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Substitution in Amino Acid 70 of Hepatitis C Virus Core Protein Changes the Adipokine Profile via Toll-Like Receptor 2/4 Signaling.

Uraki S, Tameda M, Sugimoto K, Shiraki K, Takei Y, Nobori T, Ito M - PLoS ONE (2015)

Bottom Line: The effects of toll-like receptor (TLR)2/4 inhibition on IL-6 production by 3T3-L1 induced by HCV core protein were examined.IL-6 production was significantly increased and adiponectin production was reduced without a change in triglyceride content by treatment with 70Q compared to 70R core protein in both murine and human adipocytes.IL-6 induction of 3T3-L1 cells treated by 70Q HCV core protein was significantly inhibited with anti-TLR2 antibody by 42%, and by TLR4 inhibitor by 40%.

View Article: PubMed Central - PubMed

Affiliation: First Department of Internal Medicine, Mie University School of Medicine, Tsu, Japan.

ABSTRACT

Background & aims: It has been suggested that amino acid (aa) substitution at position 70 from arginine (70R) to glutamine (70Q) in the genotype 1b hepatitis C virus (HCV) core protein is associated with insulin resistance and worse prognosis. However, the precise mechanism is still unclear. The aim of this study was to investigate the impact of the substitution at position 70 in HCV core protein on adipokine production by murine and human adipocytes.

Methods: The influence of treatment with HCV core protein (70R or 70Q) on adipokine production by both 3T3-L1 and human adipocytes were examined with real-time PCR and enzyme-linked immunosorbent assay (ELISA), and triglyceride content was also analyzed. The effects of toll-like receptor (TLR)2/4 inhibition on IL-6 production by 3T3-L1 induced by HCV core protein were examined.

Results: IL-6 production was significantly increased and adiponectin production was reduced without a change in triglyceride content by treatment with 70Q compared to 70R core protein in both murine and human adipocytes. IL-6 induction of 3T3-L1 cells treated by 70Q HCV core protein was significantly inhibited with anti-TLR2 antibody by 42%, and by TLR4 inhibitor by 40%.

Conclusions: Our study suggests that extracellular HCV core protein with substitution at position 70 enhanced IL-6 production and reduced adiponectin production from visceral adipose tissue, which can cause insulin resistance, hepatic steatosis, and ultimately development of HCC.

No MeSH data available.


Related in: MedlinePlus

Changes in protein levels of adipokines in 3T3-L1 cells treated with HCV core protein.(A) The protein level of IL-6 was significantly increased, and the protein levels of (B) adiponectin and (C) leptin were significantly reduced in cells treated with 70Q for 48 h compared with 70R and GST protein. All data are expressed as the average ± SEM of three independent experiments. The levels of mRNA are shown as ratios relative to GST treated cells.
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pone.0131346.g002: Changes in protein levels of adipokines in 3T3-L1 cells treated with HCV core protein.(A) The protein level of IL-6 was significantly increased, and the protein levels of (B) adiponectin and (C) leptin were significantly reduced in cells treated with 70Q for 48 h compared with 70R and GST protein. All data are expressed as the average ± SEM of three independent experiments. The levels of mRNA are shown as ratios relative to GST treated cells.

Mentions: First, we treated mouse 3T3-L1 cells, which differentiated into adipocytes, with HCV core recombinant protein. Since it has been reported that adipocytes secrete adipokines such as IL-6, TNF-α, adiponectin, and leptin, we examined mRNA expressions of these adipokines in 3T3-L1 cells treated with 2 pmol/l of GST, 70R, and 70Q protein by qRT-PCR. The level of IL-6 mRNA expression was significantly increased in 3T3-L1 cells treated with 70Q for 24 h compared with wild HCV core protein (Fig 1A). On the other hand, 70Q induced significant reduce in mRNA expressions of adiponectin and leptin compared to control (GST), though the differences between 70R and 70Q-treated cells did not reach statistical significance due to wide variation of data range (Fig 1B and 1C). There was no change in mRNA expression of other adipokine such as TNF-α, PAI-1 and resistin (data not shown). Next, we performed ELISA to measure the changes in protein levels of these adipokines. Similar to the changes in mRNA expression, the protein level of IL-6 was significantly increased in 70Q cells (Fig 2A), and the levels of adiponectin and leptin were significantly reduced in 70Q cells (Fig 2B and 2C). The level of TNF-α could be measured by neither RT-PCR nor ELISA because of its low expression levels. Moreover, 70Q HCV protein augmented the IL-6 production and reduced adiponectin production from 3T3-L1 cells in a dose-dependent manner (Fig 3A and 3B) and IL-6 production was higher and adiponectin production were lower in the cells treated with 70Q compared to 70R in each concentration.


