3-D finite element model of the impaction of a press-fitted femoral stem under various biomechanical environments.

BACKGROUND: Uncemented femoral stem insertion into the bone is achieved by applying successive impacts on an inserter tool called "ancillary". Impact analysis has shown to be a promising technique to monitor the implant insertion and to improve its primary stability. METHOD: This study aims to provide a better understanding of the dynamic phenomena occurring between the hammer, the ancillary, the implant and the bone during femoral stem insertion, to validate the use of impact analyses for implant insertion monitoring. A dynamic 3-D finite element model of the femoral stem insertion via an impaction protocol is proposed. The influence of the trabecular bone Young's modulus (Et), the interference fit (IF), the friction coefficient at the bone-implant interface (µ) and the impact velocity (v0) on the implant insertion and on the impact force signal is evaluated. RESULTS: For all configurations, a decrease of the time difference between the two first peaks of the impact force signal is observed throughout the femoral stem insertion, up to a threshold value of 0.23 ms. The number of impacts required to reach this value depends on Et, v0 and IF and varies between 3 and 8 for the set of parameters considered herein. The bone-implant contact ratio reached after ten impacts varies between 60% and 98%, increases as a function of v0 and decreases as a function of IF, µ and Et. CONCLUSION: This study confirms the potential of an impact analyses-based method to monitor implant insertion and to retrieve bone-implant contact properties.

Poorly Differentiated Hepatocellular Carcinoma Cells Avoid Apoptosis by Interacting with T Cells via CD40-CD40L Linkage.

Hepatocellular carcinoma (HCC) was associated with increased soluble CD40 levels in a previous study. This study aimed to investigate CD40's role in liver tumor progression. CD40 levels were examined in HCC patient tissues and various HCC cell lines, and their interaction with CD4+T cells was studied. RNA sequencing analysis was performed to explore the mechanisms of CD40 induction. Poorly differentiated HCC tumor tissues exhibited high membrane-bound CD40 expression, in contrast to nontumor areas. Poorly differentiated HCC cell lines showed high expression of membrane-bound CD40 with low CD40 promoter methylation, which was opposite of well-differentiated ones. Solely modulating CD40 expression in HCC cells exerted no direct consequences on cell growth or appearance. Interestingly, HLFs co-cultured with activated (CD40 ligand+) CD4+ T cells increased CD40 levels and showed a modest 3.2% dead cells, then increased to 10.9% underwent preneutralizing CD40 condition, whereas preblocking both CD40 and integrin α5ß1 concomitantly caused only 1.9% cell death. RNA sequencing of co-cultured HLFs with activated CD4+ T cells revealed the up-regulation of interferon and immune-response pathways. Increased interferon-γ levels in the activated T-cell media stimulated the Janus kinase/signal transducer and activator of transcription 3 pathway, resulting in increased CD40 expression in HLF. Collectively, CD40 expression in poorly differentiated HCC cells prevents cell death by interacting with CD40 ligand in activated T cells. Targeting CD40 may represent a promising anticancer therapy.

Characterization of the concentration of agar-based soft tissue mimicking phantoms by impact analysis.

In various medical fields, a change of soft tissue stiffness is associated with its physio-pathological evolution. While elastography is extensively employed to assess soft tissue stiffness in vivo, its application requires a complex and expensive technology. The aim of this study is to determine whether an easy-to-use method based on impact analysis can be employed to determine the concentration of agar-based soft tissue mimicking phantoms. Impact analysis was performed on soft tissue mimicking phantoms made of agar gel with a mass concentration ranging from 1% to 5%. An indicator Δt is derived from the temporal variation of the impact force signal between the hammer and a small beam in contact with the sample. The results show a non-linear decrease of Δt as a function of the agar concentration (and thus of the sample stiffness). The value of Δt provides an estimation of the agar concentration with an error of 0.11%. This sensitivity of the impact analysis based method to the agar concentration is of the same order of magnitude than results obtained with elastography techniques. This study opens new paths towards the development of impact analysis for a fast, easy and relatively inexpensive clinical evaluation of soft tissue elastic properties.

New insights in the targets of action of dimethyl fumarate in endothelial cells: effects on energetic metabolism and serine synthesis in vitro and in vivo.

