Interleukin-6 cytokine family member oncostatin M is a hair-follicle-expressed factor with hair growth inhibitory properties

Abstract:  The activation of receptor complexes containing glycoprotein 130 (gp130) identifies the interleukin (IL)-6 cytokine family. We examined members of this family for their expression and activity in hair follicles. Quantitative polymerase chain reaction using mRNA derived from microdissected, anagen-stage human hair follicles and comparison to non-follicular skin epithelium revealed higher levels of IL-6 (15.5-fold) and oncostatin M (OSM, 3.4-fold) in hair follicles. In contrast, expression of all mRNAs coding for IL-6 cytokine family receptors was reduced. Immunohistology suggested expression of OSM, gp130, leukaemia inhibitory factor receptor (LIFr) and IL-11r in the hair follicle root sheaths and dermal papilla, while IL-11, IL-6r and OSMr were expressed in root sheaths alone. IL-6 was expressed in the dermal papilla while cardiotrophin-1 (CT-1) and LIF were not observed. OSM and to a lesser extent CT-1 exhibited a dose-dependent growth inhibition capacity on human hair follicles in vitro . OSM and CT-1 incubated with agarose beads and injected subcutaneously at 1 μg per mouse into telogen skin of 65-day-old mice revealed no capacity to induce anagen hair growth. In contrast, injection of 65-day-old mice in which anagen had been induced by hair plucking revealed a moderate hair growth inhibitory capacity for OSM, but no significant effect for CT-1. The data identify OSM as a modulator of hair follicle growth and suggest other family members may also have some degree of hair growth inhibitory effect. In principle, increased expression of some IL-6 cytokine family members in cutaneous inflammation might contribute to the promotion of hair loss.     

毛乳头诱导毛囊形成和毛发生长信号通路的研究进展

毛乳头是位于毛囊底部的特殊成纤维细胞团块,在毛囊的周期性生长中,毛乳头细胞与周围细胞相互作用而发挥重要功能,被认为是控制毛囊形成和毛发生长信号传导的中心.近年来的研究表明,许多信号通路,包括Wnt、骨形成蛋白、Shh、Notch、成纤维细胞生长因子等信号通路,在毛乳头控制毛囊形成和毛发生长的机制中起作用.可以通过这些信号通路,促进体外培养毛乳头细胞的增殖、加强体外培养毛乳头细胞诱导毛发生长的能力,从而建立一种毛发重构技术用于临床治疗.     

Co-culture of human hair follicles and dermal papillae in a collagen matrix

Human hair follicles, either alone or in combination with dermal papillae, were cultured in a collagen matrix. When plucked hair follicles were cultured alone, spike-like structures composed of outer root sheath cells started growing around the follicle and then radiated into the gel. When isolated dermal papillae were embedded close to the follicles, spikes started growing earlier and grew more rapidly than without the papillae. In cultures of excised follicles from which the dermal papilla had been removed, epithelial cells (possibly hair bulb cells) started growing out from the bulbous portion and then also formed spikes. In the presence of a papilla, the spikes elongated toward the papilla, finally reaching and surrounding it. These findings suggest that dermal papilla cells produce a factor(s) that enhances growth of follicular epithelial cells and also attracts those cells. In cultures of whole excised follicles, two major characteristic patterns of cellular growth were recognized. When the dermal papilla remained inside the bulb in contact with the hair bulb matrix, the hair matrix cells proliferated and differentiated in the normal manner, resulting in elongation of the hair shaft and follicle. But when the papilla was detached from the hair bulb matrix, epithelial cells proliferated from the bulbous portion and finally formed hair follicle-like structures. Thus, attachment of the dermal papilla to the hair bulb matrix in the bulbous portion appears to be necessary for growth of the hair and follicle in the normal manner. Our model may be useful for examining the interaction between follicular epithelial cells and dermal papillae and for studying the growth of hair and follicles in vitro.     

