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Biomedical subjects

S Bai

Publications and source records attributed to S Bai.

At least 19 recordsLinked to original sources
PubMed2001

Effect of low dose cyclosporine and sirolimus on hepatic drug metabolism in the rat1.

BACKGROUND: We examined the effect cyclosporine (CsA) and sirolimus (SRL) alone and in combination on hepatic cytochrome P450-mediated metabolism in rats. METHODS: Rats were given 1 mg/kg of CsA or 0.4 mg/kg of SRL alone or in combination via constant intravenous infusion. Renal function was evaluated at the end of treatment. Blood samples were obtained to estimate CsA and SRL concentrations. Hepatic microsomes were prepared for immunoblotting and catalytic assays. RESULTS: CsA alone did not alter serum creatinine levels. SRL given alone or in combination with CsA produced a significant increase in urine output without changes in fluid balance. Although CsA and SRL administered alone caused damage to renal proximal tubules, the two-drug combination dramatically increased the renal structural damage. CsA alone suppressed cytochrome P450 (CYP) 3A2 protein levels by 39% (P=0.012) and catalytic activity by 30% (P=0.042). SRL alone reduced catalytic activity by 38% (P=0.012). Combination therapy reduced both CYP3A2 levels by 55% (P<0.001) and catalytic activity by 55% (P=0.001). CYP2C11 protein expression or catalytic activity were not changed in any group. CYP2A1 protein expression and catalytic activity were both significantly reduced in rats given CsA or/and SRL. Steady-state CsA levels were increased during concurrent SRL dosing, however, SRL concentrations were not changed by CsA coadministration. CONCLUSIONS: Concurrent SRL dosing increases CsA concentrations due to inhibition of hepatic CYP3A2 protein expression. Nephrotoxicity caused by combination therapy is due to CsA elevating levels of SRL or by SRL itself. Concurrent administration of CsA and SRL in transplant patients should be performed with caution.

Animals↗
PubMed2001

Hoxa-9 represses transforming growth factor-beta-induced osteopontin gene transcription.

Smad2 and Smad3 are downstream transforming growth factor-beta (TGF-beta) signaling molecules. Upon phosphorylation by its type I receptor, Smad2 or Smad3 forms a complex with Smad4 and translocates to the nucleus where the complex activates target gene transcription. In the present study, we report that Smad3 binds directly to the osteopontin (OPN) promoter and that Smad4 interacts with the Hox protein and displaces it from its cognate DNA binding site in response to TGF-beta stimulation. In gel shift assays, the glutathione S-transferase-Smad3 fusion protein was found to bind to a 50-base pair DNA element (-179 to -229) from the OPN promoter. Also, we found that both Hoxc-8 and Hoxa-9 bound to a Hox binding site adjacent to Smad3 binding sequence. Interestingly, Smad4, the common partner for both bone morphogenic protein and TGF-beta signaling pathways, inhibited the binding of Hox protein to DNA. FLAG-tagged Smad4 coimmunoprecipitated with HA-tagged Hoxa-9 from cotransfected COS-1 cells, demonstrating an interaction between Smad4 and Hoxa-9. Transfection studies showed that Hoxa-9 is a strong transcriptional repressor; it suppresses the transcription of the luciferase reporter gene driven by a 124-base pair OPN promoter fragment containing both Smad3 and Hox binding sites. Taken together, these data demonstrate a unique TGF-beta-induced transcription mechanism. Smad3 and Smad4 exhibit different functions in activation of OPN transcription. Smad3 binds directly to the OPN promoter as a sequence-specific activator, and Smad4 displaces the transcription repressor, Hoxa-9, by formation of Smad4/Hox complex as part of the transcription mechanism in response to TGF-beta stimulation.

Animals↗
PubMed2001

Syngeneic consecutive rat spleen transplantation bearing infantile testis.

Seventy-seven Lewis adult rats received autologous testicular implants in the spleen. These spleens were retransplanted in a consecutive transplant fashion as we described earlier. Some were observed as long as 26 to 27 months. Although earlier splenic-testicular transplants showed benign granulosa cell tumors, some of the long-term follow-ups showed a seminoma-like transformation.

Animals↗
PubMed2001

Functional analysis of the mouse galactose-1-phosphate uridyl transferase (GALT)promoter.

