PubMed Health⌕ Search

Biomedical subjects

B Siddiqui

Publications and source records attributed to B Siddiqui.

At least 19 recordsLinked to original sources
PubMed1990

Expression of native and deglycosylated colon cancer mucin antigens in normal and malignant epithelial tissues.

Immunohistochemical techniques were used to determine the distribution and cellular location of the mature and precursor forms of a colonic-type mucin in normal and malignant epithelial tissues. The antisera used in this study were prepared against native human colon cancer mucin (LS), partially deglycosylated mucin (HFA or GalNAc-apomucin), and fully deglycosylated mucin (HFB or apomucin). These antisera reacted with most mucin-producing cells of the normal gastrointestinal tract, salivary ductular cells, bronchial epithelial cells, some bronchial mucous glands, and squamous epithelial cells of the esophagus. Breast, endometrium, ovary, prostate, liver, and thyroid were nonreactive. In most normal organs, HFB reactivity was present in the supranuclear and perinuclear cytoplasm and LS and HFA were located primarily in goblet cell vacuoles, apical cytoplasm, and luminal secretions. These findings are consistent with the expected subcellular locations of apomucin and more "mature" mucins. LS, HFA, and HFB were frequently expressed in adenocarcinomas of the colon, stomach, pancreas, and lung. Lymphoma, sarcoma, and melanoma specimens were nonreactive. Alterations in the expression of these mucin antigens in malignant tissues included loss of subcellular compartmentalization, increased intensity of staining, and disappearance of staining. In addition, de novo expression of HFB was observed in one of five breast carcinomas and three of five ovarian mucinous cystadenocarcinomas. These data demonstrate that LS, HFA, and HFB are useful for studying the organ specificities and biosynthetic pathways of one type of mucin in normal and malignant tissues.

Adenocarcinoma↗
PubMed1989

Deglycosylation of mucin from LS174T colon cancer cells by hydrogen fluoride treatment.

Mucin from xenografts of LS174T human colon cancer cells was treated with anhydrous HF for 1 h at 0 degree C to give a product (HFA) with over 80% of the glucosamine and hexose removed, but retaining some galactosamine, and for 3 h at room temperature to give a product (HFB) devoid of carbohydrate. Rabbit antibodies against HFA bound to HFA much more than to HFB, and bound to native mucin to an intermediate extent. Antibodies to HFB bound to HFB more than to HFA, and did not bind to native mucin. Both HFA and native mucin bound a number of lectins, but HFB did not. By SDS/polyacrylamide-gel electrophoresis and size-exclusion h.p.l.c., native mucin and HFA are of apparent molecular mass greater than 400 kDa, whereas HFB is heterogeneous and of low molecular mass. On Western blots, antibody to HFA detected both high-molecular-mass mucin and a 90 kDa protein in homogenates of LS174T cells. Antibody to HFB detected a major 70 kDa band as well as higher-molecular-mass species. In tissue sections of normal colon and colon cancers, antibody to HFA showed both cytoplasmic and extracellular staining, whereas antibody to HFB generally stained only cytoplasmic antigens. These results indicate that anti-HFB antibody is specific for apo-mucin, whereas anti-HFA antibody is specific for GalNAc-apo-mucin.

Animals↗
PubMed1989

Glycoproteins and glycolipids of rat small intestinal microvillus and basolateral membranes.

Glycoprotein and glycolipid constituents were examined in purified microvillus and basolateral membranes isolated from rat small intestinal epithelial cells. SDS-polyacrylamide gel electrophoresis showed that the molecular weights of most of the major proteins from microvillus membranes were over 100 kD, whereas the majority of those from basolateral membranes tended to have lower molecular weights. Glycoprotein profiles were also examined using three labeling methods, and in each case marked differences were observed between microvillus and basolateral membranes. In both membranes, lectins with a specificity toward N-linked sugar chains bound to the majority of the glycoproteins, in contrast to those lectins which preferentially bind to O-linked sugar chains. Glycolipids were labeled in vivo and isolated from both membrane fractions. Some differences were observed in the fucolipids and neutral glycolipids suggesting a more complex pattern in microvillus membranes. These results indicate that there are differences in the glycoprotein and glycolipid compositions of microvillus and basolateral membranes that may reflect the functional polarity of intestinal epithelial cells.

