本文選題：周細胞 + 脊髓��； 參考：《基礎醫(yī)學與臨床》2015年05期

【摘要】：目的應用周細胞培養(yǎng)基(PCM)分離篩選小鼠脊髓微血管周細胞(SCMP),對其生物學功能進行評價。方法10只3周齡C57小鼠,無菌條件下取脊髓去除軟脊膜,剪碎成大約1 mm×1 mm×1 mm。兩次酶消化后用含20%牛血清白蛋白的DMEM離心獲得微血管。用內皮細胞培養(yǎng)基(ECM)培養(yǎng),傳代2次后改用PCM培養(yǎng)。倒置顯微鏡觀察細胞增殖狀況。免疫細胞化學法檢測血小板源性生長因子受體β(PDGFRβ)、神經元-膠質抗原2(NG2)、von Willebrand因子(v WF)和膠質纖維酸性蛋白(GFAP)的表達。流式檢測CD140b、CD31、CD11b和GFAP的表達。將PCM換為10%胎牛血清(FBS)的DMEM培養(yǎng)基,檢測細胞α-SMA表達的變化。通過周細胞-內皮細胞共培養(yǎng)成管實驗檢測細胞成管能力。結果接種48 h細胞爬出,7~9 d匯合,周細胞和內皮細胞伴隨狀增殖。改用PCM培養(yǎng)后內皮細胞減少,周細胞呈優(yōu)勢生長。免疫細胞化學表明PDGFRβ和NG2陽性,v WF和GFAP陰性;流式結果表明細胞PDGFRβ的陽性率為95.52%±2.55%,GFAP為0.63%±0.26%,CD31為0.80%±0.26%,CD11b為1.02%±0.35%。10%胎牛血清的DMEM促進細胞分化,α-SMA表達升高。成管實驗周細胞與內皮細胞共同形成管腔樣結構。結論通過PCM篩選法能夠成功獲得純度較高的SCMP,所獲得的細胞具備明顯的周細胞的形態(tài)和功能特征。
[Abstract]:Objective to isolate and screen mouse spinal microvascular pericyte (SCMPP) by pericyte culture medium (PCM) and evaluate its biological function. Methods the spinal cord of 10 C57 mice aged 3 weeks was removed from the spinal cord and shredded into about 1 mm 脳 1 mm under aseptic condition. Microvasculature was obtained by centrifugation with DMEM containing 20% bovine serum albumin after two enzyme digestion. Endothelial cell culture medium (ECM) was used, then subcultured for 2 times and then transferred to PCM culture. Cell proliferation was observed by inverted microscope. Immunocytochemistry was used to detect the expression of PDGFR 尾, neuron-glial antigen 2NG2An Willebrand factor (v WFV) and glial fibrillary acidic protein (GFAP) in platelet-derived growth factor receptor (尾 -PDGFR 尾) and glial fibrillary acidic protein (GFAP). The expression of CD140b, CD31, CD11b and GFAP was detected by flow cytometry. The expression of 偽 -SMA was detected by changing PCM into 10% fetal bovine serum (FBS) DMEM medium. The ability of cell tubulogenesis was measured by pericyte-endothelial cell co-culture tube-forming experiment. Results 48 h after inoculation, the cells crawled out of the cells and converged for 9 days, and the pericytes and endothelial cells were accompanied by proliferation. Endothelial cells decreased and pericytes grew preferentially after cultured with PCM. Immunocytochemistry showed that PDGFR 尾 and NG2 were positive for vWF and GFAP, and the positive rate of PDGFR 尾 was 95.52% 鹵2.55%. The positive rate of GFAP was 0.63% 鹵0.26 + 0.80% 鹵0.26 + CD11b was 1.02% 鹵0.35.10% of fetal bovine serum promoted cell differentiation, and 偽 -SMA expression increased. The endothelial cells and pericytes formed lumen-like structures. Conclusion High purity SCMP can be obtained by PCM screening method, and the obtained cells have obvious morphological and functional characteristics of pericytes.
【作者單位】： 中國醫(yī)學科學院北京協(xié)和醫(yī)學院微循環(huán)研究所;
【基金】：2014年協(xié)和青年教師基金&中央高�；究蒲袠I(yè)務費專項資金(33320140062)
【分類號】：R329.2

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本文編號：1900621