A Guide to Assessing Damage Response Pathways of the Hair Follicle: Lessons From Cyclophosphamide-Induced Alopecia in Mice
2005; Elsevier BV; Volume: 125; Issue: 1 Linguagem: Inglês
10.1111/j.0022-202x.2005.23787.x
ISSN1523-1747
AutoresSven Hendrix, Bori Handjiski, Eva M.J. Peters, Ralf Paus,
Tópico(s)melanin and skin pigmentation
ResumoAfter chemical, biological, or physical damage, growing (i.e. anagen) hair follicles develop abnormalities that are collectively called hair follicle dystrophy. Comparatively lower follicular damage induces the “dystrophic anagen” response pathway (=prolonged, dystrophic anagen, followed by severely retarded follicular recovery). More severe follicular damage induces the dystrophic catagen pathway (=immediate anagen termination, followed by a dystrophic, abnormally shortened telogen and maximally fast follicular recovery). In order to recognize these distinct damage response strategies of the hair follicle in a clinical or histopathological context, we have used the well-established C57BL/6J mouse model of cyclophosphamide-induced alopecia to define pragmatic classification criteria for hair follicle dystrophy (e.g., structure and pigmentation of the hair shaft, location, and volume of ectopic melanin granules, distension of follicular canal, number of TdT-mediated dUTP nick end labeling positive keratinocytes in the hair bulb; neural cell-adhesion molecule immunoreactivity and alkaline phosphatase activity as markers for the level of damage to the follicular papilla). These classification criteria for hair follicle dystrophy are useful not only in chemotherapy-induced alopecia models, but also in the screening of drug-treated or mutant mice in a highly standardized, accurate, sensitive, reproducible, easily applicable, and quantifiable manner. After chemical, biological, or physical damage, growing (i.e. anagen) hair follicles develop abnormalities that are collectively called hair follicle dystrophy. Comparatively lower follicular damage induces the “dystrophic anagen” response pathway (=prolonged, dystrophic anagen, followed by severely retarded follicular recovery). More severe follicular damage induces the dystrophic catagen pathway (=immediate anagen termination, followed by a dystrophic, abnormally shortened telogen and maximally fast follicular recovery). In order to recognize these distinct damage response strategies of the hair follicle in a clinical or histopathological context, we have used the well-established C57BL/6J mouse model of cyclophosphamide-induced alopecia to define pragmatic classification criteria for hair follicle dystrophy (e.g., structure and pigmentation of the hair shaft, location, and volume of ectopic melanin granules, distension of follicular canal, number of TdT-mediated dUTP nick end labeling positive keratinocytes in the hair bulb; neural cell-adhesion molecule immunoreactivity and alkaline phosphatase activity as markers for the level of damage to the follicular papilla). These classification criteria for hair follicle dystrophy are useful not only in chemotherapy-induced alopecia models, but also in the screening of drug-treated or mutant mice in a highly standardized, accurate, sensitive, reproducible, easily applicable, and quantifiable manner. alkaline phosphatase chemotherapy-induced alopecia connective tissue sheath cyclophosphamide dermal papilla inner root sheath neural cell-adhesion molecule outer root sheath TdT-mediated dUTP nick end labeling In response to damage, hair follicles undergo two distinct pathways of dystrophy, which are characterized by specific morphological abnormalities. These are particularly prominent and clinically highly relevant in the course of chemotherapy-induced alopecia (CIA). Decades ago, different types of alopecias as well as some key parameters for the recognition of defined stages of chemotherapy-induced hair follicle dystrophy in mammals were defined (Braun-Falco and Theisen, 1959Braun-Falco O. Theisen H. Histologische und histochemische Veränderungen bei temporärer Monojodacetat-Alopecie. Eine tierexperimentelle Studie.Arch Klin Exp Dermatol. 