Microsatellite instability in Bulgarian patients with endometrial cancer.

PURPOSE: Microsatellite instability (MSI) is a frequent event in different types of cancer. In several studies MSI was shown to have both clinical and prognostic value. The aim of our study was to determine the frequency of MSI in Bulgarian patients with endometrial cancer (EC) and the possible relation of this phenomenon to their clinicopathological characteristics. PATIENTS AND METHODS: A total of 33 histologically confirmed EC patients were analyzed for tumor MSI using a panel of 6 microsatellite markers. RESULTS: We identified MSI in 30% of endometrial cancer cases. Six of them had high degree of MSI (MSI-H), and 4 displayed low degree of MSI (MSI-L). CONCLUSION: The frequency of MSI in Bulgarian EC patients does not differ significantly from that reported in other European studies.

Significance of tenascin-C, fibronectin, laminin, collagen IV, alpha5beta1 and alpha9beta1 integrins and fibrotic capsule formation around liver metastases originating from cancers of the digestive tract.

The formation of a fibrotic capsule around liver metastases may functionally act as a barrier to local invasion. However, the prognostic significance of exstracellular matrix (ECM) and of some integrins' deposition around liver metastases remains unclear. An immunohistochemical investigation was carried out on 55 patients with synchronous liver metastases from colorectal and gastric cancers. Encapsulated metastases were detected in 60% of the cases. The 'non-capsular' cases showed clear immunostaining for tenascin-C, fibronectin, collagen IV, laminin, alphaSMA and integrins. On opposite, most of the cases with 'capsule' were negative for the studied ECM proteins and the two integrins. The patients with 'capsular' pattern had significantly longer median survival after the surgery compared to those with non-encapsulated metastases. The presence of tenascin, fibronectin, fibronectin receptor and laminin, as well as the strong immune signal for alphaSMA and collagen type IV in the sinusoids attached to the liver metastases was associated with a worse prognosis. The cells, forming ECM in the sinusoids attached to metastases in the 'non-capsular' pattern were alphaSMA-positive myofibroblasts. It was shown ultrastructurally that they were HSCs. The results indicate that fibrotic capsule formation is associated with longer survival after surgery. The appearance of tenascin-C and of its receptor at the periphery of liver metastases could be used as a sign of invasiveness.

Intercellular adhesion molecule-1 (ICAM-1) expression in the liver of patients with extrahepatic cholestasis.

ICAM-1 mediates the recruitment of neutrophils through the endothelium to the site of inflammation by the ICAM-1/Mac-1 and ICAM-1/LFA-1 adhesion pathways. In extrahepatic cholestasis, recruitment of neutrophils is a main feature of the inflammatory infiltrate in areas of parenchymal damage. The aim of the present study was to describe the light and electron microscopical localization of ICAM-1 expression in the liver of cholestatic patients. The peroxidase-antiperoxidase technique was used. Increased ICAM-1 expression was detected on sinusoidal endothelial and Kupffer cells. A de novo ICAM-1 expression was described on some Ito cells and the sinusoidal hepatocyte membrane in areas of parenchymal injury. In the portal areas of livers of cholestatic patients, ICAM-1 was observed on the endothelial surface of portal veins and on hepatic arteries. Occasionally, ICAM-1 was found on the surface of bile duct epithelia. It is suggested that ICAM-1 expression is up-regulated by cytokines like TNF-alpha, IL-1 and interferons released from activated Kupffer cells. The mechanisms of ICAM-1 upregulation and neutrophil recruitment in the liver during extrahepatic cholestasis are discussed.

Ultrastructural sinusoidal changes in extrahepatic cholestasis. Light and electron microscopic immunohistochemical localization of collagen type III and type IV.

Extrahepatic cholestasis causes excessive extracellular matrix formation perisinusoidally. Ito cells, transitional and endothelial cells are considered to be a source of extracellular matrix proteins in experimental cholestasis. The localization of collagens type III and type IV in human liver in extrahepatic cholestasis was investigated immunohistochemically in the present study. Immersion fixation was used after modification to be applied to surgical biopsies with commercially available kits. Sinusoidal changes were observed that indicated excessive collagen and matrix formation. Light microscopically, increased immunostaining with the two collagen antibodies was found perisinusoidally and portally. Ultrastructurally, collagen type III positive fibres were found beneath basement membranes of vessels, in collagen bundles and as a fibrillar network in the space of Disse. Collagen type IV immunostaining was located in portal tracts and near hepatocyte microvilli. Intracellular staining with collagen type IV was detected in the rough endoplasmic reticulum of some transitional cells. Immunostaining was located around transitional cells, Ito cells or endothelial cells mainly. Our study indicates that Ito cells, transitional and endothelial cells are the main source of collagens type III and IV in the space of Disse in extrahepatic cholestasis in humans.

Carcinoma-associated collagen type III and type IV immune localization and Ito cell transformation indicate tumor-related changes in sinusoids of the human liver.

