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S Rajaraman

Publications and source records attributed to S Rajaraman.

At least 55 records · Page 3Linked to original sources
PubMed1996

Detection of mitochondrial genome depletion by a novel cDNA in renal cell carcinoma.

A cDNA isolated by a subtractive hybridization procedure detected loss of mtDNA and the mRNA coding for NADH dehydrogenase subunit 3 in 8 of 13 tumor kidney tissues obtained from patients with renal cell carcinoma. Sequencing revealed a stretch of nucleotides homologous to the mitochondrial NADH dehydrogenase subunit 3 gene in the middle of the cDNA. The depletion phenomenon was also observed in five of six renal carcinoma cell lines. In the case of a benign renal oncocytoma, however, the mtDNA content was increased 200% more than that of the adjacent normal tissue. The frequency with which this phenomenon occurs in renal cell carcinomas, but not in other types of cancers, suggests that this may be an important phenotype associated with renal cell neoplastic transformation. However, the absence of any structural alterations within the mitochondrial genome suggests that the depletion may be a secondary event associated with the oncogenic transformation process.

Adult↗
PubMed1995

Nerve growth factor as a mitogen for a pancreatic carcinoid cell line.

Carcinoid tumors are a group of neuroendocrine neoplasms distributed widely throughout the body but most commonly occurring in the gut. These tumors retain many characteristics of their neural crest origin, including secretion of neuroactive peptides and responsiveness to neurotrophic substances. Nerve growth factor (NGF), a neurotrophic protein involved in maintenance and differentiation of peripheral sympathetic and sensory neurons, regulates growth of several neural tumor cells by inducing a differentiated phenotype and subsequent inhibition of cell growth rate. We examined the actions of NGF in a functioning human pancreatic carcinoid cell line (termed BON). NGF has no effect on the cytoarchitecture or constitutive secretion of bioamines in this carcinoid cell line. NGF, however, stimulates the in vitro cellular proliferation of BON cells. BON cells possess mRNA for the NGF receptors (p75LNGFR and p140trkA) and membrane-associated tyrosine kinase activity is increased in response to NGF. Both the mitogenic activity of NGF, as well as the receptor-linked tyrosine kinase activity, can be abrogated in BON cells by the trkA inhibitor K-252a and specific anti-NGF antibody. Our studies demonstrate that NGF is a mitogen for this carcinoid cell line without effect on cellular phenotype or cytoarchitecture. NGF may play a role in the development and progression of human carcinoid tumors.

Carbazoles↗
PubMed1995

Induction of apoptotic nuclei by interferon-gamma and by predesquamin in cultured keratinocytes.

Predesquamin is a glycoprotein found in the transition layer and the lower stratum corneum of human epidermis. Interferon-gamma (IFN-gamma) induces the synthesis of predesquamin by keratinocytes in culture. We now show ultrastructurally that exogenous addition of either predesquamin or IFN-gamma to cultured keratinocytes induces apoptotic nuclei with condensed chromatin. Degradation of cellular DNA is also evident as a ladder pattern in an agarose gel. After incubation with both predesquamin and IFN-gamma (but not either alone), the mobility of plasmid DNA in a gel shows retardation specific for guanine residues. This binding to the DNA may impart to it a conformational change that facilitates access by endogenous cellular nucleases. In epidermal cells cultured with IFN-gamma supplementation, we also show by RT-PCR that there is an upregulation of the genes c-myc, p53, gadd45, dsRNA-activated protein kinase, and 2'-5'-oligo(A)-dependent RNase, which have all been implicated in apoptosis in other cell types. These results are pertinent to the mechanism of occurrence of apoptosis in the epidermis in vivo, where predesquamin and IFN-gamma are endogenous. Programmed cell death is an inherent step in the terminal differentiation and desquamation of the epidermis.

Apoptosis↗
PubMed1995

Effect of ileo-jejunal transposition on the growth of the GI tract and pancreas in young and aged rats.

