Knockout of Dual-Specificity Protein Phosphatase 5 Protects Against Hypertension-Induced Renal Injury.

Dual-specificity protein phosphatase 5 (DUSP5) is a member of the tyrosine-threonine phosphatase family with the ability to dephosphorylate and inactivate extracellular signal-related kinase (ERK). The present study investigates whether knockout (KO) of Dusp5 improves renal hemodynamics and protects against hypertension-induced renal injury. The renal expression of DUSP5 was reduced, and the levels of phosphorylated (p) ERK1/2 and p-protein kinase C (PKC) α were elevated in the KO rats. KO of Dusp5 enhanced the myogenic tone of the renal afferent arteriole and interlobular artery in vitro with or without induction of deoxycorticosterone acetate-salt hypertension. Inhibition of ERK1/2 and PKC diminished the myogenic response to a greater extent in Dusp5 KO rats. Autoregulation of renal blood flow was significantly impaired in hypertensive wild-type (WT) rats but remained intact in Dusp5 KO animals. Proteinuria was markedly decreased in hypertensive KO versus WT rats. The degree of glomerular injury was reduced, and the expression of nephrin in the glomerulus was higher in hypertensive Dusp5 KO rats. Renal fibrosis and medullary protein cast formation were attenuated in hypertensive Dusp5 KO rats in association with decreased expression of monocyte chemoattractant protein 1, transforming growth factor-ß1, matrix metalloproteinase (MMP) 2, and MMP9. These results indicate that KO of Dusp5 protects against hypertension-induced renal injury, at least in part, by maintaining the myogenic tone of the renal vasculature and extending the range of renal blood flow autoregulation to higher pressures, which diminish glomerular injury, protein cast formation, macrophage infiltration, and epithelial-mesenchymal transformation in the kidney. SIGNIFICANCE STATEMENT: Dual-specificity protein phosphatase 5 (DUSP5) is a tyrosine-threonine phosphatase that inactivates extracellular signal-related kinase (ERK). We previously reported that knockout (KO) of Dusp5 enhanced the myogenic response and autoregulation in the cerebral circulation. The present study investigates whether KO of DUSP5 improves renal hemodynamics and protects against hypertension-induced renal injury. Downregulation of DUSP5 enhanced the myogenic tone of renal arteriole and artery and autoregulation of renal blood flow in association with reduced proteinuria, glomerular injury, and interstitial fibrosis after the induction of hypertension. Inhibition of ERK1/2 and protein kinase C diminished the myogenic response to a greater extent in Dusp5 KO rats. These results suggest that DUSP5 might be a viable drug target for the treatment of hypertension nephropathy.

Regulator of G Protein Signaling 2 Facilitates Uterine Artery Adaptation During Pregnancy in Mice.

Background Decreased uterine blood flow is known to contribute to pregnancy complications such as gestational hypertension and preeclampsia. Previously, we showed that the loss of regulator of G protein signaling 2 ( RGS 2), a GTP ase activating protein for Gq/11 and Gi/o class G proteins, decreases uterine blood flow in the nonpregnant state in mice. Here, we examined the effects of the absence of RGS 2 and 5 on uterine blood flow and uterine vascular structure and function at early, mid, and late gestation, as well as peripartum period in mice. Methods and Results Abdominal Doppler ultrasonography was performed on adult female wild-type, Rgs2-/-, and Rgs5-/- mice at pre-pregnancy, gestational days 10, 15, and 18, and postpartum day 3. Uterine artery structure and function were also assessed by vessel myograph studies. At mid-pregnancy, uterine blood flow decreased in both Rgs2-/- and Rgs5-/- mice, whereas resistive index increased only in Rgs2-/- mice. In uterine arteries from wild-type mice, mRNA expression of RGS 2 and 4 increased, whereas RGS 5 expression remained elevated at mid-pregnancy. These changes in gene expression were unique to uterine arteries because they were absent in mesenteric arteries and the aorta of wild-type mice. In Rgs2-/- mice, uterine artery medial cross-sectional area and G protein-coupled receptor-mediated vasoconstriction increased in mid-pregnancy, implicating a role for RGS 2 in structural and functional remodeling of uterine arteries during pregnancy. In contrast, RGS 5 absence increased vasoconstriction only in the peripartum period. Conclusions These data together indicate that RGS 2 plays a critical role in the structural and functional remodeling of uterine arteries to impact uterine blood flow during pregnancy. Targeting the signaling pathway regulated by RGS 2 may therefore be a therapeutic strategy for ameliorating utero-placental perfusion disorders during pregnancy.

