Biomedical subjects

R Lu

Publications and source records attributed to R Lu.

At least 19 recordsLinked to original sources

PubMed2001

Monocyte differentiation to macrophage requires interferon regulatory factor 7.

Interferon regulatory factors are a growing family of transcription factor that have been implicated in cellular events such as cell-growth regulation, antiviral defense, and development of the immune system. Interferon regulatory factor 7 (IRF-7) is expressed predominantly in lymphoid tissues and has been studied extensively in the context of viral infection and the induction of interferon and cytokine gene expression. In this paper, the involvement of IRF-7 in monocyte differentiation was examined in U937, HL60, and human primary macrophages. We report the induction of IRF-7 expression by 12-O-tetradecanoylphorbol-13-acetate in U937 and HL60 cells and demonstrate that this induction is essential for the monocyte differentiation to macrophages. We show that the monocyte differentiation is inhibited in cells expressing a dominant negative IRF-7 mutant, as evidenced by decreased expression of two macrophage-differentiation markers, CD11b and CD11c, and impaired phagocytic activity. In addition, we demonstrate that overexpression of IRF-7 is sufficient to trigger monocyte differentiation and to induce cell cycle arrest. The identification of IRF-7 as a key regulator in monocyte differentiation suggests a novel function of IRF-7 in innate immunity.

Blotting, Northern↗

PubMed2001

Olig bHLH proteins interact with homeodomain proteins to regulate cell fate acquisition in progenitors of the ventral neural tube.

BACKGROUND: Organizing signals such as Sonic hedgehog are thought to specify neuronal subtype identity by regulating the expression of homeodomain proteins in progenitors of the embryonic neural tube. One of these, Nkx2.2, is necessary and sufficient for the development of V3 interneurons. RESULTS: We report that Olig genes, encoding basic helix-loop-helix (bHLH) proteins, are expressed in a subset of Nkx2.2 progenitors before the establishment of interneurons and oligodendroglial precursors. Gain-of-function analysis in transgenic mouse embryos indicates that Olig genes specifically inhibit the establishment of Sim1-expressing V3 interneurons. Moreover, coexpression of Olig2 with Nkx2.2 in the chick neural tube generated cells expressing Sox10, a marker of oligodendroglial precursors. Colocalization of Olig and Nkx2.2 proteins at the dorsal extent of the Nkx2.2 expression domain is consistent with regulatory interactions that define the potential of progenitor cells in the border region. CONCLUSIONS: Interactions between homeodomain and Olig bHLH proteins evidently regulate neural cell fate acquisition and diversification in the ventral neural tube. In particular, interactions between Olig and Nkx2.2 proteins inhibit V3 interneuron development and promote the formation of alternate cell types, including those expressing Sox10.

Animals↗

PubMed2001

Delayed cardioprotection by intestinal preconditioning is mediated by calcitonin gene-related peptide.

Previous studies have shown that nitric oxide and calcitonin gene-related peptide (CGRP) are involved in mediation of the delayed cardioprotection of ischemic or pharmacological preconditioning, and nitric oxide can evoke the release of CGRP. In the present study, we examined the role of CGRP in nitric oxide-mediated delayed cardioprotection by brief intestinal ischemia in rats. The serum concentration of creatine kinase and infarct size were measured after 45-min coronary artery occlusion and 180-min reperfusion. Ischemic preconditioning was induced by six cycles of 4-min ischemia and 4-min reperfusion of the small intestine. Pretreatment with intestinal ischemic preconditioning for 24, 48, or 72 h significantly reduced infarct size and creatine kinase release, and the effects of ischemic preconditioning were completely abolished by L-nitroarginine methyl ester (L-NAME, 10 mg/kg, i.p.), an inhibitor of nitric oxide synthase, or by pretreatment with capsaicin (50 mg/kg, s.c.), which selectively depletes transmitters in capsaicin-sensitive sensory nerves. Intestinal preconditioning caused a significant increase in plasma concentrations of CGRP, and the effect was also abolished by L-NAME or capsaicin. These results suggest that the delayed cardioprotection afforded by intestinal ischemic preconditioning is mediated by endogenous CGRP via the nitric oxide pathway.