The deterioration of extracellular matrix turnover is a key event in tumor progression. It has been assumed that Ito cell transformation is stimulated by tumor-derived factors. In the present study changes in the occurrence of collagen type III and IV and Ito cell transformation are described in the sinusoids of patients with malignant gastrointestinal tumors without liver metastases, and around metastatic liver tumors using routine histology, electron microscopy as well as light microscopical and ultrastructural immunohistochemistry. Dilated sinusoids filled with lymphoid cells and variable perisinusoidal fibrosis were detected light microscopically. Collagen type III and IV immune deposits were increased perisinusoidally. Ultrastructural immunohistochemistry showed increased staining in the space of Disse and around Ito and transitional cells for both types of collagen. Ito cells were transformed into transitional cells. Pit cells appeared in the inflammatory infiltrate in sinusoids. Ito cells were significantly increased in number pericentrally and periportally. It is suggested that stimuli, which can influence Ito cellular behaviour are produced by inflammatory cells in sinusoids, resident sinusoidal cells, tumor cells or by tumor stroma. It is concluded that transformed Ito cells and increased amounts of collagen type III and IV in sinusoids of patients with malignant tumors without liver metastases or around metastatic tumors may predict tumor-related alterations of liver parenchyma, which may serve as a barrier for further outgrowths of the cancer cells.

Peptidergic nerve fibres in the human liver.

The autonomic nervous system plays a significant role in liver physiology and pathology. The aim of the present study was to investigate peptidergic nerve fibres in the liver of patients with malignant gastrointestinal tumors that are not metastasizing in this organ. Using light and electron microscopic immunohistochemistry, somatostatin (SOM)-, neuropeptide Y (NPY)-, substance P (SP)- and calcitonin gene-related peptide (CGRP)-immunoreactive (IR) nerve fibres (NF) were detected in the portal tract and perisinusoidally. Histologically, the liver showed dilated sinusoids, filled with lymphoid cells, and scarcely marked perisinusoidal fibrosis. Neuropeptide-IR NF were found in close contact with hepatic sinusoids. Numerous IR varicosities were detected in the sinusoidal wall. We discuss the origin and role of these NF in the liver. Probable quantitative changes in peptidergic NF ensue the inflammatory reaction in sinusoids in malignant gastrointestinal tumors. This could also reflect the increased exposure of the liver to toxic substances in the portal blood flow.

Immunocytochemical study on the liver innervation in patients with cirrhosis.

In the liver, the autonomic nervous system plays an important role in degenerative and inflammatory changes. The aim of the present study was to investigate the distribution of neuronal fibres containing neuropeptides in livers of 5 patients with cirrhosis by immunocytochemical localization at the light and electron microscopical level of substance P (SP), neuropeptide Y (NPY), somatostatin (SOM), and calcitonin gene-related peptide (CGRP). In patients with alcoholic cirrhosis, a decreased number of neuronal fibres was found in the portal tract and fibrous septa as well as in the sinusoids of regenerative nodules. NPY- and SP-immunoreactive neuronal fibres were more numerous than CGRP-containing fibres. They were located mainly in portal tracts. These findings led to the conclusion that peptidergic innervation plays a role in inflammatory and fibrotic changes in cirrhotic liver.

Immunohistochemical detection of collagen type III and IV in relation with transformation of Ito cells in liver sinusoids of patients with reactive biliary hepatitis.

Reactive biliary hepatitis is a defined morphological entity, which is a result of chronic diseases of the gall bladder, biliary ducts or pancreas. The aim of the present study was to describe the morphology of reactive biliary hepatitis and its significance for progression of liver fibrosis, and in particular Ito cell (fat storing cell) transformation and occurrence of collagen type III and IV in the liver. Liver tissue from 19 patients with reactive biliary hepatitis was investigated light microscopically and immunohistochemically. Histologically, the liver showed features of mild to severe portal and lobular inflammation. The number of Ito cells increased periportally and pericentrally. Deposition of collagen type III and IV was increased in portal tracts, septa and perisinusoidal spaces, mainly in periportal zones of the lobules. Ultrastructurally, collagen type III immunoreactive fibrillar networks were found to be increased in the space of Disse around transitional cells. Collagen type IV immunoreactive deposits were detected around newly proliferating bile ducts in portal stroma and in the space of Disse. Ito cells were mainly transformed into transitional and myofibroblast-like cells. We discuss here the role of Ito cells and certain cytokines in the process of fibrosis of the liver in the course of reactive biliary hepatitis. It is proposed that bile acid retention in bile ducts during non-specific reactive inflammation or a gut endotoxin may cause transformation of Ito cells and increased collagen type III and IV in this type of hepatitis.

Rat liver pit cells following administration of the glycopeptide preparation (Polyerga).

