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S Varghese

Publications and source records attributed to S Varghese.

At least 37 records · Page 2Linked to original sources
PubMed1999

Thyroid hormones regulate lipid metabolism in a teleost Anabas testudineus (Bloch).

We compared the long-term action of 3,5,3'-triiodo-L-thyronine (T3) and 3,5-diiodo-L-thyronine (T2) on lipid metabolism in a teleost Anabas testudineus. Among the six groups of animals used in this experiment, except for the control group, all received 6-propylthiouracil (6-PTU) to create a hypothyroid state in order to analyse the action of iodothyronines on lipid metabolism. Injections of 6-PTU reduced T3 concentration in the circulation by 79.6% and injections of iodothyronines enhanced the level of T3 in the plasma, and a maximum increase was observed in T3 (500 ng)-treated specimens. Analysis of lipogenic enzymes in liver and heart showed that a tissue-specific variation exists in the action of thyroid hormones and, in many cases, activity is higher in T2-treated groups. Analysis of various lipid classes showed that long-term administration of T2 is also effective in producing a comparable effect with that of T3 on lipid metabolism.

Animals↗
PubMed1999

Leukemia inhibitory factor and oncostatin M stimulate collagenase-3 expression in osteoblasts.

Leukemia inhibitory factor (LIF) and oncostatin M (OSM) have multiple effects on skeletal remodeling. Although these cytokines modestly regulate collagen synthesis in osteoblasts, their effects on collagenase expression and collagen degradation are not known. We tested whether LIF and OSM regulate the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in osteoblast-enriched cells isolated from fetal rat calvariae. LIF and OSM increased collagenase-3 (MMP-13) mRNA and immunoreactive protein levels in a time- and dose-dependent manner. LIF and OSM enhanced the rate of transcription of the collagenase gene and stabilized collagenase mRNA in transcriptionally arrested cells. LIF and OSM failed to regulate the expression of gelatinase A (MMP-2) and B (MMP-9). LIF and OSM modestly stimulated the expression of TIMP-1 but did not alter the expression of TIMP-2 and -3. In conclusion, LIF and OSM stimulate collagenase-3 and TIMP-1 expression in osteoblasts, and these effects may be involved in mediating the bone remodeling actions of these cytokines.

Animals↗
PubMed1999

Differential expression of stathmin during neoplastic conversion of human prostate epithelial cells is reversed by hypomethylating agent, 5-azacytidine.

In a variety of human tumor tissues, including those of prostate and breast, CpG hypermethylation represents one of the mechanisms downregulating the expression of specific proteins, including tumor suppressor proteins. Using 267B1-XR cells generated by ionizing radiation-induced transformation of epithelial cells, derived from neonatal human prostate and immortalized by SV40 (267B1), we now report markedly low levels of expression of the cytoplasmic phosphoprotein stathmin, in addition to several proteins of the actin microfilaments and intermediate filaments that characterize the altered phenotype. Stathmin is emerging as a relay protein integrating signals from diverse pathways during differentiation and neoplastic progression. In this in vitro prostate carcinogenesis model system, where loss of specific-protein expression is a major feature of the transformed 267B1-XR cells, we employed 5-azacytidine treatment followed by 2D-PAGE to reveal if experimental genomic hypomethylation reinstated the levels of any of the differentially expressed proteins. Our data suggest that stathmin represents one such example.

Antimetabolites, Antineoplastic↗
PubMed1998

Overexpression of Rb and E2F-1 in ataxia-telangiectasia lymphocytes.

