Research Highlights

Glucagon-like Peptide 1 Receptor Activation—Extending the Effects Beyond Weight Loss and Diabetes Ben Nasr M, Usuelli V, Dellepiane S, et al. Cell Metab. 2024;36:1302–1319. Over the many decades of pharmaceutical development, only a few prescription drugs have entered the cultural lexicon. Most recently, the advent of glucagon-like peptide 1 receptor agonists (GLP-1RA), including semaglutide (Ozempic), has both revolutionized treatment for type 2 diabetes and become a pharmacological cipher synonymous with weight loss. Glucose-dependent insulinotropic polypeptide and glucagon-like peptide-1 (GLP-1) are incretin hormones secreted (respectively) from the upper (K cells) and lower (L cells) gut. They are responsible for the “incretin effect,” where an oral glucose load promotes a 2- to 3-fold higher insulin secretory response compared with intravenous administration. Furthermore, GLP-1 production decreases gastric emptying and suppresses appetite and food intake. In patients with type 2 diabetes, the incretin effect is limited, which is the consequence of the substantially reduced effectiveness of glucose-dependent insulinotropic polypeptide on the endocrine pancreas and of the negligible physiological role of GLP-1 (which is also present in healthy individuals). However, the effects of GLP-1 are preserved such that pharmacological stimulation using GLP-1RA significantly reduces plasma glucose and improves glycemic control.1 The human GLP-1 receptor was cloned in 1993,2,3 and the sequence resembles secretin, parathyroid, and calcitonin receptors and is characteristic of G-protein coupled receptors. The N-terminal domain cross-links GLP-1, and this effect is dependent on disulfide bond integrity. The activation of the GLP-1R leads to an increase in intracellular cAMP, which drives insulin release. Cotransport of Na+ causes membrane depolarization and opening of Ca2+-gated channels, as well as ERK1/2 signaling via β-arrestin recruitment. The beneficial effects of GLP-1RA extend beyond their glucose-lowering capability with reductions in fatal and nonfatal cardiovascular events,4 heart failure,5 and rates of glomerular filtration decline,6 as well as resolution of NASH.7 There is also expanding literature on therapeutic utility in neurological conditions although limited evidence in human clinical trials.8 The progressive application of GLP-1RA to immune-driven disease is not unexpected given the initial serendipitous finding of improved psoriatic burden in diabetics.9 GLP-1R expression has been described in mouse T cells, particularly intestinal intraepithelial lymphocytes, where an absolute deficiency of these cells in integrin β7– mice protect from diet-induced obesity, diabetes, and cardiovascular pathology through unrestrained GLP-1 bioavailability.10 Further work11 has shown that the anti-inflammatory actions of GLP-1RA require GLP-1R-expressing intraepithelial lymphocytes to restrain local (intestinal) and systemic inflammation due to T-cell activation. This leads to the question of whether GLP-1R could modulate alloimmune responses. The article by Ben Nasr et al12 (Figure 1) indeed confirms GLP-1R expression in a subpopulation of both CD4+ and CD8+ effector memory T cells and regulatory (CD4+) T cells (as well as CD11b+, CD11c+, and natural killer cells). Proliferating T cells show increased GLP-1R surface expression, and even protein expression-negative CD3+ cells had an abundant intracellular pool. GLP-1R+CD3+ T cells were predominantly activated/exhausted CD8+ cells with an enhanced proliferative, migratory, and apoptotic phenotype, as well as reduced maximal and spare respiratory rate, which was confirmed with transcriptomic analysis and single-cell profiling. Phospho-proteomic analysis following GLP-1R stimulation in vitro confirmed senescent/exhaustion phenotype, which was BAZ2A dependent. BAZ2A is one of 2 noncatalytic protein subunits of chromatin remodeling complexes, crucially involved in binding to long noncoding and ribosomal RNA in the nucleoli.13 Although its function is not formally defined in T cells, the genomic stability promoted by BAZ2A appears to modify DNA template access and secure gene expression.FIGURE 1.: Known organ and newly discovered T-cell-based effects of GLP1 and GLP1-receptor agonists.Created with BioRender.GLP-1R signaling did not induce regulatort T cells, but perhaps did enhance stability through reductions in RORγT and Tbet expression. In an islet allotransplant model, acute rejection at day 14 was characterized by infiltrating GLP-1R+ T cells, with prolongation of graft survival with the agonist exendin-4, an effect potentiated by rapamycin and abolished by the GLP-1R antagonist exendin-9. This effect was replicated in cardiac allografts. Further evidence for GLP-1R modulation as an immune checkpoint inhibitor was derived from a colorectal cancer model treated with exendin-9 that enhanced CD3+ and CD44+CD62L+ T-cell tumor infiltration, which was not seen systemically. The presence of GLP-1R on human T cells (in addition to upregulation following stimulation) and their presence in rejecting renal, lung, and cardiac allografts provides a robust impetus for rapid translational application. Further work is required to delineate the role of GLP-1R in other immune subsets and how this receptor may work in conjunction with (or antagonistically to) other costimulatory pathways. Cool Organs Prevail—Understanding Cellular Responses to Cold Survival Zhang X, Ge L, Jin G, et al. Nat Commun. 