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Biomedical subjects

H Bu

Publications and source records attributed to H Bu.

At least 19 recordsLinked to original sources
PubMed2006

Interference of cyclosporine on glucose metabolism: potential role in chronic transplantation kidney fibrosis.

OBJECTIVES: To explore the fibrosis induction effects of cyclosporine (CsA) on renal tubular epithelial cells, as well as the potential mechanisms of CsA-mediated alterations of glucose metabolism in chronic allograft nephropathy (CAN). METHODS: The rat renal tubular epithelial cell line NRK-52E cells were cultured in medium with 50 mmol/L of D-glucose for 7, 14, or 28 days. The expression of TGF-beta1, CTGF, Smad3, and Smad7, which are involved in the fibrosis signal pathway, was detected by immunofluorescence and reverse-transcriptase polymerase chain reaction. Meanwhile, cells were cultured in various concentration of CsA in glucose-free medium for 24 hours followed by the addition of D-[3-(3)H]-glucose for 30 minutes; glucose uptake was detected by examining the radiation intensity. RESULTS: The expression of TGF-beta1, Smad3, Smad7, and CTGF of NRK-52E cells were up-regulated significantly (P < .05) after culture with 50 mmol/L glucose for 7 days compared with those in 25 mmol/L glucose. At 14 and 28 days, TGF-beta1 was slightly decreased compared with 7 days, but CTGF, Smad3, and Smad7 increased robustly (P < .05). CsA significantly stimulated glucose uptake at various concentrations, with an absorption peak at 60 minutes that decreased at 75 minutes. Furthermore, CsA may have dose-dependent effects on glucose uptake. CONCLUSION: High glucose concentrations may play an important role in the NRK-52E cell fibrosis during the CAN process due to an interference of CsA on glucose metabolism.

Animals↗
PubMed2006

Nuclear translocation of SMAD3 may enhance the TGF-beta/SMADS pathway in high glucose circumstances.

OBJECTIVES: Posttransplant diabetes mellitus is one of the most frequent complications after kidney transplantation. It is considered to be one cause of chronic allograft nephropathy. This study sought to investigate the effects of high glucose on the expression and nuclear translocation of Smad3, which is an important signal mediator involved in the fibrotic signal pathway. METHODS: The established rat renal mesangial cell line HBZY-1 was cultured in medium with various concentrations of glucose (4.5 mg/mL, 9.0 mg/mL, or 13.5 mg/mL), which was collected at 7, 14, or 21 days. The total expression of Smad3, including both inner and outer nucleus proteins was examined by Western blot analysis. The nuclear translocated Smad3, representing only the inner nucleus protein, was detected by immunofluorescence staining observed under a laser confocal scanning microscope. RESULTS: No significant difference in the total Smad3 expression was demonstrated by Western blot analysis among the three groups of HBZY-1 cells at various concentration of glucose after 7, 14, or 21 days. There was no fluorescence detected in the nucleus at day 7 by immunofluorescence staining; however, robust positive expression of Smad3 was detected at days 14 and 21. CONCLUSION: As a restricted Smads member, Smad3 protein might not be upregulated in the presence of high glucose. However, with prolonged culture time, Smad3 translocates from cytoplasm to nucleus, which may be a pivotal step in the fibrotic signal pathway.

Animals↗
PubMed2006

The C-terminal peptide of connective tissue growth factor blocks the full molecule binding to tubular epithelial cell.

OBJECTIVES: Interstitial fibrosis is a critical pathologic change in chronic allograft nephropathy. The cytokine connective tissue growth factor (CTGF, also CCN2) plays an important role in epithelial-mesenchymal transformation (EMT) of tubular epithelial cells to renal interstitial fibrosis. The hexadeca-peptide within the C-terminal of CTGF (named P2) contains the unique binding domain of CTGF to its potential receptor, integrin alphavbeta3. This study examined whether P2 bound preferentially to the receptor and served as an inhibitor of CTGF. METHODS: All studies used an established rat kidney tubular epithelial cell line NRK-52E. Chemically synthesized P2 was purified, and some of it labeled with FITC. The affinity of CTGF or P2 to NRK-52E cells was examined by a solid-phase cell adhesion assay. Competitive binding between P2 and CTGF to NRK-52E cells was examined with flow cytometric analysis. RESULTS: Both P2 and CTGF bound to the NRK-52E cells, mediating cell adhesion. When the cells were incubated in the mixture of P2 and CTGF, P2 bound to the cells preferentially. Furthermore, when cells were preincubated with excessive CTGF, it became difficult for subsequent P2 binding to occur. CONCLUSIONS: P2 and CTGF seemed to bind to cell membranes at the same binding domain. P2 competitively blocked CTGF binding, acting as a CTGF inhibitor.