1959; 208: 539-558Crossref PubMed Scopus (3) Google Scholar; Braun-Falco, 1961Braun-Falco O. Klinik und Pathomechanismus der Endoxan-Alopecie als Beitrag zum Wesen cytostatischer Alopecien.Arch Klin Exp Dermatol. 1961; 212: 194-216Crossref Scopus (17) Google Scholar,Braun-Falco, 1966Braun-Falco O. Dynamik des normalen und pathologischen Haarwachstums.Arch Klin Exp Dermatol. 1966; 227: 419-452PubMed Google Scholar; Zaun, 1964Zaun H. Tierexperimentelle Untersuchungen zur Pathophysiologie der “gemischten Alopecie”.Arch Klin Exp Dermatol. 1964; 221: 75-84Crossref PubMed Scopus (7) Google Scholar; Herzberg, 1966Herzberg J.J. Cytostatische Alopecien einschließlich Thallium-Alopecien.Arch Klin Exp Dermatol. 1966; 227: 452-467PubMed Google Scholar; Kostanecki et al., 1966Kostanecki W. Kwiatkowska E. Zborzil J. Die Haarmelanogenese bei der Endoxan-Alopecie und deren beeinflussung durch Corticosteroide.Arch Klin Exp Dermatol. 1966; 226: 13-20Crossref PubMed Scopus (6) Google Scholar; Homan et al., 1968Homan E.R. Zendzian R.P. Busey W.M. Rall D.P. Loss of hair in experimental animals induced by cyclophosphamide.Nature. 1968; 221: 1058-1059Crossref Scopus (13) Google Scholar). For nearly 40 y, these publications have been the only references for the classification of hair follicle dystrophy, although none of them offers a comprehensive, unified classification scheme for use in the laboratory. For this reason, we have developed a set of pragmatic classification criteria for hair follicle dystrophy, using the C57BL/6J mouse model of cyclophosphamide (CYP)-induced alopecia (Figure 1) (Paus et al., 1994cPaus R. Handjiski B. Eichmüller S. Czarnetzki B.M. Chemotherapy-induced alopecia in mice. Induction by cyclophosphamide, inhibition by cyclosporine A, and modulation by dexamethasone.Am J Pathol. 1994; 144: 719-734PubMed Google Scholar,Paus et al., 1996Paus R. Schilli M.B. Handjiski B. Menrad A. Henz B.M. Plonka P. Topical calcitriol enhances normal hair regrowth but does not prevent chemotherapy-induced alopecia in mice.Cancer Res. 1996; 56: 4438-4443PubMed Google Scholar; Slominski et al., 1996Slominski A. Paus R. Plonka P. Handjiski B. Maurer M. Chakraborty A. Mihm Jr, M.C. Pharmacological disruption of hair follicle pigmentation by cyclophosphamide as a model for studying the melanocyte response to and recovery from cytotoxic drug damage in situ.J Invest Dermatol. 1996; 106: 1203-1211Crossref PubMed Scopus (55) Google Scholar; Schilli et al., 1998Schilli M.B. Paus R. Menrad A. Reduction of intrafollicular apoptosis in chemotherapy-induced alopecia by topical calcitriol-analogs.J Invest Dermatol. 1998; 111: 598-604Crossref PubMed Scopus (56) Google Scholar; Müller-Röver et al., 2000Müller-Röver S. Rossiter H. Paus R. et al.Overexpression of Bcl-2 protects from ultraviolet B-induced apoptosis but promotes hair follicle regression and chemotherapy-induced alopecia.Am J Pathol. 2000; 156: 1395-1405Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar; Peters et al., 2001Peters E.M. Foitzik K. Paus R. Ray S. Holick M.F. A new strategy for modulating chemotherapy-induced alopecia, using PTH/PTHrP receptor agonist and antagonist.J Invest Dermatol. 2001; 117: 173-178Crossref PubMed Google Scholar). Studying this model, we had found that hair follicles undergo two distinct pathways of dystrophy when they have suffered chemical damage (here: by cytostatic drugs) (Paus et al., 1994cPaus R. Handjiski B. Eichmüller S. Czarnetzki B.M. Chemotherapy-induced alopecia in mice. Induction by cyclophosphamide, inhibition by cyclosporine A, and modulation by dexamethasone.Am J Pathol. 1994; 144: 719-734PubMed Google Scholar). These two damage-response pathways are characterized by specific morphological abnormalities, which eventually lead to hair loss and alopecia (Figure 2). Therefore, guidelines for the accurate and standardized classification of chemotherapy-induced hair follicle dystrophy are urgently needed, not the least in order to assist in the ongoing quest to combat CIA by the development of more effective alopecia-protection strategies.Figure 2Damage-response pathways of the hair follicle after chemotherapy. Schematic representation of the two different pathways of chemotherapy-induced hair follicle dystrophy: the dystrophic anagen pathway and the dystrophic