The Role of Cdc42 in the Insulin and Leptin Pathways Contributing to the Development of Age-Related Obesity.

Age-related obesity significantly increases the risk of chronic diseases such as type 2 diabetes, cardiovascular diseases, hypertension, and certain cancers. The insulin-leptin axis is crucial in understanding metabolic disturbances associated with age-related obesity. Rho GTPase Cdc42 is a member of the Rho family of GTPases that participates in many cellular processes including, but not limited to, regulation of actin cytoskeleton, vesicle trafficking, cell polarity, morphology, proliferation, motility, and migration. Cdc42 functions as an integral part of regulating insulin secretion and aging. Some novel roles for Cdc42 have also been recently identified in maintaining glucose metabolism, where Cdc42 is involved in controlling blood glucose levels in metabolically active tissues, including skeletal muscle, adipose tissue, pancreas, etc., which puts this protein in line with other critical regulators of glucose metabolism. Importantly, Cdc42 plays a vital role in cellular processes associated with the insulin and leptin signaling pathways, which are integral elements involved in obesity development if misregulated. Additionally, a change in Cdc42 activity may affect senescence, thus contributing to disorders associated with aging. This review explores the complex relationships among age-associated obesity, the insulin-leptin axis, and the Cdc42 signaling pathway. This article sheds light on the vast molecular web that supports metabolic dysregulation in aging people. In addition, it also discusses the potential therapeutic implications of the Cdc42 pathway to mitigate obesity since some new data suggest that inhibition of Cdc42 using antidiabetic drugs or antioxidants may promote weight loss in overweight or obese patients.

Circulating adiponectin levels, expression of adiponectin receptors, and methylation of adiponectin gene promoter in relation to Alzheimer's disease.

BACKGROUND: The role of adiponectin (ADIPOQ) in Alzheimer's disease (AD) has been documented, however, demonstrating controversial results. In this study, we investigated blood serum ADIPOQ levels, methylation of the adiponectin gene promoter, and adiponectin receptors (AdipoR1 and AdipoR2) expression in blood samples isolated from AD patients and healthy controls. METHODS: We performed a case-control study including 248 subjects (98 AD patients and 150 healthy controls); ADIPOQ serum levels, AdipoR1, and AdipoR2 levels in PBMC were measured by ELISA Kits, and ADIPOQ gene methylation was analyzed using methyl-specific PCR. RESULTS: Serum adiponectin levels were threefold higher in the AD group compared to the controls. We have also found a positive correlation between adiponectin and MMSE scores and high-density lipoprotein cholesterol (HDL-C) in AD patients. A significant difference in the proportion of methylation of the CpG sites at - 74 nt of the ADIPOQ gene promoter was detected in AD cases, and the levels of adiponectin in blood serum were significantly higher in methylated samples in the AD group compared to controls. The amount of AdipoR1 was significantly higher among AD subjects, while the expression of AdipoR2 did not vary between AD patients and controls. CONCLUSION: These findings may contribute to a deeper understanding of the etiological factors leading to the development of dementia and may serve as a basis for the development of predictive biomarkers of AD.

Plant Extract of Limonium gmelinii Attenuates Oxidative Responses in Neurons, Astrocytes, and Cerebral Endothelial Cells In Vitro and Improves Motor Functions of Rats after Middle Cerebral Artery Occlusion.

There are numerous publications demonstrating that plant polyphenols can reduce oxidative stress and inflammatory processes in the brain. In the present study we have investigated the neuroprotective effect of plant extract isolated from the roots of L. gmelinii since it contains a rich source of polyphenols and other biologically active compounds. We have applied an oxidative and inflammatory model induced by NMDA, H2O2, and TNF-α in human primary neurons and astrocytes, and mouse cerebral endothelial cell (CECs) line in vitro. The levels of ROS generation, NADPH oxidase activation, P-selectin expression, and activity of ERK1/2 were evaluated by quantitative immunofluorescence analysis, confocal microscopy, and MAPK assay. In vivo, sensorimotor functions in rats with middle cerebral artery occlusion (MCAO) were assessed. In neurons NMDA induced overproduction of ROS, in astrocytes TNF-α initiated ROS generation, NADPH oxidase activation, and phosphorylation of ERK1/2. In CECs, the exposure by TNF-α induced oxidative stress and triggered the accumulation of P-selectin on the surface of the cells. In turn, pre-treatment of the cells with the extract of L. gmelinii suppressed oxidative stress in all cell types and pro-inflammatory responses in astrocytes and CECs. In vivo, the treatment with L. gmelinii extract improved motor activity in rats with MCAO.

