毛囊真皮鞘细胞中过表达Noggin对皮肤及毛囊生长的作用

背景:Noggin蛋白是一个抑制BMP信号通路的重要分子,可以与BMP2/4结合形成复合物,从而阻断BMP信号,影响生物有机体的正常发育过程。研究认为真皮鞘细胞是一类能够长期自我更新的真皮干细胞,参与毛囊再生过程中真皮毛乳头和真皮鞘的形成。在毛囊发育过程中,真皮毛乳头中Noggin参与毛囊的起始,Noggin的缺失会导致毛囊数量的减少和毛囊生长缓慢,但人们对于Noggin蛋白在毛囊真皮鞘中的作用所知甚少。目的:研究Noggin蛋白在毛囊真皮鞘中的生物学功能。方法:利用真皮鞘特异的αSMA-Cre ERT2工具鼠在真皮鞘中特异过表达Noggin蛋白,分别在出生后8,9 d对实验组αSMA-CreER;pMES-Noggin小鼠和对照组αSMA-CreER小鼠注射4-羟基他莫昔芬,在出生后21,23,28 d获取皮肤组织,苏木精-伊红染色观察毛囊生长情况和皮下脂肪层厚度,免疫组化染色分析表型。结果与结论:通过苏木精-伊红染色结果发现毛囊生长并没有受到影响,但毛囊皮下脂肪组织生长发育受到严重影响,小鼠皮下脂肪层变薄。免疫组化结果说明BMP信号降低可能是导致毛囊脂肪层变薄的原因。这一发现拓展了人们对于真皮鞘细胞和Noggin蛋白的认识,也为人们更加准确理解毛囊再生和组织生长机制提供了重要依据。     

毛囊细胞--一种新的皮肤组织工程种子细胞

毛囊的上皮细胞和真皮细胞与皮肤的表皮角朊细胞和真皮成纤维细胞具有很大的相似性,但其具有更强的增殖分化能力和更多的生物学特性,并且毛囊真皮细胞具有干细胞的一些特性,作为皮肤组织工程的种子细胞具有更独特的优势,在构建带有皮肤附属器的组织工程皮肤上有潜在的前景.     

毛囊细胞——一种新的皮肤组织工程种子细胞

毛囊的上皮细胞和真皮细胞与皮肤的表皮角朊细胞和真皮成纤维细胞具有很大的相似性，但其具有更强的增殖分化能力和更多的生物学特性，并且毛囊真皮细胞具有干细胞的一些特性，作为皮肤组织工程的种子细胞具有更独特的优势，在构建带有皮肤附属器的组织工程皮肤上有潜在的前景。     

鼠胚胎期和出生后毛囊下段启动再生表达的Wnt5a

背景:Wnt5a是唯一在真皮高表达,与毛囊真皮成分发生密切相关的信号分子。目的:通过观察E12.5-E15.5胎龄的大鼠胚胎皮肤和大鼠触须毛囊上段裸鼠移植诱导毛乳头再生过程Wnt5a的表达,初步研究毛乳头胚胎发生和体外再生的启动机制。方法:将孕SD大鼠在E12.5、E13.5、E14.5、E15.5麻醉后处死,取出完整胚胎制作石蜡切片;另取单个SD大鼠触须毛囊切除毛乳头,毛囊上段拔除毛干后移植至Nu/Nu裸鼠背部皮肤;分别取以上胎龄的大鼠胚胎皮肤及裸鼠背部移植部位的毛囊标本,用酶标免疫组化法观察Wnt5a在大鼠胚胎皮肤及大鼠触须毛囊上段裸鼠移植诱导毛乳头再生过程的表达,苏木精-伊红染色观察大鼠触须毛囊上段移植后毛囊下段的再生情况。动物实验均已获得汕头大学医学院动物伦理委员会批准。结果与结论:①E12.5 Wnt5a在表皮细胞表达;E13.5 Wnt5a在表皮细胞表达增强;E14.5 Wnt5a表达由表皮转移至真皮,在真皮细胞浆明显表达;E15.5 Wnt5a表达在真皮结缔组织明显增强,在表皮的表达消失;②大鼠触须毛囊上段移植后第3天开始至第9天,毛囊上段向下形成新的毛球部,恢复完整毛囊结构,再生过程相关组织Wnt5a均呈强阳性表达;③Wnt5a在胚胎毛囊发生前后和毛囊下段再生模型毛乳头形成的部位呈现高表达,说明Wnt信号通路与毛乳头再生有密切的关系,其中Wnt5a可能扮演了非常主要的角色。     

