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Biomedical subjects

A M Shamsuddin

Publications and source records attributed to A M Shamsuddin.

At least 37 records · Page 2Linked to original sources
PubMed1992

Effects of inositol hexaphosphate on growth and differentiation in K-562 erythroleukemia cell line.

Inositol hexaphosphate (InsP6) has recently been shown to inhibit experimental cancers in vivo. Since the lower phosphorylated forms of InsP6 are important in cell growth in a wide variety of mammalian cells, we tested the efficacy of InsP6 in growth reduction of K-562 human erythroleukemia cells in vitro. We report that InsP6 decreases the K-562 cell population by 19-36% (P less than 0.001) concomitant to an increased differentiation as evidenced by ultrastructural morphology and increased hemoglobin synthesis. Pilot experiments to study the mechanism of action of InsP6 show that following treatment with InsP6, the concentration of intracellular [Ca2+] ([Ca2+]i) is increased by 57% (P less than 0.02). Likewise, a 41% increase (P less than 0.05) in InsP3 and a 26% decrease (P less than 0.02) in InsP2 were noted 1 h following treatment with InsP6. Contrary to the dogma that cell division is associated with increased [Ca2+]i, our data show that reduced cell growth and enhanced differentiation is associated with increased [Ca2+]i and increased InsP3 in the presence of InsP6.

Cell Differentiation↗
PubMed1992

Long-term culture of normal human colonic epithelial cells in vitro.

Studies of normal cellular function as well as the understanding of cellular mechanisms of carcinogenesis and other diseases of the large intestine have been limited, particularly due to the lack of long-term culture of normal human large intestinal epithelial cells (NHLIEC). Using the epithelia from surgically resected human colon, we have dissociated a sufficient number of viable NHLIEC and maintained them in in vitro culture for up to 5 months. Normal-appearing human large intestinal mucosal fragments (1 mm2) were treated with 0.01 mg/ml trypsin, 0.2 mg/ml collagenase + 0.1 mM EGTA or 0.1 mg/ml trypsin + 0.1 mM EGTA in a Stomacher laboratory blender to isolate the cells. Compared with other methods, the use of the Stomacher blender combined with low concentrations of proteolytic enzymes yielded greater numbers of cells per gram of tissue, with up to 84% viable cells. Primary and serially passaged NHLIEC were cultured in CMRL-1066, MEM with 5% serum, and serum-free KGM. These media were all supplemented with insulin, hydrocortisone, epithelial growth factor, and bovine pituitary extract. CMRL-1066 was found to be the best medium for NHLIEC. Contaminating fibroblasts were selectively removed by briefly allowing the cells to adhere to the culture vessel and adding 25 U/ml collagenase to the culture media at the first subculture treatment. The epithelial nature and secretory function of the established cells were confirmed by morphological criteria (light microscopy, phase contrast microscopy and electron microscopy), immunoreactivity to cytokeratin, and positive mucin cytochemistry. We propose that using this methodology for the culture and maintenance of NHLIEC for an extended period of time would serve as a valuable model for a variety of investigations.

Cell Division↗
PubMed1992

Detection of the tumor marker D-galactose-beta-(1-->3)-N-acetyl-D-galactosamine in colonic cancer and precancer.

The enzyme D-galactose oxidase (GO) oxidizes the carbon-6 position of the hydroxyl groups of galactose-N-acetyl galactosamine, which are commonly present in colon cancer cells and in rectal mucin of patients with colon cancer. We have studied the marker disaccharide galactose and N-acetylgalactosamine on tissue sections by the GO-Schiff reagent in normal, preneoplastic, and neoplastic human colorectal epithelial and compared it with peanut agglutinin reactivity. Fifty-seven (81.4%) of 70 carcinomas, 83.3% (10/12) of precancerous lesions, 50% (10/20) of the mucosa remote from cancer, and 58.1% (25/43) of the mucosa immediately adjacent to cancer showed a positive reaction with GO-Schiff, but the normal control mucosa was nonreactive. The GO-Schiff reagent showed an intense reactivity with mucinous adenocarcinomas and poorly differentiated adenocarcinomas. An intense reactivity was also seen in the intracellular mucus of abnormal dilated crypts (polyps, five of five cases; colitis, four of seven cases; and remote mucosa, 10 of 20 cases). Comparison of peanut agglutinin and GO-Schiff reactivity showed that the nonmucinous (glandular) adenocarcinomas less frequently reacted with the GO-Schiff sequence. Our results showed that the carbohydrate moiety detected by the two techniques may not necessarily be the same, warranting further biochemical analysis. Meanwhile, the data suggested that, like peanut agglutinin, the GO-Schiff sequence has the potential to identify the tumor marker either at the tissue level or by a mucin test for screening colorectal cancer or precancer.

