Spermidine treatment: induction of autophagy but also apoptosis?

Machado-Joseph disease (MJD), also known as spinocerebellar ataxia type 3, is a fatal neurodegenerative disease that causes loss of balance and motor co-ordination, eventually leading to paralysis. It is caused by the autosomal dominant inheritance of a long CAG trinucleotide repeat sequence within the ATXN3 gene, encoding for an expanded polyglutamine (polyQ) repeat sequence within the ataxin-3 protein. Ataxin-3 containing an expanded polyQ repeat is known to be highly prone to intraneuronal aggregation, and previous studies have demonstrated that protein quality control pathways, such as autophagy, are impaired in MJD patients and animal models of the disease. In this study, we tested the therapeutic potential of spermidine on zebrafish and rodent models of MJD to determine its capacity to induce autophagy and improve functional output. Spermidine treatment of transgenic MJD zebrafish induced autophagy and resulted in increased distances swum by the MJD zebrafish. Interestingly, treatment of the CMVMJD135 mouse model of MJD with spermidine added to drinking water did not produce any improvement in motor behaviour assays, neurological testing or neuropathology. In fact, wild type mice treated with spermidine were found to have decreased rotarod performance when compared to control animals. Immunoblot analysis of protein lysates extracted from mouse cerebellar tissue found little differences between the groups, except for an increased level of phospho-ULK1 in spermidine treated animals, suggesting that autophagy was indeed induced. As we detected decreased motor performance in wild type mice following treatment with spermidine, we conducted follow up studies into the effects of spermidine treatment in zebrafish. Interestingly, we found that in addition to inducing autophagy, spermidine treatment also induced apoptosis, particularly in wild type zebrafish. These findings suggest that spermidine treatment may not be therapeutically beneficial for the treatment of MJD, and in fact warrants caution due to the potential negative side effects caused by induction of apoptosis.

Empowering personalized pharmacogenomics with generative AI solutions.

OBJECTIVE: This study evaluates an AI assistant developed using OpenAI's GPT-4 for interpreting pharmacogenomic (PGx) testing results, aiming to improve decision-making and knowledge sharing in clinical genetics and to enhance patient care with equitable access. MATERIALS AND METHODS: The AI assistant employs retrieval-augmented generation (RAG), which combines retrieval and generative techniques, by harnessing a knowledge base (KB) that comprises data from the Clinical Pharmacogenetics Implementation Consortium (CPIC). It uses context-aware GPT-4 to generate tailored responses to user queries from this KB, further refined through prompt engineering and guardrails. RESULTS: Evaluated against a specialized PGx question catalog, the AI assistant showed high efficacy in addressing user queries. Compared with OpenAI's ChatGPT 3.5, it demonstrated better performance, especially in provider-specific queries requiring specialized data and citations. Key areas for improvement include enhancing accuracy, relevancy, and representative language in responses. DISCUSSION: The integration of context-aware GPT-4 with RAG significantly enhanced the AI assistant's utility. RAG's ability to incorporate domain-specific CPIC data, including recent literature, proved beneficial. Challenges persist, such as the need for specialized genetic/PGx models to improve accuracy and relevancy and addressing ethical, regulatory, and safety concerns. CONCLUSION: This study underscores generative AI's potential for transforming healthcare provider support and patient accessibility to complex pharmacogenomic information. While careful implementation of large language models like GPT-4 is necessary, it is clear that they can substantially improve understanding of pharmacogenomic data. With further development, these tools could augment healthcare expertise, provider productivity, and the delivery of equitable, patient-centered healthcare services.

A randomised controlled evaluation of an online perfectionism intervention for people with disordered eating - how perfect does it need to be?

Less help-seeking for an eating disorder is predicted by higher levels of denial of, and failure to perceive, illness severity. This research evaluates a "backdoor" approach to early intervention by investigating whether internet cognitive behaviour therapy for perfectionism can significantly improve disordered eating. Additionally, we investigated whether a more interactive intervention impacted outcomes. Participants were recruited worldwide online; 368 were screened, 172 (46.7%) met inclusion criteria (endorsed high shape, weight, or eating concerns) and randomised to an interactive (Focused Minds Program; FMP) or static PDF intervention (Centre for Clinical Intervention; CCI-P) or waitlisted (control condition). Participants completed assessments on disordered eating, perfectionism, and a range of secondary variables at the end of treatment, and 1- and 3-month follow-up. Intent-to-treat analyses indicated that, compared to control, FMP resulted in significantly lower levels of disordered eating at each assessment and CCI-P at the 1- and 3-month follow-up (respective 3-month follow-up between group effect sizes of 0.78 and 0.54). There were no significant differences between the two active interventions on any measure except depression and hated self. Results suggest an alternative approach to directly tackling disordered eating that is low-cost is effective, with a more interactive intervention producing a more rapid effect.Trials Registration: Australian New Zealand Clinical Trials Registry (ANZCTR) Trial Number: ACTRN12621001448831.

