Search PubMed⌕ Search

Biomedical subjects

L Shen

Publications and source records attributed to L Shen.

At least 55 records · Page 3Linked to original sources
PubMed2001

CpG island methylation in colorectal adenomas.

Methylation of cytosines in CpG islands silences gene expression. CpG island methylator phenotype (CIMP) in colorectal cancers is characterized by abnormal methylation of multiple CpG islands including those in several tumor suppressor genes such as p16, hMLH1, and THBS1. CpG island methylation has not been well characterized in adenomas. We evaluated methylation status at p16, MINT2, and MINT31 loci, which are frequently methylated in colorectal carcinomas, in 108 colorectal adenomas from a prospective study of 50 patients without cancer. Methylation at one or more loci was present in 48% (52 of 108) of adenomas with 25% (19 of 76) CIMP-high (two or more methylated loci) and 32% (24 of 76) CIMP-low (one methylated locus). The p16 gene was methylated in 27% (19 of 71) of adenomas. Methylation status of different adenomas from the same patient was not correlated (odds ratio, 0.93; P = 0.77). Adenomas with tubulovillous or villous histology were frequently methylated: 73% (17 of 26) versus 41% (35 of 85) of tubular adenomas (odds ratio, 3.46; P = 0.02). High levels of microsatellite instability were more frequent in adenomas without methylation (13% versus 2%; odds ratio, 8.48; P = 0.05). Our results indicate that methylation plays an important role early in colorectal tumorigenesis. CpG island methylation is more common in adenomas with tubulovillous/villous histology, a characteristic associated with more frequent predisposition to invasive carcinoma. Methylation is distinct from microsatellite instability and develops in individual adenomas rather than resulting from a field defect in an individual patient.

Adenoma↗
PubMed2001

Effect of human immunoglobulins on the immunogenicity of porcine bioprostheses.

BACKGROUND: Glutaraldehyde fixation (GT) is known to reduce immunologic reactions and tissue degeneration after implantation in humans. Sterilization after glutaraldehyde fixation (G-ST) improves the safety and reduces the tendency of GT valves to calcify. Intravenous immunoglobulins (IVIg) have been shown to reduce xenogeneic response against porcine tissue. We have investigated the effect of these fixation procedures combined with and without IVIg on the antigenicity of bioprostheses. METHODS: Lewis adult rats were implanted subcutaneously with a fresh, GT, or G-ST porcine heart valve pre- or posttreated with different amounts of IVIg. We followed by enzyme-linked immunosorbent assay and IgM and IgG titers against protein extracts from the porcine heart valves after implantation. Cellular reactivity was assessed in xenogeneic lymphoendothelial coculture experiments. Calcification content was also examined. RESULTS: Glutaraldehyde fixation partially decreased the humoral response against proteins of the implant but elicited a cellular xenogeneic response. Sterilization reduced these reactivities, but retained antigenicity. Intravenous immunoglobulin postincubated with GT valves before implantation reduced the antigenicity of the tissue to the same extent as G-ST valves, but had no effect on valvular tissue calcification. CONCLUSIONS: Our studies demonstrate that IVIg or the sterilization procedure (ST) reduced the cellular response against glutaraldehyde-fixed valves (GT), whereas reduced calcification was observed only with ST.

Animals↗
PubMed2001

Comparison of negative and positive ion electrospray tandem mass spectrometry for the liquid chromatography tandem mass spectrometry analysis of oxidized deoxynucleosides.

