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TNF-alpha and IL-6 inhibit apolipoprotein A-IV production induced by linoleic acid in human intestinal Caco2 cells.

Li X, Xu M, Liu M, Ji Y, Li Z - J Inflamm (Lond) (2015)

Bottom Line: LA stimulated gene and protein expression of apoA-IV in a dose and time dependent manner.Pre-treatment with the cytokines for 72 h significantly inhibited the increased expression of apoA-IV gene and protein induced by LA.Our data indicate that the pro-inflammatory cytokines may be responsible for the reduced apoA-IV production in response to fat feeding.

View Article: PubMed Central - PubMed

Affiliation: National Local Joint Engineering Research Center of Biodiagnostics and Biotherapy, The Second Affiliated Hospital of Medical College, Xi'an Jiaotong University, Xi'an, 710004 China ; Department of Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati, 45237-0507 USA.

ABSTRACT

Background: Apolipoprotein A-IV (apoA-IV) is a protein mainly synthesized by enterocytes in the intestine. Its gene expression is suppressed during fasting and stimulated during active fat absorption. Chronic feeding of a high-fat (HF) diet abolishes the differential expression between fasting and fat-feeding and therefore may contribute to diet-induced obesity since apoA-IV is a potent satiety factor. It is well established that the circulating pro-inflammatory cytokines TNF-α and IL-6 are increased by HF feeding.

Methods: To determine whether pro-inflammatory cytokines are involved in the diminished response of apoA-IV gene expression to fat-feeding, different concentrations of linoleic acid (LA), an important dietary fatty acid, was used to stimulate apoA-IV expression in human intestinal Caco2 cells. Cells were pre-treated with or without human recombinant TNF-α, IL-6 or their combination before the addition of LA. Real-time PCR and ELISA were used to detect and quantify RNA transcripts and proteins of apoA-IV and the cytokines.

Results: LA stimulated gene and protein expression of apoA-IV in a dose and time dependent manner. Pre-treatment with the cytokines for 72 h significantly inhibited the increased expression of apoA-IV gene and protein induced by LA. Furthermore, the cytokines, especially TNF-α, also positively up-regulate the cytokine themselves in Caco2 cells.

Conclusions: Our data indicate that the pro-inflammatory cytokines may be responsible for the reduced apoA-IV production in response to fat feeding. Because of apoA-IV's role in satiety, we propose the inhibitory effect of circulating pro-inflammatory cytokines on apoA-IV production contributes to diet-induced obesity.

No MeSH data available.


Related in: MedlinePlus

LA stimulated gene expression ofIl6andTnfαin Caco2 cells. Differentiated Caco2 cells were supplemented with or without 1 mM LA for the indicated times for mRNA measurement or with different amount of LA for 24 h for protein measurement. The mRNA levels of Il6 and Tnfα were measured by real time RT-PCR (A and B). Their proteins from culture medium were measured by ELISA (C and D). *P < 0.05, ** P <0.01, *** P <0.001 vs. vehicle control.
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Fig3: LA stimulated gene expression ofIl6andTnfαin Caco2 cells. Differentiated Caco2 cells were supplemented with or without 1 mM LA for the indicated times for mRNA measurement or with different amount of LA for 24 h for protein measurement. The mRNA levels of Il6 and Tnfα were measured by real time RT-PCR (A and B). Their proteins from culture medium were measured by ELISA (C and D). *P < 0.05, ** P <0.01, *** P <0.001 vs. vehicle control.

Mentions: It has been reported that long-chain fatty acid, e.g. the LA, can enhance Il6 production in rat intestinal epithelial cells [21]. To determine whether LA is able to affect gene expressions of endogenous cytokines IL-6 and TNF-α in the Caco2 cells, we measured mRNA levels of Il6 and Tnfα at the indicated times of LA treatment from Caco2 cells and their proteins released from the cells after the treatment with different amount of LA for 24 h. We found that the LA stimulated Tnfα and Il6 gene expressions sooner than apoA-IV expression. The mRNA level of Il6 started to increase by 1.03 fold after 3 h treatment, although the difference is not statistically significant, and then reached to 3.4 fold increase at 6 h (Figure 3A). The mRNA level of Tnfα was significantly increased sooner than Il6 mRNA and it reached to 3.57 fold at 2 h, 3.52 fold increases at 3 h and then reduced to 1.4 fold increases by 6 h (Figure 3B). It is noteworthy that the pattern of Tnfα gene expression was similar to Il6, and all dropped back to basal level at 12 h, but Il6 start to express again at 24 h post-treatment of LA (Figure 4A), which could be positive feedback regulation from the other factors, such as TNF-α, as a result from LA stimulation. We also measured their protein levels and found that 0.5 mM LA had the maximum effects on both IL-6 and TNF-α protein levels although 0.25 mM and 1 mM LA increased the cytokine protein productions, too (Figure 3C and D).Figure 3


TNF-alpha and IL-6 inhibit apolipoprotein A-IV production induced by linoleic acid in human intestinal Caco2 cells.