Treatment with sodium butyrate induces autophagy resulting in therapeutic benefits for spinocerebellar ataxia type 3.

Spinocerebellar ataxia type 3 (SCA3, also known as Machado Joseph disease) is a fatal neurodegenerative disease caused by the expansion of the trinucleotide repeat region within the ATXN3/MJD gene. Mutation of ATXN3 causes formation of ataxin-3 protein aggregates, neurodegeneration, and motor deficits. Here we investigated the therapeutic potential and mechanistic activity of sodium butyrate (SB), the sodium salt of butyric acid, a metabolite naturally produced by gut microbiota, on cultured SH-SY5Y cells and transgenic zebrafish expressing human ataxin-3 containing 84 glutamine (Q) residues to model SCA3. SCA3 SH-SY5Y cells were found to contain high molecular weight ataxin-3 species and detergent-insoluble protein aggregates. Treatment with SB increased the activity of the autophagy protein quality control pathway in the SCA3 cells, decreased the presence of ataxin-3 aggregates and presence of high molecular weight ataxin-3 in an autophagy-dependent manner. Treatment with SB was also beneficial in vivo, improving swimming performance, increasing activity of the autophagy pathway, and decreasing the presence of insoluble ataxin-3 protein species in the transgenic SCA3 zebrafish. Co-treating the SCA3 zebrafish with SB and chloroquine, an autophagy inhibitor, prevented the beneficial effects of SB on zebrafish swimming, indicating that the improved swimming performance was autophagy-dependent. To understand the mechanism by which SB induces autophagy we performed proteomic analysis of protein lysates from the SB-treated and untreated SCA3 SH-SY5Y cells. We found that SB treatment had increased activity of Protein Kinase A and AMPK signaling, with immunoblot analysis confirming that SB treatment had increased levels of AMPK protein and its substrates. Together our findings indicate that treatment with SB can increase activity of the autophagy pathway process and that this has beneficial effects in vitro and in vivo. While our results suggested that this activity may involve activity of a PKA/AMPK-dependent process, this requires further confirmation. We propose that treatment with sodium butyrate warrants further investigation as a potential treatment for neurodegenerative diseases underpinned by mechanisms relating to protein aggregation including SCA3.

Trained immunity in diabetes and hyperlipidemia: Emerging opportunities to target cardiovascular complications and design new therapies.

Some metabolic diseases, such as diabetes and hyperlipidemia, are associated with a state of inflammation, which adversely affects cardiovascular health. Emerging evidence suggests that long-term hyperactivation of innate immune cells and their bone marrow progenitors, termed trained immunity, functions to accelerate atherosclerosis and its complications in cardiometabolic diseases. This review will focus on how trained immunity is established, particularly through metabolic and epigenetic reprogramming, to cause persistent and deleterious changes in immune cell function, even after the original stimulus has been corrected or removed. Understanding the mechanisms driving maladaptive trained immunity and its fundamental contribution to cardiovascular disease might enable the development of novel disease-modifying therapeutics for the reduction in cardiovascular risk in diabetes, hyperlipidemia, and related cardiometabolic states.

Machado Joseph disease severity is linked with gut microbiota alterations in transgenic mice.

