Immunohistochemical localization of androgen and oestrogen receptors in canine hair follicles

Skin biopsies from seven different body sites were obtained from 21 dogs of different breeds (short, normal, long hair), which were presented for euthanasia. Commercially available polyclonal antibodies were used for the immunohistochemical detection of androgen and oestrogen receptors. Both receptors showed a similar distribution in canine skin, with specific intranuclear staining. In the epidermis, the percentage of androgen receptor (AR)-positive cells, but not that of oestrogen receptor (ER)-positive cells, was significantly higher in samples from the thorax and the flank. In the dermal papilla, the percentage of ER-positive (but not AR-positive) cells was significantly lower in biopsies from the flank. No significant difference was found for both receptors between the locations in the outer root sheath, among the three different hair types, between sex and between intact and castrated dogs.     

Follicle and fibre characteristics of Indian pashmina goats

The hair follicle and fibre characteristics of Cheghu and Changthangi pashmina goats are reported. The average numbers of primary follicles mm-2 were 6.80, 5.20, 5.67 and 4.00; secondary follicles 42.40, 27.10, 38.56 and 29.67; secondary to primary follicle ratio 6.46, 5.78, 6.79 and 7.60; total follicle population 49.20, 32.30, 44.22 and 33.67 in Cheghu females, Cheghu males, Changthangi females and Changthangi males, respectively. The breed difference was not significant for any of these traits, but the sex difference was significant for a number of secondary follicles and total density. Study of fleece characteristics of Cheghu goats in a 1 cm2 area of five body regions, namely, neck, shoulder, mid-side, flank and abdomen, showed that females had finer fibres (12.06 to 12.12 micron) than males (13.04 to 13.64 micron) in all regions. The pashmina length, pashmina diameter and fibre thickness ratio were significantly different between sexes and between body regions but hair/pashmina ratio and pashmina yield were significant for sex and body regions, respectively.     

Effects of Photoperiod on Stem Cells Activation in Cashmere Goat Hair Follicles

In order to extend the anagen of cashmere goat hair follicles and increase the production of cashmere,this study was performed with artificially shorten the daylight time among Arbas White cashmere goats. Skin tissue sections from cashmere goats were collected to compare the morphologic changes between artificial daylight and natural daylight,and immunohistochemical method was used to study the hair follicle cell proliferation and important protein expression in related signaling pathways. The results showed that strong cell proliferation occurred in cashmere goat hair follicle cells during artificial daylight,plenty of cytokeratin 15 (K15) positive signals were distributed in the outer root sheath,β-catenin protein was actively expressed in hair matrix and root sheath, indicating that the hair follicles were in the anagen growth phase;Meanwhile,cashmere goat hair follicles under natural daylight were in telogen with weak signals. Above all prove that short photoperiod played an important role in promoting hair follicle growth,the artificial short photoperiod could change hair follicle growth cycle and make hair follicles earlier enter to the anagen growth phase,causing a variety of typical gene expressions during hair follicle growth.     

光周期对于绒山羊毛囊干细胞激活的影响

为了延长绒山羊毛囊兴盛期,提高羊绒产量,本试验通过人工缩短内蒙古阿尔巴斯型绒山羊的日照时间,利用组织切片技术对比人工短光照和自然光照周期下绒山羊皮肤组织形态变化,利用免疫组织化学方法研究毛囊细胞增殖及相关信号通路重要蛋白的表达。结果显示,人工短光照周期下绒山羊毛囊细胞增殖强烈,干细胞标记角蛋白15（cytokeratin 15,K15）阳性信号大量分布于外根鞘,β-catenin信号活跃表达于毛母质及根鞘,毛囊提前进入兴盛期;而自然光照周期下绒山羊毛囊仍停留在休止期,相应蛋白信号表达较弱。综上表明,短日照对毛囊生长具有明显的促进作用,人工短光照周期可以提前激活毛囊干细胞,使其提前进入兴盛期,引起多种毛囊生长相关蛋白的表达。     

CD34 glycoprotein identifies putative stem cells located in the isthmic region of canine hair follicles

