Functional activity of intestinal epithelium demonstrated by mRNA in situ hybridization

To elucidate the synthetic activity in relation to the morphology of the epithelial cells of the small and large intestine, in situ hybridization with a poly-deoxyribothymidine (poly d(T)) probe was applied to paraffin sections of formalin-fixed blocks. This effectively displays poly-adenylated RNA and, by implication, messenger RNA (mRNA). By minimizing proteinase K pretreatment, the relative concentrations of cellular mRNA were visualized. This revealed minimal mRNA in crypt columnar cells, and maximal mRNA in proliferating cells and in cells showing terminal differentiation. The latter include surface epithelial cells, endocrine cells, Paneth cells, and maturing, but not mature, goblet cells. The goblet cells showing positive hybridization can be regarded as active cells and show characteristic morphology. Such cells are particularly evident in untreated coeliac disease, remitting ulcerative colitis, and transitional mucosa. The proliferating cells showing increased hybridization include normal mitotically active crypt epithelium, regenerating epithelium in ulcerative colitis, adenomatous epithelium, and adenocarcinomatous epithelium. The similarity of hybridization between metaplastic polyp epithelium and surface colonocytes is consistent with the concept that metaplastic polyps are formed of cells showing premature terminal differentiation.     

Anaphylactic-like reaction of small intestinal epithelium in parasitized guinea-pigs.

Antigens derived from Trichinella spiralis were used to challenge, in vitro, sensitized jejunum from infected guinea-pigs while monitoring ion transport properties of the tissue. Antigen challenge resulted in dose-dependent increases in trans-epithelial electrical potential difference and short circuit current. Both antigen-stimulated electrical alterations and Schultz-Dale contractions were demonstrated in small intestinal tissue after the passive transfer of immune serum containing anti-trichinella homocytotropic antibodies.     

Impaired growth of small intestinal epithelium by adrenalectomy in weaning rats

Functional maturation of the small intestine occurs during the weaning period in rats. It is known that this development is facilitated by glucocorticoid. However, the effect of glucocorticoid on morphological development of small intestine has yet to be clarified. The present study evaluated the morphological development and cell proliferation of the small intestine in adrenalectomized (ADX) rat pups. To further understand the mechanism of glucocorticoid effects on intestinal development, we examined the localization of the glucocorticoid receptor in the small intestine. Microscopic analysis showed that growth of villi and crypts is age-dependent, and is significantly attenuated in ADX rats compared with sham-operated rats. BrdU-positive cells, i.e. proliferating cells, were primarily observed in crypt compartments and rapidly increased in number during the early weaning period. The increase in BrdU-positive cells could be attenuated by adrenalectomy. The morphological development of small intestine may be associated with increased proliferation of epithelial cells. On the other hand, glucocorticoid receptors were found in epithelial cells of the mid- and lower villi and not in crypts where BrdU-positive cells were localized. These results indicate that the growth of small intestine is attenuated by adrenalectomy, and that glucocorticoid indirectly acts on proliferation of epithelial cells during the weaning period.     

Functional properties of lymphocytes isolated from murine small intestinal epithelium

Intraepithelial lymphocytes (IEL) were isolated from murine small intestine using a modification of previously published procedures. Analysis of IEL by immunofluorescence using monoclonal antibodies showed they were predominantly Lyt-2+ cells, with relatively few B cells or macrophages present. IEL cultured at sufficiently high cell densities proliferated in response to concanavalin A (Con A), phytohaemagglutin (PHA) and lipopolysaccharide (LPS). IEL were also capable of recognizing alloantigens in an in vivo graft-versus-host assay and in an in vitro mixed lymphocyte reaction. These studies therefore confirm that murine IEL contain cells with immunologic properties characteristic of typical T lymphocytes.     

Histochemical detection of glycoproteins in the intestinal epithelium of Columba livia

A histochemical investigation was carried out to detect glycoproteins in the epithelium of the small and large intestine, ceca and cloaca (coprodeum) of Columba livia. The results showed that the goblet cells of both crypts and villi of the various segments studied presented neutral and acid glycoproteins. Acid glycoproteins include sulphoglycoproteins and sialoglycoproteins with predominance of the sulphated components, particularly in the large intestine, ceca and coprodeum. In the striated border neutral glycoproteins were the predominant components in an inner wide band and the acid in a superficial pellicle.     