AT cells exhibit defective cell cycle regulation following DNA damage. Previous studies have shown that induction of p53 and p21 proteins are delayed in response to ionizing radiation, resulting in the failure of G1/S checkpoint in AT cells. In this study, further investigation of the molecular mechanisms underlying G1/S phase progression in AT cells was conducted. Exponentially growing normal and AT cells were exposed to 2 Gy of ionizing radiation and the expression levels and functional activities of Rb and E2F-1 proteins were determined. We observed overexpression of hyperphosphorylated Rb and E2F-1 proteins in AT cells, which was unaffected post-irradiation. Furthermore, gel shift assays showed that E2F-1-DNA binding was constitutive in AT cells, whereas it was inhibited in control cells following exposure to ionizing radiation. The data suggests that abnormalities in the function of Rb and E2F-1 proteins may also be responsible for the failure of AT cells to arrest in the G1/S checkpoint in response to DNA damage.

Adenovirus E2 Proteins↗
PubMed1997

Cytokine regulation of adult human osteoblast-like cell prostaglandin biosynthesis.

Prostaglandin (PG) biosynthesis by cytokine stimulated normal adult human osteoblast-like (hOB) cells was evaluated by thin layer chromatography, high performance liquid chromatography, and specific immunoassays. PGE2 was the predominant PG formed under all incubation conditions tested. Control samples produced measurable amounts of PGE2, and the measured level of this metabolite increased by 22-fold (from 7 to 152 ng/ml) following a 20 h treatment with the combination of TGF beta and tumor necrosis factor-alpha(TNF). The production of 6-keto-PGF1 alpha (the stable metabolite of prostacyclin) and of PGF2 alpha were each increased by about five-fold (from about 0.5 to 2.5 ng/ml) in samples treated with the cytokines. Thus, TGF beta and TNF exerted a regulation of hOB cell PG biosynthesis that was principally directed towards an increased PGE2 biosynthesis, with lesser effects on the production of other PG metabolites. COX-2 mRNA levels were increased within 2 h of cytokine stimulation, reached a maximum at 6-12 h, and levels had appreciably diminished by 24 h after treatment. Both TGF beta and TNF could independently increase COX-2 mRNA levels and PG biosynthesis. However, the increased production of PGE2 resulting from TNF stimulation was blocked by the addition of an interleukin-1 beta (IL-1 beta) neutralizing antibody, suggesting that TNF regulation of hOB cell PG synthesis was secondary to its capacity to increase hOB cell IL-1 beta production. TGF beta regulation of PG production was not affected by the addition of the neutralizing antibody. These studies support the proposition that PGs can be important autocrine/paracrine mediators of bone biology, whose production by hOB cells is responsively regulated by osteotropic cytokines.

Adult↗
PubMed1997

Transforming growth factor beta1 inhibits collagenase 3 expression by transcriptional and post-transcriptional mechanisms in osteoblast cultures.

Transforming growth factor (TGF) beta1 is an autocrine regulator of bone cell function. We demonstrated that TGF beta1 enhances bone collagen synthesis, but its effects on collagen degradation are not well characterized. We tested the effects of TGF beta1 on rat collagenase 3 expression in cultures of osteoblast-enriched cells from fetal rat calvariae (Ob cells). Treatment with TGF beta1 at 0.4 nM decreased steady state collagenase mRNA levels after 2 to 24 h. This dose-dependent effect was observed at TGF beta1 concentrations of 4 pM to 1.2 nM, and was accompanied by decreased levels of immunoreactive procollagenase. The protein synthesis inhibitor cycloheximide increased collagenase transcripts, but did not prevent the effect of TGF beta1 on collagenase mRNA levels. TGF beta1 accelerated the decay of collagenase mRNA in transcriptionally arrested Ob cells. In addition, TGF beta1 decreased the levels of collagenase heterogeneous nuclear RNA and the rate of collagenase gene transcription in Ob cells. TGF beta1 enhanced the expression of tissue inhibitors of metalloproteinases (TIMP) 1 and 3 and caused a modest decrease of TIMP 2 mRNA levels. In conclusion, TGF beta1 decreases interstitial collagenase transcripts and protease levels in Ob cells by transcriptional and post-transcriptional mechanisms, and this effect may contribute to its actions on bone matrix.

