Complexity of NAC Action as an Antidiabetic Agent: Opposing Effects of Oxidative and Reductive Stress on Insulin Secretion and Insulin Signaling.

Dysregulated redox balance is involved in the pathogenesis of type 2 diabetes. While the benefit of antioxidants in neutralizing oxidative stress is well characterized, the potential harm of antioxidant-induced reductive stress is unclear. The aim of this study was to investigate the dose-dependent effects of the antioxidant N-acetylcysteine (NAC) on various tissues involved in the regulation of blood glucose and the mechanisms underlying its functions. H2O2 was used as an oxidizing agent in order to compare the outcomes of oxidative and reductive stress on cellular function. Cellular death in pancreatic islets and diminished insulin secretion were facilitated by H2O2-induced oxidative stress but not by NAC. On the other hand, myotubes and adipocytes were negatively affected by NAC-induced reductive stress, as demonstrated by the impaired transmission of insulin signaling and glucose transport, as opposed to H2O2-stimulatory action. This was accompanied by redox balance alteration and thiol modifications of proteins. The NAC-induced deterioration of insulin signaling was also observed in healthy mice, while both insulin secretion and insulin signaling were improved in diabetic mice. This study establishes the tissue-specific effects of NAC and the importance of the delicate maintenance of redox balance, emphasizing the challenge of implementing antioxidant therapy in the clinic.

Activation of Insulin Signaling by Botanical Products.

Type 2 diabetes (T2D) is a worldwide health problem, ranked as one of the leading causes for severe morbidity and premature mortality in modern society. Management of blood glucose is of major importance in order to limit the severe outcomes of the disease. However, despite the impressive success in the development of new antidiabetic drugs, almost no progress has been achieved with regard to the development of novel insulin-sensitizing agents. As insulin resistance is the most eminent factor in the patho-etiology of T2D, it is not surprising that an alarming number of patients still fail to meet glycemic goals. Owing to its wealth of chemical structures, the plant kingdom is considered as an inventory of compounds exerting various bioactivities, which might be used as a basis for the development of novel medications for various pathologies. Antidiabetic activity is found in over 400 plant species, and is attributable to varying mechanisms of action. Nevertheless, relatively limited evidence exists regarding phytochemicals directly activating insulin signaling, which is the focus of this review. Here, we will list plants and phytochemicals that have been found to improve insulin sensitivity by activation of the insulin signaling cascade, and will describe the active constituents and their mechanism of action.

Maternal N-Acetyl Cysteine Intake Improved Glucose Tolerance in Obese Mice Offspring.

Exposure to certain environmental factors during the early stages of development was found to affect health in adulthood. Among other environmental factors, oxidative stress has been suggested to be involved in fetal programming, leading to elevated risk for metabolic disorders, including type 2 diabetes; however, the possibility that antioxidant consumption during early life may affect the development of diabetes has scarcely been studied. The aim of this study was to investigate the effects of N-acetyl-l-cysteine (NAC) given during pregnancy and lactation on the susceptibility of offspring to develop glucose intolerance at adulthood. C57bl6/J mice were given NAC during pregnancy and lactation. High fat diet (HFD) was given to offspring at an age of 6 weeks for an additional 9 weeks, till the end of the study. Isolated islets of NAC-treated offspring (6 weeks old, before HFD feeding) had an increased efficacy of glucose-stimulated insulin secretion and a higher resistance to oxidative damage. Following HFD feeding, glucose tolerance and insulin sensitivity of NAC-treated offspring were improved. In addition, islet diameter was lower in male offspring of NAC-treated mice compared to their HFD-fed littermates. NAC consumption during early life improves glucose tolerance in adulthood in mice.

Anti-diabetic activity of aerial parts of Sarcopoterium spinosum.

