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Introduction Diabetic patients are often accompanied by complications of diabetic vascular disease, which could lead to heart failure or stroke. In this work, we explored the role of miR-503/Apelin-12 in diabetic angiopathy (DA) in vitro. Methods ELISA and qPCR were applied to assess the expression of miR-503 and Apelin-12 in high glucose (HG)-treated microvascular endothelial cells (HMEC-1). The effects of miR-503 on apoptosis, inflammation and oxidative stress were assessed by flow cytometry, western blotting, qPCR, and ELISA. The interaction between miR-503 and Apelin-12 was evaluated by dual-luciferase reporter assay, qPCR and ELISA, respectively. Western blotting was performed to examine the function of miR-503/Apelin-12 on JNK and p38MAPK activation. Results MiR-503 was markedly increased and Apelin-12 was decreased in HG-treated HMEC-1 cells. MiR-503 inhibitor significantly assuaged apoptosis, inflammation and oxidative stress in HMEC-1 cells. MiR-503 could specifically bind to the 3′UTR of Apelin and inversely downregulate Apelin-12 expression. Furthermore, Apelin-12 suppressed apoptosis, inflammation and oxidative stress. Inhibition of Apelin-12 could partially reverse the decrease of p-JNK and p-p38 expression levels induced by miR-503 suppression. Conclusion In HG-induced microvascular cells injury, miR-503/Apelin-12 enhances inflammation and oxidative stress by regulating JNK and p38MAPK pathway, suggesting a potential therapeutic target for DA.

Objective Studies using the estrogen receptor alpha (ERα) knock-out (αERKO) mice have demonstrated that ERα plays a crucial role in various estrogen-mediated metabolic regulations. ERα is a ligand dependent transcription regulator and its activity is regulated by estrogenic compounds. ERα consists of two transcriptional activation domains, AF-1 and AF-2. The activities of these domains are regulated through different mechanisms; however, the specific physiological role in metabolic regulation by these domains is still unclear. Methods We utilized an ERα AF-2 mutant knock-in mouse (AF2ERKI) to evaluate the physiological functionality of ERα transactivation domains. Due to the estrogen insensitive AF-2 mutation, the phenotypes of AF2ERKI mice are seemingly identical to the global αERKO including obesity in the females. Distinct from the αERKO, the AF-1 function of AF2ERKI mice can be activated by tamoxifen (Tam). Ovariectomized (OVX) AF2ERKI and WT females were treated with Tam and fed a high-fat diet (HFD) for 10 weeks. Additionally, indirect calorimetric analysis was performed using metabolic chambers with food intake and locomotor activity recorded for Tam-treated AF2ERKI and αERKO females. Results Obesity in HFD-fed AF2ERKI females was prevented by Tam treatment; particularly, inguinal fat accumulation was strongly blocked by Tam treatment. Alterations in fat metabolism genes, however, were not found in either inguinal fat nor visceral fat to be Tam-regulated, even though fat accumulation was strongly reduced by Tam treatment. Indirect calorimetric analysis revealed that without alteration of food intake and locomotor activity Tam treatment increased energy expenditure in AF2ERKI but not αERKO females. Conclusions These results suggest that the activation of ERα AF-1 prevents fat accumulation. The prevention of obesity through AF-1 is mediated by induction of energy expenditure rather than ERα AF-1 functionality of lipid metabolism gene regulation in fat tissues., Highlights • AF-2 disrupted ERα mutant (AF2ERKI) females are obese with a pre-diabetic condition. • Activation of ERα AF-1 prevented AF2ERKI obesity. • Inguinal fat accumulation altered more than visceral fat in AF2ERKI females. • AF-1 activation improved energy expenditure without changing activity and feeding.