It is widely documented that a pool of multipotent stem cells located in humans and mice hair follicle outer root sheath (bulge region) is involved in the restoration of the whole follicular unit during each anagen phase. To the authors' knowledge, data regarding the location and characterization of hair follicle stem compartment in dogs have not been reported in the recent relevant literature. In this study, we investigated the haematopoietic stem and progenitor cell antigen CD34 as a marker of putative stem cells located in a bulge-like region of canine hair follicles. The presence of CD34 mRNA and glycoprotein was assessed on formalin-fixed, paraffin-embedded canine skin samples by in situ hybridization technique and by standard immunohistochemistry, respectively. A strong expression of CD34 mRNA and glycoprotein was observed in a well-defined area of the hair follicle isthmic region and appeared uniformly concentrated at the level of the basal layer of the outer root sheath. These findings provide compelling support to the hypothesis that in dogs, a subpopulation of basal keratinocytes located in the hair follicle isthmic region and characterized by the selective expression of CD34 is potentially associated with the stem cell compartment of this skin appendage.     

Cell proliferation kinetics in the hair root matrix of dogs with healthy skin and dogs with idiopathic seborrhea

Cell proliferation kinetic values were established for the hair root matrix of primary anagen follicles of 14 Beagles and 4 Cocker Spaniels with healthy skin and 9 Cocker Spaniels with primary idiopathic seborrhea. Indices were established by intradermal pulse labeling with tritiated thymidine, followed by cutaneous biopsy and autoradiography. The hair root matrix cell labeling index was 23.4 +/- 3.5% for Beagles, 24.4 +/- 4.0% for healthy Cocker Spaniels, and 24.9 +/- 4.3% for seborrheic Cocker Spaniels. These values indicate a rapidly proliferating cell population. Differences among these cell kinetic data for the 3 groups of dogs were not statistically significant. Although significant cell kinetic differences have been reported for other epidermal structures (interfollicular epithelium, upper hair follicle external root sheath, sebaceous glands) in seborrheic Cocker Spaniels, proliferation of hair root matrix cells apparently remains unaffected.     

The canine hair cycle - a guide for the assessment of morphological and immunohistochemical criteria

The hair follicle has a lifelong capacity to cycle through recurrent phases of controlled growth (anagen), regression (catagen) and quiescence (telogen), each associated with specific morphological changes. A comprehensive classification scheme is available for mice to distinguish the cycle stages anagen I-VI, catagen I-VIII and telogen. For dogs, such a classification system does not exist, although alopecia associated with hair cycle arrest is common. We applied analogous morphological criteria and various staining techniques to subdivide the canine hair cycle stages to the same extent as has been done in mice. Of all the staining techniques applied, haematoxylin and eosin stain, Sacpic, Masson Fontana and immunohistochemistry for vimentin and laminin proved to be most useful. To evaluate the applicability of our criteria, we investigated skin biopsies from healthy beagle dogs (n=20; biopsies from shoulder and thigh) kept in controlled conditions. From each biopsy, at least 50 hair follicles were assessed. Statistical analysis revealed that 30% of the follicles were in anagen (12% early and 18% late), 8% in catagen (2% early, 5% late and 1% not determinable) and 27% in telogen. Thirty-five per cent of hair follicles could not be assigned to a specific cycle stage because not all follicles within one biopsy were oriented perfectly. In conclusion, this guide will not only be helpful for the investigation of alopecic disorders and possibly their pathogenesis, but may also serve as a basis for research projects in which the comparison of hair cycle stages is essential, e.g. comparative analysis of gene expression patterns.     

水貂毛皮成熟期皮肤中EGF、IGF-Ⅰ及IGF-ⅠR基因的表达

为探讨水貂皮肤和毛囊发育机理,试验运用组织学和免疫组化方法对水貂毛皮成熟期皮肤及毛囊中表皮生长因子（epidermal growth factor, EGF）和胰岛素样生长因子-Ⅰ（insulin-like growth factor-Ⅰ,IGF-Ⅰ）及其受体（insulin-like growth factor-Ⅰreceptor,IGF-ⅠR）进行了研究。结果表明,水貂皮肤表皮细胞和毛囊外根鞘细胞中都有IGF-Ⅰ、IGF-ⅠR和EGF基因的强阳性表达,3种基因主要分布于细胞质中,细胞核呈空泡状未见到阳性染色,呈苏木精复染的蓝色,皱褶区域和表皮下胶原结缔组织出现了阳性染色为非特异性阳性结果。IGF-Ⅰ、IGF-ⅠR和EGF基因在水貂皮肤表皮和毛囊外根鞘细胞中广泛表达,直接参与调控皮肤和毛囊的发育。表明EGF、IGF-Ⅰ和IGF-ⅠR基因在水貂皮肤及其衍生结构的发育中发挥着重要作用。     

