Presence Of Epithelium Research Articles

Effects of sensitization on vasoactive intestinal polypeptide-induced relaxation and its concentration and binding in guinea-pig airways

We investigated the relaxant effect of vasoactive intestinal polypeptide (VIP) in trachea and lung parenchyma from normal and sensitized guinea-pigs. A technique by which drug access was restricted to either the mucosal or the adventitial surface of tracheal rings was used. In intact trachea, concentration-response curves for VIP entering from the mucosal surface (pD 2 = 6.61 ± 0.06) were displaced to the right compared with those for adventitial entry (pD 2 = 6.78 ± 0.04). Epithelium removal produced a leftward shift (≈ 2.8-fold) in the mucosal VIP concentration-response curve. Sensitization did not alter the responsiveness (maximal effect) or sensitivity (pD 2 values) of tracheal rings to VIP irrespective of the surface of drug entry and of the absence or presence of epithelium. VIP-induced relaxation of normal and sensitized lung strips was also similar. Sensitization resulted in a significant decrease in tracheal VIP content (from 2.16 ± 0.07 in normal to 0.60 ± 0.08 nmol/mg) protein in sensitized trachea; P < 0.05; n = 7 whereas the affinity of both high- and low-affinity binding sites for VIP increased as compared to that of normal trachea. Differences were not found in the binding capacities of normal and sensitized trachea. VIP content and binding did not differ in normal and sensitized lung. In conclusion, immunological sensitization produced changes in VIP tracheal content and binding but neither VIP-induced relaxation of isolated airways nor the influence of epithelium in this response was altered.

Physiologic Significance of Epithelial Removal on Guinea Pig Tracheal Smooth Muscle Response to Acetylcholine and Serotonin

We studied the modulatory effect of airway epithelium on guinea pig tracheal smooth muscle (TSM) contraction. Isometric force was measured in vivo before and after removal of the tracheal epithelium. In parallel studies, TSM contraction was also measured isometrically in epithelium-intact and epithelium-denuded TSM strips in vitro. Epithelial removal in vivo did not alter the contractile response of TSM to acetylcholine (ACh) or serotonin. In nine guinea pigs, active tension (AT) caused by 3 x 10(-7) mol/kg of intravenous ACh was 0.74 +/- 0.14 g force per longitudinal length of the segment (g/cm) in the presence of epithelium versus 0.89 +/- 0.16 g/cm after removal of airway epithelium (confirmed histologically) (p NS). The threshold response to ACh was also unchanged (-8.0 +/- 0.3 log mol/kg control versus -8.3 +/- 0.3 log mol/kg after epithelial removal, p NS). In six guinea pigs, the AT caused by 3 x 10(-8) mol/kg of intravenous serotonin was 1.92 +/- 0.63 g/cm with an intact epithelium versus 2.15 +/- 0.70 g/cm after epithelial removal in vivo (p NS). Epithelial removal in vitro increased the sensitivity of TSM contraction to ACh when the data were expressed as the percentage maximal response to ACh. The concentration of ACh causing 50% of the maximal response (EC50) was -5.74 +/- 0.25 log M in eight epithelium-intact TSM strips versus -6.37 +/- 0.16 log M after epithelial removal in controls (n = 8) (p = 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of muscarinic receptors that mediate contraction of guinea-pig isolated trachea to choline esters: effect of removing epithelium.

