Hepatic expression patterns of inflammatory and immune response genes associated with obesity and NASH in morbidly obese patients

Abstract

Background: Obesity modulates inflammation and activation of immune pathways which can lead to liver complications. We aimed at identifying expression patterns of inflammatory and immune response genes specifically associated with obesity and NASH in the liver of morbidly obese patients.

Methodology/principal findings: Expression of 222 genes was evaluated by quantitative RT-PCR in the liver of morbidly obese patients with histologically normal liver (n = 6), or with severe steatosis without (n = 6) or with NASH (n = 6), and in lean controls (n = 5). Hepatic expression of 58 out of 222 inflammatory and immune response genes was upregulated in NASH patients. The most notable changes occurred in genes encoding chemokines and chemokine receptors involved in leukocyte recruitment, CD and cytokines involved in the T cell activation towards a Th1 phenotype, and immune semaphorins. This regulation seems to be specific for the liver since visceral adipose tissue expression and serum levels of MCP1, IP10, TNFα and IL6 were not modified. Importantly, 47 other genes were already upregulated in histologically normal liver (e.g. CRP, Toll-like receptor (TLR) pathway). Interestingly, serum palmitate, known to activate the TLR pathway, was increased with steatosis.

Conclusion/significance: The liver of obese patients without histological abnormalities already displayed a low-grade inflammation and could be more responsive to activators of the TLR pathway. NASH was then characterized by a specific gene signature. These findings help to identify new potential actors of the pathogenesis of NAFLD.

Figure 1. Upregulation of 17 genes encoding proteins involved in leukocyte recruitment in liver of NASH patients.

The hepatic expression levels of genes were analyzed by real-time quantitative PCR in morbidly obese patients with normal liver histology (S0, n = 6), in morbidly obese patients with severe steatosis (S3, n = 6), and in morbidly obese patients with severe steatosis and NASH (NASH, n = 6). The mRNA levels of genes were normalized to the mRNA levels of RPLP0. Results are expressed relative to the expression levels in S0 patients and expressed as mean±SEM. *P<0.05, compared with S0 patients.

Figure 2. Specific upregulation of genes encoding Th1 cytokines and proteins involved in recognition between APC and T cells in liver of NASH patients.

A. The hepatic expression levels of genes were analyzed by real-time quantitative PCR in morbidly obese patients with normal liver histology (S0, n = 6), in morbidly obese patients with severe steatosis (S3, n = 6), and in morbidly obese patients with severe steatosis and NASH (NASH, n = 6). The mRNA levels of genes were normalized to the mRNA levels of RPLP0. Results are expressed relative to the expression levels in S0 patients and expressed as mean±SEM. *P<0.05, compared with S0patients; ^§P = 0,033. B. Scheme of recognition between APC and T cells.

Figure 3. Elevated serum levels of CCL2, IP10, IL6 and TNFα are dependent on obesity but not on liver complications.

The serum of 9 lean patients and 70 morbidly obese patients (15 S0 patients, 23 S3 patients and 23 NASH patients with severe steatosis) were used to evaluate the circulating levels of CCL2, IP10, IL6 and TNFα.

Figure 4. CRP gene expression levels in function to liver complications.

The expression levels of CRP were analyzed by real-time quantitative PCR in lean subjects (n = 6 for the liver; n = 4 for the SCAT), in morbidly obese patients with normal liver histology (S0,n = 6), in morbidly obese patients with severe steatosis (S3, n = 6), and in morbidly obese patients with severe steatosis and NASH (NASH, n = 6) in liver and subcutaneous adipose tissue. The mRNA levels of genes were normalized to the mRNA levels of RPLP0. Results are expressed relative to the expression levels in controls and expressed as mean±SEM. *P<0.05, compared with controls.

Figure 5. Elevated serum palmitate levels but not endoxin levels were present in severely steatotic obese patients.

A. The serum of 9 lean patients and 70 morbidly obese patients (15 S0 patients, 23 S3 patients and 23 NASH patients with severe steatosis) was used to evaluate the circulating levels of endotoxin. B. The abundances of palmitate as expressed relative to their values in a commercial serum sample (over 1000 individuals) were evaluated in 8 S0 patients, 9 S3 patients and 8 NASH patients.