Animals↗
PubMed2005

Muscle infarction involving muscles of abdominal and thoracic walls in diabetes.

AIMS: This paper presents two cases of muscle infarction involving four major muscles of the anterior abdominal wall (case 1) and pectoralis major (case 2) in individuals with diabetes. METHODS: Erythrocyte sedimentation rate (ESR) and creatine kinase (CK) were measured and Doppler ultrasound, an open muscle biopsy (case 1) and magnetic resonance imaging (MRI) (case 2) were performed. RESULT: The diagnosis of muscle infarction was made by histological findings and MRI images with hyper-intensive signals on a gadolinium-enhanced T2-weighted sequence, respectively. Both patients were treated with bed rest, immobilization of the involved extremities, analgesia and intensive insulin therapy. In addition, anticoagulant drugs such as low molecular weight heparin sodium and cilostazol, and some traditional Chinese medicines such as ligustrazine and salvia miltiorrhiza were administered. The symptoms of both patients resolved gradually after 3 weeks. However, muscle infarction reoccurred in case 1 on the opposite side of the abdomen and recovered after 40 days. CONCLUSIONS: This is the first report of muscle infarction involving the muscles of anterior abdominal walls and pectoralis major in diabetes. MRI is the best non-invasive technique and T2-weighted imaging is the most valuable method for the diagnosis. In addition to supportive therapy, administration of anticoagulant agents and some Chinese traditional medicine may be useful in symptom relief.

Abdominal Muscles↗
PubMed2004

Screening and analysis of porcine endogenous retrovirus in Chinese Banna minipig inbred line.

Pigs have been the most likely animal as the source of cells, tissues, and organs for xenotransplantation. But the use of pigs in xenotransplantation is associated with the risk of porcine endogenous retrovirus (PERV) transmission. Previous studies have identified that the proviruses are integrated into the genome of normal pigs and that virus particles released from the porcine cells can infect human cells in vitro. As a unique inbred pig, Banna minipig inbred (BMI) has a huge potential value for xenotransplantation and medical research. It has been the focal experimental animal for pig-to-human xenotransplantation in China, due to its clear genetic background and tiny individual differences. To evaluate whether the potential risk of PERV exists in inbred pigs, a series of screening experiments were performed herein. The results of PCR with primers specific for gag, pol, and env showed that proviruses existed in the genome of BMI, and the PERV subtypes were PERV-A and PERV-B. PERV mRNA was expressed functionally in BMI. Positive results of an RT assay identified that PERV in BMI had potential infectivity, but the concentration of PERV reverse transcriptase in BMI was almost 20 times lower than that of HIV. These results suggested that gag, pol and env genes of PERV were not lost during inbreeding, which created favorable conditions to produce viral particles that could possibly infect human cells in xenotransplantation.

Animals↗
PubMed2004

Phylogenetic analysis of porcine endogenous retrovirus variation in three Chinese pigs.

PCR amplification was performed on genomic DNA extracted from peripheral blood lymphocytes of three species of Chinese pigs (Banna minipig inbreed [BMI], Wu-Zhi-Shan pig [WZSP], and Nei jiang pig [NJP]), using primers corresponding to the highly conserved regions of polymerase (pol) gene. Extracted PCR products were then cloned in a pGEM-T vector. Phylogenetic analysis of the nucleotide sequences of BMI-PERV, NJJP-PERV, and WZSP-PERV revealed them to be a novel category of PERV. In comparison to other type C retrovirus and lentivirus, their amino acid sequenced show about 30% to 57.7% identities. Our previous research demonstrated that PERV in the three pigs was highly expressed. It appears likely that functional loci encoding these novel PERV sequences exist, but this remains to be established. The novel sequences described in this report will allow such investigations to be actively pursued.

Animals↗
PubMed2001

[Biocompatibility of HA/TCP biphasic ceramics with co-cultured human osteoblasts in vitro].

The biocompatibility of HA/TCP ceramic was evaluated by investigation of attachment and growth of osteoblasts on biomaterial, as well as monitoring the effects of biomaterial on expression of functional phenotypes of co-cultured osteoblasts in vitro. When co-cultured with HA/TCP ceramics, osteoblasts firstly attached to the surface of HA/TCP disk, then attached to notches and grew into the micropores of biomaterial during further culture period. At last, the ceramics were almost packed with osteoblasts. Additionally, osteoblasts co-cultured with HA/TCP were similar to osteoblasts cultured under normal condition in osteoblastic phenotypes; the secreted lots of collagen type I, possess strong activity of Alkaline Phosphatase and mineralized extracellular matrix. The fact that osteoblasts could grow well on HA/TCP ceramics and the biomaterial did not affect their physiological function suggest that HA/TCP ceramic is biocompatible with human osteoblasts.

