Non-transgenic guinea pig strains exhibit divergent age-related changes in hippocampal mitochondrial respiration.

AIM: Alzheimer's disease (AD) is the most common form of dementia. However, while 150+ animal models of AD exist, drug translation from preclinical models to humans for treatment usually fails. One factor contributing to low translation is likely the absence of neurodegenerative models that also encompass the multi-morbidities of human aging. We previously demonstrated that, in comparison to the PigmEnTed (PET) guinea pig strain which models "typical" brain aging, the Hartley strain develops hallmarks of AD like aging humans. Hartleys also exhibit age-related impairments in cartilage and skeletal muscle. Impaired mitochondrial respiration is one driver of both cellular aging and AD. In humans with cognitive decline, diminished skeletal muscle and brain respiratory control occurs in parallel. We previously reported age-related declines in skeletal muscle mitochondrial respiration in Hartleys. It is unknown if there is concomitant mitochondrial dysfunction in the brain. METHODS: Therefore, we assessed hippocampal mitochondrial respiration in 5- and 12-month Hartley and PET guinea pigs using high-resolution respirometry. RESULTS: At 12 months, PETs had higher complex I supported mitochondrial respiration paralleling their increase in body mass compared to 5 months PETs. Hartleys were also heavier at 12 months compared to 5 months but did not have higher complex I respiration. Compared to 5 months Hartleys, 12 months Hartleys had lower complex I mitochondrial efficiency and compensatory increases in mitochondrial proteins collectively suggesting mitochondrial dysfunction with age. CONCLUSIONS: Therefore, Hartleys might be a relevant model to test promising therapies targeting mitochondria to slow brain aging and AD progression.

Sedentary behavior in mice induces metabolic inflexibility by suppressing skeletal muscle pyruvate metabolism.

Carbohydrates and lipids provide the majority of substrates to fuel mitochondrial oxidative phosphorylation. Metabolic inflexibility, defined as an impaired ability to switch between these fuels, is implicated in a number of metabolic diseases. Here, we explore the mechanism by which physical inactivity promotes metabolic inflexibility in skeletal muscle. We developed a mouse model of sedentariness, small mouse cage (SMC), that, unlike other classic models of disuse in mice, faithfully recapitulated metabolic responses that occur in humans. Bioenergetic phenotyping of skeletal muscle mitochondria displayed metabolic inflexibility induced by physical inactivity, demonstrated by a reduction in pyruvate-stimulated respiration (JO2) in the absence of a change in palmitate-stimulated JO2. Pyruvate resistance in these mitochondria was likely driven by a decrease in phosphatidylethanolamine (PE) abundance in the mitochondrial membrane. Reduction in mitochondrial PE by heterozygous deletion of phosphatidylserine decarboxylase (PSD) was sufficient to induce metabolic inflexibility measured at the whole-body level, as well as at the level of skeletal muscle mitochondria. Low mitochondrial PE in C2C12 myotubes was sufficient to increase glucose flux toward lactate. We further implicate that resistance to pyruvate metabolism is due to attenuated mitochondrial entry via mitochondrial pyruvate carrier (MPC). These findings suggest a mechanism by which mitochondrial PE directly regulates MPC activity to modulate metabolic flexibility in mice.

A practical perspective on how to develop, implement, execute, and reproduce high-resolution respirometry experiments: The physiologist's guide to an Oroboros O2k.

The development of high-resolution respirometry (HRR) has greatly expanded the analytical scope to study mitochondrial respiratory control relative to specific tissue/cell types across various metabolic states. Specifically, the Oroboros Oxygraph 2000 (O2k) is a common tool for measuring rates of mitochondrial respiration and is the focus of this perspective. The O2k platform is amenable for answering numerous bioenergetic questions. However, inherent variability with HRR-derived data, both within and amongst users, can impede progress in bioenergetics research. Therefore, we advocate for several vital considerations when planning and conducting O2k experiments to ultimately enhance transparency and reproducibility across laboratories. In this perspective, we offer guidance for best practices of mitochondrial preparation, protocol selection, and measures to increase reproducibility. The goal of this perspective is to propagate the use of the O2k, enhance reliability and validity for both new and experienced O2k users, and provide a reference for peer reviewers.