幼年牦牛不同部位皮肤毛囊的组织学结构及TGF-β2与HIF-1α差异性表达分析

旨在探究幼年牦牛皮肤毛囊的组织学结构和TGF-β2及HIF-1α对幼年牦牛皮肤毛囊生长发育的影响，选取10头健康幼年牦牛，采集其颈部、背部、胸部、腹部、小腿部、腋下及阴囊皮肤组织，制作石蜡切片后，采用HE和Sacpic染色，对各部位皮肤组织中毛囊进行观察和计数，并筛选出多毛及少毛部位。利用qRT-PCR、Western blot及免疫组织化学法对TGF-β2和HIF-1α在幼年牦牛多毛皮肤和少毛皮肤进行定位与定量的初步研究。结果表明，幼年牦牛皮肤的毛囊分布于真皮层，常与汗腺及皮脂腺伴行，毛髓质及内根鞘结构不完整。Sacpic染色可见毛囊内根鞘为红色，外根鞘为苍绿色，结缔组织鞘为蓝绿色。腹部皮肤的毛囊数量最多[（2 085±15）个·cm^-2]，阴囊数量最少[（158±15）个·cm^-2]。免疫组织化学结果显示，TGF-β2及HIF-1α主要表达在表皮层、毛囊外根鞘、皮脂腺及汗腺。TGF-β2在阴囊和腋下的mRNA转录水平和蛋白质表达水平均显著高于腹部和背部。HIF-1α在腹部的mRNA转录水平和蛋白质表达水平均显著高于其他三个部位。幼年牦牛的毛囊处于退行期，在腹部数量最多，背部次之，阴囊最少；TGF-β2和HIF-1α在不同部位皮肤中的表达水平存在显著差异，为进一步研究TGF-β2和HIF-1α对牦牛皮肤毛囊生长发育的影响提供理论依据。     

成纤维细胞生长因子21及其受体1、2在小鼠毛囊形成期的定位和表达

本试验旨在通过研究成纤维细胞生长因子21(fibroblast growth factor 21,FGF21)及其受体FGFR1、FGFR2在胚胎小鼠毛囊形成阶段中的定位与表达,从而探究其在小鼠毛囊形成中的作用。试验采用免疫组织化学、实时荧光定量PCR及Western blotting方法研究FGF21、FGFR1、FGFR2蛋白和mRNA在胚胎期13～18d(E13～E18)小鼠皮肤中的表达。结果显示:(1)FGF21蛋白在E13主要表达于表层细胞、基底层细胞及结缔组织中,E14表达于表层细胞和基板,E15在毛芽中有微量表达,E16、E17表达于毛钉中,E18主要表达于未成熟毛囊细胞;FGFR1和FGFR2蛋白在E13～E18于表层细胞、基底层、基板、毛芽、毛钉、未成熟毛囊和结缔组织中均有表达。(2)实时荧光定量PCR及Western blotting结果显示:FGF21mRNA和蛋白在E14相对表达量最高;FGFR1mRNA及蛋白在E16～E17相对表达量较高;FGFR2mRNA及蛋白的相对表达量在E13至E18均呈上升趋势,在E18相对表达量最高。结果提示:在小鼠胚胎期毛囊形成和发育过程中,FGF21可能在诱导毛囊启动过程中发挥一定的作用;FGFR1可能促进毛钉形成;FGFR2可能在毛囊形成和成熟中发挥重要作用。     

Influence of inflammation and coat type on oestrogen receptor immunohistochemistry