Biocompatible Materials↗
PubMed2001

[Detection of methylation status of p16 tumor suppressor gene in soft tissue leiomyosarcoma].

OBJECTIVE: To observe the methylation status of p16 tumor suppressor gene in human soft tissue leiomyosarcoma (LMS) and its relationship with p16 protein expression. METHODS: Methylation-specific PCR (MSP) assay was used to analyze the methylation status of 5' CpG island promoter/exon I region of p16 tumor suppressor gene in 38 LMS, 10 leiomyomas and 5 normal smooth muscle tissues. p16 expression in these tissues were examined by immunohistochemical assay. RESULTS: Hypermethylation of p16 was observed in 23.7% (9/38) of cases, of which 7 with negative pl6 staining and 2 with faint positive staining. In the cases without protein expression, hypermethylation rate of p16 was 50% (7/14). CONCLUSIONS: Epigenetic change due to 5' CpG methylation is the main cause of inactivation of p16 (INK4) gene which may be involved in tumor pathogenesis.

CpG Islands↗
PubMed2001

[Studies of intercellular communication in human rhabdomyosarcoma cell lines of different metastatic potential].

OBJECTIVE: To investigate the relationship between intercellular communication and three human rhabdomyosarcoma (RMS) cell lines of different metastatic potential. METHODS: Indirect immunofluorescent staining and laser scanning confocal microscope (ISCM) were used to detect connexin 43 (CX43), a molecule related with gap junctional intercellular communication (GJIC). Fluorescence redistribution after photobleaching (FRAP) was used to detect function of GJIC. RESULTS: A high level of CX43 protein expression was revealed in normal human myoblasts. CX43 protein was mainly localized on the surface of cells and sometimes in cytoplasm. CX43 expression was decreased in RMS cells. Both the detection rates and fluorescent intensity of CX43 decreased when the metastatic potential of RMS increased (P < 0.05). In contrast to normal, the fluorescence recovery rates of the RMS cells decreased and there was a negative correlation between the function of GJIC and malignancy of RMS cell lines (P < 0.05). CONCLUSION: Varying degrees of GJIC inhibition may correlate with different metastatic potential of RMS. This may help to determine the malignant behavior of RMS and be used as a prognostic index of RMS.

Animals↗
PubMed2000

[The xenoantigenicity of Chinese inbred-line pigs].

OBJECTIVE: The major obstacle in pig to human transplantation is acute and hyperacute rejection (HAR) triggered mainly by alpha-galactosyl residues(alpha-Gal) in donor. Since the inbred-line Banna pig(IBNP) and Wuzhishan pig (IWZSP) are highly inbred and may be the potential donor for xenotransplantation, it is important to investigate the reaction between human serum and inbred-line pig tissues as well as the distribution of alpha-Gal in these tissues. METHODS: Samples from heart, liver, spleen, lung, kidney, pancreas, small intestine, thymus, skin, lymph node and blood vessels at all levels were collected from four 8 to 11-month-old male IBNPs and one IWZSP. Affinity-immunohistochemistry assays were conducted following routine procedures on paraffin sections with normal human sera of blood type A, B, O, AB and BSI-B4(alpha-Gal specific binding lectin) as the primary antibodies or affinity reagents. Sections digested by alpha-galactosidase were also examined as control. RESULTS: Parallel results were obtained from these pig tissues stained against human sera and BSI-B4. There was no significant difference both in the antigens recognized by sera of different blood types or BSI-B4 and in the distribution of alpha-Gal. The strongest alpha-Gal positive staining was appeared in vascular endothelial cells at all levels and partial parenchyma cells. However, tissues of cartilage, peripheral nerve and muscle were negative. After digested by alpha-Galactosidase, all samples were negative against BSI-B4 and human sera except few positions that showed different staining. CONCLUSION: The distribution of target antigen is similar in various tissues of the two kinds of pigs. Though alpha-Gal is the major xenoantigen in IBNP and IWZSP, there may be some unknown antigens related to pig to human transplantation. Possibly the level and distribution of antigen expression in pig tissues are not the first affair to be considered, and these pigs should be genetically modified in order to eliminate rejection in pig to human xenotransplantation.

Animals↗
PubMed2000

[Study on the xenoantigens of pancreatic tissues of Chinese Neijiang pig].