BACKGROUND: Sarcopoterium spinosum (S. spinosum) is used by Bedouin medicinal practitioners for the treatment of diabetes. While the anti-diabetic activity of S. spinosum root extract was validated in previous studies, the activity of aerial parts of the same plants has not been elucidated yet. The aim of this study was to clarify the glucose lowering properties of the aerial parts of the shrub. METHODS: Anti-diabetic properties were evaluated by measuring the activity of carbohydrate digesting enzymes, glucose uptake into 3 T3-L1 adipocytes, and insulin secretion. Insulin signaling cascade was followed in L6 myotubes using Western blot and PathScan analysis. RESULTS: Activity of α-amylase and α-glucosidase was inhibited by extracts of all S. spinosum organs. Basal and glucose-induced insulin secretion was measured in Min6 cells and found to be enhanced as well. Glucose uptake was induced by all S. spinosum extracts, with roots found to be the most effective and fruits the least. The effect of S. spinosum on Akt phosphorylation was minor compared to insulin effect. However, GSK3ß and PRAS40, which are downstream elements of the insulin cascade, were found to be highly phosphorylated by S. spinosum extracts. Inhibition of PI3K and Akt, but not AMPK and ERK, abrogated the induction of glucose uptake by the aerial parts of the shrub. CONCLUSION: The aerial organs of S. spinosum have anti-diabetic properties and may be used as a basis for the development of dietary supplements or to identify new agents for the treatment of type 2 diabetes.

Autocrine/paracrine function of globular adiponectin: inhibition of lipid metabolism and inflammatory response in 3T3-L1 adipocytes.

Adiponectin is an adipose-derived hormone, with beneficial effects on insulin sensitivity and inflammation. The aim of this study was to clarify the autocrine/paracrine effects of globular adiponectin (gAd) administrated during differentiation on the function of the mature adipocytes. Experiments were performed on 3T3-L1 preadipocytes treated with gAd (10 nM), starting at an early stage of differentiation. gAd treatment during differentiation was without effect on mRNA expression of adiponectin and AdipoR2, but increased AdipoR1 expression. PPARgamma, perillipin and FABP4 mRNA expressions were lower in gAd-treated adipocytes, accompanied by a reduction in lipid accumulation. While mRNA expression of HSL was not affected by gAd compared to untreated adipocytes, both ATGL and FAS were reduced, indicating that gAd regulates both lipolysis and lipogenesis. PPARα, ACOX2 and UCPs mRNA expressions were not affected by gAd, indicating that the reduction in lipid content is not attributed to an increase in fatty-acid oxidation. In accord with the lower PPARγ expression, there was reduced Glut4 mRNA expression; however, insulin-induced PKB phosphorylation was enhanced by gAd, and glucose uptake was not altered. To investigate the effect of gAd on LPS-induced inflammatory response, cells were treated with gAd either during differentiation or 24 h before induction of inflammation in differentiated adipocytes. LPS-induced inflammatory response, as indicated by increase in IL6 and MCP-1 mRNA expression. gAd given through differentiation was much more effective in abrogating LPS-dependent cytokines production than gAd given to the mature adipocyte. In conclusion, elevated gAd at differentiation of 3T3-L1 cells leads to reduced lipid content, reduced lipid metabolism and high resistance to inflammation.

Redox Balance in Type 2 Diabetes: Therapeutic Potential and the Challenge of Antioxidant-Based Therapy.

Oxidative stress is an important factor in the development of type 2 diabetes (T2D) and associated complications. Unfortunately, most clinical studies have failed to provide sufficient evidence regarding the benefits of antioxidants (AOXs) in treating this disease. Based on the known complexity of reactive oxygen species (ROS) functions in both the physiology and pathophysiology of glucose homeostasis, it is suggested that inappropriate dosing leads to the failure of AOXs in T2D treatment. To support this hypothesis, the role of oxidative stress in the pathophysiology of T2D is described, together with a summary of the evidence for the failure of AOXs in the management of diabetes. A comparison of preclinical and clinical studies indicates that suboptimal dosing of AOXs might explain the lack of benefits of AOXs. Conversely, the possibility that glycemic control might be adversely affected by excess AOXs is also considered, based on the role of ROS in insulin signaling. We suggest that AOX therapy should be given in a personalized manner according to the need, which is the presence and severity of oxidative stress. With the development of gold-standard biomarkers for oxidative stress, optimization of AOX therapy may be achieved to maximize the therapeutic potential of these agents.

