Soluble CD46 as a diagnostic marker of hepatic steatosis

Background The increasing prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) incurs substantial morbidity, mortality and healthcare costs. Detection and clinical intervention at early stages of disease improves prognosis; however, we are currently limited by a lack of reliable diagnostic tests for population screening and monitoring responses to therapy. To address this unmet need, we investigated human invariant Natural Killer T cell (iNKT) activation by fat-loaded hepatocytes, leading to the discovery that circulating soluble CD46 (sCD46) levels accurately predict hepatic steatosis. Methods sCD46 in plasma was measured using a newly developed immuno-competition assay in two independent cohorts: Prospective living liver donors (n = 156; male = 66, female = 90) and patients with liver tumours (n = 91; male = 58, female = 33). sCD46 levels were statistically evaluated as a predictor of hepatic steatosis. Findings Interleukin-4-secreting (IL-4+) iNKT cells were over-represented amongst intrahepatic lymphocytes isolated from resected human liver samples. IL-4+ iNKT cells preferentially developed in cocultures with a fat-loaded, hepatocyte-like cell line, HepaRG. This was attributed to induction of matrix metalloproteases (MMP) in fat-loaded HepaRG cells and primary human liver organoids, which led to indiscriminate cleavage of immune receptors. Loss of cell-surface CD46 resulted in unrepressed differentiation of IL-4+ iNKT cells. sCD46 levels were elevated in patients with hepatic steatosis. Discriminatory cut-off values for plasma sCD46 were found that accurately classified patients according to histological steatosis grade. Interpretation sCD46 is a reliable clinical marker of hepatic steatosis, which can be conveniently and non-invasively measured in serum and plasma samples, raising the possibility of using sCD46 levels as a diagnostic method for detecting or grading hepatic steatosis

A role for gut-associated lymphoid tissue in shaping the human B cell repertoire

PMCID: PMC3754866Rockefeller University Press grants the public the non-exclusive right to copy, distribute, or display this Work under a Creative Commons Attribution-Noncommercial-Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ and http://creativecommons.org/licenses/by-nc-sa/3.0/legalcode

Successful expansion of functional and stable regulatory T cells for immunotherapy in liver transplantation

Strategies to prevent organ transplant rejection whilst minimizing long-term immunosuppression are currently under intense investigation with regulatory T cells (Tregs) nearing clinical application. The clinical trial, ThRIL, recently commenced at King's College London, proposes to use Treg cell therapy to induce tolerance in liver transplant recipients, the success of which has the potential to revolutionize the management of these patients and enable a future of drug-free transplants. This is the first report of the manufacture of clinical grade Tregs from prospective liver transplant recipients via a CliniMACS-based GMP isolation technique and expanded using anti-CD3/CD28 beads, IL-2 and rapamycin. We report the enrichment of a pure, stable population of Tregs (>95% CD4(+)CD25(+)FOXP3(+)), reaching adequate numbers for their clinical application. Our protocol proved successful in, influencing the expansion of superior functional Tregs, as compared to freshly isolated cells, whilst also preventing their conversion to Th17 cells under pro-inflammatory conditions. We conclude with the manufacture of the final Treg product in the clinical research facility (CRF), a prerequisite for the clinical application of these cells. The data presented in this manuscript together with the much-anticipated clinical results from ThRIL, will undoubtedly inform the improved management of the liver transplant recipient

A Rapamycin-Based GMP-Compatible Process for the Isolation and Expansion of Regulatory T Cells for Clinical Trials

The concept of regulatory T cell (Treg)-based immunotherapy has enormous potential for facilitating tolerance in autoimmunity and transplantation. Clinical translation of Treg cell therapy requires production processes that satisfy the rigors of Good Manufacturing Practice (GMP) standards. In this regard, we report our findings on the implementation of a robust GMP compliant process for the ex vivo expansion of clinical grade Tregs, demonstrating the feasibility of this developed process for the manufacture of a final product for clinical application. This Treg isolation procedure ensured the selection of a pure Treg population that underwent a 300-fold expansion after 36 days of culture, while maintaining a purity of more than 75% CD4+CD25+FOXP3+ cells and a suppressive function of above 80%. Furthermore, we report the successful cryopreservation of the final product, demonstrating the maintenance of phenotype and function. The process outlined in this manuscript has been implemented in the ONE study, a multicenter phase I/IIa clinical trial in which cellular therapy is investigated in renal transplantation