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Chronic kidney disease (CKD), especially when requiring kidney replacement therapy (hemodialysis (HD) and peritoneal dialysis (PD)), is associated with extracellular water expansion with increased total body sodium. Sodium can also be accumulated in tissues independently of extracellular water. Sodium-23 magnetic resonance imaging (23Na MRI) can quantify the concentration of sodium nuclei in tissues. Applied to the human leg, quantification of tissue sodium concentrations mainly at the skin and muscle level is possible. We hypothesized that increased tissue sodium concentrations exert toxic effects in CKD and dialysis. We aimed to (1) compare tissue sodium concentrations in adults, children and adolescents with CKD, HD and PD against healthy individuals, (2) identify their predictors, (3) understand their connection to left ventricular structure in patients receiving HD and (4) observe their relationship with adverse cardiovascular events and mortality in patients receiving HD and PD. Tissue sodium concentrations were increased in adult patients receiving HD and PD, with respect to healthy individuals. In children and adolescents with CKD, tissue sodium concentrations varied depending on CKD etiology, suggesting kidney disorders can be associated with either sodium wasting or accumulation. Tissue sodium concentrations were associated with older age and overweight in healthy individuals; in patients with CKD and receiving dialysis, hypoalbuminemia was an important predictor of tissue sodium concentrations. In pediatric CKD, proteinuria was also positively associated with tissue sodium concentrations, suggesting a role of proteinuria in increased sodium reabsorption. In patients receiving HD, dialysate sodium concentration was strongly positively associated with skin sodium concentrations. In patients receiving HD, tissue sodium concentrations were associated with left ventricular hypertrophy and dilatation, with tissue sodium concentrations being highest in patients with a dilated heart. Skin sodium concentration was associated with major cardiovascular adverse events and mortality in patients receiving HD or PD. These findings expand our understanding on the toxicity of sodium in CKD and point out critical issues of current dialysis treatment practices. Future studies should explore improved technical applications of 23Na MRI and experiment with new therapies and treatment strategies to reduce tissue sodium concentration in patients with CKD and those receiving dialysis.

Fibrosis is a progressive and typically irreversible disease process characterized by the excessive deposition of collagen in organs and in tissues of the musculoskeletal (MSK) system1,2. This process, which causes loss of organ and tissue function, can be initiated by micro-traumas3, an excessive and/or prolonged immune response1, the activation and proliferation of fibrosis-inducing progenitor cells4, and a pro-fibrotic extra-cellular microenvironment5. In parallel with the events that initiate fibrosis, genetic or environmental influences may cause cells and tissues to become predisposed to fibrosis development prior to initiation. This suggests that these cells and tissues are in a state of prodromal disease6, 7, i.e. a hypersensitive state that is distinct from both active fibrosis and normal homeostasis, where cells and tissues are “primed” for fibrosis development. The ability to detect cells or tissues in this state would be extremely valuable in the clinic, as it would identify patients at risk of fibrosis and facilitate pre-emptive therapeutic interventions before irreversible tissue damage has occurred. Investigation into Duchenne Muscular Dystrophy (DMD) mice models can provide valuable insights into fibrosis. Through analysis and comparison of RNA sequence data from 8-week-old pre-fibrotic mdx mice (dystrophin-/-/utrophin+/+) and wildtype mice (dystrophin+/+/utrophin+/+) potential bio markers for fibrosis can be identified. Future research can be directed towards these bio markers in patients predisposed to or suffering from DMD.

Type 1 Diabetes Mellitus (T1DM) is an autoimmune disorder that results in insufficient endogenous insulin production. Regular exercise has numerous health benefits for individuals with T1DM, however, most insulin-dependent diabetics avoid physical activity due to the fear of exercise-induced hypoglycemia (low blood glucose/BG). The risk of hypoglycemia in this population may be partly due to lower liver glycogen stores which is a major source of blood glucose during exercise. However, the mechanism that leads to lower glycogen stores in T1DM is unknown. The purpose of this study was to determine the effect of an acute bout of moderate-intensity aerobic exercise on: Glycogen storage in the liver and skeletal muscle of T1DM rats. Content of gluconeogenic enzymes including Glucose 6-Phosphatase (G6Pase) and Phosphoenolpyruvate Carboxykinase (PEPCK) in T1DM rats.

Regular exercise has shown to benefit the health of individuals with type 1 diabetes mellitus (T1DM). However, a barrier to regular exercise for this population is the fear of low blood glucose (BG) levels, also known as hypoglycemia. Hypoglycemia can result in short and long-term side-effects, such as recurring loss of consciousness or in severe cases death. In non-diabetics, sex-related differences in fuel selection during exercise are well established. Women shift towards using fats as fuel whereas men rely mostly on sugars (i.e., carbohydrates) for energy production. Exercise during the luteal phase of the female menstrual cycle, where estrogen levels are elevated, has shown to promote fat utilization. For women with T1DM, exercise during elevated estrogen levels may potentially provide a protective mechanism against hypoglycemia as energy sources such as glycogen (i.e., stored sugar) may be preserved. This may reduce the need for the body to replete energy from the blood post-exercise, thereby keeping BG levels balanced. My summer project aimed to investigate this phenomenon.

G protein-coupled receptors (GPCRs) are proteins that are important in physiological regulatory processes within the body, and for this reason are important drug targets. When bound to an agonist, such as neurotransmitters or hormones, the receptor adopts an active state to allow these biochemical pathways to occur. However, mutations can arise within the receptor that affect its ability to bind its agonist. This natural variation found within the genome can make it difficult to design pharmaceuticals to target the receptors. To see the effects of these point substitutions on agonist-induced receptor activation, mutations were made within a negative allosteric site of the Adenosine Receptor A2A called the sodium ion binding pocket. It was hypothesized that these mutations destabilize the sodium ion, increasing the ability of an agonist to bind the Orthosteric site. These point substitutions were modelled using an online protein visualization program (PyMOL), and any shifts in residue position were noted. An in vitro analysis was then conducted to fully understand this concept, by studying the effects of the transfection of natural human variants into cells. A GloSensor cAMP assay was used, which produced luminescence activity upon receptor activation. Compared to wild type proteins, receptor activity was found to have decreased with mutation S91A. These findings suggest that receptor activation is more complex than originally thought, and that more research must be conducted to better understand these effects on pharmaceuticals.

Both metabolic and cognitive dysfunction can originate from fetal reprogramming precipitating from adverse conditions experienced in utero. Of note is the western diet (WD), which is associated with maternal energy imbalances that may hinder fetal development through altered placental function. Brain-derived neurotrophic factor (BDNF), a growth factor that supports the placenta and developing brain, is responsive to such energy imbalances. This study sought to investigate the impact of lifelong maternal WD consumption on fetoplacental development, focusing on relations between placental changes, and fetal growth and neurodevelopment in a guinea pig model. Maternal WD consumption resulting in a lean metabolically unhealthy maternal phenotype was associated with lean fetal hepatic steatosis. Placentae of these fetuses were large yet inefficient and showed reduced BDNF expression. Similar reductions in BDNF were noted in fetal brains, coinciding with decreased cell density. Such cellular changes may convey long-term cognitive deficits, although their consequences remain unknown.

