Yes / Hair fibres show wide diversity across and within all human populations, suggesting that hair fibre form and colour have been subject to much adaptive pressure over thousands of years. All human hair fibres typically have the same basic structure. However, the three-dimensional shape of the entire fibre varies considerably depending on ethnicity and geography, with examples from very straight hair with no rotational turn about the long axis, to the tightly sprung coils of African races. The creation of the highly complex biomaterials in hair follicle and how these confer mechanical functions on the fibre so formed is a topic that remains relatively unexplained thus far. We review the current understanding on how hair fibres are formed into a nonlinear coiled form and which genetic and biological factors are thought to be responsible for hair shape. We report on a new GWAS comparing low and high curl individuals in South Africa, revealing strong links to polymorphic variation in trichohyalin, a copper transporter protein CUTC and the inner root sheath component keratin 74. This builds onto the growing knowledge base describing the control of curly hair formation. / Unilever R&D

No / The hair follicle has an established role in wound re-epithelialisation, a phenomenon that has been appreciated since at least the first half of the last century. The bulge niche, one location of hair follicle epithelial stem cells has been of particular interest to researchers over recent years, with numerous studies showing its ability to directly contribute to epidermal repair. However, recent work has highlighted other progenitor regions of the hair follicle that appear to act as stem cells during epidermal repair. In addition, several studies within the last 12 months have questioned the importance of the bulge during re-epithelialisation, producing conflicting literature. Here we provide a new model to demonstrate how several important differences in experimental design between studies could account for these seemingly opposing findings, which may have implications for how future studies are conducted.

No / It is known that hair growth disorders and hair loss can cause personal distress and affect well‐being. Whilst clinical conditions remain a target for medical research, current research on hair follicle biology and hair growth control mechanisms also provides opportunities for a range of non‐medical and cosmetic interventions that have a modulating effect on the scalp and follicle function. Furthermore, an improvement of the hair fibre characteristics (cuticle structure, cortex size and integrity) could add to the overall positive visual effect of the hair array. Since phytochemicals are a popular choice because of their traditional appeal, this review provides a critical evaluation of the available evidence of their activity for hair benefit, excluding data obtained from animal tests, and offers recommendations on improving study validity and the robustness of data collection in pre‐clinical and clinical studies.

Yes / Background and Objective: Though devices for hair growth based on low levels of light have shown encouraging results, further improvements of their efficacy is impeded by a lack of knowledge on the exact molecular targets that mediate physiological response in skin and hair follicle. The aim of this study was to investigate the expression of selected light-sensitive receptors in the human hair follicle and to study the impact of UV-free blue light on hair growth ex vivo. Material and Methods: The expression of Opsin receptors in human skin and hair follicles has been characterised using RT-qPCR and immunofluorescence approaches. The functional significance of Opsin 3 was assessed by silencing its expression in the hair follicle cells followed by a transcriptomic profiling. Proprietary LED-based devices emitting two discrete visible wavelengths were used to access the effects of selected optical parameters on hair growth ex vivo and outer root sheath cells in vitro. Results: The expression of OPN2 (Rhodopsin) and OPN3 (Panopsin, Encephalopsin) was detected in the distinct compartments