Metformin, phenformin, and galegine inhibit complex IV activity and reduce glycerol-derived gluconeogenesis

MacDonald M.J., Ansari I.H., Longacre M.J., Stoker S.W.

Mitochondrial glycerol phosphate dehydrogenase (mGPD) is the rate-limiting enzyme of the glycerol phosphate redox shuttle. It was recently claimed that metformin, a first-line drug used for the treatment of type 2 diabetes, inhibits liver mGPD 30-50%, suppressing gluconeogenesis through a redox mechanism. Various factors cast doubt on this idea. Total-body knockout of mGPD in mice has adverse effects in several tissues where the mGPD level is high but has little or no effect in liver, where the mGPD level is the lowest of 10 tissues. Metformin has beneficial effects in humans in tissues with high levels of mGPD, such as pancreatic β-cells, where the mGPD level is much higher than that in liver. Insulin secretion in mGPD knockout mouse β-cells is normal because, like liver, β-cells possess the malate aspartate redox shuttle whose redox action is redundant to the glycerol phosphate shuttle. For these and other reasons, we used four different enzyme assays to reassess whether metformin inhibited mGPD. Metformin did not inhibit mGPD in homogenates or mitochondria from insulin cells or liver cells. If metformin actually inhibited mGPD, adverse effects in tissues where the level of mGPD is much higher than that in the liver could prevent the use of metformin as a diabetes medicine.

LaMoia T.E., Shulman G.I.

Abstract Metformin is a first-line therapy for the treatment of type 2 diabetes, due to its robust glucose-lowering effects, well-established safety profile, and relatively low cost. While metformin has been shown to have pleotropic effects on glucose metabolism, there is a general consensus that the major glucose-lowering effect in patients with type 2 diabetes is mostly mediated through inhibition of hepatic gluconeogenesis. However, despite decades of research, the mechanism by which metformin inhibits this process is still highly debated. A key reason for these discrepant effects is likely due to the inconsistency in dosage of metformin across studies. Widely studied mechanisms of action, such as complex I inhibition leading to AMPK activation, have only been observed in the context of supra-pharmacological (>1 mM) metformin concentrations, which do not occur in the clinical setting. Thus, these mechanisms have been challenged in recent years and new mechanisms have been proposed. Based on the observation that metformin alters cellular redox balance, a redox-dependent mechanism of action has been described by several groups. Recent studies have shown that clinically relevant (50-100 μM) concentrations of metformin inhibit hepatic gluconeogenesis in a substrate-selective manner both in vitro and in vivo, supporting a redox-dependent mechanism of metformin action. Here, we review the current literature regarding metformin’s cellular and molecular mechanisms of action.

Xie J., Ye J., Cai Z., Luo Y., Zhu X., Deng Y., Feng Y., Liang Y., Liu R., Han Z., Liang Y., Zheng Y., Mo R., Zhuo Y., Wu Y., et. al.

Abstract Metformin is an oral drug widely used for the treatment of type 2 diabetes mellitus. Numerous studies have demonstrated the value of metformin in cancer treatment. However, for metformin to elicit effects on cancer often requires a high dosage, and any underlying mechanism for how to improve its inhibitory effects remains unknown. Here, we found that low mRNA expression of glycerol-3-phosphate dehydrogenase 1 (GPD1) may predict a poor response to metformin treatment in 15 cancer cell lines. In vitro and in vivo, metformin treatment alone significantly suppressed cancer cell proliferation, a phenotype enhanced by GPD1 overexpression. Total cellular glycerol-3-phosphate concentration was significantly increased by the combination of GPD1 overexpression and metformin treatment, which suppressed cancer growth via inhibition of mitochondrial function. Eventually, increased reactive oxygen species and mitochondrial structural damage was observed in GPD1-overexpressing cell lines treated with metformin, which may contribute to cell death. In summary, this study demonstrates that GPD1 overexpression enhances the anticancer activity of metformin and that patients with increased GPD1 expression in tumor cells may respond better to metformin therapy. Significance: GPD1 overexpression enhances the anticancer effect of metformin through synergistic inhibition of mitochondrial function, thereby providing new insight into metformin-mediated cancer therapy.

Tavallaie M., Voshtani R., Deng X., Qiao Y., Jiang F., Collman J.P., Fu L.

