Biomedical subjects

R Dahiya

Publications and source records attributed to R Dahiya.

At least 109 records · Page 6Linked to original sources

PubMed1993

Pelvic plexus denervation in rats causes morphologic and functional changes of the prostate.

The effect of pelvic plexus denervation on biochemical, morphologic and functional characteristics of the rat prostate was assessed in 11 adult Sprague-Dawley rats. Unilateral denervation was performed by removing the right pelvic ganglion from the surface of the prostate; the contralateral lobe was similarly dissected, but not denervated, to serve as a control. After 15 to 18 days, the prostate was excised and specimens from both sides were used for morphologic (light and electron microscopy) and biochemical studies (sodium-dodecylsulfate-polyacrylamide gel electrophoresis). With light microscopy, histologic features of the denervated prostate showed an overall decrease in cell height and a reduction of the clear apical area of the supranuclear region. At an ultrastructural level, denervated prostatic epithelial cells showed a slight, but significant, reduction in the number of secretory granules, a decrease in the height of the supranuclear region, and fewer and less abundantly dilated apical cisternae of endoplasmic reticulum. These changes indicate a modification of the secretory activity and reflect a change in epithelial metabolism, which was further supported by the results of SDS gel electrophoresis: the denervated prostate tissue showed an increase in the expression of 108 kDa and 80 kDa protein bands and a decrease in the expression of the 45 kDa protein band. This modification in the protein content is probably temporal and would undergo further evolution with time. The finding that denervation of the prostate causes significant morphologic and functional changes of the glandular epithelium strongly suggests that prostate function is not subject solely to the regulatory influence of the hormonal milieu. It is also dependent on the anatomical and, quite possibly functional, integrity of the nervous system.

Animals↗

PubMed1993

HLA expression by benign and malignant prostatic epithelium: augmentation by interferon-gamma.

Chronic prostatitis is a poorly understood entity that is characterized by lymphocytic infiltration of benign prostatic epithelium. Previously, we and others have shown that prostatic epithelium involved by prostatitis is phenotypically different from uninvolved epithelium. In addition, we have shown that malignant prostatic epithelium is rarely, if ever, infiltrated by lymphocytes. We now report that benign prostatic epithelium expresses HLA-DR only in the presence of lymphocytic inflammation, and that benign epithelium without chronic prostatitis and malignant prostatic epithelium do not express HLA-DR. In order to determine whether HLA-DR expression is inducible on malignant prostatic epithelium and therefore, at least theoretically, susceptible to immune regulation, we studied the DU-145 cell line in culture under various conditions. DU-145 cells did not express HLA-DR under routine culture conditions. However, the addition of interferon-gamma (100 to 6000 U/ml.) resulted in HLA-DR expression by DU-145 cells at 24 hours with maximal expression by 72 hours. In contrast, other cytokines (tumor necrosis factor, interleukin-1, interleukin-2) had no effect on HLA-DR expression. These investigations show that interferon-gamma induces HLA-DR expression on the DU-145 prostatic adenocarcinoma cell line, raising the theoretical possibility that malignant prostatic cells may be induced in vivo to express HLA-DR and thus become susceptible to immune regulation.

Adenocarcinoma↗

PubMed1993

Sodium nitroprusside and neurostimulation cause increased levels of cyclic guanosine monophosphate and not cyclic adenosine monophosphate during canine penile erection.

We investigated the changes of cGMP and cAMP during penile erection induced by direct electrical stimulation of the pelvic nerve or sodium nitroprusside administration. Immediately after penile erection, the samples were put into liquid nitrogen and cyclic nucleotide content was measured. The results of these investigation suggest that both pelvic nerve stimulation and sodium nitroprusside induced penile tumescence. The total content of cGMP and not cAMP were significantly higher in treated samples as compared to control. SDS-polyacrylamide gel electrophoresis revealed several differences after treatment compared to untreated samples. This study demonstrated that the total content of cGMP and not cAMP increased after pelvic nerve stimulation and sodium nitroprusside treatment in penile tissue.

Animals↗

PubMed1993

Phenotypic and cytogenetic characterization of a human corpus cavernosum cell line (DS-1).

