蒙古马骨髓间充质干细胞的分离培养及多向分化潜能的研究

旨在探索一个高效分离和扩增蒙古马骨髓间充质干细胞的方法,并通过形态学和多向分化能力进行鉴定.抽取蒙古马骨髓标本,利用密度梯度离心和差异贴壁法分离MSCs,体外扩增后传代,进行相差显微镜形态学观察.在地塞米松、IBMX、胰岛素及罗格列酮的作用下向脂肪细胞诱导分化,以及在地塞米松、VitC、β-磷酸甘油等作用下向成骨细胞诱导分化.结果显示原代和传代细胞呈梭形成纤维细胞样外观,生长增殖能力良好.经定向诱导分化后,细胞分别呈现脂肪细胞、成骨细胞的表型特征.证明所纯化的细胞为尚未分化的基质细胞,而不是组织的成体细胞,它具有多向分化潜能.     

同源异体骨髓间充质干细胞移植治疗犬急性肝功能损伤

探讨同源异体骨髓间充质干细胞(BMMSCs)移植对犬急性肝功能损伤的治疗作用,为犬干细胞治疗相关疾病提供理论基础和技术支持。对患急性肝损伤的10只病犬经超声引导腹腔内肝门附近移植BMMSCs,移植后检测血常规、肝功能和肝脏B超,观察同期的症状体征和不良反应情况。结果显示,干细胞移植7天后可使患犬各项肝功能指标明显改善,10d后B超结果显示肝组织回声均匀,无明显异常,14d后患犬的精神、体力、食欲明显好转,干细胞移植21d后血常规各项指标恢复正常,患犬基本康复,移植干细胞的患犬均未发现有不良反应。说明相对于常规治疗,犬BMMSCs同源异体移植治疗急性肝损伤可明显缩短治疗时间,显著提高患犬生活质量。     

Production and Identification of Clone Sheep Using Bone Marrow Mesenchymal Stem Cells as Donor Cell

In order to obtain the cloned sheep using bone marrow mesenchymal stem cells（BMSCs）as the donor cells,the BMSCs of sheep were chosen and reconstructed embryos were built to transfer.10 published microsatellite markers were chosen,and the DNA samples from clone sheep,donor cells and surrogate ewes were amplified,and the relationship of father-son（RCP）was analyzed using the Quantity One for genotyping.The results showed that the reconstructed embryos were successfully built for electric fusion using sheep BMSCs as nuclear donor,and making nuclear transplantation into enucleated mature oocytes of which the fusion rate was 80.62%.20 surrogate ewe were chosen to be implanted with the reconstructed embryos at morula stage by implant surgery,and 5 lambs were born and only 3 were survived.The genotype of cloned sheep was in line with the dornor cell and the RCP were more than 99.999%.In conclusion,the first clone sheep were obtained successfully by using BMSCs as a nuclear donor in this experiment.     

骨髓间充质干细胞为核供体的克隆绵羊的生产与鉴定

本研究旨在采用骨髓间充质干细胞（BMSCs）为核供体进行体细胞核移植,以获得成体干细胞克隆绵羊。选择幼体绵羊BMSCs为核供体,构建重构胚进行克隆胚胎移植,克隆绵羊诞生后,选取IASG上公布的绵羊10个微卫星标记位点,PCR扩增克隆羊、供体细胞及代孕母羊微卫星DNA,利用Quantity One软件进行基因分型,计算供体细胞与克隆羊之间的父子关系相对机会（RCP）值。结果显示,以幼体绵羊BMSCs为核供体,与去核成熟卵母细胞进行核移植,构建重构胚进行电融合,其融合率达80.62%,重构胚发育至桑椹胚阶段,经手术植入20只代孕母羊,获得克隆绵羊5只,成活3只。克隆羊基因型与供体细胞一致,供体细胞与克隆羊之间的RCP≥99.999%,证明克隆羊来自幼体绵羊BMSCs。本试验最终成功获得了首例以BMSCs为核供体的克隆绵羊。     

Willingness of a Convenience Sample of Horse Owners to Use Stem Cell Therapy for the Treatment of Equine Tendon Injury and Factors that Influence This Decision

Stem cell therapies offer huge potential for the treatment of tendon injury in horses. However, the factors influencing clients' willingness to use these therapies have not yet been investigated. This study examines the willingness of a population of horse owners to choose stem cell therapy for the treatment of equine tendon injuries and the factors that may influence this choice including treatment cost and efficacy, veterinarian advice, client knowledge, and success stories. A short, anonymous questionnaire was delivered to a convenience sample of equine owners. Questions related to (1) respondent background; (2) prior experience with tendon injuries and their treatment; (3) willingness to use equine stem cell therapy; and (4) the importance of factors that may affect their decision to use the therapy. Responses were summarized using graphical methods and compared using Fisher exact test where appropriate. Fifty-four percent of our cohort (n = 113) reported they would be willing to use equine stem cell therapy, 6% were not willing, and 40% were unsure. When asked to rank factors that would influence their decision, scientific proof that the therapy works was ranked highest followed by veterinarian advice. This result may have been influenced by the high proportion of tertiary-educated respondents recruited in this study. This is the first reported study to investigate the willingness of a population of horse owners to use stem cell therapy for their horses. Results suggest that the majority of these equine owners may consider using stem cell therapies for their horses given a larger positive evidence base was provided for the therapy.     

Long‐Term Management with Adipose Tissue‐Derived Mesenchymal Stem Cells and Conventional Treatment in a Dog with Hepatocutaneous Syndrome

Hepatocutaneous syndrome (HS) is an uncommon skin disorder that occurs in conjunction with liver disease and is diagnosed based on decreased plasma concentrations of amino acids and the histopathology of skin lesions. The survival period generally is <6 months. A 10‐year‐old castrated male Maltese dog was presented for evaluation of lethargy, polyuria, polydipsia, and skin lesions including alopecia, erythema, and crusts. Based on increased liver enzyme activity, low plasma amino acid concentrations, and findings from liver cytology and skin biopsy, the dog was diagnosed with HS. In addition to administration of antioxidants, hepatoprotective agents, and amino acids IV, allogenic adipose tissue‐derived mesenchymal stem cells were infused 46 times over a 30‐month period: 8 times directly into the liver parenchyma guided by ultrasonography and the remainder of the times into peripheral veins. After commencing stem cell therapy, the dog's hair re‐grew and the skin lesions disappeared or became smaller. During ongoing management, the patient suddenly presented with anorexia and uncontrolled vomiting, and severe azotemia was observed. The dog died despite intensive care. On necropsy, severe liver fibrosis and superficial necrolytic dermatitis were observed. The dog survived for 32 months after diagnosis. A combination of amino acid and stem cell therapy may be beneficial for patients with HS.