Animals↗

PubMed2001

Calcitonin gene-related peptide-mediated ischemic preconditioning in the rat heart: influence of age.

In the present study, we examined whether age-related reduction of ischemic preconditioning is related to calcitonin gene-related peptide (CGRP) release in the rat heart. Thirty minutes of global ischemia and 40 min of reperfusion caused a significant decrease of cardiac function and a marked increase of creatine kinase (CK) release at 2, 6 and 20 months of age. Ischemic preconditioning and pretreatment with CGRP for 5 min significantly improved cardiac function and reduced CK release during reperfusion at 2 and 6 months of age but not at 20 months of age. The content of CGRP in the coronary effluent during ischemic preconditioning was significantly increased in the first cycle at 2, 6 months of age but not at 20 months of age. These results suggest that the protection afforded by ischemic preconditioning is decreased in aging hearts, and the age-related change may be related to reduction of the release and effect of CGRP in the rat heart.

Aging↗

PubMed2001

Zonula occludens toxin structure-function analysis. Identification of the fragment biologically active on tight junctions and of the zonulin receptor binding domain.

Zonula occludens toxin (Zot) is an enterotoxin elaborated by Vibrio cholerae that increases intestinal permeability by interacting with a mammalian cell receptor with subsequent activation of intracellular signaling leading to the disassembly of the intercellular tight junctions. Zot localizes in the bacterial outer membrane of V. cholerae with subsequent cleavage and secretion of a carboxyl-terminal fragment in the host intestinal milieu. To identify the Zot domain(s) directly involved in the protein permeating effect, several zot gene deletion mutants were constructed and tested for their biological activity in the Ussing chamber assay and their ability to bind to the target receptor on intestinal epithelial cell cultures. The Zot biologically active domain was localized toward the carboxyl terminus of the protein and coincided with the predicted cleavage product generated by V. cholerae. This domain shared a putative receptor-binding motif with zonulin, the Zot mammalian analogue involved in tight junction modulation. Amino acid comparison between the Zot active fragment and zonulin, combined with site-directed mutagenesis experiments, confirmed the presence of an octapeptide receptor-binding domain toward the amino terminus of the processed Zot.

Amino Acid Sequence↗

PubMed2001

Epiplakin, a novel member of the Plakin family originally identified as a 450-kDa human epidermal autoantigen. Structure and tissue localization.

A 450-kDa human epidermal autoantigen was originally identified as a protein that reacted with the serum from an individual with a subepidermal blistering disease. Molecular cloning of this protein has now shown that it contains 5065 amino acids and has a molecular mass of 552 kDa. As reported previously this protein, which we call epiplakin, belongs to the plakin family, but it has some very unusual features. Epiplakin has 13 domains that are homologous to the B domain in the COOH-terminal region of desmoplakin. The last five of these B domains, together with their associated linker regions, are particularly strongly conserved. However, epiplakin lacks a coiled-coil rod domain and an amino-terminal domain, both of which are found in all other known members of the plakin family. Furthermore, no dimerization motif was found in the sequence. Thus, it is likely that epiplakin exists in vivo as a single-chain structure. Epitope mapping experiments showed that the original patient's serum recognized a sequence unique to epiplakin, which was not found in plectin. Immunofluorescence staining revealed the presence of epiplakin in whole sheets of epidermis and esophagus, in glandular cells of eccrine sweat and parotid glands and in mucous epithelial cells in the stomach and colon.

Amino Acid Sequence↗

PubMed2001

Mediation of estrogen mitogenic effect in human breast cancer MCF-7 cells by PC-cell-derived growth factor (PCDGF/granulin precursor).