BACKGROUND/AIMS: Pit cells are specific liver population which has been shown to express NK activity against tumor and viral target cells. We investigated the influence of the glycopeptide preparation (Polyerga) as one of the biological response modifiers on liver pit cells of male Wistar rats. MATERIALS AND METHODS: The number of pit cells was counted by light and electron microscopy in perfusion-fixed liver sinusoids. An electron microscopical cytochemical staining for endogenous peroxidase was done. RESULTS: After a single intramuscular injection of Polyerga, a 3-to 6-fold increase in the number of pit cells was observed within a period of 1 to 7 days. Wide contact without specific junctions between pit cells and Kupffer cells was frequently detected. Granules, rod-cored vesicles and their closely related structures empty vesicles as well as hypertrophied Golgi apparatus were accumulated in the cytoplasm of pit cells next to contact surface. The number of ribosomes and polysomes in pit cell cytoplasm was significantly increased, which suggests activated protein synthesis. CONCLUSIONS: These changes are considered to represent the morphological expression of pit cell activation. It is concluded that the glycopeptide preparation Polyerga can induce proliferation of activated pit cells in the rat liver.

Ito cell morphology, alpha-smooth muscle actin and collagen type IV expression in the liver of patients with gastric and colorectal tumors.

The alteration in sinusoidal collagen type IV occurrence, and myofibroblastic (alpha-SMA-positive) Ito cellular transformation are described in the liver of patients with malignant gastric and colorectal tumors, using electron microscopy as well as light microscopical and ultrastructural immunohistochemistry. The ultrastructural finding revealed transformation of Ito cells mostly into transitional cells in highly differentiated primary tumors and into transitional and myofibroblast-like cells with expressed changes in the other sinusoidal cells in poorly differentiated tumors. Ito cell numbers increased significantly in the livers of cancer patients. A highly significant statistical association was obtained between Ito cell numbers on the one hand and collagen type IV and alpha-SMA immunoreactivity on the other hand in the pericentral zone of the liver lobule. Ultrastructural immunohistochemistry showed increased collagen IV immune deposits in the space of Disse, assembled for the most part around and inside transitional cells. Alpha-SMA immunoreactivity was detected in activated Ito cells diffuse in the lobule, with stronger expression in the intermediate and pericentral zones. It is suggested that stimuli which can influence Ito cell transformation are produced by tumor cells from the primary tumor (TGF-beta1, TNF-alpha, PDGF-beta etc.) and from the metastasizing gastric or colorectal tumor cells--matrix metalloproteinase-2 (MMP-2). It is suggested that sinusoidal extracellular matrix deterioration creates a barrier for cancer invasion on the one hand, or possibly facilitates metastasizing by ensurance of matrix for adhesion on the other hand.

Collagen type III and type IV detection in and around human hepatocellular carcinoma.

Extracellular matrix proteins participate in tumor cell growth and progression. Their role in the extratumoral liver tissue needs to be elucidated. Eight patients with hepatocellular carcinoma on noncirrhotic livers are investigated by means of light microscopical and ultrastructural immunohistochemistry for collagen type III and type IV. In the tumor collagen type III, staining is weaker, and collagen type IV is increased. It is topographically located near perisinusoidal stromal cells. In the extratumoral liver tissue, the immunostaining for the two antibodies is stronger perisinusoidally. The number of Ito cells increases significantly in the extratumoral liver tissue. A lot of transitional cells are found there. Sinusoids in the extratumoral tissue are dilated and filled with lymphoid cells and platelets. The presence of matrix proteins between tumor cells is necessary to regulate their growth and differentiation. The increase in extracellular matrix content perisinusoidally in the extratumoral tissue probably erects a protective barrier against metastasizing tumor cells. There, collagen type III and type IV accumulation is probably initiated by signals coming from tumor cells, or from inflammatory cells and platelets in sinusoids.

Tenascin immunoreactivity in the large bowel and the liver in patients with colorectal cancer.

The expression of tenascin in colorectal tumours and liver was investigated in 30 patients with colorectal adenocarcinomas. Tissue samples were immersion-fixed in 4% paraformaldehyde solution. Free-floating cryostat sections were incubated with monoclonal antibody against tenascin, and examined by light and electron microscopy. Tenascin immunostaining was positive in sub-basement membrane zones and in newly-formed connective tissue of the primary tumour and perisinusoidally in the liver. The immunoreactivity in the sub-basement membrane zones of tumour glands in well- and moderately-differentiated tumours was more intensely expressed compared to that in poorly-differentiated tumours (p = 0.007 and p = 0.001 respectively, chi2-test). Perisinusoidal tenascin deposition was more often detected in the liver of patients with well-differentiated tumours (p = 0.006, chi2-test). The presence of metastases was accompanied by low tenascin deposition (p < 0.005, Fisher's exact test). Ultrastructurally tenascin deposits were observed around single tumour cells and glands in the primary tumours, and close to hepatic stellate cells in the liver. Finally, the role of tenascin deposition in the stimulation of tumour cell proliferation and mobility is discussed.