毛囊形态学发生期的分子调控研究进展

<正>动物的毛囊(hair follicle)是皮肤表皮向真皮下陷而形成的重要附属结构,毛囊的底部可深达皮下组织内,是毛发生长起点。毛囊的生长发育能力很强,而且在动物皮肤中终生成周期性生长变化的器官。毛囊被认为是起源于神经外胚层和中胚层,并且相互作用而形成的微小器官~([1]),毛囊的形态发生起始于早期胚胎阶段,是干细胞研究及系统生物学研究的理想模型~([2])。对于动物来说,其毛发的品质及商品价值的高低主要取决于毛囊的结构和特性,因此,研究毛囊的发育生物学及其分子调控是     

ES细胞源性表皮干细胞与类真皮构建组织工程皮肤的体内分化

目的以ES细胞源性表皮干细胞为种子细胞与类真皮构建组织工程皮肤,探讨其在体内的分化。方法胎鼠皮肤成纤维细胞和大鼠骨髓基质干细胞(BMSCs),分别与复合凝胶-明胶海绵构建类真皮(类真皮Ⅰ、Ⅱ),植入小鼠全层皮肤缺损创面,以生物膜为载体,把羊膜诱导后带有核标记的表皮干细胞覆盖在类真皮上,术后1～8周连续取材,苏木精-伊红染色,β1整合素、CK15、CK19、CK10和CEA免疫荧光双标和免疫组化观察。结果两组组织工程皮肤植入皮肤缺损3～4周后,创面完全长合,较厚新生皮完全覆盖创面,基底层细胞增生,形成短的细胞柱突向真皮层。新生表皮中可见核标记的细胞呈β1整合素、CK15、CK19阳性,真皮中的管腔样结构呈核荧光和CEA免疫组化双标阳性,4～8周新生表皮基底层细胞呈CK19、CK10阳性,新生表皮下可见毛囊样、皮脂腺样结构。结论ES细胞源性表皮干细胞为种子细胞与类真皮构建的两种组织工程皮肤在体内具有修复缺损皮肤及分化为表皮及毛囊样、皮脂腺样和汗腺样等皮肤附属结构的潜能。     

一组毛囊真皮乳头细胞的单克隆抗体

目的　研究真皮乳头细胞 (DP)在毛发的生长和细胞分化中的作用。　方法　细胞培养、单克隆抗体和激光共聚焦显微技术。结果　发现一组能识别毛囊某些特异细胞类型和区域的单克隆抗体 ,其中一个为毛囊干细胞——隆突细胞的标记物。　结论　为鉴定和分离毛囊特异性的细胞群体以及对毛囊周期调控途径和毛囊干细胞的研究提供了基础     

大鼠胚胎背部皮肤基底层β-NGF和p75NTR的表达规律

目的 探讨大鼠胚胎背部皮肤β-NGF和p75NTR的表达规律.方法 使用冰冻超薄切片结合免疫组织化学染色方法和显色半定量统计方法.结果 大鼠胚胎背部皮肤呈现逐渐增厚的趋势,其中毛囊隆突区并不明显,但可明显观察到基底层细胞的表达变化.结论 形态学观察说明毛囊发生初期毛囊干细胞(hair follicle stem cell,HFSC)不仅仅定位于毛囊上段,β-NGF和p75NTR的表达有差异.背部皮肤基底层细胞具有干细胞特性,未分化为表皮细胞或者毛囊中的各种细胞.β-NGF和p75NTR的表达变化可能与毛囊干细胞以及毛囊的生物学特性有关.     

