Downregulation of miR-223 and miR-19a induces differentiation and promotes recruitment of osteoclast cells in giant-cell tumor of the bone via the Runx2/TWIST-RANK/RANKL pathway.

Giant-cell tumor (GCT) of the bone is an invasiveness and high recurrent bone tumor that is considered borderline or potentially malignant. To explore the molecular mechanism leading to bone destruction and identify novel targets for treatment, we conducted silencing of miR-223 and miR-19a in stromal giant cells and identified TWIST and Runx2 as their target genes. We investigated the impact of these microRNAs and their target genes on stromal giant cells that promote the differentiation of monocyte/macrophages into osteoclast cells and recruitment to the bone microenvironment, which in turn enhances the bone destruction capacity of GCT. MiR-223 and miR-19a were found to regulate the expression of TWIST and Runx2, influence the RANKL-RANK pathway and the expression of MCP-1, and finally regulate the pathophysiological process of osteolytic bone destruction. Our results indicate that re-expression of miR-223 and miR-19a induces an inhibitory effect on the bone destruction capacity of GCT, suggesting that re-expression of miR-223 and miR-19a can be a novel strategy for the treatment of GCT.

Carbonic anhydrase III is insufficient in muscles of myasthenia gravis patients.

Myasthenia gravis (MG) is considered as an autoimmune disease mainly mediated by antibodies against acetylcholine receptor. In recent years, other targets related to MG have been the subject of interest. Our previous research found that protein P25 was lower in muscles of MG patients using two-dimensional electrophoresis. In present study, anti-serum to P25 was prepared, immunohistochemistry and ATPase staining revealed that P25 was a muscle specific cytosolic protein and was mainly distributed in type I muscle fibers. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and precise molecular weight derived from mass spectrometer identified P25 as carbonic anhydrase III (CA III). Some members of CA family are related to autoimmune diseases and CA III is recently reported to be involved in rheumatoid arthritis. The results of immunoblot in this report showed that the level of CA III is specifically insufficient in the skeletal muscle of MG patients. The possible roles that CA III play in MG need further elucidation.

[Inhibitory effect of CVB3-VP1 siRNA on CVB3 replication].

AIM: To inhibit the expression of CVB3 VP1 protein and the replication of CVB3 with synthesized siRNAs. METHODS: According to the sequence and secondary structure of CVB3 VP1 protein, four pieces of siRNAs were designed following the requirement from Journal of Nature Cell Biology were synthesized in Shanghai GeneChem Company. Then they were transfected into HeLa cells by liposome (Lipofectamine 2000), but the non-transfected cells and non-specific siRNAs were taken as control. 48 hours later, the patho-morphous changes were observed, virus titer changes were examined by TCID50, CVB3-VP1 protein expression were detected by immunofluorescence with FITC dyeing, and CVB3-RNA level was tested by semi-quantitative RT-PCR. RESULTS: Two pieces of the four specific synthesized siRNAs (VP1-1 and VP1-2) were found to have obvious inhibitory effect on CVB3 replication and VP1 protein expression were reduced greatly. Besides, the changes of pathological cells were obviously mitigated. CONCLUSION: Specific siRNAs can effectively inhibit the expression of CVB3 VP1 protein and the replication of CVB3 in HeLa cells.

Impact of pathogen burden on in-stent restenosis in patients after coronary stent implantation.

BACKGROUND: Although some certain infectious pathogens could be detected in the patients with coronary artery disease, the roles of these infectious factors in the development of coronary artery diseases remain largely unknown. Since the number of infectious pathogens has been argued to be relative to the coronary artery diseases, we therefore examined whether there is a link between the number of infections and the incidence of in-stent restenosis after stent implantation. METHODS: One hundred and eighty-one patients were enrolled in this study. Infectious pathogens including serum anti-Chlymydia pneumoniae, cytomegalovirus, Helico pylori, human herpes simplex virus-1, human herpes simplex virus-2 antibodies and hepatitis B virus antigen were measured in all patients before coronary stent implantation. Coronary angiography was performed before, immediately after and 6 months after stent implantation. RESULTS: Restenosis rate 6 months post stent implantation was similar in patients with low pathogen burden (< 3 pathogens, 33.3%) to those with high pathogen burden (> or = 3 pathogens, 29.1%). CONCLUSIONS: Previous infections with Chlymydia pneumoniae, cytomegalovirus, Helico pylori, human herpes simplex virus-1, human herpes simplex virus-2 and hepatitis B virus do not contribute to the incidence of restenosis after stent implantation.

