Mitochondrial dysfunction associated to permeability transition pore phenomenon in lung inflammation controls mitochondria-derived vesicles biogenesis and release in cystic fibrosis

Abstract:

Although Cystic Fibrosis (CF) is considered a rare disorder, it is the most common lethal inherited disease in Caucasians worldwide. In recent decades, significant advances have been obtained in understanding the basic CF defect and in developing new therapies, termed “CFTR modulators”, which act on mutated protein to rescue in part its defective function(1). Despite these exciting therapeutic improvements, different questions remain unanswered on CF lung pathology. Increasing evidence supports the idea that the exacerbation of pulmonary inflammation in CF lung depends also on altered crosstalk between airway and immune cells. Nonetheless, this intercellular communication is still widely obscure; therefore, innovative concepts and approaches are desperately needed to uncover this aspect of CF lung pathology, emerging as a new target for the development of more effective anti-inflammatory treatments. Currently, it is well documented that in addition to traditional endocrine and paracrine communication, the cells converse via extracellular vesicles (EVs) under physio-pathological conditions(2). EVs cargo includes plasma membrane and endosomal proteins, but EVs also contain material from other cellular compartments, including mitochondria. Recent evidence suggests that mitochondria-derived vesicles (MDVs) may be transferred and that mitochondrial content may alter the metabolic and inflammatory response of recipient cells(3–5). MDVs levels and cargo may be altered when mitochondrial dysfunction occurs, indicating that MDVs biogenesis and release are conditioned by onset of several pathological conditions where mitochondrial physiology is compromised, such as in CF. This could have important implications for CF pathological progression and treatment.
Mitochondrial dysfunction affects the subset of mitochondrial constituents altering their levels and cargo. These pathophysiological processes could be supported by the mitochondrial permeability transition pore (mtPTP) phenomenon triggered by impaired bioenergetics. We will evaluate the pharmacological and genetically mtPTP modulations to understand the MDVs bioegenesis and release in healthy and CF airway cell models. To understand the molecular mechanism of MDVs effect and formation connecting the mitochondrial homeostasis will be considered the proteomic profile of MDVs and evaluate their bioenergetic structure and mitochondrial parameters. Finally, recipient immune cells obtained from donor and CF patients will be analyzed and validated for the impact of MDVs as immune modulators and their effect on the mitochondria-related process and activities of recipient immune cells.
On balance, the mitochondrial processes linking MDVs biology and mitochondrial dynamics in CF lung disease will be considered to test new and alternative therapeutic strategies.

Risultati attesi:

Cystic fibrosis (CF) lung disease is marked by chronic inflammation and persistent infections, where mitochondria have a crucial role. Among mitochondrial processes, the biogenesis and release of mitochondria-derived vesicles (MDVs) have recently attracted attention as possible mediators of intercellular communication during inflammation. However, many aspects of MDV biology, including how they form, what cargo they carry, and how they affect recipient cells remain poorly understood. Our project aims to clarify the role of the mitochondrial permeability transition pore (mtPTP) in MDV formation and function within the context of CF airway inflammation. The work is organized into three interconnected work packages (WPs), each exploring different aspects of this problem.

WP1 investigates whether mtPTP directly contributes to MDV biogenesis and release in airway epithelial cells, and whether this mechanism is affected in CF, especially during P. aeruginosa infections. We propose that mtPTP opening promotes mitochondrial dysfunction and enhances MDV release under pro-inflammatory conditions. To test this, we will analyze MDV formation and secretion in both healthy and CF airway epithelial cells, under baseline conditions and during pathogen exposure. By using pharmacological inhibitors and genetic tools to modulate mtPTP activity, we aim to clarify its role in these processes. Our expectation is to identify mitochondrial changes that drive MDV formation and to determine whether mtPTP serves as a regulatory gate in this context.

WP2 focuses on the content and functional roles of MDVs, particularly regarding mtPTP-related proteins. We hypothesize that MDVs carry components of the mtPTP complex, including ATP synthase subunits, which may influence their biological effects and serve as potential biomarkers for CF inflammation. We plan to examine MDVs isolated under the experimental conditions from WP1, studying their protein composition and secretome. A key goal is to determine whether ATP synthase actively contributes to mtPTP assembly within MDVs and to identify critical subunits involved. The findings could reveal specific protein signatures linked to mtPTP modulation, opening avenues for new diagnostic markers or therapeutic targets aimed at controlling inflammation in CF.

WP3 addresses the functional impact of MDVs on immune cells such as neutrophils and macrophages, which play central roles in CF airway inflammation. We hypothesize that MDVs influence immune cell mitochondrial function and signaling, thereby modulating inflammatory responses. To explore this, we will assess how MDVs affect mitochondrial parameters in these immune cells. We will also investigate changes in key signaling pathways downstream of mitochondrial alterations. These studies aim to determine whether MDVs possess immunomodulatory properties and how variations in their cargo composition affect immune cell physiology and the broader inflammatory environment in CF.
In summary, this integrated research will enhance our understanding of mitochondrial behavior and intercellular communication in CF lung disease. Ultimately, it may contribute to the development of innovative biomarkers and therapies targeting mitochondrial dysfunction and inflammation in chronic airway conditions.

Dettagli progetto:

Referente scientifico: Rimessi Alessandro

Fonte di finanziamento: Bando PRIN 2022 PNRR

Data di avvio: 30/11/2023

Data di fine: 30/11/2025

Contributo MUR: 137.400 €

Partner:

Università degli Studi di FERRARA (capofila)
Università degli Studi di BOLOGNA