Chemical Chaperone 4-PBA Mitigates Tumor Necrosis Factor Alpha-Induced Endoplasmic Reticulum Stress in Human Airway Smooth Muscle

Airway inflammation and pro-inflammatory cytokines such as tumor necrosis factor alpha (TNF alpha) underlie the pathophysiology of respiratory diseases, including asthma. Previously, we showed that TNF alpha activates the inositol-requiring enzyme 1 alpha (IRE1 alpha)/X-box binding protein 1 spliced (XBP1s) endoplasmic reticulum (ER) stress pathway in human airway smooth muscle (hASM) cells. The ER stress pathway is activated by the accumulation of unfolded proteins in the ER. Accordingly, chemical chaperones such as 4-phenylbutyric acid (4-PBA) may reduce ER stress activation. In the present study, we hypothesized that chemical chaperone 4-PBA mitigates TNF alpha-induced ER stress in hASM cells. hASM cells were isolated from bronchiolar tissue obtained from five patients with no history of smoking or respiratory diseases. The hASM cells' phenotype was confirmed via the expression of alpha-smooth muscle actin and elongated morphology. hASM cells from the same patient sample were then separated into three 12 h treatment groups: (1) TNF alpha (20 ng/mL), (2) TNF alpha + 4-PBA (1 mu M, 30 min pretreatment), and (3) untreated control. The expressions of total IRE1 alpha and phosphorylated IRE1 alpha (pIRE1 alpha S724) were determined through Western blotting. The splicing of XBP1 mRNA was analyzed using RT-PCR. We found that TNF alpha induced an increase in pIRE1 alpha S724 phosphorylation, which was mitigated by treatment with chemical chaperone 4-PBA. We also found that TNF alpha induced an increase in XBP1s mRNA, which was also mitigated by treatment with chemical chaperone 4-PBA. These results support our hypothesis and indicate that chemical chaperone 4-PBA treatment mitigates TNF alpha-induced ER stress in hASM cells.

Automated summary

Airway inflammation and pro-inflammatory cytokines play a significant role in the development of respiratory diseases. In this study, we investigated the potential of the chemical chaperone 4-PBA to mitigate TNFα-induced ER stress in hASM cells.