Non-Invasive Technology That Improves Cardiac Function after Experimental Myocardial Infarction: Whole Body Periodic Acceleration (pGz)

Myocardial infarction (MI) may produce significant inflammatory changes and adverse ventricular remodeling leading to heart failure and premature death. Pharmacologic, stem cell transplantation, and exercise have not halted the inexorable rise in the prevalence and great economic costs of heart failure despite extensive investigations of such treatments. New therapeutic modalities are needed. Whole Body Periodic Acceleration (pGz) is a non-invasive technology that increases pulsatile shear stress to the endothelium thereby producing several beneficial cardiovascular effects as demonstrated in animal models, normal humans and patients with heart disease. pGz upregulates endothelial derived nitric oxide synthase (eNOS) and its phosphorylation (p-eNOS) to improve myocardial function in models of myocardial stunning and preconditioning. Here we test whether pGz applied chronically after focal myocardial infarction in rats improves functional outcomes from MI. Focal MI was produced by left coronary artery ligation. One day after ligation animals were randomized to receive daily treatments of pGz for four weeks (MI-pGz) or serve as controls (MI-CONT), with an additional group as non-infarction controls (Sham). Echocardiograms and invasive pressure volume loop analysis were carried out. Infarct transmurality, myocardial fibrosis, and markers of inflammatory and anti-inflammatory cytokines were determined along with protein analysis of eNOS, p-eNOS and inducible nitric oxide synthase (iNOS).At four weeks, survival was 80% in MI-pGz vs 50% in MI-CONT (p< 0.01). Ejection fraction and fractional shortening and invasive pressure volume relation indices of afterload and contractility were significantly better in MI-pGz. The latter where associated with decreased infarct transmurality and decreased fibrosis along with increased eNOS, p-eNOS. Additionally, MI-pGz had significantly lower levels of iNOS, inflammatory cytokines (IL-6, TNF-α), and higher level of anti-inflammatory cytokine (IL-10). pGz improved survival and contractile performance, associated with improved myocardial remodeling. pGz may serve as a simple, safe, non-invasive therapeutic modality to improve myocardial function after MI

Smooth Muscle Cells of Dystrophic (mdx) Mice Are More Susceptible to Hypoxia; The Protective Effect of Reducing Ca²⁺ Influx

Duchenne muscular dystrophy (DMD) is an inherited muscular disorder caused by mutations in the dystrophin gene. DMD patients have hypoxemic events due to sleep-disordered breathing. We reported an anomalous regulation of resting intracellular Ca2+ ([Ca2+]i) in vascular smooth muscle cells (VSMCs) from a mouse (mdx) model of DMD. We investigated the effect of hypoxia on [Ca2+]i in isolated and quiescent VSMCs from C57BL/10SnJ (WT) and C57BL/10ScSn-Dmd (mdx) male mice. [Ca2+]i was measured using Ca2+-selective microelectrodes under normoxic conditions (95% air, 5% CO2) and after hypoxia (glucose-free solution aerated with 95% N2-5% CO2 for 30 min). [Ca2+]i in mdx VSMCs was significantly elevated compared to WT under normoxia. Hypoxia-induced [Ca2+]i overload, which was significantly greater in mdx than in WT VSMCs. A low Ca2+ solution caused a reduction in [Ca2+]i and prevented [Ca2+]i overload secondary to hypoxia. Nifedipine (10 µM), a Ca2+ channel blocker, did not modify resting [Ca2+]i in VSMCs but partially prevented the hypoxia-induced elevation of [Ca2+]i in both genotypes. SAR7334 (1 µM), an antagonist of TRPC3 and TRPC6, reduced the basal and [Ca2+]i overload caused by hypoxia. Cell viability, assessed by tetrazolium salt (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, was significantly reduced in mdx compared to WT VSMCs. Pretreatment with SAR7341 increases cell viability in normoxic mdx (p 2+]i overload, which appears to be mediated by increased Ca2+ entry through L-type Ca2+ and TRPC channels

Molecular Modification of Transient Receptor Potential Canonical 6 Channels Modulates Calcium Dyshomeostasis in a Mouse Model Relevant to Malignant Hyperthermia

