Immunolocalization of surfactant protein-D (SP-D) in human fetal, newborn, and adult tissues.

Immunoreactive surfactant protein-D (SP-D) was assessed in human fetal, newborn, and adult tissues. In the fetal lung, SP-D was detected on airway surfaces by 10 weeks' gestation, staining increasing in the distal airways, decreasing in the proximal conducting airways with advancing gestation. In lungs from near-term infants and adults, SP-D was detected in Type II cells, serous cells of tracheobronchial glands, and subsets of cells lining peripheral airways. Immunostaining was decreased or absent in areas of lungs of neonates after injury to Type II cells, infection, or hemorrhage and was decreased in collapsed or unseptated airways from older infants with bronchopulmonary dysplasia. SP-D was also detected in many organs at all ages. SP-D was readily detected in epithelial cells and luminal material in lacrimal glands, salivary glands, pancreas, bile ducts, renal tubules, esophageal muscle and glands, parietal cells of the stomach, crypts of Lieberkuhn, sebaceous and eccrine sweat glands, Von Ebner's glands, endocervical glands, seminal vesicles, adrenal cortex, myocardium, and anterior pituitary gland. SP-D is a widely distributed member of the "collectin" family of polypeptides secreted onto luminal surfaces by epithelial cells lining ducts of many organs, where it likely plays a role in innate host defense.     

Immunocytochemical localization of annexin 5, a calcium-dependent phospholipid-binding protein, in rat endocrine organs

Annexin 5, a unique calcium- and phospholipid-binding protein, has been investigated for its specific distribution in rat endocrine organs by immunocytochemistry with a specific antiserum to recombinant rat annexin 5. Follicular epithelial cells and parafollicular cells of the thyroid gland, adrenocortical cells of the zona fasciculata and zona reticularis, luteal cells, testicular interstitial cells, and Sertoli cells were shown to contain annexin 5. To examine whether the synthesis of annexin 5 would be affected by a change in humoral signal, the distribution of annexin 5 in the anterior pituitary was examined three weeks after ovariectomy. The withdrawal of ovarian hormones induced huge castration cells in the anterior pituitary gland, which contained abundant annexin 5. Annexin 5 was not detected in the pineal gland, the parathyroid gland, the islet of Langerhans, the adrenal medulla, zona glomerulosa cells, and granulosa cells. Since annexin 5 was shown to exist in many of the endocrine tissues examined, to be localized in specific cell types, and to be abundant in castration cells, it is suggested that annexin 5 contributes to secretory cell functions, which may be common to endocrine cells secreting chemically different hormones.     

Expression of Secretogranin III in Chicken Endocrine Cells: Its Relevance to the Secretory Granule Properties of Peptide Prohormone Processing and Bioactive Amine Content

The expression of secretogranin III (SgIII) in chicken endocrine cells has not been investigated. There is limited data available for the immunohistochemical localization of SgIII in the brain, pituitary, and pancreatic islets of humans and rodents. In the present study, we used immunoblotting to reveal the similarities between the expression patterns of SgIII in the common endocrine glands of chickens and rats. The protein–protein interactions between SgIII and chromogranin A (CgA) mediate the sorting of CgA/prohormone core aggregates to the secretory granule membrane. We examined these interactions using co-immunoprecipitation in chicken endocrine tissues. Using immunohistochemistry, we also examined the expression of SgIII in a wide range of chicken endocrine glands and gastrointestinal endocrine cells (GECs). SgIII was expressed in the pituitary, pineal, adrenal (medullary parts), parathyroid, and ultimobranchial glands, but not in the thyroid gland. It was also expressed in GECs of the stomach (proventriculus and gizzard), small and large intestines, and pancreatic islet cells. These SgIII-expressing cells co-expressed serotonin, somatostatin, gastric inhibitory polypeptide, glucagon-like peptide-1, glucagon, or insulin. These results suggest that SgIII is expressed in the endocrine cells that secrete peptide hormones, which mature via the intragranular enzymatic processing of prohormones and physiologically active amines in chickens.     