Animals↗
PubMed1997

Regulation of collagenase-3 by bone morphogenetic protein-2 in bone cell cultures.

Bone morphogenetic protein-2 (BMP-2), a member of the transforming growth factor superfamily of peptides, induces ectopic bone formation in vivo. The actions of BMP-2 on osteoblastic cells include stimulation of collagen synthesis, but the role of BMP-2 on collagen degradation is not known. We examined whether BMP-2 affects the expression of collagenase-3, an enzyme that degrades type I collagen at neutral pH, and that of tissue inhibitors of matrix metalloproteinases (TIMPs) in primary osteoblast-enriched cells from 22-day-old fetal rat calvariae. BMP-2 suppressed collagenase messenger RNA (mRNA) and immunoreactive protein levels. BMP-2 did not affect collagenase mRNA stability, but it reduced collagenase heterogeneous nuclear RNA levels and decreased the rate of transcription of the collagenase gene. BMP-2 also stimulated TIMP 1 and TIMP 3 mRNA levels, but failed to alter TIMP 2 expression. In conclusion, our studies indicate that BMP-2 suppresses collagenase-3 gene transcription and stimulates TIMP 1 and TIMP 3 expression in osteoblasts. The regulation of collagenase and TIMPs by BMP-2 in osteoblasts may play a role in osteoinduction.

Animals↗
PubMed1996

Transcriptional and posttranscriptional regulation of interstitial collagenase by platelet-derived growth factor BB in bone cell cultures.

Platelet-derived growth factor (PDGF), a bone cell mitogen, stimulates bone collagen degradation and does not enhance bone matrix apposition rates. The mechanism of the effect on collagen degradation is unknown, and it could involve changes in interstitial collagenase synthesis. We tested the effects of PDGF on interstitial collagenase expression in cultures of osteoblast-enriched cells from fetal rat calvariae (Ob cells). After 4-8 h of treatment, PDGF BB at 0.3 nM increased steady state collagenase messenger RNA (mRNA), whereas PDGF AA had no effect. The effect of PDGF BB on collagenase transcripts was dose dependent. PDGF BB increased the levels of immunoreactive collagenase after 6 h, whereas the levels were decreased after 16 h. Stimulation of collagenase mRNA by PDGF BB was dependent on de novo protein synthesis and activation of protein kinase C. PDGF BB prolonged the half-life of collagenase mRNA in transcriptionally arrested cells. PDGF BB initially increased and subsequently decreased the rate of collagenase gene transcription and the levels of collagenase heterogeneous nuclear RNA. In conclusion, PDGF BB regulates interstitial collagenase in Ob cells by transcriptional and posttranscriptional mechanisms, and this effect may contribute to its stimulatory actions on bone collagen degradation.

Animals↗
PubMed1995

Insulin-like growth factors inhibit interstitial collagenase synthesis in bone cell cultures.

Insulin-like growth factor-I (IGF-I) and IGF-II are among the most prevalent growth factors secreted by bone cells and are presumed to act as autocrine regulators of bone formation. We recently demonstrated that IGFs inhibit bone collagen degradation, and we postulated that they may either inhibit the expression of interstitial collagenase or stimulate the synthesis of tissue inhibitors of metalloproteinase-1 (TIMP-1), -2, or -3. We tested the effects of IGF-I and -II on collagenase and TIMP-1, -2, and -3 expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cells). Steady state messenger RNA (mRNA) levels were determined by Northern blot analysis, and collagenase concentrations were determined in the culture medium by a specific immunoassay. After 2-6 h of treatment, IGF-I and -II decreased collagenase transcripts by up to 80%. IGF-I was a more potent inhibitor than IGF-II, because it was active at doses as low as 10 nM, whereas a dose of 100 nM was required to observe the IGF-II effect. In addition, IGF-I and -II opposed the stimulatory effect of retinoic acid on collagenase transcripts. Immunoreactive collagenase levels were not detectable in control or IGF-treated cultures, but IGF-I and -II decreased the levels induced by retinoic acid by 70-90%. The protein synthesis inhibitor cycloheximide superinduced collagenase transcripts, and IGF-I or -II decreased this mRNA induction to levels similar to, but not lower than, those observed in control cultures. The effects of IGF-I and -II on collagenase transcripts were not modified by the DNA synthesis inhibitor hydroxyurea at 1 mM. Neither IGF-I nor IGF-II modified the expression of TIMP-1, -2, or -3 mRNA in Ob cells. TIMP protein levels were not determined, and our study does not exclude a translational or posttranslational effect of IGF. In conclusion, IGF-I and -II decrease interstitial collagenase transcripts as well as induced protease levels in Ob cells, and this effect may contribute to their inhibitory actions on bone collagen degradation.