We report here the establishment and characterization of a corpus cavernosum cell line (DS-1) from human penile tissue. This is the first cell line of its type derived from cavernosum tissue. DS-1 cells have become immortalized in culture, and show growth in monolayers. These cells have a doubling time of about 45 h in in vitro culture. Cytogenetic analysis by G-banding demonstrated a diploid karyotype with a model chromosome number of 46. The chromosome constitution of DS-1 cells was found to be male (XY), in 28/30 cells scored. Two of the 30 cells showed an extra structurally rearranged "marker" chromosome, that appeared to be a derivative of chromosome 18 with excessive chromosome on the short arm. Ploidy analysis revealed that the majority of DS-1 cells had a DNA index of one. About 35% cells were found to be in G-1 phase and 52% cells in S phase. Light and electron microscopy of DS-1 cells and original penile tissue showed typical characteristics of this tissue. Immunocytochemistry studies using antibodies to smooth muscle actin, desmin, vimentin and cytokeratin (LP34, CAM5.2) showed that the DS-1 cell line had predominantly smooth muscle cells, as these cells were positive for smooth muscle actin, desmin and vimentin.

Cell Division↗

PubMed1992

Mucin oligosaccharide biosynthesis in human colonic cancerous tissues and cell lines.

BACKGROUND: The Tn, sialosyl-Tn, and T antigens are carbohydrate-associated antigens that represent initial steps in mucin O-linked glycosylation. Previous immunohistochemical studies have shown that these three antigens are rarely, if ever, expressed in normal colonic mucosa; however, most colonic cancerous tissues express these structures. Little is known about the factors that control the expression of these antigens in colonic tissues or cell lines. One hypothesis is that cancers have increased levels of the glycosyltransferase activities responsible for synthesizing these antigens. METHODS: The current study analyzed antigen expression by immunohistochemistry and glycosyltransferase enzyme activities for Tn, sialosyl-Tn, and T antigens in colonic tissues and cell lines to (1) compare values between normal and cancerous tissues and (2) correlate these results with tumor stage, histologic findings, and location. RESULTS: All nine colonic cancer cell lines expressed Tn antigen; sialosyl-Tn and T antigens were expressed by the more mucin-producing cell lines. Sialosyl-Tn transferase activity was higher in the more mucinous cell lines; T transferase activity was higher in those with less mucin. In paired specimens of normal and cancerous tissues, levels of each of the three glycosyltransferases were similar. In cancerous tissues, enzyme activity did not correlate with tumor location, stage, or histologic type. There also was no correlation between glycosyltransferase activity and expression of the relevant antigen. CONCLUSIONS: Thus, because normal and malignant colonic tissues have comparable levels of Tn, sialosyl-Tn, and T glycosyltransferases, the absence of these antigens in normal mucosa apparently is related to other factors such as antigen masking.

Aged↗

PubMed1992

Expression and characterization of mucins associated with the resistance to methotrexate of human colonic adenocarcinoma cell line HT29.

A series of experiments were conducted to study synthesis and secretion of mucin in mucus-secreting subpopulations of HT29 human colonic adenocarcinoma cells selected by resistance to methotrexate (MTX). Mucin was quantitated by [3H]glucosamine labeling and chromatography on Sepharose CL-4B. The mucinous nature of the labeled high molecular weight glycoprotein was verified by alkaline borohydride treatment, cesium chloride density gradient ultracentrifugation, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results of these experiments demonstrated that MTX-treated cells have increased amounts of mucin in medium, cytosol, and membrane fractions. This was associated with the increase in the activities of polypeptidyl-N-acetylgalactosaminyltransferase and beta-1,3-galactosyltransferase compared to control cells. DEAE-Sephacel chromatography of [3H]glucosamine-labeled high molecular weight glycoproteins suggest that MTX-treated cells are less acidic compared to controls. Using complementary DNA probes for two distinct human intestinal mucins (MUC2 and MUC3) and one mammary mucin (MUC1), it was found that MTX-treated cells expressed more mucin messages compared to untreated cells. These results were consistent with immunoblots using anti-MRP (MUC2 repeat peptide), anti-M3P (MUC3 repeat peptide), 139H2 (MUC1 peptide), anti-T (peanut lectin), anti-Tn (91S8), and anti-sialosyl Tn (JT10e) antibodies. These data indicate that MTX-resistant HT29 cells show enhanced secretion and synthesis of mucin as well as expression of MUC1-, MUC2-, and MUC3-related mucin polypeptide epitopes.

