Biomedical subjects

Jun Li

Publications and source records attributed to Jun Li.

At least 415 records · Page 23Linked to original sources

PubMed2004

Molecular mechanism for regulation of the human mitochondrial branched-chain alpha-ketoacid dehydrogenase complex by phosphorylation.

The human mitochondrial branched-chain alpha-ketoacid dehydrogenase complex (BCKDC) is a 4 MDa macromolecular machine comprising three catalytic components (E1b, E2b, and E3), a kinase, and a phosphatase. The BCKDC overall activity is tightly regulated by phosphorylation in response to hormonal and dietary stimuli. We report that phosphorylation of Ser292-alpha in the E1b active site channel results in an order-to-disorder transition of the conserved phosphorylation loop carrying the phosphoryl serine. The conformational change is triggered by steric clashes of the phosphoryl group with invariant His291-alpha that serves as an indispensable anchor for the phosphorylation loop through bound thiamin diphosphate. Phosphorylation of Ser292-alpha does not severely impede the E1b-dependent decarboxylation of alpha-ketoacids. However, the disordered loop conformation prevents phosphorylated E1b from binding the E2b lipoyl-bearing domain, which effectively shuts off the E1b-catalyzed reductive acylation reaction and therefore completely inactivates BCKDC. This mechanism provides a paradigm for regulation of mitochondrial alpha-ketoacid dehydrogenase complexes by phosphorylation.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗

PubMed2004

Endotoxin differentially modulates the basolateral and apical sodium/proton exchangers (NHE) in enterocytes.

BACKGROUND: Maintenance of enterocyte activity during extracellular acidosis requires functional sodium/proton exchangers (NHE), which are present at both basolateral and apical surfaces. Necrotizing enterocolitis is characterized by systemic hypoperfusion, metabolic acidosis, and the apical to basolateral translocation of endotoxin (lipopolysaccharide [LPS]). We hypothesized that LPS differentially impairs NHE activity at the basolateral or apical domains of enterocytes, leading to cellular acidification, and explored the mechanisms involved. METHODS: Experimental necrotizing enterocolitis (NEC) was induced in newborn rats using a combination of gavage feeds and hypoxia. NHE isoforms were assessed in primary and cultured enterocytes by Western blot analysis and by confocal microscopy in the presence or absence of LPS. NHE activity was detected by single-cell fluorescent ratiometric imaging with the use of the pH-sensitive dye 2', 7'-bis-(2carboxyethyl) -5-(and-6)-carboxyfluorescein. RESULTS: In both NEC and control rats, NHE1 was basolateral and NHE3 was apical. A similar distribution was observed in polarized IEC-6 cells. LPS caused a dose-dependent reduction in basolateral NHE1 activity in IEC-6 cells, but had no effect on apical NHE3 activity. This effect could not be accounted for by reduced expression or impaired plasma membrane localization of NHE isoforms. Strikingly, LPS-mediated NHE1 impairment caused marked cytoplasmic acidification under conditions of extracellular acidosis, whereas functional NHE1 maintained cytoplasmic pH homeostasis in control cells. CONCLUSIONS: LPS selectively impairs basolateral NHE1 but not apical NHE3, leading to cytoplasmic acidification during extracellular acidosis. This effect could mediate impaired enterocyte function after LPS translocation and suggests a mechanism leading to barrier disruption in NEC.

Acidosis↗

PubMed2004

The proteasome inhibitor, PS-341, causes cytokeratin aggresome formation.