Transcellular transport of calcium and inorganic phosphate in the small intestinal epithelium

Transport mechanisms involved in the small intestinal handling of inorganic phosphate and calcium have been studied by different in vitro methods during the last few years. In concordance with studies on intact epithelial preparations, studies with brush-border and basal-lateral membrane vesicles isolated from the small intestinal epithelial cell revealed that transcellular calcium and inorganic phosphate fluxes are coupled to transcellular sodium flux, i.e., secondary active via coupling to the primary active sodium flux. A sodium-coupled mechanism in the brush-border membrane leads to cellular accumulation of inorganic phosphate. A sodium-coupled mechanism leads to extrusion of calcium from the cell into the serosal interstitium. A primary active transport mediated by the Ca-ATPase and located in the basal-lateral membrane also exists for calcium. Regulation of transcellular phosphate and calcium flux proceeds via altered influx rates at the luminal cell pole.     

Ultrastructure of the small intestinal epithelium of the developing pig foetus

Summary The intestinal epithelium of the developing pig foetus was studied employing electron microscopy, and included foetuses with crown to rump (C-R) lengths from 1.5 to 25 cm, corresponding to approximate foetal ages of 3–14 weeks.Marked changes in gross, microscopic and ultrastructural morphology were observed. The primitive-gut epithelium was multilayered, but became a one-layered epithelium in foetuses of approximately 8–10 cm in C-R length.The structural differentiation of the intestinal epithelium begins at an early stage of development. The first steps in this differentiation process are the most prominent: undifferentiated cells with large nuclei, few mitochondria and membranous components are replaced by cells with reduced nuclear volumes, more numerous cytoplasmic structures, and with microvilli developing in the apical parts.A comparison of the results with observations in other species revealed that the differentiation of microvilli occurred earlier in the pig than in the rat, mouse and chick, which are the only species previously studied in detail in these respects.This investigation was supported by grants from the Swedish Natural Science Research Council and the Foundation of Dir. A. Påhlsson, Malmö.The skilful technical assistance of Mrs. Anna-Greta Petersen and Mrs. Marie Adler-Maihofer is gratefully acknowledged.     

Effect of syngeneic thymocytes on proliferation of the small intestinal epithelium in mice

Bulletin of Experimental Biology and Medicine -     

Ultrastructural changes in small intestinal epithelium of neonatal pigs infected with pig rotavirus

Summary The small intestine of piglets orally infected with rotavirus was examined by electron microscopy 18, 24, 48 and 60 hours after infection. At 18 and 24 hours after infection columnar epithelial cells covered the villi. Infected epithelial cells tended to be less electron-dense than uninfected cells and were more numerous at 24 hours after infection. Two types of rotavirus particle were seen, usually within dilated cisternae of the RER: non-enveloped particles measuring 50 to 60 nm and enveloped particles measuring 65 to 75 nm. Non-membrane bound viroplasm containing electron-dense cores was encountered outside the cisternae 18 and 24 hours after infection. Tubular structures measuring 44 to 56 nm were often found in nuclei of infected cells. Single-membraned (44 to 55 nm) and double-membraned tubules (70 to 80 nm) associated with viral manufacture were found in the cytoplasm of infected cells.At 48 and 60 hours after infection a proportion of villous epithelial cells were cuboidal. Virus particles were detected in only a few epithelial cells and nuclear and cytoplasmic tubules were not seen.At all times after infection some infected cells showed a reduction in the number and size of the microvilli comprising the brush border.With 7 Figures     

Simple method for pretreatment of tissue sections for the detection of apoptosis by in situ end-labelling and in situ nick translation

Aims—To overcome the problems associated with proteolytic pretreatment of tissue sections for the detection of apoptosis.     

In situ apoptosis assay for the detection of early acute myocardial infarction.

Detection and age determination of myocardial infarction (MI) is often necessary in both clinical and pathological settings. Conventional histopathological techniques are of limited utility in the demonstration of myocardial ischemic cell death (MICD) within the first 6 hours of MI. In this study, an in situ apoptosis assay was evaluated for the determination of early MICD or early MI. Sections of formalin-fixed, paraffin-embedded archival tissue blocks from 80 hearts were stained for the presence of apoptotic cells by specific labeling of nuclear DNA fragmentation. Conventional hematoxylin and eosin stain showed acute MI (group A, n = 32), equivocal evidence for MICD or early infarction (group B, n = 35), or no abnormal findings (group C, n = 13). The sensitivity and specificity of the in situ apoptosis assay for MICD were confirmed in groups A and C patients. We showed that apoptosis of myocardial cells can occur after ischemic myocardial cell injury. Virtually all documented cases of acute MI (group A) revealed a sizeable distribution of apoptotic cells visible on gross examination of glass slides. Special attention was given to patients in group B, who were at high risk for MI and for suspected but not proved cardiac death. In this group, 34/35 cases (97%) showed focal or diffuse nuclear positivity of varying degrees for apoptosis, confirming the presence of MICD. A sizeable distribution of apoptotic cells, similar to that observed in group A, was noted in 13/35 cases (37%) of group B, suggesting acute MI in these cases. The in situ assay of DNA fragmentation can detect MICD while the histological diagnosis is still inconclusive. It is estimated that with this assay one can detect MICD as early as 2 to 4 hours.     