Emerging evidence suggests the presence of bidirectional interactions between the central nervous system and gut microbiota that may contribute to the pathogenesis of neurodegenerative diseases. However, the potential role of gut microbes in forms of spinocerebellar ataxia, such as the fatal neurodegenerative disease Machado Joseph disease (MJD), remains unexplored. Here, we examined whether gut microbiota alterations may be an early disease phenotype of MJD. We profiled the gut microbiota of male and female transgenic MJD mice (CMVMJD135) expressing human ATXN3 with expanded CAG repeats (133-143 CAG) at pre-symptomatic, symptomatic and well-established stages of the disease (7, 11 and 15 weeks of age, respectively). We compared these profiles with the gut microbiota of male and female wild-type (WT) littermate control mice at same ages. Correlation network analyses were employed to explore the relevance of microbiota changes to disease progression. The results demontrated distinct sex-dependent effects in disease development whereby male MJD mice displayed earlier motor impairments than female MJD mice. The gut microbiota community structure and composition also demonstrated sex-specific differences between MJD and WT mice. In both male and female MJD mice, the shifts in the microbiota were present by 7 weeks, before the onset of any symptoms. These pre-symptomatic microbial changes correlated with the severity of neurological impairments present at later stages of the disease. Previous efforts towards developing treatments for MJD have failed to yield meaningful outcomes. Our study reports a novel relationship between the gut microbiota and MJD development and severity. Elucidating how gut microbes are involved in MJD pathogenesis may offer new and efficacious treatment strategies for this currently untreatable disease.

Evidence and User Considerations of Home Health Monitoring for Older Adults: Scoping Review.

BACKGROUND: Home health monitoring shows promise in improving health outcomes; however, navigating the literature remains challenging given the breadth of evidence. There is a need to summarize the effectiveness of monitoring across health domains and identify gaps in the literature. In addition, ethical and user-centered frameworks are important to maximize the acceptability of health monitoring technologies. OBJECTIVE: This review aimed to summarize the clinical evidence on home-based health monitoring through a scoping review and outline ethical and user concerns and discuss the challenges of the current user-oriented conceptual frameworks. METHODS: A total of 2 literature reviews were conducted. We conducted a scoping review of systematic reviews in Scopus, MEDLINE, Embase, and CINAHL in July 2021. We included reviews examining the effectiveness of home-based health monitoring in older adults. The exclusion criteria included reviews with no clinical outcomes and lack of monitoring interventions (mobile health, telephone, video interventions, virtual reality, and robots). We conducted a quality assessment using the Assessment of Multiple Systematic Reviews (AMSTAR-2). We organized the outcomes by disease and summarized the type of outcomes as positive, inconclusive, or negative. Second, we conducted a literature review including both systematic reviews and original articles to identify ethical concerns and user-centered frameworks for smart home technology. The search was halted after saturation of the basic themes presented. RESULTS: The scoping review found 822 systematic reviews, of which 94 (11%) were included and of those, 23 (24%) were of medium or high quality. Of these 23 studies, monitoring for heart failure or chronic obstructive pulmonary disease reduced exacerbations (4/7, 57%) and hospitalizations (5/6, 83%); improved hemoglobin A1c (1/2, 50%); improved safety for older adults at home and detected changing cognitive status (2/3, 66%) reviews; and improved physical activity, motor control in stroke, and pain in arthritis in (3/3, 100%) rehabilitation studies. The second literature review on ethics and user-centered frameworks found 19 papers focused on ethical concerns, with privacy (12/19, 63%), autonomy (12/19, 63%), and control (10/19, 53%) being the most common. An additional 7 user-centered frameworks were studied. CONCLUSIONS: Home health monitoring can improve health outcomes in heart failure, chronic obstructive pulmonary disease, and diabetes and increase physical activity, although review quality and consistency were limited. Long-term generalized monitoring has the least amount of evidence and requires further study. The concept of trade-offs between technology usefulness and acceptability is critical to consider, as older adults have a hierarchy of concerns. Implementing user-oriented frameworks can allow long-term and larger studies to be conducted to improve the evidence base for monitoring and increase the receptiveness of clinicians, policy makers, and end users.

The effect of self-administered methamphetamine on GABAergic interneuron populations and functional connectivity of the nucleus accumbens and prefrontal cortex.