© 2021, The Author(s), under exclusive licence to Springer-Verlag GmbH, DE part of Springer Nature. Abstract: Glycogen synthase kinase (GSK) 3 acts to negatively regulate multiple signaling pathways, including canonical Wnt signaling. The two mammalian GSK3 proteins (alpha and beta) are at least partially redundant. While Gsk3a KO mice are viable and display a metabolic phenotype, abnormal neuronal development, and accelerated aging, Gsk3b KO animals die late in embryogenesis or at birth. Selective Gsk3b KO in bone delays development of some bones, whereas cartilage-specific Gsk3b KO mice are normal except for elevated levels of GSK3A protein. However, the collective role of these two GSK3 proteins in cartilage was not evaluated. To address this, we generated tamoxifen-inducible, cartilage-specific Gsk3a/Gsk3b KO (described as “cDKO”) in juvenile mice and investigated their skeletal phenotypes. We found that cartilage-specific Gsk3a/Gsk3b deletion in young, skeletally immature mice causes precocious growth plate (GP) remodeling, culminating in shorter long bones and hence, growth retardation. These mice exhibit inefficient breathing patterns at later stages and fail to survive. The disrupted GP in cDKO mice showed progressive loss of cellular and proteoglycan components, and immunostaining for SOX9, while BGLAP (osteocalcin) and COL2A1 increased. In addition, we observed increased osteoclast recruitment and cell apoptosis. Surprisingly, changes in articular cartilage of cDKO mice were mild compared with the GP, signifying differential regulation of articular cartilage vs GP tissues. Taken together, these findings emphasize a crucial role of two GSK3 proteins in skeletal development, in particular in the maintenance and function of GP. Key Messages: • Both GSK3 genes, together, are crucial regulators of growth plate remodeling. • Cartilage-specific deletion of both GSK3 genes causes skeletal growth retardation. • Deletion of both GSK3 genes decreases Sox9 levels and promotes chondrocyte apoptosis. • Cartilage-specific GSK3 deletion in juvenile mice culminates in premature lethality. • GSK3 deletion exhibits mild effects on articular cartilage compared to growth plate.

Maternal diabetes impairs fetal development and increases the risk of metabolic diseases in the offspring. Previously, we showed maternal dietary supplementation with 6% of olive oil prevents diabetes-induced embryo and fetal defects, in part, through the activation of peroxisome proliferator-activated receptors (PPARs). In this study we examined the effects of this diet on neonatal and adult pancreatic development in both male and female offspring. A mild diabetic model was developed by injecting neonatal rats with streptozotocin (90 mg/kg). During pregnancy these dams were fed a chow diet supplemented or not with 6% olive oil. Dual immunohistochemistry was performed to detect α and β cells in islets at post-natal day 2 and at 5 months of age. Morphometric analysis was carried out to determine the number of islets, α and β cell clusters and β-cell mass. At 5 months, male offspring of diabetic mothers had reduced β-cell mass. Moreover, this β-cell pancreatic deficit was prevented by the maternal supplementation with olive oil. While no changes in PPARα expression was detected in the pancreas, both PPARβ/δ and PPARγ expression were reduced in 5-month-old male offspring of diabetic rats. Interestingly, the reduction in PPAR β/δ expression was prevented by maternal olive-oil supplementation. To further explore the direct effects on PPARs, INS-1E (β) and αTC1-6 (α) cell lines were treated with oleic acid. Interestingly PPARβ/δ expression is highly expressed in INS-1E. Collectively, these findings suggest that olive oil supplementation in utero may prevent diabetes-induced β cell loss in postnatal life by modulating pancreatic PPARs.

© 2020, Springer Nature Limited. Research into the molecular genetics of osteoarthritis (OA) has been substantially bolstered in the past few years by the implementation of powerful genome-wide scans that have revealed a large number of novel risk loci associated with the disease. This refreshing wave of discovery has occurred concurrently with epigenetic studies of joint tissues that have examined DNA methylation, histone modifications and regulatory RNAs. These epigenetic analyses have involved investigations of joint development, homeostasis and disease and have used both human samples and animal models. What has become apparent from a comparison of these two complementary approaches is that many OA genetic risk signals interact with, map to or correlate with epigenetic mediators. This discovery implies that epigenetic mechanisms, and their effect on gene expression, are a major conduit through which OA genetic risk polymorphisms exert their functional effects. This observation is particularly exciting as it provides mechanistic insight into OA susceptibility. Furthermore, this knowledge reveals avenues for attenuating the negative effect of risk-conferring alleles by exposing the epigenome as an exploitable target for therapeutic intervention in OA.

Recent reports indicate that 7% of pregnant mothers in North America use cannabis. This is concerning given that in utero exposure to Δ9-tetrahydrocannabinol (Δ9-THC), the main psychoactive component in cannabis, causes fetal growth restriction and may alter replication and survival of pancreatic β-cells in the offspring. Accordingly, we hypothesized that maternal exposure to Δ9-THC during pregnancy will impair postnatal glucometabolic health of offspring. To test this hypothesis, pregnant Wistar rats were treated with daily intraperitoneal injections of either 3 mg/kg Δ9-THC or vehicle from gestational day 6 to birth. Offspring were subsequently challenged with glucose and insulin at 5 months of age to assess glucose tolerance and peripheral muscle insulin sensitivity. Female offspring exposed to Δ9-THC in utero were glucose intolerant, associated with blunted insulin response in muscle and increased serum insulin concentration 15 minutes after glucose challenge. Additionally, pancreata from male and female offspring were harvested at postnatal day 21 and 5 months of age for assessment of endocrine pancreas morphometry by immunostaining. This analysis revealed that gestational exposure to Δ9-THC reduced the density of islets in female, but not male, offspring at postnatal day 21 and 5 months, culminating in reduced β-cell mass at 5 months. These results demonstrate that fetal exposure to Δ9-THC causes female-specific impairments in glucose homeostasis, raising concern regarding the metabolic health of offspring, particularly females, exposed to cannabis in utero.

Epidemiological data suggest an inverse relationship between birth weight and long-term metabolic deficits, which is exacerbated by postnatal catch-up growth. We have previously demonstrated that rat offspring subject to maternal protein restriction (MPR) followed by catch-up growth exhibit impaired hepatic function and ER stress. Given that mitochondrial dysfunction is associated with various metabolic pathologies, we hypothesized that altered expression of p66Shc, a gatekeeper of oxidative stress and mitochondrial function, contributes to the hepatic defects observed in MPR offspring. To test this hypothesis, pregnant Wistar rats were fed a control (20% protein) diet or an isocaloric low protein (8%; LP) diet throughout gestation. Offspring born to control dams received a control diet in postnatal life, while MPR offspring remained on a LP diet (LP1) or received a control diet post-weaning (LP2) or at birth (LP3). At four months, LP2 offspring exhibited increased protein abundance of both p66Shc and the cis-trans isomerase Pin1. This was further associated with aberrant markers of oxidative stress (i.e., elevated 4-HNE, SOD-1 and SOD2, decreased catalase) and aerobic metabolism (i.e., increased phospho-PDH and LDHa, decreased complex II, citrate synthase and TFAM). We further demonstrated that tunicamycin-induced ER stress in HepG2 cells led to increased p66Shc protein abundance, suggesting that ER stress may underlie the programmed effects observed in vivo. In summary, because these defects are exclusive to adult LP2 offspring, it is possible that a low protein diet during perinatal life, a period of liver plasticity, followed by catch-up growth is detrimental to long-term mitochondrial function.

Piezo proteins are transmembrane ion channels, specialized in detecting mechanosensitive stimuli and transduce mechanical forces into biochemical signals. Piezo proteins research has helped understand physiological mechanisms, but the integrative role that Piezo1 plays in the regulation of the microvasculature has remained unstudied. Our main objective was to characterize ex vivo microvascular responses to the blockade of Piezo1 mechanotransduction in male (n=29) and female (n=24) Sprague-Dawley (SD) rats. Gracilis arterioles (GA) and middle cerebral arterioles (MCA) were harvested for ex-vivo vessel preparations. After vessel viability confirmation, every vessel was submitted to myogenic and flow challenges under control conditions and after Grammostola Mechanotoxin 4 (GsMTx4) incubation to blocking Piezo1 channels, to quantify the homeostatic response of arterioles before and after Piezo1 antagonism. We are able to report Piezo1 as indispensable component in vascular smooth muscle cells (VSMC) and Endothelial cells (EC) to sense and change vessel diameter based on intravascular pressure and shear stress, correspondingly. Also, we report for the first time a heterogeneous response in males and females after Piezo1 antagonism in representative resistance arterioles from the skeletal muscle and cerebral circulation.

