Injection Of Vehicle Research Articles

Nitrosyl factors play a vital role in the ventilatory depressant effects of fentanyl in freely moving guinea pigs.

An understanding of intracellular mechanisms by which fentanyl and other synthetic opioids exert adverse effects on breathing is needed. Using freely moving adult male guinea pigs, we administered the nitric oxide synthase (NOS) inhibitor, L-NAME (NG-nitro-L-arginine methyl ester), to determine whether nitrosyl factors, such as nitric oxide and S-nitrosothiols, play a role in fentanyl-induced respiratory depression. Ventilatory parameters were recorded by whole body plethysmography to determine the effects of fentanyl (75 μg/kg, IV) in guinea pigs that had received a prior injection of vehicle (saline), L-NAME or the inactive D-isomer, D-NAME (both at 50 μmol/kg, IV), 15 min beforehand. L- and D-NAME elicited minor effects on most parameters, including frequency of breathing, tidal volume and minute ventilation, although L-NAME did decrease endexpiratory pause and non-eupneic breathing index (NEBI). Subsequent injection of fentanyl in guinea pigs pre-treated with vehicle produced profound and sustained reductions in frequency, tidal volume, minute ventilation, peak inspiratory flow, and inspiratory and expiratory drives, while increasing inspiratory time, expiratory time, end inspiratory pause, and NEBI. These ventilatory depressant effects of fentanyl seen in guinea pigs pre-treated with vehicle were markedly diminished in guinea pigs pre-treated with L-NAME. Moreover, the adverse effects of fentanyl on many recorded breathing parameters were converted to stimulatory effects. In contrast, D-NAME did not alter any of the effects of fentanyl on breathing. This study is the first to characterize the role nitrosyl factors play in the intracellular mechanisms involved in fentanyl-induced respiratory depression in guinea pigs.

Melatonin lessens the susceptibility to atrial fibrillation in sleep deprivation by ameliorating Ca2+ mishandling in response to mitochondrial oxidative stress.

The antiarrhythmic effect of melatonin(MLT) has been demonstrated in several studies; however, this hypothesis has recently been contested. Our research seeks to determine if exogenous MLT supplementation can reduce atrial fibrillation (AF) susceptibility due to sleep deprivation (SD) by addressing Ca2+ mishandling and atrial mitochondrial oxidative stress. Adult rats received daily MLT or vehicle injections and were exposed to a modified water tank. We evaluated MLT's impact on AF susceptibility by analyzing atrial electrical and structural changes, calcium handling, and oxidative stress markers. Techniques used included electrophysiological recording, echocardiography, optical mapping, histopathology, and molecular assays to understand MLT's protective effects against sleep deprivation-induced AF. Our findings indicate that MLT treatment effectively mitigates SD-induced AF, safeguards against atrial structural alterations, diminishes mitochondrial oxidative stress and normalizes calcium dynamics. Notably, MLT corrected calcium transient duration (CaD), action potential duration (APD), and conduction heterogeneity, shortened calcium transient refractoriness, and improved arrhythmogenic atrial alternans and spatially discordant alternans, thereby lowering the arrhythmogenic potential of the atria during sleep deprivation. In terms of mechanisms, MLT prevents SD-induced activation of ROS/CaMKII in atrial cardiomyocytes, reversing calcium transient refractoriness and inhibiting arrhythmogenic alternans. MLT significantly decreases the susceptibility to SD-induced AF by ameliorating mitochondrial oxidative stress and Ca2+ mishandling. These findings suggest a potential therapeutic application of MLT as an antiarrhythmic intervention for SD-related AF and underscore the need for further investigation, including clinical studies, to validate these mechanisms.

Impact of Exercise on Tramadol-Conditioned Place Preference.

Tramadol (TRA) is an opioid that is used to manage moderate to severe pain. Long-term use of TRA can lead to the development of opioid use disorder. This study investigates the role of forced exercise in reducing TRA-seeking behavior. Adult male rats (240-260 g) were divided into five groups; the control group received vehicle injections, the TRA group received TRA (75 mg/kg, i.p) every other day for 8 days, and three TRA-exercise groups were forced to run on a treadmill (60 min/day, 5 days/week) for 2, 4, or 6 weeks prior to conditioning with TRA. A tramadol-conditioned place preference (CPP) procedure assessed TRA reinforcement, after which all rats were euthanized, tissue extracted, and mRNA expression for brain-derived neurotrophic factor (Bdnf) and interleukin 1 beta (Il-1β) determined in hippocampus (Hipp), prefrontal cortex (PFC), and nucleus accumbens (NAc). TRA-seeking behavior was seen in the TRA group and the 6 weeks forced exercise group. By contrast, forced exercise for 2 or 4 weeks attenuated TRA-seeking behavior. This attenuation was associated with a significant increase in Bdnf mRNA expression in the Hipp and NAc, but not the PFC. Additionally, the TRA-induced elevations in Il-1β mRNA expression were reversed by all durations of exercise in Hipp. However, only 2 and 4 weeks, but not 6 weeks, of exercise reduced elevations in PFC and NAc Il-1β expression. Forced exercise for 2 and 4 weeks attenuates TRA-seeking behavior partially through the regulation of Bdnf and Il-1β mRNA expression.

