Fascinating Biology of the Hair Follicle

Chapter 01 Fascinating Biology of the Hair Follicle F Bahar Sunay* and Sema Serter Koçoğlu Balikesir University Faculty of Medicine Department of Histology and Embryology, Turkey * Corresponding Author: F Bahar Sunay, Balikesir University Faculty of Medicine Department of Histology and Embryology, Balikesir, Turkey, Tel: +90 266 612 10 10/6879; Email: fbsunay@gmail.com First Published April 23, 2018 Copyright: 2018 F Bahar Sunay and Sema Serter Kocoglu. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source. 2 www.avidscience.com

Abstract Hair is an important feature of human and hair shaft performs different and important functions. The mini organ responsible from the production of the hair is the hair follicle. Both the structure of the hair follicle and the process of hair production are complicated and there are still unclear issues to be explained. Hair follicle is composed of three different regions. The upper infundibulum and isthmus are the permanent regions, but the lowermost part the hair bulb is regularly renewed. Embryological development of the hair follicle starts with the down growth of the surface epithelium and needs complex relations between the epithelium and underlying mesenchyme which are mostly provided by different genes and the signaling pathways. After its development completed the mature hair follicle enters into a regression and reneawal cycle which consists of anagen, katagen, telogen and exogen stages. Understanding the structure, development and renewal of the hair follicle will help the development of the new treatment methods for the hair diseases. In this chapter general features of the hair follicle, its function and, the genes and signaling pathways playing important roles for its structure and function will be presented. Hair is one of the primary characteristic features of both the male and female appearance. Hair shafts are responsible from different functions such as regulation of the body heat, collection of sensory information, protection against the trauma and hair is necessary for social communication. The estimated number of the hair follicles in an adult human is 5 million and approximately 80,000-150,000 of them are located on the scalp [1]. Despite the fact that the hair follicle has a favorable structure for the scientific examination, the details of its developmental stages and how they are regulated are still not fully enlightened. The first study about the hair was done in 1876 by a German scientist from Hamburg, Paul Gerson Unna [2,3]. The other scientist, who is particularly www.avidscience.com 3

known for his works about the developmental biology of the hair follicles, is Stöhr [3]. Engman, who is the pupil of Unna, had started the studies about the embryological development of the hair follicles and his studies were proceeded and improved by the studies of the Danforth, Trotter and Candy in 1925, Pinkus in 1958, Sengel in 1976 and the Spearman in 1977 [2]. Morphology of Human Hair The hair follicle is composed of three parts: the lower part containing the hair bulb and the hair supra bulb, the middle part containing the isthmus and the upper part containing the infundibulum (Figure-1A). In a mature, anagen hair follicle, the upper and the middle parts form the permanent, nonrenewable parts of the hair and, the lower part is the regularly renewed region. The uppermost part of the hair follicle is the infundibulum. Infundibulum is the region located between the surface opening of the hair follicle and the level of the opening of its sebaceous gland. This particular localization makes the infundibulum as a part of the pilosebaceous canal, the canal through which the sebum is transferred to the skin surface. A connective tissue layer containing stromal cells and collagen encloses all the epithelial layers of the hair follicle [4,5]. Isthmus is the region located below the infundibulum (Figure- 1B). Isthmus extends from the opening of the sebaceous gland to the level of insertion of the arrector pili muscle. The lower part of the isthmus is a particularly important part of the hair follicle because; the so-called bulge region is located here. The bulge region harbors the epithelial and melanocytic hair follicle stem cells. Arrector pili muscle, which is relevant with the sebaceous gland, is innervated by the sympathetic nervous system. Sebaceous glands brings hair follicles a waterproof and flexible structure by their secretory product; the sebum [6]. Hair bulb is the lowermost part of the hair follicle (Figure-1C). At its base, the bulb is invaginated by a highly vascularized loose con- 4 www.avidscience.com