Adverse in utero and postnatal conditions can increase susceptibility to metabolic syndrome (MS). Altered muscle respiration contributes to MS, but the effects of restricted oxygen and nutrients in utero on skeletal muscle mitochondria remain unknown. In this study guinea pig sows underwent uterine artery ablations mid-gestation, producing fetuses with low birth weight (LBW). Soleus muscle was collected near term or at four months of age, from LBW and control fetuses and offspring, where the offspring were fed either a Western Diet (WD) or a control diet (CD). Soleus muscles from LBW fetuses exhibit lower maximal respiration rates than normal birth weight (NBW) sham-surgery controls. Additionally, LBW/CD, NBW/WD and LBW/WD adult guinea pigs displayed reduced respiration compared with NBW/CD. Cultured C2C12 cells were utilized to better understand independent effects of hypoxia and fatty acid saturation upon cellular respiration. Both chronic (5 days) hypoxia and palmitate (16:0) reduced respiration compared with normoxia.

© 2019 Wiley Periodicals, Inc. Many potential pharmacological treatments for osteoarthritis can result in undesirable side effects due to the systemic administration of drugs, making the direct delivery of drugs to joints an attractive alternative. Poly(ester amide)s (PEAs) have been shown to exhibit promising properties for the development of particle-based intra-articular delivery vehicles. However, a limited range of PEA structures has been investigated. In this study, we prepared and characterized the properties of two different PEA particles composed of l-phenylalanine, sebacic acid, and either 1,4-butanediol or 1,8-octanediol (PBSe and POSe, respectively). The anti-inflammatory drug celecoxib (CXB) was encapsulated into the particles. Despite minor structural differences, PBSe and POSe exhibited different thermal and mechanical properties, and encapsulation of CXB influenced these properties. PBSe-CXB particles provided a slower release of drug in vitro relative to POSe-CXB. Toxicity studies showed that particles without drug exhibited low toxicity to ATDC5 and C2C12 cells, while the PBSe-CXB particles exhibited concentration-dependent toxicity. Host response to the particles was evaluated in an ovine model. No adverse effects were observed following intra-articular injection and it was observed that the particles diffused into the surrounding tissues. This work shows the importance of structural tuning in PEA delivery vehicles and demonstrates their potential for further development. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 1235–1243, 2019.

It is not known why there is increased risk to have seizures with increased anxiety and stress after traumatic brain injury (TBI). Stressors cause the release of corticotropin-releasing factor (CRF) both from the hypothalamic pituitary adrenal (HPA) axis and from CNS neurons located in the central amygdala and GABAergic interneurons. We have previously shown that CRF signaling is plastic, becoming excitatory instead of inhibitory after the kindling model of epilepsy. Here, using Sprague Dawley rats we have found that CRF signaling increased excitability after TBI. Following TBI, CRF type 1 receptor (CRFR1)-mediated activity caused abnormally large electrical responses in the amygdala, including fast ripples, which are considered to be epileptogenic. After TBI, we also found the ripple (120–250 Hz) and fast ripple activity (>250 Hz) was cross-frequency coupled with θ (3–8 Hz) oscillations. CRFR1antagonists reduced the incidence of phase coupling between ripples and fast ripples. Our observations indicate that pathophysiological signaling of the CRFR1increases the incidence of epileptiform activity after TBI. The use for CRFR1antagonist may be useful to reduce the severity and frequency of TBI associated epileptic seizures.

Cellular transplantation strategies have aimed to combat critical limb ischemia (CLI) by inducing endogenous blood vessel regeneration. Transplantation of mesenchymal stromal cells (MSC) for CLI has proven safe and well-tolerated but demonstrated only modestly improved clinical outcomes explained in part by a lack of MSC survival and/or potency in ischemic tissues. This thesis focuses on the development of improved transplantation strategies for patients with CLI utilizing purified pro-angiogenic subsets of MSC and endothelial colony forming cells (ECFC) seeded within decellularized adipose tissue (DAT) bioscaffolds. First, I investigated whether purification of bone marrow-derived MSC based on a conserved stem cell function, high ALDH-activity, could isolate a pro-vascular MSC subset after culture. Conditioned media generated by ALDHhi MSC promoted endothelial cell proliferation, survival, and tube forming functions under serum-free conditions, and a robust recruitment of mouse derived endothelial cells was observed after subcutaneous implantation of ALDHhi MSC within immunodeficient mice. Interestingly, isolating ECFC based on high ALDH-activity did not improve endothelial cell functions. Instead, isolation of ECFC expressing the progenitor cell surface marker, CD34 purified for ECFC with a mature endothelial cell surface phenotype and diminished colony- and tubule-forming functions in vitro. Before transplanting ECFC and MSC within human DAT scaffolds, I sought to develop robust methodologies to extract cells from DAT scaffolds, without generation of autofluorescent debris that impaired flow cytometric cellular evaluations. After protease dissociation, DAT scaffolds were centrifuged using a density gradient to produce samples with >99% purity. This method enabled accurate quantification of changes in ECFC surface marker expression alongside a decline in ECFC proliferation when co-cultured with MSC on DAT scaffolds in vitro. After intramuscular injection, or DAT scaffold implantation of ECFC and/or MSC into NOD.SCID mice with acute unilateral hindlimb ischemia, the level of blood perfusion in the ischemic limb was unaffected compared to saline injection. Importantly, implanted DAT scaffolds became vascularized and enhanced survival of ECFC or MSC. Taken together, this body of work indicates the importance of purifying pro-regenerative subsets to maximize cellular potency for clinical benefit.

Methylphenidate (MPH), commonly known as Ritalin, is the most widely prescribed drug worldwide to treat patients with attention deficit disorders. Although MPH is thought to modulate catecholamine neurotransmission in the brain, it remains unclear how these neurochemical effects influence neuronal activity and lead to attentional enhancements. Studies in rodents overwhelmingly point to the lateral prefrontal cortex (LPFC) as a main site of action of MPH. To understand the mechanism of action of MPH in a primate brain, we recorded the responses of neuronal populations using chronic multielectrode arrays implanted in the caudal LPFC of two macaque monkeys while the animals performed an attention task (N= 2811 neuronal recordings). Over different recording sessions (N= 55), we orally administered either various doses of MPH or a placebo to the animals. Behavioral analyses revealed positive effects of MPH on task performance at specific doses. However, analyses of individual neurons activity, noise correlations, and neuronal ensemble activity using machine learning algorithms revealed no effects of MPH. Our results suggest that the positive behavioral effects of MPH observed in primates (including humans) may not be mediated by changes in the activity of caudal LPFC neurons. MPH may enhance cognitive performance by modulating neuronal activity in other regions of the attentional network in the primate brain.

Fetal growth restriction (FGR) is defined by low birth weight and contributes to a variety of adult-onset diseases with different severities between males and females. However, the effects of FGR on the pulmonary surfactant are not fully elucidated. In this thesis, first, we investigated the FGR effects on the lung function and the surfactant system at the early postnatal life. It was hypothesized that FGR contributes to alterations of lung mechanics and the surfactant system during the neonatal period. Second, we assessed the FGR effects on the surfactant system in response to sepsis in adulthood. It was hypothesized that FGR contributes to the alteration of the surfactant system in response to sepsis in adulthood. Overall, the data suggest that FGR induced by maternal protein restriction affects lung compliance at the early life and predisposes the septic adults to the development of surfactant alterations in a sex-specific manner.

