Absorption of elastase through the jejunal mucosa of the rat

Summary In order to reveal the absorption process of elastase from the intestine, hog pancreatic elastase was injected into the ligated jejunum lumen of the rat, and the tissues were cytochemically observed at various times after injection. The peroxidase anti-peroxidase (PAP) method using anti-hog-elastase rabbit antibody was used for light microscopy, and the anti-elastase Fab-peroxidase conjugate was used for electron microscopy. The tissues stained by the PAP method exhibited a dense deposition of reaction products on the luminal surface of epithelial cells and a moderate deposition in the blood and lymph capillaries of the intestinal villi. Immunoelectron microscopy revealed that the reaction product was deposited on the surface of the microvilli and in their pocketing; some was found in the pinocytotic vesicles in the terminal-web area and on the inner surface of the enlarged smooth endoplasmic reticulum. Round droplets which gave a positive reaction were found in the widened intercellular cleft and the thick basement membrane lining the blood capillaries and lymphatics. The jejunum retained its normal ultrastructure. The results indicate that the elastase molecules, which were introduced into the rat jejunum lumen, were absorbed without being decomposed through healthy intestinal epithelial cells by pinocytosis and translocated into blood and lymph capillaries.     

Intracellular transport of albumin through the secretory apparatus of rat liver parenchymal cells. An immunocytochemical study

The fine structural localization of albumin in rat liver parenchymal cells was determined by an improved immunocytochemical method and serial sectioning. Albumin in the secretory apparatus of the parenchymal cells was present in segments of the rough endoplasmic reticulum, interrupted with negative segments, in transport vesicles, Golgi saccules, finely anastomosed tubules and vesicles on the trans side of the Golgi complex, and in secretion granules. Horizontally sectioned Golgi saccules contained lipoprotein particles on one side and albumin on the other side. After transport, the vesicles that contained albumin fused with the so-called rigid lamellae on the trans-side of the Golgi complex. Ultrathin serial sections revealed no true structural continuity between the endoplasmic reticulum and the cis-aspect of the Golgi complex. We concluded that secretory proteins are transported from the endoplasmic reticulum to the Golgi complex by transport vesicles that bud from the endoplasmic reticulum and fuse with the Golgi saccules. These vesicles fuse regularly with the Golgi saccules on the cis-side and occasionally with tubular elements on the trans-aspect that may belong to the so-called GERL.     

Peptide-containing innervation of the human intestinal mucosa. An immunocytochemical study on whole-mount preparations

The three-dimensional distribution of the peptide-containing innervation in the human intestinal mucosa was studied by fluorescence immunohistochemistry on whole-mount mucosal preparations. An extensive VIP-immunoreactive nerve supply was demonstrated at all levels, but was markedly increased in density in the distal intestine, where it formed a particularly rich network in close contact with the luminal epithelium. In contrast, substance P-containing nerve fibres formed a looser and evenly distributed innervation at all levels. The muscularis mucosae was richly supplied by VIP- and substance P-containing fibres. Met-enkephalin immunoreactivity was confined to a few scattered nerve bundles running in the muscularis mucosae and around the bottom of epithelial crypts.     

An immunocytochemical study of keratin reactivity during rat odontogenesis.

The cytokeratin distribution in the developing rat enamel organ from day 15 of gestation through to 11 days post partum was examined immunohistochemically using a panel of monoclonal antibodies. A temporo-spatial programme of keratin expression was observed during odontogenesis and positive reactivity of the enamel organ was seen with the pan keratin antibodies CK1 (clone LP34 - reacts with a number of keratins including 6 and 18) and AE1-3 (reacts with most acidic and basic keratins). No reactivity was observed in the enamel organ with the other antibodies examined (Ks 8.12 [reacts with keratins 13 and 16], Ks 8.60 [reacts with keratins 10 and 11) and MCA157 [reacts with rat liver antigen]), although these antibodies did stain other epithelial tissues. This study supports the view that the epithelial cells of the enamel organ synthesize a tissue-specific subset of keratins which are related to the differentiation of the cells.     

An immunocytochemical study of a rat pituitary multipotential clone.