Animals↗
PubMed1995

Basic fibroblast growth factor stimulates expression of interstitial collagenase and inhibitors of metalloproteinases in rat bone cells.

Basic fibroblast growth factor (bFGF) is a bone cell mitogen that affects osteoblastic function by suppressing type I collagen synthesis. The investigators in this study examined whether bFGF also regulates interstitial collagenase and tissue inhibitors of metalloproteinases (TIMPs) in osteoblast-enriched cells isolated from 22-day fetal rat calvariae. After exposure to 600 pM bFGF, interstitial collagenase messenger RNA (mRNA) levels, as determined by Northern hybridization analysis, increased after 2 h and were maximally stimulated to approximately 13-fold at 6 h. Exposure of osteoblast-enriched cells to 0.06-6 nM bFGF increased collagenase mRNA in a dose-dependent manner, and bFGF also increased immunoreactive collagenase measured in the culture medium by Western blot analysis. The protein synthesis inhibitor cycloheximide, as well as two inhibitors of protein kinase C, staurosporine and sangivamycin, prevented the bFGF induction of collagenase transcripts, whereas indomethacin, an inhibitor of prostaglandin synthesis, decreased the effect of bFGF on collagenase mRNA levels by about 50%. After exposure to 600 pM bFGF, levels of TIMP 1 and TIMP 3 mRNAs were also maximally stimulated to approximately 6-fold at 16 h and 4-fold at 6 h. bFGF did not modify TIMP 2 expression. In conclusion, bFGF may modulate degradation of collagenous bone matrix by inhibiting collagen as well as stimulating collagenase and TIMPs by osteoblasts.

Alkaloids↗
PubMed1994

Transformation of human epidermal keratinocytes with fission neutrons.

The biological effects of exposures to high LET radiations have particular relevance to radiation protection and risk assessment. Since most cancers are of epithelial origin, it is important to obtain a better understanding of radiation-induced oncogenic transformation in this cell type. Accordingly we have initiated studies to determine whether immortalized human epidermal keratinocytes (RHEK) can be transformed with high LET radiations. Exponentially growing RHEK cells were treated with single doses (1, 10, 25, 50 and 100 cGy) of 0.85 MeV fission neutrons from the Janus reactor. Neutron exposure led to the development of morphologically altered cells and foci formation after 6 weeks at confluence. These transformed cultures grew with an increased saturation density, exhibited anchorage-independent growth and formed tumors in athymic mice. Single-strand conformational polymorphism analysis and DNA sequencing demonstrated the absence of point mutations in codons 12/13 and 61 in the Ha-ras, Ki-ras, or N-ras genes and exons 4-9 of the p53 tumor suppressor gene. These studies demonstrate that high LET radiations (fission neutrons) can transform immortalized human epithelial cells to a malignant phenotype that does not appear to involve mutations in either the cellular p53 or ras genes.

Animals↗
PubMed1994

Regulation of interstitial collagenase expression and collagen degradation by retinoic acid in bone cells.