BACKGROUND: Ileo-jejunal transposition (IJT; transposition of the distal quarter of the small intestine into the proximal jejunum) is known to stimulate mucosal growth of the transposed ileum, but the effects on other parts of the small intestine are controversial. The effect of aging on the trophic action of IJT is not known. METHODS: We examined the trophic effect of IJT (3 weeks post-operation) on the gastrointestinal tract and pancreas, and on plasma levels of neurotensin and gastrin in three different aged groups of Fischer 344 rats (4, 12, and 24 months old). RESULTS: Three weeks after IJT, the mucosal mass, villus height, and crypt depth increased significantly in the transposed ileum as well as in the remainder of the small intestine. The weights of the colon and pancreas increased significantly after IJT. These responses were not affected by aging. In each of the three age groups, IJT did not affect plasma gastrin level, but significantly increased plasma level of neurotensin. CONCLUSIONS: The distal ileum appears to play an important role in the regulation of growth in the intestine and pancreas; this role is preserved in aged rats. Neurotensin may play an important role in this mechanism.

Aging↗
PubMed1995

Intestinal peptide YY: ontogeny of gene expression in rat bowel and trophic actions on rat and mouse bowel.

The purpose of this study was twofold: 1) to characterize the profile of colonic peptide YY (PYY) gene expression in rats and 2) to examine for potential trophic effects of PYY on the intestine in rats and mice. Expression of PYY mRNA (analyzed by Northern blotting and in situ hybridization) and PYY (analyzed by high-performance liquid chromatography and radioimmunoassay) was detected initially at day 17 of gestation in colonic extracts of Sprague-Dawley and Fischer rats. Expression of colonic PYY mRNA increased until 7 days of age and remained at its highest level (approximately twofold greater than the adult level) through the end of the nursing period. After weaning (21 days of age), PYY mRNA levels declined quickly to adult levels. Colonic PYY concentrations followed, in a coordinated manner, with some temporal delay after birth, the increase and decrease of its mRNA. Administration of PYY increased the weight and DNA content of the duodenum significantly in nursing rats and adult mice. In mice, PYY treatment also increased weight and DNA content of the ileum and colon. The trophic effects of PYY were dose related, peptide specific, and independent of species and sex. From these findings, we hypothesize that PYY plays an important role in intestinal development and dietary adaptation.

Aging↗
PubMed1995

Phorbol ester-induced alteration in the pattern of secretion and storage of chromogranin A and neurotensin in a human pancreatic carcinoid cell line.

Brief phorbol ester treatment of BON cells results in a persistent release and cellular depletion of immunoreactive chromogranin A (CGA-IR) and neurotensin (NT-IR) cell contents. The purpose of the present study was to characterize the effects of 12-O-tetradecanoyl phorbol-13-acetate (TPA) on the secretion, biosynthesis, and steady-state messenger RNA (mRNA) levels of chromogranin A (CGA) and of a coresident peptide, neurotensin, by a novel human pancreatic carcinoid cell line, called BON. Acute TPA treatment (100 nM, 1 h) of BON cells resulted in 20- and 40-fold elevations in release of CGA-IR and NT-IR, respectively; and a 70-90% depletion of CGA-IR and NT-IR cell contents. TPA treatment also increased the biosynthetic rate of CGA-IR. Steady-state mRNA levels of CGA and NT/N (neurotensin/neuromedin N) were unchanged. Cell contents of CGA-IR and NT-IR were not replenished for a period of up to 6 days; secretion of CGA-IR and NT-IR persisted. In addition, BON cells failed to release CGA in response to stimulation by ionomycin and A23187 several days after acute TPA treatment. Our data indicate that the lack of replenishment of cell contents of CGA-IR and NT-IR is not due to decreases in steady-state CGA-IR and NT-IR mRNA levels, nor is it due to a decrease in biosynthesis of CGA-IR, but it is the result of a loss in the ability of TPA-treated BON cells to store and secrete CGA-IR and NT-IR in a regulated manner. These effects of TPA are mediated through the PKC pathway.

Analysis of Variance↗
PubMed1995

Widespread expression of the parathyroid hormone-related peptide and PTH/PTHrP receptor genes in intestinal epithelial cells.

BACKGROUND: This study was designed to determine whether the genes for both parathyroid hormone-related peptide (PTHrP) and its receptor were expressed in close proximity to one another in various regions of the gut and whether they both were evident in two intestinal epithelial cell lines. The findings would test the idea that PTHrP acts as an autocrine or paracrine factor in the gut. EXPERIMENTAL DESIGN: Reverse transcription/PCR and Northern analysis were used to detect mRNA for PTHrP and its receptor in various regions of the normal rat gut, in epithelial cell populations isolated along the villus tip-crypt axis in rat jejunum, and in a rat and a human gastrointestinal epithelial cell line (IEC-6 and LoVo, respectively). Antisera to PTHrP also were used to detect the peptide in rat gastrointestinal tissues or in growth medium from cultured cells. RESULTS: Both PTHrP and PTHrP receptor mRNA were found in all regions of the rat gut, in all cell populations isolated from rat jejunum, and in the rat and human cell lines. Immunoreactive PTHrP in tissue sections was observed in rat jejunal epithelial cells all along the villus but not in crypt cells, and immunoreactive peptide was found in growth medium from the cultured intestinal epithelial cells. CONCLUSIONS: The results show that genes for PTHrP and its receptor are expressed widely in gut epithelia. Expression of the two genes in the same regions of the gut and in the same cell line implies a local autocrine/paracrine action of the peptide. Expression of the peptide in villus epithelium but not in crypt cells suggests a role in differentiating gastrointestinal epithelial cells.

