Virchows Archiv. B, Cell pathology including molecular pathology最新文献

Monolayer cultures of renal tubular (hKEC) cells were established. These cells formed empty spheroids after 2-3 weeks of culture in a collagen gel matrix. A subcellular polarity from the apex to basement was induced in these "spheroidal" hKEC cells. The weak expression of laminin at the outer surface was evident on spheroidal but not on monolayered hKEC cells. The regulation of HLA-ABC, DR, and intercellular adhesion molecule-1 (ICAM-1) antigens on hKEC cells in the gel matrix was investigated utilizing digestion of gel matrix by collagenase. Enzymatic digestion of the collagen gel did not significantly affect the surface expression of HLA-ABC and ICAM-1, but reduced HLA-DR expression as shown by flow cytometry. The MHC and ICAM-1 molecules on both spheroid-forming and monolayered hKEC cells were upregulated by adding a supernatant of mixed lymphocyte reaction (MLR) and recombinant human interferon (IFN)-gamma. HLA-DR antigen expression was inconsistently induced on the hKEC cells cultured in collagen gel without MLR supernatant or IFN-gamma. In contrast, no HLA-DR expression was found on monolayered hKEC cells in the absence of MLR supernatant or IFN-gamma. Spheroid-forming hKEC cells, when dispersed by enzymatic digestion, were more susceptible to cytolysis by lymphokine-activated killer (LAK) cells than were the enzymatically dispersed, monolayered cells in the 51Cr-release assay. The LAK cells were seen to migrate into the collagen gel and kill the hKEC cells. Thus, LAK cells may function to favor the acceleration of graft rejection.(ABSTRACT TRUNCATED AT 250 WORDS)

The distribution of high endothelial venules (HEVs) in various rat organs has been investigated immunohistochemically using two monoclonal antibodies (MoAbs), REC16-11 and REC4-1. The staining patterns observed with REC16-11 and REC4-1 were compared with those obtained with the MoAb anti-rat ICAM-1, which is a cell adhesion molecule in HEVs. REC16-11 reacted not only with HEVs but also with all vascular endothelial cells (ECs) in the tissues examined. Electronmicroscopy showed that REC16-11 and REC4-1 reacted with the cell surface antigens of ECs and immunoblot analysis of rat splenic stromal preparations showed that REC4-1 stained 42 kd and 60 kd bands. REC4-1 inhibited the binding of lymphocyte to HEVs in the mucosa-associated lymphoid tissues (MALT), but had no effects on lymphocyte binding to HEVs in peripheral (non-mucosal) lymph nodes. These findings suggested that the MoAb REC4-1 recognized the associated antigen of the lymphocyte-HEV recognition system in MALT.

The effects of ethanol (12.5-500 mM for up to 24 h) on mitochondrial structure including that of ATPase particles in cultured ventricular myocardial cells were studied using negative-stain electron microscopy. The activity of mitochondrial ATPase after ethanol treatment was also examined cytochemically and biochemically. At 5 min after the addition of all the concentrations of ethanol examined, some mitochondrial cristae were expanded and the arrangement of mitochondrial ATPase particles on these cristae was disordered. At and after 30 min the cristae decreased in number and some were expanded, vesiculated or fragmented. ATPase particles also decreased in number, particularly after the application of ethanol in concentrations of more than 50 mM. All the mitochondria had broadened and translucent cristae, and lacked ATPase particles with 200 and 500 mM ethanol at 24 h, although with 12.5 and 50 mM ethanol some mitochondria had similar negatively stained images but others had ATPase particles on broadened cristae. The enzymatic activity of the mitochondrial ATPase was unchanged with 200 and 500 mM ethanol at 24 h, compared with controls. The cytochemical technique also detected enzyme activity with all the concentrations of ethanol examined at 24 h. The discrepancy between the structural and biochemical alterations in mitochondrial ATPase induced by ethanol is discussed.

The adherence of human red blood cells (RBC) to autologous T-cells does not occur in the body, and in vitro is elicited at 4 degrees. Autologous E-rosetting at 37 degrees has not previously been described. In this work, lymphocyte-RBC adherence has been studied in mixed leukocyte-RBC cultures and in whole blood from healthy donors. Vital, cytochemical and electron microscopic studies have shown that T-cells may form stable E-rosettes with autologous RBC at 37 degrees. As in the previously reported cold-dependent reversible rosetting, stable rosetting is mediated by the erythrocyte LFA3 and lymphocyte CD2 molecules. Uniquely, this phenomenon requires both T-cell activation and an enhanced contact between the T-cell and RBC membranes. These requirements were met by exposure of cell cultures to: (1) PHAE, the erythroagglutinating component of PHAP, or (2) to either non-erythroagglutinating mitogens, PHAL, Con A, OKT3 or SEA, or to antigens of typhus group rickettsiae or salmonellae, provided that the RBC membrane was desialyted. Cultures derived from individuals seropositive to rickettsiae or vaccinated with salmonellae demonstrated the adherence phenomenon after antigen exposure when neuraminidase was present in the culture medium. The system 2 described here can be used as a diagnostic tool for defining activated T-cells and T-cell clones with the memory to antigens capable of inducing cell-mediated immunity.

