枯草芽孢杆菌guaB基因克隆及其在大肠杆菌中的表达

克隆枯草芽孢杆菌guaB基因,构建穿梭表达载体,使其在大肠杆菌中获得表达,为构建鸟苷高产工程菌奠定基础。从枯草芽孢杆菌JSIM-G-518基因组扩增出guaB基因,将其连接到pMD19-T上,得到重组质粒pMD-guaB,进行测序和Blast比对分析。该重组质粒经PstI、BamHI双酶切,获得guaB片段连接至pHP13载体上,构建穿梭表达载体pHP13-guaB,并采用双酶切电泳进行鉴定。将该载体转化入大肠杆菌BL21中,通过SDS-PAGE电泳检测guaB基因的表达效果。克隆的guaB基因长度为1 510 bp,与GenBank登录的枯草芽孢杆菌同源性为99%,编码500个氨基酸,确认为控制IMP脱氢酶的基因。穿梭载体pHP13-guaB经双酶切后产生1 510 bp和4.9 kb两个期望的目的条带。SDS-PAGE结果显示表达产物相对分子质量的大小为52.9 k,与预期的IMP脱氢酶分子量一致,且该目的蛋白的表达效果比对照组更明显。论文成功实现了枯草芽孢杆菌guaB基因在大肠杆菌中的表达。     

Detection of lymphatic invasion in resected cases of primary pancreatic cancer based on immunohistochemistry of D2-40

We investigated the presence of lymphatic invasion detected by D2-40 immunostaining compared to conventional hematoxylin-eosin (HE) staining in primary pancreatic cancer. We also compared the alkaline phosphatase-fast red detection method with the 3,3′-diaminobenzidine (DAB) detection method. We reviewed 25 cases of pancreatic cancer with lymph node (LN) metastases and 15 cases without LN metastases and evaluated the detection rate of lymphatic invasion by HE stain slide alone and D2-40 immunostains. Regarding the cases with positive LN metastasis, 4 (16%) of the 25 cases showed lymphatic invasion by HE slide alone, whereas 7 cases (28%) demonstrated positive lymphatic invasion by D2-40 immunostain. On the other hand, even in cases with negative LN metastasis, 3 of the 15 cases revealed lymphatic invasion by D2-40 immunostaining. Lymphatic invasion was easily detected by alkaline phosphatase-fast red technique, especially at the lower magnification. Regarding the location of lymphatic invasion, it was recognized not only at the peripheral portion but also in the central part of the tumors by D2-40 immunostains; this was difficult to identify by HE stain slide alone. Our study indicates that lymphatic invasion may be overlooked when only HE stain slides are used. In addition, the alkaline phosphatase-fast red detection method (vivid red color) is a distinctive advantage compared with the DAB detection method (brown color), especially in detecting lymphatic invasion at the lower magnification.     

Can cost utility evaluations inform decision making about interventions for low back pain?

Background contextLow back pain (LBP) is associated with high health-care utilization and lost productivity. Numerous interventions are routinely used, although few are supported by strong evidence. Cost utility analyses (CUAs) may be helpful to inform decision makers.PurposeTo conduct a systematic review of CUAs of interventions for LBP.Study designSystematic review.MethodsA search strategy combining medical subject headings and free text related to LBP and health economic evaluations was executed in MEDLINE. Cost utility analyses combined with randomized controlled trials for LBP were included. Studies that were published before 1998, non-English, decision analyses, and duplicate reports were excluded. Search results were evaluated by two reviewers, who extracted data independently related to clinical study design, economic study design, direct cost components, utility results, cost results, and CUA results.ResultsThe search produced 319 citations, and of these 15 met eligibility criteria. Most were from the United Kingdom (n=8), published in the past 3 years (n=12), studied chronic LBP or radiculopathy (n=13), and had a follow-up >12 months (n=13). Combined, there were 33 study groups who received a mean 2.1 interventions, most commonly education (n=17), exercise therapy (n=13), spinal manipulation therapy (n=7), surgery (n=7), and usual care from a general practitioner (n=7). Mean baseline utility was 0.57, improving to 0.67 at follow-up; the mean difference in utility improvement between study groups was 0.04. Based on available data and converted to US dollars, the cost per quality-adjusted life year ranged from $304 to $579,527, with a median of $13,015.ConclusionsFew CUAs were identified for LBP, and there was heterogeneity in the interventions compared, direct cost components measured, indirect costs, other methods, and results. Reporting quality was mixed. Currently published CUAs do not provide sufficient information to assist decision makers. Future CUAs should attempt to measure all known direct cost components relevant to LBP, estimate indirect costs such as lost productivity, have a follow-up period sufficient to capture meaningful changes, and clearly report methods and results to facilitate interpretation and comparison.     