Galactose-1-phosphate uridyltransferase (GALT) is expressed in most tissues, but the near total absence of catalytic activity in humans with the disease galactosemia leads to specific organ dysfunction, the pathophysiology of which remains an enigma. To characterize the transcriptional regulation of the mouse GALT gene, we isolated and sequenced over 3 kb of a 5'-flanking sequence and functionally characterized the region using in vitro transient transfection and in transgenic mice. A minimal promoter of 145 bp was found to function in both HepG2 cells and NS20Y mouse neuroblastoma cells. The minimal promoter contains regions of homology to the corresponding rat and human GALT genes. In transgenic mice expressing a luciferase transgene under control of a 1.9-kb fragment of the mGALT promoter region, reporter activity was found in most tissues, with higher than expected reporter levels in neonatal brain. To determine if high galactose levels in tissues could induce promoter activity, we bred the mGALT:luciferase transgene into a line of mice in which the GALT gene function has been eliminated by homologous recombination. High tissue levels of galactose and metabolites did not induce reporter activity above background. The studies show that GALT transcriptional regulation is complex and not directly induced by substrate levels.

Alleles↗
PubMed2001

Effects of matrix proteins on the expression of matrix metalloproteinase-2, -9, and -14 and tissue inhibitors of metalloproteinases in human cytotrophoblast cells during the first trimester.

The activity of matrix metalloproteinases (MMPs) specifies the ability of the trophoblast cell to degrade extracellular matrix (ECM) substrates. Usually the process of normal human placentation involves a coordinated interaction between the fetal-derived trophoblast cells and their microenvironment in the uterus. In this study, the effects of ECM proteins on the expression of MMP-2, -9, and -14 (membrane-type MMP-1); and the production of tissue inhibitors of metalloproteinase (TIMP) types -1, -2, and -3 have been investigated. Cytotrophoblast cells at 9 or 10 wk of gestation were cultured on various ECM coated dishes under serum-free conditions. Gelatin zymography analysis showed that cells grown on fibronectin (FN), laminin (LN), and vitronectin (VN) secreted more MMP-9 (about 1.5- to 3-fold more) than cells cultured on collagen I (Col I), whereas the secretion of MMP-9 by cells cultured on collagen IV (Col IV) was only half that by the cells on Col I. Northern Blot analysis gave the same results as zymography, indicating that expression of the MMP-9 gene in cytotrophoblast cells can be affected by matrix proteins. There was no significant difference in the expression of MMP-2 either at protein or mRNA levels among the cells cultured on the different matrix substrates. The expression of MMP-14 was regulated in a manner similar to that of MMP-2. Using ELISA, we detected higher levels of TIMP-1 in the culture medium of cells grown on VN, LN, and FN compared with that grown on Col I. But the expression of TIMP-3 mRNA was remarkably inhibited by VN, and ECM proteins had no effect on TIMP-1 and TIMP-2 mRNA expression. It was also observed that cultured cytotrophoblast cells expressed the corresponding receptors for the tested matrix proteins, such as integrins alpha(1), alpha(5), alpha(6), beta(1), and beta(4). Furthermore, the adhesiveness of cytotrophoblast cells on Col I, Col IV, FN, and LN was increased by 62%, 45%, 21%, and 22%, respectively, when compared with adhesiveness on VN. Isolated cytotrophoblast cells remained stationary when cultured on dishes coated with Col I and Col IV, but they assumed a more motile morphology and aggregated into a network when cultured on LN and VN. These data indicate that human trophoblast cells interact with their microenvironment to control their behavior and function.

Adult↗
PubMed2001

In vivo induction of hepatic p-glycoprotein by cyclosporine in the rat.

The objective of the present study was to investigate the regulation of P-glycoprotein by cyclosporine, a known inhibitor of CYP3A, at different dosage levels and lengths of treatment. Rats were given various doses of cyclosporine through oral administration or subcutaneous injection. Each treatment group was studied for 28 days or 28 days followed by 14 days of olive oil vehicle dosing. In each group, rats administered vehicle alone served as the controls. At the end of the study, liver microsomes were prepared and hepatic P-glycoprotein levels were quantified by Western blot analysis. Significant induction of hepatic P-glycoprotein was found in rats given cyclosporine. Rats administered 30 mg/Kg/d orally and 15 mg/Kg/d subcutaneously showed an increase in hepatic P-glycoprotein by 93% (p = 0.0011) and 136% (p < 0.001), respectively. Low doses of cyclosporine also induced P-glycoprotein but not to a significant extent, indicating a dose-dependent effect. The pattern of induction of P-glycoprotein was not, however, dependent on the route of administration. Fourteen days after the discontinuation of cyclosporine treatment, P-glycoprotein levels returned to near the control values. As a drug efflux transporter, the induction of P-glycoprotein by cyclosporine may decrease the hepatic metabolism of P-glycoprotein substrates. Therefore this induction of hepatic P-glycoprotein and suppression of hepatic CYP3A may have a coordinate effect on the metabolism of cyclosporine. These data may help explain the large variability associated with cyclosporine absorption, metabolism, and circulating blood levels.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Smad6 as a transcriptional corepressor.

