Histochemistry of complex carbohydrates in the skin of the pig snout, with special reference to eccrine glands

The histochemistry of carbohydrates has been studied in the skin of the pig snout with selected methods of light microscopy including peroxidase-labelled lectin-diaminobenzidine (PO-LT-DAB) procedures. In the snout skin the dark secretory cells and the luminal secretion of the eccrine glands contained considerable amounts of neutral glycoproteins, but only a very small amount of acidic ones. It was possible to demonstrate glycogen in the secretory cells of these glands. The other skin structures of the snout also showed positive reactions for complex carbohydrates. Most remarkable were stronger reactions of intercellular substances among the spinosum cells, particularly following the PO-LT-DAB procedures, which demonstrated such saccharide residues as beta-D-galactose and N-acetyl-D-glucosamine.     

Histochemical properties of sialic acids and antimicrobial substances in canine anal glands

The functional properties of sialic acids appear to be manifold. Additionally, antimicrobial substances serve as a non-specific defense against microorganisms. In this study, therefore, the localization of sialic acids and antimicrobial substances in the anal glands of dog was studied by sialoglycoconjugate histochemistry and immunohistochemistry. The secretory epithelium, luminal secretions and excretory ducts exhibited high levels of sialoglycoconjugates that terminated in Siaα2-6Gal/GalNAc or Siaα2-3Gal1-4GlcNAc. Additionally, O-acetylated sialic acids were detectable in these glandular structures. Antimicrobial substances, such as lysozyme, immunoglobulin A (IgA), lactoferrin and the peptide group of β-defensins, were also demonstrated as products of the anal glands. The results obtained are discussed with regard to the functional significance of the anal glands. These secretory products may create a defensive barrier against microbial invasion at the anal mucosa.     

Ultracytochemistry of glycoproteins in the eccrine nasolabial glands of the North American raccoon (Procyon lotor)

The distribution and quality of glycoproteins was studied by means of electron microscopic cytochemical methods, particularly lectin cytochemistry, in the secretory cells of the eccrine nasolabial glands of the North American raccoon (Procyon lotor). In the dark and clear glandular cells, complex glycoconjugates were demonstrable, predominantly, in secretory granules, the cisternae of the Golgi apparatus, the surface coat of the plasma membrane, and as glycogen particles. Secretory granules found in the dark cells contained a variety of saccharide residues, such as α-d-mannose, β-d-galactose, β-N-acetyl-d-glucosamine and sialic acid. Several sugars were also detectable in the surface coat of the plasma membrane and the Golgi apparatus.The results obtained may be helpful to understand the specific functions of the glandular secretions of the raccoon nasolabial glands. These could be, particularly, binding of water on the snout surface and protection against microbial hazards, to maintain the structural and functional integrity of the relatively thin snout epidermis in carnivores.     

Time course of differentiation of different cell types in 3D-reconstructed eccrine sweat glands

Epidermal basal cells invaginate into the dermis to form sweat ducts, which then grow downwards further to form secretory coils during the ontogenesis of eccrine sweat glands, but the time course of differentiation of different cell types in 3D-reconstructed eccrine sweat glands remain unclear. In this study, secretory cell-specific marker K7, clear secretory cell-specific marker CA II, dark secretory cell-specific marker GCDFP-15, myoepithelial cell-specific marker α-SMA, inner duct cell-specific marker S100P and outer duct cell-specific marker S100A2 were detected by immunofluorescence staining. The results showed that S100P and S100A2 were first detected at 2 weeks post implantation, K7 and α-SMA at 3 weeks, and GCDFP-15 and CA II at 4 weeks. The differentiation of ducts preceded secretory coils in 3D-reconstructed eccrine sweat glands. After 8 weeks post implantation, the distribution of these markers in 3D-reconstructed eccrine sweat glands was similar to that in native ones, and the percentage of the 3D-reconstructed glands expressing these markers maintained steady. We conclude that although the 3D-reconstructed and native eccrine sweat glands originated from different cells, the differentiation of different cell types in 3D-reconstructed eccrine sweat glands parallels the sequence observed during embryonic development.     