Regulated necrosis in kidney ischemia-reperfusion injury.

Ischemia-reperfusion injury (IRI) is the outcome of an inflammatory process that is triggered when an organ undergoes a transient reduction or cessation of blood flow, followed by re-establishment of perfusion. In the clinical setting, IRI contributes to significant acute kidney injury, patient morbidity and mortality, and adverse outcomes in transplantation. Tubular cell death by necrosis and apoptosis is a central feature of renal IRI. Recent research has challenged traditional views of cell death by identifying new pathways in which cells die in a regulated manner but with the morphologic features of necrosis. This regulated necrosis (RN) takes several forms, with necroptosis and ferroptosis being the best described. The precise mechanisms and relationships between the RN pathways in renal IRI are currently the subject of active research. The common endpoint of RN is cell membrane rupture, resulting in the release of cytosolic components with subsequent inflammation and activation of the immune system. We review the evidence and mechanisms of RN in the kidney following renal IRI, and discuss the use of small molecule inhibitors and genetically modified mice to better understand this process and guide potentially novel therapeutic interventions.

Intradermal delivery of modified mRNA encoding VEGF-A in patients with type 2 diabetes.

Chemically modified mRNA is an efficient, biocompatible modality for therapeutic protein expression. We report a first-time-in-human study of this modality, aiming to evaluate safety and potential therapeutic effects. Men with type 2 diabetes mellitus (T2DM) received intradermal injections of modified mRNA encoding vascular endothelial growth factor A (VEGF-A) or buffered saline placebo (ethical obligations precluded use of a non-translatable mRNA control) at randomized sites on the forearm. The only causally treatment-related adverse events were mild injection-site reactions. Skin microdialysis revealed elevated VEGF-A protein levels at mRNA-treated sites versus placebo-treated sites from about 4-24 hours post-administration. Enhancements in basal skin blood flow at 4 hours and 7 days post-administration were detected using laser Doppler fluximetry and imaging. Intradermal VEGF-A mRNA was well tolerated and led to local functional VEGF-A protein expression and transient skin blood flow enhancement in men with T2DM. VEGF-A mRNA may have therapeutic potential for regenerative angiogenesis.

Genetic ablation of NFAT5/TonEBP in smooth muscle cells impairs flow- and pressure-induced arterial remodeling in mice.

The arterial wall adapts to alterations in blood flow and pressure by remodeling the cellular and extracellular architecture. Biomechanical stress of vascular smooth muscle cells (VSMCs) in the media is thought to precede this process and promote their activation and subsequent proliferation. However, molecular determinants orchestrating the transcriptional phenotype under these conditions have been insufficiently studied. We identified the transcription factor, nuclear factor of activated T cells 5 (NFAT5; or tonicity enhancer-binding protein) as a crucial regulatory element of mechanical stress responses of VSMCs. Here, the relevance of NFAT5 for arterial growth and thickening is investigated in mice upon inducible smooth muscle cell (SMC)-specific genetic ablation of Nfat5. In cultured mouse VSMCs, loss of Nfat5 inhibits the expression of gene sets involved in the control of the cell cycle and the interaction with the extracellular matrix and cytoskeletal dynamics. In vivo, SMC-specific knockout of Nfat5 did not affect the general vascular architecture and blood pressure levels under baseline conditions. However, proliferation of VSMCs and the thickening of the arterial wall were inhibited during both flow-induced collateral remodeling and hypertension-mediated arterial hypertrophy. Whereas originally described as a hypertonicity-responsive transcription factor, these findings identify NFAT5 as a novel molecular determinant of biomechanically induced phenotype changes of VSMCs and wall stress-induced arterial remodeling processes.-Arnold, C., Feldner, A., Zappe, M., Komljenovic, D., De La Torre, C., Ruzicka, P., Hecker, M., Neuhofer, W., Korff, T. Genetic ablation of NFAT5/TonEBP in smooth muscle cells impairs flow- and pressure-induced arterial remodeling in mice.

