Epithelial integrity is maintained by a matriptase-dependent proteolytic pathway.

A pericellular proteolytic pathway initiated by the transmembrane serine protease matriptase plays a critical role in the terminal differentiation of epidermal tissues. Matriptase is constitutively expressed in multiple other epithelia, suggesting a putative role of this membrane serine protease in general epithelial homeostasis. Here we generated mice with conditional deletion of the St14 gene, encoding matriptase, and show that matriptase indeed is essential for the maintenance of multiple types of epithelia in the mouse. Thus, embryonic or postnatal ablation of St14 in epithelial tissues of diverse origin and function caused severe organ dysfunction, which was often associated with increased permeability, loss of tight junction function, mislocation of tight junction-associated proteins, and generalized epithelial demise. The study reveals that the homeostasis of multiple simple and stratified epithelia is matriptase-dependent, and provides an important animal model for the exploration of this membrane serine protease in a range of physiological and pathological processes.

Manometry and histochemistry in the diagnosis of Hirschsprung's disease.

Acetylcholine esterase histochemistry and rectal manometry have been used sequentially to evaluate constipated children and to make the diagnosis of Hirschsprung's disease. When applied together, these modalities give a reliable diagnosis of Hirschsprung's disease, restricting the use of deep rectal biopsy to those patients in whom symptoms are most suggestive of Hirschsprung's disease. Acetylcholine esterase histochemistry may be more reliable than rectal manometry in the newborn and premature periods. It is hoped that the combined use of these techniques may make deep rectal biopsy obsolete except in patients with hypoganglionosis.

Histochemical diagnosis of Hirschsprung's disease and a comparison of the histochemical and biochemical activity of acetylcholinesterase in rectal mucosal biopsies.

Three hundred and seventy-two rectal mucosal biopsies, taken from 150 children and young adults with chronic constipation, were subjected to histochemical and biochemical analysis of acetylcholinesterase to excude Hirschsprung's disease. The relative merits of the procedures were compared. The histochemical method was considered to be the most practical for laboratories handling small numbers of biopsies but the biochemical estimation of acetylcholinesterase activity was found to be a useful complementary procedure and an accurate quantitative assessment of enzyme activity.

Experience with a cholinesterase histochemical technique for rectal suction biopsies in the diagnosis of Hirschsprung's disease.

Cryostat sections from 160 rectal suction biopsies were stained for cholinesterases by the method of Karnovsky and Roots (1964) in an attempt to facilitate the diagnosis of Hirschsprung's disease. The method proved at least as reliable as experienced assessment of paraffin haematoxylin-eosin sections, and appeared to offer the advantages of reduced scanning fatigue and superior demonstration of the increased cholinesterase-positive nerves in Hirschprung's disease. Contrary to the findings of Meier-Ruge (1971) it was not possible to base a diagnosis on mucosal cholinesterase activity.

Acquired megacolon is associated with alteration of vasoactive intestinal peptide levels and acetylcholinesterase activity.

Based upon previous morphologic studies, we hypothesized that the development of acquired megacolon was associated with abnormalities of enteric neurotransmitter concentrations and enzymatic activities. Specimens were obtained at surgery from patients with normal descending-sigmoid colon (n = 13) and patients with sigmoid megacolon (n = 6; defined by radiologic measurement). Radioimmunoassays were used to measure the non-adrenergic, non-cholinergic inhibitory neuropeptide, vasoactive intestinal peptide, and the non-adrenergic, non-cholinergic excitatory neuropeptide, substance P, while spectrophotometric assays were used to quantitate acetylcholinesterase activity and choline acetyltransferase activity. There were significantly decreased concentrations of vasoactive intestinal peptide and decreased acetylcholinesterase activity in muscularis externa from patients with acquired megacolon. In megacolon, vasoactive intestinal peptide-containing nerve fibers appeared to be diminished in circular and longitudinal smooth muscle, and immunostaining of nerve cell bodies in the plexus submucosus externus appeared diminished. These results suggest the hypothesis that production of vasoactive intestinal peptide is altered allowing secondary colonic hypertrophy to develop from prolonged cholinergic nerve-mediated contractions of circular smooth muscle. As a corollary to this hypothesis, colonic dilatation might result from prolonged contraction of longitudinal smooth muscle.

Acetylcholinesterase activity and its molecular forms in rectal tissue in the diagnosis of Hirsprung's disease.

