成纤维细胞在皮肤衰老中的作用机制研究进展

在过去的20余年间,对皮肤衰老机制的探索是一个生命科学研究领域的热门话题。皮肤真皮中最重要的细胞成分是成纤维细胞,它与组织创伤修复密切相关。皮肤衰老与成纤维细胞数量减少、形态改变、分泌合成功能减弱或衰退等有关。研究成纤维细胞在皮肤衰老中的作用机制,包括ROS-PTEN-PI3K、Smad3-I型胶原蛋白、miRNAs等,为研究预防和延缓皮肤衰老提供新的思路以及指导意义。该文就成纤维细胞在皮肤衰老中的作用机制进行综述。     

皮肤衰老与微小RNA

皮肤衰老是机体衰老的表现之一,由于表皮和真皮内细胞结构、功能以及细胞外基质组分的变化所致.微小RNA是一组内源性非编码小分子RNA,研究表明,微小RNA与表皮、真皮的衰老以及紫外线诱导的皮肤衰老相关.在真皮中,微小RNA可以靶向作用于细胞外基质组分和细胞黏附分子,或调控细胞周期、端粒酶活性、细胞内信号通路及氧化应激等影响成纤维细胞的衰老.在表皮中,微小RNA可通过染色质重塑和p63途径参与角质形成细胞的衰老,或靶向作用于转化生长因子β依赖或非依赖的途径影响朗格汉斯细胞的衰老.而在紫外线诱导的皮肤衰老中,微小RNA在组蛋白甲基化、细胞周期调控因子以及转录激活因子等层面参与皮肤衰老过程.此外,有一些微小RNA如微小RNA 125b参与皮肤衰老的机制可能与表皮干细胞相关.     

皮肤衰老和细胞衰老

皮肤衰老是机体衰老的重要外在表现之一,已引起老年学和皮肤科学的重视。体外的皮肤细胞衰老可部分反映机体皮肤老化状况同时角质形成细胞在皮肤衰老中的作用也逐渐引起人们的重视,因为表皮角质形成细胞在皮肤衰老过程中的重要作用如同真皮成纤维细胞一样日益受到关注,加速有关皮肤衰老的研究可为阐明皮肤衰老机制、为老年皮肤病学的发展提供科学依据,进一步指导临床用药和皮肤疾病的防治。     

由银屑病皮肤培养的成纤维细胞活动过强:1,较快的增殖和去除血清的作用

真皮中的成纤维细胞负责维持真皮物质以及真皮的许多正常与异常性状.细胞培养允许人们在控制的条件下细致研究细胞的代谢,对病人也较为方便.本文比较了13株由寻常性银屑病皮损(PSA)、16株由患者未受累皮肤(PSB)和13株相近年龄正常人皮肤(NSF)中分离的成纤维细胞株的繁殖情况.     

体外培养毛囊真皮鞘细胞与皮肤成纤维细胞形态结构和bFGF、EGFR表达差异

Gharzi等[1]初步证实毛囊真皮鞘细胞参与皮肤损伤修复,但相关作用机制研究报道不多.成纤维细胞是皮肤创伤愈合主要效应细胞,毛囊真皮鞘细胞与皮肤成纤维细胞一样均属于间充质细胞.本研究旨在通过两种细胞形态结构及部分愈合相关蛋白表达差异研究,初步探讨毛囊真皮细胞在创伤愈合中的作用.     

皮肤间充质干细胞在促进皮肤愈合中的作用

间充质干细胞(mesenchymal stem cells,MSCs)是当前干细胞研究的热点之一。目前,皮肤愈合正逐渐受到重视。现有的研究认为骨髓间充质干细胞(BM-MSCs)能从多个方面促进皮肤愈合,如促进表皮生长、促进真皮成纤维细胞的增生等。皮肤间充质干细胞(SMSCs)和BM-MSCs均为MSCs,具有很多的相似性,且SMSCs较BM-MSCs更容易得到。所以可从目前对BM-MSCs的研究预测到SMSCs在皮肤创伤愈合中的研究前景,且将来很可能会替代BM-MSCs。     