Inherited stress resiliency prevents the development of metabolic alterations in diet-induced obese mice.

OBJECTIVE: Chronic stress promotes obesity and metabolic comorbidities. The ability of individuals to cope with stress may serve as an important parameter in the development of obesity-related metabolic outcomes. The aim of this study was to clarify whether differences in stress response affect metabolic health under obesity. METHODS: The study was performed in a selectively bred mouse model of social dominance (Dom) and submissiveness (Sub), which exhibit stress resilience or vulnerability, respectively. Mice were given a high-fat diet (HFD) or standard diet, followed by physiological, histological, and molecular analyses. RESULTS: The HFD caused hyperleptinemia, glucose intolerance, insulin resistance, steatosis of the liver and pancreas, and brown adipose tissue whitening in Sub mice, whereas Dom mice were protected from these consequences of the HFD. The HFD increased circulating levels of interleukin (IL)-1ß and induced the expression of proinflammatory genes in the liver and in epididymal white adipose tissue of Sub mice, with no changes in Dom mice. The Cox2 inhibitor celecoxib (15 mg/kg/d) reduced serum IL-1ß, improved glucose tolerance and insulin sensitivity, and prevented hepatic and brown adipose tissue whitening in HFD-fed Sub mice. CONCLUSIONS: The extent of stress resiliency is associated with inflammation and contributes to population heterogeneity in the development of healthy or unhealthy obesity.

Maintenance of redox state and pancreatic beta-cell function: role of leptin and adiponectin.

Whereas oxidative stress is linked to cellular damage, reactive oxygen species (ROS) are also believed to be involved in the propagation of signaling pathways. Studies on the role of ROS in pancreatic beta-cell physiology, in contrast to pathophysiology, have not yet been reported. In this study we investigate the importance of maintaining cellular redox state on pancreatic beta-cell function and viability, and the effects of leptin and adiponectin on this balance. Experiments were conducted on RINm and MIN6 pancreatic beta-cells. Leptin (1-100 ng/ml) and adiponectin (1-100 nM) increased ROS accumulation, as was determined by DCFDA fluorescence. Using specific inhibitors, we found that the increase in ROS levels was mediated by NADPH oxidase (Nox), but not by AMP kinase (AMPK) or phosphatidyl inositol 3 kinase (PI3K). Leptin and adiponectin increased beta-cell number as detected by the XTT method, but did not affect apoptosis, indicating that the increased cell number results from increased proliferation. The adipokines-induced increase in viability is ROS dependent as this effect was abolished by N-acetyl-L-cysteine (NAC) or PEG-catalase. In addition, insulin secretion was found to be regulated by alterations in redox state, but not by adipokines. Finally, the effects of the various treatments on activity and mRNA expression of several antioxidant enzymes were determined. Both leptin and adiponectin reduced mRNA levels of superoxide dismutase (SOD)1. Adiponectin also decreased SOD activity and increased catalase and glutathione peroxidase (GPx) activities in the presence of H2O2. The results of this study show that leptin and adiponectin, by inducing a physiological increase in ROS levels, may be positive regulators of beta-cell mass.

Integration of the Connectivity Map and Pathway Analysis to Predict Plant Extract's Medicinal Properties-The Study Case of Sarcopoterium spinosum L.

