Mitochondrial Dehydrogenase Activity Research Articles

The Novel Platinum Analog Dicycloplatin Provides Reliable Growth Inhibition of Urothelial Carcinoma Cells In Vitro.

Platinum-based chemotherapies are a component of standard-of-care regimens for urothelial carcinoma (UC). These nephrotoxic drugs are often dose-limiting, with cisplatin and carboplatin being the most commonly used. Dicycloplatin (DCP) has better solubility and stability, with comparable efficacy and better tolerability. Some suggest the use of DCP as primary treatment for non-muscle-invasive bladder cancer. We exposed UC cell lines to DCP in vitro to assess its efficacy. A high grade (IV) in vitro UC cell line (TCCSUP) was exposed to varying concentrations of cisplatin (0-600 μg/ml), carboplatin (0-600 μg/ml), oxaliplatin (0-4.0 μg/ml), and DCP (0-350 μg/ml). Grade II-IV cells were exposed to varying concentrations of DCP (0-350 μg/ml) to assess time- and concentration-dependent growth inhibition, and simulate intravesical treatment. Growth inhibition was determined following 24, 48, and 72 h of exposure, using a tetrazolium dye to assess mitochondrial dehydrogenase activity. DCP, cisplatin, and carboplatin effectively achieved >90% cell kill at 72 h. Concentrations of 325 μg/ml DCP, 50 μg/ml cisplatin, and 600 μg/ml carboplatin were sufficient for >90% cell-kill, with cisplatin demonstrating the highest efficacy at the lowest concentration/time intervals. Dose- and time-dependent cell kill were demonstrated at varying concentrations of DCP in grade II-IV cell lines, including cells exposed intravesically. In vitro, DCP demonstrates cell-killing efficacy in a time- and concentration-dependent manner in grade II-IV UC cell lines, showing promise for its intravenous, oral, and intravesical use for bladder UC in both primary and adjuvant/neoadjuvant settings.

Nutrient-sensitizing drug repurposing screen identifies lomerizine as a mitochondrial metabolism inhibitor of chronic myeloid leukemia.

In chronic myeloid leukemia (CML), the persistence of leukemic stem cells (LSCs) after treatment with tyrosine kinase inhibitors (TKIs), such as imatinib, can lead to disease relapse. It is known that therapy-resistant LSCs rely on oxidative phosphorylation (OXPHOS) for their survival and that targeting mitochondrial respiration sensitizes CML LSCs to imatinib treatment. However, current OXPHOS inhibitors have demonstrated limited efficacy or have shown adverse effects in clinical trials, highlighting that identification of clinically safe oxidative pathway inhibitors is warranted. We performed a high-throughput drug repurposing screen designed to identify mitochondrial metabolism inhibitors in myeloid leukemia cells. This identified lomerizine, a US Food and Drug Administration (FDA)-approved voltage-gated Ca2+ channel blocker now used for the treatment of migraines, as one of the top hits. Transcriptome analysis revealed increased expression of voltage-gated CACNA1D and receptor-activated TRPC6 Ca2+ channels in CML LSCs (CD34+CD38-) compared with normal counterparts. This correlated with increased endoplasmic reticulum (ER) mass and increased ER and mitochondrial Ca2+ content in CML stem/progenitor cells. We demonstrate that lomerizine-mediated inhibition of Ca2+ uptake leads to ER and mitochondrial Ca2+ depletion, with similar effects seen after CACNA1D and TRPC6 knockdown. Through stable isotope-assisted metabolomics and functional assays, we observe that lomerizine treatment inhibits mitochondrial isocitrate dehydrogenase activity and mitochondrial oxidative metabolism and selectively sensitizes CML LSCs to imatinib treatment. In addition, combination treatment with imatinib and lomerizine reduced CML tumor burden, targeted CML LSCs, and extended survival in xenotransplantation model of human CML, suggesting this as a potential therapeutic strategy to prevent disease relapse in patients.

Effect of fructose on anthracycline and HER-2 blocking agent mediated cardiotoxicity through NLRP3 and MyD88 mediated pathways: New roles of sweeteners in cardio-oncology.