Are the Variants of the Circle of Willis Determined by Genetic or Environmental Factors? Results of a Twin Study and Review of the Literature.

BACKGROUND: Anatomic variants of the circle of Willis (CW) are commonly observed in healthy subjects. Genetic and environmental factors influencing these variants remain unclear. Our aim was to assess the genetic and environmental background affecting variant CW phenotypes. METHODS: A total of 122 adult healthy twins from the Hungarian Twin Registry (39 monozygotic (MZ) and 22 dizygotic (DZ) pairs, average age 49.7 ± 13.4 years) underwent Time-of-Flight magnetic resonance angiography and transcranial Doppler sonography. We investigated the anterior and posterior CW according to morphological categories. Prevalence and concordance rates of CW variants were calculated. MZ twins discordant for CW variants were analyzed for cardiovascular risk factors and altered blood flow. RESULTS: Complete CW (45.0%) and bilaterally absent posterior communicating artery (PCoA) (22.5%) were the most prevalent variants in the anterior and posterior CW, respectively. There was no significant difference regarding the prevalence of variants across zygosity except for bilaterally hypoplastic PCoA (p = .02). DZ concordance was higher compared to MZ twins regarding morphological categories of the CW. Cardiovascular risk factors were not significantly associated with variant CW in MZ twins discordant to CW morphology. Flow parameters did not differ significantly among MZ twins discordant to CW variants. CONCLUSION: CW variants may not be determined by substantial genetic effects and are not influenced by altered blood flow in healthy individuals. Further investigations are needed to identify potential environmental factors affecting these variants.

STEEL participates in fracture healing through upregulating angiogenesis-related genes by recruiting PARP 1.

OBJECTIVE: To explore the effect of STEEL on fracture healing and its underlying mechanism. PATIENTS AND METHODS: A total of 31 patients with long bone fracture and who received reoperation because of bone nonunion, delayed union or healing disorder in the Wuxi Nine Hospital Affiliated to Soochow University from July 2016 to February 2018 were selected. The bone callus at the fracture site was collected from each patient during the reoperation. QRT-PCR (Quantitative Real-Time Polymerase Chain Reaction) was used to detect STEEL expression in the callus tissues of the treatment group (bone nonunion or delayed union) and the control group. In addition, we measured the number of blood vessels in the fracture tissues by immunohistochemistry. After the construction of tibial fracture model in mice, STEEL expression and the total number of blood vessels in the treatment group (sawing treatment) and the control group (sham operation) were detected, respectively. For in vitro experiments, CCK-8 (cell counting kit-8) assay was performed to detect cell proliferation after knockdown or overexpression of STEEL in the vascular endothelial cells. The binding condition of STEEL and its interacting proteins were detected by RIP (RNA binding protein immunoprecipitation), and the binding of PARP 1 [poly (ADP-ribose) polymerase 1] with gene promoter was observed by ChIP (chromatin immunoprecipitation assay). Western blot was used to detect the expression level of VEGF (vascular endothelial growth factor). RESULTS: STEEL expression and the vascular density in the callus tissues of the treatment group were significantly lower than those of the control group. Downregulated STEEL remarkably decreased the proliferation ability of HUVEC cells. Meanwhile, the vascular density was also significantly decreased in mice with a tibial fracture. Overexpressed STEEL obtained the opposite results. STEEL could interact with PARP 1 to regulate expressions of downstream genes. Moreover, STEEL could also promote angiogenesis by elevating VEGF expression. CONCLUSIONS: We showed that STEEL expression could partly represent the angiogenesis of fracture sites. Moreover, it promoted angiogenesis by elevating VEGF expression.

Intramuscular E-selectin/adeno-associated virus gene therapy promotes wound healing in an ischemic mouse model.