Animals↗
PubMed1989

Effect of sodium butyrate, a differentiating agent, on cell surface glycoconjugates of a human pancreatic cell line.

The effects of sodium butyrate, a differentiating agent, on growth properties and glycoconjugates of a human pancreatic carcinoma cell line (CAPAN-1) were studied. Butyrate caused marked changes of in vitro growth properties including prolongation of doubling time and greatly reduced colony forming efficiency in soft agar. Cell surface labeling revealed significant alterations in proteins and glycoproteins after treatment with sodium butyrate (appearance of glycoproteins of molecular weight 250,000, 220,000, and 70,000; reduction of glycoproteins of 164,000, 148,000, 110,000, and 66,800 sizes; increases in proteins/glycoproteins of 85,000 and 78,000). Metabolic labeling of the cells with [3H]fucose or [3H]galactose revealed that sodium butyrate treatment caused a marked reduction in the fucose-containing neutral glycolipids with six or more carbohydrate side chains and an increase in [3H]galactose-labeled neutral glycolipids, particularly GL-3a, GL-4a, and GL-5a. There was marked reduction in the labeling of a ganglioside with mobility similar to that of GM4 and of sulfogalactosylceramide. An increase in a ganglioside with mobility above GM1 was also caused by butyrate. These data indicate that sodium butyrate may be useful in the identification of differentiation or malignancy-associated glycoconjugate markers of human pancreatic cells.

Butyrates↗
PubMed1989

Comparison of metabolically labeled mucins of LS174T human colon cancer cells in tissue culture and xenograft.

Colon cancer cells in culture synthesize and secrete mucin glycoproteins, which carry a number of cancer-associated antigens. However, the structures and mechanisms of biosynthetic processing are not well understood. Mucins synthesized and secreted by LS174T human colon cancer cells were compared to those in LS174T xenografts in athymic mice. Mucins radiolabeled with glucosamine or sulfate were purified by gel filtration and cesium chloride density gradient centrifugation. The mucins were of high molecular weight and were resistant to chondroitinase ABC, hyaluronidase and HNO2 treatment. They were, however, susceptible to pronase digestion and mild alkaline treatment. Using radiochemical precursors, the cellular mucin was shown to contain fucose, galactose, N-acetylgalactosamine, N-acetylglucosamine, N-acetylneuraminic acid, and sulfate. Oligosaccharides released by beta-elimination had N-acetylgalactosaminitol as the reduced amino sugar and also unreduced galactosamine, indicating that there is N-acetyl-galactosamine O-glycosidically attached to protein core and also peripheral N-acetyl-galactosamine not directly linked to protein. DEAE-cellulose chromatography of mucins showed two major peaks with both intracellular and secreted mucins, but xenograft mucins also had more acidic components. Sulfate-labeled mucins were shifted to less acidic peaks by neuraminidase digestion, which indicates that the same mucin molecules are both sialylated and sulfated. We conclude that the intracellular mucins of cultured colon cancer cells, those secreted into the medium, and those in nude mouse xenografts are chemically similar, but differ in sialic acid and sulfate content. This experimental model system, LS174T cells maintained in culture and as nude mouse xenografts, may be useful for further biosynthetic and structural studies of colon cancer mucin.

Animals↗
PubMed1988

Isolation and characterization of colon cancer mucin from xenografts of LS174T cells.

The structure of colonic mucin, which is thought to be important in several diseases, including ulcerative colitis and colon cancer, is poorly understood. Mucin was isolated from nude mouse xenografts of the LS174T colonic adenocarcinoma cell line by gel filtration and CsCl density gradient centrifugation. The isolated mucin had a high content of threonine, serine, and proline, with 28% of the total amino acids O-glycosylated. The carbohydrates present were fucose, sialic acid, galactose, N-acetyl-glucosamine, and N-acetyl-galactosamine in the ratio of 0.4:1.5:1.0:0.9:1.4. Rabbit antibodies were prepared that recognized primarily protein-dependent determinants. By DEAE-cellulose chromatography, the purified mucin was found to be heterogeneous, with three major components that had small differences in carbohydrate composition. LS174T was antigenically and chromatographically similar to mucins in colon cancer tissue specimens and in nonmalignant colonic mucosae.