胃上皮异型增生及胃癌组织中细胞凋亡的原位观察

目的　探讨胃癌前病变和胃癌组织中细胞凋亡的情况。方法　采用TUNEL技术检测胃上皮异型增生及胃癌组织中的细胞凋亡率。结果　在不能肯定为异型增生中凋亡率为 2 1 94%± 5 4 8%;低度异型增生的凋亡率为 2 3 70 %± 4 13%;高度异型增生中凋亡率为 18 0 5 %± 3 99%,随着胃黏膜细胞异型程度的增高 ,细胞凋亡反而逐渐减少 ,而在浸润癌其凋亡率进一步减少为 8 13%± 1 98%。结论　在胃黏膜从异型增生到癌变过程中可能存在细胞凋亡的被抑制 ,应该凋亡的异常细胞得以长期生存 ,这些细胞对致突、致癌剂的易感性升高 ,基因更易发生突变 ,增加了胃黏膜细胞恶变的机会。     

Glycogen in the fetal intestinal epithelium

In the phase of differentiated epithelium the well-known phenomenon of glycogen incorporation in the supra- and infranuclear cytoplasm of the villous enterocytes was verified. Contrary to the role of glycan in the production of pentoses for the biosynthesis of nucleic acid in embryonic cells (Sasse 1968), which display a high rate of metabolism and proliferation, this enormous glycogen store is evidently of very little functional significance. It seems to be an expression of a disordered glycogenolysis. These masses of glycogen are incorporated into impressive glycogenosomes, which are the predominate cytoplasmic inclusion for a short period of time. According to the present results it is not certain whether lysosomal glycogenolysis takes place. There is no morphological or functional evidence for a reutilization of glycogen.     

E-cadherin expression in intestinal epithelium.

AIMS--To investigate E-cadherin expression in normal and inflamed intestine, in the colonic adenocarcinoma cell line HT29, in normal fetal intestine, and in a fetal gut organ culture model where a T cell mediated enteropathy can be generated; to determine whether expression of E-cadherin changes in intestinal inflammation. METHODS--Immunohistochemistry was used to determine E-cadherin expression in following tissues: frozen and paraffin wax sections of normal and inflamed intestine; HT29 colonic adenocarcinoma cell line cultured on coverslips in the presence or absence of cytokines; frozen sections of fetal small intestinal tissue (gestational age 11-22 weeks); and frozen sections of cultured fetal gut in which a T cell mediated enteropathy had been induced. RESULTS--E-cadherin was strongly and evenly expressed by the epithelium in all specimens of intestine studied. Although there was no change in inflammation generally, in some cases of Crohn's disease groups of glands with the characteristic morphology of "ulcer associated cell lineage" showed lower expression of E-cadherin. In fetal gut organ cultures epithelial expression of E-cadherin was lower when local T cells were activated with mitogens, compared with control explants. By contrast, the HT29 cell line showed low levels of expression which increased after treatment with conditioned medium from activated tonsil cells. CONCLUSIONS--E-cadherin is strongly and evenly expressed by epithelium in normal and inflamed intestine, although an increase in E-cadherin expression in cytokine treated HT29 cells was observed. E-cadherin expression is reduced in the epithelium adjacent to ulcers (ulcer associated cell lineage), possibly to assist regeneration.     

原位末端标记检测病毒性肝炎肝组织细胞凋亡相关线状断裂DNA

病毒性肝炎肝组织的细胞凋亡情况及其意义目前尚不十分清楚。线状断裂ＤＮＡ的存在是凋亡细胞的重要标志。我们应用原位末端标记技术检测了２２例病毒性肝炎肝活检标本。结果发现，多数阳性细胞为细胞核阳性，表现为核固缩胞浆嗜酸性变；少数为胞浆阳性，表现为细胞核消失，细胞膜张力减低；也可见到气球样变的细胞呈细胞核、细胞浆同时阳性。作者认为前两者符合细胞凋亡的形态特征，为肝组织的细胞凋亡；后者是否属于细胞凋亡尚需研究。与临床关系研究表明，肝组织的细胞凋亡可能与病毒性肝炎的慢性化有关。