INTRODUCTION: Methamphetamine (METH, "ice") is a potent and addictive psychostimulant. Abuse of METH perturbs neurotransmitter systems and induces neurotoxicity; however, the neurobiological mechanisms which underlie addiction to METH are not fully understood, limiting the efficacy of available treatments. Here we investigate METH-induced changes to neuronal nitric oxide synthase (nNOS), parvalbumin and calretinin-expressing GABAergic interneuron populations within the nucleus accumbens (NAc), prefrontal cortex (PFC) and orbitofrontal cortex (OFC). We hypothesise that dysfunction or loss of these GABAergic interneuron populations may disrupt the excitatory/inhibitory balance within the brain. METHODS: Male Long Evans rats (N = 32) were trained to lever press for intravenous METH or received yoked saline infusions. Following 14 days of behavioural extinction, animals were given a non-contingent injection of saline or METH (1 mg/kg, IP) to examine drug-primed reinstatement to METH-seeking behaviours. Ninety minutes post-IP injection, animals were culled and brain sections were analysed for Fos, nNOS, parvalbumin and calretinin immunoreactivity in eight distinct subregions of the NAc, PFC and OFC. RESULTS: METH exposure differentially affected GABAergic populations, with METH self-administration increasing nNOS immunoreactivity at distinct locations in the prelimbic cortex and decreasing parvalbumin immunoreactivity in the NAc. METH self-administration triggered reduced calretinin immunoreactivity, whilst acute METH administration produced a significant increase in calretinin immunoreactivity. As expected, non-contingent METH-priming treatment increased Fos immunoreactivity in subregions of the NAc and PFC. CONCLUSION: Here we report that METH exposure in this model may alter the function of GABAergic interneurons in more subtle ways, such as alterations in neuronal firing or synaptic connectivity.

The Effect of Non-Pharmacological and Pharmacological Interventions on Measures Associated with Sarcopenia in End-Stage Kidney Disease: A Systematic Review and Meta-Analysis.

This systematic review and meta-analysis provides a synthesis of the available evidence for the effects of interventions on outcome measures associated with sarcopenia in end-stage kidney disease (ESKD). Thirteen databases were searched, supplemented with internet and hand searching. Randomised controlled trials of non-pharmacological or pharmacological interventions in adults with ESKD were eligible. Trials were restricted to those which had reported measures of sarcopenia. Primary outcome measures were hand grip strength and sit-to-stand tests. Sixty-four trials were eligible (with nineteen being included in meta-analyses). Synthesised data indicated that intradialytic exercise increased hand grip strength (standardised mean difference, 0.58; 0.24 to 0.91; p = 0.0007; I2 = 40%), and sit-to-stand (STS) 60 score (mean difference, 3.74 repetitions; 2.35 to 5.14; p < 0.001; I2 = 0%). Intradialytic exercise alone, and protein supplementation alone, resulted in no statistically significant change in STS5 (−0.78 s; −1.86 to 0.30; p = 0.16; I2 = 0%), and STS30 (MD, 0.97 repetitions; −0.16 to 2.10; p = 0.09; I2 = 0%) performance, respectively. For secondary outcomes, L-carnitine and nandrolone-decanoate resulted in significant increases in muscle quantity in the dialysis population. Intradialytic exercise modifies measures of sarcopenia in the haemodialysis population; however, the majority of trials were low in quality. There is limited evidence for efficacious interventions in the peritoneal dialysis and transplant recipient populations.

Oxytocin as an adolescent treatment for methamphetamine addiction after early life stress in male and female rats.

Early life stress (ELS) is associated with perturbed neural development and augmented vulnerability to mental health disorders, including addiction. How ELS changes the brain to increase addiction risk is poorly understood, and there are no therapies which target this ELS-induced vulnerability. ELS disrupts the oxytocin system, which can modulate addiction susceptibility, suggesting that targeting the oxytocin system may be therapeutic in this ELS-addiction comorbidity. Therefore, we determined whether adolescent oxytocin treatment after ELS could: (1) reduce vulnerability to anxiety, social deficits, and methamphetamine-taking and reinstatement; and (2) restore hypothalamic oxytocin and corticotropin-releasing factor expressing neurons and peripheral oxytocin and corticosterone levels. Long Evans pups underwent maternal separation (MS) for either 15 min or 360 min on postnatal days (PND) 1-21. During adolescence (PNDs 28-42), rats received a daily injection of either oxytocin or saline. In Experiment 1, adult rats were assessed using the elevated plus-maze, social interaction procedure, and methamphetamine self-administration procedure, including extinction, and cue-, methamphetamine- and yohimbine-induced reinstatement. In Experiment 2, plasma for enzyme immunoassays and brain tissue for immunofluorescence were collected from adult rats after acute stress exposure. Adolescent oxytocin treatment ameliorated ELS-induced anxiety and reduced methamphetamine- and yohimbine-induced reinstatement in both sexes, and suppressed methamphetamine intake and facilitated extinction in males only. Additionally, adolescent oxytocin treatment after ELS restored oxytocin-immunoreactive cells and stress-induced oxytocin levels in males, and attenuated stress-induced corticosterone levels in both sexes. Adolescent oxytocin treatment reverses some of the ELS effects on later-life psychopathology and vulnerability to addiction.

