No abstract

Mitochondrial dysfunction causes increased oxidative stress and depletion of ATP, which are involved in the etiology of a variety of renal diseases, such as CKD, AKI, and steroidresistant nephrotic syndrome. Antioxidant therapies are being investigated, but clinical outcomes have yet to be determined. Recently, we reported that a newly synthesized indole derivative, mitochonic acid 5 (MA-5), increases cellular ATP level and survival of fibroblasts from patients with mitochondrial disease. MA-5 modulates mitochondrial ATP synthesis independently of oxidative phosphorylation and the electron transport chain. Here, we further investigated the mechanism of action for MA-5. Administration of MA-5 to an ischemia-reperfusion injury model and a cisplatin-induced nephropathy model improved renal function. In in vitro bioenergetic studies, MA-5 facilitated ATP production and reduced the level of mitochondrial reactive oxygen species (ROS) without affecting activity of mitochondrial complexes I-IV. Additional assays revealed that MA-5 targets the mitochondrial protein mitofilin at the crista junction of the inner membrane. In Hep3B cells, overexpression of mitofilin increased the basal ATP level, and treatment with MA-5 amplified this effect. In a unique mitochondrial disease model (Mitomice with mitochondrial DNA deletion that mimics typical human mitochondrial disease phenotype), MA-5 improved the reduced cardiac and renal mitochondrial respiration and seemed to prolong survival, although statistical analysis of survival times could not be conducted. These results suggest that MA-5 functions in a manner differing from that of antioxidant therapy and could be a novel therapeutic drug for the treatment of cardiac and renal diseases associated with mitochondrial dysfunction.

Broiler chickens are highly sensitive to high ambient temperatures due to their feathers, lack of skin sweat glands, and high productivity. Heat stress (HS) is a major concern for the poultry industry because it negatively affects growth as well as immune functions, which increase the potential risk of infectious disease outbreaks. Therefore, it is vital to elucidate HS's effect on the avian immune system, especially considering the global rise in average surface temperature. Our study identified a series of immunological disorders in heat-stressed broiler chickens. We exposed 22-day-old broiler chickens to a continuous HS condition (34.5 ± 0.5 • C) for 14 days and immunized them with a prototype bovine serum albumin (BSA) antigen. The plasma and lymphoid tissues (thymus, bursa of Fabricius, and spleen) were harvested at the end of the experiments to investigate the induction of BSA-specific immune responses. Our results revealed that plasma titers of immunoglobulin (Ig)Y, IgM, and IgA antibodies specific for BSA were lower than those of thermoneutral chickens immunized with BSA. Furthermore, the spleens of the heat-stressed broiler chickens displayed severe depression of Bu1 + B cells and CD3 + T cells, including CD4 + T cells and CD8 + T cells, and lacked a fully developed germinal center (GC), which is crucial for B cell proliferation. These immunological abnormalities might be associated with severe depression of CD4 − CD8 − or CD4 + CD8 + cells, which are precursors of either helper or killer T cells in the thymus and Bu1 + B cells in the bursa of Fabricius. Importantly, HS severely damaged the morphology of the thymic cortex and bursal follicles, where functional maturation of T and B cells occur. These results indicate that HS causes multiple immune abnormalities in broiler chickens by impairing the developmental process and functional maturation of T and B cells in both primary and secondary lymphoid tissues.

Heat stress is an environmental factor that causes oxidative stress. We found previously that acute heat stress stimulates the production of reactive oxygen species (ROS) in the skeletal muscle mitochondria of birds, and that this was accompanied by an increase of the mitochondrial membrane potential (ΔΨ) due to increased substrate oxidation by the electron transport chain. We also showed that avian uncoupling protein (avUCP) expression is decreased by the heat exposure. The present study clarifies whether ΔΨ is a major determinant of the overproduction of ROS due to acute heat stress, and if the decrease in avUCP expression is responsible for the elevation in ΔΨ. Control (24°C) and acute heat-stressed (34°C for 12 h) birds exhibited increased succinate-driven mitochondrial ROS production as indicated by an elevation of ΔΨ, with this increase being significantly higher in the heat-stressed group compared with the control group. In glutamate/malate-energized mitochondria, no difference in the ROS production between the groups was observed, though the mitochondrial ΔΨ was significantly higher in the heat-stressed groups compared with the control group. Furthermore, mitochondria energized with either succinate/glutamate or succinate/malate showed increased ROS production and ΔΨ in the heat-stressed group compared with mitochondria from the control group. These results suggest that succinate oxidation could play an important role in the heat stress-induced overproduction of mitochondrial ROS in skeletal muscle. In agreement with the notion of a decrease in avUCP expression in response to heat stress, proton leak, which was likely mediated by UCP (that part which is GDP-inhibited and arachidonic acid-sensitive), was reduced in the heat-exposed group. We suggest that the acute heat stress-induced overproduction of mitochondrial ROS may depend on ΔΨ, which may in turn result not only from increased substrate oxidation but also from a decrease in the mitochondrial avUCP content.

Mitochondrial dysfunction increases oxidative stress and depletes ATP in a variety of disorders. Several antioxidant therapies and drugs affecting mitochondrial biogenesis are undergoing investigation, although not all of them have demonstrated favorable effects in the clinic. We recently reported a therapeutic mitochondrial drug mitochonic acid MA-5 (Tohoku J. Exp. Med., 2015). MA-5 increased ATP, rescued mitochondrial disease fibroblasts and prolonged the life span of the disease model “Mitomouse” (JASN, 2016). To investigate the potential of MA-5 on various mitochondrial diseases, we collected 25 cases of fibroblasts from various genetic mutations and cell protective effect of MA-5 and the ATP producing mechanism was examined. 24 out of the 25 patient fibroblasts (96%) were responded to MA-5. Under oxidative stress condition, the GDF-15 was increased and this increase was significantly abrogated by MA-5. The serum GDF-15 elevated in Mitomouse was likewise reduced by MA-5. MA-5 facilitates mitochondrial ATP production and reduces ROS independent of ETC by facilitating ATP synthase oligomerization and supercomplex formation with mitofilin/Mic60. MA-5 reduced mitochondria fragmentation, restores crista shape and dynamics. MA-5 has potential as a drug for the treatment of various mitochondrial diseases. The diagnostic use of GDF-15 will be also useful in a forthcoming MA-5 clinical trial.