Mesenchymal Stem Cells Coated with Synthetic Bone-Targeting Polymers Enhance Osteoporotic Bone Fracture Regeneration.

Osteoporosis is a progressive skeletal disease characterized by reduced bone density leading to bone fragility and an elevated risk of bone fractures. In osteoporotic conditions, decrease in bone density happens due to the augmented osteoclastic activity and the reduced number of osteoblast progenitor cells (mesenchymal stem cells, MSCs). We investigated a new method of cell therapy with membrane-engineered MSCs to restore the osteoblast progenitor pool and to inhibit osteoclastic activity in the fractured osteoporotic bones. The primary active sites of the polymer are the N-hydroxysuccinimide and bisphosphonate groups that allow the polymer to covalently bind to the MSCs' plasma membrane, target hydroxyapatite molecules on the bone surface and inhibit osteolysis. The therapeutic utility of the membrane-engineered MSCs was investigated in female rats with induced estrogen-dependent osteoporosis and ulnar fractures. The analysis of the bone density dynamics showed a 27.4% and 21.5% increase in bone density at 4 and 24 weeks after the osteotomy of the ulna in animals that received four transplantations of polymer-modified MSCs. The results of the intravital observations were confirmed by the post-mortem analysis of histological slices of the fracture zones. Therefore, this combined approach that involves polymer and cell transplantation shows promise and warrants further bio-safety and clinical exploration.

Improvement of Neurological Function in Rats with Ischemic Stroke by Adipose-derived Pericytes.

Pericytes possess high multipotent features and cell plasticity, and produce angiogenic and neurotrophic factors that indicate their high regenerative potential. The aim of this study was to investigate whether transplantation of adipose-derived pericytes can improve functional recovery and neurovascular plasticity after ischemic stroke in rats. Rat adipose-derived pericytes were isolated from subcutaneous adipose tissue by fluorescence-activated cell sorting. Adult male Wistar rats were subjected to 90 min of middle cerebral artery occlusion followed by intravenous injection of rat adipose-derived pericytes 24 h later. Functional recovery evaluations were performed at 1, 7, 14, and 28 days after injection of rat adipose-derived pericytes. Angiogenesis and neurogenesis were examined in rat brains using immunohistochemistry. It was observed that intravenous injection of adipose-derived pericytes significantly improved recovery of neurological function in rats with stroke compared to phosphate-buffered saline-treated controls. Immunohistochemical analysis revealed that the number of blood capillaries was significantly increased along the ischemic boundary zone of the cortex and striatum in stroke rats treated with adipose-derived pericytes. In addition, treatment with adipose-derived pericytes increased the number of doublecortin positive neuroblasts. Our data suggest that transplantation of adipose-derived pericytes can significantly improve the neurologic status and contribute to neurovascular remodeling in rats after ischemic stroke. These data provide a new insight for future cell therapies that aim to treat ischemic stroke patients.

The Links Between the Gut Microbiome, Aging, Modern Lifestyle and Alzheimer's Disease.

Gut microbiome is a community of microorganisms in the gastrointestinal tract. These bacteria have a tremendous impact on the human physiology in healthy individuals and during an illness. Intestinal microbiome can influence one's health either directly by secreting biologically active substances such as vitamins, essential amino acids, lipids et cetera or indirectly by modulating metabolic processes and the immune system. In recent years considerable information has been accumulated on the relationship between gut microbiome and brain functions. Moreover, significant quantitative and qualitative changes of gut microbiome have been reported in patients with Alzheimer's disease. On the other hand, gut microbiome is highly sensitive to negative external lifestyle aspects, such as diet, sleep deprivation, circadian rhythm disturbance, chronic noise, and sedentary behavior, which are also considered as important risk factors for the development of sporadic Alzheimer's disease. In this regard, this review is focused on analyzing the links between gut microbiome, modern lifestyle, aging, and Alzheimer's disease.

Date Pit Carbon Dots Induce Acidic Inhibition of Peroxidase and Disrupt DNA Repair in Antibacteria Resistance.