Medicinal properties of plants are usually identified based on knowledge of traditional medicine or using low-throughput screens for specific pharmacological activities. The former is very biased since it requires prior knowledge of plants' properties, while the latter depends on a specific screening system and will miss medicinal activities not covered by the screen. We sought to enrich our understanding of the biological activities of Sarcopoterium spinosum L. root extract based on transcriptome changes to uncover a plurality of possible pharmacological effects without the need for prior knowledge or functional screening. We integrated Gene Set Enrichment Analysis of the RNAseq data to identify pathways affected by the treatment of cells with the extract and perturbational signatures in the CMAP database to enhance the validity of the results. Activities of signaling pathways were measured using immunoblotting with phospho-specific antibodies. Mitochondrial membrane potential was assessed using JC-1 staining. SARS-CoV-2-induced cell killing was assessed in Vero E6 and A549 cells using an MTT assay. Here, we identified transcriptome changes following exposure of cultured cells to the medicinal plant Sarcopoterium spinosum L. root extract. By integrating algorithms of GSEA and CMAP, we confirmed known anti-cancer activities of the extract and predicted novel biological effects on oxidative phosphorylation and interferon pathways. Experimental validation of these pathways uncovered strong activation of autophagy, including mitophagy, and excellent protection from SARS-CoV-2 infection. Our study shows that gene expression analysis alone is insufficient for predicting biological effects since some of the changes reflect compensatory effects, and additional biochemical tests provide necessary corrections. This study defines the advantages and limitations of transcriptome analysis in predicting the biological and medicinal effects of the Sarcopoterium spinosum L. extract. Such analysis could be used as a general approach for predicting the medicinal properties of plants.

Single probe diffuse reflectance spectroscopy to assess the effect of sarcopoterium spinosum treatment on the cerebral tissue properties of ApoE knockout mouse.

In this work, diffuse near-infrared light reflectance spectroscopy based on a single optical probe, contains central single collection fiber surrounded by a circular array of illumination fibers, was used to quantify cerebral tissue properties in ApoE knockout mice following Sarcopoterium spinosum treatment. Sarcopoterium spinosum, also known as Thorny burnet, is a Mediterranean plant widely used as a traditional therapy for the treatment of a variety of pathologies, primarily type 2 diabetes mellitus (T2D). While it's efficacy in the treatment of T2D, and of other components of metabolic syndrome, have already been validated by us, the aim of this study was to investigate the effects of Sarcopoterium spinosum extract (SSE) on dyslipidemia and vascular functions. We utilized ApoE deficient mice (ApoE-/- , Atherosclerosis-prone apolipoprotein E-deficient), who have a severe impairment in plasma lipoprotein clearance and thus develop alterations in blood lipid profile and are highly susceptible to atherogenic plaque formation. A total of 34 male mice were divided into five groups representing various genetic, dietary, and treatment configurations. Optical measurements were used to assess changes in diffused reflectance spectra, optical properties (absorption and scattering), and cerebral tissue chromophore contents. Specifically, significant improvement in cerebral hemoglobin level was observed in ApoE KO mice, fed an artherogenic diet (ATD), upon SSE treatment. Biochemical and histological analyses of ApoE-/- ATD mice showed elevated body weight and a high level of blood triglycerides, free fatty acids and cholesterol. In contrast, in SSE treated mice improvement was observed, suggesting beneficial effects of SSE. In ApoE-/- ATD mice group a higher levels of deoxyhemoglobin was monitored indicating that the rate of oxygen release to the tissue is low. This was supported by decrease in oxygen saturation. It was also shown a reduction in water content in the brain of ApoE KO. Mice fed with the atherogenic diet demonstrated increased water content as compared to STD-fed ApoE KO mice, while SSE administration reversed the effect of the diet. To our knowledge, no such study has been reported before.

Grape pomace reduces the severity of non-alcoholic hepatic steatosis and the development of steatohepatitis by improving insulin sensitivity and reducing ectopic fat deposition in mice.

While non-alcoholic fatty liver disease (NAFLD) represents the common cause of chronic liver disease, specific therapies are currently unavailable. The wine industry produces millions of tons of residue (pomace), which contains high levels of bioactive phytochemicals. The aim of this study was to clarify the potential benefits of grape pomace for the treatment of NAFLD at different levels of severity, and to clarify the mechanism of action. C57Bl/6 mice were given high fat diet (HFD) or western diet (WD) as models of obesity and hepatic steatosis or steatohepatitis, respectively, with or without pomace supplementation (50-250 mg/day). Pomace inhibited food intake, and reduced serum leptin and body weight gain. Ectopic fat deposition was reduced, while white adipose tissue mass was preserved. In addition, pomace improved glucose tolerance and insulin sensitivity, prevented the development of adipose tissue inflammation, and reduced hepatic steatosis. Higher expression of genes involved in fatty acids transport and oxidation was observed in adipose tissue, while lipogenic genes were attenuated in the liver of pomace-treated mice. In WD-fed mice, pomace reduced the severity of hepatic steatosis and inflammation and improved blood lipid profile, but was ineffective in reversing hepatic damage of advanced NASH. In conclusion, pomace improved insulin sensitivity and reduced ectopic fat deposition, leading to a healthier metabolic profile. Pomace may hold the potential as a supplement with beneficial health outcomes for the prevention and treatment of hepatic steatosis and other obesity-related pathologies.

