Ultrastructural localization of S100A3, a cysteine-rich, calcium binding protein, in human scalp hair shafts revealed by rapid-freezing immunocytochemistry.

We have characterized the subcellular distribution of S100A3, a cysteine-rich calcium binding protein, in human scalp hair shaft. This was accomplished using rapid-freezing immunocytochemistry, a technique that combines rapid-freezing, freeze-substitution fixation without chemical fixatives, and subsequent electron microscopic detection of immunocytochemical labeling. This technique preserves both the antigenicity and the ultrastructural integrity of fully keratinized tissues, which are highly unmanageable when prepared for immunoelectron microscopy. In the hair shaft, S100A3 was primarily identified in the endocuticle and was also present in the intermacrofibrillar matrix surrounding macrofibril bundles of intermediate filament keratins in cortex cells. Double immunolabeling of S100A3 and hair keratins revealed the in situ spatial relationship between them. In the endocuticle, S100A3 was present on the inner portion of the endocuticle adjacent to the cell membrane complex, whereas hair keratins were present on the outer portion. These results provide the first ultrastructural evidence that an S100 protein is localized in specific subcompartments in human hair cells. (J Histochem Cytochem 47:525-532, 1999)     

Specific citrullination causes assembly of a globular S100A3 homotetramer: a putative Ca2+ modulator matures human hair cuticle

S100A3 is a unique member of the Ca2+-binding S100 protein family with the highest cysteine content and affinity for Zn2+. This protein is highly expressed in the differentiating cuticular cells within the hair follicle and organized into mature hair cuticles. Previous studies suggest a close association of S100A3 with epithelial differentiation, leading to hair shaft formation, but its molecular function is still unknown. By two-dimensional PAGE-Western blot analyses using a modified citrulline antibody, we discovered that more than half of the arginine residues of native S100A3 are progressively converted to citrullines by Ca2+-dependent peptidylarginine deiminases. Confocal immunofluorescent microscopy showed that the cytoplasmic S100A3 within the cuticular layer is mostly co-localized with the type III isoform of peptidylarginine deiminase (PAD3) but not with PAD1. Recombinant PAD1 and PAD2 are capable of converting all 4 arginines in recombinant S100A3, whereas PAD3 specifically converts only Arg-51 into citrulline. Gel filtration analyses showed that either enzymatic conversion of Arg-51 in S100A3 to citrulline or its mutational substitution with alanine (R51A) promotes a homotetramer assembly. Fluorescent titration of R51A suggested that its potential Ca2+ binding property increased during tetramerization. A prototype structural model of the globular Ca2+-bound S100A3 tetramer with citrulline residues is presented. High concentrations of S100A3 homotetramer might provide the millimolar level of Ca2+ required for hair cuticular barrier formation.     

Protein structural changes in keratin fibers induced by chemical modification using 2-iminothiolane hydrochloride: a Raman spectroscopic investigation

For the purpose of investigating in detail the influence of chemical modification using 2-iminothiolane hydrochloride (2-IT) on keratin fibers, the structure of cross-sections at various depths of white human hair, treated with 2-IT and then oxidized, was directly analyzed without isolating the cuticle and cortex, using Raman spectroscopy. In particular, the beta-sheet and/or random coil content (beta/R) and the alpha-helix (alpha) content in human hair fibers were estimated by amide I band analysis. The S-S band intensity, amide III (unordered) band intensity, and beta/R content existing from the cuticle region to the center of cortex region of virgin white human hair remarkably increased by performing the chemical modification using 2-IT. On the other hand, not only the S-S band intensity, but also S-O band intensity existing throughout the cortex region of the bleached (damaged) white human hair increased by performing chemical modification using 2-IT. In particular, beta/R content existing throughout the cortex region of the bleached white human hair decreased, while the skeletal C-C stretch (alpha) band intensity at 935 cm(-1) and the alpha content remarkably increased. This indicates a secondary structural change from the random coil form to the alpha-helix form in the proteins existing throughout the cortex region. From these experiments, we concluded that the formation of new disulfide (-SS-) groups resulting from chemical modification using 2-IT induced the secondary structural changes of proteins existing throughout the cortex region.     