Dimethyl fumarate is an ester from the Krebs cycle intermediate fumarate. This drug is approved and currently used for the treatment of psoriasis and multiple sclerosis, and its anti-angiogenic activity was reported some years ago. Due to the current clinical relevance of this compound and the recently manifested importance of endothelial cell metabolism on the angiogenic switch, we wanted to elucidate whether dimethyl fumarate has an effect on energetic metabolism of endothelial cells. Different experimental approximations were performed in endothelial cells, including proteomics, isotope tracing and metabolomics experimental approaches, in this work we studied the possible role of dimethyl fumarate in endothelial cell energetic metabolism. We demonstrate for the first time that dimethyl fumarate promotes glycolysis and diminishes cell respiration in endothelial cells, which could be a consequence of a down-regulation of serine and glycine synthesis through inhibition of PHGDH activity in these cells. Dimethyl fumarate alters the energetic metabolism of endothelial cells in vitro and in vivo through an unknown mechanism, which could be the cause or the consequence of its pharmacological activity. This new discovery on the targets of this compound could open a new field of study regarding the mechanism of action of dimethyl fumarate.

Comprehensive isotopomer analysis of glutamate and aspartate in small tissue samples.

Stable isotopes are powerful tools to assess metabolism. 13C labeling is detected using nuclear magnetic resonance (NMR) spectroscopy or mass spectrometry (MS). MS has excellent sensitivity but generally cannot discriminate among different 13C positions (isotopomers), whereas NMR is less sensitive but reports some isotopomers. Here, we develop an MS method that reports all 16 aspartate and 32 glutamate isotopomers while requiring less than 1% of the sample used for NMR. This method discriminates between pathways that result in the same number of 13C labels in aspartate and glutamate, providing enhanced specificity over conventional MS. We demonstrate regional metabolic heterogeneity within human tumors, document the impact of fumarate hydratase (FH) deficiency in human renal cancers, and investigate the contributions of tricarboxylic acid (TCA) cycle turnover and CO2 recycling to isotope labeling in vivo. This method can accompany NMR or standard MS to provide outstanding sensitivity in isotope-labeling experiments, particularly in vivo.

Adipocytes reprogram cancer cell metabolism by diverting glucose towards glycerol-3-phosphate thereby promoting metastasis.

In the tumor microenvironment, adipocytes function as an alternate fuel source for cancer cells. However, whether adipocytes influence macromolecular biosynthesis in cancer cells is unknown. Here we systematically characterized the bidirectional interaction between primary human adipocytes and ovarian cancer (OvCa) cells using multi-platform metabolomics, imaging mass spectrometry, isotope tracing and gene expression analysis. We report that, in OvCa cells co-cultured with adipocytes and in metastatic tumors, a part of the glucose from glycolysis is utilized for the biosynthesis of glycerol-3-phosphate (G3P). Normoxic HIF1α protein regulates the altered flow of glucose-derived carbons in cancer cells, resulting in increased glycerophospholipids and triacylglycerol synthesis. The knockdown of HIF1α or G3P acyltransferase 3 (a regulatory enzyme of glycerophospholipid synthesis) reduced metastasis in xenograft models of OvCa. In summary, we show that, in an adipose-rich tumor microenvironment, cancer cells generate G3P as a precursor for critical membrane and signaling components, thereby promoting metastasis. Targeting biosynthetic processes specific to adipose-rich tumor microenvironments might be an effective strategy against metastasis.

Limiting mitochondrial plasticity by targeting DRP1 induces metabolic reprogramming and reduces breast cancer brain metastases.

Disseminated tumor cells with metabolic flexibility to utilize available nutrients in distal organs persist, but the precise mechanisms that facilitate metabolic adaptations remain unclear. Here we show fragmented mitochondrial puncta in latent brain metastatic (Lat) cells enable fatty acid oxidation (FAO) to sustain cellular bioenergetics and maintain redox homeostasis. Depleting the enriched dynamin-related protein 1 (DRP1) and limiting mitochondrial plasticity in Lat cells results in increased lipid droplet accumulation, impaired FAO and attenuated metastasis. Likewise, pharmacological inhibition of DRP1 using a small-molecule brain-permeable inhibitor attenuated metastatic burden in preclinical models. In agreement with these findings, increased phospho-DRP1 expression was observed in metachronous brain metastasis compared with patient-matched primary tumors. Overall, our findings reveal the pivotal role of mitochondrial plasticity in supporting the survival of Lat cells and highlight the therapeutic potential of targeting cellular plasticity programs in combination with tumor-specific alterations to prevent metastatic recurrences.