Animals↗
PubMed1994

Cytokines modulate terminal differentiation and the expression of predesquamin in cultured keratinocytes.

Predesquamin is a glycoprotein that we have isolated from human epidermis. Its monospecific polyclonal antibody immunolocalizes to the lower stratum corneum. We have previously found that it is not expressed in primary keratinocyte cultures unless both interferon-gamma and serum are added to the culture media. We now show similar results with interleukin-1 alpha substituted for interferon-gamma (but not with retinoic acid). Predesquamin is expressed in media supplemented with both cytokines (with or without serum), accompanied by the observation of larger outer squames and a more mature pattern of desquamation. As both cytokines are naturally present in the epidermis, this suggests that they contribute to desquamation in vivo. The predesquamin expression is demonstrated by Western blots of cell extracts with the specific antibody. Corresponding morphological changes are illustrated by phase-contrast microscopy.

Blotting, Western↗
PubMed1994

Predesquamin extends maturation of cultured keratinocytes.

Predesquamin is a glycoprotein that we have isolated from human epidermis. Its monospecific polyclonal antibody immunolocalizes to the lower stratum corneum. We have previously found that it is not expressed ordinarily in primary keratinocyte cultures, but that it is expressed in a serum-containing medium that is supplemented with interferon-gamma. Under these conditions, we observed larger outer squames and a more mature pattern of desquamation. We now demonstrate, by adding predesquamin directly to the culture medium without the cytokine, that the morphological changes are caused by predesquamin (and the role of IFN-gamma in the previous experiment was to stimulate the expression of predesquamin).

Antibodies↗
PubMed1994

Enzymatic activity of desquamin.

We have previously described the properties of desquamin, a cell adhesion molecule in the stratum corneum with lectin-like properties specific for amino sugars. We report here that desquamin is also a trypsin-like serine proteinase. It degrades several chromogenic peptides with arginine in the P1 position, with greatest activity for the tissue plasminogen activator peptide; it has no chymotrypsin-like activity. The enzymatic activity of desquamin is inhibited by aprotinin, leupeptin, and soybean trypsin inhibitor. The Km for all active substrates is in the millimole range and the pH for optimal activity is near 10. The enzymatic activity is stable in the temperature range from 37 to 80 degrees C, peaking near the upper end; it is only partially inhibited at 100 degrees C. Using zymogels with immobilized substrates, we show that desquamin degrades both casein and human keratins. Because desquamin is localized to the lipid envelopes of the stratum corneum and can function as an enzyme (and is extremely resistant to chemical and thermal degradation), it is in a position to play a crucial role in desquamation.

Amino Acid Sequence↗
PubMed1994

Prompt remission of post-renal transplant nephrotic syndrome with high-dose cyclosporine.

A 2.8-year-old girl with focal segmental glomerulosclerosis had recurrence of nephrotic syndrome within 3 days of renal transplantation and the serum creatinine increased. Renal biopsy showed cellular rejection and also complete effacement of the epithelial cell foot processes. The rejection responded to methylprednisolone therapy but massive proteinuria persisted. An increase in the dose of cyclosporine A to 14 mg/kg per day was followed by immediate remission of the proteinuria. One month later, a second renal biopsy showed only focal fusion of foot processes. She remains free of proteinuria 2 years later. We propose that the higher dose of cyclosporine caused remission of the nephrotic syndrome.

Biopsy↗
PubMed1994

Bombesin improves survival from methotrexate-induced enterocolitis.