Significance The efforts toward minimizing oxidative stress and maintaining mitochondrial energy metabolism for promoted healthy aging are at the forefront of current research on aging. We have characterized a treatment that, by reversible inhibition of mitochondrial respiration in mice through cytochrome c oxidase (CcO), markedly improves mitochondrial bioenergetics and delays the lead health concerns associated with aging in mice. We demonstrated that chronic moderate inhibition of CcO reduces ATP synthesis, promotes mitochondrial biogenesis and mitophagy, subsequently decreases ROS production and mitochondrial decay, and rectifies vital cellular energy metabolism regulators, thus, effectively refining energy homeostasis and curbing obesity and glucose irregularities linked to aging.

Di Magno L., Manni S., Di Pastena F., Coni S., Macone A., Cairoli S., Sambucci M., Infante P., Moretti M., Petroni M., Nicoletti C., Capalbo C., De Smaele E., Di Marcotullio L., Giannini G., et. al.

Summary The antidiabetic drug phenformin displays potent anticancer activity in different tumors, but its mechanism of action remains elusive. Using Shh medulloblastoma as model, we show here that at clinically relevant concentrations, phenformin elicits a significant therapeutic effect through a redox-dependent but complex I-independent mechanism. Phenformin inhibits mitochondrial glycerophosphate dehydrogenase (mGPD), a component of the glycerophosphate shuttle, and causes elevations of intracellular NADH content. Inhibition of mGPD mimics phenformin action and promotes an association between corepressor CtBP2 and Gli1, thereby inhibiting Hh transcriptional output and tumor growth. Because ablation of CtBP2 abrogates the therapeutic effect of phenformin in mice, these data illustrate a biguanide-mediated redox/corepressor interplay, which may represent a relevant target for tumor therapy.

Wang Y., Kwon H., Su X., Wondisford F.E.

Objective Fasting results in major metabolic changes including a switch from glycogenolysis to gluconeogenesis to maintain glucose homeostasis. However, the relationship between the length of fasting and the relative contribution of gluconeogenic substrates remains unclear. We investigated the relative contribution of glycogen, lactate, and glycerol in glucose production of male C57BL/6 J-albino mice after 6, 12, and 18 h of fasting. Methods We used non-perturbative infusions of 13C3 lactate, 13C3 glycerol, and 13C6 glucose combined with liquid chromatography mass spectrometry and metabolic flux analysis to study the contribution of substrates in gluconeogenesis (GNG). Results During infusion studies, both lactate and glycerol significantly label about 60% and 30–50% glucose carbon, respectively, but glucose labels much more lactate (∼90%) than glycerol carbon (∼10%). Our analyses indicate that lactate, but not glycerol is largely recycled during all fasting periods such that lactate is the largest direct contributor to GNG via the Cori cycle but a minor source of new glucose carbon (overall contribution). In contrast, glycerol is not only a significant direct contributor to GNG but also the largest overall contributor to GNG regardless of fasting length. Prolonged fasting decreases both the whole body turnover rate of glucose and lactate but increases that of glycerol, indicating that the usage of glycerol in GNG become more significant with longer fasting. Conclusion Collectively, these findings suggest that glycerol is the dominant overall contributor of net glucose carbon in GNG during both short and prolonged fasting.

McCreight L.J., Mari A., Coppin L., Jackson N., Umpleby A.M., Pearson E.R.

Wang Y., An H., Liu T., Qin C., Sesaki H., Guo S., Radovick S., Hussain M., Maheshwari A., Wondisford F.E., O’Rourke B., He L.

Impaired mitochondrial respiratory activity contributes to the development of insulin resistance in type 2 diabetes. Metformin, a first-line antidiabetic drug, functions mainly by improving patients' hyperglycemia and insulin resistance. However, its mechanism of action is still not well understood. We show here that pharmacological metformin concentration increases mitochondrial respiration, membrane potential, and ATP levels in hepatocytes and a clinically relevant metformin dose increases liver mitochondrial density and complex 1 activity along with improved hyperglycemia in high-fat- diet (HFD)-fed mice. Metformin, functioning through 5' AMP-activated protein kinase (AMPK), promotes mitochondrial fission to improve mitochondrial respiration and restore the mitochondrial life cycle. Furthermore, HFD-fed-mice with liver-specific knockout of AMPKα1/2 subunits exhibit higher blood glucose levels when treated with metformin. Our results demonstrate that activation of AMPK by metformin improves mitochondrial respiration and hyperglycemia in obesity. We also found that supra-pharmacological metformin concentrations reduce adenine nucleotides, resulting in the halt of mitochondrial respiration. These findings suggest a mechanism for metformin's anti-tumor effects.