of skin and anagen hair follicle. Treatment with 3.2 J/cm2 of blue light with 453 nm central wavelength significantly prolonged anagen phase in hair follicles ex vivo that was correlated with sustained proliferation in the light-treated samples. In contrast, hair follicle treatment with 3.2 J/cm2 of 689 nm light (red light) did not significantly affect hair growth ex vivo. Silencing of OPN3 in the hair follicle outer root sheath cells resulted in the altered expression of genes involved in the control of proliferation and apoptosis, and abrogated stimulatory effects of blue light (3.2 J/cm2; 453 nm) on proliferation in the outer root sheath cells. Conclusions: We provide the first evidence that 1) OPN2 and OPN3 are expressed in human hair follicle, and 2) 453 nm blue light at low radiant exposure exerts a positive effect on hair growth ex vivo, potentially via interaction with OPN3. / This study was supported by the European Marie-Curie Actions Programme, Grant agreement no.: 607886

Light and optical techniques have made a profound impact on modern medicine both in diagnostics and in therapy. Therapeutic action of light is based on photomechanical, photothermal, photochemical and photobiological interactions, depending on the wavelength, power density, exposure time and optical properties of tissue and cells. Last decade experienced a growing rise of commercial devices for management of hair growth, where all of them are based on low levels of light resulting into photobiological, non-thermal interaction of photons with cells, a process that recently has received an official term ‘photobiomodulation’. However, the design and analysis of the reported clinical studies are highly debated in a wider scientific community. The picture is further complicated by a virtual lack of proof about the exact molecular targets that mediate the physiological response of skin and hair follicles (HF) to low levels of light. The goal of this project was to investigate the expression of light-sensitive receptors in the human HF and to study the impact of UV-free blue light on hair growth ex vivo. The expression of Cryptochromes 1 and 2 (CRY1, 2), Opsin 2 and 3 (OPN2 and OPN3), but not other Opsins 1, 4 and 5 was detected in the distinct compartments of skin and anagen HF. Evaluation of the physiological role of detected light-sensitive receptors on hair growth was performed by the modulation of photoreceptors activity in HF ex vivo model. HFs treated with KL001, a stabilizer of CRY1 protein that lengthens the circadian period, delayed HF anagen-catagen transition; while silencing of CRY1 induced premature catagen development accompanied by reduced cell proliferation. Silencing of CRY1 in the HF outer root sheath (ORS) cells in vitro caused downregulation of ii genes involved in the control of proliferation; including the cyclin dependent kinase 6 (CDK6). OPN3 also had a positive effect on metabolic activity and proliferation of the ORS cells in vitro. OPN3 silencing resulted in the altered expression of genes involved in the control of proliferation and apoptosis. Investigated CRY1, OPN2 and 3 greatly absorb in the blue to green-region of the visible spectrum. This led us to investigate the effect of blue light on HF growth. Daily treatment with blue light (453 nm, 3.2 J/cm2, 16 nm full width half maximum) prolonged anagen phase in HF ex vivo that was associated with sustained proliferation. In addition, blue light (3.2 J/cm2) significantly stimulated proliferation of ORS cells in vitro. This effect was abrogated by silencing of OPN3. To summarize, CRY 1, OPN 2 and OPN 3 are expressed in the distinct compartments of the HF, including HF stem cells. Blue light (453 nm) at low radiant exposure exerts a positive effect on hair growth ex vivo, potentially via interaction with OPN3. The further research should be conducted to decipher interactions between blue light and the investigated receptors in the HFs. In addition, the beneficial effect of blue light at low radiant exposure on hair growth raises a possibility of increasing therapeutic efficacy when combined with topical chemistry used for management of hair growth.

