典型案例联合实践教学在儿科临床教学中的应用研究

目的　探讨典型案例联合实践教学在儿科临床教学中的应用。方法　选取济宁医学院附属医院2019年9月至2020年1月儿科专业临床实习生80名作为研究对象,按照随机数字表法将其分为观察组（n=41）与对照组（n=39）。对照组采用常规带教,观察组采用典型案例联合实践教学模式。培训16周后,比较两组学生理论知识成绩、实践操作[直接观察操作技巧（direct observation of procedural skill,DOPS）评估、迷你临床评估演练（mini-clinical evaluation exercise,Mini-CEX）]评分与总体成绩。采用SPSS 19.0进行t检验、卡方检验或秩和检验。结果　实习后,两组学生理论成绩中儿科常见疾病、疾病临床特点、发病原理、治疗方案、并发症均有提高,且观察组均高于对照组（P<0.05）;实习后,两组DOPS评分、Mini-CEX评分均上升,且观察组高于对照组（P<0.05）;实习后,观察组总体成绩优秀率为87.80%（36/41）,高于对照组的69.23%（27/39）;调查发现,观察组认为教学方案可以加深知识掌握程度、提高实习兴趣、提高综合能力、提高实习效率、提高临床思维能力的人数多于对照组。结论　典型案例联合实践教学能够有效加深实习生对临床儿科理论知识的理解,提高实践操作能力,激发学习兴趣。     

Pig-to-baboon lung xenotransplantation: Extended survival with targeted genetic modifications and pharmacologic treatments

Galactosyl transferase knock-out pig lungs fail rapidly in baboons. Based on previously identified lung xenograft injury mechanisms, additional expression of human complement and coagulation pathway regulatory proteins, anti-inflammatory enzymes and self-recognition receptors, and knock-down of the β4Gal xenoantigen were tested in various combinations. Transient life-supporting GalTKO.hCD46 lung function was consistently observed in association with either hEPCR (n = 15), hTBM (n = 4), or hEPCR.hTFPI (n = 11), but the loss of vascular barrier function in the xenograft and systemic inflammation in the recipient typically occurred within 24 h. Co-expression of hEPCR and hTBM (n = 11) and additionally blocking multiple pro-inflammatory innate and adaptive immune mechanisms was more consistently associated with survival >1 day, with one recipient surviving for 31 days. Combining targeted genetic modifications to the lung xenograft with selective innate and adaptive immune suppression enables prolonged initial life-supporting lung function and extends lung xenograft recipient survival, and illustrates residual barriers and candidate treatment strategies that may enable the clinical application of other organ xenografts.     

两种不同药物对骨质疏松性椎体骨折术后老年患者疗效的影响

目的研究丹参注射液联合仙灵骨葆胶囊对骨质疏松性椎体骨折术后老年患者血清胰岛素生长因子-1(insulin growth factor-1,IGF-1)及碱性成纤维细胞生长因子(basic fibroblast growth factor,b FGF)水平的影响。方法选取我院于2017年6月至2018年10月收治的123例老年骨质疏松性椎体骨折患者,依据随机数字表法分为观察(62例)和对照(61例)两组。两组患者均行经皮椎体成形术(PVP)治疗,对照组患者服用仙灵骨葆胶囊(持续用药3个月),观察组患者在对照组基础上使用丹参注射液(持续用药1周)。比较两组患者的治疗效果、骨折愈合情况,以及VAS评分和血清b FGF及IGF-1在不同时间点水平变化,统计两组并发症发生情况。结果观察组患者治疗有效率(95.00%)显著高于对照组(83.33%),差异有统计学意义(P0.05)。治疗后6个月两组患者骨密度显著增加,而椎体后凸Cobb角显著降低,且观察组患者改善情况显著优于对照组(P0.05)。两组患者经治疗后VAS评分显著降低,血清b FGF、IGF-1水平显著增加,其中观察组患者VAS评分、血清b FGF、IGF-1水平在治疗2周、4周、6周的改善程度均优于对照组(P0.05)。两组患者并发症发生率的差异无统计学意义(P0.05)。结论丹参注射液联合仙灵骨葆胶囊可增强PVP对老年骨质疏松性椎体骨折患者的疗效,可通过提高血清b FGF及IGF-1水平促进骨折修复,从而加快骨折愈合,降低疼痛。     

