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Elution of S100A3 from hair fiber: New model for hair damage emphasizing the loss of S100A3 from cuticle
Abstract
In hair fiber, a cysteine-rich calcium-binding S100A3 protein is segregated in the inner part of the cuticle and postulated to play an important role in the attachment to the adjacent cuticular scale. In this study, elution of S100A3 from hair fiber was examined under various conditions by means of immunoblot analyses. The exposure of hair fiber to permanent waving lotions resulted in recoveries of substantial amounts of S100A3 by elution. Ultraviolet-light radiation and perming also increased the elution of S100A3 even without reductant. The distal part of hair fiber eluted less S100A3, as compared to the proximal section, under reducing conditions. These results suggest that S100A3 is eluted preferentially by daily washing and rinsing, especially from damaged hair. Given the presence of soluble S100A3 in the inner part of cuticle, we propose a new mechanism of hair damage in which the elution of S100A3 plays a major role.
... S100A3 is a small and cysteine-rich Ca 2+ -binding protein, specifically distributed in the endocuticle. 7 It has been reported that S100A3 is released from permed hair, 8 oxidized in bleached hair, 9 and fragmented upon UV-radiation. 10 This protein is known to be citrullinated and transformed into a tetramer in the hair follicle, where its Ca 2+ -binding ability advances drastically (Fig. 1). ...
... Protein was electro-transferred onto a PVDF (polyvinylidene difluoride) membrane using Nu-PAGE transfer buffer containing 10% methanol. The protein-transferred PVDF membrane was labeled with anti-S100A3 antibody 8 and then with AlexaFluor488 labeled anti-rabbit IgG antibody (ThermoFischer Scientific, Waltham, MA, USA). Labeled S100A3 blots were detected by Typhoon 9410 (GE Healthcare UK Ltd., Buckinghamshire, England). ...
... Consequently, hair cuticles with higher S100A3 content seem to be vulnerable to grooming stress. Although loss of S100A3 from the endocuticle upon cleavage of its disulfide cross links by perming was previously suggested to cause cuticle damage, 8 the amount of S100A3 extracted from intact individual hair does not seem to directly aid in the rigidity of cuticles at all. Considering that S100A3 content in mature cuticles (0.41% in this study) is apparently lower than whole plucked beard follicle extract containing premature cuticular cells (about 3%), 11 part of the citrullinated S100A3 pro- duced in cuticular cells becomes insoluble or degraded following citrullination during later differentiation processes. ...
Background:
There are two types of damage pattern of human hair cuticle: type L, where the cell membrane complex is split and the cuticle lifts up, and type E, where the fragile substructure of the cuticle cell (endocuticle) is broken. In our previous paper, it was reported that the dominant damage pattern shifts from type L to E with the subjects' age around the 40s. Loss of the cuticle due to daily grooming stresses increases with the subjects' age and is related to the level of type E damage. It is supposed that deterioration of endocuticle advances the loss of cuticle. S100A3 protein, located at the endocuticle, was found to be citrullinated and transformed into tetramer to improve its Ca(2+) -binding ability. It is postulated that this biochemical property affects the maturation of cuticle and contributes to its reinforcement.
Aims:
This study aims to elucidate the role that S100A3 plays in age-dependent cuticle damage.
Methods:
Hair fibers collected from Japanese females were evaluated for the content and citrullination rate of S100A3, incidence of type E damage, and resistance of cuticle.
Results:
In the aged hair, the content of S100A3 was positively correlated with the level of type E damage and low resistance to stress. Hair fibers in which S100A3 is highly citrullinated, however, showed low levels of type E damage and high resistance of cuticle, even in the aged hair as well as at younger ages.
Conclusions:
S100A3 and its citrullination process are related to rigidity of endocuticle of aged hair.
... S100A3, a unique protein among all members of the calcium-binding S100 family, is specifically expressed at the inner endocuticle of human hair fibers. Upon hair damage, S100A3 is released from hair fibers and possibly destabilizes the hair tissue architecture.[25] S100A3 protein is specifically expressed in the cuticular cells of human hair shaft[26] and in the cuticle of murine pelage follicle.[27] ...
... Thus, S100A3 could possibly provide structural integrity to the hair fiber and must be associated with hair damage.[25] S100A3 is released from the cuticle of hair fibers during washing and rinsing, especially from chemically treated or UVB-irradiated fiber.[25] ...
... In human hair fiber, the protein is segregated in the endocuticle of cuticular cells and in the matrix that surrounds macrofibril bundles in cortical cells.[28] Thus, S100A3 could possibly provide structural integrity to the hair fiber and must be associated with hair damage.[25] S100A3 is released from the cuticle of hair fibers during washing and rinsing, especially from chemically treated or UVB-irradiated fiber.[25] ...
Photoaggravation of hair aging includes various chemical and physical changes in fiber properties which lead to an increase in fiber porosity, loss of mechanical strength and an increase in surface roughness. These changes come from lipid oxidation, disulfide bond cleavage, tryptophan degradation and cysteic acid formation. Hair exposed to sunlight is claimed to be more brittle, stiffer and drier than before irradiation and exhibits a reduced water-absorption capacity. Hair pigments function to provide photochemical protection to hair proteins. Hair pigments accomplish this protection by absorbing and filtering the impinging radiation and subsequently dissipating this energy as heat. However, in the process of protecting the hair proteins from light, the pigments are degraded or bleached. Dark hair is more resistant to photodegradation than light hair, because of the higher photostability of eumelanin compared to pheomelanin. Integral lipids of hair fibers are degraded by ultraviolet light, as well as by visible light, helping to explain the weakening of the cell membrane complex exposed to light radiation.
