Aimei Li

MiR-122 modulates type I interferon expression through blocking suppressor of cytokine signaling 1

MiR-122 is a liver-specific miRNA. Recent studies demonstrated that the interferon (IFN) therapy efficacy is poor in the hepatitis C virus (HCV)-infected patients with lower miR-122 abundance in the livers. The hepatocarcinoma patients also have low miR-122 levels in their livers. We previously found that the IFN expression was reduced when miR-122 was knocked down in human oligodendrocytes. The mechanism is unclear. In this study, the miR-122-abundant cell Huh7 was used to explore the regulatory mechanism of miR-122 on type I IFN expression. We found that miR-122 significantly increased the type I IFN expression in Huh7 cells, while knocking down miR-122 decreased the type I IFN expression. By screening potential miR-122 targets among the negative regulators in IFN signaling pathways, we found that there were putative miR-122 targets in the suppressor of cytokine signaling 1 (SOCS1) mRNA. Over-expressing miR-122 decreased the SOCS1 expression by 50.55% in Huh7 cells, while knocking down miR-122 increased SOCS1 expression by 62.56%. Using a green fluorescence protein (EGFP) fused SOCS1-expressing plasmid, the SOCS1-EGFP fluorescence intensity and protein were lower in miR-122 mimic-treated cells than those in mock-miRNA-treated cells, while miR-122 knockdown significantly increased the SOCS1-EGFP fluorescence intensity and protein expression. Mutations in the nt359-nt375 region abandoned the impact of miR-122 on SOCS1-EGFP expression. Taken together, SOCS1 is a target of miR-122. MiR-122 can regulate the type I IFN expression through modulating the SOCS1 expression.

Borna disease virus encoded phosphoprotein inhibits host innate immunity by regulating miR-155

It has been reported that the Borna disease virus (BDV) encoded phosphoprotein (P protein) can inhibit the activity of Traf family member-associated NF-kappaB activator (TANK)-binding kinase 1 (TBK-1), thus preventing the induction of type I interferon (IFN). However, the effects of microRNA on the regulation of BDV infection and the hosts immune response have not been characterized. miR-155 was predicted to be complementary to the BDV P mRNA by RNAhybrid software. Here, we showed that miR-155 was down-regulated in BDV persistently infected human oligodendroglial (OL/BDV) cells and that the BDV P protein, but not the X protein, directly inhibited miR-155 expression in cells. When miR-155 was over-expressed, the inhibition of type I IFNs by BDV in cells was reversed, and the expression of type I IFNs was increased. When miR-155 expression was specifically blocked, cellular IFN expression and the induction of IFN by poly I:C treatment were suppressed. Furthermore, miR-155 promoted type I IFN production by targeting suppressor of cytokine signaling 1 (SOCS1) and SOCS3. Mutations in the nt1138-nt1158 region of SOCS3 abandoned the impact of miR-155 on the expression of SOCS3-enhanced green fluorescent protein (EGFP). The levels of BDV P mRNA and protein were significantly decreased in OL/BDV cells when miR-155 was over-expressed; however, miR-155-mutation did not affect the expression of BDV P-EGFP. Thus, BDV persistent infection inhibited the expression of type I IFNs through the suppression of miR-155, and miR-155 played an important immune regulatory role in BDV persistent infection.

MAVS-mediated host cell defense is inhibited by Borna disease virus.

Viruses often have strategies for preventing host cell apoptosis, which antagonizes viral replication. Borna disease virus (BDV) is a neurotropic RNA virus that establishes a non-cytolytic persistent infection. Although BDV suppresses type I Interferon (IFN) through (TANK)-binding kinase 1 (TBK-1) associated BDV P protein, it is still unclear how BDV can survive in the host cell and establish a persistent infection. Recently, it has been recognized that mitochondria-mediated apoptosis through the mitochondrial antiviral signaling protein (MAVS) and the RIG-I-like receptor (RLR) signaling pathway is a crucial component of the innate immune response. In this work we show that BDV X protein colocalizes and interacts with MAVS in the mitochondria to block programmed cell death. BDV X protein-mediated inhibition of apoptosis was independent of type I IFN production and NF-κB activity. The reduction of BDV X expression with RNA interference (RNAi) or the mutation of BDV X enhanced MAVS-induced cell death. Collectively, our data provide novel insights into how BDV X protein inhibits antiviral-associated programmed cell death, through its action of MAVS function.

