ORIGINAL ARTICLES
XIAO Zhenyu, SUN Yangyang, YANG Yi, MA Yingkai, WANG Yongan, LUO Yuan
OBJCTIVE To evaluate the therapeutic efficacy of intranasal administration of pirfenidone in treating paraquat-induced pulmonary fibrosis in mice across treatment durations. METHODS Eight-week-old male C57BL/6 mice were randomly divided into six groups (n=8 per group): the normal control group (saline), pirfenidone control group, paraquat group, and three treatment groups receiving a combination of paraquat and pirfenidone for 15, 10 and 5 d, respectively. Except the normal and pirfenidone control groups, all the mice received intraperitoneal injection of paraquat (35 mg·kg-1) to induce pulmonary fibrosis. In the treatment groups, pirfenidone (20 mg·kg-1) was delivered intranasally once daily, beginning on days 1, 6, and 11 post-paraquat exposure, until day 15. Fifteen days after paraquat exposure, pulmonary function tests, micro-CT imaging, and arterial blood gas analysis were performed. Histopathological changes and collagen fiber deposition in lung tissues were examined using HE and Masson staining respectively.The protein expression levels of fibrosis markers, including fibronectin (FN), collagen typeⅠ(CollⅠ), E-cadherin (E-cad), vimentin (Vim), and α-smooth muscle actin (α-SMA), were detected by Western blotting. Additionally, inflammatory and pro-fibrotic cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-8, IL-1β, and connective tissue growth factor (CTGF), were quantified using ELISA. RESULTS Compared with the normal control group, mice treated with paraquat exhibited significant respiratory alterations, including prolonged expiratory time (TE), increased enhanced pause (PENH), and reduced tidal volume (TV). CT imaging revealed reticular high-density shadows and ground-glass opacities in paraquat-treated mice. Blood gas analysis showed reduced partial pressure of oxygen (PaO2), oxygen saturation of blood (SaO2), fractional oxygen saturation of hemoglobin (FO2Hb), and central venous oxygen saturation (ScvO2), along with increased partial pressure of carbon dioxide (PaCO2) and fractional deoxyhemoglobin saturation (FHHb), indicating that mice exposed to paraquat exhibited severe hypoxemia and hypercapnia. Histological evaluation highlighted pronounced lung interstitial thickening, alveolar collapse, inflammatory infiltration, and extensive fibrotic changes marked by collagen accumulation. Furthermore, exposure to paraquat significantly increased the protein levels of FN, CollⅠ, vimentin, and α-SMA, markedly reduced E-cadherin levels and elevated the levels of inflammatory cytokines (TNF-α, IL-6, IL-8 and IL-1β) as well as the pro-fibrotic cytokine CTGF. Pirfenidone treatment demonstrated time-dependent efficacy, with the 15 d group showing the most significant improvements in pulmonary function as evidenced by reduced PENH levels and increased TV, EV and MV. CT imaging revealed a decrease in high-density opacities and improved lung transparency after pirfenidone treatment. In addition, arterial blood gas measurements indicated markedly elevated levels of PaO2, SaO2 and FO2Hb. Histological analysis showed that pirfenidone alleviated lung interstitial thickening, reduced inflammatory cell infiltration, and decreased collagen deposition. At the molecular level, pirfenidone significantly reduced the protein expressions of FN, CollⅠ, Vim and α-SMA, while increasing E-cad levels. Furthermore, inflammatory cytokines, particularly IL-6 and IL-1β, were notably suppressed following pirfenidone intervention. CONCLUSION Intranasal administration of pirfenidone can exhibit potent time-dependent anti-fibrotic efficacy in paraquat-induced pulmonary fibrosis, with early interventions delivering the most substantial therapeutic benefits.