... Popular cosmetic procedures such as hair lightening deliberately induce oxidation, but also cause unintended alterations to human hair. These include modifications to the hair keratins and keratin-associated proteins at the molecular level [2,3], which lead to structural degradation such as protein backbone cleavage and imposed crosslinks, as exhibited by protein loss from the hair [3,4]. The accumulation of such perturbations in hair chemistry at the primary residue level thus translates to modification across higher structural levels, and causes changes to the appearance, texture and behaviour of hair such as brittleness, unruliness and loss of lustre, as observable to the consumer [5]. ...
... We measured these effects in bleached hair samples in a variety of ways. At a crude level, the damage to the hair's protein structure is sufficient that proteins and protein components are actually released physically from the fibres [3,4,8,12,20] when whole cells or proteinaceous material are no longer secured to the fibre by protein-protein crosslinks. Increased leachable mass must correspond with the breakage of structural crosslinks in human hair (normally, keratin proteins are very resistant to solubilization [21,22]); it thus provides a proxy measurement for overall structural protein degradation within the fibre. ...
Objective
To understand the structural and chemical effects of cosmetic peroxide bleaching on human hair. Methods
Human hair was progressively bleached using alkaline peroxide‐persulfate treatment. Proteins lost through leaching were examined using amino acid analysis and mass spectrometric sequencing. Fibre damage was assessed using transmission electron microscopy, amino acid analysis, and redox proteomics.
Results
Protein loss through leaching increased with bleaching severity. Leached proteins were not limited to the cuticle, but also included cortical intermediate filaments and matrix keratin‐associated proteins. The leached proteins were progressively oxidised as bleaching severity increased. Bleached fibres demonstrated substantial damage to the cuticle layers and to the cortex. Extensive melanin granule degradation was present after the mildest bleach treatment. Protein oxidation in bleached fibres was principally in cortical intermediate filaments – the most abundant hair proteins – and targeted the sulfur‐containing amino acids, particularly the conversion of cystine disulfide bonds to cysteic acid.
Conclusion
Peroxide chemical treatments quickly access the cortex, causing untargeted oxidative damage across the fibre in addition to the desired loss of melanin. Peroxide ingress is likely facilitated by the considerable structural degradation caused to the cuticle layers of hair fibres. The consequences of the peroxide action within the cuticle and cortex are oxidation of the proteins, and subsequent protein loss from the fibre that correlates to bleaching severity.
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... This corresponds to changes in texture and shine of hair after damage.S100A3 is a calcium-binding protein, expressed primarily inside the hair microstructure of human hair, that plays an essential role in maintaining the structural integrity of hair fibers. Chemically-treated or UV irradiated hair can readily release S100A3 and cause modifications to the hair tissue structure [35,36]. The enrichment analysis of the down-regulated proteins revealed that the damage mainly affected hair cycling, aging, and keratinization. ...
Hair is an advantageous biological sample due to its recordable, collectable, and storable nature. Hair's primary components are keratin and keratin-associated proteins. Owing to its abundance of cystine, keratin possesses impressive mechanical strength and chemical stability, formed by creating disulfide bonds as crosslinks within the protein peptide chain. Furthermore, keratin is cross-linked with keratin-associated proteins to create a complex network structure that provides the hair with strength and rigidity. Protein extraction serves as the foundation for hair analysis research. Bleaching hair causes damage to the structure between keratin and keratin-associated proteins, resulting in texture issues and hair breakage. This article outlines various physical treatment methods and lysate analysis that enhance the efficiency of hair protein extraction. The PLEE method achieves a three-fold increase in hair protein extraction efficiency when using a lysis solution containing SDS and combining high temperatures with intense shaking, compared to previous methods found in literature. We utilized the PLEE method to extract hair from both normal and damaged groups. Normal samples identified 156–157 proteins, including 51 keratin and keratin-associated proteins. The damaged group consisted of 155–158 identified proteins, of which 48–50 were keratin and keratin-associated proteins. Bleaching did not cause any notable difference in the protein identification of hair. However, it did reduce coverage of keratin and keratin-associated proteins significantly. Our hair protein extraction method provides extensive coverage of the hair proteome. Our findings indicate that bleaching damage results in subpar hair quality due to reduced coverage of protein primary sequences in keratin and keratin-associated proteins.
... Inoue et al. [26,27] suggested that UV radiation breaks the disulfide bonds from S100A3 protein, which is a cysteine-rich calcium-binding protein, located mainly in the endocuticle layer. S100A3 is cross-linked to hair keratin via disulfide bridges, thus providing structural integrity to the hair strand. ...
