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Systemic sclerosis (SSc) with interstitial lung disease (SSc-ILD) lacks curative pharmacological treatments, thus necessitating effective animal models for candidate drug discovery. Existing bleomycin (BLM)-induced SSc-ILD mouse models feature spatially limited pulmonary fibrosis, spontaneously resolving after 28 days. Here, we present an alternative BLM administration approach in female C57BL/6 mice, combining oropharyngeal aspiration (OA) and subcutaneous mini-pump delivery (pump) of BLM to induce a sustained and more persistent fibrosis, while retaining stable skin fibrosis. A dose-finding study was performed with BLM administered as 10 lg (OA) + 80 mg/kg (pump) (10 + 80), 10 + 100, and 15 + 100. Forty-two days after OA, micro-computed tomography (micro-CT) imaging and histomorphometric analyses showed that the 10 + 100 and 15 + 100 treatments induced significant alterations in lung micro-CT-derived readouts, Ashcroft score, and more severe fibrosis grades compared with saline controls. In addition, a marked reduction in hypodermal thickness was observed in the 15 + 100 group. A timecourse characterization of the BLM 15 + 100 treatment at days 28, 35, and 42, including longitudinal micro-CT imaging, revealed progressing alterations in lung parameters. Lung histology highlighted a sustained fibrosis accompanied by a reduction in hypodermis thickness throughout the explored time-window, with a time-dependent increase in fibrotic biomarkers detected by immunofluorescence analysis. BLM-induced alterations were partly mitigated by Nintedanib treatment. Our optimized BLM delivery approach leads to extensive and persistent lung fibrotic lesions coupled with cutaneous fibrotic alterations: it thus represents a significant advance compared with current preclinical models of BLM-induced SSc-ILD. NEW & NOTEWORTHY This study introduces an innovative approach to enhance the overall performance of the mouse bleomycin (BLM)-induced model for systemic sclerosis with interstitial lung disease (SSc-ILD). By combining oropharyngeal aspiration and subcutaneous mini-pump delivery of BLM, our improved model leads to sustained lung fibrosis and stable skin fibrosis in female C57BL/6 mice. The optimized 15 + 100 treatment results in extensive and persistent lung fibrotic lesions and thus represents a significant improvement over existing preclinical models of BLM-induced SSc-ILD. [ABSTRACT FROM AUTHOR]

Systemic sclerosis (SSc) is an autoimmune disease characterized by an excessive production and accumulation of collagen in the skin and internal organs often associated with interstitial lung disease (ILD). Its pathogenetic mechanisms are unknown and the lack of animal models mimicking the features of the human disease is creating a gap between the selection of anti-fibrotic drug candidates and effective therapies. In this work, we intended to pharmacologically validate a SSc-ILD model based on 1 week infusion of bleomycin (BLM) by osmotic minipumps in C57/BL6 mice, since it will serve as a tool for secondary drug screening. Nintedanib (NINT) has been used as a reference compound to investigate antifibrotic activity either for lung or skin fibrosis. Longitudinal Micro-CT analysis highlighted a significant slowdown in lung fibrosis progression after NINT treatment, which was confirmed by histology. However, no significant effect was observed on lung hydroxyproline content, inflammatory infiltrate and skin lipoatrophy. The modest pharmacological effect reported here could reflect the clinical outcome, highlighting the reliability of this model to better profile potential clinical drug candidates. The integrative approach presented herein, which combines longitudinal assessments with endpoint analyses, could be harnessed in drug discovery to generate more reliable, reproducible and robust readouts. [ABSTRACT FROM AUTHOR]