Mitochondrial dysfunction and treatment strategies

The mitochondria are essential for cellular energy production and are involved in many processes in the cells. The mitochondria contain their own DNA (mtDNA) that is vital for oxidative phosphorylation since it encodes enzymes of the respiratory chain. Mutations in the mtDNA and alterations in the mtDNA copy number are attributed to various human disorders including cancer. Mitochondrial DNA depletion syndromes (MDS) are a heterogeneous group of disorders characterized by severe depletion of the mtDNA. MDS predominantly manifests in high energy demanding tissues such as the skeletal muscle, brain and liver. Mutations in the genes that are responsible for providing precursors for the mtDNA synthesis such as thymidine kinase 2 (TK2) and deoxyguanosine kinase (dGK) are known to cause MDS. In an attempt to rescue the mtDNA depletion caused by thymidine kinase 2 (Tk2) deficiency in mice, the deoxyribonucleoside kinase from Drosophila melanogaster (Dm-dNK) was expressed in the Tk2 deficient mice (Dm-dNK+/-Tk2-/-). The Dm-dNK+/- expression was able to rescue the Tk2-/- mice and prolong their life span from 3 weeks to up to 20 months. The Dm-dNK expression driven by the CMV promoter was observed in all tissues with highest expression in skeletal muscle and lower expression in heart, liver and adipose tissues. Dm-dNK+/-Tk2-/- mice maintained normal mtDNA levels in the skeletal muscle and liver throughout the observation time of 20 months. The Dm-dNK expression resulted in highly elevated dNTP pools with dTTP pools being >100 times higher than in the wild type mice. However, the large increase in the dTTP pool did not cause mutations in the nuclear or the mitochondrial DNA. A significant reduction in total body fat (both subcutaneous and visceral fat) was observed only in the Dm-dNK+/-Tk2-/- mice compared to wild type mice, which indicates an altered fat metabolism in these mice mediated through residual Tk2 deficiency. To elucidate effective treatment strategies for TK2 deficiency, a novel mouse model with liver specific expression of Dm-dNK driven by the albumin promoter was generated. Two founder mice with high Dm-dNK expression and activity in the liver was selected for further characterization. These mice will be used to study whether Dm-dNK expression in a single tissue would be able to rescue the sever phenotype caused by Tk2 deficiency in mice. The mitochondrial dicarboxylate carrier, SLC25A10, is involved in the transport of dicarboxylates such as malate and succinate across the mitochondrial inner membrane. To understand the role of the SLC25A10 carrier in regulating cancer cell growth, metabolism and transformation, a knockdown of SLC25A10 in a lung adenocarcinoma cell line (A549) was established and characterized. The growth properties of SLC25A10 knockdown cells changed to a less malignant phenotype, with increased dependency on glutamine and altered NADPH production. An increase in expression of glutamate dehydrogenase and decrease in expression of lactate dehydrogenase indicated a metabolic shift from glycolysis to oxidative phosphorylation in the SLC25A10 knockdown cells. The study demonstrates the importance of SLC25A10 in and regulation of redox homeostasi

Expressing the multifunctional nucleoside kinase of Drosophila melanogaster in a mouse model : a strategy to reverse the depletion of mtDNA caused by nucleoside kinase deficiency

This study was initiated to investigate a possible strategy to alter an enzyme deficiency in a mouse model. The enzyme investigated is a multifunctional nucleoside kinase from Drosophila melanogaster (Dm-dNK). This enzyme has special features in that it has higher enzymatic activity than any other known nucleoside kinases and still has similar substrate specificity as the human nucleoside kinases. The deficiency where the Dm-dNK transgenic mice model will be used is a TK2 deficient model with severe phenotype caused by mitochondrial DNA depletion. The Dm-dNK transgenic mice model will be used as a way to rescue the TK2 deficient mice. The results from the present study show that Dm-dNK expression in mice results in a substantial increase of thymidine phosphorylation in several investigated tissues. The mice were otherwise normal as judged by life span, weight and behavior. The mitochondrial DNA was also detected at normal levels. In conclusion, the Dm-dNK mouse model is promising as a way to rescue the severe phenotype of the TK2 deficient mice

Genetic variants in post myocardial infarction patients presenting with electrical storm of unstable ventricular tachycardia

Electrical storm (ES) is a life threatening clinical situation. Though a few clinical pointers exist, the occurrence of ES in a patient with remote myocardial infarction (MI) is generally unpredictable. Genetic markers for this entity have not been studied. In the present study, we carried out genetic screening in patients with remote myocardial infarction presenting with ES by next generation sequencing and identified 25 rare variants in 19 genes predominantly in RYR2, SCN5A, KCNJ11, KCNE1 and KCNH2, CACNA1B, CACNA1C, CACNA1D and desmosomal genes - DSP and DSG2 that could potentially be implicated in electrical storm. These genes have been previously reported to be associated with inherited syndromes of Sudden Cardiac Death. The present study suggests that the genetic architecture in patients with remote MI and ES of unstable ventricular tachycardia may be similar to that of Ion channelopathies. Identification of these variants may identify post MI patients who are predisposed to develop electrical storm and help in risk stratification