1. The muscarinic receptor subtype that mediates contraction of guinea-pig trachea, in the presence and absence of epithelium, to acetic and carbamic acid choline esters was determined by use of preferential muscarinic receptor antagonists: pirenzepine (M1 receptor), methoctramine (M2 receptor) and 4-diphenylacetoxy-N-methylpiperidine (4-DAMP) (M3 receptor). 2. Acetylcholine (ACh), methacholine (MeCh), carbachol (CCh), bethanechol (BeCh) and oxotremorine induced concentration-dependent contraction of guinea-pig isolated tracheal strips in the presence and absence of epithelium. Contraction to acetic choline esters (ACh and MeCh) was augmented by removal of the epithelium, whereas contraction to carbamic acid choline esters (CCh and BeCh) and oxotremorine was not influenced by removal of the epithelium. 3. Pirenzepine, methoctramine and 4-DAMP caused parallel rightward displacements of the concentration-contraction curves to the muscarinic agonists. The pA2 values (determined from Arunlakshana-Schild graphs) for pirenzepine and 4-DAMP in guinea-pig trachea in the presence of epithelium were: ACh as the agonist, 7.6 and 9.0, respectively; CCh as the agonist, 7.6 and 9.1, respectively. The apparent pKB values for methoctramine with the same system were: ACh as the agonist, 5.6; CCh as the agonist, 5.6. Similar values were obtained with MeCh, BeCh and oxotremorine as the agonists. These values were agonist- and epithelium-independent. 4. It is concluded from the pA2 and apparent pKB values obtained for the muscarinic receptor antagonists used in this study that contraction of guinea-pig isolated trachea, with and without epithelium, to both acetic and carbamic acid choline esters is mediated via the muscarinic M3 receptor subtype.Differential contractile responses of guinea-pig trachea to acetic and carbamic acid choline esters upon the mechanical removal of the epithelium may not be explained by activation of different muscarinic receptor subtypes by these agonists.

Modulation of bradykinin responses in airway smooth muscle by epithelial enzymes

We have studied the effect of epithelium removal on responses of guinea pig trachea to bradykinin (BK). BK (1 nM - 10 microM) gave a concentration-dependent relaxation when epithelium was present (E+: EC50 = 10 +/- 3 nM). Epithelium removal resulted in a biphasic response to BK with relaxation at low concentrations (E-: EC50 = 3.0 +/- 1.0 nM) and a recontraction to baseline at higher concentrations (EC50 = 2.0 +/- 1 microM). Phosphoramidon (10 microM), an inhibitor of neutral endopeptidase (NEP), which cleaves BK into inactive peptides, potentiated relaxation (EC50 = 1.0 +/- 0.9 nM in E+ and E respectively) and contraction in trachea with intact epithelium (EC50 = 0.08 +/- 0.03 microM). Inhibition of cyclooxygenase by indomethacin (5 microM), inhibited relaxation to BK in E+ tracheal segments, resulting in a slight contraction (EC50 = 1.0 microM), whereas a potent contractile response was observed in E- segments (EC50 1.6 microM, maximal contraction greater than 1 g). In the presence of both indomethacin and phosphormidon BK caused contraction, even in the presence of epithelium (EC50 = 0.2 +/- 0.11 microM), and the response in the absence of epithelium was similar to the response observed in trachea with intact epithelium (EC50 = 0.25 +/- 0.1 microM). The contractile effect of BK on airway smooth muscle may be inhibited by a protective role of epithelium, due to release of relaxant prostanoids and by degradation by epithelial NEP. In asthma, bronchoconstrictor responses to BK may be partly explained by loss of airway epithelium.

The inhibitory influence of tracheal mucosa mounted in close proximity to canine trachealis

Reduced smooth muscle contractile responses to agonists occur in the presence of epithelium, perhaps due to the release of an epithelium-derived relaxing factor (EpDRF). It is not clear whether the release of EpDRF requires the direct attachment of the epithelium to the smooth muscle. In the present study, using isolated canine tracheal smooth muscle strips, we examined whether the inhibitory effects of airway mucosa require the attachment of the mucosa to smooth muscle. The smooth muscle contractile responses to acetylcholine and histamine were reduced in the presence of airway mucosa, whether the mucosa was attached or in close proximity. The inhibitory effect mediated by the airway mucosa therefore is not dependent on mucosal attachment to smooth muscle. This phenomenon appears to be due to the release of a soluble, short-acting mediator from the airway mucosa.