Globally, approximately 10-25% of women smoke during pregnancy. Since nicotine is highly addictive, women may use nicotine containing products like nicotine replacement therapies for smoking cessation, but the long-term consequences of early life exposure to nicotine remain poorly defined. Our laboratory has previously demonstrated that maternal nicotine exposed (MNE) rat offspring exhibit hypertriglyceridemia due to increased hepatic de novo lipogenesis. Hypertriglyceridemia may also be attributed to impaired white adipose tissue (WAT) lipid storage; however, the effects of MNE on WAT are not completely understood. We hypothesize that nicotine-induced alterations in adipose function (e.g. lipid storage) underlie dyslipidemia in MNE adults. Female 6 month old rats exposed to nicotine during gestation and lactation exhibited significantly decreased visceral adipocyte cell area by 40%, attributed, in part, to a 3-fold increase in adipose triglyceride lipase (ATGL) protein expression compared to vehicle. Given ATGL has antioxidant properties and in utero nicotine exposure promotes oxidative stress in various tissues, we next investigated if there was evidence of increased oxidative stress in MNE WAT. At both 3 weeks and 6 months, MNE offspring expressed 37-48% higher protein levels of SOD1 and SOD2 in WAT. Since oxidative stress can induce inflammation, we examined the inflammatory profile of WAT and found increased expression of cytokines (IL-1β, TNFα, and IL-6) by 44-61% at 6 months. Collectively, this suggests that the expression of WAT ATGL may be induced to counter MNE-induced oxidative stress and inflammation. However, higher levels of ATGL would further promote lipolysis in WAT, culminating in impaired lipid storage and long-term dyslipidemia.

Summary Objective Population-based cohort studies suggest an association between osteoarthritis (OA) and cerebrovascular disease, yet the mechanisms underlying vascular comorbidities in OA remain unclear. The purpose of this narrative review is to discuss the literature examining inflammation in OA with a focus on physiological mechanisms, and whether overlapping mechanisms exist in cerebrovascular dysfunction. Method A literature search was conducted in PubMed using combinations of search terms: osteoarthritis , cerebrovascular (disease/dysfunction/risk) , cardiovascular (disease/dysfunction/risk) , aging/ageing , inflammation , inflammatory mediators , cytokine , c-reactive protein , interleukin , advanced glycation end-products , metabolic syndrome , reactive oxidative species , cognitive impairment , (vascular-related) dementia , small cerebral vessel disease , endothelial function , blood–brain barrier , gender/sex , hypertension , peripheral vascular health , and physical activity . Reference lists of identified articles were also researched manually. Results Overlapping inflammatory factors that may contribute to onset and progression of both OA and cerebrovascular dysfunction are presented. We describe oxidative mechanisms involving pro-inflammatory cytokines and oxidative species, advanced glycation end-products, sex hormones, microvascular dysfunction and osteoprotegerin, and their specific roles in potentially contributing to OA and cerebrovascular dysfunction. Conclusion Synthesis of the current literature suggests future investigations may benefit from directly testing cerebrovascular hemodynamics and cognitive function in individuals with or at risk of OA to elucidate common physiological mechanisms.

Background: Bacterial lung infections are leading causes of death worldwide. Unfortunately, increasing resistance to antibiotics and the inflammation often accompanying these infections are leading to poor outcomes despite antibiotic intervention. Complicating treatment further, the tree-like branching structure of the lung makes drug delivery to distal sites of infection difficult. Our research aims to address these challenges by developing new therapeutics and new tools to improve and assess drug delivery, bacterial killing and inflammation. Our therapy combines host defense peptides, which have been shown to kill antibiotic-resistant bacteria and down regulate inflammation, with a pulmonary vehicle, exogenous surfactant, that can improve distribution in the lung. Methods: We have developed the Wet Bridge Transfer system, a new tool which can rapidly assess drug transport, bacterial killing and anti-inflammatory properties of compounds as they spread across a surface. Results: We anticipate that exogenous surfactant will not only increase the transport of host defense peptides, but that the mixture will effectively kill antibiotic-resistant bacteria and reduce inflammation as it spreads. Discussion & Conclusion: Utilizing host defense peptides with a spreading agent represents a novel approach to treating bacterial lung infections. Additionally, the transfer system offers the ability to rapidly screen and examine the next generation of pulmonary therapies. Interdisciplinary Reflection: This project requires interacting with clinicians treating patients with lung infections, biochemists improving the transfer system, and immunologists to understand the underlying mechanism of action for our new therapy. This interdisciplinary environment is essential to making our novel therapy a clinical reality.

Epidemiologic and clinical research has provided a large body of evidence supporting the developmental origins of health and disease (DOHaD), but there has been a relative dearth of mechanistic studies in humans due to the complexity of working with large, longitudinal cohorts. Nonetheless, animal models of undernutrition have provided substantial evidence for the potential epigenetic, metabolic, and endocrine mechanisms behind DOHaD. Furthermore, recent research has explored the interaction between the environment and the gastrointestinal system by investigating how the gut microbial ecology may impact the capacity for nutrient processing and absorption in a manner that may limit growth. This review presents a summary of current research that supports the concept of DOHaD, as well as potential mechanisms and interactions that explain how nutrition in utero and during early childhood influences lifelong health.

Cellular therapy to induce islet regeneration is emerging as a novel treatment strategy for diabetes. Umbilical cord blood (UCB)-derived hematopoietic stem/progenitor cells (HSPC) isolated by high aldehyde dehydrogenase activity (ALDHhi) reduce hyperglycemia after transplantation into streptozotocin (STZ)-treated NOD/SCID mice. However, UCB-derived ALDHhi cells are rare and expansion without the loss of regenerative function is required. We hypothesized that BMS 493, an inverse retinoic acid receptor agonist, will prevent HSPC differentiation of HSPC during expansion, generating more ALDHhi cells for therapy. ALDHhi cells expanded for 6 days with BMS 493 showed a 2.70-fold-increase in ALDHhi cells compared to untreated cells. Conditioned media from BMS 493-treated cells also increased human ß-cell proliferation in vitro. However, intrapancreatic transplantation of BMS 493-treated cells did not reduce hyperglycemia in STZ-treated NOD/SCID mice. Further characterization of HSPC expansion without differentiation is required for islet regenerative therapies.

Complete occlusion of a coronary artery causes myocardial infarction (MI), resulting in cardiomyocyte cell death. The surviving myocardium undergoes a deleterious remodeling process which causes further injury, and can ultimately result in heart failure. Despite therapeutic advances that have prolonged life, MI remains a leading cause of death worldwide and imparts a significant economic burden. The advancement of treatments to improve cardiac repair post-MI requires the discovery of new targeted treatment strategies. Epicardial-to-mesenchymal transition (EMT) occurs post-MI as a mechanism to support neovascularization and cardiac healing. However, the endogenous EMT is not enough to support sufficient repair. The transcription factor Wilms’ tumor 1 (Wt1) and the primary cilium regulate EMT. Furthermore, the long non-coding RNA (lncRNA) Malat1 is not only known to regulate EMT, but also mitigates cell death from injury. Little is known about the roles of Wt1, primary cilia, and Malat1 in relation to neonatal cardiac regeneration and adult cardiac repair post-MI. The aim of thesis was to investigate cardiac repair mechanisms related to EMT, primary cilia, and Malat1. To study the effects of deficiency of Wt1 and Malat1, a reproducible model of cardiac regeneration using coronary artery ligation to induce MI was developed in neonatal mice. We demonstrated the robust capacity for neonatal cardiac regeneration as neither Wt1 nor Malat1 deficiency affected the regenerative response. In the adult mouse, MI was induced to investigate the role of the primary cilium in EMT and repair post-MI, and the effects of Malat1 deficiency on acute MI pathology. We found that indeed epicardial cells are ciliated, and primary ciliary disassembly supports EMT post-MI, which enhances the contribution of epicardium-derived cells (EPDCs) to neovascularization. This reduced deleterious cardiac remodeling and preserved cardiac function. Contrastingly, Malat1 deficiency augmented cardiac cell death from MI. Unexpectedly, rather than affecting apoptosis, Malat1 deficiency augmented necroptotic cell death post-MI. Overall, this thesis is the first to show that in the adult heart, the primary cilium regulates epicardial EMT and the lncRNA Malat1 governs the propensity for necroptosis from ischemic injury. Both the primary cilium and Malat1 may serve as targets in the development of therapeutics to treat MI in the future.