Oxidized deoxynucleosides are widely used as biomarkers for DNA oxidation and oxidative stress assessment. Although gas chromatography mass spectrometry is widely used for the measurement of multiple DNA lesions, this approach requires complex sample preparation contributing to possible artifactual oxidation. To address these issues, a high performance liquid chromatography (HPLC)-tandem mass spectrometric (LC-MS/MS) method was developed to measure 8-hydroxy-2'-deoxyguanosine (8-OH-dG), 8-hydroxy-2'-deoxyadenosine (8-OH-dA), 2-hydroxy-2'-deoxyadenosine (2-OH-dA), thymidine glycol (TG), and 5-hydroxy-methyl-2'-deoxyuridine (HMDU) in DNA samples with fast sample preparation. In order to selectively monitor the product ions of these precursors with optimum sensitivity for use during quantitative LC-MS/MS analysis, unique and abundant fragment ions had to be identified during MS/MS with collision-induced dissociation (CID). Positive and negative ion electrospray tandem mass spectra with CID were compared for the analysis of these five oxidized deoxynucleosides. The most abundant fragment ions were usually formed by cleavage of the glycosidic bond in both positive and negative ion modes. However, in the negative ion electrospray tandem mass spectra of 8-OH-dG, 2-OH-dA, and 8-OH-dA, cleavage of two bonds within the sugar ring produced abundant S1 type ions with loss of a neutral molecule weighing 90 u, [M - H - 90]-. The signal-to-noise ratio was similar for negative and positive ion electrospray MS/MS except in the case of thymidine glycol where the signal-to-noise was 100 times greater in negative ionization mode. Therefore, negative ion electrospray tandem mass spectrometry with CID would be preferred to positive ion mode for the analysis of sets of oxidized deoxynucleosides that include thymidine glycol. Investigation of the fragmentation pathways indicated some new general rules for the fragmentation of negatively charged oxidized nucleosides. When purine nucleosides contain a hydroxyl group in the C8 position, an S1 type product ion will dominate the product ions due to a six-membered ring hydrogen transfer process. Finally, a new type of fragment ion formed by elimination of a neutral molecule weighing 48 (CO2H4) from the sugar moiety was observed for all three oxidized purine nucleosides.

Animals↗
PubMed2001

GDNF acutely modulates excitability and A-type K(+) channels in midbrain dopaminergic neurons.

Glial cell line-derived neurotrophic factor (GDNF) prevents lesion-induced death of midbrain dopaminergic neurons, but its function in normal brain remains uncertain. Here we show that GDNF acutely and reversibly potentiated the excitability of cultured midbrain neurons by inhibiting transient A-type K(+) channels. The effects of GDNF were limited to large, tyrosine hydroxylase (TH)-positive dopaminergic neurons, and were mediated by mitogen associated protein (MAP) kinase. Application of GDNF also elicited a MAP kinase-dependent enhancement of the excitability in dopaminergic neurons in midbrain slice. These results demonstrate an acute regulation of GDNF on ion channels and its underlying signaling mechanism, and reveal an unexpected role of GDNF in normal midbrain dopaminergic neurons.

4-Aminopyridine↗
PubMed2001

Comparative study of different surgical procedures using sensory nerves or neurons for delaying atrophy of denervated skeletal muscle.

To observe the effect of sensory components on muscle atrophy, 4 surgical procedures applying sensory nerves or neurons to the denervated muscle were conducted in a rat model: sensory nerve implantation (group B), sensory motor nerve crossover (group C), dorsal root ganglia implantation (group D), and implantation of preganglionically avulsed sensory nerve (group E). Rats with complete denervation served as controls (group A). The degree of muscle atrophy was evaluated after surgery by fibrillation potential amplitude, muscle wet weight, muscle fiber cross-sectional area, collagen content, and protein content. Compared with group A, the results in groups B, D, and E were superior 1 month after surgery and the results in groups B, C, and E were superior 3 months after surgery. Implantation of normal or preganglionically avulsed sensory nerve delayed atrophy. Dorsal root ganglia implantation only had a short-term trophic influence. In the animal model setting, sensory motor nerve crossover is effective only after it is maintained for at least 3 months.

Animals↗
PubMed2001

In vivo T-lymphocyte activation and transient reduction of viral replication in macaques infected with simian immunodeficiency virus.