2024;15:2859. Cold storage for organ and cellular preservation has remained the central tenet of transplantation for >50 and, in conjunction with acellular solutions, has allowed the development of trans-national programs to better serve wait-listed recipients. However, despite substantial advances in tissue typing technology, the introduction of novel induction and maintenance immunosuppression, and antiviral agents to combat opportunistic infections, there has been limited innovation in organ storage techniques. The transplant community’s inability to circumvent the fundamental nature of organ retrieval and implantation—which culminates in ischemia/reperfusion injury (IRI)—requires a shift toward understanding molecular dysregulation that accompanies both cooling and rewarming. The resurgence of machine perfusion—both hypothermic (with and without hyperoxygenation1) and normothermic2 in the kidney, as well as liver,3,4 heart,5 and lungs6—have shown excellent safety and feasibility that should support widespread application. Despite the utilitarian potential of machine perfusion to provide donor organ salvage and optimization, the impact on defined clinical variables that reflect mitigation of IRI (delayed graft function or primary nonfunction) has varied.6-8 Our understanding of how machine perfusion benefits organs remains limited. Similarly, the expanse of literature dedicated to addressing the absence of pharmacological agents that minimize or salvage IRI shows no signs of reducing in volume but has so far failed to produce a treatment that contributes effectively to standard clinical practice. It is clear there is not one answer to improving donor organ health, function, and longevity by reducing initial injury, and therefore, a multipronged approach is required. Hypothermia represents one of the most intuitive methods for reducing cellular metabolic demand to allow time-limited organ preservation, but this research area tends not to be a focus in the transplant field. Understanding the molecular response to cold will facilitate improvements in tissue storage, particularly for transplant centers where cold storage remains standard of care. A clear biological link is with hibernation, where dormancy is accompanied by profound reductions in metabolic activity. Posttranslational protein modification in the form of covalently attached small ubiquitin-related modifier (SUMO) proteins regulates diverse biological functions, including chromatin structure for RNA processing and transcription and gene expression (for DNA damage and cell cycle progression). SUMO also modifies a GTPase activator for the protein RAN that facilitates nucleocytoplasmic transport.9 SUMOylation is a feature of hibernation.10 There are some differences in mammalian hibernation based on size, but overall cytoprotective elements are preserved even outside of hibernation.11 Zhang et al12 adopted this approach to understand cellular cold (in)tolerance. Assay for transposase-accessible chromatin with sequencing, which provides information about chromatin accessibility analysis of human embryonic stem cells demonstrated upregulated FOXO1-binding motifs and concomitant nuclear translocation in response to hypothermia (at 4 °C). A FOXO1 inhibitor that sterically prevents DNA binding promoted cold-induced cell death. The authors also used a CUT&Tag assay (specific for protein-chromatin interactions) to identify novel FOXO1-binding genes not previously recognized as transcriptional targets. Rewarming of cells to 37 °C led to a rapidly changing transcriptome and overlapped with genes displaying enriched FOXO1-binding at 4 °C, which were relevant to protein phosphorylation and RHO-GTPase (required for actin cytoskeleton function, including polarity and microtubule dynamics). The authors also explored FOXO1 temperature-independent dynamics that would influence its nuclear exodus (dephosphorylation, inhibition of sirtuin) but found each condition overridden in the context of hypothermia. SUMOylated protein expression was concurrently upregulated at 4 °C, and knock-down of subunits RANBP2 or importin 7 blocked cold-induced nuclear translocation of FOXO1 (Figure 1A). Further experimental work demonstrated a cold-adaptive interactome consisting of FOXO1 cargo with SUMO-interacting motifs, RANBP2 (a SUMO ligase/nuclear pore component), and opposing transporter partners importin 7 and XPO1. This was also shown in cold-intolerant zebrafish, where suppression of foxo1a led to phenotypic abnormalities and treatment of mice with the XPO1 inhibitor KPT-330 (which stops nuclear exiting of FOXO1) reduced mortality following cold exposure. Interestingly, the transcriptome from white and brown adipose tissue, skeletal muscle, and liver showed enrichment in the immune response and cytokine signaling (Figure 1B). Different solid organs demonstrate variable responses to cold ischemic time, although in islet transplantation, the added physiological pressure of islet isolation exacerbates reduced cellular yield and peritransplant survival. Mouse islets pretreated with KPT-330 reduced cell death, for which FOXO1 nuclear entry was necessary. Longer-term storage of islets with KPT-330 preserved function glucose-stimulated insulin secretion and oxygen consumption rates (Figure 1C), and restoration of euglycemia following syngeneic transplantation. These findings provide further impetus for understanding compensatory cellular changes that support cold survival and improve donor graft quality for transplantation.FIGURE 1.: Putative role of FOXO1 in promoting cellular survival under hypothermic conditions.A, Nuclear FOXO1 expression is necessary for cell survival in hypothermic conditions. Administration of an XPO1 inhibitor promotes FOXO1 nuclear accumulation to enable murine adaptation to hypothermia (B) and prolong islet survival after prolonged cold storage (C). Created with BioRender.