Mouse skin mesenchymal stem cells (msMSCs) are dermis CD105+CD90+CD73+CD29+CD34− mesodermal precursors which, after in vitro induction, undergo chondro, adipo and osteogenesis. Extensive metabolic reconfiguration has been found to occur during differentiation, and the bioenergetic status of a cell is known to be dependent on the quality and abundance of the mitochondrial population, which may be regulated by fusion and fission. However, little is known regarding the impact of mitochondrial dynamics on the differentiation process. We addressed this knowledge gap by isolating MSCs from Swiss female mice, inducing these cells to differentiate into osteo, chondro and adipocytes and measuring changes in mass, morphology, dynamics and bioenergetics. Mitochondrial biogenesis was increased in adipogenesis, as evaluated through confocal microscopy, citrate synthase activity and mtDNA content. The early steps of adipo and osteogenesis involved mitochondrial elongation, as well as increased expression of mitochondrial fusion proteins Mfn1 and 2. Chondrogenesis involved a fragmented mitochondrial phenotype, increased expression of fission proteins Drp1, Fis1 and 2 and enhanced mitophagy. These events were accompanied by profound bioenergetic alterations during the commitment period. Moreover, knockdown of Mfn2 in adipo and osteogenesis and the overexpression of a dominant negative form of Drp1 during chondrogenesis resulted in a loss of differentiation ability. Overall, we find that mitochondrial morphology and its regulating processes of fission/fusion are modulated early on during commitment, leading to alterations in the bioenergetic profile that are important for differentiation. We thus propose a central role for mitochondrial dynamics in the maintenance/commitment of mesenchymal stem cells.
To study mitochondrial protein age dynamics, we targeted a time-sensitive fluorescent protein, MitoTimer, to the mitochondrial matrix. Mitochondrial age was revealed by the integrated portions of young (green) and old (red) MitoTimer protein. Mitochondrial protein age was dependent on turnover rates as pulsed synthesis, decreased import, or autophagic inhibition all increased the proportion of aged MitoTimer protein. Mitochondrial fusion promotes the distribution of young mitochondrial protein across the mitochondrial network as cells lacking essential fusion genes Mfn1 and Mfn2 displayed increased heterogeneity in mitochondrial protein age. Experiments in hippocampal neurons illustrate that the distribution of older and younger mitochondrial protein within the cell is determined by subcellular spatial organization and compartmentalization of mitochondria into neurites and soma. This effect was altered by overexpression of mitochondrial transport protein, RHOT1/MIRO1. Collectively our data show that distribution of young and old protein in the mitochondrial network is dependent on turnover, fusion, and transport.
PURPOSE. To examine whether diabetes-induced connexin 43 downregulation promotes retinal vascular lesions characteristic of diabetic retinopathy (DR). METHODS. Two animal models, streptozotocin-induced diabetic mice and Cx43 heterozygous knockout (Cx43(+/-)) mice, were studied to directly assess whether diabetes reduces the expression of retinal Cx43, which, in turn, contributes to retinal vascular cell loss by apoptosis. Retinal Cx43 protein levels were assessed in nondiabetic control mice, diabetic mice, and Cx43(+/-) mice by Western blot analysis, and Cx43 localization and distribution in the retinal vascular cells were studied by immunostaining of retinal trypsin digests (RTDs). In parallel, RTDs were stained with hematoxylin and periodic acid Schiff to determine pericyte loss (PL) and acellular capillaries (AC), and TUNEL assays were performed to determine retinal vascular cell apoptosis. RESULTS. Western blot analysis indicated significant reductions in retinal Cx43 protein levels in diabetic mice and Cx43(+/-) mice compared with those of nondiabetic mice. Similarly, a significant reduction in Cx43 immunostaining was observed in the retinal capillaries of diabetic mice and Cx43(+/-) mice compared with those of control mice. Both diabetic and age-matched Cx43(+/-) mice exhibited increased amount of PL, AC, and TUNEL-positive cells compared with control mice. CONCLUSIONS. Diabetes-induced inhibition of Cx43 expression contributes to vascular cell apoptosis in retinas of diabetic mice. This suggests that reduced Cx43 expression plays a critical role in the development of AC and PL associated with DR.
Mitochondrial dysfunction has been implicated in diabetic complications; however , it is unknown whether hyperglycemia affects mitochondrial morphology and metabolic capacity during development of diabetic retinopathy. We investigated high glucose (HG) effects on mitochondrial morphology, membrane potential heterogeneity, cellular oxygen consumption, extracellular acidification, cytochrome c release, and apoptosis in retinal endothelial cells. Diabetes is characterized by hyperglycemia and consequent functional failure of various target organs including the eye. In the working-age population, diabetic retinopathy is the leading cause of blindness, 1 which is triggered at least in part by hyperglycemia-induced apoptosis. While biochemical studies have implicated mitochondrial dysfunction as an underlying mechanism for inducing apoptosis, 2 the implications of mitochondrial structural changes in this process have only recently begun to be examined. In most cell types, mitochondria exist as long tubular networks that are precisely regulated by the rates of mitochondrial fusion and fission events. Disruption in this delicate balance induces altered mitochondrial membrane potential heterogeneity, 3-5 mitochondrial fragmentation, and apoptosis. 6 -8 Although oxidative stress is known to increase in diabetic retinas and trigger pro-apoptotic actions of mitochondria including the release of cytochrome c, it is currently unclear if compromised mitochondrial structure is a necessary event for high glucose (HG)-mediated apoptosis. We have shown that HG induces apoptosis in the rat retinal endothelial cells (RRECs) 9 and recent studies have indicated that HG causes mitochondrial dysfunction through oxidative damage of mitochondrial DNA and contributes to apoptosis in the human retinal endothelial cells.
Mechanistic target of rapamycin (mTOR) coordinates biosynthetic and catabolic processes in response to multiple extracellular and intracellular signals including growth factors and nutrients. This serine/threonine kinase has long been known as a critical regulator of muscle mass. The recent finding that the decision regarding its activation/inactivation takes place at the lysosome undeniably brings mTOR into the field of lysosomal storage diseases. In this study, we have examined the involvement of the mTOR pathway in the pathophysiology of a severe muscle wasting condition, Pompe disease, caused by excessive accumulation of lysosomal glycogen. Here, we report the dysregulation of mTOR signaling in the diseased muscle cells, and we focus on potential sites for therapeutic intervention. Reactivation of mTOR in the whole muscle of Pompe mice by TSC knockdown resulted in the reversal of atrophy and a striking removal of autophagic buildup. Of particular interest, we found that the aberrant mTOR signaling can be reversed by arginine. This finding can be translated into the clinic and may become a paradigm for targeted therapy in lysosomal, metabolic, and neuromuscular diseases.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.