碱性成纤维细胞生长因子在富集结直肠癌肿瘤干细胞中的作用

目的 研究不同浓度碱性成纤维细胞生长因子（bFGF）作用于结直肠癌细胞不同时间,富集肿 瘤干细胞的效果。方法? 结直肠癌DLD-1细胞分别在含5 ng/mL、10 ng/mL、20 ng/mL bFGF的无血清培 养基中悬浮培养,分为G1、G2、G3组,设置培养时间梯度为10 d、20 d、30 d,获得球细胞。采用流式 细胞术（FCM）检测细胞球中的CD44+、CD133+及CD44+CD133+双阳性的细胞表达比例,Real-time PCR 检测球细胞中的干细胞基因（KLF4、Nanog）;上皮间质转化基因（E-cadherin、Snail）;Wnt/β-catenine通路 基因（Wnt-3a）的 mRNA表达情况,成球实验检测细胞球的自我更新能力。结果 FCM检测结果：CD44+ 阳性表达的细胞表达以G2组20 d最高,CD133+及CD44+CD133+双阳性的细胞表达均以G2组20 d及G3 组10 d最高,差异有统计学意义（F=98.895、147.641、13.321,P<0.05）。 Real-time PCR检测结果：各组中 KLF4、Nanog、Snail以及Wnt-3a mRNA的表达均以G2组20 d表达最高（F=2.424、7.694、2.951、3.771, 均P<0.05）, E-cadherin基因G2组20 d mRNA表达最低（G2组30 d、G1组10 d除外）（F=10.620,P<0.05）。 成球实验结果：各组比较G1组20 d和G2组20 d成球数量明显多于其他组,差异具有统计学意义（F=14.279, P<0.01）;但 G1组20 d和G2组20 d比较,无统计学差异（t=0.605,P=0.578）。 结论 碱性成纤维细胞生长 因子无血清悬浮培养能够有效富集肿瘤干细胞,其中G2组培养20 d较其余组能更有效地富集结直肠癌肿 瘤干细胞。     

中西医治疗糖尿病周围神经病变研究进展

对于糖尿病周围神经病变的治疗,中医以"络以通为用"为治疗原则,在改善临床症状,阻止DPN进一步恶化,减少截肢率方面,都有很好的疗效。西医可以有效控制血糖、血压,弥补了中药不能快速降糖降压之不足,对防止DPN的发生和发展起了重要作用。中西医结合治疗具有一定互补优势,疗效确切,但其作用机制尚不明确,需进一步加强实验研究,明确中医药作用机理,提高临床疗效。     

Extracellular nucleotide signaling in solid organ transplantation

The role of extracellular purine nucleotides, including adenosine triphosphate (ATP) and adenosine, as modulators of posttransplantation outcome and ischemia‐reperfusion injury is becoming increasingly evident. Upon pathological release of ATP, binding and activation of P2 purinergic surface receptors promote tissue injury and inflammation, while the expression and activation of P1 receptors for adenosine have been shown to attenuate inflammation and limit ischemia‐induced damage, which are central to the viability and long‐term success of allografts. Here we review the current state of the transplant field with respect to the role of extracellular nucleotide signaling, with a focus on the sources and functions of extracellular ATP. The connection between ischemia reperfusion, purinergic signaling, and graft preservation, as well as the role of ATP and adenosine as driving factors in the promotion and suppression of posttransplant inflammation and allograft rejection, are discussed. We also examine novel therapeutic approaches that take advantage of the ischemia‐reperfusion‐responsive and immunomodulatory roles for purinergic signaling with the goal of enhancing graft viability, attenuating posttransplant inflammation, and minimizing complications including rejection, graft failure, and associated comorbidities.     