Post-Stroke Recovery in Relation to Parvalbumin-Positive Interneurons and Perineuronal Nets.

There is a critical time window of post-stroke neuroplasticity when spontaneous behavioral recovery occurs. Potential factors responsible for this heightened plasticity are the reduction of parvalbumin-immunoreactive (PV+) interneuron inhibitory signaling and the disappearance of extracellular matrix synaptic stabilizers called perineuronal net(s; PNN/PNNs). This study investigated whether behavioral recovery during this critical period following stroke is associated with changes in densities of PV+ interneurons and PNNs. Male, Sprague-Dawley rats received forelimb motor cortex stroke (n = 43) using endothelin-1, or vehicle injections (n = 44). Cohorts of rats underwent a battery of motor tests and were sacrificed within the post-stroke critical window on day 1, and 1, 2, 4, and 6 weeks. Using immunofluorescent labeling, PNNs (wisteria floribunda agglutinin; WFA+ cells), PV+ interneurons, and cells expressing both PV and PNNs were quantified in contra- and ipsilesional cortices to elucidate their spatial-temporal profiles following stroke. PV+ interneuron density decreased significantly at 1-day post-stroke in the lateral ipsilesional cortex, while the density of PNNs was significantly lower up to 4 weeks post-stroke in the lateral ipsilesional cortex and at 1 and 2 weeks post-stroke in the medial ipsilesional cortex. Reduction of combined PV+/PNN signaling coincided with spontaneous behavioral recovery. These results suggest that post-stroke behavioral recovery corresponds to an early reduction in PV+/PNN co-labeled cells in conjunction with an early temporally-dependent reduction in PV+ interneuron signaling and chronic disappearance of PNNs. Interventions targeting PNNs or PV+ interneuron signaling have significant potential for extending the critical window of recovery following stroke.

Abaloparatide Enhances Bone Regeneration in Extraction Socket Dental Implant Defects: An Experimental InVivo Study.

Abaloparatide (ABL) is a synthetic parathyroid hormone-related protein analog developed as an anabolic drug to treat osteoporosis. ABL increases bone mineral density (BMD) of the long bones and spine; however, the influence of ABL on alveolar bone regeneration remains unknown. This study assessed the effects of systemic ABL administration on tooth extraction socket healing and dental implant osseointegration in a preclinical rodent model. Sprague-Dawley rats received daily subcutaneous injection of ABL (25 μg/kg) or vehicle (VEH) at the time of maxillary first molar (M1) extraction surgery; animals underwent either: (1) unilateral M1 extraction followed by sacrifice at 10 or 42d or (2)bilateral M1 extraction followed by staged implant placement in osteotomies with standardized defects and sacrifice at 21or 28d. Micro-computed tomography (micro-CT) analysis demonstrated that ABL increased bone volume fraction (p = 0.004) and bone mineral density (BMD) (p = 0.006) of regenerated extraction sockets at 42d. Micro-CT of the femur validated systemic effects of ABL, showing greater trabecular BMD after 6 (p < 0.01), 9 (p < 0.001), and 14 (p < 0.001) weeks of treatment. Histomorphometry confirmed a higher mineralized bone area with ABL treatment in extraction sockets at 42d (p = 0.03) and in the regenerated peri-implant bone at 21d post-implant placement (p = 0.01) compared to control. Systemic ABL administration enhances osteogenesis in extraction sockets prior to implant placement and accelerates peri-implant bone formation in the early phase of healing in the present rodent model.

Safety of Human USH1C Transgene Expression Following Subretinal Injection in Wild-Type Pigs.

This study aimed to evaluate early-phase safety of subretinal application of AAVanc80.CAG.USH1Ca1 (OT_USH_101) in wild-type (WT) pigs, examining the effects of a vehicle control, low dose, and high dose. Twelve WT pigs (24 eyes) were divided into three groups: four pigs each received bilateral subretinal injections of either vehicle, low dose (3.3 × 1010 vector genomes [vg] per eye), or high dose (1.0 × 1011 vg per eye). Total retinal thickness (TRT) was evaluated using optical coherence tomography and retinal function was assessed with full-field electroretinography (ff-ERG) at baseline and two months post-surgery. After necropsy, retinal changes were examined through histopathology, and human USH1C_a1/harmonin expression was assessed by quantitative PCR (qPCR) and Western blotting. OT_USH_101 led to high USH1C_a1 expression in WT pig retinas without significant TRT changes two months after subretinal injection. The qPCR revealed expression of the human USH1C_a1 transgene delivered by the adeno-associated virus vector. TRT changes were minimal across groups: vehicle (256 ± 21 to 243 ± 18 µm; P = 0.108), low dose (251 ± 32 to 258 ± 30 µm; P = 0.076), and high dose (242 ± 24 to 259 ± 28 µm; P = 0.590). The ff-ERG showed no significant changes in rod or cone responses. Histopathology indicated no severe retinal adverse effects in the vehicle and low dose groups. Early-phase clinical imaging, electrophysiology, and histopathological assessments indicated that subretinal administration of OT_USH_101 was well tolerated in the low-dose treatment arm. OT_USH_101 treatment resulted in high expression of human USH1C_a1. Although histopathological changes were not severe, more frequent changes were observed in the high-dose group.