nective tissue. This connective tissue which enables the vascular and neuronal supply to the hair bulb is named as the dermal papilla. Dermal papilla includes firmly wrapped fibroblasts and it determines the dimensions of the hair bulb, the diameter of the hair shaft and the duration of the anagen phase [5]. The suprabulbar region is located on the upper border of the hair bulb and it extends from this point up to the arrector pili muscle. At the hair bulb, the area around the dermal papilla is known as the hair matrix (HM) and the cells located in this area are the matrix cells or matrix keratinocytes and melanocytes. Melanocytes are the cells responsible from the color of the hair. Coloration of the hair is attributable to the content and type of melanin that the hair contains [4-6]. Matrix keratinocytes, which have emigrated from the bulge region in the isthmus and become activated to generate the hair shaft, start a very fast proliferation and their number determines the dimension of the hair bulb and the diameter of the hair shaft. Matrix cells, when their proliferation is ceased and differentiation is started, form the several cell lineages of the hair shaft and the inner root sheet (IRS) [5]. The outer root sheet (ORS) is originated from different progenitor cells. While the infundibulum, isthmus, bulge and hair bulb regions are originated from the hair follicle epithelium, which is derived from ectoderm layer, dermal papilla region is originated from mesoderm. The hair follicle section contains at least eight different concentric layers (Figure-1D). From the outmost to the innermost these layers are; outer root sheet (ORS), companion layer (CL), inner root sheet (IRS) and hair shaft (HS). IRS comprises three layers; named as Henle s layer, Huxley s layer and the inner root sheet cuticle. Henle s layer forms the outermost part, the Huxley s layer stays in the middle and the inner root sheet cuticle is the innermost part that is located next to the hair shaft. The hair shaft also contains three different layers; medulla layer is located at the center, cortex layers are seen next to it on both sides of the medulla, and the hair cuticle surrounds the cortex and seen on both sides in the sections [5-7]. www.avidscience.com 5

Two different kinds of hair are seen in an adult human; the terminal hairs and the vellus hairs. Terminal hair is the long and coarse hair. Its length may reach a meter or more. The hair located on the scalp and beard is the example of the terminal hair. Vellus hair on the other hand is short and fine. They may only be visible with the aid of a magnifying lens. In infancy, childhood and adult females the body hair is the vellus hair. With the male sex hormones started to be secreted during the puberty, terminal hair replaces the vellus hair at the axillar and pubic regions of the both sexes and almost the whole body of the male. Figure 1: The histological structure of the hair follicle. This figure shows the histological structure of a growing follicle. (A) Sagittal section of the hair follicle showing its different regions. (B) Magnified view of the isthmus region. Sebaceous gland (SG), arrector pili muscle (APM), inner root sheet (IRS), outer root sheet (ORT), hair shaft (HS), basal membrane (BM) and connective tissue sheet (CTS) are seen. (C) Magnified view of the hair bulb. Matrix (M), dermal papilla (DP), CTS, IRS and ORS are seen. (D) Concentric layers of the hair follicle epithelium. From the outside to the inside: ORS, companion layer (CL), IRS and HS. IRS is composed of three layers: 6 www.avidscience.com

Henle s layer, Huxley s layer and IRS cuticle. CL constitutes the border to the Henle s layer. HS is composed of three different regions. Medulla stays in the middle. Cortex surrounds the medulla. Hair cuticle is located at the outmost part of the HS. CTS wrap the hair follicles from the outmost. The figure is taken from the Schneider et al [7]. Embryonical Development of the Hair Follicules and it s Molecular Regulation Hair development starts as a simple down growth of the surface epithelium into the developing subepithelial mesoderm. Hair follicles development starts as early as the 9 th to 12 th weeks of the intrauterine period, but they become distinguishable after the 20 th gestational week. The first event characterizing the development of hair follicles is the proliferative invasion of the germinal epidermal layer into the underlying dermis. Hence, the interaction between the epidermis, which originates from the ectoderm, and the mesenchyme derived from the mesoderm is the most important issue for the hair follicle development. The ectodermal cells in the hair placode concentrate and constitute the hair germs which are the primordial hair follicles. The epithelial cells of the hair germ produce the hair forming germinal matrix. Hair germ is also responsible from the gathering and condensation of the dermal cells. Thickening of the dermal cells results with the further differentiation of the hair germ and, it starts to grow and invaginate into the dermis and produce a rod like structure; the hair peg. Meanwhile, the dermal cells get access into the developing hair follicles through the invagination of the hair bulb and, they start to proliferate to generate the dermal papilla. While the hair follicle stems cells derived from the ectoderm turn into the epithelial components of the hair follicle like the sebaceous glands and the apocrine glands, the mesodermal cells transform into the dermal papilla and the connective tissue sheets surrounding the hair follicle. Meanwhile, neural crest cells give rise to melanocytes, and some of the mesenchyme cells surrounding the hair follicle turn into the arrector pili muscle cells [8,9]. www.avidscience.com 7