PC-cell-derived growth factor (PCDGF) is an 88-kDa glycoprotein corresponding to the granulin precursor. We have reported that PCDGF was expressed in human breast cancer cells. In estrogen-receptor positive cells, 17-beta-estradiol (E(2)) transcriptionally stimulated PCDGF expression in a dose- and time-dependent fashion. We demonstrate here that PCDGF mediates the mitogenic effect of E(2) in MCF-7 cells. PCDGF substituted for E(2) to stimulate DNA synthesis. The E(2) mitogenic effect was inhibited in a dose-dependent fashion by anti-PCDGF neutralizing antibody. Inhibition of PCDGF expression by antisense transfection also inhibited the E(2) mitogenic effect. In contrast, overexpression of PCDGF in MCF-7 cells resulted in cells that were able to proliferate in the absence of estrogen and were tamoxifen resistant. The PCDGF signaling pathway was examined. Like E(2), PCDGF stimulated mitogen-activated protein kinase activity. PCDGF could substitute for E(2) in stimulating cyclin D1 expression. The cyclin D1 stimulation by E(2) was 50% inhibited by anti-PCDGF antibody. In contrast, PCDGF did not stimulate c-myc expression, another molecular target of E(2). We conclude that autocrine PCDGF mediates the E(2) mitogenic effect via stimulation of cyclin D1. These studies provide information on estrogen action and identify an autocrine molecular target in human breast cancer cells.

Antibodies↗

PubMed2001

Purification and preliminary characterization of the zonula occludens toxin receptor from human (CaCo2) and murine (IEC6) intestinal cell lines.

In the present study, we report the preliminary characterization of the epithelial cell receptor for Vibrio cholerae zonula occludens toxin (Zot). Zot receptor was purified by ligand-affinity chromatography. Analysis of affinity-purified preparations by polyacrylamide gel electrophoresis revealed a protein of ca. 66 kDa. Partial N-terminal sequence obtained from purified murine and human Zot receptor revealed homology between the two proteins and with human alpha-1-chimaerin. Zot protein domain(s) involved in receptor binding were also analyzed by constructing several in frame deletion derivatives of a recombinant fusion Zot protein tagged with maltose binding protein. Our results suggest that Zot binding to its cellular membrane receptor requires a sequence that spans between amino acids 118 and 299.

Amino Acid Sequence↗

PubMed2001

Glutamine ameliorates mechanical obstruction-induced intestinal injury.

BACKGROUND: Glutamine has been shown to be an important dietary component for the maintenance of gut integrity. Although considered a nonessential amino acid in normal circumstances, glutamine may become conditionally essential for the bowel during episodes of severe illness and malnutrition. In this study, we employed an animal model simulating mechanical intestinal obstruction to explore the beneficial effects of glutamine on the intestine in response to obstruction-induced injury. MATERIALS AND METHODS: Rats were on three feeding regimens-standard diet and water (control group), diet and water containing 2% glutamine (glutamine group), or diet and water containing 2% arginine (arginine group)-for 3 days prior to surgical preparation of intestinal obstruction. The bowel distension, fluid accumulation, and histological alterations in the intestinal mucosa were measured 40 h after ileal ligation. RESULTS: After 3 days of drinking water intervention, the plasma glutamine levels in the glutamine group (677 +/- 12 microM) were higher than those in the control (451 +/- 27 microM) and arginine (379 +/- 25 microM) groups. The distension ratio measured 40 h after ileal ligation was significantly lower in the glutamine group (30.9 +/- 4.2%) than in the control and arginine groups (45.9 +/- 1.7 and 46.1 +/- 3.4%, respectively). Also, glutamine markedly decreased the fluid accumulation in the obstructed bowel segment (control group, 178.41 +/- 18.60 mg/cm; glutamine group, 104.97 +/- 13.17 mg/cm; arginine group, 141.4 +/- 12.85 mg/cm). Furthermore, the obstruction-induced mucosal injury was substantially improved in glutamine-fed rats. CONCLUSIONS: Our findings indicate that glutamine can significantly reduce the degree of those physiological derangements induced by mechanical intestinal obstruction.