Carbon nanodots (C-dots) are emerging as a new type of promising agent in anticancer, imaging, and new energy. Reports as well as the previous research indicate that certain C-dots can enhance targeted cancer therapy. However, in-depth mechanisms for such anticancer effect remain unclear. In this work, treatment provided by the date pit-derived C-dots, exhibits significant DNA damage; Annexin V/7-AAD-mediated apoptosis, and G2/M cell cycle arrest in prostate cancer cells. The application of C-dots to the cell generally leads to acidulation of the cell medium, cooperated with membrane compact. The date pit-derived C-dots are observed inhibiting the horseradish peroxidase. Moreover, the C-dots disrupt likely through nucleotide excision DNA repair at low dose during DNA ligation step suggesting the antimicrobial effect and targeting Pim-1, EGFR, mTOR, and DNA damage pathways in cancer cells. For the first time the detailed and novel mechanisms underlying the C-dots, derived from the date-pit, as an efficient, low-cost, and green nanomaterial are reveled for cancer therapy and anti-infection.

The Effects of Mobile Phone Radiofrequency Electromagnetic Fields on ß-Amyloid-Induced Oxidative Stress in Human and Rat Primary Astrocytes.

Amyloid beta peptide (Aß) is implicated in the development of pathological reactions associated with Alzheimer's disease (AD), such as oxidative stress, neuro-inflammation and death of brain cells. Current pharmacological approaches to treat AD are not able to control the deposition of Aß and suppression of Aß-induced cellular response. There is a growing body of evidence that exposure to radiofrequency electromagnetic field (RF-EMF) causes a decrease of beta-amyloid deposition in the brains and provides cognitive benefits to Alzheimer's Tg mice. Herein, we investigated the effects of mobile phone radiofrequency EMF of 918 MHz on reactive oxygen species (ROS) formation, mitochondrial membrane potential (MMP), activity of NADPH-oxidase, and phosphorylation of p38MAPK and ERK1/2 kinases in human and rat primary astrocytes in the presence of Aß42 and H2O2. Our data demonstrate that EMF is able to reduce Aß42- and H2O2-induced cellular ROS, abrogate Aß42-induced production of mitochondrial ROS and the co-localization between the cytosolic (p47-phox) and membrane (gp91-phox) subunits of NADPH oxidase, while increasing MMP, and inhibiting H2O2-induced phosphorylation of p38MAPK and ERK1/2 in primary astrocytes. Yet, EMF was not able to modulate alterations in the phosphorylation state of the MAPKs triggered by Aß42. Our findings provide an insight into the mechanisms of cellular and molecular responses of astrocytes on RF-EMF exposure and indicate the therapeutic potential of RF-EMF for the treatment of Alzheimer's disease.

Azelnidipine Attenuates the Oxidative and NFκB Pathways in Amyloid-ß-Stimulated Cerebral Endothelial Cells.

Cerebral amyloid angiopathy (CAA), a condition depicting cerebrovascular accumulation of amyloid ß-peptide (Aß), is a common pathological manifestation in Alzheimer's disease (AD). In this study, we investigated the effects of Azelnidipine (ALP), a dihydropyridine calcium channel blocker known for its treatment of hypertension, on oligomeric Aß (oAß)-induced calcium influx and its downstream pathway in immortalized mouse cerebral endothelial cells (bEND3). We found that ALP attenuated oAß-induced calcium influx, superoxide anion production, and phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and calcium-dependent cytosolic phospholipase A2 (cPLA2). Both ALP and cPLA2 inhibitor, methylarachidonyl fluorophosphate (MAFP), suppressed oAß-induced translocation of NFκB p65 subunit to nuclei, suggesting that cPLA2 activation and calcium influx are essential for oAß-induced NFκB activation. In sum, our results suggest that calcium channel blocker could be a potential therapeutic strategy for suppressing oxidative stress and inflammatory responses in Aß-stimulated microvasculature in AD.

Elevated levels of the small GTPase Cdc42 induces senescence in male rat mesenchymal stem cells.