Cell Differentiation↗

PubMed1992

Mucin synthesis and secretion in relation to spontaneous differentiation of colon cancer cells in vitro.

The synthesis and secretion of mucin-like high-molecular glycoprotein was studied in 2 human colon cancer cell lines that spontaneously differentiate in culture (Caco-2 and T84) and in 2 cell lines that do not spontaneously differentiate (LS174T and HT29). Mucin, quantitated by 3H-glucosamine labelling and chromatography on Sepharose CL-4B was found to be produced by all 4 cell lines. The mucinous nature of the labelled high-molecular glycoprotein was verified by enzymatic degradation treatments (heparinase, hyaluronidase, chondroitinase ABC, and N-glycanase), alkaline-borohydride treatment, inhibition of labelling by the glycosylation inhibitor benzyl-alpha-GalNAc, and by CsCl-density-gradient centrifugation. In all 4 cell lines, an inverse correlation of mucin synthesis with cell density was demonstrated. In Caco-2 cells, the spontaneous post-confluent enterocytic differentiation with increased brush-border enzyme expression was associated with a decrease in mucin synthesis and in the activities of polypeptidyl GalNAc transferase and beta 1,3-galactosyltransferase activity. Using cDNA probes for 2 distinct human intestinal mucins (MUC2 and MUC3), we found that all 4 colon cancer cell lines expressed mucin message, but the types of mucin mRNA expressed differed. These data indicate that mucin-like glycoproteins can be synthesized by cell lines derived from non-mucinous colon cancer, whether or not they undergo spontaneous differentiation in culture. These cell lines may serve as in vitro models for studying apomucin heterogeneity and control of mucin gene expression.

Cell Differentiation↗

PubMed1992

Establishment and characterization of a human primary prostatic adenocarcinoma cell line (ND-1).

A human primary prostatic adenocarcinoma cell line named ND-1 has been established in long term tissue culture. The cultured cells show growth in both monolayers and in soft agar suspension and produce subcutaneous tumors in nude mice. Cytogenetic analysis by G-banding demonstrated an aneuploid karyotype with a modal chromosome number of 62, and multiple marker chromosomes with 25-30% structural abnormalities. Ploidy analysis revealed that the majority of ND-1 cells (67%) had a DNA mass of 10.1 picogram and DNA index of 1.41. Nineteen percent of cells had a DNA mass of 21.3 picogram and DNA index of 3.0. Electron microscopic studies revealed common features of neoplastic epithelial cells such as numerous microvilli, junctional complexes, abnormal nuclei, nucleoli, and mitochondria. Due to the absence of a structurally normal Y chromosome, confirmation of the presence of a derived Y chromosome was achieved through the use of C-banding and through fluorescent in situ hybridization with a Y chromosome repeat probe. Tandem E PSA immunoenzymatic assay revealed that these ND-1 cells secrete small amounts of prostate specific antigen.

Adenocarcinoma↗

PubMed1992

Metastasis-associated alterations in phospholipids and fatty acids of human prostatic adenocarcinoma cell lines.