Histochemical study on the eccrine glands in the foot pad of the cat

Enzyme and carbohydrate histochemical methods were used to study the secretory activities and secretion properties of the eccrine tubular glands in the foot pad of the cat. The activity spectra of the different oxidative and hydrolytic enzymes investigated indicate high metabolic rates within the secretory epithelium. Additionally the enzyme reactions emphasize a double innervation of the glands by cholinergic and adrenergic nerve fibres. The carbohydrate histochemical differentiation reveals mostly neutral and very few acidic glycoproteins in the secretory cells and the secretion, respectively. Clear (basal) cells contain distinct amounts of glycogen, and dark (superficial) cells show neutral glycoproteins, which reveal after PO-lectin-DAB procedures the following saccharide residues: alpha-D-mannose, alpha-D-galactose, N-acetyl-alpha-D-glucosamine, alpha-L-fucose, beta-D-galactose-D-N-galactosamine, beta-D-galactose, and sialic acid. The results obtained confirm the view that the normal biological functions of the eccrine glands of the feline foot pad are to improve the frictional capacities of the paw and to leave typical scent marks.     

Ultrastructural immunocytochemical studies of blood group substances in human eccrine glands

We investigated the ultrastructure of blood group antigens A, B, and H in human eccrine glands by means of the immunogold labeling technique. Blood group antigens A, B, and H were found in the Golgi apparatus, secretory granules, and over the apical and basolateral cell membranes of dark cells of eccrine glands depending on the blood group phenotype of the donors. Both A and B antigens were found in the dark cells of AB donors. The labeling pattern of the Golgi stacks seemed to have a polarity whereby the anti-blood group A antibody labeled all the stacks, whereas anti-blood groups B and H bound to the trans side of the Golgi complex. These observations suggest that the blood group substances are secreted into the lumen after being processed through the Golgi apparatus and the immature and mature granules in the dark cells of human eccrine glands.     

Morphological, histochemical and immunohistochemical characterization of secretory production of the ciliary glands in the porcine eyelid

In addition to performing general histology and cytology of the ciliary glands of the miniature pig, we studied the localization of glycoconjugates and beta-defensins in these glands with the use of carbohydrate histochemical and immunohistochemical methods. The secretory cells of the glands were equipped with non-homogeneous secretory granules, a well-developed Golgi apparatus and rough endoplasmic reticulum. The secretory epithelium and luminal secretion of the glands contained large amounts of neutral and acidic glycoconjugates with various saccharide residues (alpha-L-Fuc, beta-D-Gal, alpha-D-GalNAc and sialic acid). The sebaceous glands and tarsal glands also exhibited positive reactions to most of the histochemical methods. Additionally, the antimicrobial peptide group of beta-defensins was demonstrated to be products of the ciliary glands, as well as the sebaceous glands and tarsal glands. The results obtained are discussed with regard to the specific function of the ciliary glandular secretions. These secretory products may be related to the moistening and general protection of the skin surface of the eyelid and ocular surface.     

Histochemistry of glycoconjugates in the goat nasolabial skin with special reference to eccrine glands

The histochemistry of glycoconjugates in the nasolabial skin of the goat has been studied by means of a series of selected methods of light microscopy. The epidermis of the nasolabial skin was found to contain neutral and acid glycoconjugates with different saccharide residues. The secretory epithelial cells and secretory substances of the sebaceous glands contained primarily neutral glycoconjugates, whereas those of the apocrine glands involved largely strongly acidic and neutral glycoconjugates. In the epithelial cells and secretory substances of the nasolabial eccrine glands, glycoconjugates involved were characteristically strongly neutral but weakly acidic in nature. From the present results, the histophysiological significance of glycoconjugates in the particular histologic structures of the nasolabial skin has been discussed with special reference to the functions of this particular skin type in the goat.     

Immunoelectron microscopic localization of epidermal growth factor in the eccrine and apocrine sweat glands.