Multi-seasonal water-stress memory versus temperature-driven dynamic structural changes in grapevine.

Perennial plants perpetually adapt to environmental changes in complex and yet insufficiently understood manner. We aimed to separate the intra-seasonal temperature effects on structure and function from perennial and annual water stress effects. This study focused on grapevine (Vitis vinifera L. 'Cabernet Sauvignon') petioles, which being a continuously produced organ, represent the current status of the plant. Field-grown mature plants subjected to multi-annual irrigation treatments (severe water stress, mild water stress and non-stressed) throughout the growing season were compared with greenhouse-grown plants under three temperature regimes (22, 28 and 34 °C). Physiological and functional anatomy parameters were measured. A generalized additive model (GAM) based on meteorological and lysimeter-based field data was applied to determine the relative influence of various meteorological parameters on evapotranspiration (ETc) during the growing season in the field experiment. At the beginning of the growing season, in May, petioles in the severe stress treatment showed a stress-related structure (decreased length, safer hydraulic structure and increased lignification), though having high values of stem water potential (SWP). As the season progressed and temperatures increased, all water availability treatments petioles showed similar changes, and at the end of season, in August, were structurally very similar. Those changes were independent of SWP and were comparable to high temperature-induced changes in the greenhouse. In contrast, stems hydraulic structure was strongly influenced by water availability. Regression analyses indicated a relationship between petioles xylem structure and stomatal conductance (gs), whereas gs (but not SWP) was temperature-dependent. The GAM showed that ETc was mainly dependent on temperature. Our results indicate a perennial water-stress memory response, influencing the petiole structure at the beginning of the following season. Intra-seasonally, the petiole's structure becomes independent of water status, whereas temperature drives the structural changes. Thus, ongoing climate change might disrupt plant performance by purely temperature-induced effects.

Anti-inflammatory effects of Sarcopoterium spinosum extract.

ETHNOPHARMACOLOGICAL RELEVANCE: Sarcopoterium spinosum is a Mediterranean plant, widely used by the Beduin traditional medicine. While its major use is for the treatment of diabetes, there are also documentations for its use as anti-inflammatory agent. This therapeutic potential of Sarcopoterium spinosum was not validated before. AIM OF THE STUDY: To investigate the anti-inflammatory properties of Sarcopoterium spinosum extract (SSE). MATERIALS AND METHODS: Experiments were performed on RAW264.7 macrophages and bone marrow-derived macrophages (BMDM) and the effect of SSE on markers of inflammation was investigated. In addition, the effect of SSE on the development of inflammation in adipose-tissue of obese, insulin resistant mice was measured in KK-Ay mice and high fat diet (HFD)-fed mice. RESULTS: Lipopolysaccharide (LPS) and SSE increased the viability of RAW264.7. In addition, the cells acquired distinct dendritic-like morphology, however, while LPS induced NO production and the mRNA expression of pro-inflammatory cytokines, SSE increased the mRNA expression of anti-inflammatory genes and blocked LPS effects. All three pathways of MAPK were activated by LPS and SSE, as demonstrated by the phosphorylation of ERK, p38 and JNK. NFκB was activated and Akt was phosphorylated by LPS, while SSE blocked this effects. STAT proteins were differently phosphorylated by SSE and LPS. Immunomodulatory effects of SSE were also found in BMDM. In adipose tissue of SSE-treated mice, less crown-like structures were found, and lower expression of pro-inflammatory adipocytokines was observed, although adipocytes hypertrophy was not affected. CONCLUSIONS: SSE has an immunomodulatory effects that affect macrophage function, and reduces adipose tissue inflammation. Identifying active component and clarifying its mechanism of action might support the development of new anti-inflammatory agent.