Morphological changes in hair surface are undesirable, since they cause shine loss, roughness increase and split ends. These effects occur more frequently in the cuticle, which is the outermost layer of the hair strand, and thus the most exposed to the environmental damages. Sunlight irradiation contributes significantly to these morphological alterations, which motivates the investigation of this effect on hair degradation. In this work, the influence of irradiation and hand-washing steps on the morphology of pigmented and non-pigmented hair cuticle was investigated using field emission scanning electron microscopy (FESEM) and transmission electron microscopy (TEM). To simulate daily conditions, where hair is hand-washed and light exposed, samples of dark brown and gray hair underwent three different conditions: 1) irradiation with a mercury lamp for up to 600 h; 2) irradiation with the mercury lamp combined with washes with a sodium lauryl sulphate solution; and 3) only washing. A new preparation procedure was applied for TEM samples to minimize natural variations among different hair strands: a single hair strand was cut into two neighbouring halves and only one of them underwent irradiation and washing. The non-exposed half was used as a control, so that the real effects caused by the controlled irradiation and washing procedures could be highlighted in samples that had very similar morphologies initially. More than 25 images/sample were analysed using FESEM (total of 300 images) and ca. 150 images/samples were obtained with TEM (total of 900 images). The results presented herein show that the endocuticle and the cell membrane complex (CMC) are the cuticle structures more degraded by irradiation. Photodegradation alone results in fracturing, cavities (Ø ≈ 20-200 nm) and cuticle cell lifting, while the washing steps were able to remove cuticle cells (≈ 1-2 cells removed after 60 washes). Finally, the combined action of irradiation and washing caused the most severe damages, resulting in a more pronounced cuticle extraction (≈ 1-4 cuticle cells after a 600 h irradiation and a 60 times washing). This irradiation dose corresponds to ca. 2 months of sunlight exposure (considering 5h/day) in Campinas-SP, Brazil, during the day period of maximum irradiation intensity. The combined action of irradiation and washing can be explained by the creation of fragile photodegraded spots in the endocuticle and in the CMC, where the mechanical stress associated to the washing steps are more prone to induce rupture.
... The present study demonstrated that subcutaneous injection with anti-S100A3 antibody led to hair growth retardation and a reduced number of hair follicles (Figures 3-5). Importantly, the loss of S100A3 during daily life results in the delamination of cuticles, especially in distal hair (Inoue et al., 2000). These findings suggest that S100A3 participates in the regulation of the hair cycle, and is associated with hair growth and formation. ...
Using mouse gene expression microarray analysis, we obtained dynamic expression profiles of the whole genome in a depilation-induced hair growth mouse model. S100A3 expression increased during the anagen phase and returned to normal during the telogen phase. The effects of S100A3 blockade on the hair growth cycle were examined in mice after subcutaneous injection of an anti-mouse S100A3 antibody. Protein localization of S100A3 was confined to the hair shafts during the anagen phase and the sebaceous glands during the telogen phase. S100A3 blockade delayed hair follicle entry into the anagen phase, decreased hair elongation, and reduced the number of hair follicles in the subcutis, which correlated with the downregulated expression of hair growth induction-related genes in vivo. The present study demonstrates that anti-S100A3 antibody inhibits mouse hair growth, suggesting that S100A3 can be used as a target for hair loss treatment.
... The S100A3 protein in hair has been shown to strongly bind both calcium and zinc and to be localized in the endo-cuticle with some expression in the cortex matrix [14]. Inoue and Kizawa have hypothesized that it is important for cuticle-cuticle cell adhesion [15] and it has also been demonstrated that it can be liberated by chemical treatments. PYMOL software (DeLano Scientific, Portland, Oregon, USA) was used to , bleached (middle) and UV-exposed (bottom) hair fibres. ...
Damage to hair from UV exposure has been well reported in the literature and is known to be a highly complex process involving initiation via absorption of UV light followed by formation and propagation of reactive oxygen species (ROS). The objective of this work was to understand these mechanisms, explain the role of copper in accelerating the formation of ROS and identify strategies to reduce the hair damage caused by these reactive species.
The location of copper in hair was measured by Transmission electron microscopy (TEM) x-ray energy dispersive spectroscopy (XEDS) and levels measured by ICP-OES. Protein changes were measured as total protein loss via the Lowry assay and MALDI ToF was used to identify the biomarker protein fragments. TBARS assay was used to measure lipid peroxide formation. Sensory methods and dry combing friction were used to measure hair damage due to copper and UV exposure and to demonstrate the efficacy of EDDS and histidine chelants to reduce this damage RESULTS: In this work a biomarker protein fragment formed during UV exposure is identified using mass spectrometry. This fragment originates from the calcium binding protein S100A3. Also shown is the accelerated formation of this peptide fragment in hair containing low levels of copper absorbed from hair during washing with tap water containing copper ions. Transmission electron microscopy (TEM) x-ray energy dispersive spectroscopy (XEDS) studies indicate copper is located in the sulfur-poor endo-cuticle region; a region where the S100A3 protein is concentrated. A mechanism for formation of this peptide fragment is proposed in addition to the possible role of lipids in UV oxidation. A shampoo and conditioner containing chelants (EDDS in shampoo and histidine in conditioner) is shown to reduce copper uptake from tap water, reduce protein loss and formation of S100A3 protein fragment. In addition, the long term consequences of UV oxidation and additional damage induced by copper are illustrated in a four month wear study where hair was treated with a consumer relevant protocol of hair coloring treatments, UV exposure and regular shampoo and conditioning.
The role of copper in accelerating UV damage to hair has been demonstrated as well as the ability of chelants such as EDDS and histidine in shampoo and conditioner products to reduce this damage. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
... After extracts in sample buffer containing 150 mM dithiothreitol were incubated at 90°C for 5 min, the protein extract was size-separated on a 16.5% Tricine±sodium dodecyl sulfate±polyacrylamide gel (Scha Ègger and von Jagow, 1989). The electro-transferred membrane was incubated with either anti-S100A4 or S100A6 antibody (1 mg per ml) at 4°C for 16 h, and then bound antibodies were detected as previously described (Inoue et al, 2000). ...