Additional file 2: of Inhibition of glutamate oxaloacetate transaminase 1 in cancer cell lines results in altered metabolism with increased dependency of glucose

Figure S2. Rescue of GOT1 down-regulated and GOT1-null cells by oxaloacetate. Relative cell viabilities after 8 h (a) and 24 h (b) in wild type 143B cells and 8 h (c) and 24 h (d) in wild type A549 cells. Relative cell viabilities after 8 h (e) and 24 h (f) in GOT1 siRNA knock-down A549 cells. Rescue of GOT1-null 143B cells with OAA at different concentrations upon glucose deprivation (g). Mean ± s.d. from 3 independent experiments. One-way ANOVA test was performed. *** p < 0.001; ** p < 0.01;* p < 0.05. NS: not significant. (TIF 230 kb

Additional file 3: of Inhibition of glutamate oxaloacetate transaminase 1 in cancer cell lines results in altered metabolism with increased dependency of glucose

Figure S3. Partial prevention of ischemic-like-cell-death morphological changes by NAD+. Bars indicate 25 Οm. (TIF 1353 kb

Cell-type-resolved quantitative proteomics map of interferon response against SARS-CoV-2

The commonly used laboratory cell lines are the first line of experimental models to study the pathogenicity and performing antiviral assays for emerging viruses. Here, we assessed the tropism and cytopathogenicity of the first Swedish isolate of SARS-CoV-2 in six different human cell lines, compared their growth characteristics, and performed quantitative proteomics for the susceptible cell lines. Overall, Calu-3, Caco2, Huh7, and 293FT cell lines showed a high-to-moderate level of susceptibility to SARS-CoV-2. In Caco2 cells, the virus can achieve high titers in the absence of any prominent cytopathic effect. The protein abundance profile during SARS-CoV-2 infection revealed cell-type-specific regulation of cellular pathways. Type-I interferon signaling was identified as the common dysregulated cellular response in Caco2, Calu-3, and Huh7 cells. Together, our data show cell-type specific variability for cytopathogenicity, susceptibility, and cellular response to SARS-CoV-2 and provide important clues to guide future studies

Multi-omics personalized network analyses highlight progressive disruption of central metabolism associated with COVID-19 severity

The clinical outcome and disease severity in coronavirus disease-2019 (COVID-19) are heterogeneous, and the progression or fatality of the disease cannot be explained by a single factor like age or comorbidities. In this study, we used system-wide network-based system biology analysis using whole blood RNA sequencing, immune-phenotyping by flow cytometry, plasma metabolomics, and single cell-type metabolomics of the monocytes to identify the potential determinants of COVID-19 severity at the personalized and group level. Digital cell quantification and immune-phenotyping of the mononuclear phagocytes indicated a substantial role in coordinating the immune cells that mediate the COVID-19 severity. Stratum-specific and personalized genome-scale metabolic modeling indicated monocarboxylate transporter family genes (e.g., SLC16A6), nucleoside transporter genes (e.g., SLC29A1), and metabolites such as α-ketoglutarate, succinate, malate, and butyrate, could play a crucial role in COVID-19 severity. Metabolic perturbations targeting the central metabolic pathway (TCA-cycle) can be an alternate treatment strategy in severe COVID-19

Metabolic perturbation associated with COVID-19 disease severity and SARS-CoV-2 replication

Viruses hijack host metabolic pathways for their replicative advantage. In this study, using patient-derived multiomics data and in vitro infection assays, we aimed to understand the role of key metabolic pathways that can regulate severe acute respiratory syndrome coronavirus-2 reproduction and their association with disease severity. We used multiomics platforms (targeted and untargeted proteomics and untargeted metabolomics) on patient samples and cell-line models along with immune phenotyping of metabolite transporters in patient blood cells to understand viral-induced metabolic modulations. We also modulated key metabolic pathways that were identified using multiomics data to regulate the viral reproduction in vitro. Coronavirus disease 2019 disease severity was characterized by increased plasma glucose and mannose levels. Immune phenotyping identified altered expression patterns of carbohydrate transporter, glucose transporter 1, in CD8+ T cells, intermediate and nonclassical monocytes, and amino acid transporter, xCT, in classical, intermediate, and nonclassical monocytes. In in vitro lung epithelial cell (Calu-3) infection model, we found that glycolysis and glutaminolysis are essential for virus replication, and blocking these metabolic pathways caused significant reduction in virus production. Taken together, we therefore hypothesized that severe acute respiratory syndrome coronavirus-2 utilizes and rewires pathways governing central carbon metabolism leading to the efflux of toxic metabolites and associated with disease severity. Thus, the host metabolic perturbation could be an attractive strategy to limit the viral replication and disease severity