Distribution of adenosine deaminase complexing protein (ADCP) in human tissues

The normal distribution of adenosine deaminase complexing protein (ADCP) in the human body was investigated quantitatively by ADCP-specific radioimmunoassay (RIA) and qualitatively by immunohistochemistry. In these studies we used a specific rabbit anti-human ADCP antiserum. In all 19 investigated tissues, except erythrocytes, ADCP was found by RIA in the soluble and membrane fractions. From all tissues the membrane fractions contained more ADCP (expressed per mg protein) than the soluble fractions. High membrane ADCP concentrations were found in skin, renal cortex, gastrointestinal tract, and prostate. Immunoperoxidase staining confirmed the predominant membrane-associated localization of the protein. In serous sweat glands, convoluted tubules of renal cortex, bile canaliculi, gastrointestinal tract, lung, pancreas, prostate gland, salivary gland, gallbladder, mammary gland, and uterus, ADCP immunoreactivity was found confined to the luminal membranes of the epithelial cells. These data demonstrate that ADCP is present predominantly in exocrine glands and absorptive epithelia. The localization of ADCP at the secretory or absorptive apex of the cells suggests that the function of ADCP is related to the secretory and/or absorptive process.     

In situ localization of mRNA for epidermal growth factor in the submandibular gland of the mouse

Epidermal growth factor (EGF) is a polypeptide originally isolated from the mouse submandibular gland, where it is localized immunocytochemically in cells of the granular convoluted tubules (GCT). cDNAs encoding the precursor of mouse submandibular EGF have been cloned (Scott et al. Science 221:236, 1983; Gray et al. Nature 303:722, 1983). A fragment of one of these clones, pmegf10, containing the EGF coding region, was tritium-labeled by nick-translation and used as a probe for in situ hybridization to EGF mRNA. A specific hybridization signal for EGF mRNA was seen only in mature or developing GCT cells. The intensity of the signal was stronger in glands of intact males than in females or in castrated males. In glands of castrates treated with testosterone, or of intact females treated with triiodothyronine (T3), the signal was comparable to that in intact males. In glands of males treated with T3 the intensity of the signal was stronger than in untreated males. A weak to moderate signal was seen in developing GCT cells of 20-day-old males but not females. Hybridization for 3 days gave a stronger signal than that for 1 day. No signal was seen in either sex at 10 days of age, or in control preparations exposed to labeled DNA of pBR322. The presence of EGF mRNA exclusively in GCT cells provides strong evidence that these cells are the only site of synthesis of EGF in the submandibular gland. In situ hybridization with this cDNA probe will provide a sensitive method to determine possible cellular sites of EGF production outside of the submandibular gland.     

Light and electron microscopic localization of ACTH and pro-opiomelanocortin-derived peptides in human developmental and neoplastic cells

Oncofetal aspects of ACTH and pro-opiomelanocortin (POMC)-derived peptides were studied immunohistochemically at the light and electron microscopic level in human fetal pituitary glands, pituitary adenomas, and small-cell carcinoma of the lung. ACTH, beta-endorphin, and gamma-MSH were localized in the same cells of both fetal and adult pituitary, as well as in the above-mentioned neoplastic tissues. However, alpha-MSH was observed only in the early fetal pituitary, its concentration decreasing with advancing gestational age. The adult pituitary contained only a few alpha-MSH-positive cells. By immunoelectron microscopy, ACTH in the adult pituitary was localized exclusively in the secretory granules. In fetal pituitary at 9 weeks' gestation, ACTH was localized in the perinuclear spaces (PNS), cisternae of rough endoplasmic reticulum (RER), Golgi saccules, and secretory granules. The staining pattern of ACTH in these organelles varied from cell to cell. In fetal pituitaries of greater gestational ages, ACTH was localized in secretory granules. The pituitary adenomas mimicked the staining characteristics of the adult pituitary, i.e., negative or only very occasional alpha-MSH staining and localization of ACTH in the secretory granules. The ectopic ACTH-producing tumors showed a staining pattern similar to that of the early fetal pituitary, i.e., positive staining for alpha-MSH and the presence of ACTH in PNS and cisternae of RER.     

Epidermal growth factor-like material in rat submandibular gland.

By using an antiserum specific for mouse epidermal growth factor (EGF), only the granular convoluted tubule (GCT) cells revealed immunochemical staining in rat submandibular glands. There was no regular sexual difference in the frequency or size of immunoreactive cells. Extracts of gland contained an antigen which showed a complete cross-reactivity with mouse EGF in radioimmunoassays. The relative amounts of EGF, determined by a heterologous radioimmunoassay, were not significantly different in the glands of rats of the two sexes. Administration of testosterone caused an increase, in both sexes, in the number of GCT cells stained for EGF and in the amount of EGF in the gland. There was no significant sexual differeence in these two parameters after androgen treatment.     