In order to elucidate the onset mechanisms of hair regrowth, we characterized the expression of metastasis and cell-growth-associated calcium-binding S100 proteins in the regenerating follicular epithelium. Hair-cycle-dependent expression of S100A4 and S100A6 was found in the epithelial sac regions (bulge area and hair germ) of mouse pelage follicles. Protein localization of S100A4 was confined to the bulge area, the region where the presumed follicular stem cells reside, during the catagen-telogen-anagen transitional periods, whereas S100A6 protein was distributed throughout the epithelial sac regions during anagen phase. Prior to entering anagen phase, however, both S100 mRNAs were upregulated in the epithelial sac. Despite the induction of extensive cell death in the bulge region after plucking, a new hair cycle was initiated following transcription of S100A6 in the hair germ. Upon wounding stimuli, both the outer root sheath and the basal layer of epidermis expressed S100A6 mRNA prior to hyper-proliferation. Once the epithelial sac was induced to transcribe both S100 genes, resting follicles were concomitantly rejuvenated. These results suggest that S100A4 and S100A6 might play important roles in the activation of stem cells at the onset of follicle regeneration.
Objectives
Oxidative agents used in conventional hair perming damage hair fibres. This study proposes a novel perming approach employing 1,4‐butylenediamine bismaleimide (1,4‐BB) as a cross linking agent via thiol‐Michael click chemistry to avoid oxidative harm. The perming performance and effects on hair fibre properties were investigated and compared.
Methods
1,4‐BB was synthesized via an acylation reaction and evaluated for cytotoxicity using the MTT assay. Perming efficiency was compared on reduced hair cross linked by 3 wt% H 2 O 2 , 6 wt% NaBrO 3 and 0.5 wt% 1,4‐BB, respectively. Chemical changes during different perming methods were analysed via Raman spectroscopy, fluorescence microscopy, XRD and FT‐IR characterization. Perming efficiency and durability were evaluated and explained by calculating the ‐SH connection ratio via X‐ray photoelectron spectroscopy. Mechanical properties, colour and surface properties of hair samples permed by different methods were evaluated and compared.
Results
1,4‐BB was successfully prepared and exhibited no cytotoxicity over a wide concentration range. Click perming using 1,4‐BB exhibited comparable perming efficiency and superior durability compared to oxidative perming methods. Higher α ‐helix keratin content and reduced sulfonate formation contributed to the preservation of hair mechanical properties. Click perming also benefited from the preservation of hair colour and surface properties, including morphology and hydrophobicity.
Conclusion
Thiol‐Michael click perming offers a non‐oxidative alternative with reduced hair damage and enhanced perming durability.
The proteins in human hair consist primarily of microfibrillar keratins with a molecular mass of 40-65 kDa and keratin-associated proteins (KAPs) with a molecular mass of 6-30 kDa, according to the results obtained from sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Because an effective purification procedure of KAPs has not been established, little is known about the protein chemistry of KAPs as compared with that of keratin. When hair samples were incubated in the Shindai solution containing alcohols such as methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, and 2-methyl-1-propanol, the extraction of KAPs was enhanced, while extraction of keratin was suppressed. Using ethanol, we established a selective purification procedure for KAPs and keratin. According to Tricine/SDS-PAGE, the KAPs fraction contained six polypeptides with molecular masses of 3.5, 4.4, 5.2, 7.8, 15, and 28 kDa. The keratin fraction contained two polypeptides with molecular masses of 45 and 67 kDa and was free of low-molecular-weight components. The amino acid compositions of the KAPs and keratin fractions were mostly in agreement with the values found in the literature. The recoveries of the KAPs and keratin fractions from the hair samples were approximately 10 and 50%, respectively. Scanning electron microscopy (SEM) showed that hair samples retained fine fibrous structures in the cortex after extracting the KAPs and that the additional extraction of keratin caused the fibrous structures to disappear. These results indicated that KAPs may function by surrounding the fibrous structures and supporting the keratin fibers in the cortex. In this study, we propose a novel and convenient isolation procedure for KAPs and keratin from human hair.
Hair photoaging is shown to consist of a number of concurrent processes that result in chemical and physical changes in fiber properties. Lipid oxidation, disulfide bond cleavage, tryptophan degradation, and cysteic acid formation lead to an increase in fiber porosity, loss of mechanical strength, and an increase in surface roughness. Hair exposed to sunlight is claimed to be more brittle, stiffer, and drier than before irradiation and exhibits a reduced water absorption capacity. Photochemical alteration includes the breakdown of disulfide bridges within the structural units of hair and the establishment of new intra and intermolecular cross links via reaction of carbonyl groups with protein amino groups within and between structural units, thereby decreasing structural integrity. These reactions most likely lead to a gradual increase in brittleness and a gradual loss of structural differentiation. Photodegradation of cystine occurs through the C–S fission pathway and the highest level of photodegradation occurs in the cuticular region, where cystine is present at its highest concentration. For hair damaged by sunlight, in most cases, the amino acids of the cuticle are altered to a greater extent than the amino acids of the cortex, because the outer layers of the fiber receive higher intensities of radiation. Proteins of the cuticle are degraded by ultraviolet A and ultraviolet B, but much less by visible (VIS) light. Hair pigments function to provide some photochemical protection to hair proteins. Hair pigments accomplish this protection by absorbing and filtering the impinging radiation and subsequently dissipating this energy as heat. However, in the process of protecting the hair proteins from light, the pigments are degraded or bleached. Dark hair is more resistant to photodegradation than light hair, because of the higher photostability of eumelanin when compared with pheomelanin. But, hair damages caused by ultraviolet (UV) exposure are related not only to the melanin type of each hair but also to the total amount of melanin. Pheomelanin is far more sensitive to UV light than eumelanin, though these two types of melanin are similarly sensitive to VIS light. UVA irradiation can penetrate deeply into the cortex, so photochemical changes, including cuticles and cortex together, may appear greater after UVA irradiation. On the other hand, UVB causes severe morphological damages, especially confined to the hair cuticles because of its restricted depth of penetration. Integral lipids (ILs) of hair fibers are degraded by UV light as well as by VIS light, helping to explain the weakening of the cell membrane complex (CMC) exposed to light radiation.
