NSC1122002, a TRAF-binding domain inhibitor, inhibits collagen-induced arthritis in a mouse model.

OBJECTIVES: Dysregulation of IL-12 and IL-23 is related to many autoimmune diseases including arthritis. The production of IL-12 and IL-23 were reported to be under the control of JMJD2D, whose activity and stability were promoted by the TRAF-binding domain (TRABID), a deubiquitinating enzyme that epigenetically modulated inflammatory gene expression. NSC1122002 is a novel inhibitor of TRABID, and this study aimed to examine the effects of NSC1122002 on the expression of IL-12 and IL-23 both in vitro and in vivo in the context of collagen-induced arthritis, consequently to evaluate its potential as a drug candidate for treating inflammatory disease. METHODS: Bone marrow cells were isolated to detect the effect of NSC1122002 on the development of innate immune cells and other precursor cells. Primary macrophages and osteoclasts were used to examine the impact of NSC1122002 on cytokine expression. Collagen-induced arthritis was established to determine the function of NSC1122002 in vivo. RESULTS: NSC112200 did not affect the development of innate immune cells, primary osteoclast, and haematopoietic stem cells. NSC112200 specifically downregulated the expression of IL-12 and IL-23 through promoting degradation of JMJD2D by directly inhibited the deubiquitinating activity of TRABID. Besides, NSC112200 significantly suppressed the induction of CIA in mice. CONCLUSIOINS: Our findings provided new insight into the pathological mechanism and intervention method for arthritis therapy and identified that NSC112200 could be a potential drug for treating autoimmune diseases.

Down-regulation of lncRNA PCGEM1 inhibits cervical carcinoma by modulating the miR-642a-5p/LGMN axis.

LncRNA PCGEM1 (PCGEM1) has been reported to exert essential effects on the development and progress of various tumors, while the detailed effects and possible mechanisms of PCGEM1 in cervical carcinoma remain unknown. In the present study, PCGEM1 was over-expressed in cervical carcinoma cells as evidenced by real-time quantitative polymerase chain reaction (RT-qPCR) assay. Knockdown of PCGEM1 significantly repressed proliferation, migration, and invasion, while induced G1 arrest in cervical carcinoma cells. In addition, PCGEM1 was predicted to target miR-642a-5p by bioinformatics software, which was further confirmed by luciferase reporter assay. Besides, RT-qPCR assay indicated that miR-642a-5p expression was decreased in cervical carcinoma cells and knockdown of PCGEM1 could accelerate miR-642a-5p expression. Moreover, inhibition of miR-642a-5p partly abolished the functions of PCGEM1 knockdown on proliferation, cell cycle, migration and invasion of cervical carcinoma cells. Furthermore, miR-642a-5p could bind to the 3'-UTR of LGMN, which was over-expressed in the cervical carcinoma cells. Suppression of LGMN partly restored the functions of miR-642a-5p inhibitor on proliferation, cell cycle distribution, migration and invasion in the cervical carcinoma cells treated with the PCGEM1 shRNA. Taken together, our data indicated that knockdown of PCGEM1 inhibited proliferation, migration and invasion in cervical carcinoma by modulating the miR-642a-5p/ LGMN axis.

Annexin A2 plays a critical role in epithelial ovarian cancer.

PURPOSE: This study aimed at investigating the potential role and prognostic significance of Annexin A2 in human epithelial ovarian cancer (EOC). METHODS: Western blot was used to evaluate the expression of Annexin A2 in nine fresh EOC tissues, and immunohistochemical analysis was performed on formalin-fixed paraffin-embedded sections of 119 cases of ovarian cancers. Then, we used Fisher exact test to analyze the correlation between Annexin A2 and clinicopathological parameters. Starvation refeeding was used to detect the alteration of Annexin A2 in HO8910 cell cycle. RESULTS: Annexin A2 was overexpressed in carcinoma tissues compared with normal tissue, and the expression levels gradually increased from G1 to G3. Moreover, the staining of tissue microarray was consistent with the result we got from western blot, increasing from G1 to G3 gradually, and it was related to the Figo stage (P = 0.005), histologic grade (P = 0.002), ascite (P < 0.001), malignant tumor cells (P < 0.001), residual tumor size (P = 0.044), Ki-67 (P = 0.003). Kaplan-Meier analysis revealed that high Annexin A2 expression was significantly associated with poor prognoses of the patients (P < 0.001). Multivariate analysis demonstrated that Annexin A2 was an independent prognostic indicator for overall survival. Starvation refeeding indicated that Annexin A2 was related to EOC cell proliferation. CONCLUSIONS: We could hypothesize that Annexin A2 acted a critical role in EOC cell proliferation, and may be used as a potential and novel therapeutic target for EOC. These data suggest that Annexin A2 may promote the progression of EOC and be a therapeutic target for EOC therapy.