BACKGROUND: Poor wound healing in critical limb ischemia (CLI) is attributed to impaired neovascularization and reperfusion. Optimizing the ischemic wound with adhesion molecules that enhance stem cell homing may revolutionize treatment. The purpose of this study is to test the efficacy of adhesion molecule E-selectin on wound healing in an ischemic mouse wound. METHODS: Adult FVB/NJ mice underwent unilateral femoral artery and vein ligation to induce CLI. A 4-mm punch biopsy wound was created on the anterior thigh to simulate ischemic wounds. Intramuscular injection of adeno-associated virus (AAV) carrying either E-selectin (E-selectin/AAV, n = 11) or LacZ as control (LacZ/AAV, n = 10) was performed. Gross wound size was measured for 10 d postoperatively. Ischemic hindlimb reperfusion was quantified using laser Doppler imaging. Wound tissue neovascularization was visualized using DiI perfusion and confocal microscopy. E-selectin expression in wounds was verified by immunofluorescence. RESULTS: Immunofluorescence confirmed E-selectin/AAV delivery in treatment versus control limbs. Wounds from E-selectin/AAV mice versus controls revealed surface area healing of 54% versus 20% (P < 0.01) on postoperative day (POD) 1, 78% versus 51% on POD 4 (P < 0.01), and 97% versus 84% on POD 10 (P < 0.01). Laser Doppler imaging revealed greater reperfusion in E-selectin/AAV mice versus controls by POD 10 (0.49 versus 0.27, P < 0.05). DiI perfused ligated hindlimb in E-selectin/AAV versus control mice revealed mean neovascularization intensity score of 30 versus 18 (P < 0.05) on POD 10. CONCLUSIONS: Intramuscularly injected E-selectin/AAV gene therapy in mice with CLI significantly increases wound angiogenesis and limb reperfusion, expediting overall wound healing.

Dynamic, heterogeneous endothelial Tie2 expression and capillary blood flow during microvascular remodeling.

Microvascular endothelial cell heterogeneity and its relationship to hemodynamics remains poorly understood due to a lack of sufficient methods to examine these parameters in vivo at high resolution throughout an angiogenic network. The availability of surrogate markers for functional vascular proteins, such as green fluorescent protein, enables expression in individual cells to be followed over time using confocal microscopy, while photoacoustic microscopy enables dynamic measurement of blood flow across the network with capillary-level resolution. We combined these two non-invasive imaging modalities in order to spatially and temporally analyze biochemical and biomechanical drivers of angiogenesis in murine corneal neovessels. By stimulating corneal angiogenesis with an alkali burn in Tie2-GFP fluorescent-reporter mice, we evaluated how onset of blood flow and surgically-altered blood flow affects Tie2-GFP expression. Our study establishes a novel platform for analyzing heterogeneous blood flow and fluorescent reporter protein expression across a dynamic microvascular network in an adult mammal.

Human venous valve disease caused by mutations in FOXC2 and GJC2.

Venous valves (VVs) prevent venous hypertension and ulceration. We report that FOXC2 and GJC2 mutations are associated with reduced VV number and length. In mice, early VV formation is marked by elongation and reorientation ("organization") of Prox1hi endothelial cells by postnatal day 0. The expression of the transcription factors Foxc2 and Nfatc1 and the gap junction proteins Gjc2, Gja1, and Gja4 were temporospatially regulated during this process. Foxc2 and Nfatc1 were coexpressed at P0, and combined Foxc2 deletion with calcineurin-Nfat inhibition disrupted early Prox1hi endothelial organization, suggesting cooperative Foxc2-Nfatc1 patterning of these events. Genetic deletion of Gjc2, Gja4, or Gja1 also disrupted early VV Prox1hi endothelial organization at postnatal day 0, and this likely underlies the VV defects seen in patients with GJC2 mutations. Knockout of Gja4 or Gjc2 resulted in reduced proliferation of Prox1hi valve-forming cells. At later stages of blood flow, Foxc2 and calcineurin-Nfat signaling are each required for growth of the valve leaflets, whereas Foxc2 is not required for VV maintenance.

Defining the role of corticotropin releasing factor binding protein in alcohol consumption.