OBJECTIVE: The authors determined whether bombesin could improve survival from methotrexate (MTX)-induced enterocolitis. SUMMARY BACKGROUND DATA: Bombesin prevents gut mucosal atrophy, which is produced by feeding rats an elemental diet. Administration of MTX produces a lethal enterocolitis in rats fed an elemental diet. METHODS: On treatment day 0, 60 rats were divided randomly into three groups and fed an elemental diet (Vivonex TEN, Sandoz, Minneapolis, MN) as the only source of nutrition. Groups were subdivided further to receive either saline or bombesin (10 micrograms/kg, subcutaneously, three times a day) beginning either on day 0 or day 14. Methotrexate (20 mg/kg, intraperitoneally) was given to all rats 14 days after the start of an elemental diet. RESULTS: Bombesin prevented the mucosal atrophy in the ileum produced by the elemental diet and significantly decreased mortality in rats given MTX (whether given as a pretreatment or at the time of MTX administration). CONCLUSION: Bombesin significantly improved survival in a lethal model of MTX-induced enterocolitis, possibly by maintaining gut mucosal structure. Administration of bombesin to patients receiving chemotherapy may be clinically useful in preventing the severe enterocolitis induced by various chemotherapeutic agents.

Animals↗
PubMed1994

Characterization of a human pancreatic carcinoid in vitro: morphology, amine and peptide storage, and secretion.

The study of functioning human endocrine tumors has been hampered by a lack of suitable in vitro models. We have established the first permanent cell line of a human pancreatic carcinoid tumor (BON) in culture. BON cells grow in monolayer culture and form colonies in soft agar. Injection of BON cells into nude mice produces transplantable tumors in a dose-dependent fashion. The histology of tumors in athymic mice from injection of dispersed, cultured BON cells is similar to the original histology of the resected tumor. Significant amounts of neurotensin, pancreastatin, and serotonin (5-HT) are demonstrated in the cells by radioimmunoassay (RIA) and the presence of chromogranin A, bombesin, and 5-HT is confirmed by immunocytochemistry. Numerous round and pleomorphic dense-core neurosecretory granules are present on electron microscopy. Functional receptors for acetylcholine, 5-HT, isoproterenol, and somatostatin are present on cultured cells. BON cells possess a specific transport system for uptake of 5-HT from the medium; this uptake system may be a route for regulation of autocrine effects of 5-HT on carcinoid cells. This unique human carcinoid tumor cell line should provide the opportunity for new insight into the biology of carcinoid tumors and of specific intracellular mechanisms for secretagogue action in the release of amines and peptides.

Adult↗
PubMed1994

Inhibitory effect of calcium channel blockers on growth of pancreatic cancer cells.

Calcium, which binds to calmodulin inside the cells, is an important mediator of various intracellular processes, including cell proliferation. We speculated that blockade of Ca2+ influx into the cells by Ca(2+)-channel blockers, such as phenytoin and verapamil, might affect the Ca(2+)-calmodulin pathway leading to suppression of cell growth. In this study, we examined the effect of phenytoin and verapamil on growth of two human pancreatic cancer cell lines, MIA PaCa-2 and CAV, in vitro and in vivo. Both phenytoin and verapamil inhibited growth of the two cell lines in a dose-dependent fashion. Phenytoin and verapamil each significantly prolonged doubling time of MIA PaCa-2 and the combination of the two drugs acted synergistically. The activity of ornithine decarboxylase, which is a rate-limiting enzyme of the polyamine pathway that is closely related to cell proliferation, was significantly inhibited by both drugs in a time-dependent fashion. Phenytoin, but not verapamil, inhibited growth of MIA PaCa-2 tumors xenotransplanted into nude mice, whereas both phenytoin and verapamil inhibited the growth of CAV tumors. Since phenytoin and verapamil are known to have fewer side effects than conventional antineoplastic drugs, these results suggest their possible use in novel therapeutic strategies.

Calcium Channel Blockers↗
PubMed1994

Incomplete processing of progastrin expressed by human colon cancer cells: role of noncarboxyamidated gastrins.