In osteoblasts, retinoic acid (RA) modulates the synthesis of various proteins, including collagen. However, little is known about the effects of RA on the regulation of interstitial collagenase synthesis and collagen degradation. After treatment of primary osteoblast-enriched (Ob) cells from fetal rat calvariae with 100 nM all-trans-RA (tRA), collagenase mRNA levels, as determined by Northern blotting, did not change after 2 h, increased by 13- to 18-fold after 6 h, and remained elevated until 48 h. Exposure of Ob cells to 10 nM to 1 microM tRA, 13-cis-RA, and 9-cis-RA induced collagenase mRNA in a dose-dependent manner. Collagenase mRNA induction by RA was blocked by cycloheximide. RA increased the stability of collagenase mRNA in Ob cells, suggesting posttranscriptional regulation. Exposure of Ob cells to RA induced immunoreactive procollagenase in medium, as determined by enzyme-linked immunosorbent assay and Western blotting. RA action on collagen degradation was examined in [3H]proline-pulsed intact calvariae chased with and without tRA for 72 h. The release of [3H]hydroxyproline into culture medium was increased by 64% in the presence of 10 nM to 1 microM tRA. In conclusion, RA increases collagenase synthesis and collagen degradation in bone and is likely to play an important role in bone remodeling.

Animals↗
PubMed1993

Growth factors regulate the synthesis of insulin-like growth factor-I in bone cell cultures.

Insulin-like growth factor-I (IGF-I), a prevalent growth factor secreted by bone cells, has important effects on bone remodeling. Hormones are known to regulate the synthesis of skeletal IGF-I, but there is limited information about the actions of growth factors on IGF-I synthesis. We tested the effects of basic fibroblast growth factor (bFGF), transforming growth factor-beta 1 (TGF beta 1), and platelet-derived growth factors (PDGF) AA and BB on IGF-I mRNA expression and polypeptide concentrations in cultures of osteoblast-enriched (Ob) cells from 22-day-old fetal rat calvariae. Steady state IGF-I mRNA levels were determined by Northern blot analysis, and IGF-I concentrations were determined in acidified and fractionated culture medium by a specific RIA. Treatment of Ob cells with bFGF at 0.06-6 nM, TGF beta 1 at 0.04-4 nM, and PDGF BB at 0.3-3.3 nM caused a dose-dependent decrease in steady state IGF-I mRNA. A smaller effect was observed with PDGF AA. The effect was initially observed after 6-8 h of treatment and was maximal after 16 h. Treatment with bFGF at 0.6-6 nM, TGF beta 1 at 0.4-4 nM, and PDGF BB at 0.3-3.3 nM for 24 h decreased IGF-I polypeptide concentrations by 40-80%. The effects of bFGF, TGF beta 1, and PDGF BB and AA on IGF-I mRNA were independent of protein synthesis and cell division, as they were observed in the presence and absence of cycloheximide at 3.6 microM or hydroxyurea at 1 mM. Similarly, their inhibitory actions on immunoreactive IGF-I were not prevented by hydroxyurea. In conclusion, bFGF, TGF beta 1, PDGF BB, and, to a lesser extent, PDGF AA decrease skeletal IGF-I synthesis by reducing IGF-I transcript levels, and this effect may contribute to their actions on selected aspects of Ob cell function.

Animals↗
PubMed1993

Skeletal growth factors.

Growth factors are polypeptides with important actions on the replication and differentiated function of cells. Skeletal cells synthesize a variety of growth factors, which are believed to act in an autocrine or paracrine fashion. These growth factors include platelet-derived growth factor, fibroblast growth factor 1 and 2, insulin-like growth factor I and II, transforming growth factors beta 1, 2, and 3, and selected bone morphogenetic proteins. Skeletal cells also synthesize specific binding proteins for selected growth factors. In addition, bone marrow cells synthesize a variety of cytokines known to have important actions in bone remodeling. Fibroblast growth factors and platelet-derived growth factors are, for the most part, mitogenic for skeletal cells, whereas insulin-like growth factors and transforming growth factor beta enhance the differentiated function of the osteoblast. Growth factors also modify osteoclast recruitment and function and as such, bone resorption. Skeletal growth factors can be regulated at the level of synthesis, activation, binding proteins, and receptor binding, and, as a result, their activity can be modified by exogenous agents.