The corticotropin releasing factor (CRF) exerts its effects by acting on its receptors and on the binding protein (CRFBP), and has been implicated in alcohol use disorder (AUD). Therefore, identification of the exact contribution of each protein that mediates CRF effects is necessary to design effective therapeutic strategies for AUD. A series of in vitro/in vivo experiments across different species were performed to define the biological discrete role of CRFBP in AUD. First, to establish the CRFBP role in receptor signaling, we developed a novel chimeric cell-based assay and showed that CFRBP full length can stably be expressed on the plasma membrane. We discovered that only CRFBP(10 kD) fragment is able to potentiate CRF-intracellular Ca2+ release. We provide evidence that CRHBP gene loss increased ethanol consumption in mice. Then, we demonstrate that selective reduction of CRHBP expression in the center nucleus of the amygdala (CeA) decreases ethanol consumption in ethanol-dependent rats. CRFBP amygdalar downregulation, however, does not attenuate yohimbine-induced ethanol self-administration. This effect was associated with decreased hemodynamic brain activity in the CRFBP-downregulated CeA and increased hemodynamic activity in the caudate putamen during yohimbine administration. Finally, in alcohol-dependent patients, genetic variants related to the CRFBP(10 kD) fragment were associated with greater risk for alcoholism and anxiety, while other genetic variants were associated with reduced risk for anxiety. Taken together, our data provide evidence that CRFBP may possess both inhibitory and excitatory roles and may represent a novel pharmacological target for the treatment of AUD.

High-Density Lipoproteins Rescue Diabetes-Impaired Angiogenesis via Scavenger Receptor Class B Type I.

Disordered neovascularization and impaired wound healing are important contributors to diabetic vascular complications. We recently showed that high-density lipoproteins (HDLs) enhance ischemia-mediated neovascularization, and mounting evidence suggests HDL have antidiabetic properties. We therefore hypothesized that HDL rescue diabetes-impaired neovascularization. Streptozotocin-induced diabetic mice had reduced blood flow recovery and neovessel formation in a hindlimb ischemia model compared with nondiabetic mice. Reconstituted HDL (rHDL) infusions in diabetic mice restored blood flow recovery and capillary density to nondiabetic levels. Topical rHDL application rescued diabetes-impaired wound closure, wound angiogenesis, and capillary density. In vitro, rHDL increased key mediators involved in hypoxia-inducible factor-1α (HIF-1α) stabilization, including the phosphoinositide 3-kinase/Akt pathway, Siah1, and Siah2, and suppressed the prolyl hydroxylases (PHD) 2 and PHD3. rHDL rescued high glucose-induced impairment of tubulogenesis and vascular endothelial growth factor (VEGF) A protein production, a finding associated with enhanced phosphorylation of proangiogenic mediators VEGF receptor 2 and endothelial nitric oxide synthase. Siah1/2 small interfering RNA knockdown confirmed the importance of HIF-1α stability in mediating rHDL action. Lentiviral short hairpin RNA knockdown of scavenger receptor class B type I (SR-BI) in vitro and SR-BI(-/-) diabetic mice in vivo attenuated rHDL rescue of diabetes-impaired angiogenesis, indicating a key role for SR-BI. These findings provide a greater understanding of the vascular biological effects of HDL, with potential therapeutic implications for diabetic vascular complications.

Ghrelin Promotes Functional Angiogenesis in a Mouse Model of Critical Limb Ischemia Through Activation of Proangiogenic MicroRNAs.

