Anthracycline-induced cardiotoxicity poses a threat to the long-term heart health of childhood cancer survivors; however, early risk assessment remains challenging due to limited predictive tools. While risk prediction models have been developed for chemotherapy-related hematological toxicity in pediatric cancer patients, research efforts addressing anthracycline-induced cardiotoxicity in this population remain limited. To fill this gap, we conducted a pilot study to develop a risk prediction model for anthracycline-induced cardiotoxicity in children. Using a paired-sample design, we analyzed data collected throughout treatment from 18 children receiving anthracycline-based chemotherapy. Patient demographics, clinical features, and treatment regimen served as input (explanatory) variables, while plasma concentration of high-sensitivity cardiac troponin T was used as the outcome (response), representing anthracycline-induced myocardial injury. Of the initial 33 potential variables, the top 18 were selected based on their importance scores in relation to myocardial injury. This set was further refined to 13 by removing redundancies using the caret package in R to develop a preliminary logistic regression model. A Leave-One-Patient-Out Cross-Validation identified four key predictors for the final model: sex, age at diagnosis, total cyclophosphamide dose (mg), and days since the first anthracycline dose. All were significantly associated with myocardial injury. The final logistic regression model achieved an accuracy of 85%, a sensitivity of 80%, a specificity of 88%, an area under the curve of 0.89, and a Youden's index of 0.68 for predicting myocardial injury risk. These preliminary findings suggest the potential of our predictive model to stratify risk of anthracycline-induced myocardial injury in the pediatric population.

Many oncology drugs have been found to induce cardiotoxicity in a subset of patients, which significantly limits their clinical use and impedes the benefit of lifesaving anticancer treatments. Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) carry donor-specific genetic information and have been proposed for exploring the interindividual difference in oncology drug-induced cardiotoxicity. Herein, we evaluated the inter- and intraindividual variability of iPSC-CM-related assays and presented a proof of concept to prospectively predict doxorubicin (DOX)-induced cardiotoxicity (DIC) using donor-specific iPSC-CMs. Our findings demonstrated that donor-specific iPSC-CMs exhibited greater line-to-line variability than the intraindividual variability in impedance cytotoxicity and transcriptome assays. The variable and dose-dependent cytotoxic responses of iPSC-CMs resembled those observed in clinical practice and largely replicated the reported mechanisms. By categorizing iPSC-CMs into resistant and sensitive cell lines based on their time- and concentration-related phenotypic responses to DOX, we found that the sensitivity of donor-specific iPSC-CMs to DOX may predict in vivo DIC risk. Furthermore, we identified a differentially expressed gene, DND microRNA-mediated repression inhibitor 1 (DND1), between the DOX-resistant and DOX-sensitive iPSC-CMs. Our results support the utilization of donor-specific iPSC-CMs in assessing interindividual differences in DIC. Further studies will encompass a large panel of donor-specific iPSC-CMs to identify potential novel molecular and genetic biomarkers for predicting DOX and other oncology drug-induced cardiotoxicity.

Coronavirus disease 2019 (COVID-19) has widely varying clinical severity. Currently, no single marker or panel of markers is considered standard of care for prediction of COVID-19 disease progression. The goal of this study is to gain mechanistic insights at the molecular level and to discover predictive biomarkers of severity of infection and outcomes among COVID-19 patients. This cohort study (n = 76) included participants aged 16–78 years who tested positive for SARS-CoV-2 and enrolled in Memphis, TN between August 2020 to July 2022. Clinical outcomes were classified as Non-severe (n = 39) or Severe (n = 37). LC/HRMS-based untargeted metabolomics/lipidomics was conducted to examine the difference in plasma metabolome and lipidome between the two groups. Metabolomics data indicated that the kynurenine pathway was activated in Severe participants. Significant increases in short chain acylcarnitines, and short and medium chain acylcarnitines containing OH-FA chain in Severe vs. Non-severe group, which indicates that (1) the energy pathway switched to FA β-oxidation to maintain the host energy homeostasis and to provide energy for virus proliferation; (2) ROS status was aggravated in Severe vs. Non-severe group. Based on PLS-DA and correlation analysis to severity score, IL-6, and creatine, a biomarker panel containing glucose (pro-inflammation), ceramide and S1P (inflammation related), 4-hydroxybutyric acid (oxidative stress related), testosterone sulfate (immune related), and creatine (kidney function), was discovered. This novel biomarker panel plus IL-6 with an AUC of 0.945 provides a better indication of COVID-19 clinical outcomes than that of IL-6 alone or the three clinical biomarker panel (IL-6, glucose and creatine) with AUCs of 0.875 or 0.892.