Electron transport chain inhibition increases cellular dependence on purine transport and salvage.

Cancer cells reprogram their metabolism to support cell growth and proliferation in harsh environments. While many studies have documented the importance of mitochondrial oxidative phosphorylation (OXPHOS) in tumor growth, some cancer cells experience conditions of reduced OXPHOS in vivo and induce alternative metabolic pathways to compensate. To assess how human cells respond to mitochondrial dysfunction, we performed metabolomics in fibroblasts and plasma from patients with inborn errors of mitochondrial metabolism, and in cancer cells subjected to inhibition of the electron transport chain (ETC). All these analyses revealed extensive perturbations in purine-related metabolites; in non-small cell lung cancer (NSCLC) cells, ETC blockade led to purine metabolite accumulation arising from a reduced cytosolic NAD + /NADH ratio (NADH reductive stress). Stable isotope tracing demonstrated that ETC deficiency suppressed de novo purine nucleotide synthesis while enhancing purine salvage. Analysis of NSCLC patients infused with [U- 13 C]glucose revealed that tumors with markers of low oxidative mitochondrial metabolism exhibited high expression of the purine salvage enzyme HPRT1 and abundant levels of the HPRT1 product inosine monophosphate (IMP). ETC blockade also induced production of ribose-5' phosphate (R5P) by the pentose phosphate pathway (PPP) and import of purine nucleobases. Blocking either HPRT1 or nucleoside transporters sensitized cancer cells to ETC inhibition, and overexpressing nucleoside transporters was sufficient to drive growth of NSCLC xenografts. Collectively, this study mechanistically delineates how cells compensate for suppressed purine metabolism in response to ETC blockade, and uncovers a new metabolic vulnerability in tumors experiencing NADH excess.

L-2-hydroxyglutaric aciduria - review of literature and case series.

L-2-hydroxyglutaric aciduria (L2HGA) is an autosomal recessive, slowly progressive neurodegenerative disease characterized by psychomotor delay and cerebellar dysfunction. The biochemical hallmark is increased concentrations of L2HG in body fluids. Brain MRI exhibits characteristic centripetal extension of the white matter involvement that differentiates it from other leukodystrophies. The authors report two sisters from Pakistan with L2HGA with 4 years of follow-up. The authors have also compared the clinical outcome of our patients with 45 previously reported patients with L2HGA for whom treatment and clinical outcome was reported. Case presentation: The authors report two sisters with L2HGA from Pakistan born to consanguineous parents. The 15- and 17-year-old girls presented with psychomotor delay, seizures, ataxia, intentional tremors, and dysarthria. Both had normal anthropometric measurements for age. Exaggerated tendon reflexes and bilateral sustained ankle clonus were observed in addition to cerebellar signs. Urine organic acids analysis showed marked excretion of 2-hydroxyglutaric acid, chiral differentiation of 2-hydroxyglutaric acid showed it to be L2HGA. Brain MRI of the 15-year-old showed diffuse subcortical white matter changes evident by T2/FLAIR hyperintense signals bilaterally, particularly in the frontal region in the centripetal distribution with some diffusion restriction along involvement of globus pallidus. The characteristic MRI pattern raised the suspicion of L2HGA. Targeted L2HGDH sequencing identified a homozygous pathogenic variant, c.829C>T (p.Arg227*) in L2HGDH gene in both girls. Both parents were heterozygous carriers of the familial variant. Conclusion: Neuroradiological features of centripetal subcortical leukoencephalopathy with basal ganglia and dentate nuclei involvement are rather specific to L2HGA and should lead to further biochemical investigations to look for L2HGA and L2HGDH gene sequencing.

Correction: Vu et al. Specific Changes in Arabidopsis thaliana Rosette Lipids during Freezing Can Be Associated with Freezing Tolerance. Metabolites 2022, 12, 385.

There was an error in the original publication [...].

Genomic characterization of endemic diarrheagenic Escherichia coli and Escherichia albertii from infants with diarrhea in Vietnam.