Tenascin during gut development: appearance in the mesenchyme, shift in molecular forms, and dependence on epithelial-mesenchymal interactions [published erratum appears in J Cell Biol 1989 Mar;108(3):following 1175

Tenascin, an extracellular matrix protein, is expressed in the mesenchyme around growing epithelia in the embryo. We therefore investigated whether epithelial cells can stimulate expression of tenascin in embryonic mesenchyme. Mesenchyme from the presumptive small intestine was used because it is known that reciprocal epithelial-mesenchymal interactions are important for gut morphogenesis. Rat monoclonal antibodies against mouse tenascin were raised and were found to react specifically with mouse tenascin in ELISA. In supernatants of cultured fibroblasts, the antibodies precipitated two peptides of Mr 260 and 210 kD. One of the antibodies also reacted with these tenascin chains in immunoblots of tissue extracts. We found that tenascin was absent during early stages of gut development, at stages when the mesenchyme is already in contact with the stratified epithelium of the endoderm. Rather, it appeared in the mesenchyme when the homogenous endodermal epithelium differentiated into the heterogenous absorptive epithelium. Tenascin remained present in the stroma of the adult gut, close to the migration pathways of the continuously renewing epithelium. When first detected during intestinal differentiation, the 210-kD component was predominant but at birth the relative amount of the 260-kD component had increased. The expression data suggested that the appearance of tenascin in the mesenchyme was dependent on the presence of epithelium. To test this, isolated gut mesenchymes from 13-d-old mouse embryos were cultured for 24 h either alone or together with epithelial and nonepithelial cells. Whereas mesenchyme cultured alone or in the presence of nonepithelial B16-F1 melanoma cells produced only trace amounts of tenascin, expression was strongly stimulated by the epithelial cell line, Madin-Darby canine kidney (MDCK). We propose that growing and differentiating epithelia produce locally active factors which stimulate synthesis of tenascin in the surrounding mesenchyme.

Comparison of the effects of epithelium removal and of an enkephalinase inhibitor on the neurokinin‐induced contractions of guinea‐pig isolated trachea

1. The influence of epithelium removal and/or thiorphan on the effects of neurokinins (substance P (SP), neurokinin A (NKA), neurokinin B (NKB)) and related peptides on airway contractility was investigated on the guinea-pig isolated trachea. 2. Removing the tracheal epithelium significantly enhanced the sensitivity but not the maximum contractile responses to the peptides. 3. After removal of the epithelial layer, the shifts to the left of the log concentration response curves were greater for SP and SP-OMe (1.62 and 1.94 log units, respectively) than for two SP analogues substituted in position 9 namely [Pro9]SP sulfone and [beta-Ala4, Sar9]SP(4-11) sulfone (0.66 and 0.68 log units, respectively). The leftward shifts for compounds related to NKA or NKB lay between 0.58 and 0.73 log units. 4. The leftward shifts of the log concentration-response curves for SP, SP-OMe, [Pro9]SP sulfone, [beta-Ala4, Sar9]SP(4-11) sulfone and NKA were of similar magnitude after removal of the epithelium or after pretreatment with thiorphan (10(-5) M), an enkephalinase inhibitor, in the presence of epithelium. No significant additional shift of the curves to the left was observed with thiorphan plus epithelium removal. 5. The results obtained with the selective agonists for each of the three classes of neurokinin receptor (i.e NK1, NK2, NK3) suggest that the guinea-pig trachea contains receptors for SP and NKA but few if any for NKB. 6. It was concluded that neurokinins and related peptides (especially SP and analogues not substituted in position 9) are degraded by enkephalinase mainly located in the tracheal epithelium and that the addition of thiorphan or epithelium removal results in an inhibition or loss of enkephalinase activity, thereby increasing similarly the potencies of these peptides. It was, therefore, suggested that the supersensitivity to neurokinins produced by epithelium removal was due neither to the elimination of a permeability barrier nor to reduced production of a relaxant factor, but mainly to reduced peptide degradation.

Influence of epithelium on the responsiveness of guinea-pig isolated trachea to adenosine.

