毛乳头细胞与毛囊生长周期

毛囊是一个具有生长、退化、休止三个阶段周期性循环的器官,它由上皮和真皮二部分组成。在生长期,毛囊的上皮和真皮部分增殖分化,上皮部分向真皮延伸,形成毛母质、内根鞘、毛乳头和毛干。进入退化期,毛母质向上运动,与不断缩小的毛乳头分离。在休止期毛囊内毛母质消失,毛乳头浓缩成球点状,直到下一周期的开始［１］。尽管毛囊的这些周期性变化早已为大家所熟悉,但有关控制这些变化的机制至今尚未阐明。近年来研究毛囊生长周期（Ｆｏｌｌｉｃｌｅ ｇｒｏｗｔｈ ｃｙｃｌｅ,ＦＧＣ）调控的文章较多,一些研究显示,毛乳头细胞（Ｄｅ…     

硫酸软骨素、硫酸肝素对毛乳头细胞粘附及生长的影响

目的：探讨细胞外基质在毛囊生长发育中的作用,寻持毛乳头细胞（DPC）生长的调节因素。方法：用两步酸消化法分离培养DPC,并通过免疫组化α-平滑肌肌动蛋白（Actin）染色证实。测定硫酸软骨素A,硫酸软骨素C及硫酸肝素对DPC粘附及生长的影响。结果：两步酶消化法分离培养DPC效率高,纯度高,方法简单易行。硫酸软骨素A和硫酸肝素明显促进DPC粘附及生长,而硫酸软骨素C作用不明显。结论：某些细胞外基质可调节DPC的生长,从而影响毛囊的生长发育。     

毛乳头细胞凝集性生长与细胞外基质分泌关系的组织化学研究

目的：研究毛乳头细胞的凝集性生长方式与其产生的细胞外基质之间的关系。方法：用人头皮分离的低传代毛乳头细胞与毛发上皮细胞共同培养形成的毛乳头细胞凝集性生长区及非凝集生长区的毛乳头细胞进行阿新蓝、过碘酸雪夫氏染色，观察染色反应的强弱。结果：新分离的毛乳头，阿新蓝和过碘酸雪夫氏染色均呈强阳性，培养的毛乳头细胞凝集性生长区阿新蓝－过碘酸雪夫氏染色亦显示强阳性，而非凝集区的毛乳头细胞显色呈弱阳性。结论：培养的毛乳头细胞凝集性生长后，合成分泌细胞外基质的功能较非凝集生长的毛乳头细胞明显增强。     

毛乳头及毛囊生长调控的若干研究进展

毛囊是控制毛发生长的皮肤附属器官 ,其生长周期可分为生长期 (anagen)、退行期 (catagen)和休止期 (telogen)。在毛囊生长的不同周期 ,一些活性蛋白、细胞因子及激素均可对毛囊周期或生长产生促进或抑制作用。毫无疑问 ,寻找能够影响毛发生长的成分对阐明毛囊生长周期调控机制、发展有效治疗脱发的手段具有重要意义。本文将近期有关毛囊和毛乳头生长调控的相关进展作一综述。1　毛囊的结构及其生长周期毛囊是一个复杂的皮肤附属结构 ,它由上皮 (毛母质与外根鞘 )和真皮 (毛乳头和结缔组织鞘 )两大部分组成。其中包含有 2 0余种细胞 ,包括角…     

正常人毛乳头细胞和真皮鞘细胞的生长特性观察

目的 观察毛乳头细胞和真皮鞘细胞在体外培养条件下的生长特性。方法 采用胶原酶消化法培养正常人头皮毛囊的毛乳头细胞和真皮鞘细胞,在体外进行传代培养,测定毛乳头细胞不同时相点的细胞数,并观察表皮细胞生长因子、氢化可的松＋胰岛素、粒细胞巨噬细胞集落刺激因子对毛乳头细胞和真皮鞘细胞增殖的影响。结果 毛乳头细胞、真皮鞘成纤维细胞和真皮成纤维细胞的生长呈现３个时相,即停滞期、指数生长期和缓慢生长期。３种细胞都有４ｄ的停滞期,５～１１ｄ为指数生长期,之后呈现出缓慢生长。ＥＧＦ对３种细胞都有显的促进作用（Ｐ〈０．００１）,尤其是对真皮鞘成纤维细胞的作用更强。氢化可的松和胰岛素、ＧＭ－ＣＳＦ对３种细胞的作用不明显。毛乳头细胞、真皮鞘细胞和成纤维细胞的倍增时间分别为６０、５９．２、７６．８ｈ。结论 ＥＧＦ对毛乳头细胞、真皮鞘细胞和     