Current therapeutic strategies for the treatment of critical limb ischemia (CLI) have only limited success. Recent in vitro evidence in the literature, using cell lines, proposes that the peptide hormone ghrelin may have angiogenic properties. In this study, we aim to investigate if ghrelin could promote postischemic angiogenesis in a mouse model of CLI and, further, identify the mechanistic pathway(s) that underpin ghrelin's proangiogenic properties. CLI was induced in male CD1 mice by femoral artery ligation. Animals were then randomized to receive either vehicle or acylated ghrelin (150 µg/kg sc) for 14 consecutive days. Subsequently, synchrotron radiation microangiography was used to assess hindlimb perfusion. Subsequent tissue samples were collected for molecular and histological analysis. Ghrelin treatment markedly improved limb perfusion by promoting the generation of new capillaries and arterioles (internal diameter less than 50 µm) within the ischemic hindlimb that were both structurally and functionally normal; evident by robust endothelium-dependent vasodilatory responses to acetylcholine. Molecular analysis revealed that ghrelin's angiogenic properties were linked to activation of prosurvival Akt/vascular endothelial growth factor/Bcl-2 signaling cascade, thus reducing the apoptotic cell death and subsequent fibrosis. Further, ghrelin treatment activated proangiogenic (miR-126 and miR-132) and antifibrotic (miR-30a) microRNAs (miRs) while inhibiting antiangiogenic (miR-92a and miR-206) miRs. Importantly, in vitro knockdown of key proangiogenic miRs (miR-126 and miR-132) inhibited the angiogenic potential of ghrelin. These results therefore suggest that clinical use of ghrelin for the early treatment of CLI may be a promising and potent inducer of reparative vascularization through modulation of key molecular factors.

Analysis of blood flow and local expression of angiogenesis­associated growth factors in infected wounds treated with negative pressure wound therapy.

Angiogenesis is involved in the wound healing process. Increased angiogenesis and blood flow constitute a major mechanism of negative pressure wound therapy (NPWT), which has been shown to facilitate the healing of infected wounds. However, the effect on the expression of angiogensis­related growth factor remains unknown. The goal of the current study was to investigate the angiogenic factor levels prior to and following NPWT in infected wounds. A total of 20 patients with infected wounds treated with NPWT were included in the study. Patients acted as their own control; the postoperative measurements of patients were considered as the experimental group, while preoperative measurements were considered as the controlled group. Blood flow was recorded prior to and during NPWT. A total of 10 angiogensis­related growth factors were detected using a protein biochip array to analyze the change in protein levels prior to NPWT, and on the third day during NPWT. All wounds were successfully reconstructed by skin grafting or using local flaps following NPWT. NPWT resulted in significantly increased blood flow in the wound. There was a significant increase in vascular endothelial growth factor (VEGF), EGF, platelet­derived growth factor and angiotesin­2 following NPWT, while basic fibroblast growth factor decreased significantly. NPWT affects the local expression of angiogenesis­associated growth factors, which represents another mechanism to explain how NPWT accelerates wound healing.

Connexin 36 mediates blood cell flow in mouse pancreatic islets.

The insulin-secreting ß-cells are contained within islets of Langerhans, which are highly vascularized. Blood cell flow rates through islets are glucose-dependent, even though there are no changes in blood cell flow within in the surrounding exocrine pancreas. This suggests a specific mechanism of glucose-regulated blood flow in the islet. Pancreatic islets respond to elevated glucose with synchronous pulses of electrical activity and insulin secretion across all ß-cells in the islet. Connexin 36 (Cx36) gap junctions between islet ß-cells mediate this synchronization, which is lost in Cx36 knockout mice (Cx36(-/-)). This leads to glucose intolerance in these mice, despite normal plasma insulin levels and insulin sensitivity. Thus, we sought to investigate whether the glucose-dependent changes in intraislet blood cell flow are also dependent on coordinated pulsatile electrical activity. We visualized and quantified blood cell flow using high-speed in vivo fluorescence imaging of labeled red blood cells and plasma. With the use of a live animal glucose clamp, blood cell flow was measured during either hypoglycemia (∼50 mg/dl) or hyperglycemia (∼300 mg/dl). In contrast to the large glucose-dependent islet blood velocity changes observed in wild-type mice, only minimal differences are observed in both Cx36(+/-) and Cx36(-/-) mice. This observation supports a novel model where intraislet blood cell flow is regulated by the coordinated electrical activity in the islet ß-cells. Because Cx36 expression and function is reduced in type 2 diabetes, the resulting defect in intraislet blood cell flow regulation may also play a significant role in diabetic pathology.

Genomic and metabolic disposition of non-obese type 2 diabetic rats to increased myocardial fatty acid metabolism.