1. The influence of epithelium removal on the effects of adenosine on airway contractility was investigated on the guinea-pig isolated trachea. 2. In preparations under resting tone or precontracted with histamine 10(-5) M, removal of the tracheal epithelium resulted in similar shifts to the left of the adenosine concentration-response curves (0.61 +/- 0.18 (P less than 0.05) and 0.80 +/- 0.09 (P less than 0.001) log units; n = 5), corresponding to 4.07 and 6.31 fold potentiations of the relaxant effect of adenosine. 3. In the presence of dipyridamole 10(-5) M the relaxant effects of adenosine were potentiated 85.1 fold on tracheae with epithelium; removal of the epithelium did not produce a significant additional shift to the left of the adenosine concentration-response curves (0.07 +/- 0.03 log units; n = 5; NS). 4. In the absence of dipyridamole, the theophylline-adenosine antagonism was not of the competitive type, irrespective of whether the tracheae were with or without epithelium. 5. In the presence of dipyridamole, this antagonism was likely to be of the competitive type and its characteristics were the same when the epithelium was present or absent. Regression slope and pA2 values were 0.84 and 5.07, respectively, in the presence of epithelium and 0.76 and 4.89, respectively, in its absence. 6. It is suggested that, at least in the guinea-pig isolated trachea model, the airway epithelium seems to be involved only in the uptake and metabolism of adenosine.

Nonneural components in the response of fresh human airways to electric field stimulation.

Fresh human bronchi, obtained at thoracotomy and maintained at 37 degrees C, were studied in vitro to investigate their response to electric field stimulation (EFS). We found complex responses that were not only composed of a rapid initial nerve-mediated cholinergic contraction and a non-adrenergic nerve-mediated relaxation, but, in 80% of preparations, also of a tonic contraction with a sustained time course. This sustained phase was not blocked by the nervous conductance blocker tetrodotoxin (TTX) and was therefore not neurally mediated. Controlled transient cooling to 4 degrees C in the organ bath reduced this sustained phase selectively for several hours. The leukotriene (LT) antagonist FPL 55712, dexamethasone, which inhibits phospholipase A2, and the antiasthmatic drug cromolyn all reduced the sustained phase significantly. In 20% of strips, an additional TTX-resistant contraction was seen directly after the cholinergic phase. This contraction could be inhibited by indomethacin. A similar small peak sometimes appeared after selective blocking of either the cholinergic or the sustained phases. Experiments in which the epithelium was removed from the strips suggested that this indomethacin-sensitive response, but not the sustained phase, was dependent on the presence of epithelium. These results show that EFS of fresh human bronchi stimulated cholinergic and nonadrenergic inhibitory nerves and gave rise to a partly epithelium-dependent synthesis of arachidonic acid metabolites, which caused contractile responses that interfered with the neurally mediated responses.

Proximal–distal sequence of development of the skeletal tissues in the penis of rat and the inductive effect of epithelium

The penis of adult rats comprises the corpus cavernosum penis and the proximal and distal segments of os penis which are situated proximal-distally in this order. Androgens are necessary for the phenotypic differentiation of these tissues. In the present study, genital tubercular mesenchyme of foetal rats was recombined with or without homologous or heterologous epithelia and transplanted beneath the kidney capsule of syngeneic adult male rats, and the development of the corpus cavernosum penis and the os penis in the transplants was examined. Only in the presence of epithelia can the genital tubercular mesenchyme acquire the capacity for differentiation into the corpus cavernosum penis, the proximal segment of os penis, and the distal segment of os penis in this chronological order. The inductive effect of epithelium is a permissive step in the differentiation of the os penis of rat. The epithelium seems to be necessary for the process of rudiment formation of the os penis and the corpus cavernosum penis.

The effect of airway epithelium on smooth muscle contractility in bovine trachea.

In bovine tracheal smooth muscle the presence of airway epithelium significantly reduced the sensitivity and maximum contractile response to histamine, 5-hydroxytryptamine (5-HT) or acetylcholine. Muscle contraction induced by K+ and electrical field stimulation was of similar magnitude both in the presence or absence of adherent epithelium. The effect of epithelium on smooth muscle contractility was unaffected by pretreatment with indomethacin (10(-6) M) or mepacrine (5 X 10(-5) M). The relaxant response to isoprenaline was enhanced in the presence of epithelium, although this was significant only in the case of precontraction with 5-HT. It is concluded that the bronchial epithelium may produce a relaxant factor which is not a cyclooxygenase or lipoxygenase product. The production of this factor may be reduced or lost following epithelial damage and this may be important in the pathogenesis of bronchial hyperresponsiveness in asthma.

