Immunohistochemical localization of human protein 1 in the female prostate (Skene's gland) and the male prostate

Mouse monoclonal anti-urine protein 1 antibody and the biotin-streptavid in-peroxidase technique were used for the immunohistochemical demonstration of human protein 1 in prostatic tissue of both sexes. In the female prostate (Skene's gland), like the male prostate, high expression of human protein 1 was observed on the luminal surface and in the apical cytoplasm of secretory cells of prostatic glands, as well as on the luminal surface of the epithelium of the large ducts of the female prostate and urethra. Expression was also found in the membranes of secretory and basal cells of the glands, in membranes of the urethral uroepithelium and of the female prostate ducts, in the content of glands and ducts, as well as in vascular endothelium and smooth muscle. Human protein 1 (urine protein 1) expression in the secretory cells of the male and female prostate and its incorporation into the surface of cells lining the lumina of the female urethroprostatic complex is indicative not only of the secretory role of protein 1 but also of its potential protective properties operative in shielding the uroepithelium from the aggressive urinary environment. All genito-urinary tissue, and especially the female prostate, were found to be a potential source of urine protein 1 (human protein 1), refuting the notion held so far that it is exclusively the genito-urinary prostatic tissue of the male that participates in its production. The corresponding immunohistochemical distribution of human protein 1 in the same structures of the male and female prostate provides yet another analogous functional-morphological parameter of prostatic tissue in both sexes and further evidence supporting the non-vestigial concept of the prostate in the female. This revised version was published online in November 2006 with corrections to the Cover Date.     

Immunohistochemical localization of human protein 1 in the female prostate (Skene's gland) and the male prostate

Mouse monoclonal anti-urine protein 1 antibody and the biotin-streptavid in-peroxidase technique were used for the immunohistochemical demonstration of human protein 1 in prostatic tissue of both sexes. In the female prostate (Skene's gland), like the male prostate, high expression of human protein 1 was observed on the luminal surface and in the apical cytoplasm of secretory cells of prostatic glands, as well as on the luminal surface of the epithelium of the large ducts of the female prostate and urethra. Expression was also found in the membranes of secretory and basal cells of the glands, in membranes of the urethral uroepithelium and of the female prostate ducts, in the content of glands and ducts, as well as in vascular endothelium and smooth muscle. Human protein 1 (urine protein 1) expression in the secretory cells of the male and female prostate and its incorporation into the surface of cells lining the lumina of the female urethroprostatic complex is indicative not only of the secretory role of protein 1 but also of its potential protective properties operative in shielding the uroepithelium from the aggressive urinary environment. All genito-urinary tissue, and especially the female prostate, were found to be a potential source of urine protein 1 (human protein 1), refuting the notion held so far that it is exclusively the genito-urinary prostatic tissue of the male that participates in its production. The corresponding immunohistochemical distribution of human protein 1 in the same structures of the male and female prostate provides yet another analogous functional-morphological parameter of prostatic tissue in both sexes and further evidence supporting the non-vestigial concept of the prostate in the female. This revised version was published online in November 2006 with corrections to the Cover Date.     

Immunohistochemical localization of somatostatin in the human hypothalamus

Summary In order to identify clearly the nervous structures containing somatostatin in the human hypothalamus, an immunohistochemical localization of this neurohormone was performed at light-microscopic level. Using a antiserum specific to somatostatin and the unlabeled antibody peroxidase-antiperoxidase technique, we have found somatostatin in neurons with cell bodies in an area in the anterior hypothalamus corresponding to the infundibular nucleus. Somatostatin-containing fibers were also detected in the neurovascular zone of the pituitary stalk, suggesting that somatostatin is released in that region to reach the capillaries in the pituitary portal plexus. A large bundle of somatostatin fibers extending from the anterior part of the paraventricular nucleus up to the posterior portion of the mammillary bodies has also been detected. The role of these fibers still remains to be clarified.     

Immunohistochemical localization of endothelin in human vascular endothelial cells

The cellular localization of endothelin (ET), a novel vasoconstrictor peptide, was studied in human vascular tissues by immunohistochemistry. Distinct and diffuse staining for ET-like immunoreactivity was demonstrated in the cytoplasm of vascular endothelial cells, but not in smooth muscle cells or adventitial fibroblasts. The specificity was confirmed by the negative results following immunoabsorption. These findings suggest that human vascular endothelial cells function as an endocrine and/or paracrine cells for ET secretion.     