Pregestational diabetes is a risk factor for birth defects, with diabetic women having a four times higher risk of producing children with congenital heart defects (CHDs). This is concerning because the prevalence of diabetes is growing at epidemic proportions. The purpose of this thesis is to investigate whether exercise can also reduce hyperglycemia-induced CHDs and determine the underlying mechanism. Endothelial nitric oxide synthase (eNOS) is a critical enzyme for normal cardiogenesis that can be upregulated with exercise in adult cardiovascular tissue. However, whether maternal exercise can regulate fetal eNOS and affect heart development has yet to be studied. In this thesis, a mouse model of streptozotcin-induced diabetes was used to generate a hyperglycemic gestational environment. Incidence of CHDs in offspring of diabetic mice was significantly reduced by maternal voluntary exercise from 63 to 31%. Fetal hearts also showed improved aortic and pulmonary valve remodeling compared to the non-exercised diabetic group. Diabetes-induced defects in coronary artery volume as well as capillary density were also rescued with exercise. Myocardial cell proliferation and epithelial-mesenchymal transition at E12.5 was significantly decreased with pregestational diabetes but recovered with maternal exercise. Embryonic gene expression of eNOS, as well as cardiogenic transcription factor Gata4 was restored to control levels with exercise in diabetic mice. These findings provide evidence that exercise may be a feasible intervention to lower the incidence of CHDs in the offspring of women with pregestational diabetes.

Smoking throughout pregnancy can lead to complications during gestation, parturition and neonatal development. Thus, nicotine replacement therapies are a popular alternative thought to be safer than cigarettes. However, recent studies in rodents suggest that fetal and neonatal nicotine exposure alone results in cardiac dysfunction and high blood pressure. While it is well known that perinatal nicotine exposure causes increased congenital abnormalities, the mechanisms underlying longer-term deficits in cardiac function are not completely understood. Recently, our laboratory demonstrated that nicotine impairs placental protein disulfide isomerase (PDI) triggering an increase in endoplasmic reticulum stress, leading us to hypothesize that this may also occur in the heart. At 3 months of age, nicotine-exposed offspring had 45% decreased PDI levels in the absence of endoplasmic reticulum stress. Given the association of PDI and superoxide dismutase enzymes, we further observed that antioxidant superoxide dismutase-2 levels were reduced by 32% in these offspring concomitant with a 26-49% decrease in mitochondrial complex proteins (I, II, IV and V) and tissue inhibitor of metalloproteinase-4, a critical matrix metalloprotease for cardiac contractility and health. Collectively, this study suggests that perinatal nicotine exposure decreases PDI, which can promote oxidative damage and mitochondrial damage, associated with a premature decline in cardiac function.

Up to 10% of women take selective serotonin reuptake inhibitors (SSRI) during pregnancy. Children exposed to SSRIs in utero have an increased risk of being overweight suggesting that fetal exposure to SSRIs can cause permanent metabolic changes. We have previously shown in rats that fetal and neonatal exposure to the SSRI antidepressant fluoxetine results in metabolic perturbations including increased hepatic triglyceride content; a hallmark of non-alcoholic fatty liver disease (NAFLD). Therefore, the aim of this study was to identify the mechanism(s) underlying the fluoxetine-induced increase in intrahepatic triglyceride content. Female nulliparous Wistar rats were given vehicle or fluoxetine (10 mg/kg/day) orally for 2 weeks prior to mating until weaning. At 6 months of age, we assessed whether SSRI exposure altered components of the hepatic triglyceride biosynthesis pathway in the offspring and examined the molecular mechanisms underlying these changes. Male SSRI-exposed offspring had a significant increase in the steady-state mRNA levels of Elovl6 and Dgat1 and core components of the NLRP3 inflammasome (apoptosis-associated speck-like protein containing a caspase activation recruitment domain [ASC] and caspase-1). Augmented expression of Asc in the SSRI-exposed offspring coincided with increased histone acetylation in the proximal promoter region. Given that we have previously demonstrated that antenatal exposure to SSRIs can lead to fatty liver in the offspring, this raises concerns regarding the long-term metabolic sequelae of fetal SSRI exposure. Moreover, this study suggests that elevated hepatic triglyceride levels observed in the SSRI-exposed offspring may be due, in part, to activation of the NLRP3 inflammasome and augmentation of de novo lipogenesis.

Aberrant brain development in utero accompanied by fetal growth restriction (FGR) increases the risk of neurodevelopmental disorders in later life. However there are limited non-invasive biomarkers in the brain for the early identification of said neurodevelopmental disorders in an animal model of FGR. Guinea pig sows were fed either ad libitum (Control) or 70% of the control diet pre-pregnancy, increasing to 90% at mid-pregnancy (MNR) creating appropriately grown (AGA) Control and FGR-MNR neonates, respectively. Three to four weeks corrected post-natal age, neonates were imaged using magnetic resonance imaging (MRI) and spectroscopy (MRS) techniques, and were killed 48-72 hours later for histological analysis. FGR-MNR neonates had smaller brain weights, whole brain volume, hippocampal volume and lateral ventricle volume, which correlate to histological findings. While there is a reduction in the hippocampal volume, there are no differences in hippocampus metabolite ratios between the AGA-Control and FGR-MNR neonates. Interestingly, there was a reduction in the width of the stratum oriens and stratum radiatum in the hippocampus proper, as well as the width of the polymorphic layer in the dentate gyrus, with no changes in pyramidal and granule cell number in the FGR-MNR neonates compared to AGA-Control neonates. In conclusion, MNR in guinea pigs produces FGR neonates that display catch up growth and structural differences in the brains while no changes in the metabolite levels in the hippocampal region of the brain. Together these results involve MRI and MRS as reliable imaging tools to detect the presence of brain injury for the future use of biomarkers for neurodevelopmental disorders and potential therapeutic interventions.

// James B. Greenaway 1 , Carl Virtanen 2 , Kata Osz 1 , Tamas Revay 1 , Daniel Hardy 3 , Trevor Shepherd 4 , Gabriel DiMattia 4 , Jim Petrik 5 1 Department of Biomedical Sciences, University of Guelph, Guelph, ON, N1G 2W1, Canada 2 Princess Margaret Genomics Centre, University Health Network, Toronto, ON, M5G 1L7, Canada 3 Department of Ob/Gyn and Physiology and Pharmacology, Children’s Health Research Institute, Western University, London, ON, N6A 5C1, Canada 4 Department of Ob/Gyn and Oncology, Anatomy and Cell Biology, London Regional Cancer Program, Western University, London, ON, N6A 4L6, Canada 5 Department of Biomedical Sciences, University of Guelph, Guelph, ON, N1G 2W1, Canada Correspondence to: Jim Petrik, email: jpetrik@uoguelph.ca Keywords: ovarian cancer, mevalonate pathway, simvastatin, p53 Received: January 12, 2016 Accepted: June 04, 2016 Published: June 17, 2016 ABSTRACT Epithelial ovarian cancer (EOC) is the most lethal gynecological cancer and often is not detected until late stages when cancer cells transcoelomically metastasize to the abdomen and typically become resistant to therapy resulting in very low survival rates. We utilize an orthotopic, syngeneic mouse model to study late stage disease and have discovered that the tumor cells within the abdominal ascites are irreversibly re-programmed, with an increased tumorigenicity and resistance to apoptosis. The goal of this study was to characterize the reprogramming that occurred in the aggressive ascites-derived cells (28-2 cells) compared to the original cell line used for tumor induction (ID8 cells). Microarray experiments showed that the majority of genes upregulated in the 28-2 cells belonged to the mevalonate pathway, which is involved in cholesterol biosynthesis, protein prenylation, and activation of small GTPases. Upregulation of mevalonate appeared to be associated with the acquisition of a p53 mutation in the ascites-derived cells. Treatment with simvastatin to inhibit HMG CoA reductase, the rate limiting enzyme of this pathway, induced apoptosis in the 28-2 cell line. Rescue experiments revealed that mevalonate, but not cholesterol, could inhibit the simvastatin-mediated effects. In vivo , daily intraperitoneal simvastatin treatment significantly regressed advanced stage disease and induced death of metastatic tumor cells. These data suggest that ovarian cancer cells become reprogrammed, with genetic mutations, and upregulation of the mevalonate pathway, which facilitates the development of advanced stage disease. The use of statins to inhibit HMGCR may provide novel therapeutic opportunities for the treatment of advanced stage EOC.