血管内皮细胞对成纤维细胞增殖活性的影响

目的 观察内皮细胞对皮肤成纤维细胞增殖活性的影响。方法 对培养脐静脉内皮细胞，收集内皮细胞条件培养，利用＾３Ｈ－ＴｄＲ掺入的方法检测内皮细胞条件培养液对正常人真皮成纤维细胞ＤＮＡ合成的影响，利用流式细胞仪分析内皮细胞条件培养液对正常人真皮成纤维细胞细胞周期的影响。结果 脐静脉内皮细胞条件培养液能显著增加皮肤成纤维细胞ＤＮＡ合成，细胞周期分析显示，Ｓ期成纤维细胞的比例显著增加。结论 内皮细胞通过分泌     

8-MOP/UVA诱导培养真皮成纤维细胞衰老的作用

目的　探讨以 8 MOP/UVA作用于培养真皮成纤维细胞建立皮肤光老化模型的可行性。方法　采用光镜、电镜、流式细胞仪、酶组织化学、免疫组织化学等方法检测 8 MOP/UVA对培养真皮成纤维细胞多项细胞衰老相关指标的影响。结果　 8 MOP/UVA作用后 ,培养真皮成纤维细胞迅速出现细胞衰老特征性的形态学及生物学特性改变 :细胞由分裂表型转化为无分裂活性表型 ,SA β Gal表达增加、p16蛋白表达增加。结论　 8 MOP/UVA可诱导培养真皮成纤维细胞迅速出现具有细胞衰老特征性的形态学及生物学特性改变 ,以 8 MOP/UVA作用于培养真皮成纤维细胞建立皮肤光老化模型是可行的     

小鼠真皮间充质干细胞与成纤维细胞共培养对胶原分泌和TGF-β1表达的影响

目的 探讨真皮间充质干细胞在皮肤组织修复中的作用.方法 采用低血清培养基,消化-贴壁-传代法体外培养、鉴定小鼠真皮间充质干细胞(mdMSC),并与体外分离培养的正常人皮肤成纤维细胞于transwell小室培养体系中共培养,样本碱水解法和ELISA法分别检测第4、8天培养上清液中羟脯氨酸和TGF-β1的变化.结果 共培养第8天,经mdMSC 2.5×104和mdMSC 1×104处理的正常人皮肤成纤维细胞培养上清液中羟脯氨酸含量较单独培养时明显增高(P<0.05).经mdMSC处理的各组正常人皮肤成纤维细胞培养上清液中TGF-β1含量于共培养第8天时均高于单独培养(P<0.01);经mdMSC 1×104处理的正常人皮肤成纤维细胞培养上清液中TGF-β1含量在第4天亦高于单独培养,差异有统计学意义(P<0.05).各不同细胞密度的MSC处理组的羟脯氨酸含量与TGF-β1水平无相关关系(r=0.108,P＞0.05).结论 mdMSC与正常人皮肤成纤维细胞共培养可增加羟脯氨酸和TGF-β1的分泌,可能是mdMSC促进皮肤组织修复的机制之一.     

成纤维细胞自体填充移植研究进展

成纤维细胞是真皮主要功能细胞,在皮肤抗衰老过程中起重要作用.基础研究表明,自体成纤维细胞移植具有高度的细胞存活率和良好的安全性.移植的微环境以及其他细胞、基因、蛋白、细胞因子等因素可影响成纤维细胞的活性和功能.临床研究表明,自体成纤维细胞填充移植可有效改善皱纹、治疗凹陷性瘢痕、溃疡等损害.     

诱导多潜能干细胞在皮肤病治疗中的应用

诱导多潜能干细胞拥有胚胎干细胞所有特征,包括多能性和生成各种体细胞.用皮肤细胞产生诱导多潜能干细胞,不仅起始细胞易获取,而且这些诱导多潜能干细胞更容易定向分化为角质形成细胞、黑素细胞和成纤维细胞等多种功能性皮肤细胞.患者自体来源的诱导多潜能干细胞是细胞疗法理想的细胞库,用诱导多潜能干细胞分化后的细胞治疗皮肤病,不仅细胞量充足,且可避免伦理问题和免疫排斥反应.利用回复突变体嵌合体,结合诱导多潜能干细胞技术,能获得充分的患者特异性功能性回复体细胞而用于治疗遗传性皮肤病.该技术可避免常规基因治疗中出现的免疫排斥和插入诱变.     