Preventative Cancer Screening Rates Among Uninsured Patients in Free Clinics: A Retrospective Cohort Study of Cancer Survivors and Non-cancer Survivors.

BACKGROUND: There is limited research on screening rates among uninsured cancer survivors. Uninsured cancer survivors are at higher risk of poorer health outcomes than the insured due to limited access to preventative screening for secondary cancers. This study examines the rates of surveillance and screening of uninsured cancer survivors and compares to uninsured patients without a cancer history seen in free clinics. METHODS: Data were collected retrospectively from electronic medical records and paper charts of patients from 10 free clinics between January 2016 and December 2018 in the Tampa Bay area. The prevalence of socioeconomic characteristics, cancer diagnoses, and screening practices were compared for cancer survivors and free clinic patients without a history of cancer. Study participants were determined to be eligible for cancer screenings based on the United States Preventive Services Task Force guidelines. RESULTS: Out of 13 982 uninsured patients frequenting free clinics between 2016 and 2018, 402 (2.9%) had a documented history of cancer. Out of the 285 eligible cancer survivors, 44 (15.4%) had completed age-appropriate colon cancer screening. Among the 170 female cancer survivors, 75 (44.1%) had completed breast cancer screenings, and only 5.9% (59/246) had completed cervical cancer screenings. After adjusting for age, gender, race, salary, employment status, and household size, cancer survivors were more likely to undergo colorectal cancer screening (OR: 3.59, 95% CI: 2.10-6.15) and breast cancer screening (OR: 2.13, 95% CI: 1.30-3.84) than patients without a cancer history. This difference was not seen for cervical cancer screening (OR: 0.99, 95% CI: .62-1.58). CONCLUSIONS: Uninsured cancer survivors frequenting free clinics represent a unique population that is underrepresented in the medical literature. Our results suggest that uninsured survivors use screening services at higher rates when compared to uninsured patients without a reported cancer diagnosis. However, these rates are suboptimal when compared to national screening rates of insured cancer survivors.

Low pH up-regulates interleukin-6 mRNA in L6-G8C5 rat skeletal muscle cells independent of pH sensing by SNAT2(SLC38A2) transporters.

Exercise is known to create a transient, but potent increase in skeletal muscle expression of potentially anti-inflammatory myokine interleukin-6 (IL-6). This effect may be clinically important in managing chronic inflammatory states. It has previously been proposed that lactic acidosis following exercise promotes this IL-6 up-regulation, but the mechanism of this acidosis effect is unknown. Rat skeletal muscle cell line L6-G8C5 has been used previously to model metabolic effects of acidosis, sensing low pH through the resulting inhibition of amino acid transporter SNAT2(SLC38A2). Use of ionophore ionomycin to model the rise in intracellular Ca2+ concentration occurring in contracting muscle strongly up-regulates IL-6 mRNA in L6-G8C5 myotubes. This study used this model to test the hypothesis that low extracellular pH (7.1) enhances ionomycin-induced IL-6 mRNA up-regulation by inhibiting SNAT2. Incubation of L6-G8C5 myotubes for 6 h with 0.5 µM ionomycin at control pH (7.4) resulted in a 15-fold increase in IL-6 mRNA which was further enhanced (1.74-fold) at pH 7.1. In contrast low pH had no significant effect on IL-6 mRNA without ionomycin, nor on the IL-6 mRNA increase that was induced by cyclic stretch. Even though pH 7.1 halved the transport activity of SNAT2, alternative methods of SNAT2 inhibition (JNK inhibitor SP600125; SNAT2 antagonist MeAIB; or SNAT2 silencing with siRNA) did not mimic the enhancing effect of low pH on IL-6 mRNA. On the contrary, JNK inhibition blunted the effect of pH 7.1 with ionomycin, but had no effect at pH 7.4. It is concluded that low pH promotes Ca2+/ionomycin-induced up-regulation of IL-6 mRNA through a novel SNAT2-independent JNK-dependent pH-sensing pathway not previously described in this skeletal muscle model.