Metastatic variants of human prostatic adenocarcinoma cell lines (DU-145, LNCaP, and ND-1) were studied by using soft agar colony forming efficiency, nude mice tumorigenicity, in vitro invasion assay, and type IV collagenase assay. The DU-145 and ND-1 cell line showed higher metastatic potential than LNCaP. Lipids from DU-145, ND-1, and LNCaP cells were extracted and analyzed by thin-layer chromatography and gas-liquid chromatography. The major lipids were phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, fatty acids, and cholesterol. The sphingomyelin level was significantly higher in highly metastatic cells (DU-145 and ND-1) compared with the lower metastatic variant (LNCaP). The increase in the synthetic pathway and decrease in degradation pathway of sphingomyelin in microsomal fractions was sufficient to account for the measured increase in sphingomyelin in DU-145 cells compared with LNCaP cells. The major fatty acids of these lipids were palmitic (16:0), stearic (18:0), oelic (18:1), and arachidonic acid (20:4). The arachidonic acid level was significantly decreased in DU-145 and ND-1 compared with LNCaP cells. Electron microscopic studies showed no significant changes in the morphology of DU-145, ND-1, and LNCaP cells. The results of these investigations demonstrate for the first time that sphingomyelin and arachidonic acid contents are different in high and low metastatic variants of human prostatic adenocarcinoma cell lines.

Adenocarcinoma↗

PubMed1992

Downregulation of saturated fatty acid and upregulation of unsaturated fatty acid by 13-cis-retinoic acid in human prostate cancer cells.

Retinoids play a major role in regulation of epithelial cell growth and cellular differentiation, but their mechanism(s) of action are still unclear. In the present study, we examined the effects of 13-cis-retinoic acid (13-cis-RA) on cytotoxicity, growth properties, morphology, neutral lipids, phospholipids and fatty acids in cultured human prostate cancer cell lines. The results of these experiments suggest that 13-cis-RA (10 microM) inhibits the DNA synthesis and nude mice tumorigenicity by 2- to 3-fold, compared to control. Electron microscopy revealed more differentiated phenotypes after 13-cis-RA treatment. There was a significant increase in phosphatidylcholine and decrease in sphingomyelin in 13-cis-RA treated cells compared to control. The saturated fatty acids significantly decreased whereas unsaturated fatty acids were increased after 13-cis-RA treatment in prostate cancer cells. This study demonstrates for the first time that retinoic acid mediated downregulation of saturated fatty acids and upregulation of unsaturated fatty acid in human prostate cancer cells.

Animals↗

PubMed1992

Biochemical, cytogenetic, and morphological characteristics of human primary and metastatic prostate cancer cell lines.

Metastatic properties of human prostatic cancer cell lines (ND-1 and DU-145) were examined using various biochemical techniques. DU-145 cells had a higher metastatic potential than ND-1 cells. Cytogenetic analysis by G-banding demonstrated an aneuploid karyotype with considerable structural rearrangement. ND-1 cells had a modal chromosome number range lower than DU-145 cells (45-66, compared to 54-62). Ploidy analysis revealed that DU-145 cells showed hyperdiploidy with a greater amount of proliferation than the majority of ND-1 cells. Electron microscopic studies revealed little change in the cell morphology of either line. DU-145 cells had lower phosphatidyl choline levels and higher sphingomyelin levels than ND-1. DU-145 cells had much lower arachidonic acid levels than ND-1 cells. SDS-polyacrylamide gel electrophoresis revealed protein differences between the two cell lines. This study demonstrates for the first time that lipids, proteins and cytogenetic parameters differ in human primary and secondary prostate cancer cell lines.

Adenocarcinoma↗

PubMed1991

Differential mucin gene expression in human pancreatic and colon cancer cells.

The purpose of this study was to determine the quantity and nature of the mucins synthesized and secreted by four different pancreatic cancer cell lines. Well- to moderately-differentiated SW1990 and CAPAN-2 human pancreatic cancer cells were found to produce more high-Mr glycoprotein (HMG) than less-differentiated MIA PaCa-2 and PANC-1 cells. Most of the labelled HMG was secreted within 24 h. The results of chemical and enzymic degradation, ion-exchange chromatography and density-gradient centrifugation indicated that the HMG in SW1990 and CAPAN-2 cells has the properties expected for mucins, whereas much of the HMG in MIA PaCa-2 and PANC-1 cells may not be mucin, but proteoglycan. These results are consistent with immunoblots and Northern blots showing the presence of apomucin and apomucin mRNA in SW1990 and CAPAN-2 cells, but not in MIA PaCa-2 and PANC-1 cells. The Western blots and Northern blots also show that SW1990 and CAPAN-2 cells, like breast cancer cells, have the mammary-type apomucin and mRNA coded by the MUC1 gene, but lack the intestinal type apomucin and mRNA coded by the MUC2 gene. In contrast, the colon cancer cell lines tested in culture express apomucin and mRNA coded by MUC2 but not by MUC1.

