Murgai, M. | Search bioRxiv

bioRxiv2025

β-III tubulin identifies anti-fibrotic state of pericytes in pulmonary fibrosis

Pericytes have been implicated in pulmonary fibrosis, yet their activated cellular states and functional roles remain largely unclear. Here, we identified {beta}-III tubulin (Tuj1) as a distinctive marker for fibrosis-associated lung pericytes. Most pericytes in fibrotic regions are Tuj1-positive and interact uniquely with multiple endothelial cells, localizing near collagen-producing fibroblasts and pro-fibrotic SPP1+/arginase+ macrophages. Tuj1 expression is predominantly induced in pericytes during the fibrotic phase, and Tuj1 gene (Tubb3) knockout in mice exacerbates lung fibrosis, accompanied by an increase in the neighboring pro-fibrotic fibroblasts and macrophages, suggesting an anti-fibrotic role for Tuj1-expressing pericytes. Mechanistically, the anti-fibrotic chemokine CXCL10 is upregulated in Tuj1-expressing pericytes, whereas this upregulation is not observed in Tubb3 knockout. Moreover, CXCL10 inhibits the pro-fibrotic differentiation of macrophages induced by lung fibroblasts in culture, implying that CXCL10 may mediate the anti-fibrotic effects of Tuj1-expressing pericytes. These findings underscore the role of lung pericytes in negatively regulating fibrotic process and their potential as therapeutic targets for pulmonary fibrosis patients. SummaryIn response to fibrotic stimuli, lung pericytes upregulate {beta}-III tubulin (Tuj1) expression, adopting an anti-fibrotic phenotype. This phenotype acts as a negative immune regulator of pro-fibrotic macrophage differentiation by releasing CXCL10, thereby counteracting the progression of pulmonary fibrosis.

pathology↗

bioRxiv2024

Pericytes require physiological oxygen tension to maintain phenotypic fidelity.

Pericytes function to maintain tissue homeostasis by regulating capillary blood flow and maintaining endothelial barrier function. Pericyte dysfunction is associated with various pathologies and has recently been found to aid cancer progression. Despite having critical functions in health and disease, pericytes remain an understudied population due to a lack of model systems which accurately reflect in vivo biology. In this study we developed a protocol to isolate and culture murine lung, brain, bone, and liver pericytes, that maintains their known phenotypes and functions. We demonstrate that pericytes, being inherently plastic, benefit from controlled oxygen tension culture conditions, aiding their expansion ex vivo. Primary pericytes grown in physioxia (10% O2 for lung; 5% O2 for brain, bone, and liver) also better retain pericyte phenotypes indicated by stable expression of characteristic transcriptional and protein markers. In functional tube formation assays, pericytes were observed to significantly associate with endothelial junctions. Importantly, we identified growth conditions that limit expression of the plasticity factor Klf4 to prevent spontaneous phenotypic switching in vitro. Additionally, we were able to induce pathological pericyte phenotypic switching in response to metastatic stimuli to accurately recapitulate in vivo biology. Here, we present a robust method for studying pericyte biology in both physiology and disease.

cell biology↗