Light and optical techniques have made a profound impact on modern medicine both in diagnostics and in therapy. Therapeutic action of light is based on photomechanical, photothermal, photochemical and photobiological interactions, depending on the wavelength, power density, exposure time and optical properties of tissue and cells. Last decade experienced a growing rise of commercial devices for management of hair growth, where all of them are based on low levels of light resulting into photobiological, non-thermal interaction of photons with cells, a process that recently has received an official term ‘photobiomodulation’. However, the design and analysis of the reported clinical studies are highly debated in a wider scientific community. The picture is further complicated by a virtual lack of proof about the exact molecular targets that mediate the physiological response of skin and hair follicles (HF) to low levels of light. The goal of this project was to investigate the expression of light-sensitive receptors in the human HF and to study the impact of UV-free blue light on hair growth ex vivo. The expression of Cryptochromes 1 and 2 (CRY1, 2), Opsin 2 and 3 (OPN2 and OPN3), but not other Opsins 1, 4 and 5 was detected in the distinct compartments of skin and anagen HF. Evaluation of the physiological role of detected light-sensitive receptors on hair growth was performed by the modulation of photoreceptors activity in HF ex vivo model. HFs treated with KL001, a stabilizer of CRY1 protein that lengthens the circadian period, delayed HF anagen-catagen transition; while silencing of CRY1 induced premature catagen development accompanied by reduced cell proliferation. Silencing of CRY1 in the HF outer root sheath (ORS) cells in vitro caused downregulation of ii genes involved in the control of proliferation; including the cyclin dependent kinase 6 (CDK6). OPN3 also had a positive effect on metabolic activity and proliferation of the ORS cells in vitro. OPN3 silencing resulted in the altered expression of genes involved in the control of proliferation and apoptosis. Investigated CRY1, OPN2 and 3 greatly absorb in the blue to green-region of the visible spectrum. This led us to investigate the effect of blue light on HF growth. Daily treatment with blue light (453 nm, 3.2 J/cm2, 16 nm full width half maximum) prolonged anagen phase in HF ex vivo that was associated with sustained proliferation. In addition, blue light (3.2 J/cm2) significantly stimulated proliferation of ORS cells in vitro. This effect was abrogated by silencing of OPN3. To summarize, CRY 1, OPN 2 and OPN 3 are expressed in the distinct compartments of the HF, including HF stem cells. Blue light (453 nm) at low radiant exposure exerts a positive effect on hair growth ex vivo, potentially via interaction with OPN3. The further research should be conducted to decipher interactions between blue light and the investigated receptors in the HFs. In addition, the beneficial effect of blue light at low radiant exposure on hair growth raises a possibility of increasing therapeutic efficacy when combined with topical chemistry used for management of hair growth.

The hair follicle (HF) is a cyclic biological system that progresses through stages of growth, regression and quiescence, each being characterized by unique patterns of gene activation and silencing. MicroRNAs (miRNAs) are critically important for gene silencing and delineating their role in hair cycle may provide new insights into mechanisms of hair growth control and epithelial tissue remodelling. The aims of this study were: 1) To define changes in the miRNA profiles in skin during hair cycle-associated tissue remodelling; 2) To determine the role of individual miRNAs in regulating gene expression programs that drive HF growth, involution and quiescence; 3) and to explore the role of miRNAs in mediating the effects of BMP signalling in the skin. To address Aims 1 & 2, global miRNA expression profiling in the skin was performed and revealed marked changes in miRNAs expression during distinct stages of the murine hair cycle. Specifically, miR-31 markedly increased during anagen and decreased during catagen and telogen. Administration of antisense miR-31 inhibitor into mouse skin during the early- and mid-anagen phases of the hair cycle resulted in accelerated anagen development, and altered differentiation of hair matrix keratinocytes and hair shaft formation. Microarray, qRT-PCR and Western blot analyses revealed that miR-31 negatively regulates expression of Fgf10, the components of Wnt and BMP signalling pathways Sclerostin and BAMBI, and Dlx3 transcription factor, as well as selected keratin genes. Luciferase reporter assay revealed that Krt16, Krt17, Dlx3, and Fgf10 serve as direct miR-31 targets. In addition, miR-214 was identified as a potent inhibitor of the Wnt signalling pathway in the keratinocytes. Mutually exclusive expression patterns of miR-214 and β-catenin was observed during HF morphogenesis. MiR-214 decreases the expression of β-catenin and other components of Wnt signalling pathways c-myc, cyclin D1, and Pten in the keratinocytes. Luciferase reporter assay proved that β-catenin serves as a direct target of miR-214. In addition, miR-214 prevented translocation of β-catenin into the nucleus in response to the treatment with an activator of the Wnt signalling pathway lithium chloride, and abrogated the lithium-induced increase of the expression of the Wnt target gene VI Axin2. This suggests that miR-214 may indeed be involved in regulation of skin development and regeneration at least in part, by controlling the expression of β-catenin and the activity of the Wnt signalling pathway. To address Aim 3, the role of miRNAs in mediating the effects of the bone morphogenetic protein (BMP) signalling in the skin was explored. MiRNAs were isolated from the primary mouse keratinocytes treated with BMP4 and processed for analysis of global miRNA expression using the microarray approach. Microarray and real-time PCR analysis revealed BMP4-dependent changes in the expression of distinct miRNAs, including miR-21, which expression was strongly decreased in the keratinocytes after BMP4 treatment. In contrast, miR-21 expression was substantially higher in the skin of transgenic mice over-expressing BMP antagonist Noggin. Transfection of the keratinocytes with miR-21 mimic revealed existence of two groups of the BMP target genes, which are differentially regulated by miR-21. Thus, this suggests a novel mechanism controlling the effects of BMP signalling in the keratinocytes. Thus, miRNAs play important roles in regulating gene expression programs in the skin during hair cycle. By targeting a number of growth regulatory molecules, transcription factors and cytoskeletal proteins, miRNAs are involved in establishing an optimal balance of gene expression in the keratinocytes required for the HF and skin homeostasis.