Differences in expression of specific biomarkers distinguish human beard from scalp dermal papilla cells

Androgen exposure stimulates the growth of beard hair follicles. The follicle dermal papilla appears to be the site of androgen action; however, the molecular mechanisms that regulate this process are not well understood. In an attempt to identify genes that contribute to the androgen-responsive phenotype, we compared gene expression patterns in unstimulated and androgen-treated cultured human dermal papilla cells isolated from beard (androgen-sensitive) and occipital scalp (androgen-insensitive) hair follicles. Through this analysis, we identified three genes that are expressed at significantly higher levels in beard dermal papilla cells. One of these genes, sfrp-2 has been identified as a dermal papilla signature gene in mouse pelage follicles. Two of these genes, mn1 and atp1beta1, have not been studied in the hair follicle. A fourth, fibulin-1d, was slightly upregulated in beard dermal papilla cells. The differences in the expression of these genes in cultured beard and scalp dermal papilla cells reflected similar differences in microdissected dermal papilla isolated from intact beard and scalp follicles. Our findings introduce potentially novel signaling pathways in dermal papilla cells. In addition, this study supports that cultured dermal papilla cells provide a cell-based model system that is reflective of the biology of in vivo hair follicle cells.     

Differences in hair follicle dermal papilla volume are due to extracellular matrix volume and cell number: implications for the control of hair follicle size and androgen responses

The size of a hair follicle is thought to be determined by the volume of its dermal papilla. The volume of the dermal papilla depends on the number of cells it contains and on the volume of the extracellular matrix. To establish which of these two variables is related to differences in hair follicle size we performed a stereologic study on 235 hair follicles from different sites, including male facial skin (beard), female facial skin, and scalp. In facial follicles there was a strong correlation between the area of the hair cortex and the volume of the dermal papilla. The area of the hair cortex also correlated with the number of cells in the dermal papilla and with the volume of dermal papilla per cell. In scalp hair follicles, where there was a smaller range of sizes, the correlations between these variables were weaker. In large male facial follicles the mean total dermal papilla volume was almost 40-fold higher than in vellus follicles from female facial skin. This difference was associated with a mean 17-fold greater number of cells in the dermal papilla and a 2.4-fold greater volume associated with each cell. Intermediate results were obtained in scalp follicles. In many regions of the skin hair follicles enlarge in response to androgens during adult life hair. Our results imply that the increase in the volume of the dermal papilla in these follicles is due to an increase in the number of cells, either through proliferation or through the migration of cells from the follicular dermal sheath, and to an increase in the amount of extracellular matrix per cell. As androgens are thought to act primarily on the dermal papilla, these changes may have a direct bearing on the mechanism of androgen-mediated alterations in hair follicle size.     

真皮干细胞在毛囊周期中的作用

毛囊由表皮(上皮)及真皮(间充质)组成,它们之间的相互作用在毛囊的形态发生及生长中发挥重要作用,二者之间相互作用是毛囊成功重建的关键因素.在毛发形成过程中,真皮细胞是诱导者,上皮细胞是应答者.真皮鞘和毛乳头内存在毛囊真皮干细胞,属于成体干细胞,具有慢周期、未分化、自我更新和体外增殖能力强的特点.真皮鞘中的真皮干细胞较长寿,可以历经几个毛囊周期,重建真皮鞘.在毛囊周期的生长期,真皮鞘中的干细胞产生新细胞提供给毛乳头;在退行期,真皮干细胞子代移出毛乳头或死亡.毛囊真皮细胞对于损伤和疾病之后的毛囊重建及修复具有重要意义.     