Mesenchymal stem cells (MSCs) represent a promising cell source for cellular therapy and tissue engineering and are currently being tested in a number of clinical trials for various diseases. However, like other somatic cells, MSCs age, and this senescence is accompanied by a progressive decline in stem cell function. Several lines of evidence suggest a role for the Rho family GTPase Cdc42 activity in cellular senescence processes. In the present study, we have examined aging-associated Cdc42 activity in rat adipose-derived mesenchymal stem cells (ADMSCs) and the consequences of pharmacological inhibition of Cdc42 in ADMSCs from aged rats. We demonstrate that ADMSCs show a decreased rate of cell growth and a decreased ability to differentiate into chrodrogenic, osteogenic and adipogenic cell lineages as a function of rat age. This is accompanied with an increased staining for SA-ß-Gal activity and increased levels of Cdc42 bound to GTP. Treatment of ADMSCs from 24-month old rats with three Cdc42 inhibitors significantly increased proliferation rates, decreased SA-ß-Gal staining, and reduced Cdc42-GTP. The Cdc42 inhibitor CASIN increased adipogenic and osteogenic differentiation potential in ADMSCs from 24-month old rats, and decreased the levels of radical oxygen species (ROS), p16INK4a levels, F-actin, and the activity of the ERK1/2 and JNK signaling pathways that were all elevated in these cells. These data suggest that ADMSCs show increased rates of senescence as rats age that appear to be due to elevated Cdc42 activity. Thus, Cdc42 plays important roles in MSC senescence and differentiation potential, and pharmacological reduction of Cdc42 activity can, at least partially, rejuvenate aged MSCs.

Role of ROS in Aß42 Mediated Activation of Cerebral Endothelial Cells.

INTRODUCTION: There is substantial evidence that the deposition of aggregated amyloid-beta peptide (Aß) in brain parenchyma and brain vessels is the main cause of neuronal dysfunction and death in Alzheimer's disease (AD). Aß exhibits multiple cytotoxic effects on neurons and glial cells and causes dysfunction of the blood brain barrier (BBB). In AD brains, an increased deposition of Aß in the cerebral vasculature has been found to be correlated with increased transmigration of blood-borne inflammatory cells and neurovascular inflammation. However, regulatory mediators of these processes remain to be elucidated. In this study, we examined the role of ROS in actin polymerization and expression of adhesion molecules (P-selectin) on the surface of the cerebral endothelial cells (CECs) that are activated by Aß42. MATERIALS AND METHODS: Mouse BEnd3 line (ATCC) was used in this research. BEnd3 cells respond to Aß treatment similarly to human primary CECs and are a common model to investigate CECs' function. We used immortalized bEnd3 cells as the following: controls; cells incubated with Aß42 for 10, 30, and 60 minutes; cells incubated with 30 mM of antioxidant N-acetylcysteine (NAC) for 1 hr; and, cells pre-treated with NAC followed by Aß42 exposure. We measured DHE fluorescence to investigate intracellular ROS production. Immunofluorescent microscopy of anti-P-selectin and oregon green phalloidin was used to quantify the surface P-selectin expression and actin polymerization, and Western blot analysis was used to analyze total P-selectin expression. RESULTS: The results of this study have demonstrated a significant time-dependent ROS accumulation after 10 minutes, 30 minutes, and 60 minutes of Aß42 treatment, while Aß42 stimulated ROS production in CECs was attenuated by pre-treatment with the NAC antioxidant. We also found that Aß42 increased P-selectin fluorescence at the surface of bEnd3 cells in a time dependent manner in parallel to ROS elevation. However, total expression levels of P-selectin were not changed following exposure to Aß42. Pretreatment with NAC attenuated Aß42 induced P-selectin localization, while NAC alone did not significantly affect P selectin localization. As a positive control, H2O2 also increased P-selectin expression on the cell surface, which peaked after 30 minutes of H2O2 treatment. Exposure of CECs with Aß42 promoted actin polymerization, which peaked after 10 minutes of Aß42 treatment, while no significant increase of F-actin intensity was observed when cells were pre-treated with NAC. H2O2 was able to mimic Aß42 induced oxidative stress, causing increased actin polymerization with similar timing. CONCLUSIONS: The results of our study have indicated that Aß42 induced accumulation of P-selectin on the surface of bEnd3 cells and promoted actin polymerization, and all these events were correlated with ROS generation. The rapid post-translational cell signaling response mediated by ROS may well represent an important physiological trigger of the microvascular inflammatory responses in AD and requires further investigations.

Ectopic Liver Tissue Formation in Rats with Induced Liver Fibrosis.