A dissociated glucocorticoid receptor modulator mitigates glucolipotoxicity in the endocrine pancreas and peripheral tissues: Preclinical data from a mouse model of diet-induced type 2 diabetes.

Type 2 diabetes (T2D) is a prevalent metabolic disease linked to obesity and metabolic syndrome (MS). The glucolipotoxic environment (GLT) impacts tissues causing low-grade inflammation, insulin resistance and the gradual loss of pancreatic β-cell function, leading to hyperglycemia. We have previously shown that Compound A (CpdA), a plant-derived dissociative glucocorticoid receptor-modulator with inflammation-suppressive activity, displays protective effects on β-cells in type 1 diabetes murine models. This study aimed to evaluate whether the administration of CpdA can attenuate GLT effects and improve pathophysiological parameters in a murine model of T2D/MS. Eight-week-old male C57BL/6NCrl mice were fed either a standard chow diet or a high-fat/high-sucrose diet (HFHS) for 15weeks. From week 5 of feeding, each group received i.p. injections of CpdA (2.5μg/g) or vehicle three times a week. We also examined CpdA in vitro effect against GLT using the insulinoma cell line INS-1E and naïve isolated mouse islets. CpdA administration in HFHS fed mice improved glucose homeostasis and insulin sensitivity with no apparent side effects. CpdA treatment also preserved pancreatic islet architecture and insulin expression, while reducing hepatic steatosis and visceral adipose tissue inflammation induced by HFHS diet. In vitro assays in INS-1E cells and naïve isolated mouse islets demonstrated that CpdA counteracted GLT-induced inhibition of glucose-stimulated insulin secretion and supported the expression of key β-cell identity genes under GLT conditions. These findings highlight the potential protective effect of CpdA in preserving β-cell functionality and peripheral tissue physiology in the context of T2D/MS.

Effect of galanin-like peptide on hypothalamic kisspeptin expression in female Zucker fatty rats.

Kisspeptin and galanin-like peptide (GALP) neurons in the hypothalamic arcuate nucleus (ARC) are involved in gonadotropin-releasing hormone (GnRH) neuron-mediated pulsatile luteinizing hormone (LH) secretion. Zucker fatty (ZF) rats display a leptin receptor gene abnormality and suppressed pulsatile LH secretion. ZF rats reportedly exhibit low hypothalamic GALP and kisspeptin expression, and GALP administration induces LH release in ZF rats. Therefore, we performed a histochemical analysis to determine whether GALP-induced LH release is mediated by kisspeptin neurons in ZF rats. All ZF rats were ovariectomized and subcutaneously implanted with an estradiol tube before the central injection of GALP or vehicle. GALP administration increased the plasma LH concentration. However, no significant difference was observed in the number of Kiss1 cells and the proportion of Fos-positive Kiss1 cells. The number of c-Fos-positive GnRH neurons significantly increased after GALP administration. Our results suggest that hypothalamic GALP neurons promote LH release by activating GnRH neurons without the activation of kisspeptin neurons in the ARC.

Sexual Dimorphism of Ethanol-Induced Mitochondrial Dynamics in Purkinje Cells.

The cerebellum, a key target of ethanol's toxic effects, is associated with ataxia following alcohol consumption. However, the impact of ethanol on Purkinje cell (PC) mitochondria remains unclear. To investigate how ethanol administration affects mitochondrial dynamics in cerebellar Purkinje cells, we employed a transgenic mouse model expressing mitochondria-targeted yellow fluorescent protein in Purkinje cells (PC-mito-eYFP). Both male and female PC-mito-eYFP mice received an intraperitoneal injection of ethanol or vehicle. One hour after ethanol administration, the animals were perfusion fixed or their cerebellum tissue or isolated mitochondria were collected. Cerebellum sections were analyzed using confocal microscopy to assess changes in mitochondrial length distribution. In vivo superoxide levels were measured using dihydroethidium (DHE), and mitochondrial NAD levels were determined by high-performance liquid chromatography (HPLC). Our findings revealed a sex-dependent response to ethanol administration in mitochondrial size distribution. While male Purkinje cell mitochondria exhibited no significant changes in size, female mitochondria became more fragmented after one hour of ethanol administration. This coincided with elevated phosphorylation of the fission protein Drp1 and increased superoxide production, as measured by DHE fluorescence intensity. Similarly, mitochondrial NAD levels were significantly reduced in female mice, but no changes were observed in males. Our results demonstrate that ethanol induced mitochondrial fragmentation through increased free radical levels, due to reduced NAD and increased p-Drp1, in PC cells of the female cerebellum.