Lipotoxicity of the heart has been implicated as a leading cause of morbidity in Type 2 Diabetes Mellitus (T2DM). While numerous reports have demonstrated increased myocardial fatty acid (FA) utilization in obese T2DM animal models, this diabetic phenotype has yet to be demonstrated in non-obese animal models of T2DM. Therefore, the present study investigates functional, metabolic, and genomic differences in myocardial FA metabolism in non-obese type 2 diabetic rats. The study utilized Goto-Kakizaki (GK) rats at the age of 24 weeks. Each rat was imaged with small animal positron emission tomography (PET) to estimate myocardial blood flow (MBF) and myocardial FA metabolism. Echocardiograms (ECHOs) were performed to assess cardiac function. Levels of triglycerides (TG) and non-esterified fatty acids (NEFA) were measured in both plasma and cardiac tissues. Finally, expression profiles for 168 genes that have been implicated in diabetes and FA metabolism were measured using quantitative PCR (qPCR) arrays. GK rats exhibited increased NEFA and TG in both plasma and cardiac tissue. Quantitative PET imaging suggests that GK rats have increased FA metabolism. ECHO data indicates that GK rats have a significant increase in left ventricle mass index (LVMI) and decrease in peak early diastolic mitral annular velocity (E') compared to Wistar rats, suggesting structural remodeling and impaired diastolic function. Of the 84 genes in each the diabetes and FA metabolism arrays, 17 genes in the diabetes array and 41 genes in the FA metabolism array were significantly up-regulated in GK rats. Our data suggest that GK rats' exhibit increased genomic disposition to FA and TG metabolism independent of obesity.

Control of angiogenesis dictated by picomolar superoxide levels.

Control of vascular insufficiencies due to various cardiovascular pathologies is important for developing specific and effective treatments. Fluctuations in oxidative stress significantly alter the progression of angiogenesis under physiological and pathological conditions. However, the precise amount of reactive oxygen species (ROS) required to influence subsequent signaling pathways for ischemic angiogenesis remains undefined. Here, we have determined the effect of ROS-mediated molecular mechanisms on angiogenesis in a murine model of peripheral artery disease using Gclm mutant mice (a model of compromised glutathione synthesis and therefore reduced antioxidant capacity). Left femoral artery ligation and excision were performed in Gclm WT (+/+), heterozygous (+/-), and null (-/-) mice. Blood flow (laser Doppler), angiogenic index (CD31/DAPI), and proliferation index (Ki67/DAPI) were significantly increased in Gclm(+/-) mice but not in Gclm(+/+) or Gclm(-/-) mice. Measurements of reactive oxygen species suggest that the amount of superoxide required to stimulate angiogenesis after the induction of ischemia is 9.82 pmol/mg of tissue. Protein carbonyl levels increased in a manner consistent with increasing oxidative stress. Superoxide and protein carbonyl levels were reduced by the addition of the nitroxide tempol, a known superoxide dismutase mimetic. Finally, restoration of blood flow in Gclm(+/-) mice was attenuated by a VEGF164 aptamer, verifying that slightly elevated levels of ROS restore blood flow by stimulating endothelial cell proliferation through a VEGF-dependent pathway. The results of this study reveal new information on the amount of ROS necessary for angiogenic activity and provide the foundation of critical redox parameters for vascular remodeling responses. The information obtained from this study on vascular ischemia, using a model of decreased antioxidant capacity, has provided insight into the control of revascularization and is a step forward in our ability to regulate angiogenic therapies.

Heart failure with preserved ejection fraction: chronic low-intensity interval exercise training preserves myocardial O2 balance and diastolic function.