While it is well established that cellular activation can increase human immunodeficiency virus (HIV) replication in T lymphocytes, it is also clear that both activated CD8+ and CD4+ T lymphocytes mediate anti-HIV activity. To assess the relative importance of these contrary effects on HIV replication in vivo, we evaluated the consequences of Mycobacterium bovis BCG and staphylococcal enterotoxin B (SEB) inoculation in vivo in rhesus monkeys chronically infected with simian immunodeficiency virus of macaques (SIVmac). BCG inoculation induced as much as a 2.5-log reduction of plasma and intracellular SIV RNA in SIVmac-infected monkeys. This down-regulation of virus replication persisted as long as 4 weeks after BCG inoculation. Similarly, SEB injection resulted in up to a 3-log decrease in plasma and intracellular SIV RNA in SIVmac-infected macaques. Interestingly, the short-term reduction of viremia in these monkeys correlated with the peak in vivo production of SEB- and BCG-induced cytokine responses. However, no long-term clinical benefit was observed in the SIVmac-infected macaques. These studies provide in vivo evidence that potent T-cell stimulation driven by antigens other than the virus itself can, under some circumstances, mediate short-term reduction of viremia in AIDS virus-infected individuals.

Animals↗
PubMed2001

Antiretroviral agents restore Mycobacterium-specific T-cell immune responses and facilitate controlling a fatal tuberculosis-like disease in Macaques coinfected with simian immunodeficiency virus and Mycobacterium bovis BCG.

The contribution of immune reconstitution following antiretroviral treatment to the prevention or treatment of human immunodeficiency virus-related primary or reactivation tuberculosis remains unknown. Macaque models of simian immunodeficiency virus-Mycobacterium bovis BCG (SIV/BCG) coinfection were employed to determine the extent to which anti-Mycobacterium tuberculosis immunity can be restored by antiretroviral therapy. Both SIV-infected macaques with active BCG reinfection and naive animals with simultaneous SIV/BCG coinfection were evaluated. The suppression of SIV replication by antiretroviral treatment resulted in control of the active BCG infection and blocked development of the fatal SIV-related tuberculosis-like disease. The resolution of this disease coincided with the restoration of BCG purified protein derivative (PPD)-specific T-cell immune responses. In contrast, macaques similarly coinfected with SIV/BCG but not receiving antiretroviral therapy had depressed PPD-specific primary and memory T-cell immune responses and died from tuberculosis-like disease. These results provide in vivo evidence that the restoration of anti-mycobacterial immunity by antiretroviral agents can improve the clinical outcome of an AIDS virus-related tuberculosis-like disease.

Adenine↗
PubMed2001

Intestinal satiety protein apolipoprotein AIV is synthesized and regulated in rat hypothalamus.

Apolipoprotein AIV (apo AIV) is a satiety protein secreted by the small intestine. We demonstrate for the first time that apo AIV protein and apo AIV mRNA are present in rat hypothalamus, a site intimately involved in the integration of signals for regulation of food intake and energy metabolism. We further characterized the regulation of hypothalamic apo AIV mRNA levels. Food-deprived animals showed a pronounced decrease in gene expression of apo AIV in the hypothalamus, with a concomitant decrease in the jejunum. Refeeding fasted rats with standard laboratory chow for 4 h evokes a significant increase of apo AIV mRNA in jejunum but not in hypothalamus. However, lipid refeeding to the fasted animals restored apo AIV mRNA levels both in hypothalamus and jejunum. Intracerebroventricular administration of apo AIV antiserum not only stimulated feeding, but also decreased apo AIV mRNA level in the hypothalamus. These data further confirm the central role of apo AIV in the regulation of food intake.

Animals↗
PubMed2001

Defects in development of the kidney, heart and eye vasculature in mice homozygous for a hypomorphic Notch2 mutation.