Protective effects of Mycobacterium vaccae ATCC 15483 against "Western"-style diet-induced weight gain and visceral adiposity in adolescent male mice.

The prevalence of noncommunicable inflammatory disease is increasing in modern urban societies, posing significant challenges to public health. Novel prevention and therapeutic strategies are needed to effectively deal with this issue. One promising approach is leveraging microorganisms such as Mycobacterium vaccae ATCC 15483, known for its anti-inflammatory, immunoregulatory, and stress-resilience properties. This study aimed to assess whether weekly subcutaneous administrations of a whole-cell, heat-killed preparation of M. vaccae ATCC 15483 (eleven injections initiated one week before the onset of the diet intervention), relative to vehicle injections, in adolescent male C57BL/6N mice can mitigate inflammation associated with Western-style diet-induced obesity, which is considered a risk factor for a number of metabolic and inflammatory diseases. Our results show that treatment with M. vaccae ATCC 15483 prevented Western-style diet-induced excessive weight gain, visceral adipose tissue accumulation, and elevated plasma leptin concentrations. The Western-style diet, relative to a control diet condition, decreased alpha diversity and altered the community composition of the gut microbiome, increasing the Bacillota to Bacteroidota ratio (formerly referred to as the Firmicutes to Bacteroidetes ratio). Despite the finding that M. vaccae ATCC 15483 prevented Western-style diet-induced excessive weight gain, visceral adipose tissue accumulation, and elevated plasma leptin concentrations, it had no effect on the diversity or community composition of the gut microbiome, suggesting that it acts downstream of the gut microbiome to alter immunometabolic signaling. M. vaccae ATCC 15483 reduced baseline levels of biomarkers of hippocampal neuroinflammation and microglial priming, such as Nfkbia and Nlrp3, and notably decreased anxiety-like defensive behavioral responses. The current findings provide compelling evidence supporting the potential for M. vaccae ATCC 15483 as a promising intervention for prevention or treatment of adverse immunometabolic outcomes linked to the consumption of a Western-style diet and the associated dysbiosis of the gut microbiome.

Systemic inflammation accelerates tau propagation in an experimental model of Alzheimer’s disease

AbstractBackgroundSystemic inflammation in patients with Alzheimer’s disease (AD) has been associated with an exacerbation in cognitive decline, but the underlying mechanisms remain largely unknown. In AD, intraneuronal hyperphosphorylated tau spreads through the brain via trans‐synaptic prion‐like propagation. Evidence suggests that propagation of tau pathology is linked to neuroinflammation. Here we aim to investigate if a low‐grade systemic bacterial infection drives neuroinflammation and exacerbates the spread of tau using an experimental model of ADMethodC57BL/6 mice underwent intracerebral injection of vehicle or AT8‐positive human tau lysate, isolated from post‐mortem AD tissue. Two months post‐injection, mice were given a systemic injection of saline or a life bacterial strain. Body and spleen weight was measured to confirm infection and brain tissue was collected 4 weeks post‐infection to assess tau pathology and markers of neuroinflammation.ResultWe observed tau pathology in mice exposed to human tau lysate. Pathology in the dorsal fornix and the supramammillary nuclei was observed only in mice exposed to a bacterial infection. We also examined the inflammatory markers MHCII and CD64 and detected increased expression in mice exposed to a bacterial infection. The inflammatory response was localised to regions where pathology was observed, such as the CA3‐dentate gyrus axis.ConclusionFollowing intracerebral injection of human tau lysate, propagation of tau was induced both dorsal and ventral to the injection site. The extend of pathological tau was exaggerated by the presence of low‐grade systemic inflammation. This suggests that systemic inflammatory episodes in AD patients may induce greater propagation of tau pathology.

Combination of Haloperidol With UNC9994, β-arrestin-Biased Analog of Aripiprazole, Ameliorates Schizophrenia-Related Phenotypes Induced by NMDAR Deficit in Mice.