catagen pathway. The size of the black flash () indicates how severely the follicle has been damaged, dependent on the administered dose of the cytostatic agent. In the white boxes, recognized factors are listed which promote the corresponding pathway, whereas the damage pathways shown here have, so far, only rigorously studied for cyclophosphamide-induced alopecia, both in mice and man, they likely apply also to other forms of chemical or biological damage of the hair follicle and the subsequent follicle response (cf.Braun-Falco and Theisen, 1959Braun-Falco O. Theisen H. Histologische und histochemische Veränderungen bei temporärer Monojodacetat-Alopecie. Eine tierexperimentelle Studie.Arch Klin Exp Dermatol. 1959; 208: 539-558Crossref PubMed Scopus (3) Google Scholar; Braun-Falco, 1961Braun-Falco O. Klinik und Pathomechanismus der Endoxan-Alopecie als Beitrag zum Wesen cytostatischer Alopecien.Arch Klin Exp Dermatol. 1961; 212: 194-216Crossref Scopus (17) Google Scholar; Zaun, 1964Zaun H. Tierexperimentelle Untersuchungen zur Pathophysiologie der “gemischten Alopecie”.Arch Klin Exp Dermatol. 1964; 221: 75-84Crossref PubMed Scopus (7) Google Scholar; Herzberg, 1966Herzberg J.J. Cytostatische Alopecien einschließlich Thallium-Alopecien.Arch Klin Exp Dermatol. 1966; 227: 452-467PubMed Google Scholar; Whiting, 2003Whiting D.A. Histopathologic features of alopecia areata: A new look.Arch Dermatol. 2003; 139: 1555-1559Crossref PubMed Scopus (143) Google Scholar). During the dystrophic anagen pathway the hair shaft is shed and the follicle undergoes an incomplete primary recovery. The follicle is not fully recovered before it undergoes a complete catagen–telogen transition to enter the final secondary recovery stage. In contrast, during the dystrophic catagen pathway, the follicle is more severely damaged and enters directly into dystrophic catagen and a shortened dystrophic telogen. Without passing through a primary recovery the follicle enters directly into a complete secondary recovery stage. The question mark indicates that it cannot be excluded that dystrophic anagen might also lead into the dystrophic catagen pathway.View Large Image Figure ViewerDownload (PPT) This review complements our earlier guides on the classification of murine hair follicle development (Paus et al., 1999Paus R. Müller-Röver S. van der Veen C. et al.A comprehensive guide for the recognition and classification of distinct stages of hair follicle morphogenesis.J Invest Dermatol. 1999; 113: 523-532Crossref PubMed Scopus (421) Google Scholar) and hair follicle cycling (Müller-Röver et al., 2001Müller-Röver S. Handjiski B. van der Veen C. et al.A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.J Invest Dermatol. 2001; 117: 3-15Crossref PubMed Google Scholar) so as to provide a standardized approach to the analysis of murine hair follicle dystrophy. It serves as a useful companion to a similar guide that has recently been published for the assessment of human hair follicle dystrophy (Whiting, 2003Whiting D.A. Histopathologic features of alopecia areata: A new look.Arch Dermatol. 2003; 139: 1555-1559Crossref PubMed Scopus (143) Google Scholar), and should be of particular interest to all researchers that wish to professionally assess hair follicle damage inflicted by test agents, engineered or spontaneous mutations and a wide range of diseases. Given the exquisite sensitivity of the hair follicle as a “biological damage indicator” that is negatively affected, e.g. by an enormous number of different clinically widely used drugs (cf.Litt, 2004Litt J.Z. Litt's Drug Eruption Reference Manual including Drug Interactions. 10th edn. CRC Press, Boca Raton FL2004Google Scholar) and by numerous mutations (Nakamura et al., 2002Nakamura M. Tobin D.J. Richards-Smith B. Sundberg J.P. Paus R. Mutant laboratory mice with abnormalities in pigmentation: Annotated tables.J Dermatol Sci. 2002; 28: 1-33Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar)., a professional assessment of hair follicle dystrophy along the lines indicated here exemplarily for CIA offers a simple, yet reliable comprehensive and instructive tool for obtaining novel insights into the biological effects of test agents. Based on basic histological and ultrastructural studies on human and rodent CIA (Braun-Falco, 1966Braun-Falco O. Dynamik des normalen und pathologischen Haarwachstums.Arch Klin Exp Dermatol. 