<p>Robust regulation of spindle orientation is essential for driving asymmetric cell divisions (ACDs), which generate cellular diversity within a tissue. During the development of the multilayered mammalian epidermis, mitotic spindle orientation in the proliferative basal cells is crucial not only for dictating daughter cell fate but also for initiating stratification of the entire tissue. A conserved protein complex, including LGN, Nuclear mitotic apparatus (NuMA) and dynein/dynactin, plays a key role in establishing proper spindle orientation during ACDs. Two of these proteins, NuMA and dynein, interact directly with astral microtubules (MTs) that emanate from the mitotic spindle. While the contribution of these MT-binding interactions to spindle orientation remains unclear, these implicate apical NuMA and dynein as strong candidates for the machinery required to transduce pulling forces onto the spindle to drive perpendicular spindle orientation. </p><p> In my work, I first investigated the requirements for the cortical recruitment of NuMA and dynein, which had never been thoroughly addressed. I revealed that NuMA is required to recruit the dynein/dynactin complex to the cell cortex of cultured epidermal cells. In addition, I found that interaction with LGN is necessary but not sufficient for cortical NuMA recruitment. This led me to examine the role of additional NuMA-interacting proteins in spindle orientation. Notably, I identified a role for the 4.1 protein family in stabilizing NuMA's association with the cell cortex using a FRAP (fluorescence recovery after photobleaching)-based approach. I also showed that NuMA's spindle orientation activity is perturbed in the absence of 4.1 interactions. This effect was demonstrated in culture using both a cortical NuMA/spindle alignment assay as well as a cell stretch assay. Interestingly, I also noted a significant increase in cortical NuMA localization as cells enter anaphase. I found that inhibition of Cdk1 or mutation of a single residue on NuMA mimics this effect. I also revealed that this anaphase localization is independent of LGN and 4.1 interactions, thus revealing two independent mechanisms responsible for NuMA cortical recruitment at different stages of mitosis. </p><p> After gaining a deeper understanding of how NuMA is recruited and stabilized at the cell cortex, I then sought to investigate how cortical NuMA functions during spindle orientation. NuMA contains binding domains in its N- and C-termini that facilitate its interactions with the molecular motor dynein and MTs, respectively. In addition to its known role in recruiting dynein, I was interested in determining whether NuMA's ability to interact directly with MTs was critical for its function in spindle orientation. Surprisingly, I revealed that direct interactions between NuMA and MTs are required for spindle orientation in cultured keratinocytes. I also discovered that NuMA can specifically interact with MT ends and remain attached to depolymerizing MTs. To test the role of NuMA/MT interactions in vivo, I generated mice with an epidermal-specific in-frame deletion of the NuMA MT-binding domain. I determined that this deletion causes randomization of spindle orientation in vivo, resulting in defective epidermal differentiation and barrier formation, as well as neonatal lethality. In addition, conditional deletion of the NuMA MT-binding domain in adult mice results in severe hair growth defects. I found that NuMA is required for proper spindle positioning in hair follicle matrix cells and that differentiation of matrix-derived progeny is disrupted when NuMA is mutated, thus revealing an essential role for spindle orientation in hair morphogenesis. Finally, I discovered hyperproliferative regions in the