C-reactive protein velocity to distinguish febrile bacterial infections from non-bacterial febrile illnesses in the emergency department.

INTRODUCTION: C-reactive protein (CRP) is a real-time and low-cost biomarker to distinguish febrile bacterial infections from non-bacterial febrile illnesses. We hypothesised that measuring the velocity of the biomarker instead of its absolute serum concentration could enhance its ability to differentiate between these two conditions. METHODS: We prospectively recruited adult patients (age >or= 18 years) who presented to the emergency department with fever. We recorded their data regarding the onset of fever and accompanying symptoms. CRP measurements were obtained upon admission. CRP velocity (CRPv) was defined as the ratio between CRP on admission and the number of hours since the onset of fever. Patients were diagnosed by clinical symptoms, blood cultures and imaging studies, and the diagnoses were confirmed by an infectious disease specialist. The efficacy of CRPv as a diagnostic marker was evaluated by using receiver operator curves (ROC). Excluded were patients who did not know the time fever started with certainty, patients with malignancy, patients with HIV infection and patients who had been using antibiotics upon presentation. RESULTS: Of 178 eligible patients, 108 (60.7%) had febrile bacterial infections (mean CRP: 63.77 mg/L, mean CRPv: 3.61 mg/L/hour) and 70 (39.3%) had non-bacterial febrile illnesses (mean CRP: 23.2 mg/L, mean CRPv: 0.41 mg/L/hour). The area under the curve for CRP and CRPv were 0.783 (95% confidence interval (CI) = 0.717 to 0.850) and 0.871 (95% CI = 0.817 to 0.924), respectively. In a 122-patient subgroup with a CRP level of less than 100 mg/L, the area under the curve increased from 0.689 (95% CI = 0.0595 to 0.782) to 0.842 (95% CI = 0.77 to 0.914) by using the CRPv measurements. CONCLUSIONS: CRPv improved differentiation between febrile bacterial infections and non-bacterial febrile illnesses compared with CRP alone, and could identify individuals who need prompt therapeutic intervention.

Sarcopoterium spinosum Inhibited the Development of Non-Alcoholic Steatosis and Steatohepatitis in Mice.

Non-alcoholic fatty liver disease (NAFLD) is a comorbidity of obesity, which gradually develops from hepatic steatosis into steatohepatitis (NASH) and eventually even into fibrosis or hepatic carcinoma. To date, there has been no specific and effective treatment for NAFLD. Sarcopoterium spinosum extract (SSE) was found to improve insulin sensitivity. Recognizing the intimate link between insulin resistance and NAFLD, the aim of this study was to investigate the effectivity of SSE in the prevention and management of NAFLD at various severities. SSE was given to high-fat diet (HFD)-fed mice (steatosis model) or to mice given a Western diet (WD) in the short or long term (NASH prevention or treatment, respectively). SSE reduced body weight accumulation, improved glucose tolerance and insulin sensitivity and prevented the development of hepatic steatosis. SSE also blocked the progression of liver disease toward NASH in a dose-dependent manner. The expression of genes involved in lipid metabolism, inflammation, and antioxidant machinery was regulated by SSE in both models of steatosis and NASH development. However, SSE failed to reverse the hepatic damage in the advanced model of NASH. In summary, SSE might be considered as a botanical supplement for the prevention and treatment of hepatic steatosis, and for slowing the deterioration toward NASH.

Multiparameter wide-field integrated optical imaging system-based spatially modulated illumination and laser speckles in model of tissue injuries.