We have previously reported chronic low-intensity interval exercise training attenuates fibrosis, impaired cardiac mitochondrial function, and coronary vascular dysfunction in miniature swine with left ventricular (LV) hypertrophy (Emter CA, Baines CP. Am J Physiol Heart Circ Physiol 299: H1348-H1356, 2010; Emter CA, et al. Am J Physiol Heart Circ Physiol 301: H1687-H1694, 2011). The purpose of this study was to test two hypotheses: 1) chronic low-intensity interval training preserves normal myocardial oxygen supply/demand balance; and 2) training-dependent attenuation of LV fibrotic remodeling improves diastolic function in aortic-banded sedentary, exercise-trained (HF-TR), and control sedentary male Yucatan miniature swine displaying symptoms of heart failure with preserved ejection fraction. Pressure-volume loops, coronary blood flow, and two-dimensional speckle tracking ultrasound were utilized in vivo under conditions of increasing peripheral mean arterial pressure and ß-adrenergic stimulation 6 mo postsurgery to evaluate cardiac function. Normal diastolic function in HF-TR animals was characterized by prevention of increased time constant of isovolumic relaxation, normal LV untwisting rate, and enhanced apical circumferential and radial strain rate. Reduced fibrosis, normal matrix metalloproteinase-2 and tissue inhibitors of metalloproteinase-4 mRNA expression, and increased collagen III isoform mRNA levels (P < 0.05) accompanied improved diastolic function following chronic training. Exercise-dependent improvements in coronary blood flow for a given myocardial oxygen consumption (P < 0.05) and cardiac efficiency (stroke work to myocardial oxygen consumption, P < 0.05) were associated with preserved contractile reserve. LV hypertrophy in HF-TR animals was associated with increased activation of Akt and preservation of activated JNK/SAPK. In conclusion, chronic low-intensity interval exercise training attenuates diastolic impairment by promoting compliant extracellular matrix fibrotic components and preserving extracellular matrix regulatory mechanisms, preserves myocardial oxygen balance, and promotes a physiological molecular hypertrophic signaling phenotype in a large animal model resembling heart failure with preserved ejection fraction.

Neocortical vasculature abnormalities in the Fragile X mental retardation syndrome.

The Fragile X syndrome (FXS) is the leading form of inherited mental retardation. To date, the most prominent neuronal phenotype associated with the syndrome is an abundance of long thin spines exhibiting an immature morphology. However, in addition to synaptic abnormalities, recent case studies have demonstrated that Fragile X (FX) patients also exhibit abnormal cerebral blood flow (CBF). To examine the role of the Fragile X mental retardation protein (FMRP) in altering CBF, we examined blood vessel density (BVD) in the visual cortex of Adult and Middle-aged FX mice. Analysis of Middle-aged FX mice demonstrated elevated BVD compared to wildtype controls, suggesting that FX mice exhibit a lack of age-induced BVD plasticity. However, Adult FX and wildtype mice did not exhibit consistent differences in BVD. These data demonstrate that FMRP is required for age-induced neocortical vasculature plasticity. Furthermore, these data suggest a new role for FMRP in blood vessel regulation that would have profound implications towards appropriately timed delivery of neuronal nutrients, thus contributing to or exacerbating FX cognitive and neuronal abnormalities.

Nature versus nurture in frontotemporal lobar degeneration: the interaction of genetic background and education on brain damage.

BACKGROUND: Frontotemporal lobar degeneration (FTLD) is a progressive neurodegenerative disorder with a strong genetic background. It has been reported that modifiable factors, i.e. education (E), might act as proxies for reserve capacity. OBJECTIVE: To evaluate the impact of genetic background (positive family history, FH) on reserve mechanisms, by measuring regional cerebral blood flow (rCBF) correlates in FTLD patients. METHODS: 145 FTLD patients were recruited and underwent clinical, neuropsychological, behavioral assessment, and SPECT study. The main effect of E and FH on rCBF was evaluated. To test the potential interaction between the E and rCBF in FTLD patients with or without positive FH, a difference of slope analysis in the two groups was calculated. All the analyses were controlled for disease severity (Clinical Dementia Rating Scale, FTD-CDR). RESULTS: A main effect of education (E+ < E-) in frontal regions was reported, and high genetic loading (FH+ < FH-) was associated with a greater bilateral temporoparietal hypoperfusion. Evaluating the relationship between E and rCBF, a greater hypoperfusion of cingulate region in FH+ as compared to FH- was observed. DISCUSSION: Reserve mechanisms are available also in presence of an unfavorable genetic status. However, these compensatory mechanisms are modulated by the interaction with genetic factors.