Animals↗
PubMed1992

Role of platelet derived growth factor in bone cell function.

PDGF is a mitogen for cells of the osteoblastic lineage. PDGF is present in the systemic circulation and is locally synthesized by skeletal cells. The systemic form primarily contains PDGF B chains, which are intrinsically more active than PDGF A subunits, the forms secreted by normal bone cells. PDGF AA is regulated by other growth factors and cytokines, which modulate its binding to osteoblastic receptors and its synthesis by skeletal cells. The exact role of PDGF in bone remodelling is still uncertain and current information suggests that this factor has a function in the response to inflammation and wound healing.

Animals↗
PubMed1991

Rat thymosin beta 4 gene. Intron-containing gene and multiple retroposons.

Thymosin beta 4 (T beta 4), a 43-residue peptide of uncertain function, is widely distributed in most tissues of vertebrates. Southern blot analysis demonstrated that the rat genome contains many closely related T beta 4 sequences. Two distinct T beta 4 sequences were isolated by screening a rat genomic DNA library. Analysis of these clones revealed that they represent T beta 4 retroposons; they lack introns and contain stretches of poly(A) sequence at their 3' ends and direct repeats flanking the cDNA-like sequences. In order to isolate a T beta 4 intron-containing gene, DNA fragments that include T beta 4 introns were generated by polymerase chain reaction, using rat genomic DNA as template. Subsequent Southern blot analysis of rat genomic DNA with intron-specific probes demonstrated the presence of a single T beta 4 intron-containing gene. Upon screening the rat genomic DNA library with an intron-specific probe, two overlapping clones of the T beta 4 intron-containing gene were isolated. Further characterization and sequence analysis showed that the rat T beta 4 intron-containing gene has a 2-kilobase pair-long transcription unit containing two introns. The transcription initiation site and putative regulatory elements of the promoter were identified.

Animals↗
PubMed1989

Transcriptional regulation and chromosomal assignment of the mammalian calbindin-D28k gene.

To determine whether 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] regulates transcription of the rat renal calbindin-D28k gene, the rate of calbindin-D28k mRNA synthesis was measured directly in nuclei using the in vitro nuclear transcription assay. Nuclei were prepared from kidneys of vitamin D-deficient rats at various times after a single ip injection of 1,25-(OH)2D3, and transcription was allowed to proceed in vitro in the presence of [32P]UTP for 30 min at 29 C, at which time the incorporation of UTP into trichloroacetic acid-precipitable material was optimal. Incorporation of UTP was decreased by 64.6% by alpha-amanitin, which selectively inhibits polymerase II. Purified [32P]RNA was analyzed for newly synthesized calbindin-D-28k gene transcripts by hybridization to calbindin-D28k cDNA immobilized on nitrocellulose filters. Using this assay we found that the first significant increase in calbindin-D28k gene transcription occurred at 1 h, and the peak of transcriptional activity occurred at 2 h. Within 12 h of 1,25-(OH)2D3 treatment, calbindin-D28k gene transcription returned to control levels. Using Northern blot analysis, a significant increase in calbindin-D RNA was first observed 2 h after hormone administration, reaching a maximum at 12 h. Renal calbindin-D28k protein levels are significantly increased by 3 h and reach a maximum value 48 h after hormone administration. Our results suggest that the early increase in renal calbindin-D28k may be due to transcriptional regulation. The long time lag between transcription and the peak of calbindin mRNA and calbindin protein accumulation may reflect the involvement of post-transcriptional mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