Preclinical and clinical findings suggest sexual dimorphism in cardiotoxicity induced by a chemotherapeutic drug, doxorubicin (DOX). However, molecular alterations leading to sex-related differential vulnerability of heart to DOX toxicity are not fully explored. In the present study, RNA sequencing in hearts of B6C3F(1) mice indicated more differentially expressed genes in males than females (224 vs. 19; >= 1.5-fold, False Discovery Rate [FDR] < 0.05) at 1 week after receiving 24 mg/kg total cumulative DOX dose that induced cardiac lesions only in males. Pathway analysis further revealed probable inactivation of cardiac apelin fibroblast signaling pathway (p = 0.00004) only in DOX-treated male mice that showed >= 1.25-fold downregulation in the transcript and protein levels of the apelin receptor, APJ. In hearts of DOX-treated females, the transcript levels of apelin (1.24-fold) and APJ (1.47-fold) were significantly (p < 0.05) increased compared to saline-treated controls. Sex-related differential DOX effect was also observed on molecular targets downstream of the apelin-APJ pathway in cardiac fibroblasts and cardiomyocytes. In cardiac fibroblasts, upregulation of Tgf-beta 2, Ctgf, Sphk1, Serpine1, and Timp1 (fibrosis; FDR < 0.05) in DOX-treated males and upregulation of only Tgf-beta 2 and Timp1 (p < 0.05) in females suggested a greater DOX toxicity in hearts of males than females. Additionally, Ryr2 and Serca2 (calcium handling; FDR < 0.05) were downregulated in conjunction with 1.35-fold upregulation of Casp12 (sarcoplasmic reticulum-mediated apoptosis; FDR < 0.05) in DOX-treated male mice. Drug effect on the transcript level of these genes was less severe in female hearts. Collectively, these data suggest a likely role of the apelin-APJ axis in sex-related differential DOX-induced cardiotoxicity in our mouse model.

Subclinical cardiotoxicity at low total cumulative doxorubicin (DOX) doses can manifest into cardiomyopathy in long-term cancer survivors. However, the underlying mechanisms are poorly understood. In male B6C3F1 mice, assessment of cardiac function by echocardiography was performed at 1, 4, 10, 17, and 24 weeks after exposure to 6, 9, 12, and 24 mg/kg total cumulative DOX doses or saline (SAL) to monitor development of delayed-onset cardiotoxicity. The 6- or 9-mg/kg total cumulative doses resulted in a significant time-dependent decline in systolic function (left ventricular ejection fraction (LVEF) and fractional shortening (FS)) during the 24-week recovery although there was not a significant alteration in % LVEF or % FS at any specific time point during the recovery. A significant decline in systolic function was elicited by the cardiotoxic cumulative DOX dose (24 mg/kg) during the 4- to 24-week period after treatment compared to SAL-treated counterparts. At 24 weeks after DOX treatment, a significant dose-related decrease in the expression of genes and proteins involved in sarcoplasmic reticulum (SR) calcium homeostasis (Ryr2 and Serca2) was associated with a dose-related increase in the transcript level of Casp12 (SR-specific apoptosis) in hearts. These mice also showed enhanced apoptotic activity in hearts indicated by a significant dose-related elevation in the number of apoptotic cardiomyocytes compared to SAL-treated counterparts. These findings collectively suggest that a steady decline in SR calcium handling and apoptosis might be involved in the development of subclinical cardiotoxicity that can evolve into irreversible cardiomyopathy later in life.

Cardiotoxicity is a serious adverse effect of an anticancer drug, doxorubicin (DOX), which can occur within a year or decades after completion of therapy. The present study was designed to address a knowledge gap concerning a lack of circulating biomarkers capable of predicting the risk of cardiotoxicity induced by DOX. Profiling of 2083 microRNAs (miRNAs) in mouse plasma revealed 81 differentially expressed miRNAs 1 week after 6, 9, 12, 18, or 24 mg/kg total cumulative DOX doses (early-onset model) or saline (SAL). Among these, the expression of seven miRNAs was altered prior to the onset of myocardial injury at 12 mg/kg and higher cumulative doses. The expression of only miR-34a-5p was significantly (false discovery rate [FDR] < 0.1) elevated at all total cumulative doses compared with concurrent SAL-treated controls and showed a statistically significant dose-related response. The trend in plasma miR-34a-5p expression levels during DOX exposures also correlated with a significant dose-related increase in cardiac expression of miR-34a-5p in these mice. Administration of a cardioprotective drug, dexrazoxane, to mice before DOX treatment, significantly mitigated miR-34a-5p expression in both plasma and heart in conjunction with attenuation of cardiac pathology. This association between plasma and heart may suggest miR-34a-5p as a potential early circulating marker of early-onset DOX cardiotoxicity. In addition, higher expression of miR-34a-5p (FDR < 0.1) in plasma and heart compared with SAL-treated controls 24 weeks after 24 mg/kg total cumulative DOX dose, when cardiac function was altered in our recently established delayed-onset cardiotoxicity model, indicated its potential as an early biomarker of delayed-onset cardiotoxicity.