Analysis of structural changes in bleached keratin fibers (black and white human hair) using Raman spectroscopy

To investigate the influence of bleaching treatments on keratin fibers, the structure of cross-sections at various depths of bleached human hair (black and white human hair) was directly analyzed without isolating the cuticle and cortex, using Raman spectroscopy. The S-S band intensity existing from the cuticle region to the center of cortex region of virgin white human hair decreased, while the S-O band intensity at 1040 cm(-1), assigned to cysteic acid, increased by performing the bleaching treatment. Especially, the S-O band intensity of the cuticle region increased remarkably compared with that of the cortex region. Also, the amide III (unordered) band intensity in the cortex region increased, indicating that some of the proteins existing throughout the cortex region changed to the random coil form. Moreover, it has been found that the S-S band intensity existing from the cuticle region to the center of the cortex region of the virgin black human hair decreased remarkably, while the S-O band intensity increased significantly compared with that of the virgin white human hair by performing the bleaching treatment. From these experiments, we concluded that the melanin granules including metal ions act as a decomposition accelerator for the oxidizing agent, thereby leading to a higher level of disulfide (-SS-) group cleavage in the black human hair compared with that of the white human hair.     

S100 and S100 fused-type protein families in epidermal maturation with special focus on S100A3 in mammalian hair cuticles

Epithelial Ca²⁺-regulation, which governs cornified envelope formation in the skin epidermis and hair follicles, closely coincides with the expression of S100A3, filaggrin and trichohyalin, and the post-translational modification of these proteins by Ca²⁺-dependent peptidylarginine deiminases. This review summarizes the current nomenclature and evolutional aspects of S100 Ca²⁺-binding proteins and S100 fused-type proteins (SFTPs) classified as a separate protein family with special reference to the molecular structure and function of S100A3 dominantly expressed in hair cuticular cells. Both S100 and SFTP family members are identified by two distinct types of Ca²⁺-binding loops in an N-terminal pseudo EF-hand motif followed by a canonical EF-hand motif. Seventeen members of the S100 protein family including S100A3 are clustered with seven related genes encoding SFTPs on human chromosome 1q21, implicating their association with epidermal maturation and diseases. Human S100A3 is characterized by two disulphide bridges and a preformed Zn²⁺-pocket, and may transfer Ca²⁺ ions to peptidylarginine deiminases after its citrullination-mediated tetramerization. Phylogenetic analysis utilizing current genome databases suggests that divergence of the S100A3 gene coincided with the emergence of hair, a defining feature of mammals, and that the involvement of S100A3 in epithelial Ca²⁺-cycling occurred as a result of a skin adaptation in terrestrial mammals.     

The morphogenesis of cell hairs on Drosophila wings

We describe in this paper details of morphogenesis of wing hairs in Drosophila pupae. The ultimate objective is to relate specific protein components used in hair construction to specific components produced in the rapidly changing patterns of gene expression that are characteristic for the period of hair differentiation in wing cells (H. K. Mitchell and N. S. Petersen, 1981, Dev. Biol. 85, 233-242). Hair extrusion to essentially full size occurs quite suddenly at about 34 hr (postpupariation) and this is followed by deposition of a double-layer of cuticulin during the next 4 to 5 hr. Extreme changes in shape of cells and hairs, probably related to actin synthesis, then occur for the next 5 to 6 hr. Deposition of fibers within the hairs and on hair pedestals follows. Formation of cuticle on the cell surface begins and continues until some time in the 60-hr range. It appears that cuticle is formed only on the cell surface and not in hairs or on the top of hair pedestals. The protein synthesis patterns associated with these events are described.     

Analysis of structural change in keratin fibers resulting from chemical treatments using Raman spectroscopy

In order to investigate the influence of chemical treatments (reduction, heating, and oxidation) on keratin fibers, the structure of virgin white human hair resulting from a permanent hair straightening process at various depths of cross-sectional samples was directly analyzed without isolating the cuticle and cortex, using Raman spectroscopy. The band shape of the cuticle was different from that of the cortex, and the cuticle had a more amorphous structure, compared with the cortex. The S-S band intensity existing in the hair surface remarkably decreased, while the S-S band intensity in the hair center was not changed by performing the reduction process. In the case of heating the keratin fibers after the reduction process, this tendency was unchanged. On the other hand, the amide III (unordered) band intensity in the cortex region increased, indicating that proteins existing throughout the cortex region caused a change to the random coil form. Moreover, approximately 95% of the disconnected -SS- groups were clearly reconnected by performing the oxidation process after heating (the degree of reconnection of -SS- groups was about 90%, in the case of oxidizing after reduction). From these experiments, we concluded that the heat treatment process in the permanent hair straightening treatment caused the randomization of proteins existing throughout the cortex region, thereby contributing to the acceleration of the reconnection of -SS- groups during the oxidation process.     

