Periodontitis contributes to COPD progression via affecting ferroptosis

Abstract

Background: Periodontitis has emerged as a potential risk factor for chronic obstructive pulmonary disease (COPD). However, the precise mechanism through which periodontitis influences the progression of COPD requires further investigation. Ferroptosis is one of the crucial pathogenesis of COPD and recent researches suggested that periodontitis was associated with ferroptosis. Nonetheless, the relationship among periodontitis, COPD and ferroptosis remains unclear. This study aimed to elucidate whether periodontitis contributes to COPD exacerbation and to assess the potential impact of ferroptosis on periodontitis affecting COPD.

Methods: The severity of COPD was assessed using Hematoxylin and eosin (H&E) staining and lung function tests. Iron assays, malondialdehyde (MDA) measurement and RT-qPCR were used to investigate the potential involvement of ferroptosis in the impact of periodontitis on COPD. Co-cultures of periodontitis associated pathogen Phophyromonas gingivalis (P. gingivalis) and lung tissue cells were used to evaluate the effect of P. gingivalis on inducing the ferroptosis of lung tissue via RT-qPCR analysis. Clinical Bronchoalveolar Lavage Fluid (BALF) samples from COPD patients were collected to further validate the role of ferroptosis in periodontal pathogen-associated COPD.

Results: Periodontitis aggravated the COPD progression and the promotion was prolonged over time. For the first time, we demonstrated that periodontitis promoted the ferroptosis-associated iron accumulation, MDA contents and gene expressions in the COPD lung with a time-dependent manner. Moreover, periodontitis-associated pathogen P. gingivalis could promote the ferroptosis-associated gene expression in single lung tissue cell suspensions. Clinical BALF sample detection further indicated that ferroptosis played essential roles in the periodontal pathogen-associated COPD.

Conclusion: Periodontitis could contribute to the exacerbation of COPD through up-regulating the ferroptosis in the lung tissue.

Keywords: COPD; Ferroptosis; MDA; P. gingivalis; Periodontitis.

Fig. 1

Periodontitis promoted COPD progression. Results of the earlier-stage of COPD (two weeks of daily CS exposure). (A): In the earlier-stage of COPD, lung lesions were observed and representative H&E images were shown. (B, C): The lung functions were compared in each group and the result were presented with the fold change of FEV0.05 value and FEV0.05/FVC value to control group. *: p < 0.05. B: Blank Control, P: periodontitis, C: COPD, CP: COPD with periodontitis

Fig. 2

Periodontitis exacerbated COPD through promoting ferroptosis in the lung. Results of the earlier-stage of COPD (two weeks of daily CS exposure). (A): In the earlier-stage of COPD, the accumulation of iron in the lung were observed through Perls staining and the representative images were shown. (B): The total iron content, ferrous iron content and lipid peroxides content (MDA content) in the lungs were calculated to compare the ferroptosis in each group quantitatively. (C): The relative mRNA expression of ferroptosis related genes in the lung tissues were analyzed by RT-qPCR, including Acsl4, Gpx4, Socs1, Ncoa4 and Ptgs2. *: p < 0.05, ns: not significant. B: Blank Control, P: periodontitis, C: COPD, CP: COPD with periodontitis

Fig. 3

The promotion effect of periodontitis on COPD was prolonged over time. Following, we further extended the daily smoke exposure time to 4 weeks to analyze whether the promotion effect of periodontitis on COPD was prolonged over time. (A): In the later-stage of COPD, H&E staining observation demonstrated more apparent COPD lesion in the lung, and periodontitis showed more serious promotion effect on the lesion. (B, C): The lung functions were compared among the four groups in the later-stage of COPD, and the result were presented with the fold change of FEV0.05 value and FEV0.05/FVC value to control group. (D, E): The lung function test results of COPD group and COPD with periodontitis group in the later-stage of COPD were further compared with those in corresponding earlier-stage COPD. *: p < 0.05, ns: not significant. B: Blank Control, P: periodontitis, C: COPD, CP: COPD with periodontitis

Fig. 4

The promotion effect of periodontitis on the ferroptosis in the lung was prolonged over time. Results of the later-stage of COPD (four weeks of daily CS exposure). (A): In the later-stage of COPD, the iron accumulation was increased in the lung and the representative Perls staining images were shown. (B): In the later-stage of COPD, the total iron content, ferrous iron content and MDA content in the lungs were shown quantitatively. (C): In the later-stage of COPD, the total iron content, ferrous iron content and MDA content in the lungs of COPD group and COPD with periodontitis group were further quantitatively compared with their corresponding earlier-stage groups. (D): In the later-stage of COPD, the relative mRNA expression of ferroptosis-related genes including Acsl4, Gpx4, Socs1, Ncoa4 and Ptgs2 were compared by RT-qPCR analysis. *: p < 0.05, ns: not significant. B: Blank Control, P: periodontitis, C: COPD, CP: COPD with periodontitis

Fig. 5

Periodontitis associated pathogen P. gingivalis could activate ferroptosis related genes in the lung single cell suspensions. Periodontitis associated pathogen P. gingivalis were cocultured with the lung single cell suspensions from health mice or ligature-induced periodontitis mice, and the expression levels of ferroptosis-related genes were analyzed by RT-qPCR. *: p < 0.05, ns: not significant. LC-H: Lung single cells from health mice, LC-P: Lung single cells from ligature-induced periodontitis mice

Fig. 6

Enhanced Iron and MDA levels and ferroptosis-related genes expressions in clinical COPD with periodontal pathogen BALF samples. (A): The total iron content, ferrous iron content and lipid peroxides content (MDA content) in the BALF samples were quantitatively calculated. (B): The relative mRNA expression of ferroptosis related genes in the BALF samples were compared by RT-qPCR analysis. *: p < 0.05, ns: not significant