Omics methodologies are widely used in toxicological research to understand modes and mechanisms of toxicity. Increasingly, these methodologies are being applied to questions of regulatory interest such as molecular point-of-departure derivation and chemical grouping/read-across. Despite its value, widespread regulatory acceptance of omics data has not yet occurred. Barriers to the routine application of omics data in regulatory decision making have been: 1) lack of transparency for data processing methods used to convert raw data into an interpretable list of observations; and 2) lack of standardization in reporting to ensure that omics data, associated metadata and the methodologies used to generate results are available for review by stakeholders, including regulators. Thus, in 2017, the Organisation for Economic Co-operation and Development (OECD) Extended Advisory Group on Molecular Screening and Toxicogenomics (EAGMST) launched a project to develop guidance for the reporting of omics data aimed at fostering further regulatory use. Here, we report on the ongoing development of the first formal reporting framework describing the processing and analysis of both transcriptomic and metabolomic data for regulatory toxicology. We introduce the modular structure, content, harmonization and strategy for trialling this reporting framework prior to its publication by the OECD.

The imbalance of testosterone to estradiol ratio has been related to the development of prostate diseases. Although rat models of prostate diseases induced by endocrine-disrupting chemicals (EDCs) and/or hormone exposure are commonly used to analyze gene expression profiles in the prostate, most studies utilize a single endpoint. In this study, microarray analysis was used for gene expression profiling in rat prostate tissue after exposure to EDCs and sex hormones over multiple time points (prepubertal through adulthood). We used dorsolateral prostate tissues from Sprague-Dawley rats (male offspring) and postnatally administered estradiol benzoate (EB) on postnatal days (PNDs) 1, 3, and 5, followed by treatment with additional hormones [estradiol (E) and testosterone (T)] on PNDs 90-200, as described by Ho et al. Microarray analysis was performed for gene expression profiling in the dorsolateral prostate, and the results were validated via qRT-PCR. The genes in cytokine-cytokine receptor interaction, cell adhesion molecules, and chemokines were upregulated in the EB+T+E group on PNDs 145 and 200. Moreover, early-stage downregulation of anti-inflammatory gene: bone morphogenetic protein 7 gene was observed. These findings suggest that exposure to EB, T, and E activates multiple pathways and simultaneously downregulates anti-inflammatory genes. Interestingly, these genes are reportedly expressed in prostate cancer tissues/cell lines. Further studies are required to elucidate the mechanism, including analyses using human prostate tissues.

Safety assessments of new drug candidates are an important part of the drug development and approval process. Often, possible sex-associated susceptibilities are not adequately addressed, and better assessment tools are needed. We hypothesized that hepatic transcript profiles of cytochrome P450 (P450) enzymes can be used to predict sex-associated differences in drug metabolism and possible adverse events. Comprehensive hepatic transcript profiles were generated for F344 rats of both sexes at nine ages, from 2 weeks (preweaning) to 104 weeks (elderly). Large differences in the transcript profiles of 29 drug metabolizing enzymes and transporters were found between adult males and females (8-52 weeks). Using the Pharma-Pendium data base, 41 drugs were found to be metabolized by one or two P450 enzymes encoded by sexually dimorphic mRNAs and thus were candidates for evaluation of possible sexually dimorphic metabolism and/or toxicities. Suspension cultures of primary hepatocytes from three male and three female adult rats (10-13 weeks old) were used to evaluate the metabolism of 11 drugs predicted to have sexually dimorphic metabolism. The pharmacokinetics of the drug or its metabolite was analyzed by liquid chromatography/tandem mass spectrometry using multiple reaction monitoring. Of those drugs with adequate metabolism, the predicted significant sex-different metabolism was found for six of seven drugs, with half-lives 37%-400% longer in female hepatocytes than in male hepatocytes. Thus, in this rat model, transcript profiles may allow identification of potential sex-related differences in drug metabolism. SIGNIFICANCE STATEMENT The present study showed that sex-different expression of genes coding for drug metabolizing enzymes, specifically cytochrome P450s, could be used to predict sex-different drug metabolism and, thus, provide a new tool for protecting susceptible subpopulations from possible adverse drug events.