Stem cells and the skin

Stem cells live long lives, renew themselves, and differentiate into more mature, less potent, specialized cells, such as epidermal keratinocytes and dermal fibroblasts. Stem cells can be embryonic, if derived from an embryo, or adult/somatic if derived from postembryonic tissue. By producing new skin cells, stem cell division and differentiation can potentially rejuvenate skin and restore hair. To reproduce, stem cells can undergo symmetric nondifferentiative or differentiative divisions, or asymmetric differentiative divisions. Asymmetric divisions reproduce the stem cell and provide a more differentiated, but less potent transient amplifying cell. Divisions and differentiation of transient amplifying cells regenerate tissues by producing cells of a specific lineage, for example, keratinocytes. Epidermal stem cells lie in niches in the interfollicular epidermis, sebaceous gland, and in the bulge regions of hair follicles. These epidermal stem cells renew the epidermis, the sebaceous glands, and hair follicles after mature cells die. Dermal stem cells lie in the hair papillae, around pericytes, and elsewhere among other dermal cells. These form pericytes, myoblasts, fibroblasts, chondrocytes, and other specialized dermal cells. Along with other signaling pathways, the Wnt signaling pathway controls stem cell fate. Wnt signals enlist two functionally and chemically different gene coactivators to direct the time and type of replicative divisions. Stem cells may help to heal wounds, repair damaged tissues, regenerate aged skin, and reinvigorate growth of skin, hair, nails, and mucous membranes.     

Papillary fibroblasts differentiate into reticular fibroblasts after prolonged in vitro culture

The dermis can be divided into two morphologically different layers: the papillary and reticular dermis. Fibroblasts isolated from these layers behave differently when cultured in vitro. During skin ageing, the papillary dermis decreases in volume. Based on the functional differences in vitro, it is hypothesized that the loss of papillary fibroblasts contributes to skin ageing. In this study, we aimed to mimic certain aspects of skin ageing by using high‐passage cultures of reticular and papillary fibroblasts and investigated the effect of these cells on skin morphogenesis in reconstructed human skin equivalents. Skin equivalents generated with reticular fibroblasts showed a reduced terminal differentiation and fewer proliferating basal keratinocytes. Aged in vitro papillary fibroblasts had increased expression of biomarkers specific to reticular fibroblasts. The phenotype and morphology of skin equivalents generated with high‐passage papillary fibroblasts resembled that of reticular fibroblasts. This demonstrates that papillary fibroblasts can differentiate into reticular fibroblasts in vitro. Therefore, we hypothesize that papillary fibroblasts represent an undifferentiated phenotype, while reticular fibroblasts represent a more differentiated population. The differentiation process could be a new target for anti‐skin‐ageing strategies.     

Enhanced proliferation and collagen synthesis of human dermal fibroblasts in chronically photodamaged skin

Cutaneous aging can be divided into intrinsic aging and photoaging. We investigated the influence of aging and photoaging on the proliferation and collagen synthesis of human dermal fibroblasts cultured 3-dimensionally in a collagen gel. We examined 11 human dermal fibroblast cell lines cultured from 3 newborn skins (1 day old), and both exposed and unexposed skin from 4 elderly volunteers (60, 60, 73, 76 years old), respectively. Newborn fibroblasts actively proliferated within the attached collagen gels compared with the elderly cell lines. Within the attached collagen gels in the presence of 10% fetal calf serum (FCS), the fibroblasts from exposed skin proliferated rapidly compared with fibroblasts from unexposed skin from the same individuals. In collagen gel and monolayer cultures with 1% FCS, the percentage of collagen synthesized by photoaged and aged fibroblasts decreased significantly compared with that by newborn fibroblasts. When the fibroblasts were cultured three dimensionally in attached collagen gels in the presence of 1% FCS, the relative levels of collagen synthesis by cultured fibroblasts from photoaged skin were increased significantly compared with those of aged skin fibroblasts from the same individuals. These results suggest that fibroblasts of exposed skin may be more active than those of unexposed skin and that the three-dimensional culture of fibroblast can be used as a model to investigate the influence of aging and photoaging on cell functions.     