Smad6 and Smad7, a subgroup of Smad proteins, antagonize the signals elicited by transforming growth factor-beta. These two Smads, induced by transforming growth factor-beta or bone morphogenetic protein (BMP) stimulation, form stable associations with their activated type I receptors, blocking phosphorylation of receptor-regulated Smads in the cytoplasm. Here we show that Smad6 interacts with homeobox (Hox) c-8 as a transcriptional corepressor, inhibiting BMP signaling in the nucleus. The interaction between Smad6 and Hoxc-8 was identified by a yeast two-hybrid approach and further demonstrated by co-immunoprecipitation assays in cells. Gel shift assays show that Smad6, but not Smad7, interacts with both Hoxc-8 and Hoxa-9 as a heterodimer when binding to DNA. More importantly, the Smad6-Hoxc-8 complex inhibits interaction of Smad1 with Hoxc-8- and Smad1-induced transcription activity. These data indicate that Smad6 interacts with Hox transcription factors as part of the negative feedback circuit in the BMP signaling pathway.

Bone Morphogenetic Proteins↗
PubMed2000

Effect of dietary oil intake on hepatic cytochrome P450 activity in the rat.

The objective of the present study was to investigate whether or not different dietary oils, commonly used as drug vehicles, alter hepatic microsomal drug metabolism in the rat. Male adult Sprague-Dawley rats were administered 1 mL/kg/d of either corn, olive, sesame, or soybean oil via oral gavage for 7 days. An additional rat group was given an equal volume of water each day to serve as a control. We found that the hepatic cytochrome P450 (CYP) 3A2 protein level increased by 16% (p < 0.01) in rats given soybean oil compared with control rats. In contrast, CYP2C11 protein levels decreased by 32% (p < 0.01) in the corn oil group and by 31% (p < 0. 01) in rats given olive oil. The changes in the in vitro production of 6beta- and 2alpha-hydroxytestosterone, markers of CYP3A2 and 2C11 activities, respectively, were consistent with their protein levels, although not statistically different than controls. The results demonstrate that dietary oils may have differential effects on specific hepatic CYP isoforms and may add to the variability in metabolism when xenobiotics are administered using dietary oils as vehicles.

Animals↗
PubMed2000

Plasma membrane-adjacent actin filaments, but not microtubules, are essential for both polarization and hyphal tip morphogenesis in Saprolegnia ferax and Neurospora crassa.

The organization and roles of F-actin and microtubules in the maintenance and initiation of hyphal tip growth have been analyzed in Saprolegnia ferax and Neurospora crassa. In hyphae of both species, the apex is depleted of microtubules relative to subapical regions and near-normal morphogenesis occurs in concentrations of nocodazole or MBC which remove microtubules, slow growth, and disrupt nuclear positioning. In contrast, each species contains characteristic tip-high arrays of plasma membrane-adjacent F-actin, whose organization is largely unaltered by the loss of microtubules but disruption of which by latrunculin B disrupts tip morphology. Hyphal initiation and subsequent normal morphogenesis from protoplasts of both species and spores of S. ferax are independent of microtubules, but at least in S. ferax obligatorily involve the formation of F-actin caps adjacent to the hyphal tip plasma membrane. These observations indicate an obligatory role for F-actin in hyphal polarization and tip morphogenesis and only an indirect role for microtubules.

Actins↗
PubMed2000

1H dynamic nuclear polarization in supercritical ethylene at 1.4 T.