Immunohistochemical localization of a thyroid hormone-binding protein (p55) in human tissues

We have localized p55, a thyroid hormone-binding protein found in the endoplasmic reticulum in cultured cells, in samples of normal human and monkey tissues, using a monoclonal antibody with cryostat sections and immunoperoxidase histochemistry. Large amounts of p55 were found in many tissues, generally corresponding to the amount of endoplasmic reticulum contained in each cell type. Intense localization of p55 was found in cells of the anterior and intermediate pituitary lobes, in epithelial cells of thyroid follicles, in the glandular epithelium of mammary gland, in hepatocytes, in Paneth cells and Brunner's glands in duodenum, in acinar cells of pancreas, in adrenal cortical cells, and in scattered interstitial fibroblastic cells in many tissues. These results suggest a potential role for thyroid hormone and p55 in regulating protein synthesis or secretion in multiple organs.     

Immunohistochemical localization of epidermal growth factor in rat and man

Epidermal growth factor (EGF) is a peptide which stimulates cell mitotic activity and differentiation, has a cytoprotective effect on the gastroduodenal mucosa, and inhibits gastric acid secretion. The immunohistochemical localization of EGF in the Brunner's glands and the submandibular glands is well documented. The localization of EGF in other tissues is still unclarified. In the present study, the immunohistochemical localization of EGF in tissues from rat, man and a 20 week human fetus were investigated. In man and rat, immunoreaction was found in the submandibular glands, the serous glands of the nasal cavity, Brunner's glands of the duodenum, the Paneth cells of the small intestine, and the tubular cells of the kidney. In the fetus EGF was found in the kidney and in the intestinal Paneth cells. Antisera raised against rat submandibular EGF did not recognize EGF in human tissues, whereas antisera against human urinary EGF worked in rat as well as man. EGF was found only in cells with an exocrine function.     

Different ghrelin localisation in adult human and rat endocrine pancreas

Ghrelin is an endocrine peptide that has been identified in gastric oxyntic glands and that induces growth hormone secretion in the pituitary gland. This growth hormone secretagogue is expressed in many tissues such as stomach, pituitary gland, thyroid, testis, placenta and pancreas. Initial studies of ghrelin focused on its role as a circulating orexigenic signal. However, ghrelin has also been found to be involved in the modulation of glucose homeostasis. Although a number of studies have reported ghrelin expression in developing pancreas, the location of ghrelin-immunoreactive cells in adult pancreas (epsilon cells) remains controversial. In this study, we have analysed the distribution of pancreatic epsilon cells in adult human and rat islets by immunohistochemistry and in situ hybridisation. In humans, our immunohistochemical analysis has shown that ghrelin is expressed in glucagon-secreting cells, whereas in rats, it is present in insulin-secreting cells. Similar observations have been revealed by in situ hybridisation.     

Distribution of a novel peptide in the anterior pituitary, gastric pyloric gland, and pancreatic islets of rat.

We used Western blot and immunohistochemical methods to investigate the biochemical characteristics and cellular distribution of a novel peptide (peptide 23) that was previously shown to be released from anterior pituitary cells of rat in response to growth hormone-releasing hormone. In the pituitary, peptide 23 isolated from intact cells had an Mr of 31,000, whereas that released into culture medium had an Mr of 16,000. Pancreatic islets contained a 19 KD form of the peptide. Immunohistochemically, peptide 23 in the rat pituitary gland was localized in a subpopulation of somatotropes. In pancreatic islets, the peptide was found by triple immunofluorescence labeling to be present in both insulin- and somatostatin-containing cells. In the gastrointestinal tract, peptide 23 was found only in a subpopulation of endocrine cells in the pyloric glands. This subpopulation of cells was found to be entirely separate from those containing either serotonin or somatostatin, and may represent one of the other known or as yet biochemically uncharacterized cell types in this gland. The results suggest that in response to secretagogues in vitro, an altered form of the peptide is secreted from pituitary cells and that an intracellular form of peptide 23 is expressed in some but not all somatotropes, a large proportion of islet cells, and a distinct population of pyloric cells.     