Alshawi A., Agius L.

The mechanisms by which metformin (dimethylbiguanide) inhibits hepatic gluconeogenesis at concentrations relevant for type 2 diabetes therapy remain debated. Two proposed mechanisms are 1) inhibition of mitochondrial Complex 1 with consequent compromised ATP and AMP homeostasis or 2) inhibition of mitochondrial glycerophosphate dehydrogenase (mGPDH) and thereby attenuated transfer of reducing equivalents from the cytoplasm to mitochondria, resulting in a raised lactate/pyruvate ratio and redox-dependent inhibition of gluconeogenesis from reduced but not oxidized substrates. Here, we show that metformin has a biphasic effect on the mitochondrial NADH/NAD redox state in mouse hepatocytes. A low cell dose of metformin (therapeutic equivalent:

Petersen M.C., Shulman G.I.

The 1921 discovery of insulin was a Big Bang from which a vast and expanding universe of research into insulin action and resistance has issued. In the intervening century, some discoveries have matured, coalescing into solid and fertile ground for clinical application; others remain incompletely investigated and scientifically controversial. Here, we attempt to synthesize this work to guide further mechanistic investigation and to inform the development of novel therapies for type 2 diabetes (T2D). The rational development of such therapies necessitates detailed knowledge of one of the key pathophysiological processes involved in T2D: insulin resistance. Understanding insulin resistance, in turn, requires knowledge of normal insulin action. In this review, both the physiology of insulin action and the pathophysiology of insulin resistance are described, focusing on three key insulin target tissues: skeletal muscle, liver, and white adipose tissue. We aim to develop an integrated physiological perspective, placing the intricate signaling effectors that carry out the cell-autonomous response to insulin in the context of the tissue-specific functions that generate the coordinated organismal response. First, in section II, the effectors and effects of direct, cell-autonomous insulin action in muscle, liver, and white adipose tissue are reviewed, beginning at the insulin receptor and working downstream. Section III considers the critical and underappreciated role of tissue crosstalk in whole body insulin action, especially the essential interaction between adipose lipolysis and hepatic gluconeogenesis. The pathophysiology of insulin resistance is then described in section IV. Special attention is given to which signaling pathways and functions become insulin resistant in the setting of chronic overnutrition, and an alternative explanation for the phenomenon of ‟selective hepatic insulin resistanceˮ is presented. Sections V, VI, and VII critically examine the evidence for and against several putative mediators of insulin resistance. Section V reviews work linking the bioactive lipids diacylglycerol, ceramide, and acylcarnitine to insulin resistance; section VI considers the impact of nutrient stresses in the endoplasmic reticulum and mitochondria on insulin resistance; and section VII discusses non-cell autonomous factors proposed to induce insulin resistance, including inflammatory mediators, branched-chain amino acids, adipokines, and hepatokines. Finally, in section VIII, we propose an integrated model of insulin resistance that links these mediators to final common pathways of metabolite-driven gluconeogenesis and ectopic lipid accumulation.

Hunter R.W., Hughey C.C., Lantier L., Sundelin E.I., Peggie M., Zeqiraj E., Sicheri F., Jessen N., Wasserman D.H., Sakamoto K.

Metformin is a first-line drug for the treatment of individuals with type 2 diabetes, yet its precise mechanism of action remains unclear. Metformin exerts its antihyperglycemic action primarily through lowering hepatic glucose production (HGP). This suppression is thought to be mediated through inhibition of mitochondrial respiratory complex I, and thus elevation of 5′-adenosine monophosphate (AMP) levels and the activation of AMP-activated protein kinase (AMPK), though this proposition has been challenged given results in mice lacking hepatic AMPK. Here we report that the AMP-inhibited enzyme fructose-1,6-bisphosphatase-1 (FBP1), a rate-controlling enzyme in gluconeogenesis, functions as a major contributor to the therapeutic action of metformin. We identified a point mutation in FBP1 that renders it insensitive to AMP while sparing regulation by fructose-2,6-bisphosphate (F-2,6-P2), and knock-in (KI) of this mutant in mice significantly reduces their response to metformin treatment. We observe this during a metformin tolerance test and in a metformin-euglycemic clamp that we have developed. The antihyperglycemic effect of metformin in high-fat diet–fed diabetic FBP1-KI mice was also significantly blunted compared to wild-type controls. Collectively, we show a new mechanism of action for metformin and provide further evidence that molecular targeting of FBP1 can have antihyperglycemic effects. The antidiabetic action of metformin raises heptocyte intracellular AMP levels, causing allosteric inhibition of fructose-1,6-bisphosphatase and thus reductions in gluconeogenesis.

