Behavioral and Transcriptome Profiling of Heterozygous Rab10 Knock-Out Mice.

A central question in the field of aging research is to identify the cellular and molecular basis of neuroresilience. One potential candidate is the small GTPase, Rab10. Here, we used Rab10+/- mice to investigate the molecular mechanisms underlying Rab10-mediated neuroresilience. Brain expression analysis of 880 genes involved in neurodegeneration showed that Rab10+/- mice have increased activation of pathways associated with neuronal metabolism, structural integrity, neurotransmission, and neuroplasticity compared with their Rab10+/+ littermates. Lower activation was observed for pathways involved in neuroinflammation and aging. We identified and validated several differentially expressed genes (DEGs), including Stx2, Stx1b, Vegfa, and Lrrc25 (downregulated) and Prkaa2, Syt4, and Grin2d (upregulated). Behavioral testing showed that Rab10+/- mice perform better in a hippocampal-dependent spatial task (object in place test), while their performance in a classical conditioning task (trace eyeblink classical conditioning, TECC) was significantly impaired. Therefore, our findings indicate that Rab10 differentially controls the brain circuitry of hippocampal-dependent spatial memory and higher-order behavior that requires intact cortex-hippocampal circuitry. Transcriptome and biochemical characterization of these mice suggest that glutamate ionotropic receptor NMDA type subunit 2D (GRIN2D or GluN2D) is affected by Rab10 signaling. Further work is needed to evaluate whether GRIN2D mediates the behavioral phenotypes of the Rab10+/- mice. We conclude that Rab10+/- mice described here can be a valuable tool to study the mechanisms of resilience in Alzheimer's disease (AD) model mice and to identify novel therapeutical targets to prevent cognitive decline associated with normal and pathologic aging.

Exercise increases NPY/AgRP and TH neuron activity in the hypothalamus of female mice.

Recent evidence identifies a potent role for aerobic exercise to modulate the activity of hypothalamic neurons related to appetite; however, these studies have been primarily performed in male rodents. Since females have markedly different neuronal mechanisms regulating food intake, the current study aimed to determine the effects of acute treadmill exercise on hypothalamic neuron populations involved in regulating appetite in female mice. Mature, untrained female mice were exposed to acute sedentary, low- (10 m/min), moderate- (14 m/min), and high (18 m/min)-intensity treadmill exercise in a randomized crossover design. Mice were fasted 10 h before exercise, and food intake was monitored for 48 h after bouts. Immunohistochemical detection of cFOS was performed 3 h post-exercise to determine the changes in hypothalamic neuropeptide Y (NPY)/agouti-related peptide (AgRP), pro-opiomelanocortin (POMC), tyrosine hydroxylase (TH), and SIM1-expressing neuron activity concurrent with the changes in food intake. Additionally, stains for pSTAT3tyr705 and pERKthr202/tyr204 were performed to detect exercise-mediated changes in intracellular signaling. Briefly, moderate- and high-intensity exercises increased 24-h food intake by 5.9 and 19%, respectively, while low-intensity exercise had no effects. Furthermore, increases in NPY/AgRPARC, SIM1PVN, and TH neuron activity were observed 3 h after high-intensity exercise, with no effects on POMCARC neurons. While no effects of exercise on pERKthr202/tyr204 were observed, pSTAT3tyr705 was elevated specifically in NPY/AgRP neurons 3 h post-exercise. Overall, aerobic exercise increased the activity of several appetite-stimulating neuron populations in the hypothalamus of female mice, which may provide insight into previously reported sexual dimorphisms in post-exercise feeding.

Intrinsic adaptations in OXPHOS power output and reduced tumorigenicity characterize doxorubicin resistant ovarian cancer cells.