Adenocarcinoma↗
PubMed1986

Cancer-associated colonic mucin in cultured human tumor cells and athymic (nude) mouse xenografts.

Mucins derived from colonic cancers differ immunologically and chemically from those in normal colonic epithelium. It has been demonstrated that the lectin from the peanut will bind to mucins present in colonic cancers and other neoplastic lesions but not to those from the normal colon. It was hypothesized, therefore, that in transformed colonic epithelium the glycosylation of mucins occurs differently than in normal epithelium. To rule out the possibility that the differences in oligosaccharide structure were due to postsecretory degradation, studies were designed to evaluate cancer-associated colonic mucins produced under more controlled conditions. We studied nine different cancer cell lines first in monolayer culture and then as xenografts in athymic or nude mice. Eight of the nine cell lines in monolayer culture synthesized glycoconjugates that were labeled by fluorescein-conjugated lectins. After injection into nude mice, eight of the nine cell lines produced tumors typical of human colonic cancer, and six of nine secreted mucin. The mucins produced by the xenografts were labeled at fluorescence microscopy by peanut lectin and other lectins, characteristic of what had been seen in other primary human colonic cancers. One cell line, LS174T, produced large amounts of mucin in the xenograft model. Mucin was purified from these tumors and characterized biochemically. It was demonstrated that mucin purified from the xenografts bound peanut lectin. Therefore, we have concluded that cancer-associated mucins are present in cultured colorectal tumor cells. The cancer-associated mucins are also found in nude mouse xenografts, indicating that they are not the result of postsecretory degradation by colonic flora or by tumor cell necrosis. The cell culture and xenograft can therefore be useful for studying the biosynthesis of cancer-associated mucins.

Animals↗
PubMed1984

Differential expression of ganglioside GD3 by human leukocytes and leukemia cells.

Gangliosides from normal leukocytes and the cells of 25 patients with acute and chronic leukemia were tested for the presence of the disialoganglioside II3-alpha-N-acetylneuraminosyl-alpha 2----8-N-acetylneuraminosyllactosylceramide (GD3). GD3 was detected by immunostaining thin-layer chromatographs with an anti-GD3 monoclonal antibody (AbR24). Among the myeloid cells tested, acute leukemia cells were positive for GD3, whereas chronic leukemia cells and normal neutrophils did not have detectable GD3. A range of GD3 reactivity was apparent within the acute myeloid leukemia cells; gangliosides from pure myeloid leukemia cells stained more intensely than those from leukemia cells with monocytic characteristics. All lymphocytic leukemia cells (chronic and acute) contained GD3, but this ganglioside could not be detected in extracts from normal lymphocytes. A ganglioside extract from the cells of a patient with hairy cell leukemia was also positive for GD3 immunostaining. These results demonstrate that normal leukocytes and chronic myelogenous leukemia cells are distinguished from other lymphoid and nonlymphoid leukemia cells on the basis of GD3 ganglioside expression.

Acute Disease↗
PubMed1984

Effects of sodium butyrate, dimethyl sulfoxide, and retinoic acid on glycolipids of human rectal adenocarcinoma cells.

In the present study, we have examined the neutral glycolipids, gangliosides, and sulfoglycolipids of human rectal adenocarcinoma (HRT-18) cells and the alterations produced by the differentiating agents, sodium butyrate, dimethyl sulfoxide, and retinoic acid. Thin-layer chromatography of neutral glycolipids showed that HRT-18 cells contained mono- and diglycosylceramides. Cells treated with differentiating agents had additional glycolipids which comigrated with tri- and tetraglycosylceramides. Labeling of neutral glycolipids with [3H]galactose showed that HRT-18 cells also contain glycosylceramides larger than diglycosylceramides which also were altered by treatment with differentiating agents. In studies of acidic glycolipids, GM3, the major ganglioside in untreated cells, was reduced in cells treated with retinoic acid but not in cells treated with other agents. Upon labeling with radioactive galactose, changes were seen only in the minor ganglioside components of treated cells. Differentiating agents also altered the patterns of sulfogalactolipids and fucolipids in HRT-18 cells. In the case of fucolipids, 2 new bands were observed in the cells treated with dimethyl sulfoxide and retinoic acid. The changes brought about by differentiating agents may identify glycolipids involved in the process of tumorigenicity.