毛乳头在毛囊生长周期中的变化和作用

目的 了解人毛乳头结构在毛发生长周期中的变化及其对毛发生长的调控作用.方法 将整形手术后取下的头皮在无菌条件下分离成游离的毛囊,在Williams E培养基中培养;通过横断毛囊和完整毛囊的培养来观察毛囊的生长状况和处于不同生长周期的毛囊的毛乳头结构变化.结果 分离后的完整毛囊在Williams E培养基中平均可继续生长12 d,平均生长速度为每天0.2～0.3 mm;静止期或生长初期毛囊的毛乳头呈圆形贴在毛球底部,快速生长的毛囊毛乳头变长呈锥形,结构疏松,与毛根紧密相接,当毛囊进入衰退期直至停止生长后毛乳头萎缩变小,并与毛根逐渐分离.在低位横断毛囊,毛囊可再生毛球并继续生长,但生长速度减慢;在高位横断毛囊,毛囊不能继续生长.结论 毛乳头在毛发生长周期中不断变化,生长期毛乳头体积增大,与毛母质联系紧密,它在毛囊的生长和生长周期中起着不可缺少的调控作用.     

毛囊细胞移植诱导裸鼠毛囊样结构形成的研究

目的:观察毛囊细胞移植诱导裸鼠毛发再生和毛囊重建情况.方法:采用细胞培养技术和裸鼠移植技术,将培养的毛囊毛乳头细胞、真皮鞘细胞和头皮真皮成纤维细胞按比例与毛囊上皮细胞混合,移植到裸鼠皮下,观察毛囊形成情况.结果:在毛囊毛乳头细胞与毛囊上皮细胞混合后移植到裸鼠皮下后可见毛囊样结构形成,而毛囊真皮鞘细胞和头皮成纤维细胞与毛囊上皮细胞混合则不能诱导裸鼠毛囊样结构形成.结论:低传代培养的毛乳头细胞与毛囊上皮细胞混合后在体内可诱导毛囊样结构形成.     

毛乳头细胞的研究现状

毛囊是一种皮肤附属器官,其结构的完整不仅为机体新陈代谢所需,同时对于动物的取暖、人的外表美观都有重要意义。毛囊具有自我更新和周期性生长的特点,这与毛球部存在的一群间充质细胞密切相关,即毛乳头细胞(DPC),它在毛囊发育以及生后毛囊周期过程中都发挥着重要的诱导作用。缺失DPC,无法形成毛囊;而在生后毛囊周期中,DPC通过与周围的毛囊上皮细胞相互作用,直接或间接地诱导毛囊周期性的生长。研究DPC的特性及其发挥的生物学功能,对了解毛囊的发育和再生具有重要的指导意义。     

毛囊细胞混合物移植诱导裸鼠毛囊样结构形成

目的 观察毛囊细胞复合物移植诱导裸鼠毛发再生和毛囊重建情况.方法 采用裸鼠移植技术,将混合分离出来的毛囊细胞,包括毛囊毛乳头细胞、毛囊外根鞘细胞、毛囊真皮鞘细胞和毛囊真皮成纤维细胞等,移植到裸鼠皮下,观察毛囊形成情况.结果 在裸鼠的皮肤切片中可以看到较为完整的毛囊结构形成.结论 毛囊细胞混合物可以在体内诱导出毛囊样结构形成.     

人毛乳头细胞凝集性生长差异表达基因cDNA文库的构建

目的 构建毛乳头细胞凝集性生长状态下差异表达基因的消减cDNA文库。方法 分别提取凝集性生长和非凝集性生长状态下毛乳头细胞中的总RNA，应用SMART cDNA合成技术和抑制性消减杂交技术分离毛乳头细胞凝集性生长状态下差异表达基因的cDNA片段并建立消减cDNA文库；利用PCR对随机挑选的100个白色菌落进行插入片段的验证，对其中证实有插入片段的30个克隆进行cDNA斑点杂交验证。结果 所构建的消减cDNA文库扩增后得到320个阳性克隆，随机挑选的100个阳性克隆中95％的均有长度在100—600bp的插入片段，cDNA斑点杂交验证显示27个(90．0％)克隆为阳性。结论 结合SMART cDNA合成技术，应用抑制性消减杂交成功地构建了毛乳头细胞凝集性生长差异表达基因消减cDNA文库，为进一步筛选和克隆毛乳头细胞凝集性生长相关基因奠定了基础。     

