成年鼠被毛毛乳头细胞分离方法的建立与评价

姜卫; 苗勇; 肖顺娥; 王瑾; 胡志奇

doi:10.16016/j.1000-5404.201704068

您当前的位置：

首页 >

文章列表页 >

成年鼠被毛毛乳头细胞分离方法的建立与评价

基础医学 | 更新时间：2026-01-09

- 成年鼠被毛毛乳头细胞分离方法的建立与评价
- Establishment and evaluation of separation for adult mousepelage dermal papilla
- 第三军医大学学报 2017年39卷第18期页码：1790-1795
- 作者机构：
  
  510515 广州，南方医科大学南方医院整形外科
- 作者简介：
  
  [ 胡志奇，E-mail：huzhiqidr@163.com HU Zhiqi, E-mail: huzhiqidr@163.com
- 基金信息：
  
  国家自然科学基金面上项目(31170949);国家自然科学基金面上项目(81471900);广东省自然科学基金(S2012010009454);广东省自然科学基金(2015A030311001);南方医科大学南方医院院长基金(2013Z013)
- DOI：10.16016/j.1000-5404.201704068
  中图分类号： R322.995; R329-33
- 收稿：2017-04-12，
  
  修回：2017-06-01，
  
  纸质出版：2017-09-30
- 稿件说明：
移动端阅览
姜卫, 苗勇, 肖顺娥, 等. 成年鼠被毛毛乳头细胞分离方法的建立与评价[J]. 第三军医大学学报, 2017,39(18):1790-1795.

Wei JIANG, Yong MIAO, Shun'e XIAO, et al. Establishment and evaluation of separation for adult mousepelage dermal papilla[J]. Journal of Third Military Medical University, 2017, 39(18): 1790-1795.
姜卫, 苗勇, 肖顺娥, 等. 成年鼠被毛毛乳头细胞分离方法的建立与评价[J]. 第三军医大学学报, 2017,39(18):1790-1795. DOI： 10.16016/j.1000-5404.201704068.

Wei JIANG, Yong MIAO, Shun'e XIAO, et al. Establishment and evaluation of separation for adult mousepelage dermal papilla[J]. Journal of Third Military Medical University, 2017, 39(18): 1790-1795. DOI： 10.16016/j.1000-5404.201704068.

摘要

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2 h，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(

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。

Abstract

Objective

To seek a simple and efficient method for isolating and culturing mouse pelage dermal papilla cells (DPCs).

Methods

Mouse dorsal skin was cut off from 5-week-old C57BL/6J mice

and then the hair shafts were shaved

and digested in 0.2% type Ⅰ collagenase at 37 ℃ for 2 h. The hair bulbs were obtained by scraping

and collected by Ficoll gradient centrifugation for 2 times. The pure dermal papilla (DP) were obtained and cultured

in vitro

. The adherent rate and specific markers of DPCs were detected

and the hair follicle re

construction was observed

in vivo

Results

Our method was simple and easy

and reduced the risk of contamination significantly. The adherence of obtained pelage DP and the cell emigration from DP were quite fast. qRT-PCR and cell immunofluorescence assay confirmed that the cultured pelage DPCs expressed the specific markers associated with hair follicle-inductive ability

such as ALP

β-catenin and Versican

and the expression levels had no significant differences with those of vibrissa DPCs (

0.05). The obtained pelage DPCs showed the ability to induce hair follicle regeneration after being transplanted in the back of nude mice.

Conclusion

Collagenase digestion combined with Ficoll gradient centrifugation is a simple and effective separation method to obtain mouse pelage DP.

关键词

Keywords

references

OLIVER R F. The experimental induction of whisker growth in the hooded rat by implantation of dermal papillae[J]. J Embryol Exp Morphol, 1967, 18(1): 43-51.

JAHODA C A, HORNE K A, OLIVER R F. Induction of hair growth by implantation of cultured dermal papilla cells[J]. Nature, 1984, 311(5986): 560-562. DOI: 10.1038/311560a0.

REYNOLDS A J, JAHODA C A. Cultured dermal papilla cells induce follicle formation and hair growth by transdifferentiation of an adult epidermis[J]. Development, 1992, 115(2): 587-593.

LICHTI U, ANDERS J, YUSPA S H. Isolation and short-term culture of primary keratinocytes, hair follicle populations and dermal cells from newborn mice and keratinocytes from adult mice for in vitro analysis and for grafting to immunodeficient mice[J ] . Nat Protoc, 2008, 3(5): 799-810. DOI: 10.1038/nprot.2008.50.

WARREN R, CHESTNUT M H, WONG T K, et al. Improved method for the isolation and cultivation of human scalp dermal papilla cells[J]. J Invest Dermatol, 1992, 98(5): 693-699. DOI: 10.1111/1523-1747.ep12499909.