1966; 227: 419-452PubMed Google Scholar; Herzberg, 1966Herzberg J.J. Cytostatische Alopecien einschließlich Thallium-Alopecien.Arch Klin Exp Dermatol. 1966; 227: 452-467PubMed Google Scholar), we have summarized basic as well as more advanced auxiliary criteria to define the distinct stages of the dystrophic anagen and the dystrophic catagen pathway (Figures 3 and 4) which are widely applicable to different mouse strains and mutants. In essence, this classification guide can also be utilized for staging the hair follicles of other hair-bearing animals, even though species-specific anatomic differences must be taken into account.Figure 3A comprehensive guide for the recognition and classification of distinct stages of dysotropic anagen. The left-hand column shows a computer-generated schematic drawing of healthy and dystrophic anagen stages and dystrophic catagen and telogen stages. The second column summarizes essential basic criteria for the recognition of single stages (above dotted line) and auxiliary criteria for more precise staging. The right-hand column shows representative micrographs of each dystrophy stage (lower-case letters correspond to lower-case letters used in the central column). The following staining techniques were employed: 3A, D, G, K; 4E, G, J, L: Giemsa-staining technique (Romeis, 1991Romeis B. Mikroskopische Technik. Urban & Schwarzenberg, München1991Google Scholar); 3B, E, H, J; 4A, D, K: alkaline phosphatase staining (Handjiski et al., 1994Handjiski B.K. Eichmüller S. Hofmann U. Czarnetzki B.M. Paus R. Alkaline phosphatase activity and localization during the murine hair cycle.Br J Dermatol. 1994; 131: 303-310Crossref PubMed Scopus (127) Google Scholar); 4B, H: NCAM immunoreactivity (Müller-Röver et al., 1998Müller-Röver S. Peters E.J. Botchkarev V.A. Panteleyev A. Paus R. Distinct patterns of NCAM expression are associated with defined stages of murine hair follicle morphogenesis and regression.J Histochem Cytochem. 1998; 46: 1401-1410Crossref PubMed Scopus (47) Google Scholar). Please note: upper-case letters in the left-hand corner label the image whereas lower-case letters identify tissues corresponding to the lower-case letters of the criteria listed in the central column. Please also note: the left-hand column comprehensively lists all helpful markers, although not all markers are shown in the micrographs.View Large Image Figure ViewerDownload (PPT)Figure 4A comprehensive guide for the recognition and classification of distinct stages dystrophic catagen and telogen. The left-hand column shows a computer-generated schematic drawing of healthy and dystrophic anagen stages and dystrophic catagen and telogen stages. The second column summarizes essential basic criteria for the recognition of single stages (above dotted line) and auxiliary criteria for more precise staging. The right-hand column shows representative micrographs of each dystrophy stage (lower-case letters correspond to lower-case letters used in the central column). The following staining techniques were employed: 3A, D, G, K; 4E, G, J, L: Giemsa-staining technique (Romeis, 1991Romeis B. Mikroskopische Technik. Urban & Schwarzenberg, München1991Google Scholar); 3B, E, H, J; 4A, D, K: alkaline phosphatase staining (Handjiski et al., 1994Handjiski B.K. Eichmüller S. Hofmann U. Czarnetzki B.M. Paus R. Alkaline phosphatase activity and localization during the murine hair cycle.Br J Dermatol. 1994; 131: 303-310Crossref PubMed Scopus (127) Google Scholar); 4B, H: NCAM immunoreactivity (Müller-Röver et al., 1998Müller-Röver S. Peters E.J. Botchkarev V.A. Panteleyev A. Paus R. Distinct patterns of NCAM expression are associated with defined stages of murine hair follicle morphogenesis and regression.J Histochem Cytochem. 