BACKGROUND: Diarrheagenic Escherichia coli (DEC) is a group of bacterial pathogens that causes life-threatening diarrhea in children in developing countries. However, there is limited information on the characteristics of DEC isolated from patients in these countries. A detailed genomic analysis of 61 DEC-like isolates from infants with diarrhea was performed to clarify and share the characteristics of DEC prevalent in Vietnam. PRINCIPAL FINDINGS: DEC was classified into 57 strains, including 33 enteroaggregative E. coli (EAEC) (54.1%), 20 enteropathogenic E. coli (EPEC) (32.8%), two enteroinvasive E. coli (EIEC) (3.3%), one enterotoxigenic E. coli (ETEC), and one ETEC/EIEC hybrid (1.6% each), and surprisingly into four Escherichia albertii strains (6.6%). Furthermore, several epidemic DEC clones showed an uncommon combination of pathotypes and serotypes, such as EAEC Og130:Hg27, EAEC OgGp9:Hg18, EAEC OgX13:H27, EPEC OgGp7:Hg16, and E. albertii EAOg1:HgUT. Genomic analysis also revealed the presence of various genes and mutations associated with antibiotic resistance in many isolates. Strains that demonstrate potential resistance to ciprofloxacin and ceftriaxone, drugs recommended for treating childhood diarrhea, accounted for 65.6% and 41%, respectively. SIGNIFICANCE: Our finding indicate that the routine use of these antibiotics has selected resistant DECs, resulting in a situation where these drugs do not provide in therapeutic effects for some patients. Bridging this gap requires continuous investigations and information sharing regarding the type and distribution of endemic DEC and E. albertii and their antibiotic resistance in different countries.

Recessive pathogenic variants in MCAT cause combined oxidative phosphorylation deficiency.

Malonyl-CoA-acyl carrier protein transacylase (MCAT) is an enzyme involved in mitochondrial fatty acid synthesis (mtFAS) and catalyzes the transfer of the malonyl moiety of malonyl-CoA to the mitochondrial acyl carrier protein (ACP). Previously, we showed that loss-of-function of mtFAS genes, including Mcat, is associated with severe loss of electron transport chain (ETC) complexes in mouse immortalized skeletal myoblasts (Nowinski et al., 2020). Here, we report a proband presenting with hypotonia, failure to thrive, nystagmus, and abnormal brain MRI findings. Using whole exome sequencing, we identified biallelic variants in MCAT. Protein levels for NDUFB8 and COXII, subunits of complex I and IV respectively, were markedly reduced in lymphoblasts and fibroblasts, as well as SDHB for complex II in fibroblasts. ETC enzyme activities were decreased in parallel. Re-expression of wild-type MCAT rescued the phenotype in patient fibroblasts. This is the first report of a patient with MCAT pathogenic variants and combined oxidative phosphorylation deficiency.

Detection of periprosthetic fractures around the femoral stem by resonance frequency analysis: An in vitro study.

Periprosthetic femoral bone fractures are frequent complications of Total Hip Arthroplasty (THA) and may occur during the insertion of uncemented Femoral Stems (FS), due to the nature of the press-fit fixation. Such fracture may lead to the surgical failure of the THA and require a revision surgery, which may have dramatic consequences. Therefore, an early detection of intra-operative fractures is important to avoid worsening the fracture and/or to enable a peroperative treatment. The aim of this in vitro study is to determine the sensitivity of a method based on resonance frequency analysis of the bone-stem-ancillary system for periprosthetic fractures detection. A periprosthetic fracture was artificially created close to the lesser-trochanter of 10 femoral bone mimicking phantoms. The bone-stem-ancillary resonance frequencies in the range (2-12) kHz were measured on an ancillary instrumented with piezoelectric sensors, which was fixed to the femoral stem. The measurements were repeated for different fracture lengths from 4 to 55 mm. The results show a decrease of the resonance frequencies due to the fracture occurrence and propagation. The frequency shift reached up to 170 Hz. The minimum fracture length that can be detected varies from 3.1±1.7 mm to 5.9±1.9 mm according to the mode and to the specimen. A significantly higher sensitivity (p = 0.011) was obtained for a resonance frequency around 10.6 kHz, corresponding to a mode vibrating in a plane perpendicular to the fracture. This study opens new paths toward the development of non-invasive vibration-based methods for intra-operative periprosthetic fractures detection.

Debonding of coin-shaped osseointegrated implants: Coupling of experimental and numerical approaches.