Adenocarcinoma↗
PubMed1991

Stability of fecapentaene-12 and its carcinogenicity in F-344 rats.

Carcinogenicity studies of the fecal mutagen fecapentaene-12 (FP-12) have been hampered because of its apparent instability. We report here that: (i) contrary to the popular belief, FP-12 is quite stable, particularly at micromolar to nanomolar concentration; and (ii) its characteristic spectrophotometric absorbance spectrum is a function of the solvent or vehicle. Using synchronous fluorescence spectrophotometry (SFS), we have determined that at delta lambda 36.5 nm FP-12 gives a characteristic single emission peak between 413 and 423 nm, allowing us to identify FP-12 in DNA when reacted in vitro. We also report an increased incidence (statistically not significant) of fibrosarcomas and mammary carcinomas in male F-344 rats following intrarectal instillation of FP-12. In the in vitro human colon explant model, direct addition of FP-12 results in alteration in mucin histochemical changes typical of precancer and cancer. Our results support the contention that FP-12 is a naturally occurring carcinogen and may be responsible for human cancer(s).

Animals↗
PubMed1991

Long-term explant culture of human colon and a 3-step transformation model for rat colonic epithelium.

Our previous studies have suggested that colonic epithelium from rodents pretreated in vivo with suboptimal doses of carcinogen could be more easily maintained in explant culture. Transformation of colonic epithelium from these explants may be induced by subsequent exposure to additional genotropic agents. Therefore, we describe the development of a 3-step transformation model which uses (1) in vivo pretreatment with a suboptimal dose of 1,2-dimethylhydrazine (DMH) followed by (2) in vitro organ culture and exposure to xenotropic murine sarcoma virus (X-MSV), and finally (3) xenograft maintenance in nude mice to allow sufficient time for transformation. Male Wistar rats were injected intramuscularly with 20 mg DMH/kg 10 times per week followed by the removal and explant culture of the colon, which was then treated in vitro with X-MSV, and transplanted into nude mice after 1 week of culture. All the nude mice (n = 6) transplanted with rat colon explants contained viable xenograft explant epithelium and 1 of the 6 showed transformation. Our results demonstrate that the epithelium from animals pretreated with suboptimal doses of carcinogens can be easily transformed. We also demonstrate that human colonic epithelium is viable for an extended period of time in this model. Based on these results, we hypothesize that such a 3-step transformation model is applicable for carcinogenesis studies of various organs from different species, including human if one uses dysplastic or 'pre-neoplastic initiated' tissues obtained at surgery (e.g., ulcerative colitis; Barret's esophagus, etc.).

Animals↗
PubMed1990

Dose-dependent inhibition of large intestinal cancer by inositol hexaphosphate in F344 rats.