Adenocarcinoma↗

PubMed1991

Mucin production by human colonic carcinoma cells correlates with their metastatic potential in animal models of colon cancer metastasis.

Patients with mucinous colorectal cancers characteristically present with advanced disease, however, the relationship between mucin production by colon cancer cells and their metastatic potential remains unclear. We therefore sought to define the relationship between mucin production by human colon cancer cells and metastatic ability by employing animal models of colon cancer metastasis. LS LiM 6, a colon carcinoma cell line with high liver metastasizing ability during cecal growth in nude mice produced twofold more metabolically labeled intracellular mucin and secreted four- to fivefold more mucin into the culture medium compared to poorly metastatic parental line LS174T. This was accompanied by a similar elevation in poly(A)+ RNA detected by blot hybridization with a human intestinal mucin cDNA probe, and increases in mucin core carbohydrate antigens determined immunohistochemically. Variants of LS174T selected for high (HM 7) or low (LM 12) mucin synthesizing capacity also yielded metastases after cecal growth and colonized the liver after splenic-portal injection in proportion to their ability to produce mucin. Inhibition of mucin glycosylation by the arylglycoside benzyl-alpha-N-acetyl-galactosamine greatly reduced liver colonization after splenic-portal injection of the tumor cells. These data suggest that mucin production by human colon cancer cells correlates with their metastatic potential and affects their ability to colonize the liver in experimental model systems.

Animals↗

PubMed1991

Alterations in sphingomyelin and fatty acids in human benign prostatic hyperplasia and prostatic cancer.

The present experiments were conducted to examine and characterize the lipid composition of benign prostatic hyperplasia (BPH) and prostatic cancer tissues (CAP). Lipids were extracted from these tissues and analyzed by thin layer chromatography (TLC) and gas liquid chromatography (GLC). The protein profiles of these tissues were analyzed by SDS-polyacrylamide gel electrophoresis. The results of these studies demonstrate that the principal lipids of BPH and CAP tissues are phosphatidyl choline, phosphatidyl ethanolamine, sphingomyelin, fatty acids and cholesterol. The sphingomyelin level is significantly higher in CAP tissues compared to BPH tissues. The differences in the enzymatic activities responsible for the biosynthesis and degradation of sphingomyelin appear to explain at least partially the alteration in sphingomyelin level between BPH and CAP tissues. The major fatty acids of phosphatidyl choline and phosphatidyl ethanolamine were palmitic (16:0), stearic (18:0), oleic (18:1) and arachidonic (20:4) acid. The arachidonic acid level was significantly decreased in CAP tissues compared to BPH tissues. SDS polyacrylamide gel electrophoresis revealed several protein differences between BPH and CAP tissues. The most significant difference was the decrease in 12 kDa protein in CAP tissues compared to BPH. The present data, therefore, suggests that there are significant alterations in sphingomyelin, fatty acids and protein profiles between BPH and CAP tissues.

Electrophoresis, Polyacrylamide Gel↗

PubMed1990

Cell surface sialoprotein alterations in metastatic murine colon cancer cell lines selected in an animal model for colon cancer metastasis.

