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Measuring Dengue Virus RNA in the Culture Supernatant of Infected Cells by Real-time Quantitative Polymerase Chain Reaction
Published on: November 1, 2018
Different Responses in MMP/TIMP Expression of U937 and HepG2 Cells to Dengue Virus Infection
Pannatas Seanpong1, Chanya Srisaowakarn, Anothai Thammaporn
1Department of Biology, Faculty of Science, Mahidol University.
Abstract:
Disease severities following dengue virus (DV) infection are the result of increased vascular permeability leading to hypovolemic shock. Matrix metalloproteinases (MMPs) are believed to play a key role in promoting such severities. A previous study reported that supernatants of DV-infected dendritic cells (DCs), which contained high levels of MMP-2 and MMP-9, induced vascular leakage in a mouse model. In the present study, we investigated whether hepatocytes (HepG2) and monocytes (U937) could be additional sources of MMPs during DV infection. HepG2 and U937 cells were exposed to DV serotype 2 strain 16681. The secretion of MMP-2 and MMP-9 was detected using gelatin zymography. We found that DV infection in the HepG2 cells promoted MMP-2 production while that in the U937 cells promoted MMP-9 production. Semi-quantitative RT-PCR results also confirmed that DV infection in the HepG2 cells up-regulated the expression of MMP-2 mRNA, whereas that in the U937 cells enhanced the expression of MMP-9 mRNA. We monitored the expression of endogenous TIMP-1 and TIMP-2. DV infection induced TIMP-1 expression in the U937 cells. However, lower expression of TIMP-2 was observed in the infected HepG2 cells. We believed that following DV infection, monocytes and hepatocytes can act as MMP-9 and MMP-2 producers, respectively. Their responses could be attributed to the disturbance of TIMP expression by DV in different cell types.
Insights
Dengue virus (DV) infection increases vascular permeability. This study shows that hepatocytes produce MMP-2 and monocytes produce MMP-9 during DV infection, contributing to disease severity.
Area of Science:
- Virology
- Immunology
- Biochemistry
Background:
- Dengue virus (DV) infection causes severe disease through increased vascular permeability.
- Matrix metalloproteinases (MMPs), specifically MMP-2 and MMP-9, are implicated in DV-induced vascular leakage.
- Previous research identified dendritic cells as a source of MMPs during DV infection.
Purpose of the Study:
- To investigate if hepatocytes (HepG2) and monocytes (U937) are additional sources of MMPs during DV infection.
- To analyze the expression of MMP-2, MMP-9, and their inhibitors (TIMPs) in HepG2 and U937 cells upon DV infection.
Main Methods:
- HepG2 and U937 cells were infected with DV serotype 2.
- MMP-2 and MMP-9 secretion was measured using gelatin zymography.
- mRNA expression of MMP-2 and MMP-9 was assessed via semi-quantitative RT-PCR.
- Expression of endogenous TIMP-1 and TIMP-2 was monitored.
Main Results:
- DV infection of HepG2 cells promoted MMP-2 production and MMP-2 mRNA expression.
- DV infection of U937 cells promoted MMP-9 production and MMP-9 mRNA expression.
- DV infection induced TIMP-1 in U937 cells but decreased TIMP-2 in HepG2 cells.
Conclusions:
- Hepatocytes and monocytes serve as significant producers of MMP-2 and MMP-9, respectively, during dengue virus infection.
- The differential regulation of MMPs and TIMPs by DV in these cell types contributes to disease pathogenesis.
- Understanding these cellular responses provides insights into dengue's vascular complications.