Acetylcholinesterase (AChE) activity and its molecular forms in the rectal tissue in Hirsprung's disease were studied. Total AChE activity was increased in Hirsprung's disease; the activity was separated by polyacrylamide gel electrophoresis into two molecular forms. The AChE form with higher electrophoretic mobility was not observed in healthy children, and it might be of diagnostic importance in this disease.

An improved method for the determination of acetylcholinesterase activity in rectal biopsy tissue from patients with Hirschsprung's disease.

A method is described for the assay of acetylcholinesterase in tissue samples using the selective cholinesterase inhibitor, lysivane (10-(alpha-diethylaminopropyl) phenothiazine hydrochloride). Significantly increased enzyme activity is found in rectal biopsy specimens from patients with Hirschsprung's disease (p less than 0.001) indicating the diagnostic usefulness of the assay.

Hirschsprung's disease: an appraisal of histochemically demonstrated acetylcholinesterase activity in suction rectal biopsy specimens as an aid to diagnosis.

Suction rectal biopsy specimens, from a series of 168 infants and children with constipation and other gastrointestinal problems, were stained with a sensitive acetylcholinesterase method, and the results were compared with routine histology, radiology, anorectal manometry, and the final diagnosis. In all cases of Hirschsprung's disease, there was an increase in numbers and sizes of cholinergic nerves in the lamina propria and muscularis mucosae. No false-positive or false-negative results were found. The test was found to be more reliable and consistent than other methods available.

Serum and erythrocyte acetylcholinesterase activity in Hirschsprung's disease.

Acetylcholinesterase activity was measured in the serum and erythrocytes from 22 children. The 12 patients in whom the diagnosis of Hirschsprung's disease was confirmed, had a significantly higher concentration of enzyme in both serum and erythrocytes than the 10 children in whom the diagnosis was excluded. This systemic manifestation, may prove of value in the diagnosis of Hirschsprung's disease.

Histochemical criteria for the diagnosis of Hirschsprung's disease in rectal suction biopsies by acetylcholinesterase activity.

Histochemistry to demonstrate cholinergic fibers in rectal suction biopsies is a reliable method to diagnose or exclude Hirschsprung's disease. The histochemical criterion of a positive reaction consists of the presence of many coarse discrete cholinergic fibers in the muscularis mucosae and in the immediately subjacent submucosa, best assessed under medium power with the condenser lowered. A marked increase in cholinergic fibers in the lamina propria is seen only in some cases of Hirschsprung's disease. At least some cases of Hirschsprung's disease may be the result of destruction of ganglion cells by a viral infection in the intrauterine and early postnatal life.

Sympathetic neurotransmitter metabolism in Hirschsprung's disease.

Tyrosine hydroxylase activity was measured in high speed supernatants obtained from full thickness segments of aganglionic and ganglionic colon of three children with Hirschsprung's disease. Tyrosine hydroxylase activity expressed as pmole DOPA/mg protein/min was 0.93 +/- 0.16 in ganglionic and 2.67 +/- 0.21 in aganglionic colon. Tyrosine hydroxylase activity in ganglionic colon rose to 2.29 +/- 0.11 following calcium stimulation (100 muM) but could not be further increased in aganglionic colon. Addition of norepinephrine (2 X 10(-4) M) to tissue homogenates inhibited tyrosine hydroxylase activity in ganglionic colon by 57 +/- 8% but only by 14 +/- 3% in aganglionic colon, suggesting that the enzyme present in aganglionic colon is insensitive to feedback inhibition by endogenous norepinephrine. The elevation of tyrosine hydroxylase activity in aganglionic colon and its insensitivity to calcium stimulation and norepinephrine inhibition is further evidence of sympathetic overactivity in the aganglionic colon and suggests a basic enzymatic abnormality in the pathogenesis of Hirschsprung's disease.

[Histochemical diagnosis of Hirschsprung's disease]. / Histochemická diagnostika morbus Hirschsprung.

Examination of 65 rectal biopsies and 10 resections by the histochemical detection of cholinesterase leads to the following conclusions: 1. Density of the nerve fibers in lamina propria is irregular and the examination of the cholinesterase activity may lead to falsely negative results in aganglionosis. 2. Constant is, in aganglionosis, the finding of markedly increased reaction in the nerve fibers of the muscularis mucosae and submucosa. 3. Biopsy must, therefore, include even muscularis mucosae and the choice of the instrument for biopsy removal must meet this requirement.