The first type of the hair follicle seen in the fetal period is called as lanugo. This type of hair is thinner, smoother and lightly colored as a result of their light pigmentation. They start to be seen at the end of the 12 th gestational week and their numbers increase between the 17 th to 20 th weeks. During the perinatal period normal hairs take place of the lanugo. Figure 2: Intrauterine development of the hair follicle. This schematic drawing shows the basic steps of hair development; namely the induction step during which the placode forms, the organogenesis step during which epidermal hair germ cells proliferate, dermal papilla forms and orientation and the formation of the hair shaft occurs and the cytodifferentiation stage during which the development is completed. Development of the hair follicle can be defined in three basic steps: induction, organogenesis and cytodifferentiation (Figure-2). Induction includes two stages, stage 0 and stage 1. Stage 1 is also known as the placode formation stage. Organogenesis includes four stages, stage 2, known as the germ stage, and stages 3, 4 and 5, known as the peg stage. The cytodifferentiation includes three stages, stages 6, 7 and 8, known as the bulbous peg stage. Each of these stages is regulated by the distinctive molecular signal molecules [5,7]. The main signals which play the key roles on the hair follicle development belong to 8 www.avidscience.com

the Wnt/wingless, hedgehog, transforming growth factor/bone morphogenetic protein (TGF-β/BMP), fibroblast growth factor (FGF) and the tumor necrosis factor (TNF) families [7,10]. These signals have great importance because they control the relationship between the epidermis and dermis and the interaction between the epidermis and dermis is crucial for the development of the hair follicle. The very first initiative signal of the hair follicle development is unknown, but Wnt and β-catenin genes are thought to be the active secondary signals for the beginning of the hair development [6,7,11]. The Wnt gene, which is one of the most important genes expressed by the epidermis, generates the first signal for the loosening of the epithelial cells and induces the mesenchyme cells to produce the placode. The other genes playing stimulant roles in the placode formation are the Wnt10b, EdaA1/EdaR/NF-Ƙb, Noggin/Lef-1 and P-cadherin. Contrary to this, Dkks (Dkk1, 2, and 4), BMPs [2,4,6] and Keratin 17 (KRT17) produce inhibitory effects in the same process [6,7]. Epithelial cells of the newly developed placode start to produce the required signals for the dermal condensation and dermal papilla formation. During the organogenesis step, Shh/Smo/Gli2 and Wnt (10b, 10a)/Lef1 group genes become active and induce the epidermal growth of the hair germ cells. The signals essential for the development and the maturation of the dermal papilla are the signals coming from Shh/Smo/Gli2 and PDGF-A genes. The interplay between the Shh and the Cyclin-D genes helps to the proliferation of the epithelial cells and, the placode starts to proliferate and extend into the underlying mesenchyme. Shh is the basis gene producing signals which help the formation of the hair germ cells and control all the pathways in the organogenesis step. Another factor playing significant roles during the organogenesis step is the Platelet- Derived Growth Factor (PDGF), which has great importance on the mesenchyme and epithelia relations [6,7]. At the cytodifferantiation stage of the hair follicle development; the GATA binding protein-3 (GATA3) is active on the generation of www.avidscience.com 9

the inner root sheet, and the transcription factors MSH Homebox-2 (MSX2) and Homeobox C13 (HOXC13) have important effects on the generation of the hair shaft. The KRT6 and KRT16 proteins, that are synthetized from the Henle s layer of the inner root sheet, play important roles during the differentiation of the outer root sheet [6,12,13]. During the cytodifferentiation step, Notch family genes display activity on the differentiation of the hair follicle stem cells and on the determination of the characteristics of the keratinocytes [14,15]. Wnt gene is an influential signaling pathway in the determination of the epidermis area where the hair follicle will stem [16]. While the Skleraxis (SCX), Microphthalmia-Associated Transcription Factor (MITF), Insulin-Like Growth Factor Binding Proteins (IGFBP5-6), Fibulin-1 (FBLN1), Periostin (POSTN), Tenasin-C (TNC) and FGF18 genes are effective in the differentiation of the arrector pili muscle, Wnt/β-catenin, Shh, LEF1, SMAD7, SMURF2 and KRT6 genes mediate the differentiation of the sebaceous gland [17]. Hair Follicle Cycle In order to produce new hair follicles, mature hair follicles enter into a regression and reneawal cycle. This cycle consists of four different stages namely; anagen, katagen, telogen and exogen stages [4,5]. Anagen is the stage characterized by the rapid growth and lasts between 2 to 6 years. Anagen hair follicles have long and straight appearance and, the cell cycle of their proliferative matrix cells lasts 18 hours on average. The length of the anagen stage determines the length of the hair and, it depends on the ongoing proliferation and the differentiation of the matrix cells located over the follicle base [18]. The main molecules known to control the anagen stage are; Wnt/ beta-catenin signal, BMP antagonists (Noggin), Shh signals, IGF-1, VEGF, HGF, vitamin D receptor, Hairless and retinoic acid receptor [5]. In addition to aforementioned molecules, KRT and KAP genes play roles in the production of the skeletal structure of the hair and, HOXC8 and HOXC9 have effects on the hair shaft and the localiza- 10 www.avidscience.com