Background: Pharmacologic modulation has previously shown that transient receptor potential canonical (TRPC) channels play an important role in the pathogenesis of malignant hyperthermia. This study tested the hypothesis that genetically suppressing the function of TRPC6 can partially ameliorate muscle cation dyshomeostasis and the response to halothane in a mouse model relevant to malignant hyperthermia. Methods: This study examined the effect of overexpressing a muscle-specific nonconducting dominant-negative TRPC6 channel in 20 RYR1-p.R163C and 20 wild-type mice and an equal number of nonexpressing controls, using calcium- and sodium-selective microelectrodes and Western blots. Results: RYR1-p.R163C mouse muscles have chronically elevated intracellular calcium and sodium levels compared to wild-type muscles. Transgenic expression of the nonconducting TRPC6 channel reduced intracellular calcium from 331 ± 34 nM (mean ± SD) to 190 ± 27 nM (P < 0.0001) and sodium from 15 ± 1 mM to 11 ± 1 mM (P < 0.0001). Its expression lowered the increase in intracellular Ca2+ of the TRPC6-specific activator hyperforin in RYR1-p.R163C muscle fibers from 52% (348 ± 37 nM to 537 ± 70 nM) to 14% (185 ± 11 nM to 210 ± 44 nM). Western blot analysis of TRPC3 and TRPC6 expression showed the expected increase in TRPC6 caused by overexpression of its dominant-negative transgene and a compensatory increase in expression of TRPC3. Although expression of the muscle-specific dominant-negative TRPC6 was able to modulate the increase in intracellular calcium during halothane exposure and prolonged life (35 ± 5 min vs. 15 ± 3 min; P < 0.0001), a slow, steady increase in calcium began after 20 min of halothane exposure, which eventually led to death. Conclusions: These data support previous findings that TRPC channels play an important role in causing the intracellular calcium and sodium dyshomeostasis associated with RYR1 variants that are pathogenic for malignant hyperthermia. However, they also show that modulating TRPC channels alone is not sufficient to prevent the lethal effect of exposure to volatile anesthetic malignant hyperthermia–triggering agents

Whole body periodic acceleration improves survival and microvascular leak in a murine endotoxin model.

Sepsis is a life threatening condition which produces multi-organ dysfunction with profound circulatory and cellular derangements. Administration of E.Coli endotoxin (LPS) produces systemic inflammatory effects of sepsis including disruption of endothelial barrier, and if severe enough death. Whole body periodic acceleration (pGz) is the headward-footward motion of the body. pGz has been shown to induce pulsatile shear stress to the endothelium, thereby releasing vascular and cardio protective mediators. The purpose of this study was to determine whether or not pGz performed as a pre-treatment or post-treatment strategy improves survival in a lethal murine endotoxin model.This study was designed as a prospective randomized controlled study in mice. pGz was performed in mice as pre-treatment (pGz-LPS, 3 days prior to LPS), post-treatment (LPS- pGz, 30 min after LPS) strategies or Control (LPS-CONT), in a lethal murine model of endotoxemia. Endotoxemia was induced with intraperitoneal injection of E.Coli LPS (40mg/kg). In a separate group of mice, a nonspecific nitric oxide synthase inhibitor (L-NAME) was provided in their drinking water and pGz-LPS and LPS-pGz performed to determine the effect of nitric oxide (NO) inhibition on survival. In another subset of mice, micro vascular leakage was determined. Behavioral scoring around the clock was performed in all mice at 30 min intervals after LPS administration, until 48 hrs. survival or death. LPS induced 100% mortality in LPS-CONT animals by 30 hrs. In contrast, survival to 48 hrs. occurred in 60% of pGz-LPS and 80% of LPS-pGz. L-NAME abolished the survival effects of pGz. Microvascular leakage was markedly reduced in both pre and post pGz treated animals and was associated with increased tyrosine kinase endothelial-enriched tunica interna endothelial cell kinase 2 (TIE2) receptor and its phosphorylation (p-TIE2). In a murine model of lethal endotoxemia, pGz performed as a pre or post treatment strategy significantly improved survival, and markedly reduced microvascular leakage. The effect was modulated, in part, by NO since a non-selective inhibitor of NO abolished the pGz survival effect

Antioxidant Properties of Whole Body Periodic Acceleration (pGz)