Gastrin is mitogenic for several colon cancers. To assess a possible autocrine role of gastrin in colon cancers, we examined human colon cancer cell lines for expression of gastrin mRNA and various forms of gastrin. Gastrin mRNA was not detected in the majority of colon cancer cell lines by Northern hybridization but was detected in all human colon cancer lines by the sensitive method of reverse transcriptase-polymerase chain reaction (PCR). Gastrin mRNA was quantitated by the competitive PCR method. The majority of cell lines expressed very low levels of gastrin mRNA (< 1-5 copies/cell); only one cell line expressed > 20 copies/cell. The mature carboxyamidated form of gastrin was not detected in any of the cell lines by radioimmunoassay or immunocytochemistry. Results suggested that either gastrin mRNA expressed by colon cancer cells was altered (mutated) or posttranslational processing of progastrin was incomplete. Gastrin cDNA from all the colon cancer cell lines had an identical sequence to the published sequence of human gastrin cDNA. Specific antibodies against precursor forms of gastrin were used, and significant concentrations of nonamidated (glycine-extended) and prepro forms of gastrin were measured in tumor extracts of representative colon cancer cell lines. The presence of precursor forms of gastrin suggested a lack of one or more of the processing enzymes and/or cofactors. Significant concentrations of the processing enzyme (peptidylglycine alpha-amidating monooxygenase) were detected in colon cancer cells by immunocytochemistry. Therefore, lack of other cofactors or enzymes may be contributing to incomplete processing of precursor forms of gastrin, which merits further investigation. Since low levels of gastrin mRNA were expressed by the majority of human colon cancer cell lines and progastrin was incompletely processed, it seems unlikely that gastrin can function as a viable autocrine growth factor for colon cancer cells. High concentrations of glycine-extended gastrin-17 (GG) (> 10(-6) M) were mitogenic for a gastrin-responsive human colon cancer (DLD-1) cell line in vitro. It remains to be seen if GG or other precursor forms of gastrin are similarly mitogenic in vivo, which may then lend credibility to a possible autocrine role of gastrinlike peptides in colon cancers.

Base Sequence↗
PubMed1994

Age-dependent increase of neurotensin expression in the ileum.

The proliferative activity of gut mucosa is increased with aging. Neurotensin (NT), a tridecapeptide localized to the distal small bowel, stimulates growth of gut mucosa; the effects of aging on gut NT are not known. Young (4-mo-old), adult (1-yr-old), and aged (2-yr-old) male Fischer 344 rats were killed; the ileal mucosa was scraped, weighed, and extracted for measurement of NT mRNA by slot-blot and Northern hybridization; and the relative NT transcription rate was determined by a nuclear run-on assay. In addition, NT tissue content and plasma levels were determined by radioimmunoassay, and full-thickness sections of ileum were examined for age-dependent alterations of NT endocrine cell (N-cell) number using immunohistochemical staining. Slot-blot and Northern-blot analyses showed that the steady-state levels of NT mRNA were increased threefold in the adult group and eightfold in the aged rats. In addition, NT peptide content and plasma levels were significantly increased in the aged group. The dramatic increases in the abundance of NT mRNA were not associated with increases in either NT transcription or N-cell number. In summary, we have demonstrated an age-dependent increase in the constitutive levels of ileal NT mRNA that appears not to be due to concomitant increases in transcription, suggesting that NT mRNA is increased by mechanisms involving mainly a posttranscriptional process. In addition, we have shown corresponding increases in the levels of both tissue and plasma NT with aging, indicating that expression of NT does not remain the same throughout life but actually increases with aging in the rat.

Aging↗
PubMed1994

Effect of aging on neurotensin-stimulated growth of rat small intestine.

The proliferative activity of gut mucosa is altered with aging; the potential for the aged gut to respond to trophic stimuli is not known. The purpose of this study was to determine whether there are age-related differences in the effects of the trophic gut peptide neurotensin (NT) on the structure and function of small bowel mucosa. NT (300 micrograms/kg) or saline (control) was injected subcutaneously at 8-h intervals for 5 days in rats of two age groups, young (2 mo) and aged (24 mo). On day 6, rats were killed, and the gut mucosa (proximal and distal small bowel) was scraped, weighed, and analyzed for DNA, RNA, and protein content and for disaccharidase (sucrase and maltase) activity. In a second experiment, the groups of rats and the protocol for NT administration were identical; however, when the rats were killed, the distal gut was removed for histological evaluation of crypt and villus length (mm) and density (no./cm gut segment) and bromodeoxyuridine immunohistochemistry. NT produced significant increases in mucosal growth (wt, DNA, RNA, and protein) in both age groups when compared with age-matched controls; the increase of growth measurements was the greatest in the small bowel mucosa of the aged rats. In addition, NT increased crypt density in both groups; only the aged group treated with NT demonstrated increases in crypt depth and villus height. Specific activities of sucrase and maltase did not change with NT treatment in either of the age groups. We conclude that the proliferative potential of small bowel mucosa is maintained with aging in response to administration of NT.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