Immunohistochemical localization of epidermal growth factor in rat and man

Summary Epidermal growth factor (EGF) is a peptide which stimulates cell mitotic activity and differentiation, has a cytoprotective effect on the gastroduodenal mucosa, and inhibits gastric acid secretion.The immunohistochemical localization of EGF in the Brunner's glands and the submandibular glands is well documented. The localization of EGF in other tissues is still unclarified.In the present study, the immunohistochemical localization of EGF in tissues from rat, man and a 20 week human fetus were investigated. In man and rat, immunoreaction was found in the submandibular glands, the serous glands of the nasal cavity, Brunner's glands of the duodenum, the Paneth cells of the small intestine, and the tubular cells of the kidney. In the fetus EGF was found in the kidney and in the intestinal Paneth cells.Antisera raised against rat submandibular EGF did not recognize EGF in human tissues, whereas antisera against human urinary EGF worked in rat as well as man. EGF was found only in cells with an exocrine function.     

Polyamines, molecules necessary for cell division, colocalize with peptide growth factors

The naturally occurring polyamines spermidine and spermine are necessary for cell division and growth. By restaining experiments, using three independent polyamine cytochemical methods, together with peptide immunocytochemistry, we show that substantial amounts of polyamines occur in a number of peptide growth factor-producing cell types. These include submandibular granular convoluted duct cells producing epidermal growth factor (EGF), pancreatic islet cells producing insulin and anterior pituitary cells producing growth hormone (GH). Other cell types in these tissues display only weak or no polyamine reactivity. Also blood platelets, known to contain platelet-derived growth factor (PDGF), are strongly stained for polyamines. Moreover, in EGF cells, insulin cells and blood platelets, polyamines are clearly localized in secretory granules. The possibility that polyamines may be coreleased and act in concert with peptide growth factors is discussed.     

Epidermal growth factor (EGF) expression in human salivary glands. An immunohistochemical study.

Epidermal growth factor (EGF) is a biologically active peptide involved in differentiation, growth, regeneration and repair of human and animal tissues. Quantitative biochemical studies showed in man the highest concentration of EGF in the parotid gland. The aim of the present study was to define EGF immunolocalization in the individual segments of the human major salivary glands (salivon). The material consisted of sections obtained from the surgically removed salivary glands: parotid, submaxillary and sublingual. Immunohistochemical studies were performed by PAP method using monoclonal antibody against human epidermal growth factor. EGF expression was found almost exclusively in the efferent pathways of the salivary glands, mostly in the intercalated ducts and Pflüger salivary tubules. These segments of the salivon are most developed in the parotid gland in which the staining was stronger than in other salivary glands.     

Expression of gastric gland mucous cell-type mucin in normal and neoplastic human tissues.

Gastric gland mucous cells produce class III mucin, which is also found in Brunner's glands and mucous glands along the pancreaticobiliary tract, and in metaplasia and adenocarcinomas differentiating towards gastric mucosa. Recently, we showed that class III mucin possesses GlcNAcalpha1-->4Galbeta-->R, formed by alpha1,4-N-acetylglucosaminyltransferase (alpha4GnT). Examining the tissue-specific expression of mucin epitopes is useful to clarify cell-lineage differentiation and to identify the site of origin of metastatic carcinomas in histological specimens. Formalin-fixed, paraffin-embedded tissue sections from esophagus, stomach, colon, liver, pancreas, lung, kidney, prostate, breast, and salivary gland resected for carcinoma, as well as salivary gland adenoma, colon adenoma, and metastatic adenocarcinoma of lymph nodes from stomach, pancreas, colon, and breast, were immunostained for MUC6, alpha4GnT, and GlcNAcalpha1-->4Galbeta-->R. These were all expressed in normal, metaplastic, and adenocarcinoma tissues of stomach, pancreas, and bile duct, and in pulmonary mucinous bronchioloalveolar carcinomas. Cells expressing alpha4GnT uniformly expressed GlcNAcalpha1-->4Galbeta-->R. Only MUC6 was expressed in normal salivary glands, pancreas, seminal vesicles, renal tubules, and colon adenomas, and in normal tissue and adenocarcinomas of prostate and breast. No tissues showed immunoreactivity for alpha4GnT alone. Immunohistochemistry (IHC) profiles were similar for metastatic carcinomas and primary carcinoma tissues. The IHC profiles for MUC6, alpha4GnT, and GlcNAcalpha1-->4Galbeta-->R may be diagnostically relevant.     

Cell-specific subcellular localization of soluble epoxide hydrolase in human tissues.

