人原代角质形成细胞在两种培养基中增殖能力的比较

Comparisons of the cell proliferation capacity of human primary keratinocytes in two different media

目的:比较第一代和第二代人原代角质形成细胞(human primary keratinocytes,HPK)在两种培养基[(EpiLife培养基+HKGS)和DK-SFM培养基]中增殖能力的差异。方法:通过两步法从人的皮肤组织获得第一代HPK,进行传代后获得第二代HPK,采用细胞计数仪计数,绘制生长曲线。用免疫荧光技术鉴定第二代HPK,并用流式细胞术检测其细胞周期。结果:从HPK生长曲线可见,在DK-SFM培养基中,第一代HPK的增殖速率高于(EpiLife培养基+HKGS)中,第二代HPK的增殖速率低于(EpiLife培养基+HKGS)培养中。细胞周期结果同上,第二代HPK在DK-SFM中其G2期细胞和S期细胞含量分别为8.5%和11.2%,低于在(EpiLife培养基+HKGS)中的13.9%和18.3%。结论:两种培养基培养HPK各有利弊,若想在短期获得纯度较高的HPK,选择DK-SFM培养基较好;若想相对长期培养HPK,选择(EpiLife培养基+HKGS)更有优势。

Objective, To compare the differences in proliferation abilities of the first and second generations of human pri- mary keratinocytes （HPK） in two different media [EpiLife medium ＋ HKGS and defined-keratinocyte serum-free medium（DK- SFM）]. Methods. The first generation of HPK was isolated from the skin by two-step digestion technique. The second genera- tion of HPK was obtained after passage, Cell growth curves were made by cell counter. The second generation of HPK were identified by immunofluorescence staining, and the cell cycle was determined by flow cytometry. Results： In the growth curves of HPK, the cell proliferation rate of the first generation of HPK was superior in DK-SFM than that in （EpiLife medium ＋ HKGS）. However, the proliferation rate of second generation of HPK in DK-SFM was lower than that in （EpiLife medium ＋ HKGS） . The cell cycle experiment showed that after passage, the G2 phase and S phase cell contents were 8.5% and 11.2% in DK SFM, which were lower than those in （EpiLife medium ＋ HKGS）（G2：13.9%, S：18.3 0% ）. Conclusion： Both media have their own advantages and disadvantages. If relatively pure HPK is expected in a short time, it is better to select the DK-SFM medium. On the other hand, if prolonged culture of HPK is desired, it is more advantageous to choose （EpiLife Medium＋ HKGS）.