSCC were lower than those in patients with benign tumors in head and neck. Importantly, lower melatonin levels were related to lymph node metastasis among patients with HNSCC. Moreover, melatonin significantly suppressed programmed death‐ligand 1 (PD‐L1) expression and inhibited epithelial–mesenchymal transition (EMT) of HNSCC through the ERK1/2/FOSL1 pathway in vitro and in vivo. In SCC7/C3H syngeneic mouse models, anti‐programmed death‐1 (PD‐1) antibody combined with melatonin significantly inhibited tumor growth and modulated anti‐tumor immunity by increasing CD8⁺ T cell infiltration and decreasing the regulatory T cell (Treg) proportion in the tumor microenvironment. Taken together, melatonin inhibited EMT and downregulated PD‐L1 expression in HNSCC through the ERK1/2/FOSL1 pathway and exerted synergistic effects with anti‐PD‐1 antibody in vivo, which could provide promising strategies for HNSCC treatment.     

Transperineal pelvic drainage combined with lateral position to promote perineal wound healing after abdominoperineal resection: A prospective cohort trial

Background:For the rectal cancer <5 cm from anal margin, extralevator abdominoperineal resection (eAPR) has been accepted widely by surgeons. However, the rate of perineal infection following up eAPR is approximately 70%. We did the study with the aim of evaluating the effect and safety of transperineal pelvic drainage combined with lateral position (TPDLP) on perineal wound in patients undergoing eAPR.Methods:Patients were randomly assigned to N-TPDLP group (standard arm) or TPDLP group (intervention arm). In the standard arm, surgery was completed after abdominal drainage tube was placed in pelvic. Comparatively, an additional transperineal wound drainage tube was applied in the experimental arm. Postoperatively, patients of both 2 groups were informed not to sit to reduce perineal compression until the perineal wound healed. But lateral position was demanded in the intervention arm. The primary endpoint was the rate of uncomplicated perineal wound healing defined as a Southampton wound score of <2 at 30 days postoperatively. Patients were followed for 6 months.Results:In total, 60 patients were randomly assigned to standard arm (n = 31) and intervention arm (n = 29). The mean perineal wound healing time was 34.2 (standard deviation [SD] 10.9) days in TPDLP arm, which significantly differ from 56.4 (SD 34.1) in N-TPDLP arm (P = .001). At 30 days postoperatively, 3 (10%) of 29 patients undergoing TPDLP were classified into grade 4 according to Southampton wound score, however, 16 (52%) of 31 patients were classified into grade 4 in control arm, and significantly difference was observed between randomization groups (P = .001). What''s more, perineal wound pain was assessed at 30 days postoperatively, and it is discovered that the pain degree of patients in control arm was significantly more severe than the interventive arm (P = .015).Conclusion:In the present study, we found that TPDLP generated a favorable prognosis for perineal wounds with acceptable side-effects.     

胃肠道间质瘤CD117 CD34和Ki-67的表达与临床病理因素的相关性

目的:探讨胃肠道间质瘤(gastrointestinal stromal tumor,GIST)CD117、CD34和Ki-67的表达率和表达强度及其与临床病理相关因素和危险度的关系.方法:应用免疫组化S-P法检测57例GISTs的CD117、CD34和Ki-67的表达情况,并分析上述免疫组化指标在表达率方面与临床病理相关因素(性别、年龄、发生部位)的关系.并对这些病例进行危险度分级,进一步探讨上述免疫组化指标在表达程度方面与危险度之间的关系.结果:CD117和CD34的阳性表达率分别为84.2%、71.9%.CD34阳性表达率在不同性别、年龄组之间没有显著性差异,但在肠道外GISTs阳性表达率高于肠道(P<0.05).CD117的阳性表达强度和Ki-67增殖指数在不同的危险度之间存在显著性差异(P<0.05).且CD117阳性表达强度、Ki-67增殖指数和危险度的分级呈正相关.结论:CD117和CD34在GIST中均呈高表达率,但肠道GIST诊断时,CD117的阳性表达更准确、可靠.CD117表达强度和Ki-67增殖指数和危险度的分级呈正相关,可能是判断GIST危险度的可靠指标之一.