Mitochondria to plasma membrane redox signaling is essential for fatty acid β-oxidation-driven insulin secretion.

We asked whether acute redox signaling from mitochondria exists concomitantly to fatty acid- (FA-) stimulated insulin secretion (FASIS) at low glucose by pancreatic β-cells. We show that FA β-oxidation produces superoxide/HO, providing: i) mitochondria-to-plasma-membrane redox signaling, closing K-channels synergically with elevated ATP (substituting NADPH-oxidase-4-mediated HO-signaling upon glucose-stimulated insulin secretion); ii) activation of redox-sensitive phospholipase iPLAγ/PNPLA8, cleaving mitochondrial FAs, enabling metabotropic GPR40 receptors to amplify insulin secretion (IS). At fasting glucose, palmitic acid stimulated IS in wt mice; palmitic, stearic, lauric, oleic, linoleic, and hexanoic acids also in perifused pancreatic islets (PIs), with suppressed 1st phases in iPLAγ/PNPLA8-knockout mice/PIs.

NADPH oxidase 4 in mouse β cells participates in inflammation on chronic nutrient overload.

Objective: By exposing mice carrying a deletion of NADPH oxidase isoform 4, NOX4, specifically in pancreatic β cells (βNOX4-/-) to nutrient excess stimulated by a high-fat diet (HFD), this study aimed to elucidate the role of β-cell redox status in the development of meta-inflammation within the diabetic phenotype.

Methods: The authors performed basic phenotyping of βNOX4-/- mice on HFD involving insulin and glycemic analyses, histochemistry of adipocytes, indirect calorimetry, and cytokine analyses. To characterize local inflammation, the study used caspase-1 activity assay, interleukin-1β immunochemistry, and real-time polymerase chain reaction during coculturing of β cells with macrophages.

Microenvironmental regulation of T-cells in pulmonary hypertension.

Introduction: In pulmonary hypertension (PH), pulmonary arterial remodeling is often accompanied by perivascular inflammation. The inflammation is characterized by the accumulation of activated macrophages and lymphocytes within the adventitial stroma, which is comprised primarily of fibroblasts. The well-known ability of fibroblasts to secrete interleukins and chemokines has previously been implicated as contributing to this tissue-specific inflammation in PH vessels.

Antioxidant Role and Cardiolipin Remodeling by Redox-Activated Mitochondrial Ca-Independent Phospholipase Aγ in the Brain.

Mitochondrial Ca-independent phospholipase Aγ (iPLAγ/PNPLA8) was previously shown to be directly activated by HO and release free fatty acids (FAs) for FA-dependent H transport mediated by the adenine nucleotide translocase (ANT) or uncoupling protein 2 (UCP2). The resulting mild mitochondrial uncoupling and consequent partial attenuation of mitochondrial superoxide production lead to an antioxidant effect. However, the antioxidant role of iPLAγ in the brain is not completely understood.

The Pancreatic β-Cell: The Perfect Redox System.

Pancreatic β-cell insulin secretion, which responds to various secretagogues and hormonal regulations, is reviewed here, emphasizing the fundamental redox signaling by NADPH oxidase 4- (NOX4-) mediated HO production for glucose-stimulated insulin secretion (GSIS). There is a logical summation that integrates both metabolic plus redox homeostasis because the ATP-sensitive K channel (K) can only be closed when both ATP and HO are elevated. Otherwise ATP would block K, while HO would activate any of the redox-sensitive nonspecific calcium channels (NSCCs), such as TRPM2.

Glucose-Induced Expression of DAPIT in Pancreatic β-Cells.

Transcript levels for selected ATP synthase membrane F-subunits-including DAPIT-in INS-1E cells were found to be sensitive to lowering glucose down from 11 mM, in which these cells are routinely cultured. Depending on conditions, the diminished mRNA levels recovered when glucose was restored to 11 mM; or were elevated during further 120 min incubations with 20-mM glucose. Asking whether DAPIT expression may be elevated by hyperglycemia in vivo, we studied mice with hyaluronic acid implants delivering glucose for up to 14 days.

Mitochondrial Superoxide Production Decreases on Glucose-Stimulated Insulin Secretion in Pancreatic β Cells Due to Decreasing Mitochondrial Matrix NADH/NAD Ratio.

Glucose-stimulated insulin secretion (GSIS) in pancreatic β cells was expected to enhance mitochondrial superoxide formation. Hence, we elucidated relevant redox equilibria. Unexpectedly, INS-1E cells at transitions from 3 (11 m; pancreatic islets from 5 m) to 25 m glucose decreased matrix superoxide release rates (MitoSOX Red monitoring validated by MitoB) and HO (mitoHyPer, subtracting mitoSypHer emission).

Glucose-Stimulated Insulin Secretion Fundamentally Requires HO Signaling by NADPH Oxidase 4.