Desmoglein 4 is expressed in highly differentiated keratinocytes and trichocytes in human epidermis and hair follicle

Desmosomes are critical for the tissue integrity of stratified epithelia and their appendages. Desmogleins (DSGs) and desmocollins (DSCs) are transmembrane desmosomal cadherins that interact extracellularly to link neighboring epithelial cells. We recently identified a new member of the DSG family, designated desmoglein 4, whose mutations cause hypotrichosis in human, mouse and rat. In this study, we analyzed in detail the expression domains of human desmoglein 4 protein (DSG4) in human skin relative to differentiation markers and other DSGs. Our results show that DSG4 protein is expressed in the more highly differentiated layers of the epidermis. This expression pattern in vivo is recapitulated in highly differentiated HaCaT human keratinocytes and normal human keratinocytes in vitro. In the human hair follicle, DSG4 is expressed specifically in the hair shaft cortex, the lower hair cuticle, and the upper inner root sheath (IRS) cuticle. Using a green fluorescent protein-tagged version of mouse or rat desmoglein 4 protein (Dsg4) and immuno-electron microscopy, we demonstrate that Dsg4 localizes to desmosomes both in vitro and in vivo. The highly specific expression pattern of DSG4 in the human hair follicle, combined with the phenotype of rodent models and human patients with desmoglein 4 mutations, underscores the importance of this adhesion molecule in the integrity of the hair shaft.     

THE ELECTRON MICROSCOPY OF THE HUMAN HAIR FOLLICLE : PART 3. THE INNER ROOT SHEATH AND TRICHOHYALINE

1. The three cylinders of cells, each one cell thick, which together constitute the inner root sheath, arise from the peripheral portions of the undifferentiated matrix. These cells, like the hair cuticle, are stabilised by the spread of adhesive contacts between their plasma membranes which occurs in the mid-bulb and upper bulb of the hair follicle. 2. The characteristic intracellular product of all three cell layers is trichohyaline. This substance is formed in the first place as amorphous droplets which subsequently transform into a birefringent form. 3. This transformation, involving the formation of a birefringent product from an amorphous precursor, is in contrast to the formation in the cortex of keratin which originates in a fibrous form. 4. Trichohyaline appears first and transforms first in the cells of Henle which are nearest the outer sheath and the dermal supply vessels. This transformation occurs at the level of the neck of the follicle. Synthesis and transformation in the cells of Huxley and the sheath cuticle lag behind the similar events in the cells of Henle. The transformation does not begin until the lower prekeratinous zone in the Huxley and cuticle cells. 5. The amorpous-fibrous transformation occurs rapidly cell by cell and involves the conversion of all the trichohyaline droplets. In longitudinal sections the birefringent modification can be seen extending from the droplets in both directions parallel to the axis of the hair. In cross-sections the images of the transformed material are difficult to interpret. They may be seen as sections of corrugated sheets (~100 A thick) or condensed fibrils ~100 A in width. 6. At the same time that the trichohyaline transforms, the spacing between the cell membranes increases and a dark deposit appears centrally between them. This membrane complex, and the similar complex of the hair cuticle cells described in Part 2, may be specialised formations whose purpose is to hold the hardened cells together.     

The electron microscopy of the human hair follicle. III. The inner root sheath and trichohyaline