Immunohistochemical localization of serine-protease inhibitors in the human placenta

Proteases and their inhibitors play a pivotal role in developmental and differentiative processes. In the present report we investigated the immunohistochemical localization of 1-antitrypsin, 1-antichymotrypsin and inter--trypsin inhibitor in first trimester as well as in term human placentas. For this purpose polyclonal antibodies against these serine-protease inhibitors were used. All inhibitors were expressed in the villous syncytiotrophoblast of first and last trimester placentas. Placental fibrinoid was positively stained for 1-antitrypsin and inter--trypsin inhibitor throughout gestation. 1-Antitrypsin and 1-antichymotrypsin showed a strong immunostaining in the Hofbauer cells (first trimester and full term placentas). Extravillous cytotrophoblast was negative for the three protease inhibitors throughout gestation. The presence of the three inhibitors in the syncytiotrophoblast suggests a role in coagulative, invasive and immunomodulatory processes. Fibrinoid, staining for 1-antitrypsin and inter--trypsin inhibitor, could also have an important immunoprotective function. The presence of protease inhibitors in the Hofbauer cells suggests an involvement of these cells in villous remodelling and differentiative processes.     

Immunohistochemical localization of beta-LPH in the human hypothalamus.

In order to establish the presence of β-LPH and to clearly identify the nervous structures containing β-LPH in the human hypothalamus, an immunohistochemical localization of β-LPH was performed in this tissue. The immunohistochemical technique involved use of a specific antiserum to human β-LPH and the peroxidase-antiperoxidase complex. Immunostained neuronal cell bodies were observed in the arcuate nucleus whereas β-LPH-positive nervous fibers could be detected in a large area extending rostro-caudally from the anterior part of the paraventricular nucleus up to the mammillary bodies. Staining was completely abolished by previous immunoabsorption with β-LPH while β-endorphin and ovine γ-LPH_1–47 only partially prevented immunostaining. Although it cannot be excluded that the precursor 31K molecule, β-LPH_1–58 and/or β-endorphin are detected by the immunostaining, it is likely that β-LPH is at least partly responsible for the positive reaction.     

Immunohistochemical localization of carbonyl reductase in human tissues.

Carbonyl reductase, an NADPH-dependent oxidoreductase of broad specificity, is present in many human tissues. Its precise localization, however, has remained unclear, as well as its physiological and possible pathophysiological significance. The present study reports the immunohistochemical localization of the enzyme in normal human tissues. Immunostaining was detectable in all organs investigated. The highest concentrations were found in the parenchymal cells of the liver, the epithelial cells of the stomach and small intestine, the epidermis, the proximal tubules of the kidney, neuronal and glial cells of the central nervous system, and certain cells of the anterior lobe of the pituitary gland. Consistently pronounced staining was also observed in smooth muscle fibers and the endothelium of blood vessels. The results are in agreement with a housekeeping function of carbonyl reductase in the elimination of reactive carbonyl compounds.     

Immunohistochemical localization of aquaporins in the human inner ear

We report the immunolocalization of aquaporins (AQPs) 1, 4, and 6 in the human auditory and vestibular endorgans. A rapid protocol was applied to audiovestibular endorgans microdissected from postmortem human temporal bones from six subjects (ages ranging from 75 to 97 years) with no history of audiovestibular disease. Temporal bones were fixed in formalin, and the endorgans were immediately microdissected. Cryostat sections were obtained from audiovestibular endorgans and were subjected to double-immunohistochemical staining with antibodies against AQPs and several cellular markers. In the human cochlea, AQP1 immunoreactivity was localized to the fibrocytes of the spiral ligament and the sub-basilar tympanic cells; AQP4 immunoreactivity was localized to the outer sulcus cells, Hensen’s cells, and Claudius’ cells; AQP6 immunoreactivity was localized to the apical portion of interdental cells in the spiral limbus. In the vestibular endorgans (macula utriculi and cristae), AQP1 was localized to fibrocytes and blood vessels of the underlying stroma and trabecular perilymphatic tissue; AQP4 immunoreactivity was localized to the basal pole of vestibular supporting cells; AQP6 was localized to the apical portion of vestibular supporting cells. Cochlear and vestibular hair cells and nerve fibers were not immunoreactive for any AQP. Supporting cells were identified with antibodies against glial fibrilar acidic protein. Nerve fibers and terminals were identified with antibodies against neurofilaments and Na⁺K⁺ATPase. The high degree of conservation of AQP expression in the human inner ear suggests that AQPs play a critical role in inner ear water homeostasis. The National Institutes of Health (grants AG09693-10, DC005224, 00140-02, and DC05187-01) supported this work.     