1998; 46: 1401-1410Crossref PubMed Scopus (47) Google Scholar). Please note: upper-case letters in the left-hand corner label the image whereas lower-case letters identify tissues corresponding to the lower-case letters of the criteria listed in the central column. Please also note: the left-hand column comprehensively lists all helpful markers, although not all markers are shown in the micrographs.View Large Image Figure ViewerDownload (PPT)Figure 4A comprehensive guide for the recognition and classification of distinct stages dystrophic catagen and telogen. The left-hand column shows a computer-generated schematic drawing of healthy and dystrophic anagen stages and dystrophic catagen and telogen stages. The second column summarizes essential basic criteria for the recognition of single stages (above dotted line) and auxiliary criteria for more precise staging. The right-hand column shows representative micrographs of each dystrophy stage (lower-case letters correspond to lower-case letters used in the central column). The following staining techniques were employed: 3A, D, G, K; 4E, G, J, L: Giemsa-staining technique (Romeis, 1991Romeis B. Mikroskopische Technik. Urban & Schwarzenberg, München1991Google Scholar); 3B, E, H, J; 4A, D, K: alkaline phosphatase staining (Handjiski et al., 1994Handjiski B.K. Eichmüller S. Hofmann U. Czarnetzki B.M. Paus R. Alkaline phosphatase activity and localization during the murine hair cycle.Br J Dermatol. 1994; 131: 303-310Crossref PubMed Scopus (127) Google Scholar); 4B, H: NCAM immunoreactivity (Müller-Röver et al., 1998Müller-Röver S. Peters E.J. Botchkarev V.A. Panteleyev A. Paus R. Distinct patterns of NCAM expression are associated with defined stages of murine hair follicle morphogenesis and regression.J Histochem Cytochem. 1998; 46: 1401-1410Crossref PubMed Scopus (47) Google Scholar). Please note: upper-case letters in the left-hand corner label the image whereas lower-case letters identify tissues corresponding to the lower-case letters of the criteria listed in the central column. Please also note: the left-hand column comprehensively lists all helpful markers, although not all markers are shown in the micrographs.View Large Image Figure ViewerDownload (PPT) Hair follicle cycling in young mice follows a rather precise timescale (Paus et al., 1999Paus R. Müller-Röver S. van der Veen C. et al.A comprehensive guide for the recognition and classification of distinct stages of hair follicle morphogenesis.J Invest Dermatol. 1999; 113: 523-532Crossref PubMed Scopus (421) Google Scholar; Müller-Röver et al., 2001Müller-Röver S. Handjiski B. van der Veen C. et al.A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.J Invest Dermatol. 2001; 117: 3-15Crossref PubMed Google Scholar). Nevertheless, the fine details of hair follicle cycling are dependent on the genetic background (mouse strain), the sex (e.g., female mice show a prolonged telogen; S. Müller-Röver, unpublished observation) as well as environmental factors such as time of the year (temperature, light periods) and nutritional factors. To avoid associated fluctuations, the present guide is based on the most extensively studied and best standardized hair research model, the C57BL/6J models of depilation-induced hair cycling (Chase, 1954Chase H. Growth of the hair.Physiol Rev. 1954; 1: 113-126Google Scholar; Paus et al., 1990Paus R. Stenn K.S. Link R.E. Telogen skin contains an inhibitor of hair growth.Br J Dermatol. 1990; 122: 777-784Crossref PubMed Scopus (208) Google Scholar,Paus et al., 1994aPaus R. Eichmuller S. Hofmann U. Czarnetzki B.M. Robinson P. Expression of classical and non-classical MHC class I antigens in murine hair follicles.Br J Dermatol. 1994; 131: 177-183Crossref PubMed Scopus (75) Google Scholar, Paus et al., 1994bPaus R. Handjiski B. Czarnetzki B.M. Eichmuller S. A murine model for inducing and manipulating hair follicle regression (catagen): Effects of dexamethasone and cyclosporin A.J Invest Dermatol. 1994; 103: 143-147Crossref PubMed Scopus (111) Google Scholar; Müller-Röver et al., 2001Müller-Röver S. Handjiski B. van der Veen C. et al.A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.J Invest Dermatol. 2001; 117: 3-15Crossref PubMed Google Scholar) and CYP-induced alopecia (Paus et al., 1994cPaus R. Handjiski B. Eichmüller S. Czarnetzki B.M. Chemotherapy-induced alopecia in mice. Induction by cyclophosphamide, inhibition by cyclosporine A, and modulation by dexamethasone.Am J Pathol. 