We studied the localization of the epidermal growth factor (EGF) in eccrine and apocrine sweat glands with light microscopic and electron microscopic immunohistochemistry. Anti-human EGF (anti-hEGF) polyclonal antiserum and anti-hEGF monoclonal antibody (MAb) were used for the study. Light microscopic immunohistochemistry with monoclonal and polyclonal antibodies showed that hEGF-like immunoreactivity was strongly positive in the myoepithelial cells and weakly positive in the secretory cells of eccrine sweat glands. In apocrine sweat glands, it was strongly positive in the secretory cells as well as in the myoepithelial cells. Immunoelectron microscopy with polyclonal antibody showed that hEGF-like immunoreactivity was present in secretory granules of apocrine secretory cells. These granules had mitochondrion-like internal structure. No reactivity was observed on the eccrine secretory cells by immunoelectron microscopy. Neither dark cell granules nor mitochondria in eccrine secretory cells were labeled with anti-hEGF antibody. In both eccrine and apocrine sweat glands, hEGF-like immunoreactivity was diffusely present in the cytoplasm of myoepithelial cells. However, nuclei and mitochondria of myoepithelial cells were devoid of immunoreactivity for hEGF. Our observations indicate that apocrine sweat glands may secrete more hEGF in the sweat than eccrine sweat glands.     

Cutaneous eccrine glands of the foot pads of the small Madagascan tenrec (<Emphasis Type="Italic">Echinops telfairi,</Emphasis> Insectivora,Tenrecidae): skin glands in a primitive mammal

In order to find correlations between skin gland morphology and specific ethological features, the cutaneous glands of the foot pads of the primitive mammal the Madagascan tenrec, Echinops telfairi, were studied by histological and various histochemical methods as well as by electron microscopy. In the foot pads specific eccrine skin glands occurred consisting of coiled ducts and tubular secretory portions, the lumina of which were considerably wider than in primate sweat glands. The secretory tubules were composed of branched myoepithelial cells and glandular cells. The latter contained abundant mitochondria, large amounts of glycogen particles and few secretory granules as well as individual heterolysosomes and myelin bodies. The lateral cell membrane was marked by extensive interdigitations. The apical membranes of all glandular cells contained proteoglycans with sulfated and carboxylated groups containing N-acetyl-glucosamine, N-acetyl-galactosamine, galactose and mannose. The expression pattern of cytokeratins of the glandular epithelium was variable and showed similarities to that of the human eccrine glands. Tubulin, vinculin and actin were expressed in the glandular epithelium. The secretory cells showed positive reactions with antibodies against antimicrobial peptides and IgA. A positive reaction was observed with antibodies against the androgen receptor. The PCNA and TUNEL reactions indicated that the tubular skin glands of Echinops are made up of a slowly renewing tissue. We conclude that the glands fulfill several functions: production of a fluid-rich secretory product, which may prevent slipping of the foot pads on the substrate during running or climbing, secretion of antimicrobial peptides and proteins, and playing a role in thermoregulation.We thank the Fendt Foundation for financial support     

Lectin binding pattern and proteoglycan distribution in human eccrine sweat glands

The distribution pattern of glycoconjugates in human eccrine sweat glands has been studied by the binding of newly discovered lectins and by antibodies against a chondroitin sulphate proteoglycan and chondroitin sulphate glycosaminoglycans. Mannose-specific lectins labelled large intracellular granules, part of which could be extended cisternae of the endoplasmic reticulum or Golgi apparatus. In contrast, lectins specific for terminal mannose/glucose residues predominantly labelled basement membranes and the glycocalyx. Lectins recognizing terminal N-acetylgalactosamine groups left most parts of the glands unstained, but stained some dark cells intensely. These last cells were also intensively labelled by N-acetylglucosamine-specific and by fucose-specific lectins. Sialic acid residues were preferentially located in luminal borders of secretory coils. No terminal galactose residues were detected. All antibodies against chondroitin glycoconjugates stained large granules similar to those revealed by the mannose-specific lectins in the secretory cells. The basement membrane is only stained by the proteoglycan antibody and the chondroitin-6-sulphate antibody.Thus, a complex composition of glycoconjugates exists not only in matrix elements but also in the cells of eccrine glands of the human skin. A possible secretion of glycoconjugates is discussed.     