Glutamatergic system dysfunction contributes to a full spectrum of schizophrenia-like symptoms, including the cognitive and negative symptoms that are resistant to treatment with antipsychotic drugs (APDs). Aripiprazole, an atypical APD, acts as a dopamine partial agonist, and its combination with haloperidol (a typical APD) has been suggested as a potential strategy to improve schizophrenia. Recently, an analog of aripiprazole, UNC9994, was developed. UNC9994 does not affect dopamine 2 receptor (D2R)-mediated Gi/o protein signaling but acts as a partial agonist for D2R/β-arrestin interactions. Hence, one of our objectives was to probe the behavioral effects of co-administrating haloperidol with UNC9994 in the N-methyl-D-aspartate receptor (NMDAR) mouse models of schizophrenia. The biochemical mechanisms underlying the neurobiological effects of dual haloperidol × UNC9994 action are currently missing. Hence, we aimed to explore D2R- and NMDAR-dependent signaling mechanisms that could underlie the effects of dual drug treatments. NMDAR hypofunction was induced pharmacologically by acute injection of MK-801 (NMDAR pore blocker; 0.15mg/kg) and genetically by knockdown of Grin1 gene expression in mice, which have a 90% reduction in NMDAR levels (Grin1 knockdown [Grin1-KD]). After intraperitoneal injections of vehicle, haloperidol (0.15mg/kg), UNC9994 (0.25mg/kg), or their combination, mice were tested in open field, prepulse inhibition (PPI), Y-maze, and Puzzle box. Biochemical effects on the phosphorylation of Akt, glycogen synthase kinase-3 (GSK-3), and CaMKII in the prefrontal cortex (PFC) and striatum of MK-801-treated mice were assessed by western blotting. Our findings indicate that low dose co-administration of UNC9994 and haloperidol reduces hyperactivity in MK-801-treated animals and in Grin1-KD mice. Furthermore, this dual administration effectively reverses PPI deficits, repetitive/rigid behavior in the Y-maze, and deficient executive function in the Puzzle box in both animal models. Pharmacological inhibition of NMDAR by MK-801 induced the opposite effects in the PFC and striatum on pAkt-S473 and pGSK3β-Ser9. Dual injection of haloperidol with UNC9994 reversed MK-801-induced effects on pAkt-S473 but not on pGSK3β-Ser9 in both brain structures. The dual administration of haloperidol with UNC9994 at low doses represents a promising approach to ameliorate symptoms of schizophrenia. The combined drug regimen elicits synergistic effects specifically on pAkt-S473, suggesting it as a potential biomarker for antipsychotic actions.

Histone deacetylase 6 inhibition prevents hypercholesterolemia-induced erectile dysfunction independent of changes in markers of autophagy.

Erectile dysfunction is a condition with a rapidly increasing prevalence globally with a strong correlation to the increase in obesity and cardiovascular disease rates. The aim of the current study is to investigate the potential role of tubacin, a histone deacetylase 6 (HDAC6) inhibitor, in restoring erectile function in a hypercholesterolemia-induced endothelial dysfunction model. Thirty-nine male C57Bl/6J mice were divided into 3 groups. Two groups were administered an adeno-associated virus encoding for the gain of function of proprotein convertase subtilisin/kexin type 9 (PCSK9) and placed on a high-fat diet (HFD) with 1.25% cholesterol added for 18weeks in order to induce a prolonged state of hypercholesterolemia. One of the PCSK9 groups received daily intraperitoneal injections of the HDAC6 inhibitor tubacin, while the other 2 groups received daily vehicle injections. Erectile function was assessed through measurement of intracavernosal pressure and mean arterial pressure during cavernous nerve stimulation, as well as assessment of agonist-stimulated ex vivo relaxation of the corpus cavernosum (CC). Western blotting was performed from CC tissue samples. Erectile and endothelial functions were assessed, as well as protein markers of mitochondrial dynamics, mitophagy, and autophagy. Erectile function was impaired in the HFD + PCSK9 group throughout the entire voltage range of stimulation. However, the HFD + PCSK9 mice that were treated with tubacin experienced significant restoration of erectile function at the medium and high voltages of nerve stimulation. Similarly, ex vivo CC relaxation responses to acetylcholine and the cystathionine γ-lyase (CSE) substrate L-cysteine were reduced in the vehicle-treated HFD + PCSK9 mice, both of which were restored in the HFD + PCSK9 mice treated with tubacin. Corpus-cavernosum protein expression of CSE was significantly elevated in the tubacin-treated HFD + PCSK9 mice relative to both other groups. There were no significant differences observed in any of the protein markers of mitochondrial dynamics, mitophagy, or autophagy investigated. Histone deacetylase 6 inhibition may protect against erectile and endothelial dysfunction associated with hypercholesterolemia. This was the first study to investigate HDAC6-specific inhibition for treatment of erectile dysfunction. A study limitation was the exclusive focus on the CC, rather than structure and function of the pre-penile arteries that may develop a substantial atherosclerotic plaque burden under hypercholesterolemic conditions. Tubacin may prevent hypercholesterolemia-induced erectile dysfunction through a hydrogen sulfide-related mechanism unrelated to regulation of mitophagy or autophagy.