interfollicular epidermis of these adult mutant mice, which is consistent with a loss of ACDs and perturbed differentiation. Based on these data, I propose a novel mechanism for force generation during spindle positioning whereby cortically-tethered NuMA plays a critical dynein-independent role in coupling MT depolymerization energy with cortical tethering to promote robust spindle orientation accuracy. </p><p> Taken together, my work highlights the complexity of NuMA localization and demonstrates the importance of NuMA cortical stability for productive force generation during spindle orientation. In addition, my findings validate the direct role of NuMA in spindle positioning and reveal that spindle orientation is used reiteratively in multiple distinct cell populations during epidermal morphogenesis and homeostasis.</p> / Dissertation

O pelo é uma característica dos mamíferos: na maioria das espécies, juntamente com a pele, a pelagem reveste externamente o corpo, com exceção de algumas regiões bem delimitadas. O pelo do cão e demais mamíferos apresenta formato distinto de acordo com a região do corpo. O estudo da obtenção e caracterização de células-tronco em cães poderá ampliar as possibilidades terapêuticas, mediante a utilização das células indiferenciadas na reparação capilar e lesões da pele e seus anexos. Neste trabalho foram utilizadas amostras de folículo piloso retiradas da região rostral de fetos caninos correspondentes a três fases gestacionais distintas (grupo I= 30 dias de gestação; grupo II= 35 dias de gestação; grupo III= 40 dias de gestação).Estas amostras foram cultivadas em placas de Petri de 75 cm² com 5mL de meio de cultivo DMEM, suplementado com 10% de soro fetal bovino (SFB) e 1% de antibióticos estreptomicina/penicilina. As células obtidas foram congeladas e descongeladas e utilizados nos procedimentos de, análise da expressão dos marcadores de células-tronco: STRO-1, CD 117, OCT 3/4, CD 90, Nanog, CD- 34, SSEA-4, CD -105, MCP-1, HSP- 47, CD 1 A, VEGFR1, DR4, IL- 1 &beta;, caspase3, Ki- 67, CD -45. A escolha dos anticorpos marcadores de células-tronco foi baseada em estudos que mostraram afinidade e avidez com o folículo piloso. Utilizamos também para a caracterização do folículo piloso, a imunohistoquímica e análise de PCR com o marcador S100 e a imunocitoquímica com os marcadores OCT 3/4, VEGF, STRO-1, CD117 e Nanog. Os resultados obtidos sugerem que a linhagem celular do folículo piloso de fetos de cães apresentaram crescimento satisfatório com a utilização do meio DMEM-hight suplementado com 10% de SFB inativado e 1% de antibióticos, sendo mantidas e expandidas em cultivo. O crescimento e a expansão das células-tronco do folículo piloso são peculiares, pois, sua expansão ocorre em torno da haste pilosa, utilizando-a como ancoragem. Os resultados indicaram características de pluripotência nas células-tronco do folículo piloso pela expressão dos marcadores OCT3/4, Nanog, CD105, CD90, SSEA-4, e STRO-1, em todos os grupos analisados. Considerando, a taxa de proliferação, a fase do ciclo celular, distribuição e aspectos de morte celular não foram encontradas diferenças significantes na expressão dos marcadores HSP47, DR4. Para verificar a angiogêneseforam utilizados os marcadores VEGFR1, Ki-67 e caspase-3, sendo expressos em todos os grupos. Na imunohistoquímica e análise de PCR observou-se a expressão da proteína S100, sendo maior nas células do grupo de fetos com 40 dias de gestação- grupo III. Neste estudo, podemos concluir que o conjunto de marcadores expressos nos diferentes grupos de células tronco obtidas das vibrissas de fetos caninos indicaram a expressão de marcadores de pluripotência das células-tronco do folículo piloso / Hair is a characteristic of mammals, since in most species the coat spreads