Nicotine exposure during pregnancy leads to adverse health outcomes, including compromised placental development. Although the molecular mechanisms remain elusive, recent studies identified that endoplasmic reticulum (ER) stress may underlie poor placentation. Therefore, we were interested in investigating the effects of nicotine exposure on the ER stress response in the placenta. A well-established maternal nicotine exposure rat model and Rcho-1 trophoblast giant cell model were utilized to address the research questions. Maternal nicotine exposure in vivo led to elevated ER stress in association with impaired disulfide bond formation and hypoxia. Nicotine exposure in vitro further differentiated that ER stress may be augmented directly through nicotinic acetylcholine receptor activation and indirectly through nicotine-induced hypoxia. Moreover, we relieved nicotine-induced ER stress in Rcho-1 cells in vitro using Tauroursodeoxycholic acid. In conclusion, this thesis provides novel mechanistic insight and contributes to the development of innovative therapeutic approaches to ameliorate nicotine-induced injury in pregnancy.

A nutritional mismatch in postnatal life of low birth weight offspring increases the risk of developing the metabolic syndrome. Moreover, this is associated with decreased hepatic insulin-like growth factor 1 (Igf1) expression, leading to impaired growth and metabolism. Previously we have demonstrated that the timing of nutritional restoration in perinatal life can differentially programhepatic gene expression. While micro RNAs also play an important role in silencing gene expression, to date, the impact of a nutritional mismatch in neonatal life on their long-term expression has not been evaluated. Given the complementarity of miR-29 to the 3i-UTR of Igf1, we examined how proteins restoration in maternal protein restricted rat (MPR) offspring influences hepatic miR-29 and Igf1 expression in adulthood. Pregnant Wistar rats were designated into one of four dietary regimes; 20% protein (Control), 8% protein during lactation only (LP-Lact), 8% protein during gestation only (LP1) or both (LP2). The steady-state expression of hepatic miR-29 mRNA significantly increased in LP2 offspring at postnatal day 21 and 130 and this was inversely related tohepatic Igf1 mRNA and body weight. Interestingly, this reciprocal association was stronger inLP-Lact offspring at postnatal day 21. Functional relevance of this in vivo relationship was evaluated by transfection of miR-29 mimics in neonatal clone 9 rat hepatoma cells. Transfection with miR-29suppressed Igf1 expression by 12 hours. Collectively, these findings implicate that nutritional restoration post-weaning (after liver differentiation) in MPR rat offspring fails to prevent long-term impaired growth, in part, due to miR-29 suppression of hepatic Igf1 expression.

Elongation factor 1A-1 (eEF1A-1) was previously identified as a mediator of fatty acid-induced cell death (lipotoxicity) downstream of endoplasmic reticulum (ER) stress. Furthermore, inhibition of the peptide elongation activity of eEF1A-1 with the cyclic depsipeptide didemnin B (DB) diminishes ER stress and lipotoxicity in cultured hepatocytes. Since ER stress is involved in nonalcoholic fatty liver disease (NAFLD), it was hypothesized that administration of DB to obese mice with NAFLD would reduce hepatic lipotoxicity. Treatment with DB for one week improved several parameters associated with hepatic lipotoxicity and modestly decreased food intake without evidence of illness. Liver triglycerides and protein markers of ER stress, plasma measurements of liver enzymes, plasma cholesterol, and glucose homeostasis were all improved. Of these observations, only improved plasma liver enzymes and glucose homeostasis were completely attributed to reduced food intake. It was concluded that acute intervention with DB improved hepatic lipotoxicity in obese mice with NAFLD.

Kisspeptins signal via the G-protein coupled receptor, KISS1R, and act as metastasis suppressors in numerous cancers. In estrogen receptor (ERα)-negative breast cancer cells, however, KISS1R signaling promotes cell invasion by activating the epidermal growth factor receptor (EGFR). Unfortunately, clinical success of anti-EGFR therapeutics has been limited, as patients often develop drug resistance. Recently, another receptor tyrosine kinase, AXL, has been shown to promote breast cancer drug resistance. We hypothesize that KISS1R promotes EGFR expression and induces breast cancer drug resistance. We demonstrated that KISS1R increases EGFR transcription, by increasing SP-1 binding to the EGFR promoter, as demonstrated by chromatin immunoprecipitation experiments. Additionally, we discovered that KISS1R signaling promotes drug resistance of ERα-negative breast cancer cells, since KISS1R overexpression decreased intracellular doxorubicin accumulation and decreased sensitivity. KISS1R also promotes AXL expression and AXL depletion restored doxorubicin sensitivity. Overall, our results suggest that KISS1R signaling promotes ERα-negative breast cancer drug resistance and may be a novel target to restore drug sensitivity.

Acute respiratory distress syndrome (ARDS) is a lung disease involving profound inflammation. Origins of persistent inflammation in select cases of ARDS are poorly understood, and we propose persistent inflammatory macrophages may be one of its mechanisms. Macrophages polarize to either promote inflammation, or suppress inflammation. Tissue inhibitor of metalloproteinases 3 (TIMP3) reduces the pro-inflammatory polarization in macrophages. Additionally, studies have shown TIMP3 promotes apoptosis, and its absence delays recovery from bleomycin-induced lung injury. We hypothesize that TIMP3 promotes apoptosis of murine macrophages through inhibition of metalloproteinase activity and stabilization of FAS on the cell surface. Pro-inflammatory Timp3-/- bone marrow-derived macrophages (BMDMs) have significantly higher metalloproteinase activity, and significantly lower sFASL-induced apoptosis compared to WT BMDMs measured with FLICA and Annexin V. rTIMP3 treatment rescued both metalloproteinase activity and apoptosis. In conclusion, excessive metalloproteinase activity in Timp3-/- BMDMs is associated with sFASL-induced apoptosis potentially due to metalloproteinase dependent death receptor processing.

Nicotine exposure during pregnancy through cigarette smoking, nicotine replacement therapies or e-cigarette use continues to be awidespread public health problem, impacting both fetal and postnatal health. Yet, at this time, there remains limited data regarding the safety and efficacy in using these nicotine products during pregnancy. Notably, reports assessing the effect of nicotine exposure on postnatal health outcomes in humans, including reproductive health, are severely lacking. Our current understanding regarding the consequences of nicotine exposure during pregnancy is limited to a few animal studies, which do not comprehensively address the underlying cellular mechanisms involved. This paper aims to critically review the current knowledge from human and animal studies regarding the direct and indirect effects (e.g., obesity) of maternal nicotine exposure, regardless of its source, on reproductive outcomes in pregnancy and postnatal life. Furthermore, this review highlights several key cellular mechanisms involved in these adverse reproductive deficits including oxidative stress, inflammation, and endoplasmic reticulum (ER) stress. By understanding the interplay of the cellular mechanisms involved, further strategies could be developed to prevent the reproductive abnormalities resulting from exposure to nicotine in utero and influence informed clinical guidelines for pregnant women.