Teriparatide administration is osteoanabolic but does not impact atherosclerotic plaque calcification and progression in a mouse model of menopause

Menopause exacerbates osteoporosis and increases the risk of atherosclerotic plaque rupture, leading to cardiovascular mortality. Osteoporotic women are increasingly treated with teriparatide (TPTD, 1–34 parathyroid hormone), one of the few treatments that stimulate bone formation. Despite the fact that atherosclerotic plaque calcification is a hallmark of plaque development, the impact of TPTD administration on plaque calcification remain unclear. In this context, we sought to determine the effects of TPTD administration on atherosclerosis in ovariectomized (OVX) apolipoprotein E deficient mice (ApoE−/−), as a model of postmenopausal osteoporosis. OVX ApoE−/− mice, fed a high fat, high cholesterol diet to induce atherosclerosis, received either vehicle or TPTD daily injections (40 μg/kg/d) for 4 or 10 weeks, at which points plaques are respectively weakly and heavily calcified. After sacrifice, bone remodeling was evaluated by serum markers and bone histomorphometry. Bone architectural parameters were measured by μCT. Aortic plaques were analyzed histologically, and their calcification with von Kossa staining and the calcium tracer Osteosense. Plaque inflammation and calcification markers were measured by RT-qPCR. Intermittent TPTD increased bone volume in OVX mice, due to a higher stimulation of bone formation relatively to bone resorption. These effects were not accompanied by changes in serum levels of cholesterol, triglycerides, glucose or insulin. TPTD neither significantly affected aortic plaque size, inflammation, and calcification, even if it slightly increased vascular smooth muscle cell transdifferentiation into calcifying cells. In conclusion, TPTD exhibits osteoanabolic effects in OVX ApoE−/− mice, without significantly influencing atherosclerotic plaque progression or calcification in the short term.

ZLN005 Reduces Neuroinflammation and Improves Mitochondrial Function in Mice with Perioperative Neurocognitive Disorders.

The decrease expression of PGC-1α contributes to perioperative neurocognitive disorders (PND). This study aimed to investigate the effects of the PGC-1α agonist ZLN005 in preventing PND and to explore the potential mechanism. C57BL/6 mice were randomly divided into four groups: the control group (Group C), the surgery group (Group S), the surgery and ZLN005 (5 mg/(kg⋅d)) group (Group L), and the surgery and ZLN005 (7.5 mg/(kg⋅d)) group (Group H). Except for Group C, the other three groups received intraperitoneal injections of vehicle or ZLN005 once a day from 3 days before surgery to 3 days after surgery. The open field test, novel object recognition test and fear conditioning test were performed to measure anxiety behaviors, locomotor activity and memory. The levels of IL-6 and IL-1β were measured at 24hours after surgery. ATP and ROS levels were measured at 3 days post-surgery. PGC-1α, NRF-1, Atp5d, Atp5k and Cox5a were measured at one day or three days post-surgery. ZLN005 treatment improved the cognitive function of mice in Group L and Group H compared with Group S. The expression of IL-6 and IL-1β in the hippocampus of the S group was increased after surgery, and ZLN005 reduced the expression of IL-6 and IL-1β in the hippocampus of mice one day after surgery. There were parallel decreases in the expression of PGC-1α/NRF-1 and mitochondrial function in the hippocampus of the Group S mice compared with the Group C mice. The expression of PGC-1α/NRF-1 and mitochondrial function were upregulated after ZLN005 treatment. Neuroinflammation and mitochondrial damage are involved in the occurrence of PND. ZLN005 activates PGC-1α to increase the expression of mitochondrial proteins, improve mitochondrial function, and ultimately ameliorate the cognitive status of mice after surgery.

Thyroid Hormone Supplementation Restores Cognitive Deficit, Insulin Signaling, and Neuroinflammation in the Hippocampus of a Sporadic Alzheimer's-like Disease Rat Model.

Thyroid hormones play a crucial role in the development of the central nervous system and are considered pivotal to cognitive functions in the adult brain. Recently, thyroid dysfunction has been associated with Alzheimer's disease. The aim of this study was to assess the neuroprotective effects of triiodothyronine (T3) on insulin signaling, neuroinflammation, apoptosis, and cognitive function in a streptozotocin (STZ)-induced sporadic Alzheimer's disease-like model. Male Wistar rats underwent stereotaxic surgery for intracerebroventricular injections of streptozotocin (STZ; 2 mg/kg) or vehicle in the lateral ventricles to induce an AD-like model. The animals received a daily dose of 1.5 μg of T3/100 g body weight or the same volume of vehicle for 30 days and were subdivided into four experimental groups: (1) animals receiving citrate treated with saline (Control = CTL); (2) animals receiving citrate treated with T3 (T3); (3) animals receiving STZ treated with saline (STZ); and (4) animals receiving STZ treated with T3 (STZ + T3). The novel object recognition test was used to measure cognitive function. Serum analysis, real-time RT-PCR, immunohistochemistry, and immunoblotting analyses were also carried out. Our results demonstrated that T3 treatment reversed cognitive impairment and increased Akt and GSK3 phosphorylation in the treated group, while also reducing microglial activation (Iba-1) and GFAP expression (reactive astrocytes), along with TNF-α, IL-6, and IL-1β levels in the hippocampus. Additionally, T3 treatment increased levels of the anti-apoptotic protein Bcl-2 and reduced the expression of the pro-apoptotic protein BAX in the hippocampus. Our study demonstrated that T3 could potentially protect neurons in an AD model induced by STZ.

