表皮主要由角质形成细胞(KCs)组成,KC从基底层到角质层的增殖和分化是表皮中存在的细胞层次。在这项研究中,我们探讨了人类造血干细胞(HSC)分化为KCs的能力。用角质形成细胞分化培养基(KDM)诱导培养的CD34,CD45和CD133阳性并具有显着的端粒酶活性的HSC,由牛垂体提取物(BPE)组成,表皮生长因子(EGF),胰岛素,氢化可的松,肾上腺素,转铁蛋白,氯化钙(CaCl2),骨形态发生蛋白4(BMP4),和视黄酸(RA)。通过细胞角蛋白标志物K5(角蛋白5)的表达监测分化,K14(角蛋白14),K10(角蛋白10),K1(角蛋白1),转谷氨酰胺酶1(TGM1),总蛋白(IVL),在第0天(D0)和聚丝蛋白(FLG),第6天(D6),第11天(D11),第18天(D18),第24天(D24),和第30天(D30)使用免疫细胞化学,荧光显微镜,流式细胞术,qPCR,和西方印迹。结果揭示了K5和K14基因在D6细胞(早期角质形成细胞)中的表达,D11-D18细胞(成熟角质形成细胞)中的K10和K1基因具有活性端粒酶,和FLG,IVL,和TGM1在D18-D24细胞(末端角质形成细胞)中,到D30时,KC被完全去核,类似于角质化基质。这种将HSC分化为KCs的方法解释了正常表皮中存在的细胞顺序,并为探索人类HSC在表皮分化中的用途开辟了可能性。
The epidermis is largely composed of keratinocytes (KCs), and the proliferation and differentiation of KCs from the stratum basale to the stratum corneum is the cellular hierarchy present in the epidermis. In this study, we explore the differentiation abilities of human hematopoietic stem cells (HSCs) into KCs. Cultured HSCs positive for CD34, CD45, and CD133 with prominent telomerase activity were induced with keratinocyte differentiation medium (KDM), which is composed of bovine pituitary extract (BPE), epidermal growth factor (EGF), insulin, hydrocortisone, epinephrine, transferrin, calcium chloride (CaCl2), bone morphogenetic protein 4 (BMP4), and retinoic acid (RA). Differentiation was monitored through the expression of cytokeratin markers K5 (keratin 5), K14 (keratin 14), K10 (keratin 10), K1 (keratin 1), transglutaminase 1 (TGM1),
involucrin (IVL), and filaggrin (FLG) on day 0 (D0), day 6 (D6), day 11 (D11), day 18 (D18), day 24 (D24), and day 30 (D30) using immunocytochemistry, fluorescence microscopy, flow cytometry, qPCR, and Western blotting. The results revealed the expression of K5 and K14 genes in D6 cells (early keratinocytes), K10 and K1 genes in D11-D18 cells (mature keratinocytes) with active telomerase enzyme, and FLG, IVL, and TGM1 in D18-D24 cells (terminal keratinocytes), and by D30, the KCs were completely enucleated similar to cornified matrix. This method of differentiation of HSCs to KCs explains the cellular order exists in the normal epidermis and opens the possibility of exploring the use of human HSCs in the epidermal differentiation.