Intervertebral disc (IVD) degeneration is a complex and multi-factorial process that is influenced by aging, genetic predispositions and environmental influences, such as altered mechanical loading. While recent studies have begun to characterize changes in the IVD associated with degeneration, the underlying etiology remains elusive. It is thought that the loss of notochord cells from the nucleus pulposus (NP) is one initiating factor contributing to the cascade of disc degeneration. However, questions remain about the function of notochord cells within the disc, including their capacity as tissue specific progenitor cells for the nucleus pulposus. We have demonstrated, with the use of the novel notochord-specific Cre mouse (Noto-Cre), that all cells in the adult NP are derived from the embryonic notochord (Chapter 2). Using this mouse strain, we performed transcriptional profiling paired with bioinformatic analysis of NP tissues from various time points and determined that TGF-β, PPAR and PI3K-AKT pathways are associated with NP aging, while mineralization, calcium and WNT signalling pathways are associated with NP degeneration (Chapter 3). We further characterized the proteomic signature of the healthy murine IVD to gain insight into the composition of this complex structure and provide a comprehensive catalogue of proteins enriched within the disc (Chapter 4). Lastly, we studied the effects of mechanical loading, in the form of whole- body vibration on the intervertebral disc. Whole-body vibration platforms are becoming increasingly popular in the fitness industry as well as in clinical practice for the treatment of musculoskeletal disorders. Our results demonstrate a transient beneficial effect of acute exposure to whole body vibration on the IVD. Furthermore, we demonstrate that this response is both frequency-dependent and tissue autonomous (Chapter 5). In follow up studies, we demonstrated that repeated exposure to vibration resulted in dramatic catabolic effects leading to degeneration of the IVD. These detrimental effects were also detected in tissues of the knee joint, mediated in both joints by the activation of matrix metalloproteinases (Chapter 6). In summary, this work uses novel mouse models to provide insight to intervertebral disc development, degeneration and the response of the IVD to mechanical loading.

This document reviews the literature on local brain manipulation of general anesthesia in animals, focusing on behavioral and electrographic effects related to hypnosis or loss of consciousness. Local inactivation or lesion of wake-active areas, such as locus coeruleus, dorsal raphe, pedunculopontine tegmental nucleus, perifornical area, tuberomammillary nucleus, ventral tegmental area and basal forebrain, enhanced general anesthesia. Anesthesia enhancement was shown as a delayed emergence (recovery of righting reflex) from anesthesia or a decrease in the minimal alveolar concentration that induced loss of righting. Local activation of various wake-active areas, including pontis oralis and centromedial thalamus, promoted behavioral or electrographic arousal during maintained anesthesia and facilitated emergence. Lesion of the sleep-active ventrolateral preoptic area resulted in increased wakefulness and decreased isoflurane sensitivity, but only for 6 days after lesion. Inactivation of any structure within limbic circuits involving the medial septum, hippocampus, nucleus accumbens, ventral pallidum, and ventral tegmental area, amygdala, entorhinal and piriform cortex delayed emergence from anesthesia, and often reduced anesthetic-induced behavioral excitation. In summary, the concept that anesthesia works on the sleep-wake system has received strong support from studies that inactivated/lesioned or activated wake-active areas, and weak support from studies that lesioned sleep-active areas. In addition to the conventional wake-sleep areas, limbic structures such as the medial septum, hippocampus and prefrontal cortex are also involved in the behavioral response to general anesthesia. We suggest that hypnosis during general anesthesia may result from disrupting the wake-active neuronal activities in multiple areas and suppressing an atropine-resistant cortical activation associated with movements.

Deep brain stimulation (DBS) has been shown to be effective for relief of Parkinson's disease, depression and obsessive-compulsive disorder in humans, but the effect of DBS on psychosis is largely unknown. In previous studies, we showed that inactivation of the medial septum or nucleus accumbens normalized the hyperactive and psychosis-related behaviors induced by psychoactive drugs. We hypothesized that DBS of the medial septum or nucleus accumbens normalizes the ketamine-induced abnormal behaviors and brain activity in freely moving rats. Male Long-Evans rats were subcutaneously injected with ketamine (3 mg/kg) alone, or given ketamine and DBS, or injected with saline alone. Subcutaneous injection of ketamine resulted in loss of gating of hippocampal auditory evoked potentials (AEPs), deficit in prepulse inhibition (PPI) and hyperlocomotion, accompanied by increased hippocampal gamma oscillations of 70-100 Hz. Continuous 130-Hz stimulation of the nucleus accumbens, or 100-Hz burst stimulation of the medial septum (1s on and 5s off) significantly attenuated ketamine-induced PPI deficit and hyperlocomotion. Medial septal stimulation also prevented the loss of gating of hippocampal AEPs and the increase in hippocampal gamma waves induced by ketamine. Neither septal or accumbens DBS alone without ketamine injection affected spontaneous locomotion or PPI. The results suggest that DBS of the medial septum or nucleus accumbens may be an effective method to alleviate psychiatric symptoms of schizophrenia. The effect of medial septal DBS in suppressing both hippocampal gamma oscillations and abnormal behaviors induced by ketamine suggests that hippocampal gamma oscillations are a correlate of disrupted behaviors.

Background: Cholinergic drugs are known to modulate the response of general anesthesia. However, the sensitivity of isoflurane or other volatile anesthetics after selective lesion of septal cholinergic neurons that project to the hippocampus is not known. Methods: Male Long Evans rats had 192 immunoglobulin G-saporin infused into the medial septum (n = 10), in order to selectively lesion cholinergic neurons, whereas control, sham-lesioned rats were infused with saline (n = 12). Two weeks after septal infusion, the hypnotic properties of isoflurane and ketamine were measured using a behavioral endpoint of loss of righting reflex (LORR). Septal lesion was assessed by counting choline acetyltransferase–immunoreactive cells and parvalbumin-immunoreactive cells. Results: Rats with 192 immunoglobulin G-saporin lesion, as compared with control rats with sham lesion, showed a 85% decrease in choline acetyltransferase–immunoreactive, but not parvalbumin–immunoreactive, neurons in the medial septal area. Lesioned as compared with control rats showed increased isoflurane sensitivity, characterized by a leftward shift of the graph plotting cumulative LORR percent with isoflurane dose. However, lesioned and control rats were not different in their LORR sensitivity to ketamine. When administered with 1.375% isoflurane, LORR induction time was shorter, whereas emergence time was longer, in lesioned as compared with control rats. Hippocampal 62–100 Hz gamma power in the electroencephalogram decreased with isoflurane dose, with a decrease that was greater in lesioned (n = 5) than control rats (n = 5). Conclusions: These findings suggest a role of the septal cholinergic neurons in modulating the sensitivity to isoflurane anesthesia, which affects both induction and emergence. The sensitivity of hippocampal gamma power to isoflurane appears to indicate anesthesia (LORR) sensitivity.

Waveguide Evanescent Field Scattering (WEFS) microscopy is introduced as a new and simple tool for label-free, high contrast imaging of bacteria and bacteria sensors. Bacterial microcolonies and single bacteria were discriminated both by their bright field images and by their evanescent scattering intensity. By comparing bright field images with WEFS images, the proportion of planktonic: sessile (i.e., ‘floating’: attached) bacteria were measured. Bacteria were irradiated with UV light, which limited their biofilm forming capability. A quantitative decrease in attachment of individual, sessile bacteria and in attached, microcolony occupied areas was easily determined within the apparent biofilms with increasing UV dose. WEFS microscopy is an ideal tool for providing rapid quantitative data on biofilm formation.