Primary skeletal muscle cells from chronic kidney disease patients retain hallmarks of cachexia in vitro.

BACKGROUND: Skeletal muscle wasting and dysfunction are common characteristics noted in people who suffer from chronic kidney disease (CKD). The mechanisms by which this occurs are complex, and although progress has been made, the key underpinning mechanisms are not yet fully elucidated. With work to date primarily conducted in nephrectomy-based animal models, translational capacity to our patient population has been challenging. This could be overcome if rationale developing work could be conducted in human based models with greater translational capacity. This could be achieved using cells derived from patient biopsies, if they retain phenotypic traits noted in vivo. METHODS: Here, we performed a systematic characterization of CKD derived muscle cells (CKD; n = 10; age: 54.40 ± 15.53 years; eGFR: 22.25 ± 13.22 ml/min/1.73 m2 ) in comparison with matched controls (CON; n = 10; age: 58.66 ± 14.74 years; eGFR: 85.81 ± 8.09 ml/min/1.73 m2 ). Harvested human derived muscle cells (HDMCs) were taken through proliferative and differentiation phases and investigated in the context of myogenic progression, inflammation, protein synthesis, and protein breakdown. Follow up investigations exposed HDMC myotubes from each donor type to 0, 0.4, and 100 nM of IGF-1 in order to investigate any differences in anabolic resistance. RESULTS: Harvested human derived muscle cells isolated from CKD patients displayed higher rates of protein degradation (P = 0.044) alongside elevated expression of both TRIM63 (2.28-fold higher, P = 0.054) and fbox32 (6.4-fold higher, P < 0.001) in comparison with CONs. No differences were noted in rates of protein synthesis under basal conditions (P > 0.05); however, CKD derived cells displayed a significant degree of anabolic resistance in response to IGF-1 stimulation (both doses) in comparison with matched CONs (0.4 nm: P < 0.001; 100 nM: P < 0.001). CONCLUSIONS: In summary, we report for the first time that HDMCs isolated from people suffering from CKD display key hallmarks of the well documented in vivo phenotype. Not only do these findings provide further mechanistic insight into CKD specific cachexia, but they also demonstrate this is a reliable and suitable model in which to perform targeted experiments to begin to develop novel therapeutic strategies targeting the CKD associated decline in skeletal muscle mass and function.

A Systematic Review of Handgrip Strength Measurement in Clinical and Epidemiological Studies of Kidney Disease: Toward a Standardized Approach.

In chronic kidney disease (CKD), handgrip strength (HGS) is recommended as a surrogate measure of protein-energy status and functional status. However, it is not routinely used because of inconsistencies such as the optimal timing of the HGS measurement and unclear guidance regarding technique. We aimed to determine the extent of variation in the protocols and methods of HGS assessment. We aimed to identify clinical and epidemiological studies conducted on CKD that reported on the use of HGS as an outcome. A systematic literature search identified n = 129 studies with a total participant population of n = 35,192. We identified large variations in all aspects of the methodology including body and arm position, repetitions, rest time, timing, familiarization, and how scores were calculated. The heterogeneous methodologies used reinforce the need to standardize HGS measurement. After reviewing previously employed methodology in the literature, we propose a comprehensive HGS assessment protocol for use in CKD.

Pathophysiological interplay between O-GlcNAc transferase and the Machado-Joseph disease protein ataxin-3.

Aberrant O-GlcNAcylation, a protein posttranslational modification defined by the O-linked attachment of the monosaccharide N-acetylglucosamine (O-GlcNAc), has been implicated in neurodegenerative diseases. However, although many neuronal proteins are substrates for O-GlcNAcylation, this process has not been extensively investigated in polyglutamine disorders. We aimed to evaluate the enzyme O-GlcNAc transferase (OGT), which attaches O-GlcNAc to target proteins, in Machado-Joseph disease (MJD). MJD is a neurodegenerative condition characterized by ataxia and caused by the expansion of a polyglutamine stretch within the deubiquitinase ataxin-3, which then present increased propensity to aggregate. By analyzing MJD cell and animal models, we provide evidence that OGT is dysregulated in MJD, therefore compromising the O-GlcNAc cycle. Moreover, we demonstrate that wild-type ataxin-3 modulates OGT protein levels in a proteasome-dependent manner, and we present OGT as a substrate for ataxin-3. Targeting OGT levels and activity reduced ataxin-3 aggregates, improved protein clearance and cell viability, and alleviated motor impairment reminiscent of ataxia of MJD patients in zebrafish model of the disease. Taken together, our results point to a direct interaction between OGT and ataxin-3 in health and disease and propose the O-GlcNAc cycle as a promising target for the development of therapeutics in the yet incurable MJD.