Deimination is a relatively new post-translational modification of proteins, whose recognition is ever-increasing. First linked to the pathology of rheumatoid arthritis (RA), deimination is a process by which selected positively charged arginine amino acids are converted to neutral citrulline amino acids by the peptidyl arginine deiminase (PAD) family of enzymes.
Although the medical literature is rich with articles about the possible significance of deiminated proteins in RA, Protein Deimination in Human Health and Disease is the first publication to compile this knowledge and the growing amount of new information now known about the presence of deiminated proteins in the eye, skin, hair, gums, lung and nervous system, as well. As a result, this process has now been linked to numerous additional conditions besides RA, including cancer, glaucoma, Alzheimer's disease, Parkinson's disease, multiple sclerosis, spinal cord and peripheral nerve injury, Creutzfeldt-Jakob disease, among many others.
Chronicling the earliest studies of deimination up to the present, this volume distills what is currently known about citrullination of proteins in the human body and is the first book of its kind on the topic.
This study was conducted to establish a simple efficient method for extracting the protein from human head hair materials, which can be a useful tool for the protein analysis applicable to various types of human head hairs. The method developed saves extraction time and effort considerably. The method includes four steps: cutting the hair samples into small pieces 1-2 mm in length, washing them with distilled water, incubating the hair samples in a buffer solution at 50 degrees C for 24 h and finally filtering the incubated mixtures through three layers of nylon mesh. This method is reproducible and reliable. SDS-PAGE analysis of the hair protein extracted by this method shows a clear protein profile on the gel, which is frequently observed in other hair sources. Two smaller sizes of molecular weights are also detected with the SDS-PAGE analysis. Not commonly found in other hair sources, they seem to be other types of human hair proteins.
Partial protein extracts were prepared from hair, nail, and stratum corneum in the absence of urea and interfacial surfactant. Tricine-sodium dodecyl sulfate polyacrylamide gel electrophoreses of these extracts showed low-molecular weight protein-rich patterns apparently different from those of whole protein extracts, which mainly consist of keratin bands. Several protein bands characterized each keratinized tissue or its derived species. In addition, we identified a major band of approximately 7 kDa as ubiquitin, a ubiquitously distributed protein that mediates non-lysosomal protein degradation, through direct amino acid sequence analysis of the electro-blotted protein band. The partial extraction is useful for investigation of soluble proteins retained in the keratinized tissues.
S100A3, a unique protein among all members of the calcium-binding S100 family, is specifically expressed at the inner endocuticle of human hair fibers. Upon hair damage, S100A3 is released from hair fibers and possibly destabilizes the hair tissue architecture. This study describes the purification and characterization of native S100A3 isolated from human hair fibers. We extracted native S100A3 from cuticles and purified the protein by anion-exchange chromatography. The results of 2D gel electrophoresis showed that cuticle S100A3 has a slightly lower isoelectric point compared to the recombinant protein. Tandem mass spectrometry of the peptides resulting from endoproteinase digest of cuticle S100A3 revealed that the N-terminal methionine is replaced with an acetyl group. This is the first report on biochemical characteristics of S100A3 in hair cuticle.
We previously found that certain hair proteins were soluble by means of a partial extraction method. In this study, we demonstrate that the amount of soluble proteins internally formed in permed and bleached hair, labile proteins, is a useful index for hair damage assessment. Compared to tensile property changes, this index rose in widely dynamic ranges as the time of either permanent waving or bleaching treatments increased. The amount of labile proteins was much larger than that of proteins eluted into perming and bleaching lotions. However, the labile proteins showed electrophoretic profiles similar to those of the eluted proteins. These results suggest that a portion of the stable proteins in normal hair was transformed into labile proteins upon permanent waving and bleaching treatments. Consequently, permed and bleached hair tends to release the resultant labile proteins.
Gephyrin is a bifunctional protein which is essential for both synaptic clustering of inhibitory neurotransmitter receptors in the central nervous system and the biosynthesis of the molybdenum cofactor (MoCo) in peripheral tissues. Mice deficient in gephyrin die early postnatally and display a loss of glycine receptors (GlyRs) and many GABA(A) receptor (GABA(A)R) subtypes from postsynaptic sites. In addition, the activities of the MoCo-dependent enzymes xanthine dehydrogenase and sulfite oxidase are reduced to background levels in the liver and intestine of these animals. To genetically separate the different consequences of gephyrin deficiency, we expressed a transgene of the plant homolog Cnx1, known to rescue mammalian MoCo deficiency, on the background of gephyrin knockout mice. Cnx1 partially restored sulfite oxidase activity in the liver of the transgenic animals, whereas early lethality and the loss of GlyR clustering were unaltered. Our data suggest that the loss of neurotransmitter receptor clustering at inhibitory synapses causes the early lethality of gephyrin deficient mice.