Mallory body (MB) experimental induction takes 10 weeks of drug ingestion. Therefore, it is difficult to study the dynamics and mechanisms involved in vivo. Consequently, an in vitro study was done using primary tissue culture of hepatocytes from drug-primed mice livers in which MBs had already formed. The hypothesis to be tested was that MBs are cytokeratin aggresomes, which form when hepatocytes have a defective ubiquitin-proteasome pathway by which turnover of cytokeratin proteins is prevented. To test this hypothesis, primary tissue cultures of the hepatocytes from normal and MB-forming livers were incubated with the proteasome inhibitor PS-341 and then the cytokeratin filaments and the filament connecting proteins, that is, beta-actin, and ZO1, were visualized by immunofluorescence microscopy. PS-341 caused detachment of the cytokeratins from the cell surface plasma membrane. The cytokeratin filaments retracted toward the nucleus and cytokeratin aggresomes formed. In human livers, MBs showed colocalization of cytokeratin-8 (CK-8) with ubiquitin but not with beta-actin or ZO1. Mouse hepatoma cell lines were studied using PS-341 to induce cytokeratin aggresome formation. In these cell lines, the cytokeratin filaments first retracted toward the nucleus then formed cytokeratin-ubiquitin aggresomes polarized at one side of the nucleus. At the same time, the cells became dissociated from each other, however. The results simulated MB formation. MBs differ from cytokeratin aggresomes both morphologically and in ultrastructure.

Actins↗

PubMed2004

Proteasome inhibition induces cytokeratin accumulation in vivo.

Chronic ethanol ingestion leads to inhibition of proteasomal activity. As a consequence, proteins accumulate in liver cells. Cytokeratin accumulation as seen in alcoholic hepatitis could lead to the formation of Mallory bodies. In order to study the phenomenon of cytokeratin accumulation in liver cells, rats were fed ethanol or dextrose for 1 month and some were given the proteasome inhibitor, PS-341, to augment the inhibitory effect of ethanol feeding. This was done to study the involvement of proteasome inhibition in the process of cytokeratin accumulation. There was a marked increase in the accumulation of polyubiquitinated proteins, and heat shock proteins (hsp) 25 and 70 in the liver of rats treated with PS-341. Similarly, cytokeratin-8 (CK-8) levels were markedly increased in the liver homogenates of rats fed ethanol when given PS-341. When normal mouse cultured hepatocytes were transfected with cytokeratin-18 (CK-18) tagged with red fluorescent protein (RFP), CK-18 aggresomes formed because proteasome was overloaded. These data provide new evidence that proteasome inhibition is involved in cytokeratin accumulation, when aggresomes are formed in tissue culture. Accumulation of cytokeratin in this way may ultimately lead to Mallory body formation as seen in alcoholic hepatitis.

Alcoholism↗

PubMed2004

p62 is involved in the mechanism of Mallory body formation.

p62 is a scaffolding protein that binds to polyubiquitin. It is involved in the degradation of proteins by the proteasome. To determine if p62 is critical in the development of Mallory bodies (MBs), primary culture hepatocytes from drug-primed mice were studied and the results were compared with normal hepatocytes. Gene-specific gripNA (gp62) was added to the medium of the primary cultures of the hepatocytes to inhibit the expression of p62. Overexpression of p62 was achieved by transfecting the hepatocytes with a plasmid containing green fluorescent protein (GFP) fused p62 (p62-GFP). Gp62 dramatically inhibited MB formation by 94% in drug-primed hepatocytes. The cells transfected with gp62 had decreased protein levels of p62, ubiquitin (Ub), and cytokeratin 8 (CK8). Overexpression of p62 accelerated and enhanced MB formation by 339% in drug-primed hepatocytes. Overexpression of p62 in normal mouse hepatocytes induced MB-like aggresomes that were stained by Ub but not by CK8. The results indicate that p62 is involved in the mechanism of MB formation.

Animals↗

PubMed2004

Liver hypoxia and lack of recovery after reperfusion at high blood alcohol levels in the intragastric feeding model of alcohol liver disease.