1. The three cylinders of cells, each one cell thick, which together constitute the inner root sheath, arise from the peripheral portions of the undifferentiated matrix. These cells, like the hair cuticle, are stabilised by the spread of adhesive contacts between their plasma membranes which occurs in the mid-bulb and upper bulb of the hair follicle. 2. The characteristic intracellular product of all three cell layers is trichohyaline. This substance is formed in the first place as amorphous droplets which subsequently transform into a birefringent form. 3. This transformation, involving the formation of a birefringent product from an amorphous precursor, is in contrast to the formation in the cortex of keratin which originates in a fibrous form. 4. Trichohyaline appears first and transforms first in the cells of Henle which are nearest the outer sheath and the dermal supply vessels. This transformation occurs at the level of the neck of the follicle. Synthesis and transformation in the cells of Huxley and the sheath cuticle lag behind the similar events in the cells of Henle. The transformation does not begin until the lower prekeratinous zone in the Huxley and cuticle cells. 5. The amorpous-fibrous transformation occurs rapidly cell by cell and involves the conversion of all the trichohyaline droplets. In longitudinal sections the birefringent modification can be seen extending from the droplets in both directions parallel to the axis of the hair. In cross-sections the images of the transformed material are difficult to interpret. They may be seen as sections of corrugated sheets ( approximately 100 A thick) or condensed fibrils approximately 100 A in width. 6. At the same time that the trichohyaline transforms, the spacing between the cell membranes increases and a dark deposit appears centrally between them. This membrane complex, and the similar complex of the hair cuticle cells described in Part 2, may be specialised formations whose purpose is to hold the hardened cells together.     

Effect of SDS on human hair: Study on the molecular structure and morphology

This paper presents a model study to understand the effect of surfactants on the physicochemical properties of human hair. FT‐IR ATR spectroscopy has been employed to understand the chemical changes induced by sodium dodecyl sulfate (SDS) on human scalp hair. In particular, the SDS induced changes in the secondary structure of protein present in the outer protective layer of hair, i.e. cuticle, have been investigated. Conformational changes in the secondary structure of protein were studied by curve fitting of the amide I band after every phase of SDS treatment. It has been found that SDS brings rearrangements in the protein backbone conformations by transforming β ‐sheet structure to random coil and β ‐turn. Additionally, AFM and SEM studies were carried out to understand the morphological changes induced on the hair surface. SEM and AFM images demonstrated the rupture and partial erosion of cuticle sublayers. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Structure and expression of genes for a class of cysteine-rich proteins of the cuticle layers of differentiating wool and hair follicles

The major histological components of the hair follicle are the hair cortex and cuticle. The hair cuticle cells encase and protect the cortex and undergo a different developmental program to that of the cortex. We report the molecular characterization of a set of evolutionarily conserved hair genes which are transcribed in the hair cuticle late in follicle development. Two genes were isolated and characterized, one expressed in the human follicle and one in the sheep follicle. Each gene encodes a small protein of 16 kD, containing greater than 50 cysteine residues, ranging from 31 to 36 mol% cysteine. Their high cysteine content and in vitro expression data identify them as ultra-high-sulfur (UHS) keratin proteins. The predicted proteins are composed almost entirely of cysteine-rich and glycine-rich repeats. Genomic blots reveal that the UHS keratin proteins are encoded by related multigene families in both the human and sheep genomes. Tissue in situ hybridization demonstrates that the expression of both genes is localized to the hair fiber cuticle and occurs at a late stage in fiber morphogenesis.     

Investigation of human hair cuticle structure by microdiffraction: direct observation of cell membrane complex swelling.

The cuticle of mammalian hair fibres protects the core of the fibre against physical and chemical stress. The structure and some of the properties of the cuticle have been extensively studied by electron microscopy. However, there is still a need for a less invasive structural probe. For this purpose, microdiffraction experiments have been carried out on human hair samples showing a characteristic small-angle X-ray scattering pattern for the cuticle. This pattern has been assigned to the cell membrane complex (CMC) between each cuticle scale. Using a simple model of the electron density within the CMC, values have been derived for the average thickness of the beta- and delta-layers which are close to those obtained by electron microscopy. In order to illustrate the potentialities of microdiffraction in studying the properties of the cuticle, the effect of water sorption has been monitored. Using the intensity modelling described above, a 10% swelling of the delta-layer's thickness has been observed. This study shows that structural modifications of the CMC by physical or chemical stress can be followed directly on the cuticle of human hair fibres by microdiffraction analysis.     