1994; 144: 719-734PubMed Google Scholar,Paus et al., 1996Paus R. Schilli M.B. Handjiski B. Menrad A. Henz B.M. Plonka P. Topical calcitriol enhances normal hair regrowth but does not prevent chemotherapy-induced alopecia in mice.Cancer Res. 1996; 56: 4438-4443PubMed Google Scholar; Slominski et al., 1996Slominski A. Paus R. Plonka P. Handjiski B. Maurer M. Chakraborty A. Mihm Jr, M.C. Pharmacological disruption of hair follicle pigmentation by cyclophosphamide as a model for studying the melanocyte response to and recovery from cytotoxic drug damage in situ.J Invest Dermatol. 1996; 106: 1203-1211Crossref PubMed Scopus (55) Google Scholar; Schilli et al., 1998Schilli M.B. Paus R. Menrad A. Reduction of intrafollicular apoptosis in chemotherapy-induced alopecia by topical calcitriol-analogs.J Invest Dermatol. 1998; 111: 598-604Crossref PubMed Scopus (56) Google Scholar; Müller-Röver et al., 2000Müller-Röver S. Rossiter H. Paus R. et al.Overexpression of Bcl-2 protects from ultraviolet B-induced apoptosis but promotes hair follicle regression and chemotherapy-induced alopecia.Am J Pathol. 2000; 156: 1395-1405Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar; Peters et al., 2001Peters E.M. Foitzik K. Paus R. Ray S. Holick M.F. A new strategy for modulating chemotherapy-induced alopecia, using PTH/PTHrP receptor agonist and antagonist.J Invest Dermatol. 2001; 117: 173-178Crossref PubMed Google Scholar; Ohnemus et al., 2004Ohnemus U. Unalan M. Handjiski B. Paus R. Topical estrogen accelerates hair regrowth in mice after chemotherapy-induced alopecia by favoring the dystrophic catagen response pathway to damage.J Invest Dermatol. 2004; 122: 7-13Abstract Full Text Full Text PDF PubMed Scopus (45) Google Scholar). Briefly, a wax/rosin mixture is applied on the dorsal skin of 7-wk-old mice with all dorsal skin hair follicles in the resting phase (telogen), as evidenced by the homogeneously pink back skin color. Plucking of the wax/rosin mixture removes all hair shafts and immediately induces anagen development of unparalleled homogeneity and synchrony over the entire depilated back of the mouse (Chase, 1954Chase H. Growth of the hair.Physiol Rev. 1954; 1: 113-126Google Scholar; Müller-Röver et al., 2001Müller-Röver S. Handjiski B. van der Veen C. et al.A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.J Invest Dermatol. 2001; 117: 3-15Crossref PubMed Google Scholar). Nine days after depilation, all depilated hair follicles have entered the final stage of the growth phase of the hair cycle (anagen VI). Around day 17 after depilation the follicles spontaneously start to undergo regression (catagen) to enter the resting phase (telogen) around day 20 after depilation (Figure 1a) (for further details seeMüller-Röver et al., 2001Müller-Röver S. Handjiski B. van der Veen C. et al.A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.J Invest Dermatol. 2001; 117: 3-15Crossref PubMed Google Scholar). In order to induce alopecia in this mouse model, 120–150 mg per kg body weight CYP are given once intraperitoneally at day 9 after depilation (Figure 1b). Three to seven days later the skin is harvested for further analysis (Paus et al., 1994cPaus R. Handjiski B. Eichmüller S. Czarnetzki B.M. Chemotherapy-induced alopecia in mice. Induction by cyclophosphamide, inhibition by cyclosporine A, and modulation by dexamethasone.Am J Pathol. 1994; 144: 719-734PubMed Google Scholar) (Figure 1b). In order to avoid terminological confusion, which is often caused by differences in how selected terms are used in hair research publications, we recently have summarized definitions of key terms employed in the context of this review (Müller-Röver et al., 2001Müller-Röver S. Handjiski B. van der Veen C. et al.A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.J Invest Dermatol. 