The recognition that oxidative stress is a major component of several chronic diseases has engendered numerous trials of antioxidant therapies with minimal or no direct benefits. Nanomolar quantities of nitric oxide released into the circulation by pharmacologic stimulation of eNOS have antioxidant properties but physiologic stimulation as through increased pulsatile shear stress of the endothelium has not been assessed. The present study utilized a non-invasive technology, periodic acceleration (pGz) that increases pulsatile shear stress such that upregulation of cardiac eNOS occurs, We assessed its efficacy in normal mice and mouse models with high levels of oxidative stress, e.g. Diabetes type 1 and mdx (Duchene Muscular Dystrophy). pGz increased protein expression and upregulated eNOS in hearts. Application of pGz was associated with significantly increased expression of endogenous antioxidants (Glutathioneperoxidase-1(GPX-1), Catalase (CAT), Superoxide, Superoxide Dismutase 1(SOD1). This led to an increase of total cardiac antioxidant capacity along with an increase in the antioxidant response element transcription factor Nrf2 translocation to the nucleus. pGz decreased reactive oxygen species in both mice models of oxidative stress. Thus, pGz is a novel non-pharmacologic method to harness endogenous antioxidant capacity

Physicochemical properties of chitin isolated from shell of industrial crabs of various species

Differential thermal analysis was carried out over the range of –190 ÷ 400 °С on six samples of chitin from shells of industrial crabs of various species. All samples of the chitin have several relaxation transitions of endothermal character (g1-, g2-, β- and γ-)

Antioxidant Properties of Whole Body Periodic Acceleration (pGz).

The recognition that oxidative stress is a major component of several chronic diseases has engendered numerous trials of antioxidant therapies with minimal or no direct benefits. Nanomolar quantities of nitric oxide released into the circulation by pharmacologic stimulation of eNOS have antioxidant properties but physiologic stimulation as through increased pulsatile shear stress of the endothelium has not been assessed. The present study utilized a non-invasive technology, periodic acceleration (pGz) that increases pulsatile shear stress such that upregulation of cardiac eNOS occurs, We assessed its efficacy in normal mice and mouse models with high levels of oxidative stress, e.g. Diabetes type 1 and mdx (Duchene Muscular Dystrophy). pGz increased protein expression and upregulated eNOS in hearts. Application of pGz was associated with significantly increased expression of endogenous antioxidants (Glutathioneperoxidase-1(GPX-1), Catalase (CAT), Superoxide, Superoxide Dismutase 1(SOD1). This led to an increase of total cardiac antioxidant capacity along with an increase in the antioxidant response element transcription factor Nrf2 translocation to the nucleus. pGz decreased reactive oxygen species in both mice models of oxidative stress. Thus, pGz is a novel non-pharmacologic method to harness endogenous antioxidant capacity

In vivo upregulation of nitric oxide synthases in healthy rats

Periodic acceleration (pGz), sinusoidal motion of the whole body in a head–foot direction in the spinal axis, is a novel noninvasive means for cardiopulmonary support and induction of pulsatile shear stress. pGz increases plasma nitrite levels, in vivo and in vitro. Additionally, pGz confers cardioprotection in models of ischemia reperfusion injury. We hypothesize that pGz may also confer a cardiac phenotypic change by upregulation of the expression of the various NO synthase (NOS) isoforms in vivo. pGz was applied for 1 h to awake restrained male rats at 2 frequencies (360 and 600 cpm) and acceleration (Gz) of ±3.4 m/s 2. pGz did not affect arterial blood gases or electrolytes. pGz significantly increased total nitrosylated protein levels, indicating increased NO production. pGz also increased mRNA and protein levels of eNOS and nNOS, and phosphorylated eNOS in heart. pGz increased Akt phosphorylation (p-AKT), but not total Akt, or phosphorylated ERK1/2. Inducible (i) NOS levels were undetectable with or without pGz. Immunoblotting revealed the localization of nNOS, exclusively in cardiomyocyte, and pGz increased its expression. We have demonstrated that pGz changes myocardial NOS phenotypes. Such upregulation of eNOS and nNOS was still evident 24 h after pGz. Further studies are needed to understand the biochemical and biomechanical signal transduction pathway for the observed NOS phenotype changed induced by pGz

The Effects of pGz treatment on Diabetes Induced Oxidative Stress in Cardiomyocytes.

<p>The effects of pGz treatment for 14 days on ROS in cardiomyocytes in Control (CONT-pGz) and Diabetic mice (Diab-pGz) and their respective controls without pGz treatment in non diabetic (CONT) and diabetic (Diab). ROS was measured using the method of DCF fluorescence. Diabetes significantly increased ROS in cardiomyocytes (*p < 0.001 CONT vs. Diab). Treatment with pGz significantly reduced the diabetes induced increase in ROS (*p < 0.001 Diab vs. Diab-pGz and CONT-pGz vs Diab-pGz). Optical Units = O.U.</p