Keratinolytic activity of Bacillus subtilis AMR using human hair

Aims:  To determine the ability of a novel Bacillus subtilis AMR isolated from poultry waste to hydrolyse human hair producing peptidases including keratinases and hair keratin peptides.
Methods and Results:  The Bacillus subtilis AMR was identified using biochemical tests and by analysis of 16S rDNA sequence. The isolate was grown in medium containing human hair as the sole source of carbon and nitrogen. The supplementation of hair medium (HM) with 0·01% yeast extract increased the keratinolytic activity 4·2-fold. B. subtilis AMR presented high keratinase production on the 8th day of fermentation in hair medium (HM) supplemented with 0·01% yeast extract (HMY) at pH 8·0. Keratinase yield was not correlated with increase in biomass. Zymography showed keratin-degrading peptidases migrating at c. 54, 80 and 100 kDa and gelatin-degrading bands at c. 80, 70 63, 54 32 and 15 kDa. Keratinases were optimally active at 50°C and pH 9·0 and was fully inhibited by the serine proteinase inhibitor (PMSF). Scanning electron microscopy showed complete degradation of the hair cuticle after exposure to B. subtilis AMR grown in HMY. MALDI-TOF analysis of culture supernatant containing peptides produced during enzymatic hydrolysis of hair by B. subtilis AMR revealed fragments in a range of 800–2600 Da.
Conclusions:  This study showed that B. subtilis AMR was able to hydrolyse human hair producing serine peptidases with keratinase and gelatinase activity as well as hair keratin peptides.
Significance and Impact of the Study:  This is the first report describing the production and partial characterization of keratinases by a B. subtilis strain grown in a medium containing human hair . These data suggest that peptides obtained from enzymatic hair hydrolysis may be useful for future applications on pharmaceutical and cosmetic formulations.     

Morphology and physiology of a hair plate sensory organ located on the antenna of the rock lobster Palinurus vulgaris

At the level of the J1 joint of each antenna of the rock lobster Palinurus vulgaris a hair plate sensory organ (hp) similar to those described in insects has been observed. The hp is located on the internal side of the S2 segment of the antenna, close to the soft articulating membrane of the J1 joint. It is formed by a triangular surface of cuticle about 3mm2 in area, covered with numerous hairs of different lengths (Figs. 1 and 2). Details of the hp were studied by scanning electron microscopy (Fig. 2). Physiological stimulation of the hp hairs occurs during medial movement of the J1 joint. Under this condition the soft articulating membrane rolls over the hairs and bends them progressively back onto the cuticle. Flexion of all the hairs corresponds to a medial movement of the J1 through 40 degrees. During this type of movement, the number of successively flexed hairs increases linearly (Fig. 3). Electrophysiological recordings of the hp sensory nerve correlated with selective mechanical stimulation of individual hairs demonstrated that each hair is innervated by a single sensory fiber. This sensory neurone responds phasically when the hair is flexed back onto the cuticle (as during an S2 medial movement) and when it returns to its resting position (as during an S2 lateral movement). Most of the sensory neurones are sensitive to the movement velocity of the hairs (Figs. 4 and 5). When the hair is maintained flexed its sensory neurone discharges tonically (Fig. 4). Electrical stimulation of the hp sensory nerve induced reflex actions in the external and internal rotator muscles of the segment S1. These effects were found to selectively activate the tonic motor command of these muscles (Fig. 6).     

S100a0 (αα) Protein, a Calcium-Binding Protein, Is Localized in the Slow-Twitch Muscle Fiber

We previously showed that, in contrast to the distribution of S100b (beta beta), S100a0 (alpha alpha) is mainly present in human skeletal and heart muscles at the level of 1-2 micrograms/mg of soluble protein and is universally distributed at high levels in skeletal and heart muscles of various mammals. To elucidate cellular and ultrastructural localizations of the alpha subunit of S100 protein (S100-alpha) in skeletal muscle, we used immunohistochemical and enzyme immunoassay methods. The immunohistochemical study revealed that S100-alpha is mainly localized in slow-twitch muscle fibers, whereas the beta subunit of S100 protein (S100-beta) was not detected in both types of muscle fibers, an observation indicating that the predominant form of S100 protein in the slow-twitch muscle fiber is not S100a or S100b, but S100a0. The quantitative analysis using enzyme immunoassay corroborates the immunohistochemical finding: The S100-alpha concentration of mouse soleus muscle (mainly composed of slow-twitch muscle fibers) is about threefold higher than that of mouse rectus femoris muscle (mainly composed of fast-twitch muscle fibers). At the ultrastructural level, S100-alpha is associated with polysomes, sarcoplasmic reticulum, the plasma membrane, the pellicle around lipid droplets, the outer membrane of mitochondria, and thin and thick filaments, by immunoelectron microscopy.     