The 2A8 clone, a normal diploid rat anterior pituitary cell strain, was investigated by immunocytochemistry to determine the cell types into which the clonal cells differentiated in vivo and in vitro. The in vivo study was carried out by injecting the 2A8 clone preparation either into the hypothalamic region or under the kidney capsule. After thirty days the implants were removed and studied by immunocytochemistry. In vitro, many prolactin cells and a few growth hormone cells were found. In vivo, however, prolactin, growth hormone, ACTH and TSH cells and gonadotrophs were identified. We concluded that the 2A8 clone was multipotential. Since the gonadotrophs of the implants made in the hypothalamic region were larger and more plentiful than those in the kidney implants, and since gonadotrophs were lacking in the in vitro system, it appeared that the hypophysiotrophic environment was the most conducive to gonadotrophic differentiation and maintenance, and that the factor or factors necessary for cyto-differentiation were apparently present in the general circulation of the rat but absent in the growth medium of our culture cells.     

An immunocytochemical study of keratin reactivity during rat odontogenesis

Summary The cytokeratin distribution in the developing rat enamel organ from day 15 of gestation through to 11 days post partum was examined immunohistochemically using a panel of monoclonal antibodies. A temporo-spatial programme of keratin expression was observed during odontogenesis and positive reactivity of the enamel organ was seen with the pan keratin antibodies CK1 (clone LP34 — reacts with a number of keratins including 6 and 18) and AE1-3 (reacts with most acidic and basic keratins). No reactivity was observed in the enamel organ with the other antibodies examined (Ks 8.12 [reacts with keratins 13 and 16], Ks 8.60 [reacts with keratins 10 and 11) and MCA157 [reacts with rat liver antigen]), although these antibodies did stain other epithelial tissues. This study supports the view that the epithelial cells of the enamel organ synthesize a tissuspecific subset of keratins which are related to the differentiation of the cells.     

Earliest renin containing cell differentiation during ontogenesis in the rat. An immunocytochemical study

The differentiation of renin containing cells was studied by immunocytochemistry in normal rat fetuses by the use of highly specific renin, angiotensin I and II antisera. Renin synthesizing cells were detectable as early as the 15th day of gestation outside the nephrogen territories within the walls of mesonephrotic-gonadic and renal arteries. Intrarenal differentiation began at the 17th day and progressed along the intrarenal arterial tree. AII immunostaining appeared concomitantly in the renin containing cells and developed considerably during ontogenesis, suggesting intracellular biosynthesis. It can be suggested that in the fetus newly synthesized AII may contribute to the early systemic and renal blood pressure regulation.     

Location of phosphoglycerate mutase in rat skeletal muscle. An immunocytochemical and biochemical study

The subcellular distribution of phosphoglycerate mutase was studied by immunogold techniques. With the aid of highly affinity-purified anti-phosphoglycerate mutase antibodies, the enzyme was found in both cytosol and nucleus of rat skeletal muscle. No evidence of interaction with contractile proteins was observed in cytosol. Nuclear location was also confirmed biochemically using purified nuclear preparations from rat skeletal muscle. Only one immunoreactive nuclear band was observed by Western blot experiments and corresponded to that of phosphoglycerate mutase mobility. Activity measurements from nuclear extracts showed that 25% of total specific activity is found in the nuclei.     

Indigestible material attenuated changes in apoptosis in the fasted rat jejunal mucosa

We have previously demonstrated that fasting induced apoptosis and decreased cell proliferation in the rat intestinal mucosa. The aim was to investigate the effect of expanded polystyrene as indigestible material on apoptosis and cell proliferation in rat small intestinal mucosa during fasting. Male SD rats were divided into 3 groups. The first group was fed with chow and water ad libitum. The second group fasted for 72 hrs. The third group was fasted for 24 hrs and was fed expanded polystyrene. Intestinal apoptosis was evaluated by percent fragmented DNA assay, terminal deoxynucleotidyl transferase-mediated dUDP-biotin nick end-labeling (TUNEL) staining, and caspase-3 assay. Cell proliferation was analyzed by 5-bromo-2'-deoxyuridine (5-BrdU) uptake. Truncal vagotomy was performed to evaluate a role of the central nervous system. In the 72-hr fasted rat, mucosal height of the rat jejunum was decreased to 73% of that in rats fed ad libitum, and this decrease was partly restored to 90% in rats fed expanded polystyrene. The fragmented DNA was increased in fasted rats (28.0%) when compared with that in rats fed ad libitum (2.6%). The increase in fragmented DNA in fasted rats was recovered by feeding them expanded polystyrene (8.3%). TUNEL staining confirmed this result. The effect of polystyrene on apoptosis was decreased by truncal vagotomy. Expression of cleaved caspase-3 was increased in fasted rats, which was then decreased by feeding of expanded polystyrene. In contrast to apoptosis, feeding of expanded polystyrene had no reconstructive effect on 5-BrdU uptake in the intestinal epithelium, which was decreased by fasting to 60% of that in rats fed ad libitum. In conclusion, feeding of indigestible material partly restored the decrease in intestinal mucosal length in the fasted rats through the apoptotic pathway without any influence on BrdU uptake. Further exploration focused on the mechanism of this effect of indigestible material is required.     