Moringa oleifera Lam. seed lectin (WSMoL) reduces chronic stress-induced anxiety and depression in mice by lessening inflammation and balancing brain chemicals

Phyto-based treatments for anxiety and depression are gaining attention. The efficacy of the water-soluble Moringa oleifera seed lectin (WSMoL) in reducing acute anxiolytic and depressive-like behaviors in mice has been previously demonstrated. In the present study, it was evaluated the effects of WSMoL on reducing anxiety and depressive-like symptoms in a mouse model of unpredictable chronic mild stress (UCMS). The animals were divided into groups and exposed to a four-week UCMS regimen. Following this, the mice received daily intraperitoneal injections of vehicle (non-stressed and UCMS control groups), WSMoL (2 or 4 mg/kg), or fluoxetine (10 mg/kg) for 21 days. Neurobehavioral tests included the open field test and elevated plus maze test to assess anxiety-like behavior, and the tail suspension test and sucrose preference test to evaluate depression-like behavior. Biochemical analyses measured serum corticosterone and cytokines as well brain levels of cytokines and monoamines. All tests indicated that WSMoL significantly (p < 0.05) reversed the anxiety and depression-like behaviors induced by UCMS. The stress protocol increased serum corticosterone levels and WSMoL treatment was not able to normalize corticosterone secretion. WSMoL treatment reduced serum and brain levels of IL-2, IL-6, and TNF-α, indicating reduced neuroinflammation, and increased brain levels of dopamine, serotonin, and noradrenaline. In summary, WSMoL mitigated UCMS-induced anxiety and depression-like behaviors by reducing neuroinflammation and modulating brain monoamine levels.

Signature of diabetic cardiomyopathy progression using high-fat diet and streptozotocin mouse model: from subclinical to clinical with three-hit mouse model

Abstract Background Diabetic cardiomyopathy (DbCM) is a pre-heart failure (pre-HF) condition that usually exhibits early diastolic dysfunction and evolving systolic dysfunction, along with left ventricular (LV) remodelling. Although DbCM shows distinct pathophysiology, the transition from the subclinical to the symptomatic stage is not straightforward and usually involves multifaceted processes which are not fully understood. Purpose To define the signature of DbCM progression in a novel mouse model using longitudinal echocardiography and multi-omics approaches. Methods Male mice (C57BL/6) were fed with high-fat diet (HFD) or control diet (CD) for 24 weeks, with single dose of streptozotocin (STZ) or vehicle injection at 8 weeks (n=10 each). Serial functional and metabolic parameters were measured prior to endpoint single nuclei RNA sequencing (snRNA-seq) and mass spectrometry-based proteomics analysis with comparison to cardiac remodelling to define detailed mechanisms underlying DbCM progression. Results HFD/STZ mice developed type 2 diabetes mellitus (T2DM) based on higher fasting blood glucose and HbA1C levels, insulin resistance and decreased beta-cell function (HOMA-IR and HOMA-beta, respectively) compared to controls. Progressive diastolic dysfunction was evident in HFD/STZ mice compared to CD mice based on enlarged LVPW;d from 12 weeks (P&amp;lt;0.05), reduced MV E/A ratio (P&amp;lt;0.01) and IVRT (P&amp;lt;0.05) from 16 weeks, and increased cardiac filling pressure (Left atrial (LA) area and LA volume) at 24 weeks, with preserved systolic function. This was paralleled by elevated collagen deposition (Picrosirius red), myocyte cross-sectional area and Col1a1 expression in HFD/STZ versus CD mice (all P&amp;lt;0.05). snRNA-seq analysis indicated preferential monocyte migration into diabetic hearts (P&amp;lt;0.01) whilst inflammatory pathways were activated across all cardiac cell types. Proteomics analysis revealed elevated expression of the established inflammation marker, plasma C-reactive protein (CRP), in HFD/STZ versus CD mice (P&amp;lt;0.05), which was correlated with worsening diastolic function (MV E/A ratio: r=-0.839, P&amp;lt;0.01) and LV hypertrophy (LVPW;d: r=0.861, P&amp;lt;0.01). Conclusion Our HFD/STZ mouse model offers a ‘three-hit’ approach which effectively captures key stages of clinical DbCM progression: 1) T2DM with insulin resistance, 2) progressive diastolic dysfunction with elevated chronic filling pressure and cardiac remodelling, and 3) systemic inflammation. This improved HFD/STZ mouse model is therefore appropriate to support a detailed study of mechanisms underlying DbCM progression and subsequent translation.