(1)H dynamic nuclear polarization (DNP) has been measured in supercritical ethylene in the pressure range 60-300 bar in an external field of 1.4 T. A single-cell sapphire tube was used as a high-pressure cell, and powdered 1,3-bisdiphenylene-2-phenyl allyl (BDPA) free radicals were added and distributed at the wall of the cell. At all pressures the dominant DNP mechanism was a positive Overhauser enhancement, caused by proton-electron contact interactions at the fluid/solid radical interface. The observed enhancements varied from 12 at 60 bar to 17 at 300 bar. Besides the Overhauser enhancement, small solid state and thermal mixing enhancements also were observed, indicating that part of the ethylene was adsorbed at the radical surface for a prolonged time. The impacts of the experimental conditions on the Overhauser enhancement factors are discussed, and enhancements of at least 40-60 are estimated when the EPR saturation factor and the leakage factor become maximal. These data indicate that DNP-enhanced NMR has the potential of extending the impact of NMR in research areas involving supercritical fluids.

Ethylenes↗
PubMed2000

Mechanisms of plant embryo development.

1. Evolution in plants has favored both a simpler body plan with fewer cell types and the epigenetic flexibility to regenerate, via growth, dedifferentiation, and redifferentiation, to recover from environmental insults. It has become increasingly apparent that a plant cell uses external signals to differentiate and to maintain or to change the differentiated state. A cell-cell signaling and positional information strategy seems to be the predominant mechanism employed in plant development. 2. An axis can be initiated by physical/chemical forces such as light and ion current, requiring no new gene action. Random chemical fluctuations and physicochemical forces could explain the initiation of differences among cells of equal developmental potential. Amplification of chemical polarizing events may lead to biochemical differences, new gene expression, and finally shoot/root axis establishment. 3. Radial and axial patterning may be governed by a mechanism involving polar auxin transport. 4. Because the meristems and the three fundamental tissues formed during embryogenesis are renewed and extended throughout the life of the plant, with some exceptions, most genes expressed in the embryo are also expressed during postgermination development. 5. Embryogenic competence is acquired during reproductive development. While the zygote is determined for embryogenesis, the developing embryo and often the seedling remain embryogenic. Embryogenic potential declines during vegetative development. The embryogenic strength of a tissue is correlated with its developmental distance from the zygote.

Embryonic Induction↗
PubMed2000

Unusual deregulation of cell cycle components in early and frank estrogen-induced renal neoplasias in the Syrian hamster.

There is strong evidence that estrogens are involved in the etiology, promotion and progression of a variety of cancers, including the cancers of the breast and endometrium. The Syrian hamster estrogen-induced, estrogen-dependent renal neoplasm is a well-established animal model used to elucidate the cellular and molecular mechanisms involved in solely estrogen-induced carcinogenic processes. G(1) cell cycle progression was studied in estrogen-induced early renal tumor foci and in large kidney tumors of castrated male hamsters. Levels of cyclin D1, cyclin E and retinoblastoma (pRb) proteins were higher in these renal neoplasias than in adjacent uninvolved renal tissue and kidneys from untreated, age-matched animals. Of particular interest is the presence of a predominant 35 kDa cyclin E protein variant form in primary renal tumors. In addition, amounts of the phosphorylated forms of cyclin-dependent kinases (cdk) 2 and 4 were decreased, and both RNA and protein levels of p27(kip1) (p27), a cyclin-dependent kinase inhibitor, were markedly higher in early and frank renal tumors than in adjacent uninvolved renal tissue and kidneys of untreated, age-matched animals. These changes in cell cycle components coincided with a rise in renal tumor cell proliferation. Binding of the elevated p27 protein to cyclin E, cdk2 and cdk4, however, was not impaired, suggesting that this cell cycle suppressor protein is functional. In addition, cyclin D1-, cdk2-, cdk4- and cyclin E-associated kinase activities were also lower in these estrogen-induced renal neoplasms than in untreated, age-matched kidneys. Interestingly, when compared with untreated kidney tissue, early and frank renal neoplasms had less of the 62 kDa native form of E2F1 and contained a 57 kDa variant form. Thus we have characterized an unusual deregulation of the cell cycle during estrogen-induced renal tumorigenesis in Syrian hamsters which still allows for estrogen-driven kidney tumor cell proliferation and may contribute to the early genomic instability found.

Animals↗
PubMed2000

[Reactivation of denatured lysozyme with immobilized molecular chaperones GroE].

The molecular chaperones GroEL and GroES were expressed in recombinant E. coli and purified by anion exchange chromatography. The renaturation of the denatured lysozyme with the free and immobilized GroEL/ES or GroEL was studied. We show here that using free GroEL alone could reactive the denatured lysozyme up to a relative activity of over 90%. The immobilized GroEL was also effective for promoting lysozyme refolding. Moreover, the optimal temperature (i.e., 37 degrees C) and (pH(i.e., 6 to 8) for the immobilizde GroEL-facilitated lysozyme refolding operation were determined. Under the optimal condition, the activity of lysozyme could be recovered up to 85%. In addition, the immobilized GroEL was repeatedly used five times without loss of its renaturation ability, indicating its potentiality to be used in practical downstream bioprocesses.