Administration, Oral↗

PubMed2001

Alterations of telomerase activity and terminal restriction fragment in gastric cancer and its premalignant lesions.

AIMS: In order to explore the role of alterations of telomerase activity and terminal restriction fragment (TRF) length in the development and progression of gastric cancer. METHODS: Telomerase activity was detected in 176 specimens of gastric mucosa obtained through an operation or endoscopical biopsy by using the telomeric repeat amplification protocol (TRAP) assay. Meanwhile, the mean length of TRF was measured with the use of a Southern blot in part of those samples. RESULTS: Telomerase activity was detected in 14 of 57 (24.6%) chronic atrophy gastritis patients, six of 18 (33.3%) intestinal metaplasia patients, three of eight (37.5%) dysplasia patients and 60 of 65 (92.3%) gastric cancer patients, respectively. Normal gastric mucosa revealed no telomerase activity. No association was found between telomerase activity and any clinicopathological parameters. The mean TRF length was decreased gradually with age in normal mucosa and in gastric cancer tissue. Regression analysis demonstrated that the reduction rate in these tissues was 41 +/- 12 base pairs/year. Among 35 gastric cancers, TRF length was shown to be shorter in 20 cases (57.1%), similar in 12 cases (34.3%) and elongated in three cases (7.6%), compared to the corresponding adjacent tissues. The mean TRF length tended to decrease as the mucosa underwent chronic atrophy gastritis, intestinal metaplasia, dysplasia and into gastric cancer. The mean TRF length in gastric cancer was not statistically correlated with clinicopathological parameters and telomerase activity. CONCLUSIONS: Our results suggest that telomerase is expressed during the early stage of gastric carcinogenesis, and that the clinical significance of TRF length appears to be limited in gastric cancer.

Adult↗

PubMed2001

Human immunodeficiency virus type 1 replication in the absence of integrase-mediated dna recombination: definition of permissive and nonpermissive T-cell lines.

Functional retroviral integrase protein is thought to be essential for productive viral replication. Yet, previous studies differed on the extent to which integrase mutant viruses expressed human immunodeficiency virus type 1 (HIV-1) genes from unintegrated DNA. Although one reason for this difference was that class II integrase mutations pleiotropically affected the viral life cycle, another reason apparently depended on the identity of the infected cell. Here, we analyzed integrase mutant viral infectivities in a variety of cell types. Single-round infectivity of class I integration-specific mutant HIV-1 ranged from <0.03 to 0.3% of that of the wild type (WT) across four different T-cell lines. Based on this approximately 10-fold influence of cell type on mutant gene expression, we examined class I and class II mutant replication kinetics in seven different cell lines and two primary cell types. Unexpectedly, some cell lines supported productive class I mutant viral replication under conditions that restricted class II mutant growth. Cells were defined as permissive, semipermissive, or nonpermissive based on their ability to support the continual passage of class I integration-defective HIV-1. Mutant infectivity in semipermissive and permissive cells as quantified by 50% tissue culture infectious doses, however, was only 0.0006 to 0.005% of that of WT. Since the frequencies of mutant DNA recombination in these lines ranged from 0.023 to <0.093% of the WT, we conclude that productive replication in the absence of integrase function most likely required the illegitimate integration of HIV-1 into host chromosomes by cellular DNA recombination enzymes.

CD4-Positive T-Lymphocytes↗

PubMed2001

Genome segment S8 of grass carp hemorrhage virus encodes a virion protein.

The complete nucleotide sequence of the genome segment S8 of grass carp hemorrhage virus (GCHV) was determined from cDNA corresponding to the viral genomic RNA. It is 1,287 nucleotides in length and contains a large open reading frame that could encode a protein of 409 amino acids with a predicted molecular mass of 44 kD. The S8 was expressed using the pET fusion protein vector and detected by Western blotting analysis using the chicken egg IgY against intact GCHV particles, indicating that S8 encodes a virion protein. Amino acid sequence comparisons revealed that the protein encoded by S8 is closely related to protein sigma2 of mammalian reovirus, suggesting that the deduced protein of S8 is an inner capsid protein.