The purpose of this study was to test for the presence of liver hypoxia and recovery after reperfusion when blood alcohol levels (BAL) are high. Male rats were fed ethanol intragastrically at a constant rate for 1 month. The pO(2) levels were then measured on the liver surface of these rats, in vivo during laparatomy under isoflurane anesthesia. To measure the response to acute hypoxia, the hepatic blood flow was clamped off at the porta hepatis. When the clamp was released, recovery from hypoxia was measured. A number of hypoxic-inducible genes in the liver were analyzed by means of quantitative RT-PCR as a measure of increased activation of hypoxia initiated transcription. The mRNA levels of genes for adrenomedullin, adrenergic receptor alpha, 1a and 1d, CDK inhibitor 1a, and erythropoietin were all significantly higher at the peaks than troughs. Expression of these same genes in the livers of control rats fed dextrose was lower than at the troughs. Although the mRNA level of the hypoxia-inducible factor (HIF-1alpha) was higher at the trough than at the peak, its protein concentration in the nuclear fraction was not increased at the troughs compared with the peaks. In fact, the nuclear protein level of HIF-1alpha at the peak was significantly higher than in control samples, which is consistent with the presence of hypoxia at the peaks. Further analysis of the HIF-alpha degradation regulation revealed that prolyl 4-hydroxylase (P4ha1) and von Hippel-Lindau syndrome homolog (Vhl) were both up-regulated at the troughs compared with the peaks. The liver surface oxygen levels at the peaks were reduced compared with the control samples. The pO(2) levels fell abruptly when the vessels at the porta hepatis were clamped. When the clamp was removed, allowing reperfusion of the liver, pO(2) returned to baseline levels in the control, and at the troughs but not at the peaks. These results support the hypothesis that hypoxia occurs at the peaks of the BAL cycle and recovery from ischemia is impaired at the peaks.

Administration, Oral↗

PubMed2004

Sesterterpenoids from the marine sponge Hyrtios erectus.

Four new sesterterpenes, namely, (+)-20-formylhyrtiosal (1), (+)-16-O-acetyl-20-formylhyrtiosal (2), 12-alpha-O-acetylhyrtiolide (3), and 5,10-dihydroxyfurospinulosine-1 (4), together with seven known sesterterpenes (5-11) were isolated from the marine sponge Hyrtios erectus collected at Hainan, China. The structures were elucidated on the basis of extensive spectroscopic analysis involving mainly one- and two-dimensional NMR as well as mass spectroscopy.

Animals↗

PubMed2004

Chemical constituents from the mangrove plant, Aegiceras corniculatum.

From the stems and twigs of the mangrove plant, Aegiceras corniculatum, seven new compounds, namely, 2-methoxy-3-nonylresorcinol (1), 5-O-ethylembelin (2), 2-O-acetyl-5-O-methylembelin (3), 3,7-dihydroxy-2,5-diundecylnaphthoquinone (4), 2,7-dihydroxy-8-methoxy-3,6-diundecyldibenzofuran-1,4-dione (5), 2,8-dihydroxy-7-methoxy-3,9-diundecyldibenzofuran-1,4-dione (6), and 10-hydroxy-4-O-methyl-2,11-diundecylgomphilactone (7), were isolated together with three known compounds, 5-O-methylembelin (8), 3-undecylresorcinol, and 2-dehydroxy-5-O-methylembelin. The structures of 1-7 were determined by spectroscopic methods. Compound 2 and 5-O-methylembelin showed in vitro cytotoxicity against the HL-60, Bel(7402), U937, and Hela cell lines.

Antineoplastic Agents, Phytogenic↗

PubMed2004

Isomalabaricane-type compounds from the marine sponge Rhabdastrella aff. distincta.

Fractionation of the marine sponge Rhabdastrella aff. distincta led to the isolation and characterization of four new isomalabaricane analogues, isogeoditin A (1), 13-(E)-isogeoditin A (2), isogeoditin B (3), and 22,23-dihydrostellettin B (4), along with seven known isomalabaricane derivatives (5-11). Methylation of 10 and 11 afforded methyl esters 12 and 13. The structures of compounds 1-4 were determined on the basis of spectroscopic data analysis. All compounds were tested against a small panel of human tumor cell lines.