Although calcification seldom occurs in pleomorphic adenoma, it often occurs in salivary glands, and so we decided to investigate the possible role of calcium in this difference. A histochemical method using glyoxal bis(2-hydroxyanil) demonstrated a small amount of calcium outlining lumina and separated cells of epithelial structures and associated with cells of myxoid and chondroid regions in pleomorphic adenoma, and a conspicuous amount in the acini of the associated salivary glands. A biochemical method using dry ashing demonstrated a significantly higher level of calcium in the glands than in pleomorphic adenoma. The results indicate that the calcium is mainly associated with secretory granules, which are scarce in pleomorphic adenoma, and with proteoglycan present intercellularly and in stromal regions of pleomorphic adenoma. The calcium in secretory granules is of possible importance in calcification in lumina and epithelium, and that bound to proteoglycan is possibly released following necrosis to be of importance in stromal calcification. However, the overall low level of calcium in pleomorphic adenoma is the likely explanation for the usual lack of calcification.

The expression of cerebral type aldolase C was investigated immunohistochemically in six varieties of neuroendocrine (n = 57) and six types of non-endocrine tumor (n = 76) using the avidin-biotin complex method. Aldolase C expression in the neuroendocrine tumors was also compared with those of chromogranin and gamma enolase. Aldolase C was detected in all the islet cell (7/7) and carcinoid tumors (10/10), thyroid medullary carcinomas (7/7), and pheochromocytomas (10/10), as well as in the majority of neuronal tumors (8/10) and bronchial small cell carcinomas (10/13). Chromogranin immunoreactivity was restricted to the tumors with abundant neuroendocrine granules. Gamma enolase positivity was generally similar to that of aldolase C, but there were some differences. Amongst the bronchial small cell carcinomas, three tumors negative for gamma enolase were positive for aldolase C, while another three tumors were positive for gamma enolase only. However all the small cell carcinomas were positive for at least one of these two enzymes. Aldolase C was detected in 28 (37%) of the 76 non-endocrine tumors and tended to be expressed preferentially in the differentiated portions of these tumors. Although aldolase C was expressed in many bronchial squamous cell carcinomas, the immunoreactivity was localized mainly in keratinizing foci and the less differentiated parts of these tumors expressed the enzyme only occasionally. Thus aldolase C, in conjunction with other neuroendocrine-associated markers, may be of value in identifying tumors of neuroendocrine type.

The ultrastructure of granular lymphocytes (GLs) in the peripheral blood of 12 patients with granular lymphocyte-proliferative disorders (GLPDs) was studied. Autophagolysosomes exhibiting so-called "lysosomophagy" were seen in the GLs of four of these patients but not in normal individuals. Because rod-shaped lysosomes beginning to enclose other lysosomes were seen, the autophagolysosomes exhibiting lysosomophagy in GLs were thought to be formed by the same wrapping mechanism. As far as we know, this is the first report of lysosomophagy in lymphocytes.

A total of 26 thymomas and thymic carcinomas were studied by immunohistochemistry to determine the presence and distribution of intratumoural B-cells. Double staining experiments revealed two distinct B-cell populations in the thymic epithelial tumours. One was found within the perivascular space (PVS), which is separated from the neoplastic epithelium by a basement membrane. In all tumours the PVS contained lymphocytes with the immunophenotype of peripheral B-cells. Large numbers of B-cells with germinal centre formation were found almost exclusively in myasthenia gravis (MG)-associated tumours, mainly in cortical thymomas and well differentiated thymic carcinomas. A second population of B-cells was located in the neoplastic epithelial meshwork, mostly in areas of organoid medullary differentiation characterized by epidermoid cells or Hassall's corpuscules. This population frequently comprised large, CD23+ cells with dendritic features resembling the special type of intramedullary B-cells of the normal human thymus. In contrast, B-cells were uncommon in areas of mixed thymoma showing spindle celled medullary differentiation, and were almost completely absent from tumour areas composed of cortical type epithelium. Hence a medullary microenvironment with epidermoid cells corresponding to Hassall's corpuscules seems to be necessary for specific intrathymic B-cell homing.

The cytoskeleton of alveolar type II cells on different matrices has been examined, and the bioelectric properties of these cells grown as monolayers in primary culture has be measured using Ussing-type chambers, to determine whether the extracellular matrix affects the cytoskeletal organization of alveolar type II cells and whether any such interactions influence their physiological functions. Alveolar type II cells cultured on a fibronectin substratum spread slowly over a 6-day period to produce cells of extremely large diameter. The cytoskeletal structure of these cells was characterized by a more marked accumulation of large bundles of actin and a finer network of keratin than cells grown on a collagen substratum. The transepithelial resistances of monolayers grown on a fibronectin substratum were much higher than those on a collagen substratum. These results indicate that alveolar type II cells cultured on fibronectin can form tighter, better organized and more polarized monolayers in primary culture, which suggests that fibronectin may have a physiologically important role in the maintenance of the alveolar wall.

We have established tumor cell lines from a rare juxtaglomerular cell tumor (JGCT) of the kidney. The original tumor contained both neoplastic cells and a tubular component. The tumor cell line (JX-G) maintained a capacity for renin production after the 20th passage. Cells at the fourth and 16th passage strongly reacted immunocytochemically with an antihuman renin antibody. Western blot analysis at the 9th passage revealed a positive reaction for renin at M(r) 45,000 and M(r) 62,000 which corresponded to renin precursors. However, the total amount of active renin secreted into the culture medium was high in the primary culture, but undetectable after the 4th passage. These findings indicate that the JGCT cell lines produced both active and inactive renin and the primary cultured JGCT cells secreted active renin, but that the secretion of renin had diminished after several passages. Additional stimuli thus appear necessary for maturation of renin and its subsequent secretion. These findings and the presence of a tubular component in the original tumor suggest that some signal transduction system is maintained in JGCTs. The physiological relationship of JGCT to the juxtaglomerular apparatus (JGA) and possible applications of the new cell lines are also discussed.