e24020 Background: Combinatorial therapy based on anthracyclines and HER-2 blocking agents increases the risk of cardiomyopathy in women with breast cancer. Fructose is a monosaccharide associated with high risk of diabetes, non-alcoholic fatty liver disease, atherosclerosis and visceral obesity. A high fructose diet is a potential cardiometabolic risk factor and may increase cardiac risk in women with breast cancer treated with cardiotoxic drugs. In this study, we highlighted the role of sweeteners fructose in the worsening of cardiotoxicity induced by combinatorial therapy anthracycline-HER2-blocking agents. Methods: Human cardiomyocytes were pre-exposed with doxorubicin combined to trastuzumab (at 300 and 1000 nM) for 48 and 72h alone or combined to fructose at 0.25, 1 and 2.5 mM. After the incubation period, we performed the following tests: determination of cell viability, through mitochondrial dehydrogenase activity, study of lipid peroxidation (quantifying cellular Malondialdehyde and 4-hydroxynonenal), intracellular Ca2+ homeostasis. Moreover, pro-inflammatory studied were also performed (activation of NLRP3 inflammasome; expression of peroxisome proliferator-activated receptor-α; transcriptional activation of p65/NF-κB and secretion of cytokines involved in cardiotoxicity (Interleukins 1β, 8, 6, CXCL-12). Results: High fructose increases significantly cell mortality of cardiomyocytes exposed to doxorubicin and trastuzumab through the activation of NLRP-3 and Myd-88 expression. Intracellular Ca++ levels were also increased of 4 times compared to non-fructose exposed cardiomyocytes; MDA and 4-HNA levels were increased of 48,3 and 51,2 % compared to non-fructose exposed cells (p<0.001). Fructose increases IL-1, IL-6 and CXCL-12 expression in cardiomyocytes in a concentration dependent manner, compared to doxorubicin-trastuzumab group (p<0.05). Conclusions: Data of the present study indicate that fructose induces a pro-inflammatory phenotype in human cardiac cells exposed to anticancer cardiotoxic drugs, providing even more data on the harmful role of fructose as a key player of cardiotoxic phenomena.

Anti-cancer activities based on ZnII complex of potassium 5-thiophen-2-yl-[1,3,4]-oxadiazole-2-thiolate: Synthesis, crystal structure, photoluminescence study and Hirshfeld analysis

Recent advancements in developing the most suitable drug for humankind to cure cancer have paved the way to synthesize an alternative to platinum-based drugs. Herein, the synthesis of novel polymeric complex [Zn(thot)2]n (1) derived from potassium 5-thiophen-2-yl-[1,3,4]-oxadiazole-2-thiolate (Kthot) is reported. The synthesized ligand and complex were characterized by infrared, NMR, Mass and UV–vis. spectrometry. Moreover, single-crystal X-ray analysis revealed the polymeric nature of the [Zn(thot)2]n (1). Complex 1 is stabilized via various types of intermolecular interactions. The intermolecular interaction found in complex 1 was further investigated through Hirshfeld surface analysis. Emission spectra data showed that complex [Zn(thot)2]n (1) exhibits higher fluorescent intensity than Kthot. The cytotoxicity of complex 1 and Kthot was evaluated against MDA-MB-231 breast cancer cells, which was determined by measuring mitochondrial dehydrogenase activity using MTT as substrate. The result showed that they both suppressed the viability of cancer cells. Moreover, complex 1 was found to be more effective than ligand salt Kthot. The IC50 value for complex 1 was found around 37.43 μM whereas for Kthot it is 69.24 μM which suggests that complex 1 reduces the cell viability more effectively in comparison to the ligand salt.

FRUCTOSE EXACERBATES ANTHRACYCLINE AND HER–2 BLOCKING AGENT MEDIATED CARDIOTOXICITY THROUGH NLRP3 AND MYD88 MEDIATED PATHWAYS: NEW ROLES OF SWEETENERS IN CARDIO–ONCOLOGY

Abstract Background Combinatorial therapy based on anthracyclines and HER–2 blocking agents increases the risk of cardiomyopathy in women with breast cancer. Fructose is a monosaccharide associated with high risk of diabetes, non–alcoholic fatty liver disease, atherosclerosis and visceral obesity. A high fructose diet is a potential cardiometabolic risk factor and may increase cardiac risk in women with breast cancer treated with cardiotoxic drugs. Purpose In this study, we highlighted the role of sweeteners fructose in the worsening of cardiotoxicity induced by combinatorial therapy anthracycline–HER2–blocking agents. Methods Human cardiomyocytes were pre–exposed with doxorubicin combined to trastuzumab (at 300 and 1000 nM) for 48 and 72h alone or combined to fructose at 0.25, 1 and 2.5 mM. After the incubation period, we performed the following tests: determination of cell viability, through mitochondrial dehydrogenase activity, study of lipid peroxidation (quantifying cellular Malondialdehyde and 4–hydroxynonenal), intracellular Ca2+ homeostasis. Moreover, pro–inflammatory studied were also performed (activation of NLRP3 inflammasome; expression of peroxisome proliferator–activated receptor–α; transcriptional activation of p65/NF–κB and secretion of cytokines involved in cardiotoxicity (Interleukins 1β, 8, 6, CXCL–12). Results High fructose increases significantly cell mortality of cardiomyocytes exposed to doxorubicin and trastuzumab thorugh the activation of NLRP–3 and Myd–88 expression. Intracellular Ca++ levels were also increased of 4 times compared to non–fructose exposed cardiomyocytes; MDA and 4–HNA levels were increased of 48,3 and 51,2 % compared to non–fructose exposed cells (p<0.001). Fructose increases IL–1, IL–6 and CXCL–12 expression in cardiomyocytes in a concentration dependent manner, compard to doxorubicin–trastuzumab group (p<0.05). Conclusion Data of the present study indicate that fructose induces a pro–inflammatory phenotype in human cardiac cells exposed to anticancer cardiotoxic drugs, providing even more data on the harmful role of fructose as a key player of cardiotoxic phenomena.