骨髓基质细胞培养基对体外脊髓运动神经元的影响

目的:体外检测骨髓基质细胞(bone marrow stromal cells,BMSCs)的条件培养基对脊髓运动神经元的影响。方法:体外培养SD大鼠的BMSCs,并收集其条件培养基。实验组脊髓运动神经元用条件培养基培养,对照组用NB培养液。观察两组运动神经元形态学指标,测量细胞突起长度。MTT法检测两组细胞活力。结果:实验组运动神经元的细胞活力、突起长度与对照组相比差异均有统计学意义(P<0.01)。结论:BMSCs合成和分泌神经营养活性物质对脊髓运动神经元有维持存活和促进突起生长的作用。     

The effects of heparin releasing hydrogels on vascular smooth muscle cell phenotype

Poly(ethylene glycol) diacrylate (PEGDA) hydrogel scaffolds were engineered to promote contractile smooth muscle cell (SMC) phenotype via controlled release of heparin. The scaffold design was evaluated by quantifying the effects of free heparin on SMC phenotype, engineering hydrogels to provide controlled release of heparin, and synthesizing cell-adhesive, heparin releasing hydrogels to promote contractile SMC phenotype. Heparin inhibited SMC proliferation and up-regulated expression of contractile SMC phenotype markers, including smooth muscle α-actin, calponin, and SM-22α, in a dose-dependent fashion (6 μg/ml to 3.2 mg/ml). Heparin release from PEGDA hydrogels was controlled by altering PEGDA molecular weight (MW 1000–6000) and concentration at polymerization (10–30% w/w), yielding release profiles ranging from hours to weeks in duration. Heparin released from PEGDA gels, formulated for optimized heparin loading and release kinetics (30% w/w PEGDA, MW 3000), stimulated SMCs to up-regulate contractile marker mRNA. A cell-instructive scaffold construct was prepared by polymerizing a thin hydrogel film, with pendant RGD peptides for cell attachment, over the optimized hydrogel depots. SMCs seeded on these constructs had elevated levels of contractile marker mRNA after 3 d of culture compared with SMCs on control constructs. These results indicate that RGD-modified, heparin releasing PEGDA gels can act as cell-instructive scaffolds that promote contractile SMC phenotype.     

Asystole due to trigemino-cardiac reflex: A rare complication of trans-sphenoidal surgery for pituitary adenoma

The trigemino-cardiac reflex (TCR) is a well-known reflexive response in which bradycardia, hypotension, and gastric hypermotility are induced by stimulation of a peripheral or central portion of the trigeminal nerve. This reflex occurs during craniofacial surgery and other operations on or near the cerebellopontine angle, petrosal sinus, orbit and trigeminal ganglion. TCR is a well-known, although not well documented, phenomenon commonly observed during trans-sphenoidal surgery for resection of pituitary adenomas. We report a case in which asystole occurred during trans-sphenoidal surgery on a pituitary adenoma that was infiltrating the right cavernous sinus. When the anesthesiologist reported asystole, the team stopped manipulation and administered intravenous atropine. Intra-operative MRI showed a small tumour remnant in the right cavernous sinus. The operation was terminated but subsequent radiosurgery was planned for the residual tumor. Although TCR is rare and usually self-limiting, this case led us to change our treatment strategy. Surgeons who perform trans-sphenoidal surgery should be aware of this potential problem. Invasive pituitary adenomas should be removed gently and the risk of triggering TCR should be kept in mind.     

口腔鳞癌腮腺淋巴结转移的临床病理特征与预后分析

目的:探讨口腔鳞癌患者腮腺淋巴结转移的临床病理特征及预后影响因素.方法:回顾性分析2003年1月—2017年12月上海交通大学医学院附属第九人民医院收治的60例伴或不伴腮腺淋巴结转移的口腔鳞癌患者的临床病理资料及影像学资料,分析其临床病理和影像学特征及其与预后的相关性.采用SPSS 23.0软件包对数据进行统计学分析....     