INTRODUCTION: The possible alternative approach to whole-organ transplantation is a cell-based therapy, which can also be used as a "bridge" to liver transplantation. However, morphological and functional changes in the liver of patients suffering from chronic liver fibrosis and cirrhosis restrict the effectiveness of direct cell transplantation. Therefore, extra hepatic sites for cell transplantation, including the spleen, pancreas, peritoneal cavity, and subrenal capsule, could be a useful therapeutic approach for compensation of liver functions. However, a method of transplantation of hepatocytes into ectopic sites is needed to improve hepatocyte engraftment. Previously published data has demonstrated that mouse lymph nodes can support the engraftment and proliferation of hepatocytes as ES and rescue Fah mice from lethal liver failure. Thus, the aim of the study was to evaluate the engraftment of i.p. injected allogeneic hepatocytes into extra hepatic sites in albino rats with chemically induced liver fibrosis (LF). MATERIALS AND METHODS: Albino rats were randomly divided into 4 groups: (1) intact group (n = 18); (2) rats with induced LF (n = 18); (3) rats with induced LF and transplanted with hepatocytes (n = 18); (4) as a control, rats were treated with cyclosporine A only (n = 18). In order to prevent an immune response, groups 2 and 3 were subjected to immunosuppression by cyclosporine A (25 mg/kg per day). LF was induced using N-nitrosodimethylamine (NDMA), i.p., 10 mg/kg, three times a week for 4 weeks and confirmed by histological analysis of the liver samples. Hepatocytes transplantation (HT) was performed two days after NDMA exposure cessation by i.p. injection of 5×106 freshly isolated allogeneic hepatocytes. Liver function was assessed by quantifying blood biochemical parameters (ALT, AST, GGT, total protein, bilirubin, and albumin) at 1 week, 1 month, and 2 months after hepatocytes transplantation (HT). To confirm a hepatocytes' engraftment, we conducted immunohistochemical staining against HepPar1. RESULTS: We observed a 30% mortality rate among rats with LF within 1 week after NDMA exposure cessation, while 100% of animals with HT survived. ALT, AST, and GGT activities and bilirubin levels were markedly elevated in blood samples of LF rats compared to the control animals. However, HT significantly improved ALT, AST, and GGT activity as well as bilirubin levels. We also observed decreased levels of total protein and albumin in the blood serum of rats with LF, while HT normalized these parameters. At the same time, we have not detected any statistical differences of the studied parameters in the group 4, which was treated with Cyclosporine A only, compared with the intact animals. HepPar1 immunohistochemical staining of the different tissue sections demonstrated the presence of engrafted hepatocytes, mainly within enlarged Peyer's patches (aggregated lymphoid nodules in the lowest portion of the small intestine). CONCLUSION: The results of our study provide evidence that HT improves animal survival and liver functions. One potential reason for these results is that ectopic hepatic mass inside the Peyer's patches can rescue rats from liver failure.

Effects of Amyloid Beta Peptide on Neurovascular Cells.

Alzheimer's disease (AD) is a chronic neurodegenerative disorder, which is characterized by the accumulation of amyloid plaques and neurofibrillary tangles in specific regions of the brain, accompanied by impairment of the neurons, and progressive deterioration of cognition and memory of affected individuals. Although the cause and progression of AD are still not well understood, the amyloid hypothesis is dominant and widely accepted. According to this hypothesis, an increased deposition of amyloid-ß peptide (Aß) in the brain is the main cause of the AD's onset and progression. There is increasing body of evidence that blood-brain barrier (BBB) dysfunction plays an important role in the development of AD, and may even precede neuron degeneration in AD brain. In the early stage of AD, microvasculature deficiencies, inflammatory reactions, surrounding the cerebral vasculature and endothelial dysfunctions are commonly observed. Continuous neurovascular degeneration and accumulation of Aß on blood vessels resulting in cerebral amyloid angiopathy is associated with further progression of the disease and cognitive decline. However, little is known about molecular mechanisms that underlie Aß induced damage of neurovascular cells. In this regards, this review is aimed to address how Aß impacts the cerebral endothelium. Understanding the cellular pathways triggered by Aß leading to alterations in cerebral endothelial cells structure and functions would provide insights into the mechanism of BBB dysfunction and inflammatory processes in Alzheimer's, and may offer new approaches for prevention and treatment strategies for AD.