人毛乳头细胞在不同传代时期某些细胞因子表达的变化

目的 观察毛乳头细胞在体外培养条件下的生长特性,为毛囊重建提供参考资料.方法 采用二步酶消化法分离正常人头皮毛囊的毛乳头细胞,在体外进行传代培养,用免疫组化方法观察不同传代毛乳头细胞生长因子表达的差异.结果 高代培养毛乳头细胞逐渐丧失其凝集性生长的特性,生长因子的表达逐渐消失.结论 毛乳头细胞的生长及其特性的保持受许多因素的影响,体外培养的毛乳头细胞与体内有差异.低代毛乳头条件培养基可恢复毛乳头细胞部分生长特性.     

Biological characterization of cultured dermal papilla cells and hair follicle regeneration in vitro and in vivo

Background Dermal papilla cells （DPC） are a group of mesenchyme-derived cells at the base of the hair follicle, where they regulate and control hair follicle growth through the expression and secretion of cytokines. Nevertheless, the role of DPC derived chemokines and other cytokines in the hair follicle biology remain speculative. In this study, we investigated the expression of basic fibroblast growth factor （bFGF）, endothelin-1 （ET-1） and stem cell factor （SCF） in different passages of cultured DPC and their effects on the biological behaviour of DPC. Methods The expression of bFGF, ET-1 and SCF in different passages of cultured DPC and their possible effects on the biological behavior of DPC are investigated using in sire hybridization and immunochemistry. In addition, we performed transplantation of hair follicle cells into nude mice. The cultured DPC, dermal sheath cells and fibroblast of human scalp, respectively, were mixed with cells of the hair follicle epithelium in different ratios, and then were cultured in hair follicle organotypic cultures or implanted into the subcutis of nude mice. Results The expression of ET-1 and SCF in early passages of cultured DPC became stronger, but turned weaker and even negative in late passages （〉6 passages）. Hair follicle-like structures were formed after DPC combined with the cells of hair follicle epithelium cells in hair follicle organotypic cultures. When hair follicle organotypic cultures were implanted into the subcutis of nude mice, the relative intact hair follicles were formed. After the transplantation of hair follicle cells into the nude mice, the hair follicle-like structure was formed in the group that contained DPC mixed with hair follicle epithelium cells. However, no hair follicles were formed in the other two groups. It was found that the higher the expression of ET-1 and SCF in DPC, the stronger the ability of DPC to induce hair follicle regeneration. Conclusions The cultured DPC can induce hair follicle regeneration and sustain hair growth in vivo and in vitro. Moreover, the expression of ET-1 and SCF is correlated with the ability of DPC inducing hair follicle regeneration.     

Biological characterization of cultured dermal papilla cells and hair follicle regeneration in vitro and in vivo

Background Dermal papilla cells (DPC) are a group of mesenchyme-derived cells at the base of the hair follicle, where they regulate and control hair follicle growth through the expression and secretion of cytokines. Nevertheless, the role of DPC derived chemokines and other cytokines in the hair follicle biology remain speculative. In this study, we investigated the expression of basic fibroblast growth factor (bFGF), endothelin-1 (ET-1) and stem cell factor (SCF) in different passages of cultured DPC and their effects on the biological behaviour of DPC.Methods The expression of bFGF, ET-1 and SCF in different passages of cultured DPC and their possible effects on the biological behavior of DPC are investigated using in situ hybridization and immunochemistry. In addition, we performed transplantation of hair follicle cells into nude mice. The cultured DPC, dermal sheath cells and fibroblast of human scalp, respectively, were mixed with cells of the hair follicle epithelium in different ratios, and then were cultured in hair follicle organotypic cultures or implanted into the subcutis of nude mice.Results The expression of ET-1 and SCF in early passages of cultured DPC became stronger, but turned weaker and even negative in late passages (>6 passages). Hair follicle-like structures were formed after DPC combined with the cells of hair follicle epithelium cells in hair follicle organotypic cultures. When hair follicle organotypic cultures were implanted into the subcutis of nude mice, the relative intact hair follicles were formed. After the transplantation of hair follicle cells into the nude mice, the hair follicle-like structure was formed in the group that contained DPC mixed with hair follicle epithelium cells. However, no hair follicles were formed in the other two groups. It was found that the higher the expression of ET-1 and SCF in DPC, the stronger the ability of DPC to induce hair follicle regeneration.Conclusions The cultured DPC can induce hair follicle regeneration and sustain hair growth in vivo and in vitro. Moreover, the expression of ET-1 and SCF is correlated with the ability of DPC inducing hair follicle regeneration.     