The purpose of this study was to evaluate oestrogen receptor alpha staining in a variety of breeds and skin conditions. The influence of inflammation and coat type on the presence and intensity of oestrogen receptor alpha staining was evaluated. Approximately 1700 haematoxylin and eosin (H&E)-stained slides of skin biopsies were screened for presence of small hair bulbs. Slides from 94 cases were submitted for oestrogen receptor alpha immunohistochemistry. H&E-stained skin biopsy tissues were examined for inflammation and hair follicle stages. Oestrogen receptor alpha staining characteristics of telogen follicles, flame follicles, large anagen bulbs, small hair bulbs and early anagen hairs (capped bulbs) were recorded. To assess the influence of inflammation and coat type on oestrogen receptor staining of hair follicle types, chi-square tests, Fisher's exact tests and logistic regression models were performed. Slides were classified as inflammatory (65) and noninflammatory (29). There were no statistically significant differences in oestrogen receptor staining when comparing inflammatory to noninflammatory skin biopsies or skin biopsies from dogs with different coat types. A subset of 13 noninflammatory biopsies from alopecic skin was identified. There was a statistically significant increase in the number of flame follicles in this subset as compared to all others. Comparison of oestrogen receptor staining of hair follicle types from these biopsies and all other biopsies revealed a statistically significant increase in the number of mature telogen hair follicles stained in this subset. No statistical difference in staining of early follicle stages was noted. Therefore, the oestrogen receptor is unlikely to be the controlling factor for the transition from telogen to anagen in the dog.     

不同水平肌醇对獭兔皮肤中β-连环蛋白和激素敏感脂酶表达的影响

本试验旨在研究饲粮中添加不同水平的肌醇对獭兔皮肤中β-连环蛋白(β-catenin)和激素敏感脂酶(HSL)表达的影响。选取(40±1)日龄的同期断奶獭兔120只(公母各占1/2),随机分成4组,每组30只。在4组獭兔的饲粮中分别添加0、25、50、75 mg/kg的肌醇,试验期为3个月。在试验的第30天(2月龄)、第60天(3月龄)和第90天(4月龄),分别取腹部、背中部、臀部皮肤,采用免疫组织化学和蛋白质印迹(Western blot)法对β-catenin和HSL的蛋白质表达和定位进行检测。结果表明:β-catenin在毛囊中广泛表达,在毛根鞘细胞和毛乳头均有棕黄色阳性反应细胞。HSL在毛根鞘细胞,尤其是在内根鞘细胞内,呈现非常明显的棕黑色强阳性表达。饲粮中添加50 mg/kg肌醇可以极显著增加2~4月龄獭兔背中部皮肤毛囊中β-catenin和HSL阳性表达细胞的平均灰度值(P0.01),极显著增加4月龄獭兔背中部、腹部和臀部皮肤毛囊中β-catenin和HSL阳性表达细胞的平均灰度值(P0.01)。结果提示,肌醇能够通过上调4月龄獭兔毛囊中β-catenin和HSL的表达来促进毛囊的发育,在本试验中,饲粮肌醇水平达到50 mg/kg时效果最佳。     

Histology of the hair cycle in male beagle dogs

A detailed histologic study of the hair follicles of male Beagle dogs was made to determine changes in microscopic structure associated with the three stages in the hair cycle. Observed changes are that the bulbs of hair follicles in anagen are more closely associated with adipose tissue than they are in telogen. This is due to the deeper penetration of the follicles into the subcutaneous tissue during anagen. The hair germ cells of telogen were presumed to arise from the stratum basale of the matrix cells rather than from the outer root sheath cells. During telogen, the dermal papilla is separated from the club hair, but remains in close proximity to it. There is no connecting stalk as is reported for other animals.     

Clear cell trichoblastomas in two dogs

Two cases of clear cell trichoblastomas were diagnosed in young dogs. The tumour had ribbons of basaloid cells as seen in ribbon trichoblastomas, as well as differentiation to external root sheath of the hair follicle and few cells with sebaceous differentiation. This is the first report of clear cell trichoblastoma in dogs.     

Skin morphology of thyroidectomized rats

We induced hypothyroidism in rats by conducting a thyroidectomy (TD) and investigated subsequent changes in the morphology of the skin, especially that of the epidermis and hair follicles. The 6 rats in the TD group seemed less active than the 3 rats in the control group and had cold, dry paws. All of the rats in the TD group exhibited retarded hair growth 12 weeks after surgery. Histologically, all of the rats in the TD group exhibited epidermal thinning from 12 weeks after surgery. Many hair follicles were in the telogen phase: the bulbs and papillae were involuted and had migrated towards the epidermis. Hair follicle atrophy involving thinning of the outer root sheath and the inner root sheath was often observed. The immunoreactivities of antithyroid hormone receptors alpha and beta in the outer root sheaths of 5 of the TD rats were weaker than those of control rats. Cell proliferation in hair follicles of TD rats was weaker than in follicles of control rats 4 weeks after surgery. It is suggested that decreased expression of TRs and decreased cell proliferation activity in the hair follicles of rats is associated with a lack of thyroid hormone and results in retardation of hair growth.     