Although the development of chemoresistance is multifactorial, active chemotherapeutic efflux driven by upregulations in ATP binding cassette (ABC) transporters are commonplace. Chemotherapeutic efflux pumps, like ABCB1, couple drug efflux to ATP hydrolysis and thus potentially elevate cellular demand for ATP resynthesis. Elevations in both mitochondrial content and cellular respiration are common phenotypes accompanying many models of cancer cell chemoresistance, including those dependent on ABCB1. The present study set out to characterize potential mitochondrial remodeling commensurate with ABCB1-dependent chemoresistance, as well as investigate the impact of ABCB1 activity on mitochondrial respiratory kinetics. To do this, comprehensive bioenergetic phenotyping was performed across ABCB1-dependent chemoresistant cell models and compared to chemosensitive controls. In doxorubicin (DOX) resistant ovarian cancer cells, the combination of both increased mitochondrial content and enhanced respiratory complex I (CI) boosted intrinsic oxidative phosphorylation (OXPHOS) power output. With respect to ABCB1, acute ABCB1 inhibition partially normalized intact basal mitochondrial respiration between chemosensitive and chemoresistant cells, suggesting that active ABCB1 contributes to mitochondrial remodeling in favor of enhanced OXPHOS. Interestingly, while enhanced OXPHOS power output supported ABCB1 drug efflux when DOX was present, in the absence of chemotherapeutic stress, enhanced OXPHOS power output was associated with reduced tumorigenicity.

Circulating α-klotho regulates metabolism via distinct central and peripheral mechanisms.

Emerging evidence implicates the circulating α-klotho protein as a prominent regulator of energy balance and substrate metabolism, with diverse, tissue-specific functions. Despite its well-documented ubiquitous role inhibiting insulin signaling, α-klotho elicits potent antidiabetic and anti-obesogenic effects. α-Klotho facilitates insulin release and promotes ß cell health in the pancreas, stimulates lipid oxidation in liver and adipose tissue, attenuates hepatic gluconeogenesis, and increases whole-body energy expenditure. The mechanisms underlying α-klotho's peripheral functions are multifaceted, including hydrolyzing transient receptor potential channels, stimulating integrin ß1➔focal adhesion kinase signaling, and activating PPARα via inhibition of insulin-like growth factor receptor 1. Moreover, until recently, potential metabolic roles of α-klotho in the central nervous system remained unexplored; however, a novel α-klotho➔fibroblast growth factor receptor➔PI3kinase signaling axis in the arcuate nucleus of the hypothalamus has been identified as a critical regulator of energy balance and glucose metabolism. Overall, the role of circulating α-klotho in the regulation of metabolism is a new focus of research, but accumulating evidence identifies this protein as an encouraging therapeutic target for Type 1 and 2 Diabetes and obesity. This review analyzes the new literature investigating α-klotho-mediated regulation of metabolism and proposes impactful future directions to progress our understanding of this complex metabolic protein.

Energy Status Differentially Modifies Feeding Behavior and POMCARC Neuron Activity After Acute Treadmill Exercise in Untrained Mice.

Emerging evidence identifies a potent role for aerobic exercise to modulate activity of neurons involved in regulating appetite; however, these studies produce conflicting results. These discrepancies may be, in part, due to methodological differences, including differences in exercise intensity and pre-exercise energy status. Consequently, the current study utilized a translational, well-controlled, within-subject, treadmill exercise protocol to investigate the differential effects of energy status and exercise intensity on post-exercise feeding behavior and appetite-controlling neurons in the hypothalamus. Mature, untrained male mice were exposed to acute sedentary, low (10m/min), moderate (14m/min), and high (18m/min) intensity treadmill exercise in a randomized crossover design. Fed and 10-hour-fasted mice were used, and food intake was monitored 48h. post-exercise. Immunohistochemical detection of cFOS was performed 1-hour post-exercise to determine changes in hypothalamic NPY/AgRP, POMC, tyrosine hydroxylase, and SIM1-expressing neuron activity concurrent with changes in food intake. Additionally, stains for pSTAT3tyr705 and pERKthr202/tyr204 were performed to detect exercise-mediated changes in intracellular signaling. Results demonstrated that fasted high intensity exercise suppressed food intake compared to sedentary trials, which was concurrent with increased anorexigenic POMC neuron activity. Conversely, fed mice experienced augmented post-exercise food intake, with no effects on POMC neuron activity. Regardless of pre-exercise energy status, tyrosine hydroxylase and SIM1 neuron activity in the paraventricular nucleus was elevated, as well as NPY/AgRP neuron activity in the arcuate nucleus. Notably, these neuronal changes were independent from changes in pSTAT3tyr705 and pERKthr202/tyr204 signaling. Overall, these results suggest fasted high intensity exercise may be beneficial for suppressing food intake, possibly due to hypothalamic POMC neuron excitation. Furthermore, this study identifies a novel role for pre-exercise energy status to differentially modify post-exercise feeding behavior and hypothalamic neuron activity, which may explain the inconsistent results from studies investigating exercise as a weight loss intervention.