tion of the hair follicle during the anagen stage. In general, HOX13, MSX2, Delta, BMP2 and Ntrk are the factors that decelerate the development of the hair follicle in the anagen stage. On the contrary, PDGF, FGF5, Wnt10b, FRZB, TGF-β and Nanog signaling pathways accelerate it [6]. Katagen is the regression stage which provides the transition between the anagen and telogen stages and lasts about 2 or 3 weeks [5,18,19]. During the katagen stage, the cells located in the part of the hair follicles with a lower cyclic activity, meaning the cells located on the lower 2/3 part of the hair follicle, in the bulb, in the IRS and in the ORS undergo apoptosis [20]. With the start of the apoptosis thickening of the hair shaft stops, hair follicle starts to shorten and, the connection between the dermal papilla and epithelia continuous. One of the most important activators of the katagen phase is FGF [7,21]. In mice with a damaged FGF gene, anagen phase is prolonged and this results with the development of the angora type hair. Other factors known to be the activators of the katagen stage are IGF-I, HGF, BCL, IAP, TNFβ1, interleukin-1 beta, neurotrophins (NT3, NT4, and BDNF) and BMP274 [22,23]. The genes which play active role during the apoptosis process or blocking it are BCL and IAP genes. Additionally, TNFβ1 gene expression increases during the katagen stage and it plays role during the apoptosis [23]. As the result of the decreased proliferation rate and biochemical activity of the hair follicle cells during the katagen phase, the telogen stage which lasts about three months and known as the silent stage of the hair cycle starts. The shedding of the old hair follicle finishes, apoptosis goes down, dermal papilla stem cells reach to their niche and the follicle becomes silent. Briefly, telogen phase is a resting phase during which insignificant levels of proliferation, differentiation and apoptosis take place [18]. Even though being a silent stage, important changes occur in the expression levels of some genes like estrogen during the telogen phage [5,24]. In 2008, it has been determined that the telogen phase responds to hair follicle stimulus at two different www.avidscience.com 11

levels, namely the unresponsive stage and the competent stage [5]. This discovery showed that telogen phase is not a silent phase and, it responds in a different way to the hair follicle stimulants. During the telogen phase NF1C signaling activates the Shh, Wnt5a, LEF1 and TGF-β, and the TGF-β1 signaling increases the keratinocyte proliferation. Wnt/β-catenin signaling is the main signaling pathway that leads to the start of the anagen phase [6]. During the transition from the telogen to the anagen phase the follicle stem cells become activated to create a new hair follicle. The cells around the dermal papilla proliferate and a new hair bulb develops. After the hair bulb formation, the IRS/HS differentiation starts [18]. Exogen stage is a newly defined stage during which the shedding of the old hair shaft, the club, occurs. Formerly it was believed that the shedding of the club has occurred passively and with the effects of the mechanical forces. But, recent studies suggest that this stage is also an active stage controlled by different molecular pathways and it is started to be called as the exogen stage. The molecular pathways controlling the exogen stage are unknown at the moment [1,3]. One factor changing the duration of the different stages of the hair cycle is the localization of the hair. Scalp hair follicles for example have an anagen phase longer than 2 years and a telogen phase of a few months. As a result the scalp hair can grow to a great length. Just the opposite is true for the pubic hair, coarse trunk hair, eyelashes and eyebrows. The short anagen phase and long telogen phase of the hair in these areas results in a much shorter hair length [25]. Hair Biology Hair content consists of fusiform keratinized cells. The major part of the hair shaft is formed by the cortex, which is formed by the tightly packed spindle shaped cells. A narrow gap, which is a proteinbased intercellular lamella, forms the cell borders. Cortex specifies the mechanical features of the hair shaft. The medulla region of the hair follicle is composed of the trichohylin protein, glycogen and mela- 12 www.avidscience.com