The Notch gene family encodes large transmembrane receptors that are components of an evolutionarily conserved intercellular signaling mechanism. To assess the in vivo role of the Notch2 gene, we constructed a targeted mutation, Notch2(del1). Unexpectedly, we found that alternative splicing of the Notch2(del1) mutant allele leads to the production of two different in-frame transcripts that delete either one or two EGF repeats of the Notch2 protein, suggesting that this allele is a hypomorphic Notch2 mutation. Mice homozygous for the Notch2(del1) mutation died perinatally from defects in glomerular development in the kidney. Notch2(del1)/Notch2(del1 )mutant kidneys were hypoplastic and mutant glomeruli lacked a normal capillary tuft. The Notch ligand encoded by the Jag1 gene was expressed in developing glomeruli in cells adjacent to Notch2-expressing cells. We show that mice heterozygous for both the Notch2(del1) and Jag1(dDSL) mutations exhibit a glomerular defect similar to, but less severe than, that of Notch2(del1)/Notch2(del1 )homozygotes. The co-localization and genetic interaction of Jag1 and Notch2 imply that this ligand and receptor physically interact, forming part of the signal transduction pathway required for glomerular differentiation and patterning. Notch2(del1)/Notch2(del1 )homozygotes also display myocardial hypoplasia, edema and hyperplasia of cells associated with the hyaloid vasculature of the eye. These data identify novel developmental roles for Notch2 in kidney, heart and eye development.

Alleles↗
PubMed2001

OxLDL induces macrophage gamma-GCS-HS protein expression: a role for oxLDL-associated lipid hydroperoxide in GSH synthesis.

Oxidized LDL (oxLDL) produced a rapid depletion of intracellular glutathione (GSH) followed by an adaptive increase in J774 A.1 macrophages. OxLDL also induced a transient increase in the levels of gamma-glutamylcysteine synthetase heavy subunit (gamma-GCS-HS), representing the catalytic subunit of the rate-limiting enzyme for de novo GSH synthesis. The induction took place within 3 h, with maximum levels observed by 10 h of treatment. Pretreatment of oxLDL with ebselen inhibited GSH depletion and attenuated the gamma-GCS-HS induction. OxLDL-associated lipid hydroperoxides and their decomposition product aldehydes are two major components thought to account for GSH depletion in macrophages. Ebselen pretreatment had only a minor effect on malondialdehyde levels, whereas peroxide content was essentially abolished, suggesting that oxLDL-associated hydroperoxides may mediate both GSH depletion and gamma-GCS-HS induction. Acetylated LDL (AcLDL) also caused a moderate induction of gamma-GCS-HS protein along with a 30% transient increase in GSH by 3;-6 h, suggesting a minor involvement of scavenger receptor-mediated signaling of GSH synthesis. However, the level of gamma-GCS induction by AcLDL was insufficient to cause a sustained increase in GSH. Macrophages with higher glutathione peroxidase (GPx) activity experienced a more rapid and extensive depletion of GSH when treated with oxLDL under similar conditions, along with greater resistance to oxLDL- or peroxide-induced cytotoxicity. We conclude that oxLDL-associated peroxides are primarily responsible for GSH depletion, creating an oxidizing environment required for gamma-GCS induction and compensatory GSH synthesis. This is facilitated in cells expressing high GPx activity through a rapid depletion of GSH in the face of a peroxide challenge.

Antioxidants↗
PubMed2001

[Adverse effects of exposure to p-nonylphenol on reproductive system of young male rats].

OBJECTIVE: To study the adverse effects of exposure to p-nonylphenol (NP) during gestation and lactation on the reproductive system of young male rats. METHODS: Pregnant rats were administrated with p-nonylphenol, at doses of 0, 50, 100 [symbol: see text] 200 mg/kg of body weight, respectively, by gavage from the first day of gestation to the day of weaning, i.e., the 21st day after birth. The young male rats were killed at the age of 70 days and some indices related to their reproductive system (i.e., weight of the gonad, sperm counts of the epididymis, histology of the testes, et al.) were analyzed. RESULT: Weights of the testes and prostate gland decreased with the increase doses of exposed NP, their weights of the testes of 2.86, 2.98, 2.59 and 2.44 g, and those of the prostrate gland of 0.26, 0.23, 0.20 and 0.19 g, respectively. The same trend existed in daily production of sperm in the testes and sperm counts of the epididymis, with 22.46, 18.46, 17.43 and 17.26 x 10(6) sperm per gram of the testis, with sperm counts of 46.85, 39.74, 35.57 and 31.36 x 10(6) per gram of the epididymis tail. Separation of prepuce occurred on the 47.83th days and 46.31th days in the rats exposed to 200 mg/kg of NP and in the control group, respectively. CONCLUSION: NP could decrease their ability of spermatogenesis in the rats without morphological changes of spermatogenic cells of the testes and epididymis.