Chaperonin 10↗
PubMed2000

Effect of simulated weightlessness on phase II drug metabolism in the rat.

BACKGROUND: Exposure to weightlessness is known to alter physiological processes in humans and animals. As a result of these changes, hepatic drug metabolism may be altered as well. Indeed, short term simulated weightlessness in the rat has been shown to increase oxidative metabolism. HYPOTHESIS: Simulated weightlessness will increase Phase II drug metabolism in the rat during short-term tail suspension. METHODS: The tail-suspended rat model was used to simulate weightlessness. Rats were subjected to 1, 3, 7, or 10 d of tail-suspension in order to mimic the effect of exposure to a microgravity environment. One additional rat group was not suspended and served as a control. On the final day of the study, rats we administered a single intravenous bolus dose of acetaminophen 25 mg x kg(-1) through an implanted jugular catheter and serial blood samples were taken for 90 min. Serum acetaminophen concentrations were measured by high-performance liquid chromatography. Pharmacokinetic parameters were determined by using standard model independent methods. RESULTS: The results show that simulated weightlessness in the rat has no effect on Phase II drug metabolism, using acetaminophen as a marker compound. CONCLUSIONS: These data support the hypothesis that simulated weightlessness in the rat modulates oxidative metabolism, but not drug conjugation to glucuronide or sulfate metabolites. These data offer insight into the physiological changes and variability seen in hepatic metabolic profiles in humans and animals following actual spaceflight.

Acetaminophen↗
PubMed1999

Simple and rapid assay for acetaminophen and conjugated metabolites in low-volume serum samples.

The use of marker compounds for estimating drug metabolic capacity or pharmacokinetic parameters is common in the biological sciences. Often small laboratory animals are used and thus sample size is a limiting concern. In this report, we describe an assay we developed for measuring the concentration of acetaminophen and its conjugated metabolites in low-volume serum samples. Acetaminophen and metabolites were removed from 10 microl serum samples by a single-step 6% (v/v) perchloric acid deproteination using theophylline as internal standard. Samples were separated in a pH 2.2 sodium sulfate-acetonitrile mobile phase at a flow-rate of 1.5 ml/min on a 15 cm octadecylsilyl column at room temperature. Analytes were detected at a wavelength of 254 nm. The resulting chromatograms showed no interfering peaks from endogenous serum components. The concentration ranges measured were 1.56-200 microg/ml for acetaminophen and acetaminophen sulfate and 3.91-500 microg/ml for acetaminophen glucuronide. The assay was linear in the range of concentrations analyzed. The intra-day and inter-day coefficient of variation ranged from 0.4 to 8.2% and 0.2 to 12.3% for acetaminophen, 0.5 to 12.9% and 0.3 to 16.1% for acetaminophen glucuronide, and 0.4 to 8.1% and 0.2 to 14.3% for acetaminophen sulfate, respectively. Results from the experiments show that acetaminophen and its conjugated metabolites can easily and reproducibly be measured in low-volume serum samples and thus may offer an additional method to measure these compounds when the volume of biological samples may be limited.

Acetaminophen↗
PubMed1999

Tentative novel mechanism of the bystander effect in glioma gene therapy with HSV-TK/GCV system.

Although many works support gap junctional intercellular communication (GJIC) having a close relation to bystander cell killing in herpes simplex virus thymidine kinase (HSV-TK) gene and ganciclovir (GCV) treatment, our previous work suggested that other factors involved in bystander effect besides GJIC exist. To confirm our primary work, we evaluated the mode of the bystander cell (C6) co-cultured with TK-positive cells (TF10.2) in our designed "insert plates" in which two cell lines could be separated but share the same medium. Another method that we used was adding the supernatant from the medium of GCV-treated TF10.2 cells to the wild type C6. Growth inhibition of the bystander cells was observed despite the absence of GJIC. In addition, apoptotic cell death of TK+ cells and bystander cells was obvious. These studies suggested that other pathways besides cell-cell contacts may play a role in bystander cell killing; the factors released from TK-positive cells could induce apoptosis of bystander cells.

Apoptosis↗