Modulation of miR‑122 expression affects the interferon response in human hepatoma cells

Type I interferon (IFN) is believed to play significant roles in limiting tumor growth. It has been revealed that the induction of endogenous IFN expression is one of the key mechanisms for successful IFN therapy. However, recent studies have shown that the efficacy of type I IFN therapy has limitations in the clinical treatment of certain tumors, including hepatocellular carcinoma (HCC). It has been revealed that the expression of miR‑122 is significantly decreased in HCC and that restoration of miR‑122 expression may improve the prognosis of this condition. Previous studies also showed that patients with low miR‑122 levels in the liver responded poorly to the IFN therapy. We previously identified that the IFN expression was reduced when miR‑122 was suppressed in human oligodendrocytes. Based on these studies, it was hypothesized that the expression of miR‑122 may modulate the endogenous IFN expression and subsequently affect the treatment outcome of IFN therapy for HCC. The results of the present study showed that miR‑122‑abundant Huh7 cells responded more significantly than miR‑122‑deficient HepG2 cells when treated with exogenous IFN. Upregulation of miR‑122 significantly increased the ability of exogenous IFN‑induced IFN expression, while downregulation of miR‑122 decreased this ability. These data indicate that a high level of miR‑122 expression may promote the expression of type I IFN induced by exogenous IFNs and further contribute to IFN therapy for HCC.

Using functional magnetic resonance imaging to explore the possible mechanism of the action of acupuncture at Dazhong (KI 4) on the functional cerebral regions of healthy volunteers.

Acupuncture at right Dazhong (KI 4) mostly affects functional magnetic resonance imaging signal in the right inferior frontal gyrus, right insular lobe, right thalamus, right middle frontal gyrus and right orbitofrontal cortex, which are associated with governing executive functions, emotional activities and social behaviour.

Over-expression of mitochondrial antiviral signaling protein inhibits coxsackievirus B3 infection by enhancing type-I interferons production

BackgroundRecent studies have revealed that Mitochondrial Antiviral Signaling (MAVS) protein plays an essential role in the inhibition of viral infection through type I interferon (IFN) pathway. It has been shown that 3C (pro) cysteine protease of coxsackievirus B3 (CVB3) cleaves MAVS to inhibit type I IFNs induction. Other workers also found that MAVS knock-out mice suffered CVB3 susceptibility and severe histopathological change. Accordingly,our experiments were designed to explore the protection of over-expressing MAVS against CVB3 infection and the possible mechanism.ResultsIn this study, HeLa cells (transfected with MAVS constructs pre- or post- exposure to CVB3) were used to analyze the function of exogenous MAVS on CVB3 infection. The results revealed that though CVB3 infection induced production of type I IFNs, viral replication and cell death were not effectively inhibited. Similarly, exogenous MAVS increased type I IFNs moderately. Morever, we observed robust production of type I IFNs in CVB3 post-infected HeLa cells thereby successfully inhibiting CVB3 infection, as well formation of cytopathic effect (CPE) and cell death. Finally, introduction of exogenous MAVS into CVB3 pre-infected cells also restricted viral infection efficiently by greatly up-regulating IFNs.ConclusionsIn summary, exogenous MAVS effectively prevents and controls CVB3 infection by modulating and promoting the production of type I IFNs. The IFNs level in MAVS over-expressing cells is still tightly regulated by CVB3 infection. Thus, the factors that up-regulate MAVS might be an alternative prescription in CVB3-related syndromes by enhancing IFNs production.

Chlorpromazine inhibits mitochondrial apoptotic pathway via increasing expression of tissue factor

Chlorpromazine (CPZ) is a widely used antipsychotic drug with antagonistic effect on dopamine receptors. Accumulating evidence has shown that CPZ plays a neuroprotective role in various models of toxicity and apoptosis. However, the underlying mechanism contributing to this protective effect remains unclear. Here, we evaluate the effect of CPZ on mitochondrial apoptotic pathway in the neuron system. Higher levels of B-cell lymphoma-2 (Bcl-2) and tissue factor (TF) but lower apoptotic rate were found in hippocampus of CPZ-treated schizophrenic patients compared with non-antipsychotic treated controls. Additionally, both short-term and long-term treatment of CPZ in rats could up-regulate the levels of Bcl-2 and TF with no cytotoxic effects. In the in vitro experiment, expression of Bcl-2 was up-regulated in the C6 glioma cells transfected with pEGFP-N1-TF recombinant plasmid. Furthermore, in another independent rat model of apoptosis, compared with the group administrated with alcohol only, the brains of the CPZ-pretreated rats showed lower expression of cleaved caspase-3, cytochrome c and Bax, but higher expression of Bcl-2 and TF. Our data demonstrate that CPZ exerts its neuronal protective effects through inhibiting the activation of mitochondrial apoptotic pathway by up-regulating TF expression, thus providing new insight into CPZ function and application.

