Smoking cessation only partially reverses cardiac metabolic and structural remodeling in mice.

AIMS: Active cigarette smoking is a major risk factor for chronic obstructive pulmonary disease that remains elevated after cessation. Skeletal muscle dysfunction has been well documented after smoking, but little is known about cardiac adaptations to cigarette smoking. The underlying cellular and molecular cardiac adaptations, independent of confounding lifestyle factors, and time course of reversibility by smoking cessation remain unclear. We hypothesized that smoking negatively affects cardiac metabolism and induces local inflammation in mice, which do not readily reverse upon 2-week smoking cessation. METHODS: Mice were exposed to air or cigarette smoke for 14 weeks with or without 1- or 2-week smoke cessation. We measured cardiac mitochondrial respiration by high-resolution respirometry, cardiac mitochondrial density, abundance of mitochondrial supercomplexes by electrophoresis, and capillarization, fibrosis, and macrophage infiltration by immunohistology, and performed cardiac metabolome and lipidome analysis by mass spectrometry. RESULTS: Mitochondrial protein, supercomplex content, and respiration (all p < 0.03) were lower after smoking, which were largely reversed within 2-week smoking cessation. Metabolome and lipidome analyses revealed alterations in mitochondrial metabolism, a shift from fatty acid to glucose metabolism, which did not revert to control upon smoking cessation. Capillary density was not different after smoking but increased after smoking cessation (p = 0.02). Macrophage infiltration and fibrosis (p < 0.04) were higher after smoking but did not revert to control upon smoking cessation. CONCLUSIONS: While cigarette-impaired smoking-induced cardiac mitochondrial function was reversed by smoking cessation, the remaining fibrosis and macrophage infiltration may contribute to the increased risk of cardiovascular events after smoking cessation.

Diving into cancer OXPHOS - The application of metabolic control analysis to cell and tissue research.

Knowing how the oxidative phosphorylation (OXPHOS) system in cancer cells operates differently from that of normal cells would help find compounds that specifically paralyze the energy metabolism of cancer cells. The first experiments in the study of mitochondrial respiration using the metabolic control analysis (MCA) method were done with isolated liver mitochondria in the early 80s of the last century. Subsequent studies have shown that the regulation of mitochondrial respiration by ADP in isolated mitochondria differs significantly from a model of mitochondria in situ, where the contacts with components in the cytoplasm are largely preserved. The method of selective permeabilization of the outer membrane of the cells allows the application of MCA to evaluate the contribution of different components of the OXPHOS system to its functioning while mitochondria are in a natural state. In this review, we summarize the use of MCA to study OXPHOS in cancer using permeabilized cells and tissues. In addition, we give examples of how this data fits into cancer research with a completely different approach and methodology.

Wolframin deficiency is accompanied with metabolic inflexibility in rat striated muscles.

The protein wolframin is localized in the membrane of the endoplasmic reticulum (ER), influencing Ca2+ metabolism and ER interaction with mitochondria, but the exact role of the protein remains unclear. Mutations in Wfs1 gene cause autosomal recessive disorder Wolfram syndrome (WS). The first symptom of the WS is diabetes mellitus, so accurate diagnosis of the disease as WS is often delayed. In this study we aimed to characterize the role of the Wfs1 deficiency on bioenergetics of muscles. Alterations in the bioenergetic profiles of Wfs1-exon-5-knock-out (Wfs1KO) male rats in comparison with their wild-type male littermates were investigated using high-resolution respirometry, and enzyme activity measurements. The changes were followed in oxidative (cardiac and soleus) and glycolytic (rectus femoris and gastrocnemius) muscles. There were substrate-dependent alterations in the oxygen consumption rate in Wfs1KO rat muscles. In soleus muscle, decrease in respiration rate was significant in all the followed pathways. The relatively small alterations in muscle during development of WS, such as increased mitochondrial content and/or increase in the OxPhos-related enzymatic activity could be an adaptive response to changes in the metabolic environment. The significant decrease in the OxPhos capacity is substrate dependent indicating metabolic inflexibility when multiple substrates are available.

Correction: A line-broadening free real-time 31P pure shift NMR method for phosphometabolomic analysis.

Correction for 'A line-broadening free real-time 31P pure shift NMR method for phosphometabolomic analysis' by Karl Kristjan Kaup et al., Analyst, 2021, 146, 5502-5507, DOI: 10.1039/D1AN01198G.

A line-broadening free real-time 31P pure shift NMR method for phosphometabolomic analysis.

Phosphometabolomics by 31P NMR can be challenging, since overlapping multiplets of homonuclear coupled phosphorus nuclei complicate spectral analysis. Pure shift NMR allows to simplify such spectra by collapsing multiplets into singlets, but most pure shift methods require substantially elongated measurement times or cause disturbing spectral line broadening. Herein, we combine established pure shift NMR and artefact suppression techniques to record 31P pure shift NMR spectra without penalties in measurement time or line width. Examples are demonstrated in resolution of a mixture of nucleotide triphosphates and a biological sample of 18O labelled ATP isotopomers.

Energy Metabolic Plasticity of Colorectal Cancer Cells as a Determinant of Tumor Growth and Metastasis.

Metabolic plasticity is the ability of the cell to adjust its metabolism to changes in environmental conditions. Increased metabolic plasticity is a defining characteristic of cancer cells, which gives them the advantage of survival and a higher proliferative capacity. Here we review some functional features of metabolic plasticity of colorectal cancer cells (CRC). Metabolic plasticity is characterized by changes in adenine nucleotide transport across the outer mitochondrial membrane. Voltage-dependent anion channel (VDAC) is the main protein involved in the transport of adenine nucleotides, and its regulation is impaired in CRC cells. Apparent affinity for ADP is a functional parameter that characterizes VDAC permeability and provides an integrated assessment of cell metabolic state. VDAC permeability can be adjusted via its interactions with other proteins, such as hexokinase and tubulin. Also, the redox conditions inside a cancer cell may alter VDAC function, resulting in enhanced metabolic plasticity. In addition, a cancer cell shows reprogrammed energy transfer circuits such as adenylate kinase (AK) and creatine kinase (CK) pathway. Knowledge of the mechanism of metabolic plasticity will improve our understanding of colorectal carcinogenesis.