TNF-α expression in Schwann cells is induced by LPS and NF-κB-dependent pathways.

Lipopolysaccharide (LPS) is recognized by Toll-like receptor 4 and activates mitogen-activated protein kinase, which leads to the induction of proinflammatory cytokine gene expression. In vivo, Schwann cells (SCs) at the site of injury may also produce tumor necrosis factor-α (TNF-α). However, the precise mechanism that regulates TNF-α synthesis is still not clear. The nuclear transcription factor-κB (NF-κB) is an important transcription factor which is involved in the regulation of host immune responses. In the present study, we found that LPS possessed a comparable specific activity for activation of NF-κB-dependent gene expression in SCs. We also observed IκB-α/IκB-ß degradation and the nuclear translocation of P65 due to LPS treatments. LPS-elicited TNF-α production in SCs was also drastically suppressed by SN50 (NF-κB inhibitor).

Upregulated miR-146a Expression in Peripheral Blood Relates to Th17 and Treg Imbalance in Elder Rheumatoid Arthritis Patients.

BACKGROUND: The expression level of microRNA-146a (miR-146a) increased in peripheral blood and synovialis tissue of rheumatoid arthritis (RA) patient, and it may play an important role in the pathological process of RA. We investigated its possibility as a diagnostic marker and the correlation with T helper 17 (Th17) and Treg cells in elder RA patients. METHODS: Blood samples were collected from 38 active RA patients, 38 inactive RA patients, and 40 healthy controls. RNA expression levels of miR-146a were detected from the peripheral blood samples. The proportion of Th17 and Treg cells were analyzed, as well as their cell-specific transcription factor retinoic acid-related orphan receptor variant 2 (RORc) and forkhead box protein 3 (FOXP3). Furthermore, secretion of pre-inflammatory and anti-inflammatory factors was detected. Correlations between miR-146a and these factors were also analyzed. RESULTS: Compared with healthy control, expression levels of miR-146a in inactive and active groups were significantly higher, with the highest level in active group. The expression of miR-146a and the RA severity, Th17 cell ratio, RORc expression, IL-17 level showed a significant positive correlation, while it showed a significantly negative correlation with Treg cell ration, FOXP3 expression, and TGF-ß1 secretion. CONCLUSIONS: These results suggested that miR-146a may be used as a disease progression marker in the peripheral blood of elder RA patients.

miR­145 suppresses ovarian cancer progression via modulation of cell growth and invasion by targeting CCND2 and E2F3.

MicroRNAs (miRNA/miRs) have been demonstrated to be critical post­transcriptional modulators of gene expression during tumorigenesis. Numerous miRNAs have been revealed to be downregulated in human epithelial ovarian cancer (EOC). In the present study, it was observed that the expression of miR­145 was decreased in EOC tissues and cell lines. Overexpression of miR­145 inhibited the proliferation, migration and invasion of EOC cells. The D­type cyclin 2, cyclin D2 (CCND2), and E2F transcription factor 3 (E2F3) were confirmed to be targets of miR­145. In addition, restoration of these 2 genes significantly reversed the tumor suppressive effects of miR­145. Collectively, the results indicated that miR­145 serves a critical role in suppressing the biological behavior of EOC cells by targeting CCND2 and E2F3. Therefore, miR­145 was suggested to be a potential miRNA­based therapeutic target in ovarian cancer.

Antibody Modified Nanoparticle-Mediated Delivery of miR-124 Regulates Apoptosis via Repression the Stat3 Signal in Mycobacterial-Infected Microglia.