Inhibitory autocrine factors produced by the mesenchyme-derived hair follicle dermal papilla may be a key to male pattern baldness

BACKGROUND: Androgenetic alopecia, or male pattern baldness, is a common, progressive disorder where large, terminal scalp hairs are gradually replaced by smaller hairs in precise patterns until only tiny vellus hairs remain. This balding can cause a marked reduction in the quality of life. Although these changes are driven by androgens, most molecular mechanisms are unknown, limiting available treatments. The mesenchyme-derived dermal papilla at the base of the mainly epithelial hair follicle controls the type of hair produced and is probably the site through which androgens act on follicle cells by altering the regulatory paracrine factors produced by dermal papilla cells. During changes in hair size the relationship between the hair and dermal papilla size remains constant, with alterations in both dermal papilla volume and cell number. This suggests that alterations within the dermal papilla itself play a key role in altering hair size in response to androgens. Cultured dermal papilla cells offer a useful model system to investigate this as they promote new hair growth in vivo, retain characteristics in vitro which reflect their parent follicle's response to androgens in vivo and secrete mitogenic factors for dermal papilla cells and keratinocytes. OBJECTIVES: To investigate whether cultured dermal papilla cells from balding follicles secrete altered amounts/types of mitogenic factors for dermal papilla cells than those from larger, normal follicles. We also aimed to determine whether rodent cells would recognize mitogenic signals from human cells in vitro and whether factors produced by balding dermal papilla cells could alter the start of a new mouse hair cycle in vivo. METHODS: Dermal papilla cells were cultured from normal, balding and almost clinically normal areas of balding scalps and their ability to produce mitogenic factors compared using both human and rat whisker dermal papilla cells as in vitro targets and mouse hair growth in vivo. RESULTS: Normal scalp cells produced soluble factors which stimulated the growth of both human scalp and rat whisker dermal papilla cells in vitro, demonstrating dose-responsive mitogenic capability across species. Although balding cells stimulated some growth, this was much reduced and they also secreted inhibitory factor(s). Balding cell media also delayed new hair growth when injected into mice. CONCLUSIONS: Human balding dermal papilla cells secrete inhibitory factors which affect the growth of both human and rodent dermal papilla cells and factors which delay the onset of anagen in mice in vivo. These inhibitory factor(s) probably cause the formation of smaller dermal papillae and smaller hairs in male pattern baldness. Identification of such factor(s) could lead to novel therapeutic approaches.     

Rat hair follicle growth in vitro

Pelage hair follicles were isolated by gentle microdissection from 8-12-day-old rats, and maintained in supplemented Williams E medium. Length measurements made on freshly isolated hair follicles, and at 24-h intervals, showed a significant increase in hair follicle length over 48 h, after which time no further significant increase in length was observed. Photomicrographs of maintained follicles showed that this increase in hair follicle length could be attributed to the production of a keratinized hair shaft. Histology and [methyl-3H] thymidine autoradiography of freshly isolated hair follicles showed the dermal papilla to be elongated, with thymidine uptake located predominantly in the matrix cells of the hair follicle bulb adjacent to the dermal papilla. This pattern remained unaltered for the first 48 h of maintenance, but after 72 h the dermal papilla had rounded into a tight ball of cells, with very little thymidine uptake occurring in the adjacent matrix cells. On maintenance, fetal calf serum (FCS), epidermal growth factor (EGF) and 12-o-tetradecanoyl phorbol 13-acetate (TPA) all significantly stimulated [methyl-3H] thymidine and [U-14C] leucine uptake, but inhibited hair follicle elongation. Insulin-like growth factor-1 (IGF-1) had no significant effect on rates of hair follicle elongation and [methyl-3H] thymidine uptake, but significantly stimulated rates of [U-14C] leucine uptake. Transforming growth factor-beta 1 (TGF-beta 1) significantly inhibited both the rate of [methyl-3H] thymidine uptake and hair follicle elongation.     

层黏连蛋白、纤连蛋白在人毛囊生长周期中的表达

目的 探讨层黏连蛋白(LM)、纤连蛋白(FN)在毛囊生长周期中的作用.方法 免疫组化S-P法检测LM、FN在毛囊生长期、退行期、休止期的表达.结果 ①生长期毛囊:LM均匀的表达于毛乳头处,在基底膜处呈明显的线状表达,外根鞘处表达也呈明显的阳性;FN均匀表达于毛乳头、基底膜、真皮鞘.②退行期毛囊:LM在毛乳头处少量表达,在基底膜处呈线状表达;FN在毛乳头和基底膜处表达均较生长期明显减少,但仍为阳性.③休止期毛囊:毛乳头浓缩成球点状,LM在毛乳头的表达为阴性,但在基底膜处仍然呈明显的线状表达,且较退行期更明显;FN在毛乳头、真皮鞘、基底膜处表达均为阴性.结论 LM、FN在毛囊的生长期、退行期及休止期的表达随毛囊生长周期而发生周期性变化,LM、FN可能参与毛囊生长周期的调节.     