绵羊FGF10基因克隆、序列分析及其在毛囊发育中的表达分析

本试验旨在获得中国美利奴羊成纤维细胞生长因子10（fibroblast growth factor 10，FGF10）基因的编码区（CDS）全长序列并进行生物信息学分析，随后对FGF10基因在中国美利奴羊毛囊发育过程中的表达特征进行分析，明确其在中国美利奴羊毛囊发育过程中的表达模式，为进一步研究FGF10 mRNA表达水平与中国美利奴羊毛囊生长发育的表达调控机制奠定理论基础。采用PCR扩增获得中国美利奴羊FGF10基因CDS，并克隆到zero PCR@^TM-Blunt进行测序验证；利用实时荧光定量PCR技术检测FGF10在中国美利奴羊毛囊发育过程中的表达差异。结果表明，绵羊FGF10基因CDS长度为696 bp（序列上传GenBank，获得登录号：MT872422），编码231个氨基酸，与牛和山羊的氨基酸序列同源性达100%，存在1个信号肽和1个跨膜结构域，其为分泌通路信号蛋白；实时荧光定量PCR分析表明，FGF10基因在中国美利奴羊毛囊发育过程中均表达，在毛囊发育第85天表达最高，显著高于其他毛囊发育时期（P<0.05）。本研究获得中国美利奴羊FGF10基因完整的编码区序列和毛囊发育过程中的表达特征，生物信息学分析发现，FGF10基因编码区序列具有物种间的保守性，同时FGF10在绵羊毛囊不同发育阶段的皮肤组织中表达，由此表明，FGF10基因可能在绵羊毛囊的生长发育过程中发挥重要的生物学作用。     

A study of normal hair growth in the dog

The rate of hair growth in the forehead, shoulder and flank regions of four normal-coated, cross-bred, male dogs maintained under the same controlled conditions of photoperiod, ambient temperature and diet, was measured during two six-week periods, one during summer and the other in winter. It was found that the rate of hair growth varied between individual dogs and also between the different regions in the same dog. Growth rate was most rapid in the shoulder region, followed by the flank and then the forehead regions, and was slightly more rapid in the shoulder and flank regions of three of the dogs during summer than in winter. Possible explanations for these variations in hair growth rate are discussed.     

Oestrogen receptor evaluation in Pomeranian dogs with hair cycle arrest (alopecia X) on melatonin supplementation

The role of oestrogen receptors in dogs with hair cycle arrest (alopecia X) was investigated by immunohistochemistry. The purpose of this study was to determine if hair regrowth in dogs with hair cycle arrest treated with melatonin was associated with a decrease in follicular oestrogen receptors. Fifteen Pomeranians (excluding intact females) with hair cycle arrest were enrolled. Two biopsies were obtained from alopecic areas of the trunk before and after 3 months on melatonin. Haematoxylin and eosin-stained tissues were examined and oestrogen receptor-alpha was demonstrated immunohistochemically. Common histopathological findings included hyperkeratosis, follicular keratosis, excessive tricholemmal keratinization (flame follicles), thin epidermis, few small anagen bulbs, epidermal pigmentation and melanin aggregates within follicular keratin. Melanin aggregates within basal cells and hair were an occasional finding. After 3 months, 40% (six) dogs had mild to moderate hair regrowth. Biopsies from six dogs showed histological evidence of an increase in anagen hairs and eight dogs had a decrease in epidermal pigmentation. Moderate to marked staining intensity of oestrogen receptor-alpha was noted in all sebaceous gland basal cells, all small hair bulbs and follicular epithelium of telogen hairs. There was no oestrogen receptor-alpha staining of nuclei within the epidermis, apocrine glands or dermal fibroblasts. Large anagen hair bulbs had minimal to no oestrogen receptor staining. Hair regrowth was not associated with a change in oestrogen receptor-alpha staining.     

The effects of thyroxine on hair growth in the dog

The effects of orally and topically administered thyroxine on hair growth were studied in normal, intact, cross-bred male dogs maintained under the same controlled conditions of temperature and photoperiod. It was found that, irrespective of the route of administration, thyroxine produced an increase in both the rate of hair growth and in the number of hair follicles entering the anagen phase of the hair cycle. With orally administered thyroxine, the changes in hair growth were most marked in the flank region.