Sulforaphane (SFN) has been shown to protect the brain vascular system and effectively reduce ischemic injuries and cognitive deficits. Given the robust cerebrovascular protection afforded by SFN, the objective of this study was to profile these effects in vitro using primary mouse brain microvascular endothelial cells and focusing on cellular redox, metabolism and detoxification functions. We used a mouse MitoChip array developed and validated at the FDA National Center for Toxicological Research (NCTR) to profile a host of genes encoded by nuclear and mt-DNA following SFN treatment (0–5 µM). Corresponding protein expression levels were assessed (ad hoc) by qRT-PCR, immunoblots and immunocytochemistry (ICC). Gene ontology clustering revealed that SFN treatment (24 h) significantly up-regulated ~50 key genes (>1.5 fold, adjusted p < 0.0001) and repressed 20 genes (<0.7 fold, adjusted p < 0.0001) belonging to oxidative stress, phase 1 & 2 drug metabolism enzymes (glutathione system), iron transporters, glycolysis, oxidative phosphorylation (OXPHOS), amino acid metabolism, lipid metabolism and mitochondrial biogenesis. Our results show that SFN stimulated the production of ATP by promoting the expression and activity of glucose transporter-1, and glycolysis. In addition, SFN upregulated anti-oxidative stress responses, redox signaling and phase 2 drug metabolism/detoxification functions, thus elucidating further the previously observed neurovascular protective effects of this compound.

2-hydroxy-4-methoxybenzophenone (HMB) is an ultraviolet light-absorbing compound that is used in sunscreens, cosmetics and plastics. HMB has been reported to have weak estrogenic activity by in vivo and in vitro studies, making it a chemical with potential reproductive concern. To explore if prenatal and lactational HMB exposure alters gene expression profiles of the developing reproductive organs, we performed microarray analysis using the prostate and testis of postnatal day (PND) 30 male Sprague-Dawley rats offspring exposed to 0, 3000, or 30,000 ppm of HMB from gestational day 6 through PND 21. Gene expression profiles of the prostate and testis were differentially affected by HMB dose with significant alterations observed at the 30,000 ppm HMB group. Tissue-specific gene expression was also identified. These genes, whose expression was altered by HMB exposure, may be considered as candidate biomarker(s) for testicular or prostatic toxicity; however, further studies are necessary to explore this potential.

Cardiotoxicity is a serious adverse effect of doxorubicin (DOX) treatment in cancer patients. Currently, there is a lack of sensitive biomarkers to predict the risk of DOX-induced cardiotoxicity. Using SOMAmer-based proteomic technology, 1129 proteins were profiled to identify potential early biomarkers of cardiotoxicity in plasma from male B6C3F1 mice given a weekly intravenous dose of 3 mg/kg DOX or saline (SAL) for 2, 3, 4, 6, or 8 weeks (6, 9, 12, 18, or 24 mg/kg cumulative DOX doses, respectively). Also, a group of mice received the cardio-protectant, dexrazoxane (DXZ; 60 mg/kg; intraperitoneal) 30 min before a weekly DOX or SAL dose. Proteomic analysis in plasma collected a week after the last dose showed a significant ≥1.2-fold change in level of 18 proteins in DOX-treated mice compared to SAL-treated counterparts during 8-week exposure. Of these, neurogenic locus notch homolog protein 1 (NOTCH1), von Willebrand factor (vWF), mitochondrial glutamate carrier 2, Wnt inhibitory factor 1, legumain, and mannan-binding lectin serine protease 1 were increased in plasma at 6 mg/kg cumulative DOX dose, prior to the release of myocardial injury marker, cardiac troponin I at 12 mg/kg and higher cumulative doses. These six proteins also remained significantly elevated following myocardial injury or pathology at 24 mg/kg. Pretreatment of mice with DXZ significantly attenuated DOX-induced elevated levels of only NOTCH1 and vWF with mitigation of cardiotoxicity. This suggests NOTCH1 and vWF as candidate early biomarkers of DOX cardiotoxicity, which may help in addressing a clinically important question of identifying cancer patients at risk for cardiotoxicity.