Thakur S., Daley B., Gaskins K., Vasko V.V., Boufraqech M., Patel D., Sourbier C., Reece J., Cheng S., Kebebew E., Agarwal S., Klubo-Gwiezdzinska J.

Abstract Purpose: Mitochondrial glycerophosphate dehydrogenase (MGPDH) is the key enzyme connecting oxidative phosphorylation (OXPHOS) and glycolysis as well as a target of the antidiabetic drug metformin in the liver. There are no data on the expression and role of MGPDH as a metformin target in cancer. In this study, we evaluated MGPDH as a potential target of metformin in thyroid cancer and investigated its contribution in thyroid cancer metabolism. Experimental Design: We analyzed MGPDH expression in 253 thyroid cancer and normal tissues by immunostaining and examined its expression and localization in thyroid cancer–derived cell lines (FTC133, BCPAP) by confocal microscopy. The effects of metformin on MGPDH expression were determined by qRT-PCR and Western blot analysis. Seahorse analyzer was utilized to assess the effects of metformin on OXPHOS and glycolysis in thyroid cancer cells. We analyzed the effects of metformin on tumor growth and MGPDH expression in metastatic thyroid cancer mouse models. Results: We show for the first time that MGPDH is overexpressed in thyroid cancer compared with normal thyroid. We demonstrate that MGPDH regulates human thyroid cancer cell growth and OXPHOS rate in vitro. Metformin treatment is associated with downregulation of MGPDH expression and inhibition of OXPHOS in thyroid cancer in vitro. Cells characterized by high MGPDH expression are more sensitive to OXPHOS-inhibitory effects of metformin in vitro and growth-inhibitory effects of metformin in vitro and in vivo. Conclusions: Our study established MGPDH as a novel regulator of thyroid cancer growth and metabolism that can be effectively targeted by metformin. Clin Cancer Res; 24(16); 4030–43. ©2018 AACR.

Madiraju A.K., Qiu Y., Perry R.J., Rahimi Y., Zhang X., Zhang D., Camporez J.G., Cline G.W., Butrico G.M., Kemp B.E., Casals G., Steinberg G.R., Vatner D.F., Petersen K.F., Shulman G.I.

Metformin, the universal first-line treatment for type 2 diabetes, exerts its therapeutic glucose-lowering effects by inhibiting hepatic gluconeogenesis. However, the primary molecular mechanism of this biguanide remains unclear, though it has been suggested to act, at least partially, by mitochondrial complex I inhibition. Here we show that clinically relevant concentrations of plasma metformin achieved by acute intravenous, acute intraportal or chronic oral administration in awake normal and diabetic rats inhibit gluconeogenesis from lactate and glycerol but not from pyruvate and alanine, implicating an increased cytosolic redox state in mediating metformin’s antihyperglycemic effect. All of these effects occurred independently of complex I inhibition, evidenced by unaltered hepatic energy charge and citrate synthase flux. Normalizing the cytosolic redox state by infusion of methylene blue or substrates that contribute to gluconeogenesis independently of the cytosolic redox state abrogated metformin-mediated inhibition of gluconeogenesis in vivo. Additionally, in mice expressing constitutively active acetyl-CoA carboxylase, metformin acutely decreased hepatic glucose production and increased the hepatic cytosolic redox state without altering hepatic triglyceride content or gluconeogenic enzyme expression. These studies demonstrate that metformin, at clinically relevant plasma concentrations, inhibits hepatic gluconeogenesis in a redox-dependent manner independently of reductions in citrate synthase flux, hepatic nucleotide concentrations, acetyl-CoA carboxylase activity, or gluconeogenic enzyme protein expression. Using 13C-labeled substrates in vivo, this group shows that metformin inhibits mG3PDH to reduce hepatic gluconeogenesis and lower glycemia by altering the redox potential of the cytosol of hepatocytes rather than affecting substrate availability.