Daily Subcutaneous injections of Relaxin is a therapy for heart failure with preserved ejection fraction

Abstract Background and Significance: Relaxin (RLX), a hormone of pregnancy has engendered a great deal of excitement as a therapy for cardiovascular diseases. Clinical trials treated patients with acute decompensated Heart Failure (HF) with RLX (i.v. 30µg/kg/day-2 days) and reported a 37% decrease in mortality during a 6-months follow up. This led the FDA to declare RLX a ‘break-through’ therapy. However, a larger trial failed to meet the sought-after end points. Here, we propose that periodic subQ RLX pulses are more effective. We measured the pharmacokinetics of serum RLX following subQ injections of RLX in Sprague Dawley rats. Further, we treated rats with confirmed HFpEF with daily subQ RLX (2-weeks) to reverse the diastolic dysfunction (DD), arrhythmia and fibrotic profile. Methods Sprague-Dawley (250 g, n=20, either sex) received 2-subQ injections 12 hrs apart of RLX (from Relaxera) or the vehicle. Serial blood samples were drawn following each injection to measure [RLX]. ZSF1 rats (9-weeks old, either sex) were placed on a high fat diet (HFD) and serial echos tracked the DD. Once severe DD was established (25-35 weeks of HFD), rats received daily subQ injections of RLX (100µg/kg) or vehicle. After 2-weeks, hearts were perfused to optically map action potentials and Ca2+ transients, and analyze the arrhythmia profile. IF and Westerns were used to measure changes in fibrosis (collagen 1), Nav1.5, connexin 43, Wnt1 and β-catenin. Results After an injection, RLX reached a peak in ~60 min, fell to 50% in 5-6 hours. Injections of 0, 30, 100 or 500 µg/kg yielded peak levels of 0, 11.26±3.52, 58.33±16.10, and 209.42±29.04 ng/ml and residual levels after 24-hrs of 0, 4.9, 45.1 and 156 pg/ml, respectively. The 30µg/kg injections had no effect and 100 µg/kg increased Nav1.5 (25%), Cx43 (30%) and β-catenin (90%). The 500 µg/kg injections increased Nav1.5 and Cx43 but upregulated β-catenin, only slightly. In ZSF1 HFpEF rats that received the vehicle (n=12), a premature stimulus (at 30 ms) elicited arrhythmia in 67% of the hearts. RLX injections reversed DD, suppressed sustained atrial and ventricular arrhythmias (n=0/12) but not self-terminating arrhythmias, lasting &amp;lt; 10sec. RLX improved the conduction velocity (CV), particularly at fast rates (100ms) from 0.74 to 0.9 m/s (p&amp;lt;0.001, n=12). RLX significantly increased Cx43 expression, Nav1.5 and β-catenin at intercalated disks. RLX reduced collagen deposition in HFpEF rats, to normal levels and increased Wnt1 compared to control HFpEF rats. Conclusions The ZSF1 diabetic rat on a HFD recapitulates human HFpEF, high blood pressure, fibrosis, DD, AF and VF. SubQ RLX injections reversed DD i.e. enlarged Left Atrium, E/e’ ratio, fibrosis, and the arrhythmia vulnerability by increasing CV, Cx43, and Nav1.5. Daily subQ RLX injections were highly effective as a therapy for HFpEF.Echo parameterHFpEF + RLX

Inhibition of CB1R in the Hypothalamic Paraventricular Nucleus Ameliorates Hypertension Through Wnt/β-Catenin/RAS Pathway.

The hypothalamic paraventricular nucleus (PVN), as an important integrating center, plays a prominent role in the pathogenesis of hypertension, in maintaining the stability of cardiovascular activity through peripheral sympathetic nervous activity and secretion of various humoral factors. Acknowledging that the mechanistic targets of the endocannabinoid type 1 receptor (CB1R) are the key signaling systems involved in the regulation of hypertension, we sought to clarify whether inhibition of CB1R within the PVN ameliorates hypertension through Wnt/β-catenin/RAS pathway. Spontaneously hypertensive rats (SHRs) and Wistar Kyoto rats were randomly assigned to different groups and treated with bilateral PVN injections of AM251 (CB1R antagonist, 10µg/h) or vehicle (artificial cerebrospinal fluid, aCSF) for four weeks. Bilateral PVN injections of AM251 significantly decreased the heart rate, the body weight and the mean arterial pressure in SHRs. AM251 lowered the expression of CB1R, Wnt3, active-β-catenin, p-IKKβ, RAS components, pro-inflammatory cytokines and elevated the expression level of Glycogen synthase kinase3β and Superoxide Dismutase in the PVN of hypertensive rats. Our findings suggest that inhibition of CB1R in the PVN ameliorates hypertension through Wnt/β-catenin/RAS pathway and broaden our current understanding of the pathological mechanism and clinical treatment of hypertension.