Amino Acid Sequence↗

PubMed2001

Transgenic mice overexpressing leptin accumulate adipose mass at an older, but not younger, age.

Sensitivity to leptin is associated with a normal regulation of the adipose mass, whereas decreased leptin sensitivity results in elevated adipose tissue stores. To address whether the effects of chronic hyperleptinemia are sustained with age, we generated transgenic mice that overexpress leptin under the control of the fat specific aP2 promoter/enhancer. At 6-9 weeks of age, transgenic mice overexpressed 5-fold more human leptin than endogenous mouse levels and had consistently low body weights, with reduced brown and white fat depots characterized by adipocytes either devoid of or containing minute lipid droplets. However, at 33-37 weeks, despite continuous secretion of human leptin, the transgenic mice showed a rebound effect characterized by an increase in body weight, accumulation of adipose mass, and lipid-filled adipocytes. Thus, this mouse model exhibits a two-stage phenotype, with respect to fat accumulation. In addition, plasma glucose, triglycerides, and cholesterol levels were markedly depressed in young, but not older, transgenic mice. A detrimental consequence of early hyperleptinemia was a failure of the transgenic mice to acclimatize to the cold, as a result of depleted fat stores within their brown adipocytes. Cold exposure was tolerated after a 2-week high-fat diet or at an older age when fat depots had naturally accumulated. Treatment of the older transgenic mice with large doses of leptin stimulated weight loss, demonstrating that the leptin pathway still responds to pharmacological levels of leptin. Overall, these studies show that moderate hyperleptinemia in normal mice results in a sensitivity of the adipose mass to leptin at a younger (but not older) age. The mechanisms that lead to the accumulation of fat at an older age remain largely unknown, and this hyperleptinemic mouse model will allow the uncovering of at least some of these mechanisms.

Adipocytes↗

PubMed2001

[Single particle source apportionment of atmospheric aerosol based on proton microprobe].

A proton microprobe with high resolution and high sensitivity was applied to analyze atmospheric aerosol in the people's park in Shanghai urban for single particle source apportionment. The results showed that the sum of the average contribution rates from the sources of soil dust, building construction dust, vehicle exhaust and metallurgical industry, which are the main pollution sources of atmospheric aerosol in Shanghai urban and surpasses 80%. Besides, about 13% of the particles could not be identified. On the basis of cluster analysis of these particles, they are preliminary known belonging to two different individual sub-pollution sources and a new unidentified pollution source.

Aerosols↗

PubMed2001

[Studies on cloning and expression of hepatitis B virus x gene].

OBJECTIVE: To express and purify the x gene of hepatitis B virus (HBV), and then the purity of the protein could specifically be analysed by the human antibodies against HBV x antigen (protein). METHODS: HBV x gene was amplified from the complete genome by PCR, and cloned in the pGEX-2T fused expression vector. After transforming the plasmid into E.coli, the recombinant with HBx gene was obtained. The fusion protein was generated with E.coli fused expression system and purified with affinity chromatography. RESULTS: The HBx protein could be applied to detecting specific antibody of hepatitis B virus patients. CONCLUSIONS: The purity of the HBx protein can specifically be recognized by the human antibodies against HBV x antigen.

Base Sequence↗

PubMed2001

[Different serum lipid adjustment drugs for the treatment of hyperlipidemic fatty liver].