Immunohistochemical localization of human carbonic anhydrase isoenzyme C

Summary Methods for immunohistochemical localization of human carbonic anhydrase isoenzyme C (HCA C) with indirect fluorescent antibody and immunoperoxidase techniques are described. Both methods revealed large amounts of this high activity isoenzyme in the mucosae of human stomach and appendix. With the indirect immunofluorescent method the presence of the enzyme in human erythrocyte cytoplasm was also demonstrated. Correlations of present findings with those obtained with the traditional histochemical methods for demonstration of carbonic anhydrase activity are discussed.     

Immunohistochemical localization of Zn-alpha 2-glycoprotein in normal human tissues

The Zn-alpha 2-glycoprotein (Zn-alpha 2-GP) is present at a high concentration in the seminal plasma and at significant levels in other human body fluids. Its precise localization, however, has remained unclear, as well as its physiological and pathological significance. The present study reports the immunohistochemical localization of this protein in normal adult human tissues. Localization of the reactive product to anti-human plasma Zn-alpha 2-GP antibody was demonstrated in the following cells: luminal and basal cells of the prostate gland, luminal epithelial cells of the acini and of some ducts of the mammary glands, luminal cells of the secretory portion of the eccrine and apocrine sweat glands, serous cells of the salivary, tracheal, and bronchial glands, acinar cells of the esophageal glands, exocrine acinar cells of the pancreas, hepatocytes of the liver, and epithelial cells of the proximal and distal tubules in the kidney. The present results suggest that Zn-alpha 2-GP exerts some unknown but fairly widespread exocrine function and may be produced in the various epithelial cells tested. Hepatocytes are also suggested to be a source of the protein in the blood plasma.     

Immunohistochemical localization of truncated midkine in developing human bile ducts

Midkine (MK) is a heparin-binding growth factor whose gene has been identified in embryonal carcinoma cells in early stages of retinoic acid-induced differentiation. In the present study, we investigated the developmental localization of truncated MK protein in human bile ducts. Thirty specimens of the livers from 25 fetuses (from 9 to 40 gestational weeks) and from five neonates less than 4 weeks old were examined. Immunohistochemical analysis was performed using a mouse IgG2b monoclonal antibody against recombinant-truncated MK. Truncated MK was expressed moderately in the fetal liver from 9 to 15 gestational weeks. The immunoreactivities were found in the primitive hepatocytes, ductal plates, migrating biliary cells and immature bile ducts. The reaction products were localized in the cytoplasm heterogeneously. The intensity of immunostaining was weak from 15 gestational weeks to 26 gestational weeks. After 27 gestational weeks, truncated MK was not detected in the fetal livers. It was suggested that primitive hepatocytes, ductal plates and immature bile ducts produced truncated MK transiently during human bile ducts development.     

Immunohistochemical localization of heparan sulfate proteoglycan in human gastrointestinal tract

Heparan sulfate proteoglycans (HS-PGs) are associated with important cell functions, for example, cell motility, cell adhesion, and oncogenesis. We examined the localization of HS-PGs in normal and carcinoma tissues of the gastrointestinal tract to help elucidate their roles in these organs. Fresh surgical materials from 134 patients with carcinoma of the stomach or large intestine and 26 patients with various diseases of the small intestine were immunostained after fixation with 10E4 (an antibody against the HS of HS-PG) as a primary antibody. Immunoelectron microscopy (immunogold method) was also performed. The basolateral surfaces of normal tissues of the large and small intestines were strongly stained with antibody confirmed by electron microscopy. In the stomach, lesions with intestinal metaplasia showed the same staining as the intestines, although normal gastric tissue showed staining only in some parts of the basal layer of fundic and pyloric glands. Carcinoma tissues in all cases examined showed staining with antibody. Better results were obtained after fixation in acetic alcohol or zinc-containing solutions than in ordinary formalin. These characteristic localizations of HS-PG in intestines and stomachs suggest that this kind of HS-PG staining could be a hallmark characteristic of the intestine.     

Immunohistochemical localization of kallikrein in human pancreas and salivary glands

The localization of kallikrein in human exocrine organs was studied with a direct immunofluorescence method. In the submandibular and parotid salivary glands, kallikrein was found apically in the striated duct cells whereas it was absent from the main excretory ducts or present only as a weak luminal rim. Kallikrein was not found in the acinar cells or in cells of the intercalated ducts. In the pancreas, kallikrein-specific fluorescence was seen in the granular portion of the acinar cells, whereas the islets of Langerhans and ductal cells were unstained.     