TRPA1 up-regulation mediates oxidative stress in a pulpitis model in vitro.

Pulpitis is associated with tooth hypersensitivity and results in pulpal damage. Thermosensitive transient receptor potential (TRP) ion channels expressed in the dental pulp may be key transducers of inflammation and nociception. We aimed at investigating the expression and role of thermo-TRPs in primary human dental pulp cells (hDPCs) in normal and inflammatory conditions. Inflammatory conditions were induced in hDPC cultures by applying polyinosinic:polycytidylic acid (poly(I:C)). Gene expression and pro-inflammatory cytokine release were measured by RT-qPCR and ELISA. Functions of TRPA1 channels were investigated by monitoring changes in intracellular Ca2+ concentration. Mitochondrial superoxide production was measured using a fluorescent substrate. Cellular viability was assessed by measuring the activity of mitochondrial dehydrogenases and cytoplasmic esterases. TRPA1 activity was modified by agonists, antagonists, and gene silencing. Transcripts of TRPV1, TRPV2, TRPV4, TRPC5, and TRPA1 were highly expressed in control hDPCs, whereas TRPV3, TRPM2, and TRPM3 expressions were much lower, and TRPM8 was not detected. Poly(I:C) markedly up-regulated TRPA1 but not other thermo-TRPs. TRPA1 agonist-induced Ca2+ signals were highly potentiated in inflammatory conditions. Poly(I:C)-treated cells displayed increased Ca2+ responses to H2O2, which was abolished by TRPA1 antagonists. Inflammatory conditions induced oxidative stress, stimulated mitochondrial superoxide production, resulted in mitochondrial damage, and decreased cellular viability of hDPCs. This inflammatory cellular damage was partly prevented by the co-application of TRPA1 antagonist or TRPA1 silencing. Pharmacological blockade of TRPA1 channels may be a promising therapeutic approach to alleviate pulpitis and inflammation-associated pulpal damage.

2D TiO2 Nanosheets Decorated Via Sphere-Like BiVO4: A Promising Non-Toxic Material for Liquid Phase Photocatalysis and Bacterial Eradication.

An in-depth investigation was conducted on a promising composite material (BiVO4/TiO2), focusing on its potential toxicity, photoinduced catalytic properties, as well as its antibiofilm and antimicrobial functionalities. The preparation process involved the synthesis of 2D TiO2 using the lyophilization method, which was subsequently functionalized with sphere-like BiVO4 through wet impregnation. Finally, we developed BiVO4/TiO2 S-scheme heterojunctions which can greatly promote the separation of electron-hole pairs to achieve high photocatalytic performance. The evaluation of concentration- and time-dependent viability inhibition was performed on human lung carcinoma epithelial A549 cells. This assessment included the estimation of glutathione levels and mitochondrial dehydrogenase activity. Significantly, the BiVO4/TiO2 composite demonstrated minimal toxicity towards A549 cells. Impressively, the BiVO4/TiO2 composite exhibited notable photocatalytic performance in the degradation of rhodamine B (k=0.135 min-1) and phenol (k=0.016 min-1). In terms of photoinduced antimicrobial performance, the composite effectively inactivated both gram-negative E. coli and gram-positive E. faecalis bacteria upon 60 minutes of UV-A light exposure, resulting in a significant log 6 (log 10 CFU/mL) reduction in bacterial count. In addition, a 49 % reduction of E. faecalis biofilm was observed. These promising results can be attributed to the unique 2D morphology of TiO2 modified by sphere-like BiVO4, leading to an increased generation of (intracellular) hydroxyl radicals, which plays a crucial role in the treatments of both organic pollutants and bacteria. This research has significant potential for various applications, particularly in addressing environmental contamination and microbial infections.

