Biology subjects

Dickens, J. A.

Publications and source records attributed to Dickens, J. A..

3 recordsLinked to original sources

bioRxiv2025

Both terminal misfolding and polymerisation contribute to disease-relevant phenotypic changes in cell models of α1-antitrypsin deficiency-associated liver disease

Polymerisation of 1-antitrypsin within hepatocytes is considered central to the pathogenesis of 1-antitrypsin deficiency-associated liver fibrosis, most commonly in homozygotes for the Z (p.Glu342Lys) allele. Polymerisation proceeds via self-association of monomeric intermediate states. In parallel, >50% of synthesised Z 1-antitrypsin is instead recognized as terminally-misfolded and degraded. It is unclear whether this contributes to Z 1-antitrypsin deficiency-associated liver disease. We characterised the relationships between polymer formation, terminal misfolding and their cellular consequences, using label-free proteomics mass spectrometry (MS), light and electron microscopy, and cellular assays. Proteomic analyses of well-established CHO cell models of hepatocyte handling of 1-antitrypsin variants indicated that cellular responses to the Z mutation were surprisingly similar to those seen with the NullHongKong variant (NHK), which can only misfold terminally and cannot polymerise. A minor set of proteins showed increases associated with Z and not NHK 1-antitrypsin expression, consistent with a polymer-specific response, characterized by association with increased organellar organization and vesicle-mediated transport. Conversely, proteostatic and pro-fibrotic integrin-associated pathways increased with the degree of terminal misfolding of the expressed 1-antitrypsin variant. Bioenergetic pathway changes indicated concomitant switching from oxidative to glycolytic metabolism. Cell studies further correlated fibrosis-associated behaviours with terminal misfolding rather than polymerisation. Terminal misfolding, as well as polymerisation behaviour, may therefore be important for pro-fibrotic responses including metabolic reprogramming and senescence in Z 1-antitrypsin deficiency. Molecular therapies may prove most efficacious for associated liver disease if they address terminal misfolding as well as polymerisation.

cell biology↗

bioRxiv2025

Endogenous gene editing of alveolar organoids reveals that expression of pathogenic variant SFTPC-I73T disrupts endosomal function, epithelial polarity and wound healing

BackgroundIdiopathic pulmonary fibrosis is a fatal lung disease of progressive lung parenchymal scarring caused by the aberrant response of an alveolar epithelium repeatedly exposed to injury. Understanding epithelial dysfunction has been hampered by the lack of physiological alveolar type 2 (AT2) cell models and defined disease triggers. Monogenic forms of familial pulmonary fibrosis (FPF) caused by toxic gain-of-function variants provide an opportunity to investigate early pathogenic events. One such variant, surfactant protein C (SFTPC)-I73T, abnormally localises within AT2 cells and causes their dysfunction. MethodsWe used base editing of fetal lung-derived AT2 (fdAT2) organoids to create a heterozygous disease model of endogenous SFTPC-I73T expression. We also created an inducible overexpression system to interrogate temporal changes associated with SFTPC-I73T expression. We cultured fdAT2 both in 3D culture and at air-liquid interface to understand the importance of polarity cues and air exposure on disease phenotypes. ResultsIn our heterozygous endogenous expression system, we found that fdAT2 expressing SFTPC-I73T grew without a lumen and were unable to correctly polarise. SFTPC-I73T accumulated with time and caused gross enlargement of early endosomes, preventing correct apico-basal trafficking of multiple endosomally trafficked cargoes including polarity markers and cell adhesion proteins. This phenotype was exacerbated by air exposure and led to loss of epithelial monolayer integrity and abnormal wound healing after injury. ConclusionUsing endogenous gene editing for the first time in differentiated alveolar organoids, we have demonstrated that the pathogenic effects of SFTPC-I73T are mediated through endosomal dysfunction and abnormal epithelial organisation. This has important implications for AT2 function in vivo.

cell biology↗

bioRxiv2023

A novel human fetal lung-derived alveolar organoid model reveals mechanisms of surfactant protein C maturation relevant to interstitial lung disease

Alveolar type 2 (AT2) cells maintain lung health by acting as stem cells and producing pulmonary surfactant1-3. AT2 dysfunction underlies many lung diseases including interstitial lung disease (ILD), in which some inherited forms result from mislocalisation of surfactant protein C (SFTPC) variants4,5. Disease modelling and dissection of mechanisms remains challenging due to complexities in deriving and maintaining AT2 cells ex vivo. Here, we describe the development of expandable adult AT2-like organoids derived from human fetal lung which are phenotypically stable, can differentiate into AT1-like cells and are genetically manipulable. We use these organoids to test key effectors of SFTPC maturation identified in a forward genetic screen including the E3 ligase ITCH, demonstrating that their depletion phenocopies the pathological SFTPC redistribution seen for the SFTPC-I73T variant. In summary, we demonstrate the development of a novel alveolar organoid model and use it to identify effectors of SFTPC maturation necessary for AT2 health.

cell biology↗