小鼠自然毛囊生长周期及β连环蛋白的表达

目的 观察昆明系小鼠自然毛囊生长周期的组织学特点,探讨β连环蛋白(β-catenin)在小鼠自然毛囊生长周期中的表达规律及其与毛囊生长更替的关系。 方法 采用石蜡切片HE染色、Western blotting以及免疫组织化学方法,研究小鼠（90只）毛囊的周期性变化,检测β-catenin在小鼠毛囊生长不同时期皮肤组织中的蛋白表达水平,并对其进行组织定位。 结果 处于不同生长时期的毛囊具有各期的特殊结构;Western blotting结果显示,在小鼠皮肤组织粗蛋白提取物中存在分子量约为85kD且与兔抗β-catenin多克隆抗体发生免疫阳性反应的蛋白条带;免疫组织化学结果显示,β-catenin在不同毛囊生长时期的表皮、皮脂腺、根鞘和真皮乳头均有不同程度的表达。 结论 昆明系小鼠毛囊生长呈周期性变化,β-catenin在毛囊生长周期中的阳性反应与毛囊的周期性变化存在相关性,表明β-catenin在昆明系小鼠自然毛囊生长周期中起维持、促进毛囊生长的作用。     

表皮干细胞与毛囊干细胞生物学特性的比较

背景：获取更合适的组织工程皮肤种子细胞可使皮肤功能得到更好的修复。 目的：分离、培养表皮干细胞和毛囊干细胞,并比较两种细胞的生物学特性。 方法：体外分离培养2月龄新西兰兔表皮干细胞和毛囊干细胞,取生长良好的第2,3,6代细胞观察其生物学特性。 结果与结论：毛囊干细胞较表皮干细胞贴壁快,具有更高的增殖活性。免疫组化及荧光定量PCR检测显示毛囊干细胞β1整合素、角蛋白19蛋白及mRNA表达均明显高于表皮干细胞。提示作为皮肤组织工程的种子细胞,毛囊干细胞较表皮干细胞更具优势。     

Embryonic development of hair follicle pluripotent stem (hfPS) cells

Our laboratory discovered nestin-expressing hair follicle stem cells and demonstrated their pluripotency. We have shown that nestin-positive and K15-negative multipotent hair follicle stem cells are located above the hair follicle bulge, and we termed these cells hair follicle pluripotent stem (hfPS) cells. We have previously shown that hair follicle stem cells can regenerate peripheral nerve and spinal cord. In the present study, we describe the embryonic development of the hair follicle stem cell area (hfPSCA), which is located above the bulge and below the sebaceous glands in the adult mouse. At embryonic day 16.5 (E16.5) of nestindriven GFP (ND-GFP) transgenic mice, which express nestin in hfPS cells, the ND-GFP hair follicle stem cells are located in mesenchymal condensates. At postnatal day 0 (P0), the ND-GFP-expressing cells are migrating to the upper part of the hair follicle from the dermal papilla. At P3, keratin 15 (K15)-positive cells, derived from ND-GFP dermal papilla cells, are located in the outer-root sheath and basal layer of the epidermis. By P10, the ND-GFP have formed the K15-positive outer-root sheath as well as the ND-GFP hfPSA. These results suggest that ND-GFP hfPS cells in the dermal papilla form nestin-expressing hair follicle stem cells in the first hair cycle. These observations provide new insight into the origins of hfPS cells and the hfPSCA.     