In Vitro Hepatotoxicity of Routinely Used Opioids and Sedative Drugs.

A hepatocyte cell line was used to determine the hepatotoxicity of sedatives and opioids, as the hepatotoxicity of these drugs has not yet been well characterized. This might pose a threat, especially to critically ill patients, as they often receive high cumulative doses for daily analgosedation and often already have impaired liver function due to an underlying disease or complications during treatment. A well-established biosensor based on HepG2/C3A cells was used for the determination of the hepatotoxicity of commonly used sedatives and opioids in the intensive care setting (midazolam, propofol, s-ketamin, thiopental, fentanyl, remifentanil, and sufentanil). The incubation time was 2 × 3 days with clinically relevant (Cmax) and higher concentrations (C5× and C10×) of each drug in cell culture medium or human plasma. Afterward, we measured the cell count, vitality, lactate dehydrogenase (LDH), mitochondrial dehydrogenase activity, cytochrome P 450 1A2 (CYP1A2), and albumin synthesis. All tested substances reduced the viability of hepatocyte cells, but sufentanil and remifentanil showed more pronounced effects. The cell count was diminished by sufentanil in both the medium and plasma and by remifentanil only in plasma. Sufentanil and remifentanil also led to higher values of LDH in the cell culture supernatant. A reduction of mitochondrial dehydrogenase activity was seen with the use of midazolam and s-ketamine. Microalbumin synthesis was reduced in plasma after its incubation with higher concentrations of sufentanil and remifentanil. Remifentanil and s-ketamine reduced CYP1A2 activity, while propofol and thiopental increased it. Our findings suggest that none of the tested sedatives and opioids have pronounced hepatotoxicity. Sufentanil, remifentanil, and s-ketamine showed moderate hepatotoxic effects in vitro. These drugs should be given with caution to patients vulnerable to hepatotoxic drugs, e.g., patients with pre-existing liver disease or liver impairment as part of their underlying disease (e.g., hypoxic hepatitis or cholestatic liver dysfunction in sepsis). Further studies are indicated for this topic, which may use more complex cell culture models and global pharmacovigilance reports, addressing the limitation of the used cell model: HepG2/C3A cells have a lower metabolic capacity due to their low levels of CYP enzymes compared to primary hepatocytes. However, while the test model is suitable for parental substances, it is not for toxicity testing of metabolites.

Activity-based protein profiling technology reveals malate dehydrogenase as the target protein of cinnamaldehyde against Aspergillus niger

Cinnamaldehyde displays strong antifungal activity against fungi such as Aspergillus niger, but its precise molecular mechanisms of antifungal action remain inadequately understood. In this investigation, we applied chemoproteomics and bioinformatic analysis to unveil the target proteins of cinnamaldehyde in Aspergillus niger cells. Additionally, our study encompassed the examination of cinnamaldehyde's effects on cell membranes, mitochondrial malate dehydrogenase activity, and intracellular ATP levels in Aspergillus niger cells. Our findings suggest that malate dehydrogenase could potentially serve as an inhibitory target of cinnamaldehyde in Aspergillus niger cells. By disrupting the activity of malate dehydrogenase, cinnamaldehyde interferes with the mitochondrial tricarboxylic acid (TCA) cycle, leading to a significant decrease in intracellular ATP levels. Following treatment with cinnamaldehyde at a concentration of 1 MIC, the inhibition rate of MDH activity was 74.90 %, accompanied by an 84.5 % decrease in intracellular ATP content. Furthermore, cinnamaldehyde disrupts cell membrane integrity, resulting in the release of cellular contents and subsequent cell demise. This study endeavors to unveil the molecular-level antifungal mechanism of cinnamaldehyde via a chemoproteomics approach, thereby offering valuable insights for further development and utilization of cinnamaldehyde in preventing and mitigating food spoilage.

Mitochondrial Impairment and Oxidative Stress Are Essential Mechanisms Involved in the Pathogenesis of Acute Kidney Injury