Qi H., Nielsen P.M., Schroeder M., Bertelsen L.B., Palm F., Laustsen C.

Metformin inhibits hepatic mitochondrial glycerol phosphate dehydrogenase, thereby increasing cytosolic lactate and suppressing gluconeogenesis flux in the liver. This inhibition alters cytosolic and mitochondrial reduction–oxidation (redox) potential, which has been reported to protect organ function in several disease states including diabetes. In this study, we investigated the acute metabolic and functional changes induced by metformin in the kidneys of both healthy and insulinopenic Wistar rats used as a model of diabetes. Diabetes was induced by intravenous injection of streptozotocin, and kidney metabolism in healthy and diabetic animals was investigated 4 weeks thereafter using hyperpolarised 13C-MRI, Clark-type electrodes and biochemical analysis. Metformin increased renal blood flow, but did not change total kidney oxygen consumption. In healthy rat kidneys, metformin increased [1-13C]lactate production and reduced mitochondrial [1-13C]pyruvate oxidation (decreased the 13C-bicarbonate/[1-13C]pyruvate ratio) within 30 min of administration. Corresponding alterations to indices of mitochondrial, cytosolic and whole-cell redox potential were observed. Pyruvate oxidation was maintained in the diabetic rats, suggesting that the diabetic state abrogates metabolic reprogramming caused by metformin. This study demonstrates that metformin-induced acute metabolic alterations in healthy kidneys favoured anaerobic metabolism at the expense of aerobic metabolism. The results suggest that metformin directly alters the renal redox state, with elevated renal cytosolic redox states as well as decreased mitochondrial redox state. These findings suggest redox biology as a novel target to eliminate the renal complications associated with metformin treatment in individuals with impaired renal function.

Iversen A.B., Horsman M.R., Jakobsen S., Jensen J.B., Garm C., Jessen N., Breining P., Frøkiær J., Busk M.

The anti-diabetic biguanide drugs metformin (METF) and phenformin (PHEN) may have anti-cancer effects. Biguanides suppress plasma growth factors, but nonetheless, the view that these mitochondrial inhibitors accumulate in tumor tissue to an extent that leads to severe energetic stress or alleviation of hypoxia-induced radioresistance is gaining ground. Our cell studies confirm that biguanides inhibits cell proliferation by targeting respiration, but only at highly suprapharmacological concentrations due to low drug retention. Biodistribution/PET studies of 11C-labeled metformin (11C-METF) revealed that plasma bioavailability remained well below concentrations with metabolic/anti-proliferative in vitro effects, following a high oral dose. Intraperitoneal administration resulted in higher drug concentrations, which affected metabolism in normal organs with high METF uptake (e.g., kidneys), but tumor drug retention peaked at low levels comparable to plasma levels and hypoxia was unaffected. Prolonged intraperitoneal treatment reduced tumor growth in two tumor models, however, the response did not reflect in vitro drug sensitivity, and tumor metabolism and hypoxia was unaffected. Our results do not support that direct inhibition of tumor cell respiration is responsible for reduced tumor growth, but future studies using 11C-METF-PET are warranted, preferably in neoplasia’s originating from tissue with high drug transport capacity, to investigate the controversial idea of direct targeting.

Xiang J., An Y., Sun J., Xu J., Xiong Y., Wang S., Xia Q.

Roeb E.

Iheagwam F.N., Iheagwam O.T.

McAleese C., Joudah G., Salt I.P., Petrie J.R., Leiper J.M., Dowsett L.B.