Mitochondrial Respiration in KRAS and BRAF Mutated Colorectal Tumors and Polyps.

This study aimed to characterize the ATP-synthesis by oxidative phosphorylation in colorectal cancer (CRC) and premalignant colon polyps in relation to molecular biomarkers KRAS and BRAF. This prospective study included 48 patients. Resected colorectal polyps and postoperative CRC tissue with adjacent normal tissue (control) were collected. Patients with polyps and CRC were divided into three molecular groups: KRAS mutated, BRAF mutated and KRAS/BRAF wild-type. Mitochondrial respiration in permeabilized tissue samples was observed using high resolution respirometry. ADP-activated respiration rate (Vmax) and an apparent affinity of mitochondria to ADP, which is related to mitochondrial outer membrane (MOM) permeability, were determined. Clear differences were present between molecular groups. KRAS mutated CRC group had lower Vmax values compared to wild-type; however, the Vmax value was higher than in the control group, while MOM permeability did not change. This suggests that KRAS mutation status might be involved in acquiring oxidative phenotype. KRAS mutated polyps had higher Vmax values and elevated MOM permeability as compared to the control. BRAF mutated CRC and polyps had reduced respiration and altered MOM permeability, indicating a glycolytic phenotype. To conclude, prognostic biomarkers KRAS and BRAF are likely related to the metabolic phenotype in CRC and polyps. Assessment of the tumor mitochondrial ATP synthesis could be a potential component of patient risk stratification.

Adaptation of striated muscles to Wolframin deficiency in mice: Alterations in cellular bioenergetics.

BACKGROUND: Wolfram syndrome (WS), caused by mutations in WFS1 gene, is a multi-targeting disease affecting multiple organ systems. Wolframin is localized in the membrane of the endoplasmic reticulum (ER), influencing Ca2+ metabolism and ER interaction with mitochondria, but the exact role of the protein remains unclear. In this study we aimed to characterize alterations in energy metabolism in the cardiac and in the oxidative and glycolytic skeletal muscles in Wfs1-deficiency. METHODS: Alterations in the bioenergetic profiles in the cardiac and skeletal muscles of Wfs1-knock-out (KO) male mice and their wild type male littermates were determined using high resolution respirometry, quantitative RT-PCR, NMR spectroscopy, and immunofluorescence confocal microscopy. RESULTS: Oxygen consumption without ATP synthase activation (leak) was significantly higher in the glycolytic muscles of Wfs1 KO mice compared to wild types. ADP-stimulated respiration with glutamate and malate was reduced in the Wfs1-deficient cardiac as well as oxidative and glycolytic skeletal muscles. CONCLUSIONS: Wfs1-deficiency in both cardiac and skeletal muscles results in functional alterations of energy transport from mitochondria to ATP-ases. There was a substrate-dependent decrease in the maximal Complex I -linked respiratory capacity of the electron transport system in muscles of Wfs1 KO mice. Moreover, in cardiac and gastrocnemius white muscles a decrease in the function of one pathway were balanced by the increase in the activity of the parallel pathway. GENERAL SIGNIFICANCE: This work provides new insights to the muscle involvement at early stages of metabolic syndrome like WS as well as developing glucose intolerance.

Altered mitochondrial metabolism in the insulin-resistant heart.

Obesity-induced insulin resistance and type 2 diabetes mellitus can ultimately result in various complications, including diabetic cardiomyopathy. In this case, cardiac dysfunction is characterized by metabolic disturbances such as impaired glucose oxidation and an increased reliance on fatty acid (FA) oxidation. Mitochondrial dysfunction has often been associated with the altered metabolic function in the diabetic heart, and may result from FA-induced lipotoxicity and uncoupling of oxidative phosphorylation. In this review, we address the metabolic changes in the diabetic heart, focusing on the loss of metabolic flexibility and cardiac mitochondrial function. We consider the alterations observed in mitochondrial substrate utilization, bioenergetics and dynamics, and highlight new areas of research which may improve our understanding of the cause and effect of cardiac mitochondrial dysfunction in diabetes. Finally, we explore how lifestyle (nutrition and exercise) and pharmacological interventions can prevent and treat metabolic and mitochondrial dysfunction in diabetes.

Tubulin ßII and ßIII Isoforms as the Regulators of VDAC Channel Permeability in Health and Disease.

In recent decades, there have been several models describing the relationships between the cytoskeleton and the bioenergetic function of the cell. The main player in these models is the voltage-dependent anion channel (VDAC), located in the mitochondrial outer membrane. Most metabolites including respiratory substrates, ADP, and Pi enter mitochondria only through VDAC. At the same time, high-energy phosphates are channeled out and directed to cellular energy transfer networks. Regulation of these energy fluxes is controlled by ß-tubulin, bound to VDAC. It is also thought that ß-tubulin‒VDAC interaction modulates cellular energy metabolism in cancer, e.g., switching from oxidative phosphorylation to glycolysis. In this review we focus on the described roles of unpolymerized αß-tubulin heterodimers in regulating VDAC permeability for adenine nucleotides and cellular bioenergetics. We introduce the Mitochondrial Interactosome model and the function of the ßII-tubulin subunit in this model in muscle cells and brain synaptosomes, and also consider the role of ßIII-tubulin in cancer cells.