Cardiac gene therapy treats diabetic cardiomyopathy and lowers blood glucose.

Diabetic cardiomyopathy, an increasingly global epidemic and a major cause of heart failure with preserved ejection fraction (HFpEF), is associated with hyperglycemia, insulin resistance, and intracardiomyocyte calcium mishandling. Here we identify that, in db/db mice with type 2 diabetes-induced HFpEF, abnormal remodeling of cardiomyocyte transverse-tubule microdomains occurs with downregulation of the membrane scaffolding protein cardiac bridging integrator 1 (cBIN1). Transduction of cBIN1 by AAV9 gene therapy can restore transverse-tubule microdomains to normalize intracellular distribution of calcium-handling proteins and, surprisingly, glucose transporter 4 (GLUT4). Cardiac proteomics revealed that AAV9-cBIN1 normalized components of calcium handling and GLUT4 translocation machineries. Functional studies further identified that AAV9-cBIN1 normalized insulin-dependent glucose uptake in diabetic cardiomyocytes. Phenotypically, AAV9-cBIN1 rescued cardiac lusitropy, improved exercise intolerance, and ameliorated hyperglycemia in diabetic mice. Restoration of transverse-tubule microdomains can improve cardiac function in the setting of diabetic cardiomyopathy and can also improve systemic glycemic control.

Phytochemical compound PB125 attenuates skeletal muscle mitochondrial dysfunction and impaired proteostasis in a model of musculoskeletal decline.

Impaired mitochondrial function and disrupted proteostasis contribute to musculoskeletal dysfunction. However, few interventions simultaneously target these two drivers to prevent musculoskeletal decline. Nuclear factor erythroid 2-related factor 2 (Nrf2) activates a transcriptional programme promoting cytoprotection, metabolism, and proteostasis. We hypothesized daily treatment with a purported Nrf2 activator, PB125, in Hartley guinea pigs, a model of musculoskeletal decline, would attenuate the progression of skeletal muscle mitochondrial dysfunction and impaired proteostasis and preserve musculoskeletal function. We treated 2- and 5-month-old male and female Hartley guinea pigs for 3 and 10 months, respectively, with the phytochemical compound PB125. Longitudinal assessments of voluntary mobility were measured using Any-MazeTM open-field enclosure monitoring. Cumulative skeletal muscle protein synthesis rates were measured using deuterium oxide over the final 30 days of treatment. Mitochondrial oxygen consumption in soleus muscles was measured using high resolution respirometry. In both sexes, PB125 (1) increased electron transfer system capacity; (2) attenuated the disease/age-related decline in coupled and uncoupled mitochondrial respiration; and (3) attenuated declines in protein synthesis in the myofibrillar, mitochondrial and cytosolic subfractions of the soleus. These effects were not associated with statistically significant prolonged maintenance of voluntary mobility in guinea pigs. Collectively, treatment with PB125 contributed to maintenance of skeletal muscle mitochondrial respiration and proteostasis in a pre-clinical model of musculoskeletal decline. Further investigation is necessary to determine if these documented effects of PB125 are also accompanied by slowed progression of other aspects of musculoskeletal dysfunction. KEY POINTS: Aside from exercise, there are no effective interventions for musculoskeletal decline, which begins in the fifth decade of life and contributes to disability and cardiometabolic diseases. Targeting both mitochondrial dysfunction and impaired protein homeostasis (proteostasis), which contribute to the age and disease process, may mitigate the progressive decline in overall musculoskeletal function (e.g. gait, strength). A potential intervention to target disease drivers is to stimulate nuclear factor erythroid 2-related factor 2 (Nrf2) activation, which leads to the transcription of genes responsible for redox homeostasis, proteome maintenance and mitochondrial energetics. Here, we tested a purported phytochemical Nrf2 activator, PB125, to improve mitochondrial function and proteostasis in male and female Hartley guinea pigs, which are a model for musculoskeletal ageing. PB125 improved mitochondrial respiration and attenuated disease- and age-related declines in skeletal muscle protein synthesis, a component of proteostasis, in both male and female Hartley guinea pigs.