AbstractEndothelial cells (ECs) are highly glycolytic, with mitochondria primarily serving a signalling function. Metabolic disruptions are early contributors to endothelial dysfunction, a primary feature of diabetic vascular complications, such as retinopathy, impaired wound healing and cerebral small vessel disease. The degree to which metabolism varies amongst such different vascular beds is unknown. Mitochondrial function was therefore characterised in human aortic, dermal, retinal and cerebral ECs in vitro, aiming to determine whether basal metabolism influences the response and susceptibility of vascular beds experimental hyperglycaemia (HG). Furthermore, the potential of metformin to maintain endothelial function independent of glycaemic control was assessed. Using a Seahorse analyser, metabolic function of human primary ECs from different vascular beds was compared under basal conditions, as well as HG and metformin treatment. ECs differed significantly in respiratory profile and glycolytic function. For example aortic ECs were preferentially aerobic, whereas dermal ECs were glycolytic. HG significantly lowered mitochondrial network area but elicited modest effects upon respiratory function at the same time as influencing glycolytic function in a manner that was possibly conditional upon basal utilisation. Metformin inhibited basal respiratory function at the same time as significantly enhancing glycolysis in retinal and brain ECs. These data suggest that EC responses to HG and metformin are influenced by the basal metabolic profile, highlighting the potential of targeting EC metabolism to preserve function in a diabetic condition. A nuanced approach is needed to address diabetic vascular complications and endothelial metabolic health in diabetes, both in the investigation of pathophysiology and in prospective therapeutics. imageKey points Endothelial dysfunction is an early feature of diabetes‐associated cardiovascular complications Endothelial cells (ECs) are highly glycolytic, with mitochondria serving a signalling function ECs are known to be heterogeneous in function, but how this is reflected in metabolism is not fully understood, in addition to how this influences their response to hyperglycaemia Using experimental hyperglycaemia (HG) in vitro, we demonstrate that ECs differed significantly in respiratory profile and glycolytic function. Their response to HG is possibly contingent upon this basal utilisation. These results suggest a nuanced approach is needed when investigating diabetic vascular complications, both in the investigation of pathophysiology and in prospective therapeutics.

Gillespie A., Mehdorn A., Lim T.Q., Wang T., Mooney B.A., Ovens A.J., Orang A., Oakhill J.S., Michael M.Z., Petersen J.

Abstract Metformin is a well-tolerated drug frequently prescribed for managing type 2 diabetes. Extended metformin use has been linked to a significant decrease in cancer incidence across both diabetic and non-diabetic populations. Here we investigate the anti-proliferative effects of metformin on fission yeast S. pombe. Our findings demonstrate that metformin’s inhibitory impact on cell proliferation is effective in the absence of AMP-activated protein kinase (AMPK). Using an unbiased genetic screen we identified the plasma membrane signalling scaffold Efr3, critical for phosphatidylinositol signalling and the generation of PI4Ps, as a key determinant of resistance to the anti-proliferative effect of metformin. Deletion of efr3 resulted in both AMPK-dependent and AMPK-independent resistance to metformin. We show that Efr3 does not influence cell proliferation by controlling Ras1 activity or its cellular localization in yeast. We observe that dnm1 (DRP1) mutants with elongated mitochondria are also resistant to the anti-proliferative effect of metformin and that metformin treatment promotes mitochondrial fusion. Metabolic measurements after prolonged metformin exposure demonstrated a reduction in respiration in both wild type and the efr3 deletion, however, that reduction is less pronounced in the efr3 deletion, which also contained elongated mitochondria. It is likely that mitochondrial fusion enhances yeast fitness in response to metformin exposure. Together we provide a new perspective on the cellular response to metformin.

Yi Y., Wang G., Zhang W., Yu S., Fei J., An T., Yi J., Li F., Huang T., Yang J., Niu M., Wang Y., Xu C., Xiao Z.J.

AbstractGlucose deprivation, a hallmark of the tumor microenvironment, compels tumor cells to seek alternative energy sources for survival and growth. Here, we show that glucose deprivation upregulates the expression of mitochondrial-cytochrome c oxidase II (MT-CO2), a subunit essential for the respiratory chain complex IV, in facilitating glutaminolysis and sustaining tumor cell survival. Mechanistically, glucose deprivation activates Ras signaling to enhance MT-CO2 transcription and inhibits IGF2BP3, an RNA-binding protein, to stabilize MT-CO2 mRNA. Elevated MT-CO2 increases flavin adenosine dinucleotide (FAD) levels in activating lysine-specific demethylase 1 (LSD1) to epigenetically upregulate JUN transcription, consequently promoting glutaminase-1 (GLS1) and glutaminolysis for tumor cell survival. Furthermore, MT-CO2 is indispensable for oncogenic Ras-induced glutaminolysis and tumor growth, and elevated expression of MT-CO2 is associated with poor prognosis in lung cancer patients. Together, these findings reveal a role for MT-CO2 in adapting to metabolic stress and highlight MT-CO2 as a putative therapeutic target for Ras-driven cancers.