Immunohistochemical localization of NAD glycohydrolase in human and rabbit tissues

NAD glycohydrolase (NADase) is present in many organisms from bacteria to mammals. In any given organism, this enzyme is ubiquitous in many tissues. However, its precise localization and its physiological significance have not been defined. We have determined the distribution of NADase in normal human and rabbit tissues by immunoblotting and immunohistochemistry, using a polyclonal antibody raised in goats. Immunoblot analyses revealed that NADase was highly expressed in the heart, lung, stomach, and liver tissues of the rabbit. From immunohistochemical studies of NADase, high concentrations in both human and rabbit tissues were found in hepatocytes and sinusoidal lining cells, sinus histiocytes of the lymph node, spleen and thymus, glomerular capillary endothelial cells of the kidney, cardiac muscle, endothelium of blood vessles, and erythrocytes.     

Immunohistochemical localization of complement regulatory proteins in the human retina

Age-related macular degeneration (AMD) is a complex disease. Genetic studies have found strong associations between AMD and variants of several complement pathway-associated genes. The regulation of the complement cascade seems to be critical in the pathogenesis of AMD. In 45 human donor eyes immunohistochemistry was performed using antibodies directed against major regulators of the complement system: complement factor H (CFH), decay accelerating factor (DAF/CD55), complement receptor 1 (CR1/CD35), and membrane cofactor protein (MCP/CD46). All eyes were classified in AMD and controls. 11 eyes were graded as early AMD. 34 eyes were controls. In all eyes staining was found in intercapillary pillars of choroid adjacent to Bruch's membrane for CFH, at the basal surface of RPE cells for MCP, and at the apical side of the retinal pigment epithelium for CR1. DAF immunoreactivity was increased along the inner segments of rod and cone photoreceptor cells at the level of the external limiting membrane Labeling of soft drusen was found for CFH and CR1. In addition, DAF and CR1 showed staining of ganglion cells in all eyes. CFH and particularly MCP showed decreased or absent staining in eyes with early AMD adjacent to Bruch's membrane. The overlapping expression of regulators at the level of Bruch's membrane and the retinal pigment epithelium shows the importance of this site for control of the complement system. Decreased and therefore unbalanced expression of regulators, as shown in this study for CFH and MCP, may ultimately lead to AMD.     

Immunohistochemical localization of smooth muscle myosin in normal human tissues

Immunohistochemical localization of smooth muscle myosin, an immunologically distinct contractile protein, was achieved using rabbit anti-human uterine smooth muscle myosin antibodies. In immunodiffusion studies and in cryostat sections, these antibodies were highly specific and reacted with smooth muscle myosin but not with platelet, skeletal muscle, or cardiac muscle myosin. To evaluate comprehensively the structural profile of smooth muscle elements in normal human tissues, an indirect immunoperoxidase technique (peroxidase-antiperoxidase) was applied to a wide variety of specimens. Parallel studies comparing cryostat sections with fixed (10% formalin, B5, Bouin's, or Zenker's solution) paraffin-embedded tissues revealed optimal immunoreactivity, sensitivity, and specificity of staining for smooth muscle myosin using frozen tissues. Strong immunoreactivity was present in muscular tissues such as blood vessels and the muscularis of gastrointestinal and genitourinary tracts. Distinct delineation of smooth muscle elements, including individual smooth muscle cells, and their specific patterns of alignment and organization, were observed, e.g., cells comprising the muscularis mucosae and extending into the lamina propria of the gastrointestinal tract, and myoepithelial cells of skin, exocrine glands, and breast. This method provides excellent morphologic preservation and readily permits unambiguous identification of individual cells containing smooth muscle myosin.     

Immunohistochemical localization of 2-Cys peroxiredoxins in human ciliary body

2-Cys peroxiredoxins (PRDX) are novel antioxidant enzymes that eliminate the hydrogen peroxide in cells to protect the cellular components from reactive oxygen species. To evaluate whether 2-Cys PRDX family plays a role in human ciliary body, the expression of PRDX I, II and III on normal human ciliary body was investigated. Three normal human ciliary body tissues obtained from three donor eyeballs were examined by an immunohistochemistry using light microscopy and fluorescent microscopy with antibodies directed against the PRDX I, II and III. In the normal human ciliary body, PRDX I, II and III were immunolocalized to the non-pigmented epithelial cells and ciliary muscle fibers. It suggests that 2-Cys PRDXs may have physiological functions to protect cells in human ciliary body.