人毛囊外根鞘隆起、真皮鞘和毛乳头细胞高效同步分离培养的方法

目的 寻求一种快速高效同步分离培养人头皮毛囊外根鞘隆起(Bulge)细胞.真皮鞘细胞和毛乳头细胞的方法.方法 将人头皮标本剪成小块,中性蛋白酶中37℃孵育2h后镜下将带外根鞘的毛干从真皮鞘中拔出,组织块法培养外根鞘隆起(Bulge)细胞;从真皮一皮下组织交界处横断头皮,拔出含毛乳头的真皮鞘,胶原酶D 37℃孵育6～8h,多次低速离心,毛乳头沉淀重悬后培养,收集的真皮鞘细胞上清液经高速离心后重悬培养;培养的细胞分别用K19或α-平滑肌动蛋白组化鉴定.结果 同一标本可获得纯化的外根鞘Bulge细胞,真皮鞘细胞和毛乳头细胞.结论 将现有的分离方法改进和综合运用.可以从同一毛囊标本同步高效获取3种成分细胞.     

毛囊细胞移植法诱导毛囊的初步研究

目的 构建一个可靠有效的移植毛囊细胞诱导毛发发育的模型,以治疗脱发.方法 取自愿捐献的成人头皮标本,联用显微分离与免疫磁珠法获得人毛囊干细胞;消化法获得毛乳头细胞.培养后混合植入裸鼠皮下,观察毛囊形成情况.结果 在裸鼠的皮肤切片中可以看到较为完整的毛囊结构形成.结论 毛囊细胞移植法可以在体内诱导出毛囊样结构,为将来治疗脱发奠定了基础.

Abstract：

Objective To establish a convenient and reliable method for inducing hair regeneration by follicular cell implantation for the treatment of alopecia. Methods The human hair follicle stem cells were separated and purified by micromanipulation and magnetic cell sorting, and human scalp dermal papilla cells were isolated by enzyme digestion. The two cells were mixed and implanted subcutaneously in nude mice to observe the regeneration of the hair follicles. Results Formation of intact hair follicle-like structures was observed in the skin sections of the recipient nude mice. Conclusion Follicular cell implantation can induce hair follicle-like structures in nude mice, which provides a means for efficient hair regeneration for treatment of hair loss.     

毛乳头细胞的培养与鉴定

目的：建立毛乳头细胞的培养方法,以进一步研究毛乳头细胞在毛发再生中的作用和意义．方法：分离得到完整毛囊,依次采用Dispase酶和胶原酶消化得到毛乳头,再进行毛乳头细胞的培养．结果：成功进行了毛乳头细胞的原代和传代培养,所获细胞经细胞化学染色证实细胞具有明显的异染特性．毛乳头贴壁后5～6d周围即可见细胞长出,细胞生长快,12～16d左右细胞即可融合．在倒置显微镜下细胞呈成纤维细胞样,有聚集生长特性,至少可以传15代．冻存后复苏细胞贴壁率在80％以上．结论：该方法是体外培养毛乳头细胞的理想方法,操作较为简单,降低了工作强度,所获细胞纯度高,可以长期传代、冻存．     

成年鼠被毛毛乳头细胞分离方法的建立与评价

目的 寻求一种简单高效的分离培养小鼠被毛毛乳头细胞的方法.方法 取5周龄C57小鼠背部皮肤,刮去毛发后予0.2％Ⅰ型胶原酶37℃消化2h,刮下毛球部,经过2次Ficoll密度梯度离心后,得到纯净的毛乳头进行培养.检测毛乳头贴壁率、毛乳头细胞特异性标记表达及其体内毛囊重建能力.结果 该分离方法能显著降低工作强度,减少污染机会,获得的毛乳头贴壁快、细胞迁出快.qRT-PCR、细胞免疫荧光实验结果均证实体外培养的小鼠被毛毛乳头细胞可表达与毛囊诱导能力相关的特异性标记物ALP、β-catenin及Versican,且与触须毛乳头细胞相比差异无统计学意义(P＞0.05).体外培养的小鼠被毛毛乳头细胞具有诱导毛囊再生的能力.结论 胶原酶消化结合Ficoll梯度离心是一种简单、有效地分离获取小鼠背部毛乳头的方法.