Mechanism of Jinshui Huanxian Component Formula Ⅱ Combined with Matrix Softening in Ameliorating Pulmonary Fibrosis Through Inhibition of Epithelial-mesenchymal Transition
10.13422/j.cnki.syfjx.20260761
- VernacularTitle:金水缓纤组分方Ⅱ联合基质软化抑制上皮间质转化改善肺纤维化的作用机制
- Author:
Xiaojie WU
1
;
Mingyan LI
1
;
Shiyu KUANG
1
;
Xiling PENG
1
;
Xiaorong HOU
1
;
Han LIU
1
Author Information
1. Henan Key Laboratory of Chinese Medicine for Respiratory Disease， Collaborative Innovation Center for Chinese Medicine and Respiratory Diseases Co-constructed by Henan Province & Ministry of Education of China， Academy of Chinese Medical Sciences， Henan University of Chinese Medicine，Zhengzhou 450046，China
- Publication Type:Journal Article
- Keywords:
Jinshui Huanxian prescription;
component formula;
pulmonary fibrosis;
matrix hardness;
integrin β1;
epithelial-mesenchymal transition
- From:
Chinese Journal of Experimental Traditional Medical Formulae
2026;32(18):108-121
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo determine whether Jinshui Huanxian component formula Ⅱ （ECC-JHF Ⅱ） and matrix hardness interfere with the process of epithelial-mesenchymal transition （EMT） during pulmonary fibrosis by regulating integrin β1 （ITG β1）， and to elucidate the underlying mechanism. MethodsA pulmonary fibrosis mouse model was established by a single intratracheal instillation of bleomycin （BLM）. To observe the disease progression at various time points， the mice were sacrificed before modeling and on days 7， 14， 21， 28 and 42 after modeling for assessment of relevant indicators. To evaluate the therapeutic effect of drug intervention， the mice were given ECC-JHF Ⅱ and pirfenidone （PFD） by gavage starting on day 29 after modeling for 14 days of treatment， after which the mice were sacrificed. Pulmonary function， pathological changes， collagen deposition， lung tissue hardness and EMT markers were detected. Polyethylene glycol （PEG） composite hydrogels with different hardness were prepared in vitro using biomaterials such as eight-arm polyethylene glycol thiol （PEG-SH） and eight-arm polyethylene glycol maleimide （PEG-MAL）. Human type Ⅱ alveolar epithelial cells cultured in hard matrix culture dishes were treated with low， medium， and high doses of ECC-JHF Ⅱ （15.31， 30.63， 61.25 mg·L-1） to observe the intervention effect of ECC-JHF Ⅱ on EMT induced by transforming growth factor beta 1 （TGF-β1）. Human type Ⅱ alveolar epithelial cells cultured on soft （3.8 kPa） and hard （culture dishes， ~ GPa） matrices were treated with ECC-JHF Ⅱ， respectively. The mRNA and fluorescence expression levels of the epithelial marker E-cadherin （CDH1）， the mesenchymal marker N-cadherin （CDH2） and vimentin （VIM）， as well as ITG β1， were detected by real-time quantitative polymerase chain reaction （Real-time PCR） and immunofluorescence. Western blot was used to detect the protein expression of CDH1， CDH2 and ITG β1， and the biomechanical mechanism of ECC-JHF Ⅱ on EMT and pulmonary fibrosis was further verified by knocking down and specific activation of ITG β1. ResultsECC-JHF Ⅱ treatment improved BLM-induced pulmonary fibrosis in mice， as evidenced by significantly increased minute ventilation （MV） and 50% expiratory flow at tidal volume （EF50） （P<0.05）， and significantly increased tidal volume （TV） after treatment （P<0.01） compared with the model group. Immunohistopathological analysis revealed attenuated inflammation and alveolar injury， reduced deposition of type Ⅰ collagen （Col-Ⅰ）， and significantly decreased lung tissue stiffness （P<0.01）， thereby alleviating disease progression. The real-time PCR results showed that， compared with the model group， the ECC-JHF Ⅱ group showed increased CDH1 expression and decreased CDH2 and VIM expression in mouse lung tissues （P<0.05， P<0.01）， indicating that ECC-JHF Ⅱ inhibited the EMT process to ameliorate pulmonary fibrosis. In vitro experimental results showed that both ECC-JHF Ⅱ and matrix softening reversed the EMT phenotype switch. Specifically， compared with the hard matrix group， the cells cultured on the soft matrix exhibited significantly upregulated expression of the epithelial marker CDH1 （P<0.01） and significantly downregulated expression of the mesenchymal markers CDH2 and VIM （P<0.05）. Furthermore， ECC-JHF Ⅱ inhibited the expression of ITG β1 both in vitro and in vivo. Knockdown and specific activation of ITG β1 further confirmed that ITG β1 is involved in the regulatory process by which ECC-JHF Ⅱ inhibits EMT and ameliorates pulmonary fibrosis. ConclusionECC-JHF Ⅱ and matrix softening can block the EMT process and ameliorate pulmonary fibrosis， and the underlying mechanism may be associated with the inhibition of ITG β1-mediated mechanotransduction. This study reveals a novel mechanism of ECC-JHF Ⅱ intervention in pulmonary fibrosis from a biomechanical perspective.