NADPH facilitates glucose-stimulated insulin secretion (GSIS) in pancreatic islets (PIs) of β-cells through an as yet unknown mechanism. We found NADPH oxidase isoform 4 (NOX4) to be the main producer of cytosolic HO, which is essential for GSIS; an increase in ATP alone was insufficient for GSIS. The fast GSIS phase was absent from PIs from NOX4-null, β-cell-specific knockout mice (NOX4βKO) (though not from NOX2 knockout mice) and from NOX4-silenced or catalase-overexpressing INS-1E cells.

Potential of Mitochondria-Targeted Antioxidants to Prevent Oxidative Stress in Pancreatic -cells.

Pancreatic -cells are vulnerable to oxidative stress due to their low content of redox buffers, such as glutathione, but possess a rich content of thioredoxin, peroxiredoxin, and other proteins capable of redox relay, transferring redox signaling. Consequently, it may be predicted that cytosolic antioxidants could interfere with the cytosolic redox signaling and should not be recommended for any potential therapy. In contrast, mitochondrial matrix-targeted antioxidants could prevent the primary oxidative stress arising from the primary superoxide sources within the mitochondrial matrix, such as at the flavin (I) and ubiquinone (I) sites of superoxide formation within respiratory chain complex I and the outer ubiquinone site (III) of complex III.

Mitochondrial Nucleoids: Superresolution microscopy analysis.

The mitochondrion owns an autonomous genome. Double-stranded circular mitochondrial DNA (mtDNA) is organized in complexes with a packing/stabilizing transcription factor TFAM, having multiple roles, and proteins of gene expression machinery in structures called nucleoids. From hundreds to thousands nucleoids exist distributed in the matrix of mitochondrial reticulum network.

Metabolic and Proliferative State of Vascular Adventitial Fibroblasts in Pulmonary Hypertension Is Regulated Through a MicroRNA-124/PTBP1 (Polypyrimidine Tract Binding Protein 1)/Pyruvate Kinase Muscle Axis.

Background: An emerging metabolic theory of pulmonary hypertension (PH) suggests that cellular and mitochondrial metabolic dysfunction underlies the pathology of this disease. We and others have previously demonstrated the existence of hyperproliferative, apoptosis-resistant, proinflammatory adventitial fibroblasts from human and bovine hypertensive pulmonary arterial walls (PH-Fibs) that exhibit constitutive reprogramming of glycolytic and mitochondrial metabolism, accompanied by an increased ratio of glucose catabolism through glycolysis versus the tricarboxylic acid cycle. However, the mechanisms responsible for these metabolic alterations in PH-Fibs remain unknown.

Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering.

The relationship of the inner mitochondrial membrane (IMM) cristae structure and intracristal space (ICS) to oxidative phosphorylation (oxphos) is not well understood. Mitofilin (subunit Mic60) of the mitochondrial contact site and cristae organizing system (MICOS) IMM complex is attached to the outer membrane (OMM) via the sorting and assembly machinery/topogenesis of mitochondrial outer membrane β-barrel proteins (SAM/TOB) complex and controls the shape of the cristae. ATP synthase dimers determine sharp cristae edges, whereas trimeric OPA1 tightens ICS outlets.

Constitutive Reprogramming of Fibroblast Mitochondrial Metabolism in Pulmonary Hypertension.

Remodeling of the distal pulmonary artery wall is a characteristic feature of pulmonary hypertension (PH). In hypoxic PH, the most substantial pathologic changes occur in the adventitia. Here, there is marked fibroblast proliferation and profound macrophage accumulation.

Distribution of mitochondrial nucleoids upon mitochondrial network fragmentation and network reintegration in HEPG2 cells.

Mitochondrial DNA (mtDNA) is organized in nucleoids in complex with accessory proteins, proteins of mtDNA replication and gene expression machinery. A robust mtDNA genome is represented by hundreds to thousands of nucleoids in cell mitochondrion. Detailed information is lacking about the dynamics of nucleoid distribution within the mitochondrial network upon physiological and pathological events.

Disrupted ATP synthase activity and mitochondrial hyperpolarisation-dependent oxidative stress is associated with p66Shc phosphorylation in fibroblasts of NARP patients.

p66Shc is an adaptor protein involved in cell proliferation and differentiation that undergoes phosphorylation at Ser36 in response to oxidative stimuli, consequently inducing a burst of reactive oxygen species (ROS), mitochondrial disruption and apoptosis. Its role during several pathologies suggests that p66Shc mitochondrial signalling can perpetuate a primary mitochondrial defect, thus contributing to the pathophysiology of that condition. Here, we show that in the fibroblasts of neuropathy, ataxia and retinitis pigmentosa (NARP) patients, the p66Shc phosphorylation pathway is significantly induced in response to intracellular oxidative stress related to disrupted ATP synthase activity and mitochondrial membrane hyperpolarisation.