Steady and temporary expressions of smooth muscle actin in hair, vibrissa, arrector pili muscle, and other hair appendages of developing rats

The hair erection muscle, arrector pili, is a kind of smooth muscle located in the mammalian dermis. The immunohistochemical study using an antibody against smooth muscle alpha actin (SMA) showed that the arrector pili muscle develops approximately 1-2 weeks after birth in dorsal and ventral skin, but thereafter they degenerate. The arrector pili muscle was not detected in the mystacial pad during any stage of development, even in the neighboring pelage-type hair follicle. A strong signal of SMA in the skin was located in the dermal sheath as well as in some outer root sheath cells in the hair and vibrissal follicles. Positive areas in the dermal and outer root sheaths were restricted to a lower moiety, particularly areas of similar height, where keratinization of the hair shaft occurs. This rule is valid for both pelage hair follicles and vibrissal follicles. At medium heights of the follicle, SMA staining in the dermal sheath was patchy and distant from the boundary between dermis and epidermis. In contrast to SMA, vimentin was expressed over the entire height of the dermal sheath. Unlike the arrector pili muscle, the expression of SMA in the dermal sheath was observed during fetal, neonatal, and adult stages. The presence of actin-myosin and vimentin fibers in supporting cells is thought to be beneficial for the hair follicle to cope with the movement of the hair shaft, which may be caused by physical contacts with outside materials or by the contraction of internal muscles.     

Comparative characterization of hair follicle dermal stem cells and bone marrow mesenchymal stem cells

We compared the growth and differentiation characteristics of hair follicle-derived dermal stem cells with bone marrow mesenchymal stem cells (MSCs). Follicular dermal cells were isolated from whisker hairs of Wistar rats and bone marrow MSCs were isolated from femora of the same animals. The adherent hair follicle dermal cells showed a fibroblastic morphology in serum-containing culture medium, were CD44(+), CD73(+), CD90(+), and CD34(), and had a population doubling time of 27 h. MSCs isolated from the bone marrow showed a similar morphology and population doubling time and expressed the same cell-surface markers. Following exposure to appropriate induction stimuli, both cell populations had the capacity to differentiate into various mesenchymal lineages, such as osteoblasts, adipocytes, chondrocytes, and myocytes and expressed neuroprogenitor cell markers. The rate and extent of differentiation were remarkably similar for both hair follicleand bone marrow-derived cells, whereas interfollicular dermal cells failed to differentiate. We identified telomerase activity in follicle dermal stem cells and marrow MSCs and demonstrated that they were capable of clonal expansion. In ex vivo analyses, we identified the presence of putative dermal stem cells in the dermal sheath and dermal papillae of the hair follicle. Consequently, the hair follicle may represent a suitable, accessible source for MSCs.     

Nitric oxide in the human hair follicle: constitutive and dihydrotestosterone-induced nitric oxide synthase expression and NO production in dermal papilla cells

The free radical nitric oxide, generated by different types of epidermal and dermal cells, has been identified as an important mediator in various physiological and pathophysiological processes of the skin, such as regulation of blood flow, melanogenesis, wound healing, and hyperproliferative skin diseases. However, little is known about the role of NO in the human hair follicle and in hair cycling processes. Here we demonstrate for the first time that dermal papilla cells derived from human hair follicles spontaneously produce NO by measuring nitrate and nitrite levels in culture supernatants. This biomolecule is apparently formed by the endothelial isoform of nitric oxide synthase, which was detected at the mRNA and protein levels. Remarkably, basal NO level was enhanced threefold by stimulating dermal papilla cells with 5alpha-dihydrotestosterone (DHT) but not with testosterone. Addition of N-[3-(aminomethyl)benzyl]acetamidine (1400W), a highly selective inhibitor of inducible nitric oxide synthase, restrained the elevation in NO level induced by DHT. Analyses of DHT-stimulated cells at the mRNA and protein levels confirmed the expression of inducible nitric oxide synthase. These findings suggest NO as a signaling molecule in human dermal papilla cells and implicate basal and androgen-mediated NO production to be involved in the regulation of hair follicle activity.     