While cementless implants are now widely used clinically, implant debonding still occur and is difficult to anticipate. Assessing the biomechanical strength of the bone-implant interface can help improving the understanding of osseointegration phenomena and thus preventing surgical failures. A dedicated and standardized implant model was considered. The samples were tested using a mode III cleavage device to assess the mechanical strength of the bone-implant interface by combining experimental and numerical approaches. Four rough (Sa = 24.5 µm) osseointegrated coin-shaped implants were left in sheep cortical bone during 15 weeks of healing time. Each sample was experimentally rotated at 0.03°/sec until complete rupture of the interface. The maximum values of the torque were comprised between 0.48 and 0.72 N m, while a significant increase of the normal force from 7-12 N to 31-43 N was observed during the bone-implant interface debonding, suggesting the generation of bone debris at the bone-implant interface. The experimental results were compared to an isogeometric finite element model describing the adhesion and debonding phenomena through a modified Coulomb's law, based on a varying friction coefficient to represent the transition from an unbroken to a broken bone-implant interface. A good agreement was found between numerical and experimental torques, with numerical friction coefficients decreasing from 8.93 to 1.23 during the bone-implant interface rupture, which constitutes a validation of this model to simulate the debonding of an osseointegrated bone-implant interface subjected to torsion.

Mitochondrial metabolism in primary and metastatic human kidney cancers.

Most kidney cancers display evidence of metabolic dysfunction1-4 but how this relates to cancer progression in humans is unknown. We used a multidisciplinary approach to infuse 13C-labeled nutrients during surgical tumour resection in over 70 patients with kidney cancer. Labeling from [U-13C]glucose varies across cancer subtypes, indicating that the kidney environment alone cannot account for all metabolic reprogramming in these tumours. Compared to the adjacent kidney, clear cell renal cell carcinomas (ccRCC) display suppressed labelling of tricarboxylic acid (TCA) cycle intermediates in vivo and in organotypic slices cultured ex vivo, indicating that suppressed labeling is tissue intrinsic. Infusions of [1,2-13C]acetate and [U-13C]glutamine in patients, coupled with respiratory flux of mitochondria isolated from kidney and tumour tissue, reveal primary defects in mitochondrial function in human ccRCC. However, ccRCC metastases unexpectedly have enhanced labeling of TCA cycle intermediates compared to primary ccRCCs, indicating a divergent metabolic program during ccRCC metastasis in patients. In mice, stimulating respiration in ccRCC cells is sufficient to promote metastatic colonization. Altogether, these findings indicate that metabolic properties evolve during human kidney cancer progression, and suggest that mitochondrial respiration may be limiting for ccRCC metastasis but not for ccRCC growth at the site of origin.

Influence of the biomechanical environment on the femoral stem insertion and vibrational behavior: a 3-D finite element study.

The long-term success of cementless surgery strongly depends on the implant primary stability. The femoral stem initial fixation relies on multiple geometrical and material factors, but their influence on the biomechanical phenomena occurring during the implant insertion is still poorly understood, as they are difficult to quantify in vivo. The aim of the present study is to evaluate the relationship between the resonance frequencies of the bone-implant-ancillary system and the stability of the femoral stem under various biomechanical environments. The interference fit IF, the trabecular bone Young's modulus [Formula: see text] and the bone-implant contact friction coefficient [Formula: see text] are varied to investigate their influence on the implant insertion phenomena and on the system vibration behavior. The results exhibit for all the configurations, a nonlinear increase in the bone-implant contact throughout femoral stem insertion, until the proximal contact is reached. While the pull-out force increases with [Formula: see text], IF and [Formula: see text], the bone-implant contact ratio decreases, which shows that a compromise on the set of parameters could be found in order to achieve the largest bone-implant contact while maintaining sufficient pull-out force. The modal analysis on the range [2-7] kHz shows that the resonance frequencies of the bone-implant-ancillary system increase with the bone-implant contact ratio and the trabecular bone Young's modulus, with a sensitivity that varies over the modes. Both the pull-out forces and the vibration behavior are consistent with previous experimental studies. This study demonstrates the potential of using vibration methods to guide the surgeons for optimizing implant stability in various patients and surgical configurations.

Modeling the debonding process of osseointegrated implants due to coupled adhesion and friction.