WU J J, LIU R Q, LU Y G, et al. Enzyme digestion to isolate and culture human scalp dermal papilla cells: a more efficient method[J]. Arch Dermatol Res, 2005, 297(2): 60-67. DOI: 10.1007/s00403-005-0554-z.

FUKUDA J. A pure, monolayer culture of Merkel cells from sinus hair follicles of the rat[J]. Neurosci Lett, 1996, 216(2): 73-76. DOI: 10.1016/0304-3940(96)12945-1.

KIM J N, KOH K S, LEE E, et al. The morphology of the rat vibrissal follicle-sinus complex revealed by three-dimensional computer-aided reconstruction[J]. Cells Tissues Organs (Print), 2011, 193(3): 207-214. DOI: 10.1159/000319394.

COHEN J. The transplantation of individual rat and guineapig whisker papillae[J]. J Embryol Exp Morphol, 1961, 9: 117-127.

OLIVER R F. Whisker growth after removal of the dermal papilla and lengths of follicle in the hooded rat[J]. J Embryol Exp Morphol, 1966, 15(3): 331-347.

MESSENGER A G. The culture of dermal papilla cells from human hair follicles[J]. Br J Dermatol, 1984, 110(6): 685-689. DOI: 10.1111/j.1365-2133.1984.tb04705.x.

MAGERL M, KAUSER S, PAUS R, et al. Simple and rapid method to isolate and culture follicular papillae from human scalp hair follicles[J]. Exp Dermatol, 2002, 11(4): 381-385. DOI: 10.1034/j.1600-0625.2002.110414.x.

伍津津, 刘荣卿, 叶庆佾, 等.毛乳头细胞高效培养方法探索[J].中华皮肤科杂志, 1997, 30(6): 383-385.

WU J J, LIU R Q, LIU Q Y, et al. An Efficient method for isolation and cultivation of human scalp dermal papilla cells[J]. Chin J Dermatol,1997, 30(6): 383-385.

李宇, 李国强, 纪影畅, 等.一步酶消化法高效快速分离培养人头皮毛乳头细胞[J].中国美容医学, 2004, 13(3): 264-266.

LI Y, LI G Q, JI Y C, et al. An efficient and rapid method to isolate and culture dermal papilla cells from human hair follicles by one step digestive treatment[J]. Chin J Aesth Med, 2004, 13(3): 264-266.

MORGAN B A. The dermal papilla: an instructive niche for epithelial stem and progenitor cells in development and regeneration of the hair follicle[J]. Cold Spring Harb Perspect Med, 2014, 4(7): a015180. DOI: 10.1101/cshperspect.a015180.

MESSENGERA G, ELLIOTT K, WESTGATE G E, et al. Distribution of extracellular matrix molecules in human hair follicles[J]. Ann N Y Acad Sci, 1991, 642: 253-262. DOI:10.1111/j.1749-6632.1991.tb24392.x.

MCELWEE K J, KISSLING S, WENZEL E, et al. Cultured peribulbar dermal sheath cells can induce hair follicle development and contribute to the dermal sheath and dermal papilla[J]. J Invest Dermatol, 2003, 121(6): 1267-1275. DOI:10.1111/j.1523-1747.2003.12568.x.

LEE S H, YOON J, SHIN S H, et al. Valproic Acid Induces Hair Regeneration in Murine Model and Activates Alkaline Phosphatase Activity in Human Dermal Papilla Cells[J]. PLoS One, 2012, 7(4): e34152. DOI: 10.1371/journal.pone.0034152.

DU CROS D L, LEBARON R G, COUCHMAN J R. Association of versican with dermal matrices and its potential role in hair follicle development and cycling[J]. J Invest Dermatol, 1995, 105(3): 426-431. DOI: 10.1111/1523-1747.ep12321131.

RABBANI P, TAKEO M, CHOU W, et al. Coordinated activation of Wnt in epithelial and melanocyte stem cells initiates pigmented hair regeneration[J]. Cell, 2011, 145(6): 941-955. DOI: 10.1016/j.cell.2011.05.004.

WODARZ A, NUSSE R. Mechanisms of Wnt signaling in development[J]. Annu Rev Cell Dev Biol, 1998, 14: 59-88. DOI: 10.1146/annurev.cellbio.14.1.59.

SHIMIZU H, MORGAN B A. Wnt signaling through the beta-catenin pathway is sufficient to maintain, but not restore, anagen-phase characteristics of dermal papilla cells[J]. J Invest Dermatol, 2004, 122(2): 239-245. DOI: 10.1046/j.0022-202X.2004.22224.x.

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

Frizzled4在小鼠毛囊周期中的表达

JNK在小鼠毛囊周期中的动态表达

TPA促进小鼠毛囊中TCF3的表达

gasdermin3在小鼠同步化毛囊周期中的表达

GasderminA3基因突变对小鼠毛囊β-catenin表达的影响