Predictive Accuracy of the Nelson Equation via BodPod Compared to Commonly Used Equations to Estimate Resting Metabolic Rate in Adults.

Indirect calorimetry (IC) is considered the gold standard for assessing resting metabolic rate (RMR). However, many people do not have access to IC devices and use prediction equations for RMR estimation. Equations using fat free mass (FFM) as a predictor have been developed to estimate RMR, as a strong relationship exists between FFM and RMR. One such equation is the Nelson equation which is used by the BodPod (BP). Yet, there is limited evidence whether the Nelson equation is superior to other common equations to predict RMR. To examine the agreement between predicted RMR from common RMR equations and the BP, and RMR measured via IC. Data from 48 healthy volunteers who completed both the BP and IC were collected. Agreement between RMR measured by BP, common regression equations, and indirect caloriometry was evaluated using repeated measures ANOVA, Bland-Altman analysis and root mean square error (RMSE). Predicted RMR values from common equations and BP were significantly different from IC with the exception of the World Health Organization (WHO) equation. Large limits of agreement and RMSE values demonstrate a large amount of error at the individual level. Despite the use of FFM, the Nelson equation does not appear to be superior to other common RMR equations. Although the WHO equation presented the best option within our sample, all equations performed poorly at the individual level. Clinicians should be aware that prediction equations may significantly under- or overestimate RMR compared to IC and when an accurate value of RMR is required, IC is recommended.

Centrally circulating α-klotho inversely correlates with human obesity and modulates arcuate cell populations in mice.

OBJECTIVE: Our laboratory recently identified the centrally circulating α-klotho protein as a novel hypothalamic regulator of food intake and glucose metabolism in mice. The current study aimed to investigate novel molecular effectors of central α-klotho in the arcuate nucleus of the hypothalamus (ARC), while further deciphering its role regulating energy balance in both humans and mice. METHODS: Cerebrospinal fluid (CSF) was collected from 22 adults undergoing lower limb orthopedic surgeries, and correlations between body weight and α-klotho were determined using an α-klotho enzyme-linked immunosorbent assay (ELISA) kit. To investigate the effects of α-klotho on energy expenditure (EE), 2-day intracerebroventricular (ICV) treatment was performed in diet-induced obesity (DIO) mice housed in TSE Phenomaster indirect calorimetry metabolic cages. Immunohistochemical staining for cFOS and patch clamp electrophysiology were used to determine the effects of central α-klotho on proopiomelanocortin (POMC) and tyrosine hydroxylase (TH) neurons. Additional stainings were performed to determine novel roles for central α-klotho to regulate non-neuronal cell populations in the ARC. Lastly, ICV pretreatment with fibroblast growth factor receptor (FGFR) or PI3kinase inhibitors was performed to determine the intracellular signaling involved in α-klotho-mediated regulation of ARC nuclei. RESULTS: Obese/overweight human subjects had significantly lower CSF α-klotho concentrations compared to lean counterparts (1,044 ± 251 vs. 1616 ± 218 pmol/L, respectively). Additionally, 2 days of ICV α-klotho treatment increased EE in DIO mice. α-Klotho had no effects on TH neuron activity but elicited varied responses in POMC neurons, with 44% experiencing excitatory and 56% experiencing inhibitory effects. Inhibitor experiments identified an α-klotho→FGFR→PI3kinase signaling mechanism in the regulation of ARC POMC and NPY/AgRP neurons. Acute ICV α-klotho treatment also increased phosphorylated ERK in ARC astrocytes via FGFR signaling. CONCLUSION: Our human CSF data provide the first evidence that impaired central α-klotho function may be involved in the pathophysiology of obesity. Furthermore, results in mouse models identify ARC POMC neurons and astrocytes as novel molecular effectors of central α-klotho. Overall, the current study highlights prominent roles of α-klotho→FGFR→PI3kinase signaling in the homeostatic regulation of ARC neurons and whole-body energy balance.