Animals↗

PubMed2004

Microelectronic DNA assay for the detection of BRCA1 gene mutations.

Mutations in BRCA1 are characterized by predisposition to breast cancer, ovarian cancer and prostate cancer as well as colon cancer. Prognosis for this cancer survival depends upon the stage at which cancer is diagnosed. Reliable and rapid mutation detection is crucial for the early diagnosis and treatment. We developed an electronic assay for the detection of a representative single nucleotide polymorphism (SNP), deletion and insertion in BRCA1 gene by the microelectronics microarray instrumentation. The assay is rapid, and it takes 30 minutes for the immobilization of target DNA samples, hybridization, washing and readout. The assay is multiplexing since it is carried out at the same temperature and buffer conditions for each step. The assay is also highly specific, as the signal-to-noise ratio is much larger than recommended value (72.86 to 321.05 vs. 5) for homozygotes genotyping, and signal ratio close to the perfect value 1 for heterozygotes genotyping (1.04).

Adult↗

PubMed2004

A 771726, the active metabolite of leflunomide, inhibits TNF-alpha and IL-1 from Kupffer cells.

This study was conducted to investigate the importance of direct contact between Kupffer cells (KCs) and hepatocytes (HCs) during the hepatic inflammatory responses, and the effect of leflunomide's active metabolite, A771726, on cytokines in KCs and HCs (DC cocultures) and KC cultures using an in vitro approach. Lipopolysaccharide (LPS)-induced inflammatory response in monocultures of rats HCs and KCs were compared with DC cocultures. Tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1) concentrations of different culture supernatants were measured with ELISA. TNF-alpha and IL-1 mRNA in KCs of inflammatory liver injury was analyzed with reverse transcription polymerase chain reaction (RT-PCR). Our data showed that DC cocultures exhibited the highest production of TNF-alpha and IL-1 compared with other cultures, and these cytokines were mainly produced by KCs, in particular activated KCs. Time course studies revealed an increased production of TNF-a preceding the IL-1 production, suggesting that increased TNF-alpha levels could be involved in the increased IL-1 production. Leflunomide's active metabolite, A771726, has significantly inhibitory effect on TNF-a and IL-1 at protein and transcription levels, and reduced production of IL-1 by A771726 was associated with inhibitory action of A771726 on TNF-alpha. These results provided evidence that leflunomide significantly inhibited TNF-alpha and IL-1 from KCs.

Aniline Compounds↗

PubMed2004

Gender-specific gene expression in post-mortem human brain: localization to sex chromosomes.

Gender differences in brain development and in the prevalence of neuropsychiatric disorders such as depression have been reported. Gender differences in human brain might be related to patterns of gene expression. Microarray technology is one useful method for investigation of gene expression in brain. We investigated gene expression, cell types, and regional expression patterns of differentially expressed sex chromosome genes in brain. We profiled gene expression in male and female dorsolateral prefrontal cortex, anterior cingulate cortex, and cerebellum using the Affymetrix oligonucleotide microarray platform. Differentially expressed genes between males and females on the Y chromosome (DBY, SMCY, UTY, RPS4Y, and USP9Y) and X chromosome (XIST) were confirmed using real-time PCR measurements. In situ hybridization confirmed the differential expression of gender-specific genes and neuronal expression of XIST, RPS4Y, SMCY, and UTY in three brain regions examined. The XIST gene, which silences gene expression on regions of the X chromosome, is expressed in a subset of neurons. Since a subset of neurons express gender-specific genes, neural subpopulations may exhibit a subtle sexual dimorphism at the level of differences in gene regulation and function. The distinctive pattern of neuronal expression of XIST, RPS4Y, SMCY, and UTY and other sex chromosome genes in neuronal subpopulations may possibly contribute to gender differences in prevalence noted for some neuropsychiatric disorders. Studies of the protein expression of these sex-chromosome-linked genes in brain tissue are required to address the functional consequences of the observed gene expression differences.