The mechanism of Mycobacterium tuberculosis (M.tb) evasion of host cell remains elusive. Several microRNAs that are involved in this complex process were identified. miRNA interference-based therapeutics represents an attractive challenge and shows huge potential for disorder treatment. In this study, we found that miR-124-3p expression is significantly decreased in microglia after Mycobacterium marinum (M.m) infection. To achieve better target transfection effect, a CD11b antibody and PEI modified nanoparticles-based Nano platform had been developed. This system was equipped by conjugation of miRNA-124-3p onto the surface of nanoparticles with a PEI/CD11b antibody coating. Transfection with miR-124-3p promoted microglia apoptosis through upregulation of Caspase3 or downregulation of Bcl-2 and Bcl-xl. More importantly, transfection with miR-124-3p inhibitor increases the mycobacterium proliferation in microglia. Based on the above, we further found miRNA-124-3p to bind to 3'untranslated region of Stat3, resulting in the downregulation of its protein to trigger cells apoptosis through Stat3-related pathway. As such, our research might provide new insights towards target delivering miRNA through the bold-brain barrier (BBB) and exploiting highly effective anti-tuberculous meningitis drugs. Taken together, our findings suggest how Mycobacterium can manipulate host miRNA expression to regulate cell survival for its own proliferation, and highlight the importance to develop novel therapeutic strategies against tuberculous meningitis.

Mycobacterium marinum Infection in Zebrafish and Microglia Imitates the Early Stage of Tuberculous Meningitis.

Mycobacterium tuberculosis (M. tuberculosis) invading and activating microglia causes the most serious subtypes of tuberculosis called tubercular meningitis. However, the developmental process of tubercular meningitis, especially the early phase, is poorly understood due to lacking well-established and well-accepted visible models in vitro and in vivo. Here, consistent with one recent report, we found Mycobacterium marinum (M. marinum) invade the zebrafish brain and subsequently cause granuloma-like structures. We further showed that M. marinum, which shares similar characteristics with M. tuberculosis, can invade microglia and replicate in microglia, which subsequently promote the secretion of pro-inflammatory cytokines such as IL-1ß, IL-6, and TNF-α. M. marinum infection in microglia can also promote autophagy, which conversely limits the replication of M. marinum. Thus, pharmacological activation of autophagy by rapamycin could prevent M. marinum replication. Our study provides in vivo and in vitro models to study underlying pathogenic mechanisms of tubercular meningitis by using M. marinum. Our results also showed that activation of autophagy could be a meaningful way to prevent tubercular meningitis.

Resveratrol improves treatment outcome and laboratory parameters in patients with Takayasu arteritis: A randomized double-blind and placebo-controlled trial.

Tumor necrosis factor (TNF) inhibitors have exhibited certain clinical efficacy in treating refractory Takayasu arteritis (TA), albeit with severe adverse effects. We aimed to explore the anti-TNF function of resveratrol, a natural compound, in the treatment of TA. A total of 271 patients diagnosed of acute TA were enrolled in this clinical trial, who were then randomized to be administered 250mg resveratrol or placebo on a daily basis for a period of 3 months, and revisited biweekly to assess treatment outcomes. Primary treatment outcome was defined as the disease activity, determined using the Birmingham Vascular Activity Score (BVAS). Secondary outcome was defined by laboratory parameters, including erythrocyte sedimentation rate (ESR), plasma levels of C-reactive protein (CRP) and TNF-α. BVAS score and laboratory parameters of patients receiving resveratrol treatment exhibited a steady decline throughout the study. In contrast, outcomes remained practically unchanged in placebo-treated patients. Strong linear correlations were also observed between TNF-α with BVAS scores, ESR and plasma levels of CRP. Resveratrol could greatly improve treatment outcome and laboratory parameters in acute TA patients, likely due to its anti-TNF property.

Inhibitory Effect of a Novel Antirheumatic Drug T-614 on the IL-6-Induced RANKL/OPG, IL-17, and MMP-3 Expression in Synovial Fibroblasts from Rheumatoid Arthritis Patients.

T-614 (also named as iguratimod), a novel antirheumatic drug, could attenuate joint inflammation and articular damage in rheumatoid arthritis (RA) patients, providing a new therapy for RA. Here, we tested the role T-614 on the IL-6-induced receptor activator of nuclear factor κB ligand (RANKL)/osteoprotegerin (OPG), IL-17, and MMP-3 expression in synovial fibroblasts from rheumatoid arthritis (RASFs) patients. T-614 decreased RANKL expression and RANKL/OPG ratio in IL-6-induced RASFs. We confirmed this effect by a decrease of the mRNA and protein RANKL and mRNA RANKL/OPG in RASFs exposed in vitro to T-614 or MTX. Markedly decreased levels of IL-17, retinoid-related orphan receptor C (RORc), and MMP-3 mRNA expression were also observed in IL-6-induced RASFs in the presence of T-614 or MTX compared with those in its absence. Furthermore, T-614 blocked expression of p-ERK1/2 protein without affecting ERK1/2 expression, indicating that the way that T-614 regulated RANKL expression might be ERK1/2 pathway. Our results suggest that T-614 yields a strong improvement in arthritis via exact suppression of RANKL/OPG, IL-17, and MMP-3 expression in RASFs.