Animals↗
PubMed2001

Expression, Purification, Characterization of Amphioxus Insulin-like Peptide and Preparation of Polyclonal Antibody to It.

To elucidate the origination and evolution as well as structure-function relationship of insulin and IGF-1, the gene of amphioxus insulin-like peptide (ILP), a common ancestor of insulin and IGF-1, was chemically synthesized and cloned into the expression vector pVT102-U. In the recombinant ILP, the C-terminus of B-domain and the N-terminus of A-domain of amphioxus ILP deduced from cDNA were linked together by a tripeptide, Ala-Ala-Lys, and its B28Arg residue was also replaced by Lys to facilitate its conversion to a double-chain form by Lys-C cleavage. The expression vector was transformed into yeast cells and the recombinant ILP was expressed efficiently. The purified single-chain and double-chain ILP was obtained by fermentation, purification and enzymatic cleavage. Molecular weight measurement and amino acid composition analysis showed that the primary structure of ILP was correct. Circular dichroism analysis showed that the secondary and tertiary structure of double-chain ILP was similar to that of insulin, while double-chain ILP had no measurable insulin activity in insulin receptor binding assay. Additionally, New Zealand Rabbits were immunized with single-chain ILP and high titre polyclonal antibody was obtained. This work was important for further investigating the molecular evolution of insulin and IGF-1 and the distribution and existence of ILP in amphioxus.

Journal Article↗
PubMed2001

Expression, Purification and Functional Identification of Extracellular Part of Discoidin Domain Receptor 2.

Discoidin domain receptor 2 (DDR2) is a new type of receptor tyrosine kinases, and was thought to be involved in the metastasis of some tumors. Its ligand is fibrillar collagen. The activation of DDR2 induced by collagen mediates the over-expression of matrix metalloproteinase 1 (MMP-1) in cells. A specific inhibitor of DDR2 was necessary for the study of DDR2 function. Theoretically, a soluble receptor could possibly be used as specific inhibitor for the native receptor on cell membrane. In this report, a fragment (DB) of extracellular part of DDR2 was cloned and expressed for the use as potential inhibitor. This DB fragment corresponded to the polypeptide from the 23rd amino acid residue to the 293rd amino acid residue of DDR2. The fragment was amplified by RT-PCR from human lung cancer tissue, and the product was cloned into pMD18-T vector. After identification by sequence analysis, the fragment was sub-cloned into pGEX-4T-1 vector. Fusion protein of GST-DB was expressed in JM109 E.coli cells as expected and the soluble part accounted for about 13% of the total fusion protein. The soluble fusion protein was then purified with glutathione affinity resin, and GST-DB with purity of 86.1% was obtained. Competitive combination inhibitory test showed that the purified GST-DB inhibited the interaction between collagen II and DDR2 on the surface of RA synovial fibroblasts. Zymography analysis showed that the level of MMP-1 of both NIH 3T3 cell and RA synovial fibroblasts with collagen II-stimulation decreased after adding GST-DB fusion protein. The results indicated that the fusion protein GST-DB could inhibit the function of DDR2 on cells, and DDR2 might mediate collagen II-induced over-expression of MMP-1 in these cells.

Journal Article↗
PubMed2001

Fusion Expression and Purification of OSBP PH Domain and Preliminary Analysis of Its Second Structure.