This article describes the current understanding of the effect of ultraviolet (UV) radiation and visible light on the structure and integrity of human hair fibers; furthermore, it discusses current and past approaches to the protection of hair from UV rays. Relevant literature is reviewed.
The existence of holes in the endocuticle, in the inter-macrofibrilar matrix and in the cell membrane complex of hair fibers is described. In this paper, we show that these holes are absent in the hair follicle. However, cell remnants found in young hair inside the follicle are located in the same regions and are of the same dimensions as the holes observed in mature fibers. Since holes in the endocuticle have been described to appear as a result of daily care actions, it is probable that the development of holes in the inter-macrofibrilar matrix and in the cell membrane complex is related to the removal of extractable substances. We discuss hole formation in terms of the nature of the extractable substances and the presence of cell remnants in the young fibers.
Fluorescence spectroscopic measurement of the amino acid tryptophan on the hair fibre surface was extended to include in situ fibre irradiation and a novel cyclical wash off, reapplication protocol. When applied to the investigation of a new damage prevention active, it was shown that the active was preferentially degraded in a sacrificial manner and that the underlying fibre surface was maintained in good condition. In addition, tensile strength measurements were performed to assess the mechanical properties of the treated and untreated fibres following UV and sunlight exposure and the results demonstrate the damage prevention effectiveness of the active.
Pilomatrixoma is a common benign cutaneous tumour containing differentiated hair matrix cells. This tumour is mainly composed of basophilic, transitional, shadow and squamoid cells. Although some S100 proteins are expressed in a tissue-specific manner in the hair follicle (e.g. S100A2 in the outer root sheath, S100A3 in the cortex and cuticle, and S100A6 in the inner root sheath), little information is available concerning their distribution in the aberrantly differentiated tissues of pilomatrixoma.
To characterize the disordered epithelial elements of pilomatrixoma by localizing S100A2, S100A3 and S100A6 proteins.
Immunohistochemistry and dual-immunofluorescence microscopy were performed on 22 pilomatrixoma specimens using antibodies specific to the three proteins.
Tissue-specific distribution of the S100 proteins investigated was preserved in the morphologically disordered tumour tissues. Anti-S100A2 antibody stained squamoid cells and putative outer root sheath cells; basophilic and potential hair matrix cells were occasionally stained. S100A3 staining was found in transitional cells and putative cortical cells, and was strong in both dispersed cells and hair-like structures surrounding cells which were presumably cuticular cells. Anti-S100A6 antibody labelled some S100A3-negative transitional cell strands, potentially inner root sheath cells.
The epithelial elements of pilomatrixoma can be characterized using S100 proteins as biochemical markers. Our results show that pilomatrixomas retain a certain degree of differentiation indicative of distinct hair-forming cells.
Hair treatment chemicals induce sudden and severe hair damage. In this study, we examined cuticles from untreated, permed, and bleached hair that were mechanically discriminated by shaking in water. Both perming and bleaching treatments are prone to easily delaminate cuticles. Confocal microscopy revealed that the cuticles of permed hair were delaminated with larger pieces than untreated ones. On the other hand, the cuticles of bleached hair tend to fragment into small peptides. At the minimum concentration of thioglycolate required to elute S100A3 protein from the endocuticle into the reductive permanent waving lotion, enlarged delaminated cuticle fragments were observed. Although S100A3 is retained in bleached hair, S100A3 is irreversibly oxidized upon bleaching treatment. It is likely that the oxidative cleavage of disulfide bonds between cuticle-constituting proteins, including S100A3, results in the fragile property of cuticles. Here we present a more comprehensive model of hair damage based on a diverse mechanism of cuticle delamination.
The human genome contains large regions that are highly structured. Sequence-related members of multigene families are often found in a clustered organization. Here we describe a previously unrecognized gene cluster composed of genes coding for calcium-binding proteins of the S100 family. The linkage of six S100 genes was established by pulsed-field gel electrophoresis, and a contiguous DNA sequence of 15 kilobases containing the full coding region of four different S100 genes was characterized. This is the tightest mammalian gene cluster discovered so far to our knowledge. Two additional S100 genes are located within the cluster, both of which exhibit unique structural features when compared with other S100 genes. The product of S100E is cysteine-rich, whereas that of S100D contains a long hydrophobic N-terminal tail. The gene cluster was assigned to chromosome 1q21, one of the bands showing rearrangements in neoplasms at high frequency. The deregulated expression of some S100 genes in the cluster during tumor progression suggests that chromosomal abnormalities may influence the expression of S100 genes in late stages of cancer, particularly in association with the formation of metastases.
A protein determination method which involves the binding of Coomassie Brilliant Blue G-250 to protein is described. The binding of the dye to protein causes a shift in the absorption maximum of the dye from 465 to 595 nm, and it is the increase in absorption at 595 nm which is monitored. This assay is very reproducible and rapid with the dye binding process virtually complete in approximately 2 min with good color stability for 1 hr. There is little or no interference from cations such as sodium or potassium nor from carbohydrates such as sucrose. A small amount of color is developed in the presence of strongly alkaline buffering agents, but the assay may be run accurately by the use of proper buffer controls. The only components found to give excessive interfering color in the assay are relatively large amounts of detergents such as sodium dodecyl sulfate, Triton X-100, and commercial glassware detergents. Interference by small amounts of detergent may be eliminated by the use of proper controls.