throughout the body, except for some well-defined regions. Mammals in general, including dogs, have different characteristics in their coat according to the region of the body. The study of obtainment and characterization of stem cells from the hair follicle could contribute to new therapeutic possibilities, particularly in the treatment of hair, skin and it appendices injuries. In this study, hair follicle samples were harvestedin the rostral region of canine fetuses, divided into three distinct groups, according to the stage of pregnancy, as follows: Group I = 30 days, Group II = 35 days, Group III = 40 days. These samples were grown in 75 cm² Petri dishes with 5 mL of DMEM culture medium supplemented with 10% fetal bovine serum (FBS) and antibiotics 1% streptomycin / penicillin. Cells were frozen and thawed and used in procedures for analysis of expression of markers of stem cells: STRO-1, CD 117, OCT 3/4, CD 90, Nanog, CD- 34,SSEA -4,CD- 105, MCP-1, HSP-47, CD 1 A, VEGF-R1, DR4, IL-1 &beta;, caspase-3, Ki-67, CD -45. The choice of antibodies was based on studies that showed affinity and avidity to the hair follicle. In addition, for purposes of characterization of the hair follicle, procedures for immunohistochemistry and PCR analysis with the marker S100 and immunocytochemistry with the markers OCT3/4, VEGF, STRO-1, CD117 e Nanog were used. The results suggested that hair follicle cell line of fetal dogs showed satisfactory growth using the DMEM high medium supplemented with 10% inactivated FBS and 1% antibiotics and could be maintained and expanded in culture. The growth and expansion of stem cells of the hair follicle are unique because they occur around the hair shaft using it as anchorage. Results also indicated pluripotency features in the hair follicle stem cells through the positive expression of Oct3/4, Nanog, CD105, CD90, SSEA-4 and STRO-1, in all groups. Regarding the proliferation rate and cell cycle phase, distribution and cell death aspects, HSP47, DR4 were used. To verify angiogenesis, VEGFR1, Ki67 and Caspase-3 were expressed in all groups. In Immunohistochemistry procedures we observed the expression of S100, and in mRNA on RT-PCR. We concluded in this study that in group III the expression of S100 was higher than in the two other groups. In addition, we found that the pluripotency of the stem cells was indicated by the expression of the markers OCT3/4, Nanog, CD105, CD90, SSEA-4 and STRO-1

A alopecia difusa não-cicatricial (ADNC) acomete 7% dos pacientes HIV positivos. O objetivo deste trabalho foi comparar os achados histológicos e imunohistoquímicos (citoqueratina 19/TUNEL e caspase 3 clivada) em cortes transversais de couro cabeludo de 15 pacientes HIV-1 positivos com ADNC e de 12 controles sadios. A apoptose de células tronco-foliculares e amplificadoras transitórias na protuberância folicular foi demonstrada pela dupla marcação TUNEL/CK19 em 80% dos casos e em 25% dos controles e, pelo anticorpo anti-caspase 3 clivada em 61% dos casos e 16% dos controles. Não houve correlação entre a incidência de apoptose e o grau de imunodeficiência dentre os casos / Seven percent of HIV positive patients present diffuse non-cicatricial alopecia (DNCA). The aim of this study was to compare the histological and the immunohistochemical (cytokeratin 19/TUNEL and cleaved caspase 3) findings in transverse scalp sections of 15 HIV-1 positive patients with DNCA and 12 healthy controls. Follicular stem and transit amplifying cells apoptosis in the bulge was demonstrated by double labeling TUNEL/CK19 in 80% of cases and 25% of controls, and by the anti-cleaved caspase 3 antibody in 61% of cases and 16% of the controls. There was no relation between the incidence of apoptosis and the degree of immunodeficiency among cases