Adenocarcinoma↗
PubMed1982

Membrane-associated, fucose-containing glycoproteins and glycolipids of cultured epithelial cells from human colonic adenocarcinoma and fetal intestine.

Fucose-containing glycoproteins and glycolipids were compared in three human colon cancer cell lines and five human fetal intestinal epithelial cell lines. Cells were labeled by culturing cells in the presence of L-[3H]-fucose. Fucose was incorporated into both the membrane and cytoplasmic fractions of all three colon cancer cell lines to a much lesser extent than into fetal cells. When the relative fucose labeling of glycolipids and glycoproteins were examined, a much greater proportion of fucose labeling in the membrane was associated with lipid in colon cancer cells (11.6-16.7%) compared to fetal intestinal cells (1.3-2.5%). Fluorographic analysis of SDS-polyacrylamide gel electrophoresis of fucose-labeled glycoproteins revealed a rather uniform labeling pattern of fetal intestinal cells which was distinct from those of colon cancer cells. Thin-layer chromatographic analysis of fucose labeled glycolipids of all three colon cancer cell lines indicated the presence of fucose-containing glycolipids with carbohydrate chain lengths greater than five sugars. Glycolipids of the SKCO-1 cells in particular appear to consist predominantly of complex fucose-containing glycolipids. These results indicate that significant qualitative differences in the fucose-containing glycoproteins and glycolipids exist between the membranes of human colon cancer cells and fetal intestinal cells and that complex fuco-glycolipids with long carbohydrate side chains are present in the three human colon cancer cell lines.

Adenocarcinoma↗
PubMed1980

Effects of sodium butyrate and dimethylsulfoxide on biochemical properties of human colon cancer cells.

Sodium butyrate and dimethylsulfoxide (DMSO) have marked effects on the growth, morphology, and biochemistry of two human colonic adenocarcinoma cell lines in culture. Doubling times were increased between 18% and 660% while cell viability was unaffected. Both cell lines formed colonies in soft agar in the absence of butyrate of DMSO, but no colonies were observed in the presence of these agents. However, no differences in in vivo tumorigenicities, when cells were implanted in athymic mice, were seen following treatment. Gross morphological alterations including cell enlargement, process formation, and cellular flattening occurred during culture in butyrate or DMSO. Acrylamide gel electrophoresis in sodium dodecyl sulfate revealed no change in membrane protein constituents, but autoradiographic analysis of membrane glycoproteins demonstrated differences between treated and untreated cells. Ganglioside compositions were altered, and a sialyltransferase required for the synthesis of GM3 ganglioside was elevated by butyrate. Although cytoplasmic aminooligopeptidase remained unaffected by butyrate or DMSO, brush border-associated activity was enhanced by butyrate. Alkaline phosphatase also rose dramiatically during culture in butyrate but was not enhanced by DMSO.

Adenocarcinoma↗
PubMed1972

Structures of ceramide tetrasaccharides from various sources: uniqueness of rat kidney ceramide tetrasaccharide.

Methylation analysis of ceramide tetrasaccharide isolated from human erythrocytes gave acetates of 2,3,6-tri-O-methylgalactitol and 2,4,6-tri-O-methylgalactitol in a ratio of 1:1, and about 50% of the galactose was oxidized by periodate. Rat kidney ceramide tetrasaccharide gave, in contrast, a much larger proportion of the acetates of 2,4,6-tri-O-methylgalactitol (ratio 1:0.3), and less than 20% of the galactose was oxidized by periodate. Sequential degradation by beta-N-acetylhexosaminidase, alpha-galactosidase, and beta-galactosidase showed ceramide tetrasaccharides to have identical carbohydrate sequences and anomeric structures. The major part of ceramide trihexoside derived from rat kidney ceramide tetrasaccharide migrated on thin-layer chromatography more slowly than that derived from other ceramide tetrasaccharides. The structure of a major part of rat kidney ceramide tetrasaccharide was thus determined to be GalNAcbeta(1-->3)Galalpha(1-->3)Galbeta(1-->4)Glcbeta(1-->1)Cer, whereas other ceramide tetrasaccharides have Galalpha(1-->4) structure at the penultimate residue.

Animals↗