2001; 117: 3-15Crossref PubMed Google Scholar). Please note that the term “proximal” here refers to those parts of the hair follicle, which are located close to the subcutaneous muscle layer, the panniculus carnosus, whereas “distal” refers to those parts, which are located close to the epidermis. This guide describes general principles, i.e. the two possible damage-response pathways (dystrophic anagen, dystrophic catagen) (Figure 2) and lists detailed criteria for precise staging of chemotherapy-induced hair follicle dystrophy (Figures 3 and 4). Table I lists auxiliary methods, while Table II summarizes general indicators of hair follicle dystrophy, which are illustrated in detail in Figure 3 (four stages of the dystrophic anagen pathway) and Figure 4 (four stages of the dystrophic catagen pathway).Table ISynopsis of auxiliary methods used in this guideAuxiliary methods/markersCompartmentReferenceAlkaline phosphatase stainingDermal papillaHandjiski et al., 1994Handjiski B.K. Eichmüller S. Hofmann U. Czarnetzki B.M. Paus R. Alkaline phosphatase activity and localization during the murine hair cycle.Br J Dermatol. 1994; 131: 303-310Crossref PubMed Scopus (127) Google ScholarTUNEL stainingHair matrix, ORS, IRSLindner et al., 1997Lindner G. Botchkarev V.A. Botchkareva N.V. Ling G. van der Veen C. Paus R. Analysis of apoptosis during hair follicle regression (catagen).Am J Pathol. 1997; 151: 1601-1617PubMed Google ScholarNCAMDermal papilla, perifollicular CTS and CTS tailMüller-Röver et al., 1998Müller-Röver S. Peters E.J. Botchkarev V.A. Panteleyev A. Paus R. Distinct patterns of NCAM expression are associated with defined stages of murine hair follicle morphogenesis and regression.J Histochem Cytochem. 1998; 46: 1401-1410Crossref PubMed Scopus (47) Google ScholarThese methods provide additional help in determining the stage-specific morphology of distinct hair follicle compartments such as the dermal papilla or the trailing connective tissue sheath.ORS, outer root sheath; IRS, inner root sheath; TUNEL, TdT-mediated dUTP nick end labeling; NCAM, neural cell-adhesion molecule; CTS, connective tissue sheath. Open table in a new tab Table IIGeneral signs of chemotherapy-induced hair follicle dystrophyEctopic melanin (intrafollicular, perifollicular)Irregular banding pattern of the hair shaft, fragmented hair shaftWide-open hair canalFollicular distortion, hair shaft distortionIrregular diameter of hair bulbs Open table in a new tab These methods provide additional help in determining the stage-specific morphology of distinct hair follicle compartments such as the dermal papilla or the trailing connective tissue sheath. ORS, outer root sheath; IRS, inner root sheath; TUNEL, TdT-mediated dUTP nick end labeling; NCAM, neural cell-adhesion molecule; CTS, connective tissue sheath. In order to illustrate the key parameters for recognizing the distinct stages of chemotherapy-induced hair follicle dystrophy, Figures 3 and 4 are structured as follows: the left-hand column shows computer-generated schematic drawings of the distinct stages of hair follicle dystrophy. The central column provides a list of basic and auxiliary classification criteria, which are separated from each other (top: basic, bottom: auxiliary criteria). The former are recognizable by routine light microscopy, whereas the latter require additional staining methods. The basic criteria are applicable to all pigmented mouse strains. The auxiliary criteria require histochemical techniques (i.e. alkaline phosphatase (AP) staining;Handjiski et al., 1994Handjiski B.K. Eichmüller S. Hofmann U. Czarnetzki B.M. Paus R. Alkaline phosphatase activity and localization during the murine hair cycle.Br J Dermatol. 1994; 131: 303-310Crossref PubMed Scopus (127) Google Scholar and TdT-mediated dUTP nick end labeling (TUNEL) staining;Lindner et al., 1997Lindner G. Botchkarev V.A. Botchkareva N.V. Ling G. van der Veen C. Paus R. Analysis of apoptosis during hair follicle regression (catagen).Am J Pathol. 1997; 151: 1601-1617PubMed Google Scholar) or immunohistochemistry for the adhesion receptor neuronal cell-adhesion molecule (NCAM) (Müller-Röver et al., 1998Müller-Röver S. Peters E.J. Botchkarev V.A. Panteleyev A. Paus R. Distinct patterns of NCAM expression are associated with defined stages of murine hair follicle morphogenesis and regression.J Histochem Cytochem. 1998; 46: 1401-1410Crossref PubMed Scopus (47) Google Scholar). The right-hand column illustrates the criteria listed in the central column with three repr
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