[Relationship between infection burden and atherosclerosis and plaque feature].

OBJECTIVE: To evaluate the relationship between infection burden and coronary atherosclerosis and the plaque feature. METHODS: One hundred and eighty two patients underwent coronary angiography in Zhongshan Hospital from 2002 - 2003. Atherosclerosis and vulnerable plaque were determined by intravascular ultrasound (IVUS). Seropositivity of cytomegalovirus, helicobacter pylori, chlamydia pneumonia, hepatitis B virus, EB virus, CoxB virus, influenza A virus, influenza B virus and mycobacterium tuberculosis were determined by ELISA. The serum hs-CRP was detected by Dade Behring prospect (Immuno-nehelomitery). Patients were divided into three groups according to the pathogen burden: group A, n or= 6. RESULTS: The pathogen burden was independent of the C-reactive protein level. Increasing pathogen burden was significantly associated with increasing atherosclerosis risk, the prevalence of atherosclerosis was 44.4%, 70.6% and 76.7% in group A, B and C. The risk associated with elevated pathogen burden was much higher when CRP was also elevated (> 5.0 mg/L) (43.8%, 70.0%, 70.8%) vs (45.5%, 63.7%, 96.8%). The positively of vulnerable plaque increased significantly when the pathogen burden was high (n > 5) (33.3%, 32.4% and 51.7% P < 0.05). CONCLUSION: Our data suggested that infection burden was associated with prevalence of coronary atherosclerosis, and it was particularly important when C-reactive protein was elevated. The high level infection burden could predict vulnerable plaque.

Hepatitis B virus infection and coronary atherosclerosis: results from a population with relatively high prevalence of hepatitis B virus.

AIM: To investigate the possible association between hepatitis B virus (HBV) infection and angiographically proven coronary artery disease (CAD) in a population with relatively high prevalence of HBV. METHODS: Sera from 434 patients who underwent coronary angiography were tested for HBV antigens (HBsAg, HBeAg) and antibodies (Anti-HBs, Anti-HBc and Anti-HBe) by ELISA. RESULTS: Seventy-seven percent (224/291) of the patients with CAD and 73.4% (105/143) of the patients without angiographic evidence of atherosclerosis were seropositive for HBV (P > 0.05). However, C-reactive protein (CRP) levels were significantly higher in patients with CAD (P = 0.008), while lower in HBV seropositive population (P = 0.043 and P = 0.021 after adjustment for conventional risk factors). CONCLUSION: Our results suggested HBV infection negatively correlates with CRP levels, but seems not to be associated with coronary atherosclerosis.

[Protective immunoresponse to CVB3 induced by gene immunization with pcDNA3-VP1].

AIM: To induce Coxsackie virus B type 3 (CVB3)-specific immune response by using a DNA vaccine containing CVB3-VP1 and to observe its protection against CVB3 challenge. METHODS: The gene coding for VP1 was obtained by RT-PCR and then was cloned into plasmid pcDNA3 to construct pcDNA3-VP1. In-vitro expression of VP1 was performed by transfection of pcDNA3-VP1 into Hela cells. Expressed product was detected by ELISA. BALB/c mice were immunized intramuscularly with 50 microg DNA three times, and challenged by 5xLD(50) CVB3 four weeks after the last immunization. RESULTS: pcDNA3-VP1 had been constructed and the expression product was detected in the culture supernatant of Hela cells 24 hours after transfection. CVB3-specific IgM and IgG elicited in the mice immunized with pcDNA3-VP1 were significantly higher than those in the control mice immunized with pcDNA3. Specific proliferation of the splenic lymphocytes and activity of CVB3-specific CTLs from the pcDNA3-VP1 immunized mice were much stronger than those in the controls. pcDNA3-VP1 could protect 33.3% mice from lethal CVB3 challenge, while control mice only survived 6.7 days. Infiltration of inflammatory cells or unusual proliferation of connective tissue, indicating ongoing myocarditis or fibrosis, were not found in pcDNA3-VP1 immunized mice, but did exist in control mice. CONCLUSION: Intramuscular immunization with pcDNA3-VP1 may be a promising approach against CVB3 infection.