The C terminus (amino acids 75-94) and the linker region (amino acids 42-54) of the Ca2+-binding protein S100A1 differentially enhance sarcoplasmic Ca2+ release in murine skinned skeletal muscle fibers

S100A1, a Ca2+-binding protein of the EF-hand type, is most highly expressed in striated muscle and has previously been shown to interact with the skeletal muscle sarcoplasmic reticulum (SR) Ca2+ release channel/ryanodine receptor (RyR1) isoform. However, it was unclear whether S100A1/RyR1 interaction could modulate SR Ca2+ handling and contractile properties in skeletal muscle fibers. Since S100A1 protein is differentially expressed in fast- and slow-twitch skeletal muscle, we used saponin-skinned murine Musculus extensor digitorum longus (EDL) and Musculus soleus (Soleus) fibers to assess the impact of S100A1 protein on SR Ca2+ release and isometric twitch force in functionally intact permeabilized muscle fibers. S100A1 equally enhanced caffeine-induced SR Ca2+ release and Ca2+-induced isometric force transients in both muscle preparations in a dose-dependent manner. Introducing a synthetic S100A1 peptide model (devoid of EF-hand Ca2+-binding sites) allowed identification of the S100A1 C terminus (amino acids 75-94) and hinge region (amino acids 42-54) to differentially enhance SR Ca2+ release with a nearly 3-fold higher activity of the C terminus. These effects were exclusively based on enhanced SR Ca2+ release as S100A1 influenced neither SR Ca2+ uptake nor myofilament Ca2+ sensitivity/cooperativity in our experimental setting. In conclusion, our study shows for the first time that S100A1 augments contractile performance both of fast- and slow-twitch skeletal muscle fibers based on enhanced SR Ca2+ efflux at least mediated by the C terminus of S100A1 protein. Thus, our data suggest that S100A1 may serve as an endogenous enhancer of SR Ca2+ release and might therefore be of physiological relevance in the process of excitation-contraction coupling in skeletal muscle.     

The crystal structure of metal-free human EF-hand protein S100A3 at 1.7-A resolution

S100A3 is a unique member of the EF-hand superfamily of Ca(2+)-binding proteins. It binds Ca(2+) with poor affinity (K(d) = 4-35 mm) but Zn(2+) with exceptionally high affinity (K(d) = 4 nm). This high affinity for Zn(2+) is attributed to the unusual high Cys content of S100A3. The protein is highly expressed in fast proliferating hair root cells and astrocytoma pointing toward a function in cell cycle control. We determined the crystal structure of the protein at 1.7 A. The high resolution structure revealed a large distortion of the C-terminal canonical EF-hand, which most likely abolishes Ca(2+) binding. The crystal structure of S100A3 allows the prediction of one putative Zn(2+) binding site in the C terminus of each subunit of S100A3 involving Cys and His residues in the coordination of the metal ion. Zn(2+) binding induces a large conformational change in S100A3 perturbing the hydrophobic interface between two S100A3 subunits, as shown by size exclusion chromatography and CD spectroscopy.     

Analysis of structural changes in permanent waved human hair using Raman spectroscopy

To investigate the mechanism leading to the reduction in tensile strength of permanent waved human hair, the structure of cross-sections at various depths of permanent waved white human hair was directly analyzed without isolating the cuticle and cortex, using Raman spectroscopy. The beta-sheet and/or random coil content (beta/R) and the Amide III(unordered) band intensity existing throughout the cortex region of virgin white human hair remarkably increased, while the alpha-helix (alpha) content slightly decreased by performing the permanent waving treatment. This suggests a secondary structural change from the alpha-helix form to the random coil form in the proteins existing in the microfibril of the cortex region. On the other hand, the S-S band intensity existing in the matrix of the cortex region almost did not change, despite the reduction in the tensile strength of the white human hair following the permanent waving treatment. Moreover, the transmission electron microscope observation shows that the macrofibril (the microfibril and matrix) existing in the cortex region of the virgin white human hair was remarkably disturbed, while the cuticle region was almost unchanged by performing the permanent waving treatment. From these experiments, the authors concluded that some of proteins existing in the cortex region (the microfibril and matrix) of the virgin white human hair were changed, thereby leading to the remarkable reduction in the tensile strength of the white human hair after the permanent waving treatment.