MiR-122 Directly Inhibits Human Papillomavirus E6 Gene and Enhances Interferon Signaling through Blocking Suppressor of Cytokine Signaling 1 in SiHa Cells

Human Papillomavirus (HPV) 16 infection is considered as one of the significant causes of human cervical cancer. The expression of the viral oncogenes like E6 and E7 play an important role in the development of the cancer. MiR-122 has been reported to exhibit a strong relationship with hepatitis viruses and take part in several tumor development, while the effects of miR-122 on HPV infection and the HPV viral oncogenes expression still remain unexplored. In this study, using RNAhybrid software, the potential binding sites between miR-122 and HPV16 E6 and E7 mRNAs were identified. Over and loss of miR-122 function showed that miR-122 could directly bind with HPV16 E6 mRNA and significantly inhibit its expression in SiHa cells, which was further confirmed by constructing the miR-122-E6-mu to eliminate the miR-122 binding effects with E6. The increase of the expression of type I interferon (IFN) and its classical effective molecules and the phosphorylation of signal transducers and activators of transcription (STAT1) protein indicated that miR-122 might enhance type I interferon in cervical carcinoma cells, which explained the significant reduction of HPV16 E7 and E6*I mRNA expression. This might be due to the binding between miR-122 and suppressor of cytokine signaling 1 (SOCS1) mRNA, which is the suppressor of interferon signaling pathway. Moreover, it was identified that the miR-122 binding position was nt359-nt375 in SOCS1 mRNA. Taken together, this study indicated that HPV16 could be effectively inhibited by miR-122 through both direct binding with E6 mRNA and promoting SOCS1-dependent IFN signaling pathway. Thus, miR-122 may serve as a new therapeutic option for inhibiting HPV infection.

Infection of Borna disease virus in healthy animals in northern China

Borna disease virus (BDV) is a non-segmented, single negative stranded RNA virus. It is highly neurotropic, but the infection is not cytopathic to the infected cells. BDV is widely infected in vertebrates, causing abnormal behavior and multiple pathological changes. Horses and sheep are the main natural hosts of BDV. Borna disease (BD) is characterized by progressive mononuclear encephalomyelitis. The BD clinical manifestations include ataxia, depression, rotation movement, mandatory standing, hitting objects with body and paralysis, and they usually last for 1-3 weeks. BDV infection causes a significant health problem in animals and economic loss for the society. However, BDV natural infection frequency in the raised animals in northern China, is poorly understood, but is required for understanding BDV infection epidemiology. BDV infection can be diagnosed through detecting viral proteins, antibodies and nucleic acid. The fluorescence based quantitative PCR is commonly used for BDV diagnosis because of the superior sensitivity. The assays for detecting plasma BDV specific antibodies include immunofluorescence, Western Blot and ELISA. Western Blot offers comparatively high specificity, which can detect different antibodies on the same blot/strip, and is often used as the confirmatory assay. This study investigated the infection prevalence of BDV in the meadow that raised horses and sheep in northern China. Blood samples were collected from a herd of healthy horses and sheep that were meadow raised in Hailin City, Heilongjiang Province, China. The uncoagulated blood samples were used to isolate peripheral lymphocytes and plasma. We selected Mongolian horses which are stocky, with relatively short but strong legs and a large head. The selected Han sheep is relatively small, has short tail and weighs 35-45 kg on average. BDV p40 mRNA was detected by Taqman based quantitative RT-PCR in lymphocytes of horses and sheep. BDV His-phosphoprotein (P) and His-nucleoprotein (N) fusion proteins were isolated with Histidine coated beads and the purity and immunological activity of the isolated viral proteins were assessed and verified by SDS-PAGE and Western Blot. Plasma BDV antibodies in horses and sheep were detected using the purified fusion proteins as antigens. Quantitative PCR assay showed that 50.00% (16/32) of the tested horse samples were positive for BDV p40 mRNA, and 20.83% (5/24) positive for BDV p40 mRNA in the sheep samples. The purified BDV His-P fusion protein in SDS-PAGE electrophoresis migrated at expected 24 kD size. A known specific BDV P antibody verified the immunogenicity of the purified BDV P in Western Blot, and BDV persistently infected human oligodendrocytes as positive control. The purified BDV His-N fusion protein appeared at the expected 40 kD position and the immunogenicity was also verified. The positive rates of BDV P antibody and N antibody were 11.11% (4/36) and 5.56% (2/36) in the tested horses, respectively. The positive rates of BDV P antibody and N antibody were 13.16% (5/38) and 7.89% (3/38) in the tested sheep, respectively. In conclusion, we utilized the expressed BDV His-P and His-N fusion proteins as antigen and established a modified Western Blot assay for detecting plasma BDV antibodies. Our data showed the direct evidence that natural BDV infection occurred in the meadow raised healthy horses and sheep in northern China.

A functional magnetic resonance imaging study of the neuronal specificity of an acupoint: acupuncture at Rangu (KI 2) and its sham point

The neuronal specificity of acupoints has not been entirely supported by the results of previous functional magnetic resource imaging studies. This study tested the specificity of an acupoint using right Rangu (KI 2) and its sham acupoint. The results showed specific cerebral response patterns and thus provided the evidence of the existence of acupoint neuronal specificity.