Actins↗

PubMed2004

A practical method to experimentally evaluate the Hausdorff dimension: an alternative phase-transition-based methodology.

We introduce a methodology to estimate numerically the Hausdorff dimension of a geometric set. This practical method has been conceived as a subsequent tool of another context study, associated to our concern to distinguish between various fractal sets. Its conception is natural since it can be related to the original idea involved in the definitions of Hausdorff measure and Hausdorff dimension. It is based on the critical behavior of the measure spectrum functions of the set around its Hausdorff dimension value. We illustrate on several well-known examples, the ability of this method to accurately estimate the Hausdorff dimension. Also, we show how the transition property, exhibited by the quantities used as substitutes of the Hausdorff measure in the corresponding fractal dimension relationships, can be used to accurately estimate the fractal dimension. To show the potential of our method, we also report the results of Hausdorff dimension measurements on some typical examples, compared to a direct application of the scaling relation involved in the box-counting dimension definition.

Journal Article↗

PubMed2004

Cerebroside analogues from marine-derived fungus Aspergillus flavipes.

From the mycelium of the marine-derived fungus Aspergillus flavipes, isolated from the sea anemone Anthopleura xanthogrammica, two new cerebroside analogues, namely flavicerebrosides A (1): [(2S,2'R,3R,4E,8E)-N-2'-hydroxyoctadecanoyl-1-O-beta--galactopyranosyl-9-methyl-4,8-sphingadienine], and B (2): [(2S,2'R,3R,3'E,4E,8E)-N-2'-hydroxy-3'-octadecenoyl-1-O-beta--galactopyranosyl-9-methyl-4,8-sphingadienine], together with two known glycosphingolipids cerebrosides D (3) and C (4), were isolated. Their structures were identified by means of extensive spectroscopic analysis (IR, UV, 2D NMR, MS, CD) and chemical degradation. All four compounds showed cytotoxic activity against the KB cell line.

Antineoplastic Agents↗

PubMed2004

Identification of genetic networks.

In this report, we propose the use of structural equations as a tool for identifying and modeling genetic networks and genetic algorithms for searching the most likely genetic networks that best fit the data. After genetic networks are identified, it is fundamental to identify those networks influencing cell phenotypes. To accomplish this task we extend the concept of differential expression of the genes, widely used in gene expression data analysis, to genetic networks. We propose a definition for the differential expression of a genetic network and use the generalized T2 statistic to measure the ability of genetic networks to distinguish different phenotypes. However, describing the differential expression of genetic networks is not enough for understanding biological systems because differences in the expression of genetic networks do not directly reflect regulatory strength between gene activities. Therefore, in this report we also introduce the concept of differentially regulated genetic networks, which has the potential to assess changes of gene regulation in response to perturbation in the environment and may provide new insights into the mechanism of diseases and biological processes. We propose five novel statistics to measure the differences in regulation of genetic networks. To illustrate the concepts and methods for reconstruction of genetic networks and identification of association of genetic networks with function, we applied the proposed models and algorithms to three data sets.

Algorithms↗

PubMed2004

The effect of blocking a nutritional pathway to the intervertebral disc in the dog model.

