Circumventing the Crabtree effect in cell culture: A systematic review.

Metabolic reprogramming and mitochondrial dysfunction are central elements in a broad variety of physiological and pathological processes. While cell culture established itself as a versatile technique for the elaboration of physiology and disease, studying metabolism using standard cell culture protocols is profoundly interfered by the Crabtree effect. This phenomenon refers to the adaptation of cultured cells to a glycolytic phenotype, away from oxidative phosphorylation in glucose-containing medium, and questions the applicability of cell culture in certain fields of research. In this systematic review we aim to provide a comprehensive overview and critical appraisal of strategies reported to circumvent the Crabtree effect.

Results of an explorative clinical evaluation suggest immediate and persistent post-reperfusion metabolic paralysis drives kidney ischemia reperfusion injury.

Delayed graft function is the manifestation of ischemia reperfusion injury in the context of kidney transplantation. While hundreds of interventions successfully reduce ischemia reperfusion injury in experimental models, all clinical interventions have failed. This explorative clinical evaluation examined possible metabolic origins of clinical ischemia reperfusion injury combining data from 18 pre- and post-reperfusion tissue biopsies with 36 sequential arteriovenous blood samplings over the graft in three study groups. These groups included living and deceased donor grafts with and without delayed graft function. Group allocation was based on clinical outcome. Magic angle NMR was used for tissue analysis and mass spectrometry-based platforms were used for plasma analysis. All kidneys were functional at one-year. Integration of metabolomic data identified a discriminatory profile to recognize future delayed graft function. This profile was characterized by post-reperfusion ATP/GTP catabolism (significantly impaired phosphocreatine recovery and significant persistent (hypo)xanthine production) and significant ongoing tissue damage. Failing high-energy phosphate recovery occurred despite activated glycolysis, fatty-acid oxidation, glutaminolysis and autophagia, and related to a defect at the level of the oxoglutarate dehydrogenase complex in the Krebs cycle. Clinical delayed graft function due to ischemia reperfusion injury associated with a post-reperfusion metabolic collapse. Thus, efforts to quench delayed graft function due to ischemia reperfusion injury should focus on conserving metabolic competence, either by preserving the integrity of the Krebs cycle and/or by recruiting metabolic salvage pathways.

Leukocyte Dynamics during the Evolution of Human Coronary Atherosclerosis: Conclusions from a Sevenfold, Chromogen-Based, Immunohistochemical Evaluation.

Atherosclerosis is a complex process with strong inflammatory component. We developed a straightforward sevenfold staining protocol for simultaneous assessment of dominant leukocyte classes, vascularization, and expression of the putative foam cell maker CD36. The method was applied on human coronaries covering the full spectrum of atherosclerotic disease. Results confirm the progressive association of macrophages and T cells with the process and a global presence of mast cells. B cells are exclusively present in adventitial follicles that accompany the process plaque destabilization (thin cap and ruptured lesions) and are otherwise absent. Neutrophils are only present as part of the hemorrhage that accompanies plaque rupture. This study does not classify CD36 as a key factor in foam cell formation. Observed macrophage accumulation in the neointima of stabilized fibrous calcified plaques is consistent with a process of neoatherosclerosis. This study on human coronaries shows a progressive association of macrophage and T-cell abundance with plaque progression. Follicle-like structures are transiently present during the process of plaque destabilization. Plaque healing is accompanied by cessation of the inflammatory response but followed by a new cycle of atherosclerosis.