血管生成素在人毛囊中的表达及其对毛发生长的影响

目的 探讨血管生成素在人毛囊中的表达及其意义。方法 分离完整的人生长期毛囊,提取总RNA,RT-PCR法检测血管生成素mRNA的表达,同时应用免疫组化法检测血管生成素蛋白在人毛囊中的表达。在此基础上,在体外培养的人毛囊中加入不同浓度的重组人血管生成素（0 ～ 200 ng/mL）,培养6 d后测量毛囊的生长长度。利用两步酶法分离培养人毛乳头细胞,加入不同浓度的重组人血管生成素（0 ～ 200 ng/mL）,48 h后,MTT法检测细胞增殖,流式细胞仪检测细胞周期。结果 RT-PCR显示人毛囊表达血管生成素mRNA,免疫组化法发现人毛囊的毛乳头和真皮鞘表达血管生成素蛋白。25 ～ 200 ng/mL重组人血管生成素呈浓度依赖性促进体外培养的人毛囊生长（P < 0.05）,而12.5 ～ 200 ng/mL重组人血管生成素能够明显促进体外培养的人毛乳头细胞增殖（P < 0.05）。流式细胞仪检测12.5 ～ 200 ng/mL重组人血管生成素能够显著增加S期细胞比率和细胞增殖指数（P < 0.05）。结论 血管生成素可能是一种促进毛发生长的因子。     

Androgens and hair growth

Hair's importance in human communication means that abnormalities like excess hair in hirsutism or hair loss in alopecia cause psychological distress. Androgens are the main regulator of human hair follicles, changing small vellus follicles producing tiny, virtually invisible hairs into larger intermediate and terminal follicles making bigger, pigmented hairs. The response to androgens varies with the body site as it is specific to the hair follicle itself. Normally around puberty, androgens stimulate axillary and pubic hair in both sexes, plus the beard, etc. in men, while later they may also inhibit scalp hair growth causing androgenetic alopecia. Androgens act within the follicle to alter the mesenchyme-epithelial cell interactions, changing the length of time the hair is growing, the dermal papilla size and dermal papilla cell, keratinocyte and melanocyte activity. Greater understanding of the mechanisms of androgen action in follicles should improve therapies for poorly controlled hair disorders like hirsutism and alopecia.     

Plasticity of hair follicle dermal cells in wound healing and induction

The capacity of adult hair follicle dermal cells to participate in new follicle induction and regeneration, and to elicit responses from diverse epithelial partners, demonstrates a level of developmental promiscuity and influence far exceeding that of interfollicular fibroblasts. We have recently suggested that adult follicle dermal cells have extensive stem or progenitor cell activities, including an important role in skin dermal wound healing. Given that up to now tissue engineered skin equivalents have several deficiencies, including the absence of hair follicles, we investigated the capacity of follicle dermal cells to be incorporated into skin wounds; to form hair follicles in wound environments; and to create a hair follicle-derived skin equivalent. In our study, we implanted rat follicle dermal cells labelled with a vital dye into ear and body skin wounds. We found that they were incorporated into the new dermis in a manner similar to skin fibroblasts, but that lower follicle dermal sheath also assimilated into hair follicles. Using different combinations of follicle dermal cells and outer root sheath epithelial cells in punch biopsy wounds, we showed that new hair follicles were formed only with the inclusion of intact dermal papillae. Finally by combining follicle dermal sheath and outer root sheath cells in organotypic chambers, we created a skin equivalent with characteristic dermal and epidermal architecture and a normal basement membrane - the first skin to be produced entirely from hair follicle cells. These data support the hypothesis that follicle dermal cells may be important in wound healing and demonstrate their potential usefulness in human skin equivalents and skin substitutes. While we have made progress towards producing skin equivalents that contain follicles, we suggest that the failure of cultured dermal papilla cells to induce follicle formation in wounds illustrates the complex role the follicle dermis may play in skin. We believe that it demonstrates a genuine dichotomy of activity for follicle cells within skin.     