BACKGROUND: The hypothesis that injecting bone cement adjacent to one or both endplates would bring about degeneration in the intervening disc was tested. METHODS: In 11 dogs, bone cement was injected just below the superior endplates of L1, L2, and L3 to block the nutritional supply through these endplates to the three intervertebral discs T13-L1, L1-L2, and L2-L3. In one other dog, both the superior and the inferior endplates of the same discs (T13-L1, L1-L2, and L2-L3) were blocked with bone cement. All 12 dogs were euthanized between 31 and 70 weeks after the surgery. The three experimental discs (T13-L1, L1-L2, and L2-L3) and two control discs (T12-T13 and L4-L5) were excised and assessed using enzyme-linked immunosorbent assay (ELISA) and histology. RESULTS: Radiographs of the lumbar spine at the time of death did not show any signs of disc bulging, disc space narrowing, or peripheral osteophyte formation in any of the 12 dogs. The experimental discs as well as the control discs appeared normal in every dog. After the discs were bisected, they were carefully inspected for any visible signs of degeneration. The experimental discs showed no clear signs of disc degeneration and were not distinguishable from the control discs on a gross level. The numerical results from the ELISA showed that in the experimental discs as opposed to the control discs, there were significant increases in proteoglycan content in both the nucleus (P = 0.033) and annulus (P = 0.01) and clear histologic changes in some of the discs. CONCLUSION: The results show that injecting bone cement adjacent to one or both endplates for up to 70 weeks does not produce degeneration in any visible form in the intervening disc. There were no disc bulging, no apparent annular fissures, and no disc spacing narrowing. There were, however, increases in protoglycan content in both the nucleus and the annulus and clear histologic changes in some of the discs.

Animals↗

PubMed2004

Effect of bone morphogenetic protein-2 (BMP-2) on matrix production, other BMPs, and BMP receptors in rat intervertebral disc cells.

OBJECTIVE: An in vitro experiment study using rat disc cells was carried out to determine the effect of bone morphogenetic protein-2 (BMP-2) on extracellular matrix production, other BMPs, and BMP receptors (BMPRs). METHODS: Cells from the anulus fibrosus and transition zone were harvested and cultured. When the cells reached 80% confluence, BMP-2 was added to reach a final concentration of 200 ng/mL. Three days later, the culture media were collected for the assay of sulfated glycosaminoglycans (sGAG) and collagen types I and II. The cells were harvested for RNA extraction to determine the genes expressed. All experiments were performed at least three times to ensure repeatability. RESULTS: BMP-2 significantly increased aggrecan and collagen type II mRNA expression 8.30 and 4.61 times, respectively, and decreased versican mRNA expression 0.54 times as compared with control. Collagen type I production and mRNA level were not changed. BMP-2 significantly increased transforming growth factor-beta1 (TGFbeta1) and BMP-7 mRNA expression 2.32 and 2.45 times, respectively, compared with control. There was no significant change in BMP-6 mRNA expression. BMPR type IB and II mRNA expressions were increased and BMPR type 1A mRNA expression was decreased, but none of these differences was significant. CONCLUSIONS: The results of this study show that in rat intervertebral disc cells, BMP-2 increases aggrecan and collagen type II mRNA expression and decreases versican gene expression. BMP-2 also up-regulates mRNA expression for BMP-7 and TGFbeta but has no significant effect on the BMPRs.

Aggrecans↗

PubMed2004

Association of norepinephrine transporter gene with methylphenidate response.

OBJECTIVE: This study aimed to explore the association between alleles of the norepinephrine transporter gene and the methylphenidate response. METHOD: Chinese Han youths with attention-deficit/hyperactivity disorder recruited in the Outpatient Department of the Institute of Mental Health from 2001 to 2004 were treated with methylphenidate in doses of 0.45 to 0.60 mg/kg per day. Behavior changes were measured by the ADHD Rating Scale-IV when optimal doses were reached. The single-nucleotide polymorphic allele of norepinephrine transporter gene G1287A was used as the genetic marker, and the genotypes were identified by restriction fragment length polymorphism analysis. RESULTS: A significant association was found between norepinephrine transporter gene G1287A genotypes and response to methylphenidate for hyperactive-impulsive subscale scores (mean score reduction was 7.15 and 6.94 for G/G and G/A genotype, respectively, and 2.13 for A/A; p = .012) but not inattentive scores. CONCLUSIONS: These findings suggest an association between the G1287A polymorphism of the norepinephrine transporter gene and variation in methylphenidate response. Further work is needed to confirm this finding and to assess its generalization to other ethnic groups.

Adolescent↗