This work was concerned with the characterization of nonkeratinous and keratinous proteins of human hair eluted by permanent wave lotion I (PWL-I). Polyacrylamide gel electrophoresis, infrared spectroscopy and amino acid analysis method were applied.From these examinations, the following results were obtained. (1) Nothing was eluted from human hair by hot water (60°C, 3hr treatment), whereas some extracts were obtained using alkaline solution, surfactant solution, or PWL-I (permanent wave lotion I). (2) The mass of the proteins extracted by PWL-1 accounted for about 1% of the total mass of the hair and tended to increase with an increase in pH. (3) It was confirmed that the molecular weights ranged from 5, 000 to 65, 000 by SDS PAGE. (4) The amino acid composition of the PWL-1 extracts was similar to that of colagenous proteins which was unexpected. The composition was diffrent from that of intercuticular material, δ, eluted by formic acid. Therefore, we suspected that the PWL-1 extract was eluted from the δ layer among the cortical cells.In addition, it was clarified that a part of keratinous protein was extracted from hair which was damaged by repeated treatments of PWL or breached by hydrogen peroxide and ammonia water. It is thought to be eluted from the matrix in the cortical cell which is composed of amorphous keratin of high sulfer content.These results will help clarify the mechanism of the elution of intercellar material of humann hair and the mechanism of hair damage by external treatments.
A simple and sensitive procedure amenable to visual or spectrofluorometric quantitation has been developed to detect certain types of hair damage. We have found that hair treated with the fluorescent dye 1-dimeth- ylamino-naphthalene-5-sulfonyl chloride (dansyl chloride) acquires fluorescence under ultraviolet light. The fluorescence intensity is a function of the amount of dansyl chloride binding to the hair and can be measured in a spectrofluorometer at very low levels. Our studies have shown that changes in the fluores- cence emission of dansylated hair indicate hair damage. Using this procedure, we were able to demon- strate statistically significant differences in hair damaged due to (i) weathering, (ii) ultraviolet light, and (iii) chemical bleaching.
We have developed a new method of fractionating hair components in order to analyze the damaged components and the degree of damage due to perming. We found that the amount of constituent proteins extracted by an anionic surfactant with reductant was influenced by the concentration of the reductant. Using this method, the matrix and the microfibril protein could be easily separated and quantified. Applying this method to the analysis of individual hairs, we found a significant decrease in the "intact" microfibril protein on the tip end of permed hair.
This article describes a simple yet sensitive technique to assess surface damage to hair. It is based on the hypothesis that the damaged hair surface is more susceptible to abrasion/erosion than undamaged hair, and involves shaking hair in water and quantitatively measuring the amount of protein abraded/eroded from the hair using a colorimetric procedure capable of detecting as little as 5 Ixg of protein per mi. With this procedure, we were able to demonstrate significant differences in hair damaged due to bleaching, permanent wave treatments, and Suprox (a diperisophthalic acid based oxidant), and even after extended treatment with different surfactants. Furthermore, hair with exposed cortex was found to be more susceptible to protein loss by surfactants and/or water than hair with intact cuticle.
In a recent publication, Kaplin et al. described the dissolution of ultrastructural components in hair fibers as a direct result of various cosmetic treatments. These observations were based on a subjective assessment of holes or voids observed in electron micrographs of hair fiber cross sections. We have further investigated these effects using electron microscopy in conjunction with image analysis. Cross sections of proximal and distal ends of intact hair fibers were examined before and after repeated shampooings. The total number of holes, total projected areas, mean areas, and size distributions were determined in the cuticle and cortex regions of hair fibers using a Quantimer 900 Image Analyzer. All measured parameters indicated that ultrastructural disruption increased from the proximal to the distal end of hair fibers. The impact of shampooing was limited to the cuticle region of weathered hair fibers.
Synopsis--Some PHYSICO-CHEMICAL PROPERTIES of CUTICLE are described and cor- related to the function of the latter in the overall STRUCTURE of HAIR. Chemical analy- sis of the cuticular layer revealed significant differences in the amino acid composition of the cuticle and the cortex of the hair KERATIN. The cuticle is extensively crosslinked by cystine, is more hydrophobic than the cortex, and contains large quantities of serine, glycine, and proline. All this satisfactorily accounts for the function of the cuticle as a molecular sieve and a chemical barrier. which as an industrial commodity commands the resources necessary for extensive biochemical and physical investigations. The similarity in morphology and chemical properties of mammalian fibers provides a sci- entifically acceptable rationale for regarding the results on wool as ap- plicable to all of them, although such generalization from a specific case can be misleading. For example, it has become customary to focus one's attention almost exclusively on the ultrafine structure and properties of the cortex, which is the dominant structural element of the wool fiber. However, the cor- tex appears to be more important in wool than it is in hair. Conse- quently, when one is concerned with the latter, the properties of the re- maining morphological components (epicuticle, cuticle, cell inembranes, medulla) should be taken more fully into account. In an attempt to do this, we are presenting here a brief discussion concerning the contribu- tion of the cuticle to some of the physico-chemical parameters of human hair.
Synopsis---The CUTICLE of human HAIR has been isolated in bulk by a new method involving vigorous agitation of fibres in water. The cuticle fractions have been shown to be of high morpho- logical purity using various techniques of ELECTRON MICROSCOPY. The significance of AMINO ACID ANALYSES is discussed.