Stably Expressed Genes (SEGs) whose expression varies within a narrow range may be involved in core cellular processes necessary for basic functions. To identify such genes, we re-analyzed existing RNA-Seq gene expression profiles across 11 organs at 4 developmental stages (from immature to old age) in both sexes of F344 rats (n = 4/group; 320 samples). Expression changes (calculated as the maximum expression / minimum expression for each gene) of >19000 genes across organs, ages, and sexes ranged from 2.35 to >109-fold, with a median of 165-fold. The expression of 278 SEGs was found to vary ≤4-fold and these genes were significantly involved in protein catabolism (proteasome and ubiquitination), RNA transport, protein processing, and the spliceosome. Such stability of expression was further validated in human samples where the expression variability of the homologous human SEGs was significantly lower than that of other genes in the human genome. It was also found that the homologous human SEGs were generally less subject to non-synonymous mutation than other genes, as would be expected of stably expressed genes. We also found that knockout of SEG homologs in mouse models was more likely to cause complete preweaning lethality than non-SEG homologs, corroborating the fundamental roles played by SEGs in biological development. Such stably expressed genes and pathways across life-stages suggest that tight control of these processes is important in basic cellular functions and that perturbation by endogenous (e.g., genetics) or exogenous agents (e.g., drugs, environmental factors) may cause serious adverse effects.

Physiological factors such as age and sex have been shown to be risk factors for adverse effects in the liver, including liver diseases and drug-induced liver injury. Previously, we have reported age- and sex-related significant differences in hepatic basal gene expression in rats during the life span that may be related to susceptibility to such adverse effects. However, the underlying mechanisms of the gene expression changes were not fully understood. In recent years, increasing evidence for epigenetic mechanisms of gene regulation has fueled interest in the role of microRNAs (miRNAs) in toxicogenomics and biomarker discovery. We therefore proposed that significant age and sex differences exist in baseline liver miRNA expression, and that comprehensive profiling of miRNAs will provide insights into the epigenetic regulation of gene expression in rat liver.

Identification of early biomarkers of cardiotoxicity could help initiate means to ameliorate the cardiotoxic actions of clinically useful drugs such as doxorubicin (DOX). Since DOX has been shown to target mitochondria, transcriptional levels of mitochondria-related genes were evaluated to identify early candidate biomarkers in hearts of male B6C3F1 mice given a weekly intravenous dose of 3mg/kg DOX or saline (SAL) for 2, 3, 4, 6, or 8 weeks (6, 9, 12, 18, or 24 mg/kg cumulative DOX doses, respectively). Also, a group of mice was pretreated (intraperitoneally) with the cardio-protectant, dexrazoxane (DXZ; 60 mg/kg) 30 min before each weekly dose of DOX or SAL. At necropsy a week after the last dose, increased plasma concentrations of cardiac troponin T (cTnT) were detected at 18 and 24 mg/kg cumulative DOX doses, whereas myocardial alterations were observed only at the 24 mg/kg dose. Of 1019 genes interrogated, 185, 109, 140, 184, and 451 genes were differentially expressed at 6, 9, 12, 18, and 24 mg/kg cumulative DOX doses, respectively, compared to concurrent SAL-treated controls. Of these, expression of 61 genes associated with energy metabolism and apoptosis was significantly altered before and after occurrence of myocardial injury, suggesting these as early genomics markers of cardiotoxicity. Much of these DOX-induced transcriptional changes were attenuated by pretreatment of mice with DXZ. Also, DXZ treatment significantly reduced plasma cTnT concentration and completely ameliorated cardiac alterations induced by 24 mg/kg cumulative DOX. This information on early transcriptional changes during DOX treatment may be useful in designing cardioprotective strategies targeting mitochondria.

