Rajagopalan, Pavithra

Background Eosinophilic oesophagitis (EoE) is a food allergen-induced inflammatory disorder characterised by interleukin (IL)-13-mediated oesophageal inflammation and epithelial basal cell hyperplasia (BCH). The role of mitochondria in EoE pathogenesis remains elusive. Design Prompted by single cell transcriptomics data, we interrogated the role of mitochondria in EoE pathobiology using patient biopsies, EoE-mouse models and oesophageal epithelial cells grown in monolayer and three-dimensional (3D) organoid cultures treated with EoE-relevant cytokines. 3D organoids and EoE-bearing mice were treated with omeprazole—a proton-pump inhibitor used as first-line EoE therapy. We performed CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) interference in mouse organoids to identify the key mitochondrial regulatory genes whose depletion may lead to BCH. We analysed mitochondrial membrane potential, mass and superoxide production by flow cytometry, cellular oxygen consumption by respirometry, mitochondrial structures and perturbation of cellular energy homeostasis by immunoblotting. Results Mitochondrial dysfunction appeared to be a hallmark of EoE-related BCH where mitochondrial structural damage was associated with impaired oxidative respiratory capacity, elevation of mitochondrial superoxide and decreased adenosine triphosphate (ATP) production, as corroborated by activation of the adenosine monophosphate (AMP) -activated protein kinase and suppression of mammalian target-of-rapamycin signalling. Depletion of PGC1A, the master regulator of mitochondria biogenesis, recapitulated EoE-related BCH, suggesting that mitochondrial dysfunction drives BCH. Further, omeprazole alleviated mitochondrial damage and dysfunction in EoE-related BCH modelled in mice and patient-derived organoids. Conclusion Mitochondrial dysfunction is tightly linked to perturbation of redox homeostasis in EoE-related BCH, which is promoted by IL-13 and reversible with omeprazole treatment.

Abstract Primary immunodeficiencies (PIDs) are a rare and heterogeneous group of inherited genetic disorders that are characterized by an absent or impaired immune system. In this report, we describe the use of next-generation sequencing to investigate a male infant with clinical and immunological manifestations suggestive of a PID. Whole-exome sequencing of the infant along with his parents revealed a novel nucleotide variant (cytosine to adenine substitution at nucleotide position 252) in the coding region of the interleukin 2 receptor subunit gamma ( IL2RG ) gene. The mother was found to be a carrier. These findings are consistent with a diagnosis of X-linked severe combined immunodeficiency and represent the first such reported mutation in an Indian family. This mutation leads to an asparagine to lysine substitution ( p.Asn84Lys ) located in the extracellular domain of IL2RG, which is predicted to be pathogenic. Our study demonstrates the power of next-generation sequencing in identifying potential causative mutations to enable accurate clinical diagnosis, prenatal screening, and carrier female detection in PID patients. We believe that this approach, which is not a current routine in clinical practice, will become a mainstream component of individualized medicine in the near future.