Pham L., Arroum T., Wan J., Pavelich L., Bell J., Morse P.T., Lee I., Grossman L.I., Sanderson T.H., Malek M.H., Hüttemann M.

Barroso E., Jurado-Aguilar J., Wahli W., Palomer X., Vázquez-Carrera M.

Abnormally increased hepatic gluconeogenesis is a significant contributor to hyperglycemia in the fasting state in patients with type 2 diabetes mellitus (T2DM) due to insulin resistance. Metformin, the most prescribed drug for the treatment of T2DM, is believed to exert its effect mainly by reducing hepatic gluconeogenesis. Here, we discuss how increased hepatic gluconeogenesis contributes to T2DM and we review newly revealed mechanisms underlying the attenuation of gluconeogenesis by metformin. In addition, we analyze the recent findings on new determinants involved in the regulation of gluconeogenesis, which might ultimately lead to the identification of novel and targeted treatment strategies for T2DM.

Kiyuna L.A., Krishnamurthy K.A., Homan E.B., Langelaar-Makkinje M., Gerding A., Bos T., Oosterhuis D., Overduin R.J., Schreuder A.B., de Meijer V.E., Olinga P., Derks T.G., van Eunen K., Bakker B.M., Oosterveer M.H.

Fasting hypoglycemia is a severe and incompletely understood symptom of various inborn errors of metabolism (IEM). Precision-cut liver slices (PCLS) represent a promising model for studying glucose production ex vivo. This study quantified the net glucose production of human and murine PCLS in the presence of different gluconeogenic precursors. Dihydroxyacetone-supplemented slices from the fed mice yielded the highest rate, further stimulated by forskolin and dibutyryl-cAMP. Moreover, using 13C isotope tracing, we assessed the contribution of glycogenolysis and gluconeogenesis to net glucose production over time. Pharmacological inhibition of the glucose 6-phosphate transporter SLC37A4 markedly reduced net glucose production and increased lactate secretion and glycogen storage, while glucose production was completely abolished in PCLS from glycogen storage disease type Ia and Ib patients. In conclusion, this study identifies PCLS as an effective ex vivo model to study hepatic glucose production and opens opportunities for its future application in IEM research and beyond. Ex vivo precision-cut liver slices allow glucose production quantification to model glycogen storage disease and inborn errors of metabolism.

LaMoia T.E., Hubbard B.T., Guerra M.T., Nasiri A., Sakuma I., Kahn M., Zhang D., Goodman R.P., Nathanson M.H., Sancak Y., Perelis M., Mootha V.K., Shulman G.I.

To examine the roles of mitochondrial calcium Ca

Yang Y., Lu X., Liu N., Ma S., Zhang H., Zhang Z., Yang K., Jiang M., Zheng Z., Qiao Y., Hu Q., Huang Y., Zhang Y., Xiong M., Liu L., et. al.

In a rigorous 40-month study, we evaluated the geroprotective effects of metformin on adult male cynomolgus monkeys, addressing a gap in primate aging research. The study encompassed a comprehensive suite of physiological, imaging, histological, and molecular evaluations, substantiating metformin's influence on delaying age-related phenotypes at the organismal level. Specifically, we leveraged pan-tissue transcriptomics, DNA methylomics, plasma proteomics, and metabolomics to develop innovative monkey aging clocks and applied these to gauge metformin's effects on aging. The results highlighted a significant slowing of aging indicators, notably a roughly 6-year regression in brain aging. Metformin exerts a substantial neuroprotective effect, preserving brain structure and enhancing cognitive ability. The geroprotective effects on primate neurons were partially mediated by the activation of Nrf2, a transcription factor with anti-oxidative capabilities. Our research pioneers the systemic reduction of multi-dimensional biological age in primates through metformin, paving the way for advancing pharmaceutical strategies against human aging.

Sirtori C.R., Castiglione S., Pavanello C.