Nontransgenic Guinea Pig Strains Exhibit Hallmarks of Human Brain Aging and Alzheimer's Disease.

Older age is the primary risk factor for most chronic diseases, including Alzheimer's disease (AD). Current preclinical models to study brain aging and AD are mainly transgenic and harbor mutations intended to mirror brain pathologies associated with human brain aging/AD (eg, by increasing production of the amyloid precursor protein, amyloid beta [Aß], and/or phosphorylated tau, all of which are key pathological mediators of AD). Although these models may provide insight on pathophysiological processes in AD, none completely recapitulate the disease and its strong age-dependence, and there has been limited success in translating preclinical results and treatments to humans. Here, we describe 2 nontransgenic guinea pig (GP) models, a standard PigmEnTed (PET) strain, and lesser-studied Dunkin-Hartley (DH) strain, that may naturally mimic key features of brain aging and AD in humans. We show that brain aging in PET GP is transcriptomically similar to human brain aging, whereas older DH brains are transcriptomically more similar to human AD. Both strains/models also exhibit increased neurofilament light chain (NFL, a marker of neuronal damage) with aging, and DH animals display greater S100 calcium-binding protein B (S100ß), ionized calcium-binding adapter molecule 1 (Iba1), and Aß and phosphorylated tau-which are all important markers of neuroinflammation-associated AD. Collectively, our results suggest that both the PET and DH GP may be useful, nontransgenic models to study brain aging and AD, respectively.

EFHD1 ablation inhibits cardiac mitoflash activation and protects cardiomyocytes from ischemia.

Altered levels of intracellular calcium (Ca2+) are a highly prevalent feature in different forms of cardiac injury, producing changes in contractility, arrhythmias, and mitochondrial dysfunction. In cardiac ischemia-reperfusion injury, mitochondrial Ca2+ overload leads to pathological production of reactive oxygen species (ROS), activates the permeability transition, and cardiomyocyte death. Here we investigated the cardiac phenotype caused by deletion of EF-hand domain-containing protein D1 (Efhd1-/-), a Ca2+-binding mitochondrial protein whose function is poorly understood. Efhd1-/- mice are viable and have no adverse cardiac phenotypes. They feature reductions in basal ROS levels and mitoflash events, both important precursors for mitochondrial injury, though cardiac mitochondria have normal susceptibility to Ca2+ overload. Notably, we also find that Efhd1-/- mice and their cardiomyocytes are resistant to hypoxic injury.

The Dunkin Hartley Guinea Pig Is a Model of Primary Osteoarthritis That Also Exhibits Early Onset Myofiber Remodeling That Resembles Human Musculoskeletal Aging.

Skeletal muscle dysfunction, articular cartilage degeneration, and bone loss occur essentially in parallel during aging. Mechanisms contributing to this systemic musculoskeletal decline remain incompletely understood, limiting progress toward developing effective therapeutics. Because the progression of human musculoskeletal aging is slow, researchers rely on rodent models to identify mechanisms and test interventions. The Dunkin Hartley guinea pig is an outbred strain that begins developing primary osteoarthritis by 4 months of age with a progression and pathology similar to aging humans. The purpose of this study was to determine if skeletal muscle remodeling during the progression of osteoarthritis in these guinea pigs resembles musculoskeletal aging in humans. We compared Dunkin Hartley guinea pigs to Strain 13 guinea pigs, which develop osteoarthritis much later in the lifespan. We measured myofiber type and size, muscle density, and long-term fractional protein synthesis rates of the gastrocnemius and soleus muscles in 5, 9, and 15-month-old guinea pigs. There was an age-related decline in skeletal muscle density, a greater proportion of smaller myofibers, and a decline in type II concomitant with a rise in type I myofibers in the gastrocnemius muscles from Dunkin Hartley guinea pigs only. These changes were accompanied by age-related declines in myofibrillar and mitochondrial protein synthesis in the gastrocnemius and soleus. Collectively, these findings suggest Dunkin Hartley guinea pigs experience myofiber remodeling alongside the progression of osteoarthritis, consistent with human musculoskeletal aging. Thus, Dunkin Hartley guinea pigs may be a model to advance discovery and therapeutic development for human musculoskeletal aging.