Acute kidney injury (AKI) is an emergency condition that requires restrictive and appropriate clinical interventions. Identifying mechanisms of organ injury is a critical step in developing clinical interventions. Unilateral ureter obstruction (UUO) is widely used as an animal model for investigating AKI. The current study was designed to evaluate the role of mitochondrial impairment and oxidative stress in the pathogenesis of renal injury in UUO model. Mice underwent UUO surgery. Then, kidney tissue histopathological changes, plasma biomarkers of renal injury, oxidative stress, and different renal mitochondrial indices were evaluated at scheduled time intervals (3, 7, 14, and 21 days after UUO surgical procedure). Significant increase in plasma creatinine and blood urea nitrogen levels was evident in UUO mice. The UUO surgery induced severe kidney tissue histopathological alterations, including necrosis, severe tubular atrophy, and interstitial inflammation. Moreover, kidney biomarkers of oxidative stress included reactive oxygen species formation, lipid peroxidation, protein carbonylation, decreased glutathione reservoirs (GSH), and increased oxidized glutathione (GSSG) observed in UUO mice. On the other hand, significant mitochondrial depolarization, decreased mitochondrial dehydrogenases activity, mitochondrial permeabilization, and decreased adenosine triphosphate and GSH/GSSG levels were discovered in mitochondria isolated from the kidneys of UUO mice. The data obtained from the current study demonstrated a pivotal and interconnected role for oxidative stress and mitochondrial dysfunction in the pathogenesis of renal injury in UUO model. Therefore, these directions could serve as therapeutic targets in animal models or patients of acute renal failure.

Kinetic modelling of β-cell metabolism reveals control points in the insulin-regulating pyruvate cycling pathways.

Insulin, a key hormone in the regulation of glucose homoeostasis, is secreted by pancreatic β-cells in response to elevated glucose levels. Insulin is released in a biphasic manner in response to glucose metabolism in β-cells. The first phase of insulin secretion is triggered by an increase in the ATP:ADP ratio; the second phase occurs in response to both a rise in ATP:ADP and other key metabolic signals, including a rise in the NADPH:NADP+ ratio. Experimental evidence indicates that pyruvate-cycling pathways play an important role in the elevation of the NADPH:NADP+ ratio in response to glucose. The authors developed a kinetic model for the tricarboxylic acid cycle and pyruvate cycling pathways. The authors successfully validated the model against experimental observations and performed a sensitivity analysis to identify key regulatory interactions in the system. The model predicts that the dicarboxylate carrier and the pyruvate transporter are the most important regulators of pyruvate cycling and NADPH production. In contrast, the analysis showed that variation in the pyruvate carboxylase flux was compensated by a response in the activity of mitochondrial isocitrate dehydrogenase (ICDm ) resulting in minimal effect on overall pyruvate cycling flux. The model predictions suggest starting points for further experimental investigation, as well as potential drug targets for the treatment of type 2 diabetes.

Evaluation of in vitro effects of ifosfamide drug on mitochondrial functions using isolated mitochondria obtained from vital organs.

Mitochondrial toxicity has been shown to contribute to a variety of organ toxicities such as, brain, heart, kidney, and liver. Ifosfamide (IFO) as an anticancer drug, is associated with increased risk of neurotoxicity, cardiotoxicity nephrotoxicity, hepatotoxicity, and hemorrhagic cystitis. The aim of this study was to evaluate the direct effect of IFO on isolated mitochondria obtained from the rat brain, heart, kidney, and liver. Mitochondria were isolated with mechanical lysis and differential centrifugation from different organs and treated with various concentrations of IFO. Using biochemical and flowcytometry assays, we evaluated mitochondrial succinate dehydrogenase (SDH) activity, mitochondrial swelling, lipid peroxidation, reactive oxygen species (ROS) production, and mitochondrial membrane potential (MMP). Our data showed that IFO did not cause deleterious alterations in mitochondrial functions, mitochondrial swelling, lipid peroxidation ROS formation, and MMP collapse in mitochondria isolated from brain, heart, kidney, and liver. Altogether, the data showed that IFO is not directly toxic in mitochondria isolated from brain, heart, kidney, and liver. This study proved that mitochondria alone does not play the main role in the toxicity of IFO, and suggests to reduce the toxicity of this drug, other pathways resulting in the production of toxic metabolites should be considered.

Safranal'ın Servikal Kanseri C-4 I Hücre Hattı Üzerindeki Antiproliferatif Etkisinin Değerlendirilmesi

Aim: Safranal is a bioactive compound responsible for the aroma propriety of Crocus sativus. Many studies have shown the antioxidant activity of safranal besides some pharmacological properties, including its anti-inflammatory effect. Our study aimed on identifying the cytotoxic effects of safranal on the C-4 I cell line of cervical cancer. 
 Methods: The cytotoxic effect of safranal on the C-4 I cell line was determined after incubating the cells for certain durations (2 to 72 hours) and concentrations (25 to 800 µM). After incubation, cell viability and anti-proliferation effect of safranal were determined respectively by Mitochondrial Dehydrogenase and Lactate dehydrogenase enzymes activities assays. Additionally, morphological changes occurring during incubation in cells were examined with inverted and optical microscope using Giemsa staining. 
 Results: According to the results, compared to Control group, the % viability of treated cells was decreased depending on concentration and the incubation time, and safranal significantly inhibited the growth of C-4 I cells (p

Streptococcal Arginine Deiminase Inhibits T Lymphocyte Differentiation In Vitro.