Synthesis of glycosaminoglycans in corneal organ cultures

Incorporation of [ 3H]-glucosamine into glycosaminoglycans in rabbit corneas in tissue culture has been investigated. Autoradiography showed that the label was available to the stroma even in the presence of epithelium and endothelium. The acid glycosaminoglycans were isolated by protease digestion and ECTEOLA-chromatography and were identified by sequential degradation with specific enzymes and nitrous acid. The main part of the glycosaminoglycans could be accounted for as chondroitin sulphates, dermatan sulphate, N-sulphated and N-acetylated heparan sulphate, and keratan sulphate but an undegraded fraction remained. This contained galactosamine and corresponded to about 10–15% of the total incorporation. The pattern of synthesis changed with incubation time. The proportion of labelled keratan sulphate decreased and the proportion of labelled chondroitin sulphate and dermatan sulphate increased when incubations were extended from 2 to 48 hr. This change in biosynthesis was not due to swelling of the corneas during the incubation since it still occurred when swelling was prevented by the presence of dextran in the medium. The total incorporation and distribution of label between the different glycosaminoglycans in 48-hr incubations were approximately the same in intact corneas and in corneas that had been gently freed from epithelium, endothelium or both, thus indicating that the glycosaminoglycans are synthesized mainly in the stroma. A deep dissection of epithelium and/or endothelium which also injured the subepithelial layer and Descemet's membrane had a pronounced effect on the incorporation pattern. When both limiting layers were removed there was a drastic decrease in total incorporation. Injury to the antierior layer led to an increased proportion of labelled chondroitin sulphate and dermatan sulphate and a decreased proportion of the keratan sulphate-rich fraction. Injury to the posterior layer had the opposite effect. This asymmetry in the biosynthesis was confirmed in experiments where anterior and posterior parts of the cornea were investigated separately. The anterior part synthesized mostly the keratan sulphate-rich fraction and the posterior part mostly chondroitin sulphate and dermatan sulphate. Human corneas with keratitis and to a certain degree rabbit corneas with keratoconus-like syndrome showed incorporation patterns similar to that displayed by corneas with injury to the anterior layer. Incubation of corneas with 10 −7 m-10 −5 m ouabain led to a dose-dependent drop in total incorporation. At the highest drug concentration an increase in the proportion of hyaluronate and a decrease in the proportion of dermatan sulphate were observed. Thus this drug, which is frequently used to inhibit transport in the limiting cell layers also interferes with the normal functions of the stromal cells.

Microscopic evaluation of pocket depth measurements performed with six different probing forces in dogs.

This study investigates the relationship between probing force and probe penetration into the periodontal tissues in pockets with overt gingivitis and in pockets with minimal gingival inflammation. In eight dogs experimental periodontal tissue breakdown was induced during a period of 14 weeks. The dogs were then distributed over two groups of four dogs each. In group the dogs were subjected to meticulous tooth cleaning once every day for a period of 3 weeks. The dogs in the second group did not receive any oral hygiene measures. Next, wooden probes were inserted, mesial and distal to each premolar. In each dog six different probing forces were used. Microscopic examination showed that, in the brushed group, epithelium was always present between the probe tips and the connective tissue. In the nonbrushed group presence of epithelium was found in 21 of 23 specimen. In both groups of dogs it was found that, with increasing probing forces, the location of the tips of the probes changed from a position occlusal to the most coronal connective tissue fibers to a position apical to the most coronal connective tissue fibers. With light probing forces it appeared that in the nonbrushed group the tips of the probes were located more apically. When greater probing forces were used, no differences between the brushed and the nonbrushed group were found. It is suggested that, using a probe of 0.63 mm in diameter, the optimal force level for clinical pocket measurements is about 0.75 N, or in other words, the optimal probing pressure is about 240 N/cm2.