毛发生长周期对小鼠皮肤创伤愈合的影响

目的　探讨处于不同毛发生长周期的C57BL/6小鼠皮肤创伤愈合速度。 方法　制备小鼠皮肤创伤模型,计算术后0、3、7 d创面愈合率,愈合率=（原始创面面积－未愈合的创面面积）/原始创面面积×100%,比较毛发静止期（Hair telogen stages）小鼠和毛发生长期（Hair anagen stages）小鼠伤口愈合速度。采用HE染色比较伤口愈合的组织形态结构差异,利用BrdU检测伤口周围细胞增殖。 结果　毛发生长期的小鼠皮肤伤口愈合率显著高于毛发静止期的小鼠伤口愈合率。HE染色显示毛发生长期小鼠伤口周围表皮细胞层较多,且表皮细胞向伤口迁移增强;BrdU检测显示毛发生长期小鼠皮肤伤口周围表皮BrdU+ 细胞数多于毛发静止期小鼠。 结论　毛发生长期的小鼠皮肤创伤愈合率高于毛发静止期小鼠,这一结果为进一步探讨毛囊在创伤愈合过程中的作用提供研究基础,也为选择皮肤创伤愈合动物模型提供指导。     

Structural changes in hair follicles and sebaceous glands of hairless mice following exposure to sulfur mustard

Sulfur mustard (SM) is a bifunctional alkylating agent causing skin inflammation, edema and blistering. A hallmark of SM-induced toxicity is follicular and interfollicular epithelial damage. In the present studies we determined if SM-induced structural alterations in hair follicles and sebaceous glands were correlated with cell damage, inflammation and wound healing. The dorsal skin of hairless mice was treated with saturated SM vapor. One to seven days later, epithelial cell karyolysis within the hair root sheath, infundibulum and isthmus was apparent, along with reduced numbers of sebocytes. Increased numbers of utriculi, some with connections to the skin surface, and engorged dermal cysts were also evident. This was associated with marked changes in expression of markers of DNA damage (phospho-H2A.X), apoptosis (cleaved caspase-3), and wound healing (FGFR2 and galectin-3) throughout pilosebaceous units. Conversely, fatty acid synthase and galectin-3 were down-regulated in sebocytes after SM. Decreased numbers of hair follicles and increased numbers of inflammatory cells surrounding the utriculi and follicular cysts were noted within the wound 3–7 days post-SM exposure. Expression of phospho-H2A.X, cleaved caspase-3, FGFR2 and galectin-3 was decreased in dysplastic follicular epidermis. Fourteen days after SM, engorged follicular cysts which expressed galectin-3 were noted within hyperplastic epidermis. Galectin-3 was also expressed in basal keratinocytes and in the first few layers of suprabasal keratinocytes in neoepidermis formed during wound healing indicating that this lectin is important in the early stages of keratinocyte differentiation. These data indicate that hair follicles and sebaceous glands are targets for SM in the skin.     

Dynamic signals for hair follicle development and regeneration

Hair follicles form during embryonic development and, after birth, undergo recurrent cycling of growth, regression, and relative quiescence. As a functional mini-organ, the hair follicle develops in an environment with dynamic and alternating changes of diverse molecular signals. Over the past decades, genetically engineered mouse models have been used to study hair follicle morphogenesis and significant advances have been made toward the identification of key signaling pathways and the regulatory genes involved. In contrast, much less is understood in signals regulating hair follicle regeneration. Like hair follicle development, hair follicle regeneration probably relies on populations of stem cells that undergo a highly coordinated and stepwise program of differentiation to produce the completed structure. Here, we review recent advances in the understanding of the molecular signals underlying hair follicle morphogenesis and regeneration, with a focus on the initiation of the primary hair follicle structure placode. Knowledge about hair follicle morphogenesis may help develop novel therapeutic strategies to enhance cutaneous regeneration and improve wound healing.