Pathogenic microbes use arginine-metabolizing enzymes as an immune evasion strategy. In this study, the impact of streptococcal arginine deiminase (ADI) on the human peripheral blood T lymphocytes function in vitro was studied. The comparison of the effects of parental strain (Streptococcus pyogenes M49-16) with wild type of ArcA gene and its isogenic mutant with inactivated ArcA gene (Streptococcus pyogenes M49-16delArcA) was carried out. It was found that ADI in parental strain SDSC composition resulted in a fivefold decrease in the arginine concentration in human peripheral blood mononuclear cell (PBMC) supernatants. Only parental strain SDSCs suppressed anti-CD2/CD3/CD28-bead-stimulated mitochondrial dehydrogenase activity and caused a twofold decrease in IL-2 production in PBMC. Flow cytometry analysis revealed that ADI decreased the percentage of CM (central memory) and increased the proportion of TEMRA (terminally differentiated effector memory) of CD4+ and CD8+ T cells subsets. Enzyme activity inhibited the proliferation of all CD8+ T cell subsets as well as CM, EM (effector memory), and TEMRA CD4+ T cells. One of the prominent ADI effects was the inhibition of autophagy processes in CD8+ CM and EM as well as CD4+ CM, EM, and TEMRA T cell subsets. The data obtained confirm arginine's crucial role in controlling immune reactions and suggest that streptococcal ADI may downregulate adaptive immunity and immunological memory.

Coordination of the NF-κB signaling pathway and lymphocyte metabolism in children with autoimmune diseases

Metabolic aberrations underlie many chronic diseases, including autoimmune diseases (AUD). Immune metabolism is an area of immunological research that is actively developing and studying the processes of metabolic reprogramming in immune cells. The regulation of the nuclear factor kappa B (NF-κB) activity, which is involved in the coordination of innate and adaptive immunity, inflammatory reactions and other processes, is being actively studied. The studies on immune metabolism and regulation of NF-κB is a promising direction in searching for new therapeutic approaches in the AUD treatment. The aim of the present study was to evaluate the informative value of NF-κB and the activity of intracellular lymphocyte succinate dehydrogenase (SDH) and glycero-3-phosphate dehydrogenase (GPDH) determined in children with immune-dependent disorders. 350 children with autoimmune diseases were examined: 97 patients with IBD, 72 children with relapsing-remitting multiple sclerosis (MS), 83 pediatric patients with psoriasis vulgaris (PS), and 97 children with autoimmune hepatitis (AIH). The comparison group consisted of 100 conditionally healthy children. Activity of mitochondrial dehydrogenases, i.e., SDH and GPDH, was evaluated by immunocytochemical method. The levels of NF-κB translocation (per cent of cells with NF-κB translocation from cytoplasm to cell nucleus) was determined by flow cytometry, with visualization. Statistical evaluation and plotting were carried out using the Statistica 13.0 software. The highest activity of SDH and GPDH was detected in the population of cytotoxic T lymphocytes and T helper cells, and the lowest activity of the enzymes was registered in the population of B lymphocytes, both in children with AUD and in comparison group. In children with AUD, there was a significant decrease in SDH activity in T lymphocytes, cytotoxic T lymphocytes, B lymphocytes and NK cells against the comparison group (p 0.01). In children with PS, AIH and IBD, a decrease in SDH activity was revealed in Treg and Th17 cells. The most pronounced decrease in GPDH was characteristic of patients with AH (in T cells, cytotoxic T lymphocytes, B cells, NK cells and Tregs against the comparison group). In children with PS, the activity of GPDH was reduced only in Tregs (p 0.05). For children with multiple sclerosis, a decrease in GPDH was revealed in populations of T lymphocytes, B lymphocytes and activated T helpers (p 0.01). In the group of patients with IBD, there were no significant differences in the activity of GPDG relative to the comparison group. A significant increase in the level of NF-κB translocation in T helpers was revealed in all children with AUD relative to the comparison group. In children with AIH and PS, a significant increase in the level of NF-κB translocation was revealed in Treg, Thact and Th17 cells, in children with MS it was found in Treg cells, in patients with IBD, it was registered in Thact against the comparison group (p 0.05). An inverse correlation was found between the levels of NF-κB translocation in lymphocyte populations, and activity of mitochondrial dehydrogenases in the lymphocytes. The most significant dependencies are characteristic of NK cells and T cell populations, and these correlations are valid for all groups of AUD patientsh. In the course of in vitro experiments with a drug of metabolic action, a decreased number of cells with NF-κB translocation and an increased SDH activity was observed; the degree of SDH activation depended on the cell population type, the greatest changes were detectable in the population of T lymphocytes (by 61%), in B lymphocytes (by 30%), in NK cells (by 19%). The study of the metabolic activity of lymphocytes and the NF-κB signaling pathway allows us to assess the general mechanisms of immunopathological processes in children with autoimmune diseases of various etiologies. As based on the inverse correlation between the level of translocation of NF-κB and the activity of SDH in lymphocytes, one may consider the use of an available immunocytochemical method being an analogue for assessing activity of the NF-κB transcription factor. The studies of immune metabolic correction of immunocompetent cells are a promising direction in the AUD treatment.

