The potential of mesenchymal stem cells (MSCs) to differentiate into nonmesodermal cells such as pancreatic beta cells has been reported. New cell-based therapy using MSCs for diabetes mellitus is anticipated as an alternative treatment option to insulin injection or islet transplantation in both human and veterinary medicine. Several protocols were reported for differentiation of MSCs into insulin-producing cells (IPCs), but no studies have reported IPCs generated from canine MSCs. The purpose of this study was to generate IPCs from canine adipose tissue-derived MSCs (AT-MSCs) in vitro and to investigate the effects of IPC transplantation on diabetic mice in vivo. Culturing AT-MSCs with the differentiation protocol under a two-dimensional culture system did not produce IPCs. However, spheroid-like small clusters consisting of canine AT-MSCs and human recombinant peptide μ-pieces developed under a three-dimensional (3D) culture system were successfully differentiated into IPCs. The generated IPCs under 3D culture condition were stained with dithizone and anti-insulin antibody. Canine IPCs also showed gene expression typical for pancreatic beta cells and increased insulin secretion in response to glucose stimulation. The blood glucose levels in streptozotocin-induced diabetic mice were decreased after injection with the supernatant of canine IPCs, but the hyperglycemic states of diabetic mice were not improved after transplanting IPCs subcutaneously or intramesenterically. The histological examination showed that the transplanted small clusters of IPCs were successfully engrafted to the mice and included cells positive for insulin by immunofluorescence. Several factors, such as the transplanted cell number, the origin of AT-MSCs, and the differentiation protocol, were considered potential reasons for the inability to improve the hyperglycemic state after IPC transplantation. These findings suggest that canine AT-MSCs can be differentiated into IPCs under a 3D culture system and IPC transplantation may be a new treatment option for dogs with diabetes mellitus.

中文翻译：

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从犬脂肪组织间充质干细胞中产生胰岛素的细胞的生成

已经报道了间充质干细胞（MSC）分化为非中胚层细胞，例如胰腺β细胞的潜力。预期将新的基于MSC的基于细胞的疗法用于糖尿病，作为人和兽药中胰岛素注射或胰岛移植的替代治疗选择。据报道，有几种方案可将MSC分化为胰岛素产生细胞（IPC），但尚无研究报道由犬MSC产生的IPC。这项研究的目的是在体外从犬脂肪组织来源的MSC（AT-MSC）生成IPC，并研究IPC移植在体内对糖尿病小鼠的影响。在二维培养系统下使用分化方案培养AT-MSC不会产生IPC。然而，由犬AT-MSC和人重组肽μ组成的类球状小簇在三维（3D）培养系统下开发的生物碎片已成功地分化为IPC。用双硫zone和抗胰岛素抗体对在3D培养条件下生成的IPC进行染色。犬IPC还显示出胰腺β细胞特有的基因表达，并响应葡萄糖刺激而增加了胰岛素分泌。注射犬IPCs上清液后，链脲佐菌素诱导的糖尿病小鼠的血糖水平降低，但皮下或肠系膜内移植IPCs后，糖尿病小鼠的高血糖状态并未改善。组织学检查表明，已移植的IPC小簇已成功移植到小鼠体内，并通过免疫荧光检测了胰岛素阳性细胞。几个因素，例如移植的细胞数，AT-MSC的起源和分化方案被认为是IPC移植后无法改善高血糖状态的潜在原因。这些发现表明，犬AT-MSC可以在3D培养系统下分化为IPC，而IPC移植可能是糖尿病犬的一种新的治疗选择。