Donor Graft MicroRNAs: A Newly Identified Player in the Development of New-onset Diabetes After Liver Transplantation

Abstract

New-onset diabetes after liver transplantation (NODALT) is a frequent complication with an unfavorable outcome. We previously demonstrated a crucial link between donor graft genetics and the risk of NODALT. We selected 15 matched pairs of NODALT and non-NODALT liver recipients using propensity score matching analysis. The donor liver tissues were tested for the expression of 10 microRNAs (miRNAs) regulating human hepatic glucose homeostasis. The biological functions of potential target genes were predicted using gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Both miR-103 and miR-181a were significantly highly expressed in the NODALT group as compared to the non-NODALT group. The predicted target genes (e.g. Irs2, Pik3r1, Akt2, and Gsk3b) were involved in glucose import and the insulin signaling pathway. We also observed dysregulation of miRNAs (e.g. let-7, miR-26b, miR-145, and miR-183) in cultured human hepatocytes treated with tacrolimus or high glucose, the two independent risk factors of NODALT identified in this cohort. The hepatic miRNA profiles altered by tacrolimus or hyperglycemia were associated with insulin resistance and glucose homeostatic imbalance as revealed by enrichment analysis. The disease susceptibility miRNA expressive pattern could be imported directly from the donor and consolidated by the transplant factors.

Keywords: calcineurin inhibitor: tacrolimus; diabetes: new onset/posttransplant; donors and donation: donor evaluation; immunosuppressant; liver allograft function/dysfunction; liver transplantation/hepatology; microarray/gene array; translational research/science.

Figure 1

Donor grafts miR‐103 and miR‐181a were associated with NODALT . (A) Donor grafts miR‐103 and miR‐181a were significantly differentially expressed between 15 propensity‐matched patients with and without NODALT (both p < 0.05). (B) Donor graft miR‐103 and miR‐181a expressions were significantly correlated with fasting glucose levels at 1 week, 1 month and 3 months following liver transplantation. NODALT, new‐onset diabetes after liver transplantation; LT, liver transplantation.

Figure 2

TAC‐induced dysregulation of mi RNA s and their potential targets. (A) The expressions of let‐7a, miR‐26b, and miR‐183 were significantly increased after different concentrations of TAC treatment (5 or 20 ng/mL vs. 0 ng/mL). (B) The expressions of let‐7a, miR‐26b, and miR‐183 did not significantly change after different culture times (48 or 72 h vs. 24 h) in physiological concentrations of TAC treatment (5 ng/mL). (C) The expressions of let‐7a, miR‐26b, and miR‐183 were increased in a time‐related manner (48 or 72 h vs. 24 h) in extremely high concentrations of TAC treatment (20 ng/mL). (D) The selected glucose metabolism‐associated targets (Akt, Irs2, Gsk3, Tcf7l2, and Foxo1) of let‐7, miR‐26b, and miR‐183 were significantly decreased after TAC treatment. The protein content of p‐AKT, AKT, IRS2, TCF7L2, and FoxO1 decreased after TAC treatment. HepG2 and HUH7 cells were seeded at 2.5 × 10⁵ cells per well in six‐well plates and treated with different concentrations of TAC (0, 5, or 20 ng/mL). Protein was extracted after 72 h of culture. *p < 0.05 versus control group. TAC, tacrolimus; miRNA, microRNA.

Figure 3

High glucose led to changed mi RNA levels in a dose‐ and time‐dependent manner. (A) The expressions of miR‐145, miR‐183, and miR‐29a significantly increased after high‐glucose culture. HepG2 and HUH7 cells were seeded at 2.5 × 10⁵ cells per well in six‐well plates and treated with different concentrations of glucose (5.5, 10, or 30 mM). (B) The expressions of miR‐145 and miR‐183 elevated in a time‐dependent manner when cultured with 10 mM of glucose. RNA was extracted after 24, 48, and 72 h of culture. *p < 0.05 versus control group. miRNA, microRNA.