Notes on Technic: Differential staining of Fungus and host cells Using a modifciation of Pianeze IIIB

Intercellular secretory capillaries in parotid glands, eccrine sweat glands and intracellular secretory capillaries in parietal cells of gastric glands were demonstrated histo-chemically by the use of the Wachstein-Meisel adenosinetriphosphatase (ATPase) technique in the rabbit, rat and guinea pig. However, with the Wachstein-Meisel 5-nucleotidase technique, secretory capillaries were not stained. For parotid glands, optimal incubation in ATPase substrate mixture was: in rabbit, 15 min; in rat, 2.5 hr; and in guinea pig, 2 hr. For eccrine sweat glands, optimal incubation was 15 min in rabbit, 30 min in rat and 15 min in guinea pig. For parietal cells of gastric glands, optimal incubation was 3 hr for all three species. Secretory capillaries were best demonstrated in the parotid by using rabbit tissue; in eccrine sweat glands, with rat tissue, and in parietal cells, guinea pig tissue. Since ATPase activity in cell membranes of secretory cells may play a part in the mechanism of transport of secretory products from their place of formation in the acini to the excretory ducts, the Wachstein-Meisel ATPase technique can therefore be used successfully for staining secretory capillaries in many of the exocrine glands of laboratory mammals.     

Immunohistochemical localization of activated EGF receptor in human eccrine and apocrine sweat glands.

Epidermal growth factor (EGF) is secreted into sweat from secretory cells of human sweat glands. The function of EGF in sweat is poorly understood. The biological function of EGF is exerted by the binding of EGF to the receptor (EGFR) and its activation. Therefore, we immunohistochemically localized the activated form of EGFR in human eccrine and apocrine sweat glands to assess the functional importance of the EGF-EGFR system in human sweat glands. Frozen sections of human skin were stained with a monoclonal antibody (MAb) specific for tyrosine-phosphorylated (activated) EGFR and with an MAb that stains both activated and non-activated EGFR. In the secretory portion of eccrine sweat glands, nuclei of the secretory cells were stained with the anti-activated EGFR MAb. In coiled and straight portions of eccrine sweat ducts, nuclei of luminal and peripheral cells were stained with the antibody specific for activated EGFR. Luminal cell membranes and luminal cytoplasm of inner ductal cells possessed non-activated EGFR. In the secretory portion of apocrine sweat glands, activated EGFRs were present in cytoplasm and nuclei of secretory cells. These data suggest that EGF, already known to be present in the cytoplasm of secretory cells in eccrine and apocrine sweat glands, activates EGFR in the nuclei of secretory cells themselves in an intracrine manner. Because ductal cells do not express EGF, EGF in the sweat secreted from the secretory cells should activate EGFR in the ductal cells in a paracrine manner. (J Histochem Cytochem 49:597-601, 2001)     

The histochemistry of complex carbohydrates in the scrotum of the boar

Summary In the scrotal skin of the boar, the histochemistry of complex carbohydrates has been studied by means of a series of selected methods of light microscopy. The epidermis of the scrotal skin was found to contain neutral and acidic complex carbohydrates with different saccharide residues. The secretory epithelial cells and secretory substances of the saccular apocrine sweat glands contained sulfated, other acidic and neutral complex carbohydrates, whereas the secretory epithelial cells and secretory substances of the tubular apocrine sweat glands involved largely neutral complex carbohydrates. The two types of complex carbohydrates from the both glands were shown to contain commonly substantial amounts of various saccharide residues but were devoid of notable amounts of sialic acid residues. In addition, complex carbohydrates in the smooth muscle cells were reacted for relatively small amounts of saccharide residues. From the present results, the histophysiological significanses of complex carbohydrates in the particular histologic structures of the scrotum have been discussed with special reference to the functions of the skin in the boar.A major part of this work has been presented at the 6th International Histochemistry and Cytochemistry Congress, Brighton, United Kingdom, in 1980     

Staining Secretory Capillaries of Exocrine Glands with Techniques for Specific Phosphatases

Intercellular secretory capillaries in parotid glands, eccrine sweat glands and intracellular secretory capillaries in parietal cells of gastric glands were demonstrated histo-chemically by the use of the Wachstein-Meisel adenosinetriphosphatase (ATPase) technique in the rabbit, rat and guinea pig. However, with the Wachstein-Meisel 5-nucleotidase technique, secretory capillaries were not stained. For parotid glands, optimal incubation in ATPase substrate mixture was: in rabbit, 15 min; in rat, 2.5 hr; and in guinea pig, 2 hr. For eccrine sweat glands, optimal incubation was 15 min in rabbit, 30 min in rat and 15 min in guinea pig. For parietal cells of gastric glands, optimal incubation was 3 hr for all three species. Secretory capillaries were best demonstrated in the parotid by using rabbit tissue; in eccrine sweat glands, with rat tissue, and in parietal cells, guinea pig tissue. Since ATPase activity in cell membranes of secretory cells may play a part in the mechanism of transport of secretory products from their place of formation in the acini to the excretory ducts, the Wachstein-Meisel ATPase technique can therefore be used successfully for staining secretory capillaries in many of the exocrine glands of laboratory mammals.     