Alterations in cell surface proteins and glycoproteins may play a key role in determining the metastatic behavior of tumor cells. The cell surface proteins of a series of related murine colon cancer cells selected in an animal model for colon cancer metastasis (R. S. Bresalier et al., Cancer Res., 47: 1398-1406, 1987) were therefore compared by a variety of biochemical methods. Lactoperoxidase-catalyzed iodination of cell surface proteins followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated quantitative and qualitative differences in the cell surface protein profiles of parental cell line 51B (low metastatic potential) and its metastatic derivatives 51B LiM 5 and 51B LiM 6. Labeling of sialic acid-containing proteins suggested that, in the case of at least four of these proteins (Mr 170,000, 120,000, 95,000, and 55,000), this represented an increase in radioactive labeling of sialoglycoproteins from the metastatic lines. Affinity chromatography of solubilized 125I-labeled cell membrane proteins revealed a 2- to 3-fold increase in wheat germ agglutinin and Sambucus nigra lectin binding associated with the metastatic lines, compared to the poorly metastatic parent. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of material eluted from these columns demonstrated enhancement of proteins from the metastatic cells corresponding in molecular weight to the previously identified major sialoglycoproteins. Neuraminidase-releasable membrane-associated sialic acid and sialyltransferase activities were 2- to 3-fold higher in the metastatic cell lines compared to the parental line. Liver colonization after intrasplenic injection of the various lines into syngeneic mice was dramatically reduced by prior removal of cell surface sialic acid. Immunohistochemical staining of primary and metastatic tumors formed after cecal injection of parental 51B suggested selective metastasis by wheat germ agglutinin-binding tumor cells. These results further support the concept that cell membrane sialylation is important in determining the metastatic potential of cancer cells.

Animals↗

PubMed1990

Blood group antigen synthesis and degradation in normal and cancerous colonic tissues.

ABH antigens are expressed by colonic epithelial cells throughout the colon during fetal life but only in proximal segments during adulthood. Malignant and premalignant colonic tumors frequently exhibit ABH reappearance (distal lesions) or ABH deletion (proximal lesions) and occasionally express incompatible A or B substances. Mechanisms governing these developmental and cancer-associated alterations are unknown. Therefore, experiments were performed to assess the activities of biosynthetic (glycosyltransferase) and degradative (glycosidase) enzymes in normal and cancerous tissues of the proximal and distal colon. In normal colonic mucosa, A, B, and H transferase activities were similar in proximal and distal segments. Analysis of enzyme substrate affinities and product characterization confirmed that the ABH transferases in colonic tissues were similar to the gene-specified transferases in human serum. Glycosidase enzyme activities were also comparable in proximal and distal normal colon. Cancers had lower A and B transferase but similar H transferase activities compared with paired normal mucosa. Thus, the absence of ABH antigen expression in normal distal colon is not caused by insufficient glycosyltransferase activity or excessive glycosidase activity.

ABO Blood-Group System↗

PubMed1990

Effect of testosterone on the glycosphingolipid composition of the rat kidney.

A series of experiments was conducted to examine the effect of subcutaneous administration of synthetic testosterone (500 micrograms/100 g body weight) on the acidic and neutral glycolipids. The kidney glycosphingolipids were separated by using Sep-Pak cartridge column. Florisil column, and high performance thin layer chromatography, and analyzed by gas liquid chromatography. Testosterone administration increased the total content of ceramide, acidic and neutral glycolipids. The increase in total neutral glycolipids was mainly due to an increase in glucosyl- and globotetraosylceramide. The increase in total gangliosides was due to the increase in GM3 (hematoside) and "others". The relative percentage of N-glycolylneuraminic acid in GM3 was increased whereas the relative percentage of sphingoside and phytosphingosine were increased and decreased, respectively, in GM3, glucosyl- and globotetraosylceramide. The relative percentage and chain length of hydroxy and nonhydroxy fatty acids of major glycosphingolipids were altered in this organ.

Animals↗

PubMed1990

Inhibition of lipoprotein lipase activity by high density lipoprotein & apolipoproteins C, E & A in rhesus monkeys.

Experiments were conducted to study the effects of high density lipoprotein (HDL) and apolipoproteins C, E, and A on lipoprotein lipase activity in rhesus monkeys. The lipoprotein lipase activity was inhibited up to 32 +/- 6 per cent by monkey HDL. This inhibition was considerably decreased (2 +/- 0.02%) by using apolipoprotein-poor HDL. Apolipoproteins C and E inhibited the hydrolysis of activated intralipid by monkey lipoprotein lipase to a maximum of 83 +/- 7 and 57 +/- 5 per cent respectively. Apolipoprotein A produced little inhibition of lipoprotein lipase activity. The results of these studies demonstrate that HDL and apolipoproteins compete with the substrate for the binding to lipoprotein lipase in rhesus monkeys.

Animals↗