The present study aimed to identify molecular markers of early stages of cardiotoxicity induced by a potent chemotherapeutic agent, doxorubicin (DOX). Male B6C3F(1) mice were dosed with 3mgkg(-1) DOX or saline via tail vein weekly for 2, 3, 4, 6 or 8weeks (cumulative DOX doses of 6, 9, 12, 18 or 24mgkg(-1), respectively) and euthanized a week after the last dose. Mass spectrometry-based and nuclear magnetic resonance spectrometry-based metabolic profiling were employed to identify initial biomarkers of cardiotoxicity before myocardial injury and cardiac pathology, which were not noted until after the 18 and 24mgkg(-1) cumulative doses, respectively. After a cumulative dose of 6mgkg(-1), 18 amino acids and four biogenic amines (acetylornithine, kynurenine, putrescine and serotonin) were significantly increased in cardiac tissue; 16 amino acids and two biogenic amines (acetylornithine and hydroxyproline) were significantly altered in plasma. In addition, 16 acylcarnitines were significantly increased in plasma and five were significantly decreased in cardiac tissue compared to saline-treated controls. Plasma lactate and succinate, involved in the Krebs cycle, were significantly altered after a cumulative dose of 6mgkg(-1). A few metabolites remained altered at higher cumulative DOX doses, which could partly indicate a transition from injury processes at 2weeks to repair processes with additional injury happening concurrently before myocardial injury at 8weeks. These altered metabolic profiles in mouse heart and plasma during the initial stages of injury progression due to DOX treatment may suggest these metabolites as candidate early biomarkers of cardiotoxicity. Published 2016. This article is a U.S. Government work and is in the public domain in the USAMetabolomics analyses of plasma and heart tissue from male B6C3F(1) mice dosed weekly with 3mgkg(-1) doxorubicin or saline for 2, 3, 4, 6 or 8weeks (cumulative doses of 6, 9, 12, 18 and 24mgkg(-1), respectively) identified potential early stage injury biomarkers of doxorubicin-induced cardiotoxicity. Select metabolites were altered after the 6mgkg(-1) cumulative dose whereas myocardial injury and cardiac pathology were not noted until after the 18 and 24mgkg(-1) cumulative doses, respectively.

Sex is a risk factor for development of cardiotoxicity, induced by the anti-cancer drug, doxorubicin (DOX), in humans. To explore potential mechanisms underlying differential susceptibility to DOX between sexes, 8-week old male and female B6C3F1 mice were dosed with 3mg/kg body weight DOX or an equivalent volume of saline via tail vein once a week for 6, 7, 8, and 9 consecutive weeks, resulting in 18, 21, 24, and 27mg/kg cumulative DOX doses, respectively. At necropsy, one week after each consecutive final dose, the extent of myocardial injury was greater in male mice compared to females as indicated by higher plasma concentrations of cardiac troponin T at all cumulative DOX doses with statistically significant differences between sexes at the 21 and 24mg/kg cumulative doses. A greater susceptibility to DOX in male mice was further confirmed by the presence of cytoplasmic vacuolization in cardiomyocytes, with left atrium being more vulnerable to DOX cardiotoxicity. The number of TUNEL-positive cardiomyocytes was mostly higher in DOX-treated male mice compared to female counterparts, showing a statistically significant sex-related difference only in left atrium at 21mg/kg cumulative dose. DOX-treated male mice also had an increased number of γ-H2A.X-positive (measure of DNA double-strand breaks) cardiomyocytes compared to female counterparts with a significant sex effect in the ventricle at 27mg/kg cumulative dose and right atrium at 21 and 27mg/kg cumulative doses. This newly established mouse model provides a means to identify biomarkers and access potential mechanisms underlying sex-related differences in DOX-induced cardiotoxicity.

Cardiovascular disease (CVD) is the leading cause of mortality worldwide. Moreover, sex and age are considered major risk factors in the development of CVDs. Mitochondria are vital for normal cardiac function, and regulation of mitochondrial structure and function may impact susceptibility to CVD. To identify potential role of mitochondria in sex-related differences in susceptibility to CVD, we analyzed the basal expression levels of mitochondria-related genes in the hearts of male and female rats. Whole genome expression profiling was performed in the hearts of young (8-week), adult (21-week), and old (78-week) male and female Fischer 344 rats and the expression of 670 unique genes related to various mitochondrial functions was analyzed. A significant (p<0.05) sexual dimorphism in expression levels of 46, 114, and 41 genes was observed in young, adult and old rats, respectively. Gene Ontology analysis revealed the influence of sex on various biological pathways related to cardiac energy metabolism at different ages. The expression of genes involved in fatty acid metabolism was significantly different between the sexes in young and adult rat hearts. Adult male rats also showed higher expression of genes associated with the pyruvate dehydrogenase complex compared to females. In young and adult hearts, sexual dimorphism was not noted in genes encoding oxidative phosphorylation. In old rats, however, a majority of genes involved in oxidative phosphorylation had higher expression in females compared to males. Such basal differences between the sexes in cardiac expression of genes associated with energy metabolism may indicate a likely involvement of mitochondria in susceptibility to CVDs. In addition, female rats showed lower expression levels of apoptotic genes in hearts compared to males at all ages, which may have implications for better preservation of cardiac mass in females than in males.