Recombinant human ADAMTS13 treatment and anti‐NET strategies enhance skin allograft survival in mice

Enhancing skin allograft longevity lessens the need for new allografts before optimal intervention is available. Reduced activity of ADAMTS13 (an enzyme that cleaves the pro‐thrombotic and proinflammatory von Willebrand factor) and presence of neutrophil extracellular traps (NETs) have been implicated in liver and lung allograft failures. The effect of ADAMTS13 treatment and the impact of NETs on skin allografts, however, remain unexplored. Here, we adopted a murine model of complete mismatch full‐thickness skin transplant by grafting dorsal skin from BALB/c mice to C57BL/6J background mice. Recombinant human ADAMTS13 (rhADAMTS13) treatment of graft recipients increased allograft survival. Western blot and immunofluorescence microscopy revealed the presence of NETs in allografts of vehicle, but surprisingly, not in rhADAMTS13‐treated mice, 3 days after surgery. Recapitulating the observations in mice, NETs were also observed in all the examined allografts from burn patients. Intriguingly, knocking out peptidylarginine deiminase 4 (PAD4, a key enzyme for NET formation) or DNase 1 treatment (which cleaves NETs) also prolonged allograft survival. In summary, rhADAMTS13 lessens inflammation in allografts by reducing NET burden, resulting in enhanced allograft survival. RhADAMTS13 and anti‐NET treatments could be new therapeutic strategies to promote skin allograft longevity and, hence, the survival of patients with severe burns.     

Extracellular vesicles derived from injured vascular tissue promote the formation of tertiary lymphoid structures in vascular allografts

Tertiary lymphoid structures (TLS) accumulate at sites of chronic injury where they function as an ectopic germinal center, fostering local autoimmune responses. Vascular injury leads to the release of endothelial‐derived apoptotic exosome‐like vesicles (ApoExo) that contribute to rejection in transplanted organs. The purpose of the study was to evaluate the impact of ApoExo on TLS formation in a model of vascular allograft rejection. Mice transplanted with an allogeneic aortic transplant were injected with ApoExo. The formation of TLS was significantly increased by ApoExo injection along with vascular remodeling and increased levels of antinuclear antibodies and anti‐perlecan/LG3 autoantibodies. ApoExo also enhanced allograft infiltration by γδT17 cells. Recipients deficient in γδT cells showed reduced TLS formation and lower autoantibodies levels following ApoExo injection. ApoExo are characterized by proteasome activity, which can be blocked by bortezomib. Bortezomib treated ApoExo reduced the recruitment of γδT17 cells to the allograft, lowered TLS formation, and reduced autoantibody production. This study identifies vascular injury‐derived extracellular vesicles (ApoExo), as initiators of TLS formation and demonstrates the pivotal role of γδT17 in coordinating TLS formation and autoantibody production. Finally, our results suggest proteasome inhibition with bortezomib as a potential option for controlling TLS formation in rejected allografts.     

Triple (GGTA1, CMAH,B2M) modified pigs expressing an SLA class Ilow phenotype—Effects on immune status and susceptibility to human immune responses

Porcine xenografts lacking swine leukocyte antigen (SLA) class I are thought to be protected from human T cell responses. We have previously shown that SLA class I deficiency can be achieved in pigs by CRISPR/Cas9‐mediated deletion of β₂‐microglobulin (B2M). Here, we characterized another line of genetically modified pigs in which targeting of the B2M locus did not result in complete absence of B2M and SLA class I but rather in significantly reduced expression levels of both molecules. Residual SLA class I was functionally inert, because no proper differentiation of the CD8⁺ T cell subset was observed in B2M^low pigs. Cells from B2M^low pigs were less capable in triggering proliferation of human peripheral blood mononuclear cells in vitro, which was mainly due to the nonresponsiveness of CD8⁺ T cells. Nevertheless, cytotoxic effector cells developing from unaffected cell populations (eg, CD4⁺ T cells, natural killer cells) lysed targets from both SLA class I⁺ wildtype and SLA class I^low pigs with similar efficiency. These data indicate that the absence of SLA class I is an effective approach to prevent the activation of human CD8⁺ T cells during the induction phase of an anti‐xenograft response. However, cytotoxic activity of cells during the effector phase cannot be controlled by this approach.