Curcumin induces mitophagy by promoting mitochondrial succinate dehydrogenase activity and sensitizes human papillary thyroid carcinoma BCPAP cells to radioiodine treatment

Thyroid cancer is one of the most common endocrine malignancies. Differentiated thyroid cancer (DTC) treatment is based on the ability of thyroid follicular cells to accumulate radioactive iodide (RAI). DTC generally has a good prognosis. However, tumor dedifferentiation or defect in certain cell death mechanism occurs in a subset of DTC patients, leading to RAI resistance. Therefore, developing novel therapeutic approaches that enhance RAI sensitivity are still warranted. We found that curcumin, an active ingredient in turmeric with anti-cancer properties, rapidly accumulated in the mitochondria of thyroid cancer cells but not normal epithelial cells. Curcumin treatment triggered mitochondrial membrane depolarization, engulfment of mitochondria within autophagosomes and a robust decrease in mitochondrial mass and proteins, indicating that curcumin selectively induced mitophagy in thyroid cancer cells. In addition, curcumin-induced mitophagic cell death and its synergistic cytotoxic effect with radioiodine could be attenuated by autophagy inhibitor, 3-methyladenine (3-MA). Interestingly, the mechanism of mitophagy-inducing potential of curcumin was its unique mitochondria-targeting property, which induced a burst of SDH activity and excessive ROS production. Our data suggest that curcumin induces mitochondrial dysfunction and triggers lethal mitophagy, which synergizes with radioiodine to kill thyroid cancer cells.

Effects of OsAOX1a Deficiency on Mitochondrial Metabolism at Critical Node of Seed Viability in Rice.

Mitochondrial alternative oxidase 1a (AOX1a) plays an extremely important role in the critical node of seed viability during storage. However, the regulatory mechanism is still poorly understood. The aim of this study was to identify the regulatory mechanisms by comparing OsAOX1a-RNAi and wild-type (WT) rice seed during artificial aging treatment. Weight gain and time for the seed germination percentage decreased to 50% (P50) in OsAOX1a-RNAi rice seed, indicating possible impairment in seed development and storability. Compared to WT seeds at 100%, 90%, 80%, and 70% germination, the NADH- and succinate-dependent O2 consumption, the activity of mitochondrial malate dehydrogenase, and ATP contents all decreased in the OsAOX1a-RNAi seeds, indicating that mitochondrial status in the OsAOX1a-RNAi seeds after imbibition was weaker than in the WT seeds. In addition, the reduction in the abundance of Complex I subunits showed that the capacity of the mitochondrial electron transfer chain was significantly inhibited in the OsAOX1a-RNAi seeds at the critical node of seed viability. The results indicate that ATP production was impaired in the OsAOX1a-RNAi seeds during aging. Therefore, we conclude that mitochondrial metabolism and alternative pathways were severely inhibited in the OsAOX1a-RNAi seeds at critical node of viability, which could accelerate the collapse of seed viability. The precise regulatory mechanism of the alternative pathway at the critical node of viability needs to be further analyzed. This finding might provide the basis for developing monitoring and warning indicators when seed viability declines to the critical node during storage.

Alleviative effects of pinostrobin against cadmium-induced renal toxicity in rats by reducing oxidative stress, apoptosis, inflammation, and mitochondrial dysfunction.

