Single-cell RNA sequencing distinguishes dendritic cell subsets in the rat, allowing advanced characterization of the effects of FMS-like tyrosine kinase 3 ligand.

Tissue-resident dendritic cells (DCs) are essential for immunological homeostasis and hold promise for a variety of therapeutic interventions. The rare nature of tissue-resident DCs and their suboptimal description in the lab rat model has limited their characterization. To address this limitation, FMS-like tyrosine kinase 3 ligand (FLT3L) has been utilized to expand these population in vitro and in vivo for investigative or therapeutic purposes. However, conflicting reports have suggested that FLT3L can either promote immune tolerance or enhance immunogenicity, necessitating clarification of the effects of FLT3L on DC phenotype and functionality. We first paired single-cell RNA sequencing with multicolour spectral flow cytometry to provide an updated strategy for the identification of tissue-resident classical and plasmacytoid DCs in the rat model. We then administered FLT3L to Lewis rats in vivo to investigate its effect on tissue-resident DC enumeration and phenotype in the liver, spleen, and mesenteric lymph nodes. We found that FLT3L expands classical DCs (cDCs) 1 and 2 in a dose-dependent manner and that cDC1 and cDC2 in secondary lymphoid organs had altered MHC I, MHC II, CD40, CD80, CD86, and PD-L1 cell-surface expression levels following FLT3L administration. These changes were accompanied by an increase in gene expression levels of toll-like receptors 2, 4, 7, and 9 as well as inflammatory cytokines IL-6 and TNF-α. In conclusion, FLT3L administration in vivo increases cDC enumeration in the liver, spleen, and mesenteric lymph nodes accompanied by a tissue-restricted alteration in expression of antigen presentation machinery and inflammatory mediators.

Machine perfusion organ preservation: Highlights from the American Transplant Congress 2021.

The American Transplant Congress 2021 was a virtual meeting and occurred between June 4 and June 9 through an online platform. We highlighted abstracts discussing machine perfusion preservation, a hot topic that may become the gold standard of organ preservation in the future. A total of 33 abstracts on organ machine preservation (3 for heart, 4 for lungs, 18 for liver, and 8 for kidneys) were presented at the meeting. We selected 23 abstracts that showed advances including new approaches to organ preservation, promising treatments and biomarkers, cellular therapy, and novel research areas. Here, we summarize the new developments concerning machine perfusion in both experimental and clinical studies.

Modifying organs with gene therapy and gene modulation in the age of machine perfusion.

PURPOSE OF REVIEW: This review aims to highlight current advances in gene therapy methods, describing advances in CRISPR-Cas9 gene editing and RNA interference in relevance to liver transplantation, and machine perfusion. RECENT FINDINGS: In order to minimize rejection, increase the donor pool of available organs, and minimize the effects of ischemia-reperfusion injury, gene therapy and gene modification strategies are, thus, required in the context of liver transplantation. SUMMARY: Gene therapy has been used successfully in a diverse array of diseases, and, more recently, this technique has gained interest in the field of organ transplantation. Biological and logistical challenges reduce the rate of successful procedures, increasing the waiting list even more. We explore the exciting future implications of customized gene therapy in livers using machine perfusion, including its potential to create a future in which organs destined for transplant are individualized to maximize both graft and recipient longevity.

Interleukin-10 and Transforming Growth Factor-ß Cytokines Decrease Immune Activation During Normothermic Ex Vivo Machine Perfusion of the Rat Liver.

Normothermic ex vivo liver perfusion (NEVLP) is a novel system for organ preservation that may improve over static cold storage clinically and offers the chance for graft modification prior to transplantation. Although recent studies have shown the presence of inflammatory molecules during perfusion, none have yet shown the effects of NEVLP on liver-resident immune cell activation. We investigated the effects of NEVLP on liver-resident immune cell activation and assessed the ability of anti-inflammatory cytokines interleukin 10 (IL10) and transforming growth factor ß (TGF-ß) to improve organ function and reduce immune activation during perfusion. Rat livers were perfused for 4 hours at 37°C with or without the addition of 20 ng/mL of each IL10 and TGF-ß (n = 7). Naïve and cold storage (4 hours at 4°C) livers served as controls (n = 4). Following preservation, gene expression profiles were assessed through single-cell RNA sequencing; dendritic cell and macrophage activation was measured by flow cytometry; and cytokine production was assessed by enzyme-linked immunosorbent assay. NEVLP induced a global inflammatory gene expression signature, most notably in liver-resident macrophages and dendritic cells, which was accompanied by an increase in cell-surface levels of major histocompatibility complex (MHC) II, CD40, and CD86. Immune activation was partially ameliorated by IL10 and TGF-ß treatment, but no changes were observed in inflammatory cytokine production. Overall levels of liver damage and cellular apoptosis from perfusion were low, and liver function was improved with IL10 and TGF-ß treatment. This is the first study to demonstrate that liver-resident immune cells gain an activated phenotype during NEVLP on both the gene and protein level and that this activation can be reduced through therapeutic intervention with IL10 and TGF-ß.

What Is Hot and New in Basic and Translational Science in Liver Transplantation in 2023? Report of the Basic and Translational Research Committee of the International Liver Transplantation Society.

The 2023 Joint Annual Congress of the International Liver Transplantation Society, European Liver and Intestine Transplant Association, and Liver Intensive Care Group of Europe were held in Rotterdam, the Netherlands, from May 3 to 6, 2023. This year, all speakers were invited to attend the Congress in person for the first time since the COVID-19 pandemic. The congress was attended by 1159 registered delegates from 54 countries representing 5 continents, with the 10 countries comprising the bulk of the delegates. Of the 647 abstracts initially submitted, 542 were eventually presented at the meeting, coming from 38 countries (mainly North America, Europe, and Asia) and 85% of them (462 abstracts) came from only 10 countries. Fifty-three (9.8%) abstracts, originated from 17 countries, were submitted under the Basic/Translational Scientific Research category, a similar percentage as in 2022. Abstracts presented at the meeting were classified as (1) ischemia and reperfusion injury, (2) machine perfusion, (3) bioengineering and liver regeneration, (4) transplant oncology, (5) novel biomarkers in liver transplantation, (6) liver immunology (rejection and tolerance), and (7) artificial intelligence and machine learning. Finally, we evaluated the number of abstracts commented in the Basic and Translational Research Committee-International Liver Transplantation Society annual reports over the past 5 y that resulted in publications in peer-reviewed journals to measure their scientific impact in the field of liver transplantation.