Morphometric, metabolic, and inflammatory markers across a cohort of client-owned horses and ponies on the insulin dysregulation spectrum.

In human metabolic syndrome and type II diabetes, methylglyoxal (MG), D-lactate, and several cytokines have been recognized as biomarkers of important metabolic and inflammatory processes. Equine metabolic syndrome (EMS) shares many similarities with these human counterparts. The objectives of this cross-sectional study were to compare body condition score (BCS), cresty neck score (CNS), resting insulin, MG, D-lactate, L-lactate, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and monocyte chemoattractant protein-1 (MCP-1) between horses with and without insulin dysregulation, as classified via combined glucose and insulin test (CGIT). 32 client-owned horses were included. History and morphometric data such as BCS and CNS were recorded. Subjects with abnormalities on physical examination or CBC, elevated ACTH or incomplete information were excluded. Baseline serum or plasma concentrations of biomarkers were tested via commercial ELISA or colorimetric assays. Characteristics of insulin dysregulated and insulin sensitive horses were compared by univariate analysis and forward logistic regression. 12 (38%) of the 32 horses were classified as insulin dysregulated. No significant difference between the 2 groups was found for age, BCS, baseline glucose, triglycerides, MG, D-lactate, L-lactate, TNF-α, IL-6, and MCP-1. Baseline insulin was significantly associated with insulin dysregulation in univariate analysis (P = 0.02), but not in the final model. Horses with CNS ≥ 3 had 11.3 times higher odds of having insulin dysregulation (OR 11.3, 95% C.I. 2.04 - 63.08, P = 0.006). In this population, horses with mild-moderate signs of EMS presented similar metabolic and inflammatory profiles to non-insulin dysregulated controls.

A Novel Calpain Inhibitor Compound Has Protective Effects on a Zebrafish Model of Spinocerebellar Ataxia Type 3.

Spinocerebellar ataxia type 3 (SCA3) is a hereditary ataxia caused by inheritance of a mutated form of the human ATXN3 gene containing an expanded CAG repeat region, encoding a human ataxin-3 protein with a long polyglutamine (polyQ) repeat region. Previous studies have demonstrated that ataxin-3 containing a long polyQ length is highly aggregation prone. Cleavage of the ataxin-3 protein by calpain proteases has been demonstrated to be enhanced in SCA3 models, leading to an increase in the aggregation propensity of the protein. Here, we tested the therapeutic potential of a novel calpain inhibitor BLD-2736 for the treatment of SCA3 by testing its efficacy on a transgenic zebrafish model of SCA3. We found that treatment with BLD-2736 from 1 to 6 days post-fertilisation (dpf) improves the swimming of SCA3 zebrafish larvae and decreases the presence of insoluble protein aggregates. Furthermore, delaying the commencement of treatment with BLD-2736, until a timepoint when protein aggregates were already known to be present in the zebrafish larvae, was still successful at removing enhanced green fluorescent protein (EGFP) fused-ataxin-3 aggregates and improving the zebrafish swimming. Finally, we demonstrate that treatment with BLD-2736 increased the synthesis of LC3II, increasing the activity of the autophagy protein quality control pathway. Together, these findings suggest that BLD-2736 warrants further investigation as a treatment for SCA3 and related neurodegenerative diseases.

A pilot randomised controlled trial of a structured, home-based exercise programme on cardiovascular structure and function in kidney transplant recipients: the ECSERT study design and methods.