Previous studies have shown a significant increase in the mitochondrial generation of hydrogen peroxide (H2O2) and other peroxides in recently denervated muscle fibers. The mechanisms for generation of these peroxides and how the muscle responds to these peroxides are not fully established. The aim of this work was to determine the effect of denervation on the muscle content of proteins that may contribute to mitochondrial peroxide release and the muscle responses to this generation. Denervation of the tibialis anterior (TA) and extensor digitorum longus (EDL) muscles in mice was achieved by surgical removal of a small section of the peroneal nerve prior to its entry into the muscle. An increase in mitochondrial peroxide generation has been observed from 7 days and sustained up to 21 days following denervation in the TA muscle fibers. This increased peroxide generation was reduced by incubation of skinned fibers with inhibitors of monoamine oxidases, NADPH oxidases or phospholipase A2 enzymes and the muscle content of these enzymes together with peroxiredoxin 6 were increased following denervation. Denervated muscle also showed significant adaptations in the content of several enzymes involved in the protection of cells against oxidative damage. Morphological analyses indicated a progressive significant loss of muscle mass in the TA muscle from 7 days up to 21 days following denervation due to fiber atrophy but without fiber loss. These results support the possibility that, at least initially, the increase in peroxide production may stimulate adaptations in an attempt to protect the muscle fibers, but that these processes are insufficient and the increased peroxide generation over the longer term may activate degenerative and atrophic processes in the denervated muscle fibers., Graphical abstract Image 1, Highlights • Skeletal muscle denervation results in mitochondrial generation of peroxides in muscle fibers. • This increased peroxide generation was reduced by inhibitors of monoamine oxidases, NADPH oxidases or phospholipase A2 enzymes. • The increased peroxide generation was associated with an increase in the muscle content of peroxiredoxin 6 and phospholipase A2 that may mediate phospholipid breakdown and are also potentially involved in activation of NADPH oxidase. • The content of key antioxidant enzymes and Heat Shock Proteins involved in the protection against oxidative damage were also increased in muscle following denervation. • Data are compatible with the proposal that the denervation-induced increase in mitochondrial peroxide production initially stimulates adaptive and repair processes in muscle, but sustained peroxide generation leads to degeneration and muscle atrophy.

The serpinopathies are human pathologies caused by mutations that promote polymerisation and intracellular deposition of proteins of the serpin superfamily, leading to a poorly understood cell toxicity. The dementia FENIB is caused by polymerisation of the neuronal serpin neuroserpin (NS) within the endoplasmic reticulum (ER) of neurons. With the aim of understanding the toxicity due to intracellular accumulation of neuroserpin polymers, we have generated transgenic neural progenitor cell (NPC) cultures from mouse foetal cerebral cortex, stably expressing the control protein GFP (green fluorescent protein), or human wild type, G392E or delta NS. We have characterised these cell lines in the proliferative state and after differentiation to neurons. Our results show that G392E NS formed polymers that were mostly retained within the ER, while wild type NS was correctly secreted as a monomeric protein into the culture medium. Delta NS was absent at steady state due to its rapid degradation, but it was easily detected upon proteasomal block. Looking at their intracellular distribution, wild type NS was found in partial co-localisation with ER and Golgi markers, while G392E NS was localised within the ER only. Furthermore, polymers of NS were detected by ELISA and immunofluorescence in neurons expressing the mutant but not the wild type protein. We used control GFP and G392E NPCs differentiated to neurons to investigate which cellular pathways were modulated by intracellular polymers by performing RNA sequencing. We identified 747 genes with a significant upregulation (623) or downregulation (124) in G392E NS-expressing cells, and we focused our attention on several genes involved in the defence against oxidative stress that were up-regulated in cells expressing G392E NS (Aldh1b1, Apoe, Gpx1, Gstm1, Prdx6, Scara3, Sod2). Inhibition of intracellular anti-oxidants by specific pharmacological reagents uncovered the damaging effects of NS polymers. Our results support a role for oxidative stress in the cellular toxicity underlying the neurodegenerative dementia FENIB., Graphical abstract Image 1, Highlights • A cell model system for FENIB was generated using mouse neural progenitor cells. • Cells expressing polymerogenic neuroserpin upregulated several anti-oxidant genes. • Inhibition of the anti-oxidant defences led to apoptosis of these cells. • Oxidative stress and lipid oxidation may contribute to neurodegeneration in FENIB.