STUDIES ON PROTEIN METABOLISM OF THE CORNEA: 2. THE INCORPORATION OF C14-LABELED AMINO ACIDS INTO COMPONENTS OF THE RABBIT CORNEA

Preliminary studies were made on the incorporation of C14-labeled glycine and proline into tissue cultures of rabbit stromal cells, excised rabbit corneas, and rabbit corneas in vivo. Some evidence has been presented that this incorporation reflects synthesis, chiefly into proteins. With rabbit corneas in vitro, no evidence was found that the presence of epithelium increased the incorporation of amino acid into constituents of the stroma. During in vivo incorporation it was found that the proteins of the endothelium were about twice and the proteins of the epithelium about four times as active as those of the stroma.

The effects of desalivation on periodontal tissues of the Syrian hamster

Sixty hamsters were used to study the effects of desalivation on the incidence of periodontal lesions under three different dietary regimens. The desalivated animals consistently showed a greater number and extent of periodontal lesions in the soft and calcified tissues than their controls. Highly statistically significant differences were observed between the lesions of the control animals and those of the desalivated animals. The consistency and composition of diet seemed to play a role in the production of periodontal lesions in the hamster. High-carbohydrate, finely powdered, cariogenic ration 4 produced a greater number and extent of soft-and calcified-tissue lesions than either the laboratory chow or the ground laboratory chow. Ground laboratory chow produced a greater number and extent of lesions than laboratory chow in pellet form. The effects of the diets on the soft tissues appeared to be more severe than on the calicified tissues. The microscopic findings substantiated the gross observations. The desalivated animals showed histologic changes of greater severity than the intact controls. The pronounced changes in the desalivated animals were apical migration of the epithelial attachment and the presence of epithelium at the interradicular area. The control and desalivated animals fed ration 4 showed severe osteoporosis, possibly due to the nutritional inadequacy of the diet.

Activation of corneal stromal cells to take up the vital dye neutral red

1. 1. A procedure is described for producing in vitro the activation of corneal stromal cells to transport the vital dye neutral red. The amount of dye taken up by the corneal stromal cells was quantitated by extraction of the dye from the cornea and measurement of the amount of dye extracted. 2. 2. Normal corneal stromal cells do not take up the dye, but cells activated by injury concentrate the dye approximately 150–250-fold. 3. 3. The transport of neutral red by corneal stromal cells may be broken down into 2 steps: ( a) activation of the corneal stromal cells to transport the dye, and ( b) the actual transport of the dye by the activated cells. 4. 4. Some factor (or factors) given off by the epithelium appears to be necessary for the activation of the cells. If the epithelium is removed atraumatically activation of the cells does not occur and dye is not taken up. In the presence of mildly injured (by cutting) epithelium, activation occurs during the 4th post-injury hour. Severe injury, produced by removing the epithelial cells by crushing them, leads to activation within 1 hour. Direct trauma to the corneal stromal cells in the absence of the epithelium does not activate them to transport neutral red. 5. 5. Proteolytic activity in the injured epithelium apparently leads to the liberation from the epithelium of the factor (or factors) which activates the corneal stromal cells. Soybean trypsin inhibitor prevents activation of the corneal stromal cells in the presence of epithelium but not in the absence of epithelium (removed traumatically). Trypsin is an effective activator of the neutral red transport by corneal stromal cells. Trypsin, however, causes the reaction only in the presence, but not in the absence, of epithelium.

A survey of seventy-five cases

The odontogenic tumors are classified into three groups: Epithelial, mesenchymal, and mixed. The dentinoma, a pure mesenchymal tumor, is composed of connective tissue in which denticles or islands of irregularly formed dentine are present. The odontogenic mixed tumors consist of epithelial and mesodermal elements which are in combination in various proportions and arrangements. Three types are recognized: Soft, soft and calcified, and calcified. The soft type has been differentiated from the solid adamantoblastoma. There is evidence of the inductive influences of one tissue on another in the odontogenic mixed tumors. It is noted that epithelium in these tumors seems to stimulate dentine formation, but that the presence of epithelium is not necessary for the production of dentine. Also, dentine is formed in the presence of epithelial cells not differentiated into ameloblasts. Neoplastic adamantine tissue and enamel-forming ameloblasts have been distinguished. The presence of these two types accounts, in part, for the formation of the soft and calcified odontogenic mixed tumors.