We have previously reported that inositol hexaphosphate (InsP6) inhibits mitosis and large intestinal cancer (LIC) in F344 rats and CD1 mice when given as 1 or 2% solution in drinking water at the unadjusted pH of 11.3. The purpose of this study was to determine whether InsP6 (i) shows a dose-response inhibition of LIC, and (ii) retains its anti-neoplastic effect at physiological pH. Since InsP6 is known to be a chelator of divalent cations, in preparation for putative clinical trials in humans, we also looked at the mineral bioavailability. F344 rats were fed 0.1% (pH 10.8), 1% (pH 11.3) and 1% (pH 7.4) Na-InsP6 in drinking water. Two weeks following the beginning of InsP6 supplementation, rats were given six injections of azoxymethane (AOM) at a dose of 8 mg/kg body wt/week and were killed 30 weeks following the last injection. Compared to the untreated control rats injected with AOM, 1% InsP6 (pH 11.3) reduces tumor prevalence by 52.2% (P less than 0.01), tumor frequency by 55.8% (P = 0.001) and tumor size by 62.3% (P = 0.001); 0.1% InsP6 showed a lesser reduction in tumor prevalence (21%) but a greater reduction in tumor size 71% (P = 0.001). While there was no significant difference in tumor prevalence and frequency between the two pH groups, the tumor size following 1% InsP6 (pH 7.4) was the smallest (65% smaller than those of pH 11.3, P less than 0.005). There was no significant difference in the serum Mg2+, Ca2+, Fe2+ and Zn2+ level between control rats and those treated with 1% InsP6. We therefore demonstrate that InsP6 (i) is consistently anti-neoplastic for LIC in a dose-dependent manner, (ii) retains its anti-neoplastic activity at physiological pH and (iii) has no demonstrable toxic effect on long-term administration as evident by body wt data and serum mineral levels.

Animals↗
PubMed1989

Inositol hexaphosphate inhibits large intestinal cancer in F344 rats 5 months after induction by azoxymethane.

A treatment regimen of 2% Na-InsP6 in drinking water was effective in significantly reducing large intestinal cancer in F344 rats even when the treatment was begun 5 months after carcinogenic induction with azoxymethane (AOM 8 mg/kg/wk X 6). Compared to untreated (AOM-only) rats, animals on InsP6 had 27% fewer tumors (P less than 0.02). The tumors were approximately two-thirds smaller in size (P less than 0.01) and percentage mitotic rate in the non-neoplastic epithelium was less than half (1.0 +/- 0.1, compared to 2.3 +/- 0.2 of AOM-only animals, significant at P less than 0.001). We postulate that InsP6 may exert its antineoplastic effect by way of regulating cellular proliferation even after effective carcinogenic stimuli and thus may be an important candidate for chemointervention.

Animals↗
PubMed1989

Inositol and inositol hexaphosphate suppress cell proliferation and tumor formation in CD-1 mice.

In previous studies, we have shown that inositol hexaphosphate (InsP6), a constituent of cereal diet, inhibited azoxymethane-induced experimental large intestinal cancer (LIC) in Fischer 344 rats. We now report a similar antineoplastic action of InsP6 in CD-1 mice injected with 1,2-dimethylhydrazine (DMH). We had hypothesized that InsP6 may bring about this effect by undergoing dephosphorylation to lower phosphorylated forms; the ready availability of Ins, to react with phosphates, may increase the total amount of the lower phosphorylated Ins and potentiate the action of InsP6. LIC induced by DMH (15 mg/kg/week x 13) in mice given a mixture of 1% InsP6 + 1% Ins show a significant reduction (P less than 0.005) in LIC prevalence over InsP6 treatment. Surprisingly, Ins, an in vitro growth promoting agent also caused a significant (P less than 0.001) suppression of LIC. InsP6 +/- Ins also showed a concomitant reduction in the mitotic rate in the non-neoplastic epithelium. Body weight data did not suggest any overt toxic effect of long-term administration of InsP6, Ins or InsP6 + Ins. Since InsP6 is antineoplastic in two species of experimental animals, it should, in combination with Ins, be considered in our strategies for prevention of large intestinal cancer.

1,2-Dimethylhydrazine↗
PubMed1989

Inositol-phosphate-induced enhancement of natural killer cell activity correlates with tumor suppression.