nosomes and its function is not totally known [2]. Hair cuticle is the protective layer surrounding both the cortex and the medulla and fibers constitute about 10% of its weight. Cuticle itself has three different layers; the A-layer, exocuticle layer and the endocuticle layer. A-layer contains huge amounts of highly sulphated proteins and protects the hair from any kind of physical or chemical damage [2]. The physical properties of the hair shaft are related with its geometrical shape and organization. Cortex is the region responsible from the strength of the hair shaft. It needs a strong cuticle structure in order to resist any kind of mechanical force applied from the outside. Elasticity of the hair is its most important mechanical power. The water content of the hair follicle is very important in terms of its physical and cosmetic features. The ratio of pores in a hair shaft is about 20%, and about 12-18% increase is observed in its weight after it is immersed into the water. Also, wet and straight hair causes more friction than the dry and straight hair and, wet hair provides an excellent electrical transmission [1]. Every hair follicle completes about 10 to 30 cycles during its whole life. Normally, the number of the hair follicles in human does not change after the birth, but the hair fibers undergo some morphological changes [26]. Conclusion Because of its unique behavioral features, the hair follicle is accepted as a mini organ with a unique and complex functioning pattern. Until today, hundreds of genes that play key roles in the development of the hair follicles have been detected and, as with the proceeding research more of them come to been known. But, the very first signaling responsible from the start of the hair follicle morphogenesis is still unknown. As a result, more studies searching for the initiative signal of the hair follicle development are needed. Wnt/β- Catenin signaling is crucial because it generates regulatory effects on the EDA/EDAR/Nuclear Factor Kappa-B (NF-kB) pathways and it plays a key role in the development of the hair follicle placode. But the www.avidscience.com 13

gene or the signaling pathway regulating the Wnt gen and its working mechanism are still not clarified. According to us, this puts the Wnt signaling mechanism to the center of the molecular research about the development of the hair follicles. Hair follicle is also important for being an important and rich source of the stem cells. Stem cells located in the bulge area of the hair follicle has the ability to differentiate into the different cell lineages and are very important for the clinical applications. The reason underlying the hair damage is the application of the false chemical products to it. So having a good knowledge about the structure and the biology of the hair is important for the development of the new methods and treatments for the protection of the hair from different types of chemicals. For instance determination and development of the substances are needed to improve the hair shaft function. Clarification of the molecules and the signaling pathways taking part in the hair follicle morphogenesis and cycle will provide us more knowledge about the skin and hair biology and, will provide new treatment options for the hair defects and disorders. The new information about the biology of the hair follicle will especially offer new treatments for the hair follicle derived diseases like androgenic alopecia, telogen effluvium and hirsutism. References 1. Araujo R, Fernandes M, Cavaco-Paulo A, Gomes A. Advances in Biochemical Engineering/Biotechnology, First Edition. New York: Springer Press. 2011; 121-143. 2. Draelos ZD. The Biology of Hair Care. Dermatol Clin. 2000; 18: 651-658. 3. Schneider MR. The First Description of The Hair Follicle Bulge by Franz Von Leydig. Dermatology. 2011; 223: 29-31. 4. Veijouye SJ, Yari A, Heidari F. Bulge Region as a Putative Hair Follicle Stem Cells Niche : A Brief Review. Iran J Health. 2017; 46: 1167-1175. 14 www.avidscience.com

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17. Niemann C. Differentiation of the Sebaceous Gland. Dermatoendocrinol. 2009; 1: 64-67. 18. Alonso L, Fuchs E. The Hair Cycle. J Cell Sci. 2006; 119: 391-393. 19. Müller-Röver S, Handjiski B, van der Veen C, Eichmüller S, Foitzik K, et al. A Comprehensive Guide For The Accurate Classification of Murine Hair Follicles in Distinct Hair Cycle Stages. J Invest Dermatol. 2001; 117: 3-15. 20. Lindner G, Botchkarev VA, Botchkareva NV, Ling G, van der Veen C, et al. Analysis of Apoptosis During Hair Follicle Regression (catagen). Am J Pathol. 1997; 151: 1601-1617. 21. Hébert JM, Rosenquist T, Götz J, Martin GR. FGF5 as a Regulator of The Hair Growth Cycle: Evidence from Targeted and Spontaneous Mutations. Cell. 1994; 78: 1017-1025. 22. Paus R, Foitzik K. In Search of the & Quot; Hair Cycle Clock & Quot: A Guided Tour. Differentiation. 2004; 72: 489-511. 23. Stenn KS, Paus R. Controls of Hair Follicle Cycling. Physiol Rev. 2001; 81: 449-494. 24. Lin KK, Chudova D, Hatfield GW, Smyth P, Andersen B. Identification of Hair Cycle-Associated Genes From Time- Course Gene Expression Profile Data By Using Replicate Variance. Proc Natl Acad Sci U S A. 2004; 101: 15955-15960. 25. Young B, O Down G, Woodford P. Wheater s Functional Histology, Sixth edition. London: Churchill Livingstone Press. 2013; 169-171. 26. Goodier M, Hordinsky M. Normal and Aging Hair Biology and Structure Aging and Hair. Curr Probl Dermatol. 2015; 47: 1-9. 16 www.avidscience.com