BACKGROUND: Cardiovascular disease (CVD) is a major cause of morbidity and mortality in kidney transplant recipients (KTRs). CVD risk scores underestimate risk in this population as CVD is driven by clustering of traditional and non-traditional risk factors, which lead to prognostic pathological changes in cardiovascular structure and function. While exercise may mitigate CVD in this population, evidence is limited, and physical activity levels and patient activation towards exercise and self-management are low. This pilot study will assess the feasibility of delivering a structured, home-based exercise intervention in a population of KTRs at increased cardiometabolic risk and evaluate the putative effects on cardiovascular structural and functional changes, cardiorespiratory fitness, quality of life, patient activation, healthcare utilisation and engagement with the prescribed exercise programme. METHODS AND ANALYSIS: Fifty KTRs will be randomised 1:1 to: (1) the intervention; a 12week, home-based combined resistance and aerobic exercise intervention; or (2) the control; usual care. Intervention participants will have one introductory session for instruction and practice of the recommended exercises prior to receiving an exercise diary, dumbbells, resistance bands and access to instructional videos. The study will evaluate the feasibility of recruitment, randomisation, retention, assessment procedures and the intervention implementation. Outcomes, to be assessed prior to randomisation and postintervention, include: cardiac structure and function with stress perfusion cardiac MRI, cardiorespiratory fitness, physical function, blood biomarkers of cardiometabolic health, quality of life and patient activation. These data will be used to inform the power calculations for future definitive trials. ETHICS AND DISSEMINATION: The protocol was reviewed and given favourable opinion by the East Midlands-Nottingham 2 Research Ethics Committee (reference: 19/EM/0209; 14 October 2019). Results will be published in peer-reviewed academic journals and will be disseminated to the patient and public community via social media, newsletter articles and presentations at conferences. TRIAL REGISTRATION NUMBER: NCT04123951.

Flow cytometry allows rapid detection of protein aggregates in cellular and zebrafish models of spinocerebellar ataxia 3.

Spinocerebellar ataxia 3 (SCA3, also known as Machado-Joseph disease) is a neurodegenerative disease caused by inheritance of a CAG repeat expansion within the ATXN3 gene, resulting in polyglutamine (polyQ) repeat expansion within the ataxin-3 protein. In this study, we have identified protein aggregates in both neuronal-like (SHSY5Y) cells and transgenic zebrafish expressing human ataxin-3 with expanded polyQ. We have adapted a previously reported flow cytometry methodology named flow cytometric analysis of inclusions and trafficking, allowing rapid quantification of detergent insoluble forms of ataxin-3 fused to a GFP in SHSY5Y cells and cells dissociated from the zebrafish larvae. Flow cytometric analysis revealed an increased number of detergent-insoluble ataxin-3 particles per nuclei in cells and in zebrafish expressing polyQ-expanded ataxin-3 compared to those expressing wild-type human ataxin-3. Treatment with compounds known to modulate autophagic activity altered the number of detergent-insoluble ataxin-3 particles in cells and zebrafish expressing mutant human ataxin-3. We conclude that flow cytometry can be harnessed to rapidly count ataxin-3 aggregates, both in vitro and in vivo, and can be used to compare potential therapies targeting protein aggregates. This article has an associated First Person interview with the first author of the paper.

Sodium valproate increases activity of the sirtuin pathway resulting in beneficial effects for spinocerebellar ataxia-3 in vivo.

Machado-Joseph disease (MJD, also known as spinocerebellar ataxia type 3) is a fatal neurodegenerative disease that impairs control and coordination of movement. Here we tested whether treatment with the histone deacetylase inhibitor sodium valproate (valproate) prevented a movement phenotype that develops in larvae of a transgenic zebrafish model of the disease. We found that treatment with valproate improved the swimming of the MJD zebrafish, affected levels of acetylated histones 3 and 4, but also increased expression of polyglutamine expanded human ataxin-3. Proteomic analysis of protein lysates generated from the treated and untreated MJD zebrafish also predicted that valproate treatment had activated the sirtuin longevity signaling pathway and this was confirmed by findings of increased SIRT1 protein levels and sirtuin activity in valproate treated MJD zebrafish and HEK293 cells expressing ataxin-3 84Q, respectively. Treatment with resveratrol (another compound known to activate the sirtuin pathway), also improved swimming in the MJD zebrafish. Co-treatment with valproate alongside EX527, a SIRT1 activity inhibitor, prevented induction of autophagy by valproate and the beneficial effects of valproate on the movement in the MJD zebrafish, supporting that they were both dependent on sirtuin activity. These findings provide the first evidence of sodium valproate inducing activation of the sirtuin pathway. Further, they indicate that drugs that target the sirtuin pathway, including sodium valproate and resveratrol, warrant further investigation for the treatment of MJD and related neurodegenerative diseases.