Cementless implants have become widely used for total hip replacement surgery. The long-term stability of these implants is achieved by bone growing around and into the rough surface of the implant, a process called osseointegration. However, debonding of the bone-implant interface can still occur due to aseptic implant loosening and insufficient osseointegration, which may have dramatic consequences. The aim of this work is to describe a new 3D finite element frictional contact formulation for the debonding of partially osseointegrated implants. The contact model is based on a modified Coulomb friction law by Immel et al. (2020), that takes into account the tangential debonding of the bone-implant interface. This model is extended in the direction normal to the bone-implant interface by considering a cohesive zone model, to account for adhesion phenomena in the normal direction and for adhesive friction of partially bonded interfaces. The model is applied to simulate the debonding of an acetabular cup implant. The influence of partial osseointegration and adhesive effects on the long-term stability of the implant is assessed. The influence of different patient- and implant-specific parameters such as the friction coefficient [Formula: see text], the trabecular Young's modulus [Formula: see text], and the interference fit [Formula: see text] is also analyzed, in order to determine the optimal stability for different configurations. Furthermore, this work provides guidelines for future experimental and computational studies that are necessary for further parameter calibration.

An interactive web application for exploring human plasma and fibroblast metabolomics data from patients with inborn errors of metabolism.

Metabolomic profiling is instrumental in understanding the systemic and cellular impact of inborn errors of metabolism (IEMs), monogenic disorders caused by pathogenic genomic variants in genes involved in metabolism. This study encompasses untargeted metabolomics analysis of plasma from 474 individuals and fibroblasts from 67 subjects, incorporating healthy controls, patients with 65 different monogenic diseases, and numerous undiagnosed cases. We introduce a web application designed for the in-depth exploration of this extensive metabolomics database. The application offers a user-friendly interface for data review, download, and detailed analysis of metabolic deviations linked to IEMs at the level of individual patients or groups of patients with the same diagnosis. It also provides interactive tools for investigating metabolic relationships and offers comparative analyses of plasma and fibroblast profiles. This tool emphasizes the metabolic interplay within and across biological matrices, enriching our understanding of metabolic regulation in health and disease. As a resource, the application provides broad utility in research, offering novel insights into metabolic pathways and their alterations in various disorders.

In vivo characterization of glutamine metabolism identifies therapeutic targets in clear cell renal cell carcinoma.

Targeting metabolic vulnerabilities has been proposed as a therapeutic strategy in renal cell carcinoma (RCC). Here, we analyzed the metabolism of patient-derived xenografts (tumorgrafts) from diverse subtypes of RCC. Tumorgrafts from VHL-mutant clear cell RCC (ccRCC) retained metabolic features of human ccRCC and engaged in oxidative and reductive glutamine metabolism. Genetic silencing of isocitrate dehydrogenase-1 or isocitrate dehydrogenase-2 impaired reductive labeling of tricarboxylic acid (TCA) cycle intermediates in vivo and suppressed growth of tumors generated from tumorgraft-derived cells. Glutaminase inhibition reduced the contribution of glutamine to the TCA cycle and resulted in modest suppression of tumorgraft growth. Infusions with [amide-15N]glutamine revealed persistent amidotransferase activity during glutaminase inhibition, and blocking these activities with the amidotransferase inhibitor JHU-083 also reduced tumor growth in both immunocompromised and immunocompetent mice. We conclude that ccRCC tumorgrafts catabolize glutamine via multiple pathways, perhaps explaining why it has been challenging to achieve therapeutic responses in patients by inhibiting glutaminase.

Metabolon formation regulates branched-chain amino acid oxidation and homeostasis.

The branched-chain aminotransferase isozymes BCAT1 and BCAT2, segregated into distinct subcellular compartments and tissues, initiate the catabolism of branched-chain amino acids (BCAAs). However, whether and how BCAT isozymes cooperate with downstream enzymes to control BCAA homeostasis in an intact organism remains largely unknown. Here, we analyse system-wide metabolomic changes in BCAT1- and BCAT2-deficient mouse models. Loss of BCAT2 but not BCAT1 leads to accumulation of BCAAs and branched-chain α-keto acids (BCKAs), causing morbidity and mortality that can be ameliorated by dietary BCAA restriction. Through proximity labelling, isotope tracing and enzymatic assays, we provide evidence for the formation of a mitochondrial BCAA metabolon involving BCAT2 and branched-chain α-keto acid dehydrogenase. Disabling the metabolon contributes to BCAT2 deficiency-induced phenotypes, which can be reversed by BCAT1-mediated BCKA reamination. These findings establish a role for metabolon formation in BCAA metabolism in vivo and suggest a new strategy to modulate this pathway in diseases involving dysfunctional BCAA metabolism.