Many neurodegenerative diseases, including Alzheimer’s Disease, are associated with ER stress. It is suggested that plasma membrane channels contribute to the increased ionic influx and subsequent cell death in response to ER stress. Pannexin channels, which have been implicated in various pathophysiological conditions, are a suitable candidate for facilitating this response. However, mechanisms of pannexin channel activation are poorly defined. I investigated the potential regulation of pannexin activity by the ER stress sensor, STIM. It was hypothesized that pannexin channel activation during ER stress is contingent on the recruitment of STIM proteins. In neurons, pannexin activation was observed in response to pharmacological ER stressors as well as Aβ42. Moreover, I identified STIM as a key signaling component promoting pannexin activation under ER stress in HEK cells, though not in neurons. In conclusion, pannexin activation is coupled to ER stress in neurons; however, the underlying molecular mechanism remains unknown.

Electrolytic lesion of the medial septum, a basal forebrain nucleus that projects to the hippocampus, prolonged the emergence from general anesthesia in rats. Septal lesioned rats required a longer time to recover from a loss of righting reflex (LORR) and a loss of tail-pinch response after injectable (20 mg/kg i.p. pentobarbital, 5 mg/kg i.v. propofol) or volatile (1.5% halothane, 2% isoflurane) anesthetic. When incremental doses of propofol were given i.p., septal lesioned rats as compared to control rats showed LORR at a lower dose of propofol. Similarly, when the rats were exposed to increasing concentrations of isoflurane, the percent of rats showing LORR was leftward shifted for lesioned rats as compared to control rats. Septal lesioned rats as compared to control rats showed decreased locomotor activity when exposed to 1.5% halothane. Lesion of the medial septum was confirmed by thionin-stained histological sections as well as loss of acetylcholinesterase (AchE) staining in the hippocampus, indicating a depletion of septohippocampal cholinergic afferents. Medial septal lesion resulted in a near complete loss of hippocampal theta rhythm during walking and a general decrease in power of the hippocampal EEG at all frequencies (0–100 Hz), during walking or immobility. It is concluded that lesion of medial septum, in part through a loss of septohippocampal cholinergic afferents, increased the anesthesia response to volatile and injectable general anesthetics, during both induction and emergence. It is suggested that the septohippocampal system participates in many components of general anesthesia including hypnosis, immobility, and analgesia.

While nicotine replacement therapy (NRT) is presumed to be a safer alternative to smoking in pregnancy, the long-term consequences in offspring are still largely unknown. Animal studies now suggest that maternal nicotine exposure during pregnancy and lactation (MNE-PL) leads to a wide variety of adverse outcomes for the offspring, including increased adiposity. The focus of this study was to investigate how MNE-PL in rats may lead to liver dysfunction long-term in offspring through alterations in gene expression and epigenetic modifications. Postnatal day 180 (PND180) offspring exposed to nicotine during pregnancy and lactation (1mg/kg/day) exhibited increased circulating and hepatic triglycerides concomitant with increased expression of fatty acid synthase (FAS), an enzyme involved in hepatic de novo fatty acid synthesis. Furthermore, we demonstrate that MNE-PL offspring displayed increased protein expression of the Liver X Receptor a (LXRa), a key regulator of FAS. Chromatin immunoprecipitation revealed enriched binding of LXRa to the putative LXRE element on the FAS promoter in PND180 male offspring. This was associated with enhanced acetylation of histone H3 [K9,14] surrounding the FAS promoter, a hallmark of chromatin activation. Collectively, these findings suggest that nicotine exposure during pregnancy and lactation leads to increased circulating and hepatic triglyceride levels long-term via changes in transcriptional and epigenetic regulation of the hepatic lipogenic pathway.

Stomatin-like Protein 2 (SLP-2) has been identified as a stress-inducible transcript and has been shown to interact with and stabilize mitochondrial proteins. Since mitochondria are critical for neuronal function, we hypothesized that SLP-2 regulates neuron survival in response to stressful stimuli. A conditional SLP-2 knockout mouse (deletion) and the SN56 cell line (upregulation) were employed to study the role of SLP-2 in mitochondrial dynamics and neuron survival. SLP-2 deficient primary cortical neurons displayed significantly decreased levels of various mitochondrial respiratory chain proteins, indicating SLP-2 contributes to maintenance of mitochondrial membrane integrity. SLP-2 was up-regulated in response to oxidative stress and DNA damage, but was not required for stress-induced mitochondrial biogenesis. The level of SLP-2 expression had a significantly different effect on neuron survival, depending on the target of the insult. The results of this study improve our understanding of the role of SLP-2 in mitochondrial stress responses, bioenergetics, and survival in neurons.

We present a biosensor based on periodic arrays of gold nanodisks patterned on top of a glass substrate. The sensor's resonance wavelength, peak linewidth and figure of merit were studied both in experiments and in simulations. We analyzed the dependence of the sensor's resolution on the inherent figure of merit of the sensor structure and the signal to noise ratio of the detection system. The best achieved refractive index resolution is 1.5 × 10 −4 RIU and the detection limit on antigen–antibody binding is better than 1 ng/mL.

We study and analyze various spectral features of gold nano-disk arrays coupled with the evanescent field of an optical waveguide using finite difference time domain simulations for comparison to our previous work on this topic (Jiang et al, Journal of Lightwave Technology 27, no.13, (2009): 2264-2270). We find that the quadrupolar and dipolar peaks are both heavily dependent on the coherent interactions of the periodic array and show comparable performance for refractive index sensing applications. We also find sharp extinction peaks and dips in the simulated spectra, but they are not sensitive to the index change of the surface-bound layer. A detailed analysis on these sharp features reveals that the sharp extinction peaks are grating-induced quadrupolar modes and the dips represent the suppressed plasmon resonances caused by the photonic band gap.

We describe the growth of surface immobilized gold nanoparticles with organometallic chemical vapour deposition (OMCVD) on amine terminated surfaces, utilizing (trimethylphospine)methylgold ((CH3)3PAuCH3) precursor. Samples fabricated using different deposition times were characterized by UV-Vis spectroscopy and scanning electron microscopy. Particle stability on the samples was tested by washing and rinsing treatments with various organic solvents. A biotin-streptavidin scheme was applied to demonstrate the biosensing capabilities of the samples. The size, interparticle distance, and shape of the gold nanoparticles demonstrated that OMCVD is a simple, economic, and fast way to fabricate surface bonded and stable gold nanoparticles. The plasmonic properties, the stability of the particles and the biotin-streptavidin test showed that these OMCVD-grown gold nanoparticles are suitable for reproducible, low noise and highly sensitive biosensing applications.

Seizures are relatively common in children and are a risk factor for subsequent temporal lobe epilepsy. To investigate whether early-life seizures themselves are detrimental to the proper function of the adult brain, we studied whether dendritic excitation and inhibition in the hippocampus of adult rats were altered after hyperthermia-induced seizures in immature rats. In particular, we hypothesized that apical dendritic inhibition in hippocampal CA1 pyramidal cells would be disrupted following hyperthermia-induced seizures in early life. Seizure rats were given three hyperthermia-induced seizures per day for three days from postnatal day (PND) 13 to 15; control rats were handled similarly but not heated. At PND 65-75, paired-pulse inhibition in area CA1 was evaluated under urethane anesthesia, using CA3 and medial perforant path (MPP) stimulation to excite the proximal and distal apical-dendrites, respectively, and the evoked field potentials were analyzed by current source density. There was no difference in the CA1 response to single-pulse stimulation of CA3 or MPP. In control rats, a high-intensity CA3 stimulus inhibited a subsequent MPP-evoked CA1 distal dendritic excitatory sink, and the inhibition at 150-200 ms was blocked by a GABA(B) receptor antagonist. Seizure as compared to control rats showed a decrease in a CA3-evoked inhibition of the CA1 distal dendritic excitation, 30-400 ms after the CA3 stimulus. In addition, seizure as compared to control rats showed a reduced early (20-80 ms) inhibition of a CA1 mid-apical dendritic sink following paired-pulse CA3 stimulation. In conclusion, long-term alterations in dendritic inhibition in CA1 were found following early-life seizures.