Tissue-Specific Approaches Reveal Diverse Metabolic Functions of Rho-Kinase 1.

Rho-kinase 1 (ROCK1) has been implicated in diverse metabolic functions throughout the body, with promising evidence identifying ROCK1 as a therapeutic target in diabetes and obesity. Considering these metabolic roles, several pharmacological inhibitors have been developed to elucidate the mechanisms underlying ROCK1 function. Y27632 and fasudil are two common ROCK1 inhibitors; however, they have varying non-specific selectivity to inhibit other AGC kinase subfamily members and whole-body pharmacological approaches lack tissue-specific insight. As a result, interpretation of studies with these inhibitors is difficult, and alternative approaches are needed to elucidate ROCK1's tissue specific metabolic functions. Fortunately, recent technological advances utilizing molecular carriers or genetic manipulation have facilitated discovery of ROCK1's tissue-specific mechanisms of action. In this article, we review the tissue-specific roles of ROCK1 in the regulation of energy balance and substrate utilization. We highlight prominent metabolic roles in liver, adipose, and skeletal muscle, in which ROCK1 regulates energy expenditure, glucose uptake, and lipid metabolism via inhibition of AMPK2α and paradoxical modulation of insulin signaling. Compared to ROCK1's roles in peripheral tissues, we also describe contradictory functions of ROCK1 in the hypothalamus to increase energy expenditure and decrease food intake via leptin signaling. Furthermore, dysregulated ROCK1 activity in either of these tissues results in metabolic disease phenotypes. Overall, tissue-specific approaches have made great strides in deciphering the many critical metabolic functions of ROCK1 and, ultimately, may facilitate the development of novel treatments for metabolic disorders.

Accuracy of Commonly Used Age-Predicted Maximal Heart Rate Equations.

Age-predicted maximal heart rate (APMHR) is an essential measure for healthcare professionals in determining cardiovascular response to exercise testing, exertion, and prescription. Although multiple APMHR prediction equations have been validated for specific populations, the accuracy of each within a general population requires testing. We aimed to determine which APMHR equation (Fox, Gellish, Gulati, Tanaka, Arena, Astrand, Nes, Fairbarn) most accurately predicts max heart rate (HRmax) in a general population. HRmax from 99 graded treadmill exercise tests (GXT) were measured. GXTs ended upon volitional fatigue and were only included for analysis if RER > 1.10. Individual paired t-test were performed to determine if significant differences existed between measured and predicted HRmax, along with root mean square errors for each equation. Bland-Altman plots were constructed to determine agreement between equations and measured HRmax. Significant differences between measured and predicted HRmax were found for the Gulati, Astrand, Nes, and Fairbarn (male) equations (p < 0.05). Bland-Altman plots revealed wide limits of agreement for all nine APMHR equations, suggesting poor agreement between measured and predicted HRmax. Proportional bias indicates that prediction equations under and overestimated HRmax in individuals with lower and higher measured HRmax, respectively, with the exception of the Fox equation. All equations used in this study show poor agreement between measured HRmax and APMHR. The Fox equation may represent the best option for a general population as it is less likely to under or overestimate based on individual HRmax. Individuals should use data from GXTs to determine HRmax when applicable to ensure accuracy.