Soluble epoxide hydrolase (sEH) is a phase-I xenobiotic metabolizing enzyme having both an N-terminal phosphatase activity and a C-terminal epoxide hydrolase activity. Endogenous hydrolase substrates include arachidonic acid epoxides, which have been involved in regulating blood pressure and inflammation. The subcellular localization of sEH has been controversial. Earlier studies using mouse and rat liver suggested that sEH may be cytosolic and/or peroxisomal. In this study we applied immunofluorescence and confocal microscopy using markers for different subcellular compartments to evaluate sEH colocalization in an array of human tissues. Results showed that sEH is both cytosolic and peroxisomal in human hepatocytes and renal proximal tubules and exclusively cytosolic in other sEH-containing tissues such as pancreatic islet cells, intestinal epithelium, anterior pituitary cells, adrenal gland, endometrium, lymphoid follicles, prostate ductal epithelium, alveolar wall, and blood vessels. sEH was not exclusively peroxisomal in any of the tissues evaluated. Our data suggest that human sEH subcellular localization is tissue dependent, and that sEH may have tissue- or cell-type-specific functionality. To our knowledge, this is the first report showing the subcellular localization of sEH in a wide array of human tissues.     

Age-related decrease of urinary excretion of human epidermal growth factor (hEGF)

Epidermal growth factor (EGF) has been first isolated from the submandibular glands of the male mouse and recently from human urine. Despite its potent mitogenic effect in a variety of tissues, the physiological functions of EGF in human still remain undetermined. In order to study the effect of age on urinary human EGF (hEGF), we have evaluated urinary excretion of hEGF in normal subjects over a wide range of age (20–79 yr.) using homologous radioimmunoassay (RIA) for hEGF. Urinary excretion of hEGF expressed as a function of creatinine significantly decreased with increasing age, age, while females excreted significantly more hEGF than males. These data suggest that urinary excretion of hEGF decreases with age in normal subjects which may be due to reduced synthesis and/or secretion of hEGF.     

Postnatal development of female sheep pineal gland under natural inhibitory photoperiods: an immunocytochemical and physiological (melatonin concentration) study

The purpose of this study was to determine structural and immunocytochemical changes taking place during the day and at night in developing sheep pineal gland under natural non-stimulatory photoperiods (summer solstice). Additionally, the diurnal cycle of plasma melatonin levels was charted and differences between diurnal and nocturnal pineal melatonin concentrations were analyzed. 36 ewes of three different ages were examined: infants (1-6 months old), pubertal and early fertile age (9-24 months old) and adults (36-60 months old). Plasma and pineal gland melatonin levels were higher in pubertal sheep than in infants or adults. Pubertal sheep pineal glands were also heavier, contained a larger number of pinealocytes and interstitial cells and displayed more evident innervation and vascularisation than infants or adults. There was no difference in the number of pinealocytes and interstitial cells between animals killed during daylight or at night. Gland weight, pinealocyte nuclear profile areas and plasma melatonin concentrations were all significantly higher at night than during the day.     

Organogenesis of the pituitary, adrenal, and lung at birth in the wallaby, Macropus rufogriseus

The ultrastructure of the pituitary, the adrenal, and the lung was examined in the newborn wallaby, Macropus rufogriseus. Tissue from six wallaby neonates (less than 8 hr of age), two near-term fetuses (26 days after removal of suckling pouch young [RPY]), and a two-day-old pouch young was examined; and tissue levels of cortisol in the adrenal glands of five neonates and a near-term fetus (26 days) were measured by radioimmunoassay. At birth the adenohypophysis comprised the bulk of the pituitary gland. The pars distalis was well vascularized and many cells contained electron-dense, membrane-bound granules. The adrenal glands lacked specific zones but comprised two distinct populations of cells. The cytoplasm of one cell type contained electron-dense, membrane-bound granules, similar to those observed inside catecholamine-secreting cells of the adrenal medulla; the other cell type possessed large amounts of smooth endoplasmic reticulum and mitochondria with tubulo-vesicular cristae. These features are characteristic of cells which are actively synthesizing steroid hormones. The concentration of cortisol was 0.58 ng/adrenal in the wallaby at birth. The fetal lungs near term were at the glandular stage of development, and epithelial differentiation of type I and type II pneumocytes was imminent although attenuation was not evident. The canalicular neonatal lung did not contain true alveoli, but type II pneumocytes contained osmiophilic lamellar inclusions of surfactant. The fetal pituitary and adrenal are functional at birth and are thus capable of initiating parturition and of influencing lung maturation in the fetus.