Li X, Xu M, Liu M, Ji Y, Li Z - J Inflamm (Lond) (2015)

LA stimulated gene expression ofIl6andTnfαin Caco2 cells. Differentiated Caco2 cells were supplemented with or without 1 mM LA for the indicated times for mRNA measurement or with different amount of LA for 24 h for protein measurement. The mRNA levels of Il6 and Tnfα were measured by real time RT-PCR (A and B). Their proteins from culture medium were measured by ELISA (C and D). *P < 0.05, ** P <0.01, *** P <0.001 vs. vehicle control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4389805&req=5

Fig3: LA stimulated gene expression ofIl6andTnfαin Caco2 cells. Differentiated Caco2 cells were supplemented with or without 1 mM LA for the indicated times for mRNA measurement or with different amount of LA for 24 h for protein measurement. The mRNA levels of Il6 and Tnfα were measured by real time RT-PCR (A and B). Their proteins from culture medium were measured by ELISA (C and D). *P < 0.05, ** P <0.01, *** P <0.001 vs. vehicle control.
Mentions: It has been reported that long-chain fatty acid, e.g. the LA, can enhance Il6 production in rat intestinal epithelial cells [21]. To determine whether LA is able to affect gene expressions of endogenous cytokines IL-6 and TNF-α in the Caco2 cells, we measured mRNA levels of Il6 and Tnfα at the indicated times of LA treatment from Caco2 cells and their proteins released from the cells after the treatment with different amount of LA for 24 h. We found that the LA stimulated Tnfα and Il6 gene expressions sooner than apoA-IV expression. The mRNA level of Il6 started to increase by 1.03 fold after 3 h treatment, although the difference is not statistically significant, and then reached to 3.4 fold increase at 6 h (Figure 3A). The mRNA level of Tnfα was significantly increased sooner than Il6 mRNA and it reached to 3.57 fold at 2 h, 3.52 fold increases at 3 h and then reduced to 1.4 fold increases by 6 h (Figure 3B). It is noteworthy that the pattern of Tnfα gene expression was similar to Il6, and all dropped back to basal level at 12 h, but Il6 start to express again at 24 h post-treatment of LA (Figure 4A), which could be positive feedback regulation from the other factors, such as TNF-α, as a result from LA stimulation. We also measured their protein levels and found that 0.5 mM LA had the maximum effects on both IL-6 and TNF-α protein levels although 0.25 mM and 1 mM LA increased the cytokine protein productions, too (Figure 3C and D).Figure 3

Bottom Line: LA stimulated gene and protein expression of apoA-IV in a dose and time dependent manner.Pre-treatment with the cytokines for 72 h significantly inhibited the increased expression of apoA-IV gene and protein induced by LA.Our data indicate that the pro-inflammatory cytokines may be responsible for the reduced apoA-IV production in response to fat feeding.

View Article: PubMed Central - PubMed

Affiliation: National Local Joint Engineering Research Center of Biodiagnostics and Biotherapy, The Second Affiliated Hospital of Medical College, Xi'an Jiaotong University, Xi'an, 710004 China ; Department of Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati, 45237-0507 USA.

ABSTRACT

Background: Apolipoprotein A-IV (apoA-IV) is a protein mainly synthesized by enterocytes in the intestine. Its gene expression is suppressed during fasting and stimulated during active fat absorption. Chronic feeding of a high-fat (HF) diet abolishes the differential expression between fasting and fat-feeding and therefore may contribute to diet-induced obesity since apoA-IV is a potent satiety factor. It is well established that the circulating pro-inflammatory cytokines TNF-α and IL-6 are increased by HF feeding.

Methods: To determine whether pro-inflammatory cytokines are involved in the diminished response of apoA-IV gene expression to fat-feeding, different concentrations of linoleic acid (LA), an important dietary fatty acid, was used to stimulate apoA-IV expression in human intestinal Caco2 cells. Cells were pre-treated with or without human recombinant TNF-α, IL-6 or their combination before the addition of LA. Real-time PCR and ELISA were used to detect and quantify RNA transcripts and proteins of apoA-IV and the cytokines.

Results: LA stimulated gene and protein expression of apoA-IV in a dose and time dependent manner. Pre-treatment with the cytokines for 72 h significantly inhibited the increased expression of apoA-IV gene and protein induced by LA. Furthermore, the cytokines, especially TNF-α, also positively up-regulate the cytokine themselves in Caco2 cells.

Conclusions: Our data indicate that the pro-inflammatory cytokines may be responsible for the reduced apoA-IV production in response to fat feeding. Because of apoA-IV's role in satiety, we propose the inhibitory effect of circulating pro-inflammatory cytokines on apoA-IV production contributes to diet-induced obesity.

No MeSH data available.


Related in: MedlinePlus