OBJECTIVE: To research the effect of different serum lipid adjustment drugs, Simvastatin and Fenofibrate, on hyperlipidemic fatty liver. METHODS: The rat models of hyperlipidemic fatty liver induced by high fat diet were treated with Fenofibrate and Simvastatin respectively to investigate the change of the liver index, liver function, the serum and liver lipid, MDA, and pathologic histology. RESULTS: The liver index and the content of triglyceride (TC1.80 +/-0.20/2.10+/-0.33), total cholesterol (TG 0.76+/-0.18/1.09+/-0.31), MDA both in the serum and the liver in the model group were significantly higher than those in control group. Slight or moderate hepatic steatosis was observed in model group. Fenofibrate decreased serum triglyceride, total cholesterol, and MDA both in serum and liver significantly, but increased the ALT, ALP, liver index, liver lipids, and finally leading to severe hepatic steatosis. Simvastatin lowered the level of lipid and MDA both in serum and liver, and liver histology was improved nearly to normal. Moreover, little side effect on the liver was observed. CONCLUSIONS: Simvastatin can be applied for prevention and treatment of hyperlipidemic fatty liver safely and effectively, while Fenofibrate can not.

Animals↗

PubMed2000

Adeno-associated viral vector-mediated vascular endothelial growth factor gene transfer induces neovascular formation in ischemic heart.

Vascular endothelial growth factor (VEGF) plays important roles in physiological and pathological angiogenesis. Recent studies have demonstrated that direct injection of VEGF protein, plasmid DNA, or an adenoviral vector encoding the VEGF gene into ischemic myocardium or limb can induce collateral blood vessel formation and improve perfusion of the ischemic areas. However, these approaches have limitations ranging from a short-lasting effect to angioma formation. In this study, we investigated the feasibility of using adeno-associated viral (AAV) vectors to deliver VEGF genes to mouse myocardium. A cytomegalovirus promoter was used to drive genes for a human VEGF isoform, VEGF(165), and LacZ. A mouse myocardial ischemic model was generated by ligation of the anterior descending coronary artery. Approximately 10(11) copies of the AAV-VEGF vector mixed with 10(10) copies of AAV-LacZ were injected to one site of normal myocardium and a total of 10(11) copies of AAV-VEGF were injected to multiple sites of myocardium around the ischemic region. LacZ gene expression was observed up to 3 months after the vector inoculation. After AAV-VEGF inoculation, neoangiogenesis was observed in the ischemic heart model but not in normal heart tissue. An inflammatory-cell infiltration was not observed in the AAV-VEGF- and AAV-LacZ-inoculated hearts, and angioma-like structure was not observed. These results indicated that injection of the AAV vector directly to myocardium could mediate efficient gene transfer and transgene expression and that VEGF gene delivered by AAV vector can induce angiogenesis in ischemic myocardium.

Animals↗

PubMed2000

Regulation of the promoter activity of interferon regulatory factor-7 gene. Activation by interferon snd silencing by hypermethylation.

The molecular mechanism by which virus induces expression of the early inflammatory genes has not yet been completely elucidated. Previous studies indicated that the virus-mediated transcription of type I interferon (IFN) genes required activation of two members of IFN regulatory factor (IRF) family, IRF-3 and IRF-7, where the expression of IRF-7 was found to be indispensable for the induction of IFNA genes. To determine the factors that regulate expression of IRF-7 gene, as well as its inducibility by type I IFNs, we have isolated and characterized the promoter and first intron of the human IRF-7 gene. This region shows a presence of two potential interferon-sensitive response elements (ISRE/IRF-E). However, only the ISRE present in the first intron was functional and conferred interferon inducibility in a transient transfection assay. Using a pull-down assay with an oligodeoxynucleotide corresponding to this ISRE immobilized to magnetic beads, we have demonstrated that this ISRE binds ISGF3 complex and IRF-1 from the extract of IFN-treated cells but not from the untreated cells. We have further shown that the previously observed lack of expression of IRF-7 in 2fTGH fibrosarcoma cell line, correlated with hypermethylation of the CpG island in the human IRF-7 promoter. The repression of the promoter activity was relieved by treatment with DNA methyltransferase inhibitor 5-aza-deoxycytidine. In vitro methylation of IRF-7 promoter silenced IRF-7 directed expression of luciferase gene in HeLa cells that express endogenous IRF-7 gene. Whether silencing of IRF-7 by methylation is instrumental for the process of tumorigenesis remains to be determined.

Azacitidine↗