Immune privilege in hair growth.

Immunostaining techniques were used to investigate the relationship between immune cells, proteoglycan, and class I MHC distribution in skin during the hair cycle in rats. The growth stage, anagen, was characterized by absence of class I MHC staining on most cells of the lower follicle and presence of chondroitin proteoglycan in the follicle sheath and dermal papilla. Immune cells were few in number and not associated with follicles. Dramatic changes were observed during regression in catagen; class I MHC was expressed on all follicle epithelium, large numbers of activated macrophages aggregated around the follicles, and the chondroitin proteoglycans disappeared from the follicle sheath and dermal papilla. During the resting stage, telogen, class I MHC remained on cells of the secondary germ, but macrophages and chondroitin proteoglycans were absent. These observations lead us to propose a hypothesis of immune privilege in hair growth.     

Hair follicle stem cell marker nestin expression in regenerating hair follicles of patients with alopecia areata

Cells that are nestin positive and keratin 15 (K15) negative are located in the hair follicle pluripotent stem cell (hfPS) area (hfPSA). The hfPSA is located within the root of the sebaceous glands, in a region just above the hair follicle bulge area. In the current study, we investigated the expression pattern of the stem cell marker nestin in the hair follicle cycling of patients with alopecia areata. In the normal human scalp, the majority of hair follicles are in the anagen phase of development. While it is often difficult to identify nestin expression in late anagen phases, nestin-expressing cells are easily identified in proliferating cells located in the hfPSA of the growing early and middle anagen phase hair follicles. In patients exhibiting alopecia areata, the middle anagen hair follicles with growing cells were found to be nestin positive and K15 negative. In contrast, the hair follicles undergoing degradation in alopecia areata patients demonstrated lymphocytic infiltration within the nestin- and K15-negative dermal papilla cells. Both the nestin-positive hfPSA and K15-positive hair follicle bulge areas were not damaged in all phases. In addition, the regenerating early anagen hair follicles demonstrated nestin-positive and K15-negative cells within the dermal papilla and in the area surrounding the hair bulb. These results suggest that the nestin-positive cells play an important role not only in the hfPSA, but also in the dermal papilla in the regenerating hair follicle.     

A comparison of the culture and growth of dermal papilla cells from hair follicles from non-balding and balding (androgenetic alopecia) scalp

Male pattern baldness is a common, androgen-dependent skin problem in adult men which is not well understood, although androgens are believed to act on the hair follicle via the mesenchyme-derived dermal papilla situated in the middle of the hair follicle bulb. Since dermal papilla cells retain specific characteristics in culture, such as hair-growth promoting ability and appropriate features of the mechanism of androgen action, dermal papilla cells from follicles undergoing androgen-stimulated miniaturization may provide a useful in vitro model system. Therefore, dermal papilla cells have been derived from intermediate follicles from balding and nearly clinically normal sites of men with androgenetic alopecia. Balding dermal papillae were much smaller than non-balding ones and grew much less well under normal growth conditions. Supplementing the medium with human serum, rather than fetal calf serum, increased both the yield of established cultures and the number and health of the dermal papilla cells produced. Non-balding cells also grew faster in human serum. Balding cells retained the normal fibroblastic shape and aggregative behaviour of dermal papilla cells, but always grew less well than non-balding cells. Nearly clinically normal dermal papillae were similar, or slightly smaller, in size to non-balding ones, but their growth resembled balding cells. Since balding dermal papilla cells can be cultured, though with much greater difficulty than non-balding ones, and exhibit differing growth characteristics to non-balding cells, they merit further investigation which may Increase our understanding of. and ability to control, androgenetic alopecia.     