CAP1 is overexpressed in human epithelial ovarian cancer and promotes cell proliferation.

Adenylate cyclase-associated protein 1 (CAP1) regulates both actin filaments and the Ras/cAMP pathway in yeast, and has been found play a role in cell motility and in the development of certain types of cancer. In the present study, we investigated CAP1 gene expression in human epithelial ovarian cancer (EOC). Western blot analysis and immunohistochemistry were performed using EOC tissue samples and the results revealed that CAP1 expression increased with the increasing grade of EOC. In the normal ovarian tissue samples however, CAP1 expression was barely detected. Using Pearson's χ2 test, it was demonstrated that CAP1 expression was associated with the histological grade and Ki-67 expression. Kaplan-Meier analysis revealed that a higher CAP1 expression in patients with EOC was associated with a poorer prognosis. In in vitro experiments using HO-8910 EOC cells, the expression of CAP1 was knocked down using siRNA. The proliferation of the HO-8910 cells was then determined by cell cycle analysis and cell proliferation assay using the cell counting kit-8 and flow cytometry. The results revealed that the loss of CAP1 expression inhibited cell cycle progression. These findings suggest that a high expression of CAP1 is involved in the pathogenesis of EOC, and that the downregulation of CAP1 in tumor cells may be a therapeutic target for the treatment of patients with EOC.

Adiponectin exacerbates collagen-induced arthritis via enhancing Th17 response and prompting RANKL expression.

We previously reported adiponectin (AD) is highly expressed in the inflamed synovial joint tissue and correlates closely with progressive bone erosion in Rheumatoid arthritis (RA) patients. Here, we investigate the role of adiponectin in regulating Th17 response and the expression of receptor activator of nuclear factor-κB ligand (RANKL) in mice with CIA mice by intraarticularly injection of adiponectin into knee joints on day 17, day 20 and day 23 post first collagen immunization. The increased adiponectin expression was found in inflamed joint tissue of collagen-induced arthritis (CIA) mice. Adiponectin injection resulted in an earlier onset of arthritis, an aggravated arthritic progression, more severe synovial hyperplasia, bone erosion and osteoporosis in CIA mice. CD4(+)IL-17(+) Th17 cells, IL-17 mRNA and RANKL mRNA expression were markedly increased in the joint tissue of adiponectin treated CIA mice. Moreover, adiponectin treatment markedly enhanced Th17 cell generation from naive CD4(+) T cells in vitro, which accompanied by the high expression of Th17 transcription factor ROR-γt, and Th17 cytokine genes included IL-22 and IL-23. This study reveals a novel effect of adiponectin in exacerbating CIA progression by enhancing Th17 cell response and RANKL expression.

Expression and clinical role of TCTP in epithelial ovarian cancer.

The aim of this study is to investigate the potential role and prognostic significance of translationally controlled tumor protein (TCTP) in human epithelial ovarian cancer (EOC). Western blot was used to evaluate the expression of TCTP in eight fresh EOC tissues. Immunohistochemistry was performed on formalin-fixed paraffin-embedded sections of 119 cases of ovarian cancers. Kaplan-Meier method indicated the relation between TCTP and EOC patients' overall survival rate. Starvation and re-feeding was used to assess cell cycle. Knocking down of TCTP and CCK8 assay showed the role of TCTP in HO8910 cell cycle. We found that TCTP was overexpressed in carcinoma tissues compared with normal tissues. Immunohistochemistry revealed that TCTP expression was significantly associated with clinicopathologic variables. Kaplan-Meier analysis revealed that high TCTP expression was significantly related to poor prognosis of the patients. Starvation and re-feeding suggested TCTP played a critical role in HO8910 cell proliferation. Interference of TCTP and CCK8 assay showed that the TCTP-siRNA treated HO8910 cells grew more slowly than the control group. CCK-8 assays and terminal-deoxynucleoitidyl transferase mediated nick end labeling assays were also performed to demonstrate the cisplatin could inhibit the survival of HO8910 cells and promote their apoptosis. All the experiments we have done showed that TCTP could promote the progression of EOC and reduce the sensitiveness of HO8910 cells to cisplatin.