Dysregulation of circulating CD4 + CXCR5 + PD-1+ T cells in diabetic retinopathy

AimsWe aimed to determine an association between follicular helper T (Tfh) cells and Bcl-6 and CXCL13 levels and determine the role of Tfh cells, Bcl-6, and CXCL13 serum levels in the pathogenesis of diabetic retinopathy (DR) since Tfh cells have an important role in type 1 diabetes; however, their role in type 2 diabetes-related DR requires exploration.MethodsBlood samples were collected from 24 patients with non-proliferative diabetic retinopathy (NPDR), 20 with proliferative diabetic retinopathy (PDR), and 18 age- and sex-matched healthy volunteers. Flow cytometry detected CD4 + CXCR5 + PD1+ Tfh cells. Serum Bcl-6 and CXCL13 levels were determined using enzyme-linked immunosorbent assay.ResultsCD4 + CXCR5 + PD-1+ Tfh cell percentages in peripheral blood and serum levels of Bcl-6 and CXCL13 in the non-proliferative DR (NPDR) and proliferative DR (PDR) groups' were significantly higher than those in healthy individuals. The proportion of Tfh cells in DR patients' peripheral blood positively correlated with Bcl-6 and CXCL13 serum levels, DR course severity, Fasting blood glucose, glycosylated hemoglobin and body mass index.ConclusionsThe increased circulating Tfh cells, serum Bcl-6 levels, and CXCL13 levels of DR patients with type 2 diabetes suggested that circulating Tfh cells and the germinal center response may have a role in the occurrence and development of DR.     

Associations of apolipoproteinA1, high density lipoprotein cholesterol with hemoglobin glycation index and triglyceride-glucose index in Chinese adults with coronary artery disease

AimsScarce data explored the associations of apolipoproteins with hemoglobin glycation index (HGI) and triglyceride-glucose (TyG) index. This study determined associations of serum apolipoproteinA1 (ApoA1) and high density lipoprotein cholesterol (HDL-C) with HGI and TyG index in coronary artery disease (CAD) patients.MethodsA total of 10,803 CAD patients were included in this cross-sectional pilot study. Serum concentrations of ApoA1 and HDL-C were measured. Analyses of covariance were used to compare the mean differences in glucose metabolism indices (e.g., HGI, TyG index, hemoglobin glycation [HbA1c], fasting blood glucose [FBG]) among the quartiles of ApoA1, HDL-C and HDL-C/ApoA1 ratio.ResultsIn multivariate analysis, higher ApoA1, HDL-C and HDL-C/ApoA1 ratio were associated with significantly lower HGI (Quartile [Q]4 vs. Q1: −0.032 % vs. 0.017 % for ApoA1; −0.072 % vs. 0.079 % for HDL-C; −0.083 % vs. 0.085 % for HDL-C/ApoA1 ratio). Intermediate ApoA1 level was inversely associated with TyG index (Q2 vs. Q1: 296.278 vs. 306.794). The mean TyG index were significantly decreased with increased HDL-C and HDL-C/ApoA1 ratio (Q4 vs. Q1: 298.584 vs. 309.221 for HDL-C; 300.405 vs. 315.218 for HDL-C/ApoA1 ratio). Moreover, the inverse associations of ApoA1, HDL-C and HDL-C/ApoA1 ratio with HbA1c and FBG also were observed. In path analysis, the associations of HDL-C and HDL-C/ApoA1 ratio with TyG index were mediated by obesity.ConclusionThis study provided further support for the hypoglycemic effects of ApoA1 and HDL-C in patients with CAD. Replication of these findings is warranted in further longitudinal studies in different populations.     

Inflammation biomarkers and inflammatory genes expression in metabolically healthy obese patients

Background and aimsObesity without metabolic alterations (Metabolically Healthy Obesity, MHO) is a condition with a risk of death and cardiovascular disease lower than that of obesity associated with metabolic alterations (Metabolically Unhealthy Obesity, MUO) and similar to that of healthy non obese individuals. Inflammation is considered as a key risk factor mediating the adverse health outcomes in obesity.Methods and resultsWe compared circulating levels of thirteen major cytokines and adipokines and the expression profiles of fifteen pro-inflammatory and two anti-inflammatory genes in visceral and subcutaneous adipose tissue in a series of 16 MHO patients and in 32 MUO patients that underwent bariatric surgery. MHO was defined according to the most applied definition in current literature.Serum levels of a large set of major cytokines and adipokines did not differ between MHO and MUO patients (p ≥ 0.15). Analyses of the expression profile of pro-inflammatory and anti-inflammatory genes in subcutaneous and visceral adipose tissue failed to show differences between MHO and MUO patients (p ≥ 0.07). Sensitivity analyses applying two additional definitions of MHO confirmed the results of the primary analysis.ConclusionIn a series of metabolically healthy obese patients neither circulating levels of major cytokines and adipokines nor the gene expression profile of a large set of pro-inflammatory and anti-inflammatory genes in subcutaneous and visceral fat differed from those in metabolically unhealthy obese patients.