The expression and distribution of xenoantigens in pancreatic tissue of Neijiang pig were investigated with immunohistochemistry technique. BSIB4, normal human serum and anti-human A, B, H monoclonal antibodies were used as the primary antibody to examine the expression of xenoantigens in the pancreatic tissues from 10 Neijiang pigs. The results demonstrate that compared with the negative results of human blood B and H type antigens, the alpha-Gal antigen was highly expressed on the pancreatic vascular endothelial cells and the duct epithelium. The human blood A type antigen was highly expressed on some islet cells, some exocrine secretory cells and all the duct epithelium. Since there are pre-exited xenoantibodies against pig pancreas cells in human serum, cross-matching test and graft pre-treatment are needed before clinical transplantation of pancreas. The expression of xenoantigens in Neijiang pig is different from other pig strains reported by others, which suggests that looking for a pig strain that contains less xenoantigens is possible.

ABO Blood-Group System↗
PubMed2000

[Immune reconstitution in immunosuppressed pigs with human immune competent cells].

To reconstitute the immune system of the pig with human immune cells after destroying or suppressing it, so that the transplant immunoreactivity between human and pig could be monitored in vivo. Fifteen Chinese Neijiang pigs were divided into 3 groups. Group A (n = 5) received immunosuppression through intravenous infusion of methotrexate (MTX), myleran and cyclophosphamide (CTX). Group B received intraperitoneal injection of human bone marrow and spleen cells (HBSC) after the same treatment as group A. Group C (n = 5) received intravenous infusion of HBSC after the same treatment as group A. Blood routines and biochemical analysis were monitored. The chimerisms of human cells were supervised through immunohistochemistry of cytospin of peripheral blood from recipient pigs and flow cytometry of the same specimen and immunohistochemistry of frozen tissue sections and paraffin sections. All pigs in group A survived definitely. Group B and C survived 3 days to 54 days and 15 hours to 60 days respectively. The peripheral white blood cell count (WBCC) of group A, B and C decreased to the bottom 7 days after drug administration, about 1%-10% of the normal level. After transplantation, the WBCC of group B and C recovered more slowly than those of group A. Human cells were found in the recipient pigs of group B and C after transplantation through immunohistochemistry of peripheral blood cytospin and flow cytometry. Infiltration of human cells was found in the spleens and lungs of group C and D through immunohistochemistry. The chimerism of human cells in recipient pigs could be established by transplanting human bone marrow and spleen cells into immunosuppressed pigs. It is possible that XGVHD occurs in the recipient, which offers a novel animal model for studies on immunoreactivity of pig-to-human xenotransplantation. The proportions of human cells in peripheral blood of recipient pigs are low long after transplantation through both intraperitoneal injection and intravenous infusion.

Animals↗
PubMed2000

[The RYR1 genotype of Chinese inbred pigs].

OBJECTIVE: Porcine stress syndrome (PSS) is one kind of molecular genetics defect diseases of pig which will cause malignant hyperthermia syndrome (MHS) and is the first index should be excluded in screening of a pig species for xenotransplantation. It was reported that mutation of pig rynodine receptor(RYR1) gene is the main reason for PSS. In this study, RYR1 genotypes of the Chinese Banna mini pig inbred line and inbreeding closed colony Wuzhishan pig were investigated with polymerase chain reaction-restriction endonuclease fragment length polymorphism (PCR-RFLP) technique. METHODS: Antevenocaval whole blood samples were collected from 50 Banna mini-pig inbred-line(BMI), 15 inbreeding Wuzhishan pig (WZSP) and 25 Neijiang pigs (NJP) as negative control, the primer were designed and synthesized, PCR reaction was conducted following the sequence of 94 degrees C (1 min), 58 degrees C (1 min) and 72 degrees C (1 min) for 30 cycles. The PCR products were digested with restriction endonuclease HhaI and then electrophoresis check. RESULTS: A 659 bp DNA fragment was amplified with these two primers, the HALNN sample fragment was cut into fragments as 493 bp and 166 bp individually after the digestion, indicates no point mutation at site 1,843 in RYR1 gene in all tested BMI pig and WZSP. Namely, the RYR1 genotype of 50 cases of BMI and 15 cases of WZSP were HALNN, therefore their phenotype is PSS negative. CONCLUSION: It indicates that the genotype of Banna mini pig inbred line and inbreeding Wuzhishan pig are HALNN therefore PSS absolutely negative, the group penetrance is 0. This is consistent with experimental observation. It suggests that Banna mini pig inbred line and inbreeding Wuzhishan pig may be the alternative donor for xenotransplantation.

Animals↗