In recent studies, we have demonstrated that inositol hexaphosphate (InsP6) inhibits experimental colon carcinogenesis. Since natural killer (NK) cells are involved in tumor cell destruction, we investigated the effect of InsP6 on murine NK cell activity. We show that; (i) 1,2-dimethylhydrazine (DMH), a colon carcinogen, depresses NK activity; (ii) in vivo treatment of mice with InsP6 enhances baseline NK activity and reverses DMH-induced depressed NK activity with an inverse correlation (r = -0.9811) with tumor incidence, (iii) short-term in vitro treatment of spleen cells and NK-enriched fraction with InsP6 also enhances NK cytotoxicity in a dose-dependent manner, (iv) inositol potentiates the action of InsP6. Our data suggest yet another important role of inositol phosphates in the regulation of cellular activity.

1,2-Dimethylhydrazine↗
PubMed1988

Suppression of large intestinal cancer in F344 rats by inositol hexaphosphate.

Epidemiological data demonstrate correlations between dietary factors and the incidence of large intestinal cancer (LIC). Certain high-fiber diets are associated with a lower risk of LIC; these high-fiber diets are also rich in inositol hexaphosphate (IP6 or phytic acid). In a pilot study, we have used F344 rats to investigate the effect of sodium inositol hexaphosphate (Na-IP6) prior to (experiment I) and following injections of the carcinogen azoxymethane (AOM) (experiment II). In experiment I, rats started on 1% Na-IP6 in drinking water 1 week prior to the carcinogen treatment showed a 34.7% decrease (P less than 0.01) in LIC compared to control carcinogen treatment group. A similar reduction in the incidence of LIC was also observed in experiment II, wherein Na-IP6 supplementation was started 2 weeks following the last dose of the carcinogen. Comparison of the incidence of mitosis in the colonic crypts of the animals in different groups show that animals on AOM + IP6 demonstrate a significantly lower (P less than 0.001) mitotic rate than those receiving AOM only. Pilot studies of free radical generation demonstrate a reduction in .OH radical formation by Na-IP6. Further studies to expand this pilot data and to understand the mechanism of IP6 mediated LIC suppression are needed for it may have significance in our strategies for LIC control.

Animals↗
PubMed1987

Metastatic renal oncocytic neoplasm with benign histologic appearance.

We present a case of malignant renal oncocytoma which displayed a benign histologic appearance in the primary renal tumor and in metastases to the spine and liver. The case demonstrates the difficulty that may be encountered in distinguishing benign renal oncocytomas from malignant renal oncocytomas. Whereas, the accepted criteria will enable a distinction in the majority of instances, occasional instances may be encountered in which a benign histology does not provide an accurate reflection on the clinical course.

Adenoma↗
PubMed1987

Expression of carcinoembryonic antigen, T-antigen, and oncogene products as markers of neoplastic and preneoplastic colonic mucosa.

Several crypt abnormalities have been demonstrated in the mucosa of neoplastic and preneoplastic lesions of the large intestine. In addition, certain tumor markers are expressed in large intestinal carcinoma but not in normal mucosa. To determine whether any correlation exists between tumor marker expression and crypt abnormalities and at what stage markers are expressed, we studied specimens of large intestinal mucosa from 13 patients with preneoplastic conditions (adenomatous polyp, familial polyposis, Crohn's disease, and ulcerative colitis). The tumor markers examined include carcinoembryonic antigen (CEA), the ras gene products p21 and p21ser (mutated form), and beta-D-galactosyl-(1----3)-alpha-N-acetyl-D-galactosamine (gal--gal NAc, also known as T-antigen). Results were compared to those of five cases of adenocarcinoma of colon and three control cases of colonic mucosa obtained at immediate autopsy. All four markers were expressed in three of the five cases of adenocarcinoma, but none were expressed in the control cases. Variable expression of each marker was demonstrated in the dilated, distorted crypts of preneoplastic lesions. CEA and gal--gal NAc appeared to be expressed most frequently, suggesting that these are common markers or are expressed at an earlier stage in the neoplastic process than p21 or p21ser. Demonstration of such markers in preneoplastic conditions may be of use in determining the malignant potential and in monitoring these lesions.

Antigens, Neoplasm↗
PubMed1987

Localization of aflatoxin B1--nucleic acid adducts in mitochondria and nuclei.