In this report, an integrated optical platform based on spatial illumination together with laser speckle contrast technique was utilized to measure multiple parameters in live tissue including absorption, scattering, saturation, composition, metabolism, and blood flow. Measurements in three models of tissue injury including drug toxicity, artery occlusion, and acute hyperglycemia were used to test the efficacy of this system. With this hybrid apparatus, a series of structured light patterns at low and high spatial frequencies are projected onto the tissue surface and diffuse reflected light is captured by a CCD camera. A six position filter wheel, equipped with four bandpass filters centered at wavelengths of 650, 690, 800 and 880 nm is placed in front of the camera. Then, light patterns are blocked and a laser source at 650 nm illuminates the tissue while the diffusely reflected light is captured by the camera through the two remaining open holes in the wheel. In this manner, near-infrared (NIR) and laser speckle images are captured and stored together in the computer for off-line processing to reconstruct the tissue's properties. Spatial patterns are used to differentiate the effects of tissue scattering from those of absorption, allowing accurate quantification of tissue hemodynamics and morphology, while a coherent light source is used to study blood flow changes, a feature which cannot be measured with the NIR structured light. This combined configuration utilizes the strengths of each system in a complementary way, thus collecting a larger range of sample properties. In addition, once the flow and hemodynamics are measured, tissue oxygen metabolism can be calculated, a property which cannot be measured independently. Therefore, this merged platform can be considered a multiparameter wide-field imaging and spectroscopy modality. Overall, experiments demonstrate the capability of this spatially coregistered imaging setup to provide complementary, useful information of various tissue metrics in a simple and noncontact manner, making it attractive for use in a variety of biomedical applications.

Decreased cerebral blood flow and hemodynamic parameters during acute hyperglycemia in mice model observed by dual-wavelength speckle imaging.

In this study, we use dual-wavelength optical imaging-based laser speckle technique to assess cerebral blood flow and metabolic parameters in a mouse model of acute hyperglycemia (high blood glucose). The effect of acute glucose levels on physiological processes has been extensively described in multiple organ systems such as retina, kidney, and others. We postulated that hyperglycemia also alters brain function, which in turn can be monitored optically using dual-wavelength laser speckle imaging (DW-LSI) platform. DW-LSI is a wide-field, noncontact optical imaging modality that integrates the principles of laser flowmetry and oximetry to obtain macroscopic information such as hemoglobin concentration and blood flow. A total of eight mice (C57/BL6) were used, randomized into two groups of normoglycemia (control, n = 3) and hyperglycemia (n = 5). Hyperglycemia was induced by intraperitoneal injection of a commonly used anesthetic drug combining ketamine and xylazine (KX combo). We found that this KX combo increases blood glucose (BG) levels from 150 to 350 mg/dL, approximately, when measured 18 minutes post-administration. BG continues to increase throughout the test period, with BG reaching an average of 463 ± 20.34 mg/dL within 60 minutes. BG levels were measured every 10 minutes from tail blood using commercially available glucometer. Experimental results demonstrated reductions in cerebral blood flow (CBF) by 55%, tissue oxygen saturation (SO2 ) by 15%, and cerebral metabolic rate of oxygen (CMRO2 ) by 75% following acute hyperglycemia. The observed decrease in these parameters was consistent with results reported in the literature, measured by a variety of experimental techniques. Measurements with laser Doppler flowmetry (LDF) were also performed which confirmed a reduction in CBF following acute hyperglycemia. In summary, our findings indicate that acute hyperglycemia modified brain hemodynamic response and induced significant changes in blood flow and metabolism. As far as we are aware, the implementation of the DW-LSI to monitor brain hemodynamic and metabolic response to acute hyperglycemia in intact mouse brain has not been previously reported.

Activation of Insulin Signaling in Adipocytes and Myotubes by Sarcopoterium Spinosum Extract.

Sarcopoterium spinosum (S. spinosum) is a medicinal plant, traditionally used as an antidiabetic remedy. Previous studies demonstrated its beneficial properties in the treatment of insulin resistance. The aim of this study was to further clarify the effect of S. spinosum extract (SSE) on insulin signaling. Phosphoproteomic analysis, performed in 3T3-L1 adipocytes treated with SSE, revealed the activation of insulin receptor pathways. SSE increased Glut4-facilitated glucose uptake in adipocytes, with an additive effect between SSE and insulin. While the maximal effect of insulin on glucose uptake was found at days 15-16 of differentiation, SSE-induced glucose uptake was found at an earlier stage of differentiation. Inhibition of PI3K and Akt blocked SSE-dependent glucose uptake. Western blot analysis, performed on 3T3-L1 adipocytes and L6 myotubes, showed that in contrast to insulin action, Akt was only marginally phosphorylated by SSE. Furthermore, GSK3ß and PRAS40 phosphorylation as well as glucose uptake were increased by the extract. SSE also induced the phosphorylation of ERK similar to insulin. In conclusion, SSE activates insulin signaling, although the upstream event mediating its effects should be further clarified. Identifying the active molecules in SSE may lead to the development of new agents for the treatment of insulin resistance.