Liporotein particles in the jejunal mucosa of fetal rats.

The jejunal mucosa of fetal rats contains particles of the same size, electron density and distribution as particles identified by others in adult jejunum as very low density lipoprotein particles. These particles are first seen in 19-day fetuses at which time the jejunal mucosa is still relatively undifferentiated and formation of villi is just beginning. They become progressively more abundant during the last 3 days of gestation as differentiation proceeds and are seen in virtually all absorptive cells in the apical third of jejunal villi of fetuses at term.     

Expression of epidermal growth factor in the rat kidney. An immunocytochemical and in situ hybridization study

The renal localization and the site of synthesis of epidermal growth factor (EGF) were investigated in the rat kidney by immunohistochemistry and in situ hybridization techniques. EGF was localized in the cells of the thick ascending limb of Henle (TAL) and distal convoluted tubule (DCT). At the ultrastructural level, EGF immunoreactivity was distributed on the apical membrane and trans-Golgi complex of the TAL and DCT cells. These segments of the rat nephron also hybridized to prepro-EGF cRNA probes in a specific manner, indicating that TAL and DCT are the sites of EGF synthesis in the rat kidney.     

Role of multipotent fibroblasts in the healing colonic mucosa of rabbits. Ultrastructural and immunocytochemical study.

Light- and electron microscopy and immunocytochemistry were used to study the healing colonic mucosa of rabbits after experimental excision. Between 3 and 5 days, abundant young fibroblasts which retained many features of mesenchymal cells invaded the growing capillaries into the loose connective tissue of the healing colonic mucosa. Our electron microscopy revealed the transformation of these young fibroblasts into smooth muscle cells, into histiocyte-like cells involved in phagocytotic activity, and into vasoformative cells incorporated into the growing capillaries. The mitotic proliferation of pre-existing smooth muscle cells at the ulcer margin did not seem to be the major reason for re-establishment of the muscular tissue. The present immunocytochemistry revealed an active production of fibronectin in rough endoplasmic reticulum in the young fibroblasts. This may mean that this glycoprotein is involved in the re-establishment of both connective and muscular tissues by enhancement of adhesion and chemoattractant activities of such cells. In addition, the immunoreaction of endothelial cells of the growing capillaries suggests a role of this glycoprotein in the acceleration of the neocapillarization.     

Localization of triphosphoinositide in the cochlea. An electronmicroscopic immunocytochemical study

Triphosphoinositide (TPI) has been demonstrated to be a receptor for aminoglycosides in the cochlea and may regulate ionic permeability by its binding with Ca++. This phospholipid was localized by a protein A-gold technique in the cochlea at the electronmicroscopic level. TPI was prepared by a neomycin column and antibodies to it were raised in rabbits. The antibody used in this study reacted virtually only to TPI among the tested lipids. TPI was localized mainly at stereocilia, cuticular plates, head plates of Deiters' cells, plasma membrane, and mitochondria of various cells in the organ of Corti. In the vascular stria, TPI was found mainly at the plasma membrane of basal infoldings of the marginal cells. Possible physiological and pathophysiological roles of TPI in the cochlea are briefly discussed.     

On the myoepithelium of human salivary glands. An immunocytochemical study

This study investigated the immunocytochemical characteristics of normal myoepithelial cells (MECs) of human major and minor salivary glands using the LSAB method. Other human exocrine glands were used as controls. Immunoreactivity of MECs was observed exclusively with fully differentiated smooth muscle antibodies (a-SMA; SMMS-1; CALP; hCD) and with epithelial markers (cytokeratins) Ck14 and Ck17. This epithelial-muscular immunophenotype was similarly expressed in the MECs of other human exocrine glands used as control. In the salivary MECs, we did not observe evidence for neuroectodermic phenotype (S-100 protein, GFAP, NSE). On the contrary, positivity was observed for S-100 protein in Mecs of control glands (mammary, bronchial and sweat glands). Immunoreaction for extracellular matrix markers (fibronectin, laminin and collagen IV) and vimentin always were negative. Our data show that in normal "non transformed" MECs of the salivary glands the smooth muscle phenotype is in a state of complete differentiation, while the epithelial phenotype expresses only Ck14 and Ck17. This double and simultaneous immunoreactivity represents a differential marker of MECs from the epithelial basal cell (EBCs).