Localization of sex steroid receptors in human skin

Sex steroid hormones are involved in regulation of skin development and functions as well as in some skin pathological events. To determine the sites of action of estrogens, androgens and progestins, studies have been performed during the recent years to accurately localize receptors for each steroid hormone in human skin. Androgen receptors (AR) have been localized in most keratinocytes in epidermis. In the dermis, AR was detected in about 10% of fibroblasts. In sebaceous glands, AR was observed in both basal cells and sebocytes. In hair follicles, AR expression was restricted to dermal papillar cells. In eccrine sweat glands, only few secretory cells were observed to express AR. Estrogen receptor (ER) alpha was poorly expressing, being restricted to sebocytes. In contrast, ERbeta was found to be highly expressed in the epidermis, sebaceous glands (basal cells and sebocytes) and eccrine sweat glands. In the hair follicle, ERbeta is widely expressed with strong nuclear staining in dermal papilla cells, inner sheath cells, matrix cells and outer sheath cells including the buldge region. Progesterone receptors (PR) staining was found in nuclei of some keratinocytes and in nuclei of basal cells and sebocytes in sebaceous glands. PR nuclear staining was also observed in dermal papilla cells of hair follicles and in eccrine sweat glands. This information on the differential localization of sex steroid receptors in human skin should be of great help for future investigation on the specific role of each steroid on skin and its appendages.     

Three-dimensional reconstructed eccrine sweat glands with vascularization and cholinergic and adrenergic innervation

Functional integrity of the regenerated tissues requires not only structural integrity but also vascularization and innervation. We previously demonstrated that the three-dimensional (3D) reconstructed eccrine sweat glands had similar structures as those of the native ones did, but whether the 3D reconstructed glands possessing vascularization and innervation was still unknown. In the study, Matrigel-embedded eccrine sweat gland cells were implanted under the inguinal skin. Ten weeks post-implantation, the vascularization, and innervation in the 10-week reconstructed eccrine sweat glands and native human eccrine sweat glands were detected by immunofluorescence staining. The results showed that the fluorescent signals of general neuronal marker protein gene product 9.5, adrenergic nerve fiber marker tyrosine hydroxylase, and cholinergic nerve fiber markers acetylcholinesterase and vasoactive intestinal peptide embraced the 3D reconstructed glands in circular patterns, as the signals appeared in native eccrine sweat glands. There were many CD31- and von Willebrand factor-positive vessels growing into the plugs. We demonstrated that the 3D reconstructed eccrine sweat glands were nourished by blood vessels, and we for the first time demonstrated that the engineering sweat glands were innervated by both cholinergic and adrenergic fibers. In conclusion, the 3D reconstructed eccrine sweat glands may have functions as the native ones do.     

Aspects of glycoconjugate production and lysozyme- and defensins-expression of the ceruminous glands of the horse (Equus przewalskii f. dom.)

The distribution of complex glycoconjugates and antimicrobial substances in the ceruminous glands of the horse (Equus przewalskii f. dom., type: pony) was studied using carbohydrate histochemical and immunohistochemical methods. The epithelial cells and luminal secretion of these glands exhibited considerable amounts of glycoconjugates with various saccharide residues, such as alpha-D-mannose, alpha-L-fucose, beta-D-galactose, beta-N-acetyl-D-glucosamine and sialic acid, including O-acetylated sialic acid. Several sugars (alpha-D-mannose, alpha-L-fucose, and beta-D-galactose) were also detectable in the secretion of sebaceous glands present. Additionally, lysozyme and the peptide group of beta-defensins are demonstrated as products of the apocrine ceruminous glands and sebaceous glands. The results obtained are discussed with regard to the functional significance of the glandular secretions. It is suggested that the complex carbohydrates, lysozyme and beta-defensins found in the ceruminous gland secretions are involved in the function of cerumen as a general antimicrobial protective agent in the external auditory canal.