N-Acetyl-l-Cysteine Supplement in Early Life or Adulthood Reduces Progression of Diabetes in Nonobese Diabetic Mice.

BACKGROUND: Oxidative stress contributes to the pathologic process leading to the development, progression, and complications of type 1 diabetes (T1D). OBJECTIVE: The aim of this study was to investigate the effect of the antioxidant N-acetyl-l-cysteine (NAC), supplemented during early life or adulthood on the development of T1D. METHODS: NAC was administered to nonobese diabetic (NOD) female mice during pregnancy and lactation, and the development of diabetes was followed in offspring. In an additional set of experiments, offspring of untreated mice were given NAC during adulthood, and the development of T1D was followed. Morbidity rate, insulitis and serum cytokines were measured in the 2 sets of experiments. In addition, markers of oxidative stress, glutathione, lipid peroxidation, total antioxidant capacity and activity of antioxidant enzymes, were followed. RESULTS: Morbidity rate was reduced in both treatment protocols. A decrease in interferon γ, tumor necrosis factor α, interleukin 1α, and other type 1 diabetes-associated proinflammatory cytokines was found in mice supplemented with NAC in adulthood or during early life compared with control NOD mice. The severity of insulitis was higher in control NOD mice than in treated groups. NAC administration significantly reduced oxidative stress, as determined by reduced lipid peroxidation and increased total antioxidant capacity in serum and pancreas of mice treated in early life or in adulthood and increased pancreatic glutathione when administrated in adulthood. The activity of antioxidant enzymes was not affected in mice given NAC in adulthood, whereas an increase in the activity of superoxide dismutase and catalase was demonstrated in the pancreas of their offspring. CONCLUSION: NAC decreased morbidity of NOD mice by attenuating the immune response, presumably by eliminating oxidative stress, and might be beneficial in reducing morbidity rates of T1D in high-risk individuals.

Related to testes-specific, vespid, and pathogenesis protein-1 (RTVP-1) is overexpressed in gliomas and regulates the growth, survival, and invasion of glioma cells.

In this study, we examined the expression and functions of related to testes-specific, vespid, and pathogenesis protein 1 (RTVP-1) in glioma cells. RTVP-1 was expressed in high levels in glioblastomas, whereas its expression in low-grade astrocytomas and normal brains was very low. Transfection of glioma cells with small interfering RNAs targeting RTVP-1 decreased cell proliferation in all the cell lines examined and induced cell apoptosis in some of them. Overexpression of RTVP-1 increased astrocyte and glioma cell proliferation and the anchorage-independent growth of the cells. In addition, overexpression of RTVP-1 rendered glioma cells more resistant to the apoptotic effect of tumor necrosis factor-related apoptosis-inducing ligand and serum deprivation. To delineate the molecular mechanisms involved in the survival effects of RTVP-1, we examined the expression and phosphorylation of various apoptosis-related proteins. We found that overexpression of RTVP-1 decreased the phosphorylation of c-Jun-NH2-kinase and increased the expression of Bcl2 and that the protective effect of RTVP-1 was partially mediated by Bcl2. Finally, we found that RTVP-1 regulated the invasion of glioma cells as was evident by their enhanced migration through Matrigel and by their increased invasion in a spheroid confrontation assay. The increased invasive potential of the RTVP-1 overexpressors was also shown by the increased activity of matrix metalloproteinase 2 in these cells. Our results suggest that the expression of RTVP-1 is correlated with the degree of malignancy of astrocytic tumors and that RTVP-1 is involved in the regulation of the growth, survival, and invasion of glioma cells. Collectively, these findings suggest that RTVP-1 is a potential therapeutic target in gliomas.