Fetal growth is critically dependent on energy metabolism in the placenta, which drives active exchange of nutrients. Placental oxygen levels are therefore vital, and chronic hypoxia during pregnancy impairs fetal growth. Here we tested the hypothesis that placental hypoxia alters mitochondrial electron transport chain (ETS) function, and sought to identify underlying mechanisms. We cultured human placental cells under different oxygen concentrations. Mitochondrial respiration was measured, alongside levels of ETS complexes. Additionally, we studied placentas from sea-level and high-altitude pregnancies. After 4 d at 1% O2 (1.01 KPa), complex I-supported respiration was 57% and 37% lower, in trophoblast-like JEG3 cells and fibroblasts, respectively, compared with controls cultured at 21% O2 (21.24 KPa); complex IV-supported respiration was 22% and 30% lower. Correspondingly, complex I levels were 45% lower in placentas from high-altitude pregnancies than those from sea-level pregnancies. Expression of HIF-responsive microRNA-210 was increased in hypoxic fibroblasts and high-altitude placentas, whilst expression of its targets, iron-sulfur cluster scaffold (ISCU) and cytochrome c oxidase assembly protein (COX10), decreased. Moreover, protein synthesis inhibition, a feature of the high-altitude placenta, also suppressed ETS complex protein levels. Our results demonstrate that mitochondrial function is altered in hypoxic human placentas, with specific suppression of complexes I and IV compromising energy metabolism and potentially contributing to impaired fetal growth.

Preeclampsia (PE) is a dangerous complication of pregnancy, especially when it presents at <34 wk of gestation (PE < 34 wk). It is a major cause of maternal and fetal morbidity and mortality and also increases the risk of cardiometabolic diseases in later life for both mother and offspring. Placental oxidative stress induced by defective placentation sits at the epicenter of the pathophysiology. The placenta is susceptible to activation of the unfolded protein response (UPR), and we hypothesized this may affect mitochondrial function. We first examined mitochondrial respiration before investigating evidence of mitochondrial UPR (UPRmt) in placentas of PE < 34 wk patients. Reduced placental oxidative phosphorylation (OXPHOS) capacity measured in situ was observed despite no change in protein or mRNA levels of electron transport chain complexes. These results were fully recapitulated by subjecting trophoblast cells to repetitive hypoxia–reoxygenation and were associated with activation of a noncanonical UPRmt pathway; the quality-control protease CLPP, central to UPRmt signal transduction, was reduced, while the cochaperone, TID1, was increased. Transcriptional factor ATF5, which regulates expression of key UPRmt genes including HSP60 and GRP75, showed no nuclear translocation. Induction of the UPRmt with methacycline reduced OXPHOS capacity, while silencing CLPP was sufficient to reduce OXPHOS capacity, membrane potential, and promoted mitochondrial fission. CLPP was negatively regulated by the PERK-eIF2α arm of the endoplasmic reticulum UPR pathway, independent of ATF4. Similar changes in the UPRmt pathway were observed in placentas from PE < 34 wk patients. Our results identify UPRmt as a therapeutic target for restoration of placental function in early-onset preeclampsia.

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The regeneration in the peripheral nervous system is often incomplete and the treatment of severe lesions with nerve tissue loss is primarily aimed at recreating nerve continuity. Guide tubes of various types, filled with Schwann cells, stem cells, or nerve growth factors are attractive as an alternative therapy to nerve grafts. In this study, we evaluated whether skin-derived stem cells (SDSCs) can improve peripheral nerve regeneration after transplantation into nerve guides. We compared peripheral nerve regeneration in adult rats with sciatic nerve gaps of 16 mm after autologous transplantation of GFP-labeled SDSCs into two different types of guides: a synthetic guide, obtained by dip coating with a L-lactide and trimethylene carbonate (PLA-TMC) copolymer and a collagen-based guide. The sciatic function index and the recovery rates of the compound muscle action potential were significantly higher in the animals that received SDSCs transplantation, in particular, into the collagen guide, compared to the control guides filled only with PBS. For these guides the morphological and immunohistochemical analysis demonstrated an increased number of myelinated axons expressing S100 and Neurofilament 70, suggesting the presence of regenerating nerve fibers along the gap. GFP positive cells were found around regenerating nerve fibers and few of them were positive for the expression of glial markers as S-100 and glial fibrillary acidic protein. RT-PCR analysis confirmed the expression of S100 and myelin basic protein in the animals treated with the collagen guide filled with SDSCs. These data support the hypothesis that SDSCs could represent a tool for future cell therapy applications in peripheral nerve regeneration.

Purpose:We investigated the ability of the combinatorial administration of different inhibitors with activities on glioma angiogenesis, migration, and proliferation to produce a prolonged inhibition of glioma growth. Experimental Design: We combined inhibitors affecting solely tumor angiogenesis (PF-4/CTF, cyclo-VEGI) or inhibitors affecting both angiogenesis and invasion together (PEX, PF-4/DLR).Results: When administered in combination, these drugs produced a prolonged and increased inhibition of glioma growth independently from the type of inhibitor used. The combinatory administration was more effective than the administration of a single inhibitor alone, and a strong therapeutic response was reached with a significantly lower amount of protein. The strongest inhibition was observed when human PEX and PF-4/DLR, which affect both glioma angiogenesis and invasion by separate mechanisms, were combined.Conclusions: This supports the concept that prolonged glioma growth inhibition can be achieved by simultaneous delivery of molecules that target both tumor and endothelial cells and acting by separate mechanisms.

Duchenne muscular dystrophy is an inherited fatal genetic disease characterized by mutations in dystrophin gene, causing membrane fragility leading to myofiber necrosis and inflammatory cell recruitment in dystrophic muscles. The resulting environment enriched in proinflammatory cytokines, like IFN-γ and TNF-α, determines the transformation of myofiber constitutive proteasome into the immunoproteasome, a multisubunit complex involved in the activation of cell-mediate immunity. This event has a fundamental role in producing peptides for antigen presentation by MHC class I, for the immune response and also for cytokine production and T-cell differentiation. Here, we characterized for the first time the presence of T-lymphocytes activated against revertant dystrophin epitopes, in the animal model of Duchenne muscular dystrophy, the mdx mice. Moreover, we specifically blocked i-proteasome subunit LMP7, which was up-regulated in dystrophic skeletal muscles, and we demonstrated the rescue of the dystrophin expression and the amelioration of the dystrophic phenotype. The i-proteasome blocking lowered myofiber MHC class I expression and self-antigen presentation to T cells, thus reducing the specific antidystrophin T cell response, the muscular cell infiltrate, and proinflammatory cytokine production, together with muscle force recovery. We suggest that i-proteasome inhibition should be considered as new promising therapeutic approach for Duchenne muscular dystrophy pathology.

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