During the extension of keratin fibers, their two major morphological components, the cuticula and the cortex, accommodate the stresses imposed on the fiber each in a totally different fashion. While the latter extends by mechanisms that have been discussed extensively and appear to be well understood, the cuticle cells are essentially inextensible and have to move relative to one another. In the multilayer structure of the cuticular sheath of human hair fibers, this relative movement has to be accommodated by the various layers within each cuticle cell and by the bonding layers between cells, and finally causes the lifting of surface scale edges at higher strain levels. It is proposed that extension mainly causes shear stresses between layers of different composition and extensibility within the cuticle cell. This leads to failure in the weak endocuticular layer and results in "delammation" and lifting of the outer layers of the surface cuticle. The damage is irreversible upon release of the fiber and immersion in water, as reflected in the onset of scale lifting at considerably lower strain levels during a second extension. Scale lifting was not observed during the extension of wool fibers, which appears to be a reflection of the higher rigidity of the cuticle cells of wool.
A large-scale procedure was developed for the anaerobic purification of the human recombinant Ca2+- and Zn2+-binding protein S100A3 for spectroscopic studies. S100A3 eluted as a non-covalently bound dimer (20.8 kDa). It contained 7.5±0.1 free thiol groups/monomer, and bound Ca2+ with a Kd of ≈4 mM, which corresponds to a tenfold increase in affinity compared to the aerobically purified protein. The transition metal ions Co2+, Zn2+ and Cd2+ were used as spectroscopic probes to investigate the role of the 10 cysteine residues per monomer S100A3 in metal binding. Spectrophotometric titrations suggest the formation of dinuclear thiolate-bridged clusters consisting of a Me2+(SCys)4 and a Me2+(SCys)3(NHis) site as described for zinc finger proteins. A three-dimensional structural model of S100A3 was proposed on the basis of the NMR structure of the structurally related rabbit S100A6 protein, and taking into account the structural influence of cysteine residues.
Synopsis
Some previously unreported fine variations in the form of normal hairs are described as they are observed in the scanning electron microscope (SEM). They all arise in the course of surface frictional wear and the chipping away of the hair surface scale edges and include remanent surface impressions of cuticle scale edges, ‘false’ scale edges, granular surface remnants and highly irregular (chevron) scale patterns. Mechanisms are proposed for the way in which each of these different fine features arise.
The paper also contains deliberations on the conditions for operating the SEM consistent with obtaining the best information about the architecture of hair surfaces. The correctness of viewing orientation of scanning electron micrographs is also emphasized to avoid misinterpretation of features on the hair surface.
A discontinuous sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) system for the separation of proteins in the range from 1 to 100 kDa is described. Tricine, used as the trailing ion, allows a resolution of small proteins at lower acrylamide concentrations than in glycine-SDS-PAGE systems. A superior resolution of proteins, especially in the range between 5 and 20 kDa, is achieved without the necessity to use urea. Proteins above 30 kDa are already destacked within the sample gel. Thus a smooth passage of these proteins from sample to separating gel is warranted and overloading effects are reduced. This is of special importance when large amounts of protein are to be loaded onto preparative gels. The omission of glycine and urea prevents disturbances which might occur in the course of subsequent amino acid sequencing.
Analyses on sodium dodecyl sulfate-polyacrylamide gel electrophoreses showed that the human hair cuticle extracts mainly consist of a 7-kDa component and keratin proteins. The S-carboxymethylation of the cuticle extracts made the 7-kDa band shift to the 15-kDa position. After electroblotting of the S-carboxymethyl derivative, the membrane pieces carrying the 15-kDa band were treated with trypsin and the released peptides were separated by reverse-phased HPLC. Amino acid sequence analyses revealed that the peptides corresponded to the partial sequences deduced from human genome coding for S100A3, a cysteine-rich calcium binding protein. The anti S100A3 serum, prepared by immunizing a synthetic peptide antigen, reacted with the 7-kDa and 15-kDa bands in immunoblotting analyses. Immunofluorescence microscopy showed intense labeling to the cuticular layer with the anti S100A3 serum. These results indicated that S100A3 was highly expressed in the human hair cuticle.
We have previously identified a cysteine-rich calcium binding protein S100A3 present in the cuticle of human hair fiber. In this study, we cloned a cDNA for mouse S100A3, identified its gene location, and elucidated the expression profile throughout hair follicle development. The mouse S100A3 gene was clustered with other S100 family members on chromosome 3, and specifically expressed in dorsal skin containing hair follicles. The level of S100A3 mRNA was elevated during the anagen phase of the hair growth cycle, and sharply declined from the regression phase on. In situ hybridization revealed that the S100A3 gene was prominently expressed in cuticular cells of the hair follicle, and mRNA levels were highest in the keratogenous zone over the entire cuticular layer. Expression was also observed to a lesser extent in differentiated cortical cells; however, expression was not observed in any other component of the hair follicle or dorsal tissues. Immunohistochemical analysis showed that the S100A3 protein accumulated in cuticular and cortical cells undergoing terminal differentiation. These results indicate that the S100A3 gene is exclusively expressed, and the translation product retained, in follicular cells differentiating into major components of the hair shaft. It seems likely that S100A3 plays an important role in calcium-dependent processes leading to hair shaft formation.
A soluble protein characteristic of the nervous system Gene expression of mouse S100A3, a cysteine-rich calcium binding protein, in developing hair follicle Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa
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Dissolution of proteins from hair. II: The analysis of proteins dissolved into permanent waving agent and the evaluation of hair damage A sensitive fluorescence technique using dansyl chloride to assess hair damage
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Analysis of the damaged components of permed hair using biochemical technique Probing the structure of the human Ca 2+ and (21) I Effects of cosmetic treatments on the ultrastructure of hair, (22) C. Dubief, Experiments with hair photodegradation
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