In vivo cytokine and receptor gene expression during the rat hair growth cycle

Abstract A number of cytokines have previously been localised within the developing and adult hair follicle, however, the role they play in producing a mature hair follicle remains unknown. In an attempt to identify dermal papilla specific cytokines and thus those that may have an important controlling role, cytokine gene expression profiles, obtained by reverse transcriplase-polymerase chain reaction (RT-PCR), were compared between whole anagen rat hair follicles, passage 2 dermal papillae (a cell type with hair inductive capacity), and footpad fibroblasts (a non-hair inducing cell type). Based on this qualitative data, we were unable to identify a dermal papilla specific gene. The analysis of the pattern and timing of cytokine gene expression during the hair cycle is likely to be more informative. A semi-quantitative RT-PCR technique was therefore developed for studying trends in the level of in vivo expression of the following cytokines and their receptors from early anagen to early catagen in the rat hair growth cycle: insulin-like growth factor I, transforming growth factor β1, tumour necrosis factor, and basic fibroblast growth factor. These genes were found to be differentially expressed and this was correlated with their possible functions in controlling the hair growth cycle, providing valuable insights into the role of cytokines in regulating the hair growth process.     

Ultrastructural abnormalities in the dermal papillae of both lesional and clinically normal follicles from alopecia areata scalps

Summary Alopecia areala is a form of balding whose aetiology is uncertain. Although the dermal papilla in the hair bulb regulates the iollicie and may play a part in the pathogenesis of alopecia areata. Its ultrastructure has not been well described. As clinically normal, i.e. non-balding, follicles from alopecia areata scalps show abnormalities at the light microscope level, it would be expected that they should exhibit the earliest pathological changes involved in the dysfunction of the follicle. This study was designed to investigate the ultrastructure of normal human scalp follicular dermal pap-illae and to see if changes occurred in the ultrastructure of dermal papillae from either lesional or non-balding regions of alopecia areata. Normal dermal papillae contained well formed fibroblast-like cells with large, oval nuclei and well-developed endoplasmic reticulum; the cells were separated from each other by extracellular matrix containing small pieces of collagen and basal lamina-like material. Dermal papille from both clinically normal and lesional alopecia areata follicles were less well organized and the dermal papilla cells exhibited signs of cell injury and contained abnormal amounts of pigment; an increased concentration of fibrous material in the extracellular matrix and thickening of the dermal papilla-epithelial junction were also seen. Follicles from lesional areas showed more pronounced changes than clinically normal ones. Ultrastructural abnormalities in the dermal papillae of clinically normal scalp follicles support the study of these follicles as a prime research target. The changes detected suggest that dermal papilla cells in alopecia areata would be less able to synthesize regulatory factors and that these may have more difficulty crossing into the epithelial compartment. They are consistent with an early pathological role for the dermal papilla in alopecia areata, but do not distinguish whether this is a primary aetiological role or a secondary response to an insult elsewhere in the follicle.     

Effect of androgens on the growth of cultured human dermal papilla cells derived from beard and scalp hair follicles

Androgens stimulate hair growth in some areas, e.g., beard, but may cause regression and baldness on the scalp. The mesenchyme-derived dermal papilla is believed to regulate many aspects of hair growth. It is probable that androgens exert their effect on hair growth via the dermal papilla. In this study the effect of androgens on the growth of cultured dermal papilla cells from beard and non-balding scalp was assessed. Dermal papilla cells from beard hair follicles and non-balding scalp were cultured in vitro in the presence and absence of different concentrations of testosterone or the synthetic, non-metabolizable androgen, mibolerone. Cell growth was reflected by the incorporation of 3H-thymidine. The presence of either androgen did not significantly alter DNA synthesis at any of the concentrations examined in either type of cell line. These results do not mean that dermal papilla cells do not respond to androgens in vitro, but that the measurement of cell growth is an inappropriate method of assessment. Androgens may well stimulate the synthesis of specific proteins that could influence the hair follicle.