Oxysterol binding protein (OSBP) is a regulator of oxysteroid metabolism. To investigate the function and the structure-function relationship of OSBP, the recombinant vector OSBP PH-pRSET-A was transformed into E.coli JM109(DE3), and the strain highly expressing soluble 6His-OSBP PH domain in minimal medium were obtained. The fusion protein was purified by Ni(2 )-NTA agarose beads. The secondary structure of the purified 6His-OSBP PH domain fusion protein was analysed by circular dichronism. The results indicated the PH domain was composed of alpha-helix 7.2%, beta-pleated sheets 71.1% and radom coil 21.7%.

Journal Article↗
PubMed2001

[Effect of dietary protein on the function and morphology of diaphragm in rats with experimental emphysema].

The effects of dietary protein on the function and morphology of diaphragm of rats with emphysema were observed in 50 SD rats, 10 rats in each group. The emphysema in groups E(emphysema control), HP(emphysema with high protein diet) and LP(emphysema with low protein diet) was induced by intra-tracheal instillation of elastase (750 U/kg BW). The control rat (C1, C2) were instilled with normal saline. After 6 weeks of experiment, the total lung volume and the average area of alveolus was significantly increased in the group E compared with group C1 (P < 0.05) and the number of alveolus per unit area was also reduced obviously (P < 0.05). The average contractile force of diaphragm at a stimulus rate of 20 Hz expressed as a ratio of the maximal contractile force (F20/Fmax) was increased significantly in group E compared with group C (P < 0.01). The cross-sectional area of slow-twitch fibers increased significantly in group E. After 10 weeks of oral supplementation with 24% casein(HP) or 8% casein (LP) to the rats with emphysema, the contractility of the diaphragm in group LP was lower than that in the control group and the HP group. It was concluded that high protein diet might be beneficial to maintain the function of diaphragm in patients with emphysema.

Animals↗
PubMed2001

[Clonning and identification of the gene encoding specific IgE antibody-related antigen of Schistosoma japonicum].

OBJECTIVE: To obtain the gene encoding specific IgE-related antigen of Schistosoma japonicum from the cDNA library of adult Schistosoma japonicum. METHODS: The pooled sera from 15 individuals with high levels of specific IgE antibody against SWAP were absorbed with Protein-G and used for screening of IgE-related antigen from the adult worm cDNA library of Schistosoma japonicum. The inserted cDNA was amplified by PCR and sequenced. According to the first reading frame of the sequence, a pair of new probes, in which EcoR I and Not I sites were incorporated respectively, were designed and used to amplify the target gene. Then, the gene was cloned into vector pGEM-T and subcloned into expression vector pGEX-6p-1. The fusion protein was expressed, analysed by SDS-PAGE and identified by Western blotting with the specific IgE antibody, respectively. RESULTS: The inserted cDNA fragment from the positive clone was about 1,200 bp, with the ORF of 507 bp which encoded 169 amino acids. The deduced molecular weight of the recombinant protein was 19.3 kDa. The homology between the target gene (Sj43B) and other known DNA sequences was less than 40%. The fusion protein expressed by the recombinant vector pGEX-6p-1/Sj43B could be recognized by schistosome specific IgE antibody. CONCLUSION: Sj43B may encode the specific IgE-related antigen of Schistosoma japonicum. The successful construction of recombinant plasmid pGEX-6p-1/Sj43B lay the groundwork for further studies on immunological characteristics and protection immunity of the recombinant protein.

Animals↗
PubMed2001

[Spectral interference correction software and its application in ICP-4 spectroscopic instrument].

In this paper, the spectral interference correction software based on Kalman filtering and its application in ICP-4 spectroscopic instrument are introduced. The principle, algorithm and experimental results of the software are discussed. The three analyte lines P (213.618 nm), P(214.914 nm) and Mo(268.323 nm) interforced by Cu(213.598 nm), Cu(214.897 nm) and V(268.309 nm) respectively were chosen as typical examples. The experiment results prove the validity of the software.

Algorithms↗