The reaction of chemical carcinogens with cellular macromolecules, particularly DNA, is considered to be an important event during chemical carcinogenesis. Until recently, most studies have dealt primarily with nuclear DNA (nDNA). Mitochondrial DNA (mtDNA) is increasingly being investigated for its possible role during chemical carcinogenesis. Using monoclonal antibodies against aflatoxin B1 (AFB1)-modified guanosine and ultrastructural immunocytochemistry, we have localized the AFB1-guanosine adducts predominantly in mitochondria and nuclei. Morphometric analysis of the electron micrographs demonstrates that localization of the AFB1-guanosine adducts is several-fold greater in mitochondria than in the nuclei. However, biochemical analysis demonstrates that 77% of the covalent binding was detected in nucleic acids of the mitochondria relative to the nuclear fractions. In RNA-free preparations, the extent of covalent modification of mt circular DNA was less than 50% of that of nuclear DNA. Our results indicate that 67% of AFB1 binding to mitochondrial nucleic acids is associated primarily with mtRNA.

Aflatoxin B1↗
PubMed1987

Fecal mutagen fecapentaene-12 damages mammalian colon epithelial DNA.

Fecapentaenes are fecal mutagens that are naturally produced in the human colon and have been shown to be highly mutagenic in the Ames assay system. However to date no studies have been reported regarding the effects of fecapentaene in the target epithelium. In vivo studies with fecapentaene-12 (FP-12) using Fischer 344 rats in our laboratory indicate that a concentration of 10(-6) M FP-12 is capable of inducing a 2.7-fold increase (P less than 0.001) in [3H]-thymidine incorporation into DNA. Autoradiographic studies demonstrate a similar (2.6-fold) increase in the labelling index but an 8.8-fold reduction in the mitotic rate in colonic epithelial cells. Results of DNA single-strand breakage measurements show that in vivo treatment with FP-12 at concentrations of 1 microM introduces a 16-fold increase (P less than 0.001) in the number of alkali-labile sites over controls. Similar studies in in vitro assays indicate a linear trend in the number of alkali-labile sites over a range of concentrations varying from 1 nM to 1 microM. These findings indicate that the fecal mutagen FP-12 induces damage in situ to nucleic acids and thus may play a role in neoplastic transformation of the colon.

Animals↗
PubMed1986

Lewisx- and sialylated Lewisx-related antigen expression in human malignant and nonmalignant colonic tissues.

Biochemical studies have revealed that some normal cells express the LeX trisaccharide Gal beta 1----4(Fuc alpha 1----3)GlcNAc either on short-chain fucolipids or as a single immunodeterminant on glycolipid oligosaccharide side chains. Cancer cells, including those from colonic adenocarcinomas, express this antigen on longer type 2 blood group side chains as difucosylated or trifucosylated fucolipids. Moreover, sialylated forms of difucosylated LeX also accumulate in colon cancer but not in normal colonic mucosa. In the present study, six monoclonal antibodies which selectively recognize the various LeX-related antigens were used for immunohistochemical examination of these antigens in serial sections of human colonic tissue. All of these antigens were oncodevelopmental in human colon. Monoclonal antibodies anti-SSEA-1 and AH8-183, directed against short-chain, monofucosylated LeX, were unable to discriminate well between normal and malignant colonic tissue. However, the other four antibodies were much better at distinguishing cancer from normal tissue. FH6 was the most specific in that no normal tissues bound this antibody. However, FH6 failed to stain poorly differentiated cancers and some colloid-type carcinomas. FH4, which was also highly specific, stained almost all cancers, regardless of the degree of differentiation. FH4 primarily stained cancer cell cytoplasm, whereas the sialylated antigen defined by FH6 predominantly stained cell membranes. Differences were noted between the expression of LeX-related antigens in autopsied normal mucosa compared to mucosa of benign colonic diseases. Monoclonal antibodies recognizing long-chain polyfucosylated and sialylated LeX-related antigens appear to be useful tools for detection of colon cancer.

Antibodies, Monoclonal↗