The metformin molecule dates back to over a century, but its clinical use started in the '50s. Since then, its use in diabetics has grown constantly, with over 150 million users today. The therapeutic profile also expanded, with improved understanding of novel mechanisms. Metformin has a major activity on insulin resistance, by acting on the insulin receptors and mitochondria, most likely by activation of the adenosine monophosphate-activated kinase. These and associated mechanisms lead to significant lipid lowering and body weight loss. An anti-cancer action has come up in recent years, with mechanisms partly dependent on the mitochondrial activity and also on phosphatidylinositol 3-kinase resistance occurring in some malignant tumors. The potential of metformin to raise life-length is the object of large ongoing studies and of several basic and clinical investigations. The present review article will attempt to investigate the basic mechanisms behind these diverse activities and the potential clinical benefits. Metformin may act on transcriptional activity by histone modification, DNA methylation and miRNAs. An activity on age-associated inflammation (inflammaging) may occur via activation of the nuclear factor erythroid 2 related factor and changes in gut microbiota. A senolytic activity, leading to reduction of cells with the senescent associated secretory phenotype, may be crucial in lifespan prolongation as well as in ancillary properties in age-associated diseases, such as Parkinson's disease. Telomere prolongation may be related to the activity on mitochondrial respiratory factor 1 and on peroxisome gamma proliferator coactivator 1-alpha. Very recent observations on the potential to act on the most severe neurological disorders, such as amyotrophic lateral sclerosis and frontotemporal dementia, have raised considerable hope.

Lefrançois G., Lavallée E., Rowell M., Bourdeau V., Mohebali F., Berthomeu T., Tyers M., Gravel S., Schmitzer A.R., Ferbeyre G.

AbstractHere we identify the subunit e of F1Fo-ATP synthase (ATP5I) as a target of medicinal biguanides. ATP5I maintains the stability of F1Fo-ATP synthase dimers which is crucial for shaping cristae morphology. Although its roles have been mainly studied in yeast models, its function in cellular energy metabolism within the context of cancer remains poorly characterized. In this study, we demonstrate that ATP5I interacts with a biguanide analoguein vitroand disabling its expression by CRISPR-Cas9 in pancreatic cancer cells leads to the same phenotype as biguanide treated cells including a decrease in the levels of some respiratory complex subunits, mitochondrial morphology alterations, inhibition of oxidative phosphorylation (OXPHOS) and a compensatory increase in glycolysis. Moreover, ATP5I knockout (KO) cells exhibit resistance to the antiproliferative effects of biguanides, but reintroduction of ATP5I rescues the metabolic and anti-proliferative effects of metformin and phenformin. These findings highlight ATP5I as a significant antineoplastic mitochondrial target of medicinal biguanides, opening new opportunities for the development of mitochondrial-targeted therapies.

Yao L., Wang L., Zhang R., Soukas A.A., Wu L.

Metformin, an oral antihyperglycemic drug that has been in use for over 60 years, remains a first-line therapy for type 2 diabetes (T2D). Numerous studies have suggested that metformin promotes health benefits beyond T2D management, including weight loss, cancer prevention and treatment, and anti-aging, through several proposed mechanistic targets. Here we discuss the established effects of metformin and the progress made in identifying its direct targets. Additionally, we emphasize the importance of elucidating the structural bases of the drug and its direct targets. Ultimately, this review aims to highlight the current state of knowledge regarding metformin and its related emerging discoveries, while also outlining critical future research directions.

Radosavljevic T., Brankovic M., Samardzic J., Djuretić J., Vukicevic D., Vucevic D., Jakovljevic V.

Metabolic dysfunction-associated steatotic liver disease (MASLD), formerly known as nonalcoholic fatty liver disease (NAFLD), encompasses a range of liver conditions from steatosis to nonalcoholic steatohepatitis (NASH). Its prevalence, especially among patients with metabolic syndrome, highlights its growing global impact. The pathogenesis of MASLD involves metabolic dysregulation, inflammation, oxidative stress, genetic factors and, notably, mitochondrial dysfunction. Recent studies underscore the critical role of mitochondrial dysfunction in MASLD’s progression. Therapeutically, enhancing mitochondrial function has gained interest, along with lifestyle changes and pharmacological interventions targeting mitochondrial processes. The FDA’s approval of resmetirom for metabolic-associated steatohepatitis (MASH) with fibrosis marks a significant step. While resmetirom represents progress, further research is essential to understand MASLD-related mitochondrial dysfunction fully. Innovative strategies like gene editing and small-molecule modulators, alongside lifestyle interventions, can potentially improve MASLD treatment. Drug repurposing and new targets will advance MASLD therapy, addressing its increasing global burden. Therefore, this review aims to provide a better understanding of the role of mitochondrial dysfunction in MASLD and identify more effective preventive and treatment strategies.