ABSTRACT
Objective:To investigate the effects of macrophage migration inhibitory factor (MIF) on AMPK/ERK/mTOR autophagy signaling pathway in primary human umbilical vein endothelial cells (HUVEC) after dengue virus type 2 (DENV2) infection.Methods:The virulence of DENV2 to C6/36 cells was assessed with 50% tissue culture infectious dose (TCID 50). NS1 gene fragments in DENV2-infected HUVEC were detected by RT-PCR. Transmission electron microscopy was used to detect autophagosomes. Western blot was performed to detect the effects of DENV2 infection on the expression of autophagy-related protein LC3-Ⅱ and MIF in HUVEC. The correction of MIF with LC3-Ⅱ was then analyzed. HUVEC were pretreated with MIF inhibitor (ISO-1) or pathway inhibitor (Compound C or U0126), and then the changes in the expression of MIF, adenosine 5′-monophosphate-activated protein kinase (AMPK) pathway-related proteins and LC3-Ⅱ after DENV2 infection were detected by Western blot to reveal the correlation between MIF and AMPK autophagy pathway. Results:The TCID 50 to C6/36 cells was 10 -9.09/ml in this experiment. NS1 gene fragments were detected in DENV2-infected HUVEC. Autophagosomes or autophagolysosomes were observed in the infected HUVEC and there were differences in autophagy induced by different doses of DENV2. The mRNA levels of MIF and LC3-Ⅱ in HUVEC were positively correlated after DENV2 infection. MIF inhibitor affected AMPK, ERK and LC3-Ⅱ levels, but had no significant influence on MIF expression at protein level. Conclusions:MIF could affect autophagy through regulating the AMPK/ERK/mTOR signaling pathway in HUVEC during DENV2 infection.
ABSTRACT
Background: Dengue is one of the most important mosquito-borne viral diseases in the world. The emergence and spread of four dengue viruses (DENVs) (serotypes) represent a global pandemic. The four distinct serotypes are, namely, DENV-1, DENV-2, DENV-3 and DENV-4. Very few dengue serotyping studies have been reported from Andhra Pradesh. In this context, the present study focuses on the circulating serotypes of dengue in South-Eastern Andhra Pradesh. Methodology: Study was done at Sri Venkateswara Institute of Medical Sciences, a teaching hospital in Tirupati, Andhra Pradesh. Acute phase dengue serum samples were collected and tested for NS1 antigen and anti-human IgM antibodies by enzyme-linked immunosorbent assay (ELISA). NS1-positive samples were further serotyped by reverse transcriptase real-time polymerase chain reaction (rRT-PCR). Results: A total of 398 serum samples were received from clinically suspected dengue fever cases. Of these, 150 (37.7%) samples were positive for NS1 and/or IgM ELISA. The 96 NS1 antigen-positive samples were further processed for serotyping, of which 36 were negative by rRT-PCR. DENV-2 (41%) was the predominant serotype, followed by DENV-4 (37%), DENV-3 (12%) and DENV-1 (10%) in descending order. Conclusion: This study reports the all four dengue serotypes' co-circulation. This is the first report from South Eastern Andhra Pradesh. Amongst four, DENV-2 was predominant followed by DENV-4. The information of predominant serotypes can guide in forecasting dengue outbreaks and improving control measures of vectors thus may be helpful in the prevention of outbreaks.