Oxytocin alleviates cellular senescence through oxytocin receptor-mediated extracellular signal-regulated kinase/Nrf2 signalling

Intrinsic skin aging is skin aging affected by natural factors within the body, such as hormones, rather than by external factors like UV light or smoking. Skin ageing causes the appearance of wrinkles, skin sagging and reduced elasticity. Like elsewhere in the body, skin ageing involves an increase in the proportion of cells undergoing cellular senescence (a type of deterioration due to age). Senescent skin cells called dermal fibroblasts have several characteristics that are different to cells that are not senescent, including having what is known as a senescence-associated secretory phenotype (SASP). Oxytocin (OT) is a neuropeptide hormone that has many beneficial effects in the body, including protection against age-related disorders. However, less is known about the role of OT in intrinsic skin aging. The authors looked into the role of oxytocin in prevention against senescence in skin cells called normal human dermal fibroblasts (NHDFs), taken from female donors of different ages. They found that OT suppressed SASP-induced senescence in NHDFs derived from young but not old female donors. The authors were also able to learn more about the processes within the body by which OT suppresses SASP and why it affects younger women differently to older women. Their results demonstrate that oxytocin may be effective in the prevention of skin aging.     

Generation of melanocytes from induced pluripotent stem cells

Epidermal melanocytes have an important role in protecting skin from UV rays, and are implicated in a variety of skin diseases. Here, we developed an efficient method for differentiating induced pluripotent stem cells (iPSCs) into melanocytes. We first generated iPSCs from adult mouse tail-tip fibroblasts (TTFs) using retroviral vectors or virus-free piggyBac transposon vectors carrying murine Sox2, Oct3/4, c-Myc, and Klf4. The TTF-derived iPSC clones exhibited similar morphology and growth properties as mouse embryonic stem (ES) cells. The iPSCs expressed ES cell markers, displayed characteristic epigenetic changes, and formed teratomas with all three germ layers. The iPSCs were used to generate embryonic bodies and were then successfully differentiated into melanocytes by treatment with growth factors. The iPSC-derived melanocytes expressed characteristic melanocyte markers and produced melanin pigment. Electron microscopy showed that the melanocytes contained mature melanosomes. We manipulated the conditions used to differentiate iPSCs to melanocytes and discovered that Wnt3a is not required for mouse melanocyte differentiation. This report shows that melanocytes can be readily generated from iPSCs, providing a powerful resource for the in vitro study of melanocyte developmental biology and diseases. By inducing iPSCs without viruses, the possibility of integration mutagenesis is alleviated, and these iPSCs are more compatible for cell replacement therapies.     

毛乳头细胞体外诱导毛囊上皮细胞分化的实验研究

目的:探讨间质细胞对毛囊上皮细胞分化的调节作用,研究毛囊上皮细胞的分化特性。方法:分别用团块状的毛乳头细胞、皮肤成纤维细胞制成间质细胞胶原凝胶,表面接种毛囊上皮细胞,进行气-液界面培养。结果:毛囊上皮细胞有向毛乳头细胞移动集结的趋势;团块状的毛乳头细胞诱导毛囊上皮细胞形成球形结构;皮肤成纤维细胞诱导毛囊上皮细胞形成表皮样层化结构。结论:(1)毛囊上皮细胞具有双向分化特性,它既能分化形成毛囊,也能分化形成表皮结构;(2)毛乳头细胞对毛囊上皮细胞有趋化作用;(3)间质细胞的种类及分布在毛囊上皮细胞分化的调节中起着重要的作用。     

Towards the development of a pragmatic technique for isolating and differentiating nestin‐positive cells from human scalp skin into neuronal and glial cell populations: generating neurons from human skin?

Nestin+ hair follicle-associated cells of murine skin can be isolated and differentiated in vitro into neuronal and glial cells. Therefore, we have asked whether human skin also contains nestin+ cells, and whether these can be differentiated in vitro into neuronal and/or glial cell populations. In this methodological pilot study, we show that both are indeed the case - employing purposely only very simple techniques for isolating, propagating, and differentiating nestin+ cells from normal human scalp skin and its appendages that do not require selective microdissection and tissue compartment isolation prior to cell culture. We show that, it is in principle, possible to maintain and propagate human skin nestin+ cells for extended passage numbers and to differentiate them into both neuronal (i.e. neurofilament+ and/or PGP9.5+) and glial (i.e. GFAP+, MBP+ and/or O4+) cell populations. Therefore, human scalp skin can serve as a highly accessible, abundant, and convenient source for autologous adult stem cell-like cells that offer themselves to be exploited for neuroregenerative medicine purposes.