Substitution in Amino Acid 70 of Hepatitis C Virus Core Protein Changes the Adipokine Profile via Toll-Like Receptor 2/4 Signaling.

Uraki S, Tameda M, Sugimoto K, Shiraki K, Takei Y, Nobori T, Ito M - PLoS ONE (2015)

Changes in protein levels of adipokines in 3T3-L1 cells treated with HCV core protein.(A) The protein level of IL-6 was significantly increased, and the protein levels of (B) adiponectin and (C) leptin were significantly reduced in cells treated with 70Q for 48 h compared with 70R and GST protein. All data are expressed as the average ± SEM of three independent experiments. The levels of mRNA are shown as ratios relative to GST treated cells.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4487891&req=5

pone.0131346.g002: Changes in protein levels of adipokines in 3T3-L1 cells treated with HCV core protein.(A) The protein level of IL-6 was significantly increased, and the protein levels of (B) adiponectin and (C) leptin were significantly reduced in cells treated with 70Q for 48 h compared with 70R and GST protein. All data are expressed as the average ± SEM of three independent experiments. The levels of mRNA are shown as ratios relative to GST treated cells.
Mentions: First, we treated mouse 3T3-L1 cells, which differentiated into adipocytes, with HCV core recombinant protein. Since it has been reported that adipocytes secrete adipokines such as IL-6, TNF-α, adiponectin, and leptin, we examined mRNA expressions of these adipokines in 3T3-L1 cells treated with 2 pmol/l of GST, 70R, and 70Q protein by qRT-PCR. The level of IL-6 mRNA expression was significantly increased in 3T3-L1 cells treated with 70Q for 24 h compared with wild HCV core protein (Fig 1A). On the other hand, 70Q induced significant reduce in mRNA expressions of adiponectin and leptin compared to control (GST), though the differences between 70R and 70Q-treated cells did not reach statistical significance due to wide variation of data range (Fig 1B and 1C). There was no change in mRNA expression of other adipokine such as TNF-α, PAI-1 and resistin (data not shown). Next, we performed ELISA to measure the changes in protein levels of these adipokines. Similar to the changes in mRNA expression, the protein level of IL-6 was significantly increased in 70Q cells (Fig 2A), and the levels of adiponectin and leptin were significantly reduced in 70Q cells (Fig 2B and 2C). The level of TNF-α could be measured by neither RT-PCR nor ELISA because of its low expression levels. Moreover, 70Q HCV protein augmented the IL-6 production and reduced adiponectin production from 3T3-L1 cells in a dose-dependent manner (Fig 3A and 3B) and IL-6 production was higher and adiponectin production were lower in the cells treated with 70Q compared to 70R in each concentration.

Bottom Line: The effects of toll-like receptor (TLR)2/4 inhibition on IL-6 production by 3T3-L1 induced by HCV core protein were examined.IL-6 production was significantly increased and adiponectin production was reduced without a change in triglyceride content by treatment with 70Q compared to 70R core protein in both murine and human adipocytes.IL-6 induction of 3T3-L1 cells treated by 70Q HCV core protein was significantly inhibited with anti-TLR2 antibody by 42%, and by TLR4 inhibitor by 40%.

View Article: PubMed Central - PubMed

Affiliation: First Department of Internal Medicine, Mie University School of Medicine, Tsu, Japan.

ABSTRACT

Background & aims: It has been suggested that amino acid (aa) substitution at position 70 from arginine (70R) to glutamine (70Q) in the genotype 1b hepatitis C virus (HCV) core protein is associated with insulin resistance and worse prognosis. However, the precise mechanism is still unclear. The aim of this study was to investigate the impact of the substitution at position 70 in HCV core protein on adipokine production by murine and human adipocytes.

Methods: The influence of treatment with HCV core protein (70R or 70Q) on adipokine production by both 3T3-L1 and human adipocytes were examined with real-time PCR and enzyme-linked immunosorbent assay (ELISA), and triglyceride content was also analyzed. The effects of toll-like receptor (TLR)2/4 inhibition on IL-6 production by 3T3-L1 induced by HCV core protein were examined.

Results: IL-6 production was significantly increased and adiponectin production was reduced without a change in triglyceride content by treatment with 70Q compared to 70R core protein in both murine and human adipocytes. IL-6 induction of 3T3-L1 cells treated by 70Q HCV core protein was significantly inhibited with anti-TLR2 antibody by 42%, and by TLR4 inhibitor by 40%.

Conclusions: Our study suggests that extracellular HCV core protein with substitution at position 70 enhanced IL-6 production and reduced adiponectin production from visceral adipose tissue, which can cause insulin resistance, hepatic steatosis, and ultimately development of HCC.

No MeSH data available.


Related in: MedlinePlus