Cadmium (Cd) is a highly toxic heavy metal that can be found everywhere in the environment and can have harmful effects on both human and animal health. Pinostrobin (PSB) is a bioactive natural flavonoid isolated from Boesenbergia rotunda with several pharmacological properties, such as antiinflammatory, anticancer, antioxidant, and antiviral. This investigation was intended to assess the therapeutic potential of PSB against Cd-induced kidney damage in rats. In total, 48 Sprague Dawley rats were divided into four groups: a control, a Cd (5 mg/kg), a Cd + PSB group (5 mg/kg Cd and 10 mg/kg PSB), and a PSB group (10 mg/kg) that received supplementation for 30 days. Exposure to Cd led to a decrease in the activities of catalase (CAT), glutathione reductase (GSR), superoxide dismutase (SOD), and glutathione peroxidase (GSH-PX), whereas levels of reactive oxygen species (ROS) and malondialdehyde (MDA) increased. Cd exposure also caused a substantial increase in urea, kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL), and creatinine levels. Moreover, a noticeable decline was noticed in creatinine clearance. Moreover, Cd exposure considerably increased the levels of inflammatory indices, including interleukin-1b (IL-1b), tumor necrosis factor-a (TNF-a), interleukin-6 (IL-6), nuclear factor kappa-B (NF-kB), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) activity. Cd treatment decreased the expression of the antiapoptotic markers (Bcl-2) while increasing the expression of apoptotic markers (Bax and Caspase-3). Furthermore, Cd treatment substantially reduced the TCA cycle enzyme activity, such as alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, and isocitrate dehydrogenase. Moreover, mitochondrial electron transport chain enzymes, succinatedehydrogenase, NADH dehydrogenase, cytochrome c-oxidase, and coenzyme Q-cytochrome reductase activities were also decreased following Cd exposure. PSB administration substantially reduced the mitochondrial membrane potential while inducing significant histological damage. However, PSB treatment significantly reduced Cd-mediated renal damage in rats. Thus, the present investigation discovered that PSB has ameliorative potential against Cd-induced renal dysfunction in rats.

Cardiotoxicity of chloroquine and hydroxychloroquine through mitochondrial pathway

BackgroundMedical therapies can cause cardiotoxicity. Chloroquine (QC) and hydroxychloroquine (HQC) are drugs used in the treatment of malaria and skin and rheumatic disorders. These drugs were considered to help treatment of coronavirus disease (COVID-19) in 2019. Despite the low cost and availability of QC and HQC, reports indicate that this class of drugs can cause cardiotoxicity. The mechanism of this event is not well known, but evidence shows that QC and HQC can cause cardiotoxicity by affecting mitochondria and lysosomes.MethodsTherefore, our study was designed to investigate the effects of QC and HQC on heart mitochondria. In order to achieve this aim, mitochondrial function, reactive oxygen species (ROS) level, mitochondrial membrane disruption, and cytochrome c release in heart mitochondria were evaluated. Statistical significance was determined using the one-way and two-way analysis of variance (ANOVA) followed by post hoc Tukey to evaluate mitochondrial succinate dehydrogenase (SDH) activity and cytochrome c release, and Bonferroni test to evaluate the ROS level, mitochondrial membrane potential (MMP) collapse, and mitochondrial swelling.ResultsBased on ANOVA analysis (one-way), the results of mitochondrial SDH activity showed that the IC50 concentration for CQ is 20 µM and for HCQ is 50 µM. Based on two-way ANOVA analysis, the highest effect of CQ and HCQ on the generation of ROS, collapse in the MMP, and mitochondrial swelling were observed at 40 µM and 100 µM concentrations, respectively (p < 0.05). Also, the highest effect of these two drugs has been observed in 60 min (p < 0.05). The statistical results showed that compared to CQ, HCQ is able to cause the release of cytochrome c from mitochondria in all applied concentrations (p < 0.05).ConclusionsThe results suggest that QC and HQC can cause cardiotoxicity which can lead to heart disorders through oxidative stress and disfunction of heart mitochondria.

The mitochondrion of Plasmodium falciparum is required for cellular acetyl-CoA metabolism and protein acetylation

Coenzyme A (CoA) biosynthesis is an excellent target for antimalarial intervention. While most studies have focused on the use of CoA to produce acetyl-CoA in the apicoplast and the cytosol of malaria parasites, mitochondrial acetyl-CoA production is less well understood. In the current study, we performed metabolite-labeling experiments to measure endogenous metabolites in Plasmodium falciparum lines with genetic deletions affecting mitochondrial dehydrogenase activity. Our results show that the mitochondrion is required for cellular acetyl-CoA biosynthesis and identify a synthetic lethal relationship between the two main ketoacid dehydrogenase enzymes. The activity of these enzymes is dependent on the lipoate attachment enzyme LipL2, which is essential for parasite survival solely based on its role in supporting acetyl-CoA metabolism. We also find that acetyl-CoA produced in the mitochondrion is essential for the acetylation of histones and other proteins outside of the mitochondrion. Taken together, our results demonstrate that the mitochondrion is required for cellular acetyl-CoA metabolism and protein acetylation essential for parasite survival.