Department of Translational Research and Cellular Therapeutics, Diabetes and Metabolism Research Institute, Beckman Research Institute of City of Hope, 1500 E. Duarte Rd, Duarte, CA, 91010, USA.
Department of Diabetes Immunology, Diabetes and Metabolism Research Institute, Beckman Research Institute of City of Hope, Duarte, CA, USA.
Stem Cell Res Ther. 2019 Nov 15;10(1):322. doi: 10.1186/s13287-019-1435-2.
Human pancreata contain many types of cells, such as endocrine islets, acinar, ductal, fat, and mesenchymal stromal cells (MSCs). MSCs are important and shown to have a promising therapeutic potential to treat various disease conditions.
We investigated intra-pancreatic tissue-derived (IPTD) MSCs isolated from tissue fractions that are routinely discarded during pancreatic islet isolation of human cadaveric donors. Furthermore, whether pro-angiogenic and anti-inflammatory properties of these cells could be enhanced was investigated.
IPTD-MSCs were expanded in GMP-compatible CMRL-1066 medium supplemented with 5% human platelet lysate (hPL). IPTD-MSCs were found to be highly pure, with > 95% positive for CD90, CD105, and CD73, and negative for CD45, CD34, CD14, and HLA-DR. Immunofluorescence staining of pancreas tissue demonstrated the presence of CD105 cells in the vicinity of islets. IPTD-MSCs were capable of differentiation into adipocytes, chondrocytes, and osteoblasts in vitro, underscoring their multipotent features. When these cells were cultured in the presence of a low dose of TNF-α, gene expression of tumor necrosis factor alpha-stimulated gene-6 (TSG-6) was significantly increased, compared to control. In contrast, treating cells with dimethyloxallyl glycine (DMOG) (a prolyl 4-hydroxylase inhibitor) enhanced mRNA levels of nuclear factor erythroid 2-related factor 2 (NRF2) and vascular endothelial growth factor (VEGF). Interestingly, a combination of TNF-α and DMOG stimulated the optimal expression of all three genes in IPTD-MSCs. Conditioned medium of IPTD-MSCs treated with a combination of DMOG and TNF-α contained higher levels of pro-angiogenic (VEGF, IL-6, and IL-8) compared to controls, promoting angiogenesis of human endothelial cells in vitro. In contrast, levels of MCP-1, a pro-inflammatory cytokine, were reduced in the conditioned medium of IPTD-MSCs treated with a combination of DMOG and TNF-α.
The results demonstrate that IPTD-MSCs reside within the pancreas and can be separated as part of a standard islet-isolation protocol. These IPTD-MSCs can be expanded and potentiated ex vivo to enhance their anti-inflammatory and pro-angiogenic profiles. The fact that IPTD-MSCs are generated in a GMP-compatible procedure implicates a direct clinical application.
人类胰腺包含多种细胞类型，如内分泌胰岛、腺泡、导管、脂肪和间充质基质细胞（MSCs）。MSCs 非常重要，并且显示出在治疗各种疾病方面有很大的应用潜力。
我们研究了从常规胰岛分离过程中丢弃的人尸体供体胰腺组织部分中分离出的胰腺内组织衍生（IPTD）MSCs。此外，还研究了这些细胞的促血管生成和抗炎特性是否可以增强。
IPTD-MSCs 在符合 GMP 的 CMRL-1066 培养基中扩增，该培养基中添加了 5%的人血小板裂解物（hPL）。结果发现 IPTD-MSCs 高度纯净，＞95%为 CD90、CD105 和 CD73 阳性，而 CD45、CD34、CD14 和 HLA-DR 阴性。对胰腺组织的免疫荧光染色显示，胰岛附近存在 CD105 细胞。IPTD-MSCs 能够在体外分化为脂肪细胞、软骨细胞和成骨细胞，突出了其多能性特征。当这些细胞在低剂量 TNF-α存在的情况下培养时，与对照组相比，肿瘤坏死因子-α刺激基因-6（TSG-6）的基因表达显著增加。相比之下，用二甲氧乙醯甘氨酸（DMOG）（脯氨酰 4-羟化酶抑制剂）处理细胞可增强核因子红细胞 2 相关因子 2（NRF2）和血管内皮生长因子（VEGF）的 mRNA 水平。有趣的是，TNF-α和 DMOG 的组合刺激了 IPTD-MSCs 中所有三种基因的最佳表达。用 DMOG 和 TNF-α 组合处理的 IPTD-MSCs 的条件培养基中含有更高水平的促血管生成（VEGF、IL-6 和 IL-8），与对照组相比，可促进人内皮细胞的血管生成。相反，用 DMOG 和 TNF-α 组合处理的 IPTD-MSCs 的条件培养基中，促炎细胞因子 MCP-1 的水平降低。
结果表明，IPTD-MSCs 存在于胰腺中，并且可以作为标准胰岛分离方案的一部分进行分离。这些 IPTD-MSCs 可以在体外进行扩增和增强，以增强其抗炎和促血管生成特性。事实是，IPTD-MSCs 是在符合 GMP 的程序中产生的，这暗示了直接的临床应用。
