The Eye of the Storm: Investigating the Long-Term Cardiovascular Effects of COVID-19 and Variants.

COVID-19 had stormed through the world in early March of 2019, and on 5 May 2023, SARS-CoV-2 was officially declared to no longer be a global health emergency. The rise of new COVID-19 variants XBB.1.5 and XBB.1.16, a product of recombinant variants and sub-strains, has fueled a need for continued surveillance of the pandemic as they have been deemed increasingly infectious. Regardless of the severity of the variant, this has caused an increase in hospitalizations, a strain in resources, and a rise of concern for public health. In addition, there is a growing population of patients experiencing cardiovascular complications as a result of post-acute sequelae of COVID-19. This review aims to focus on what was known about SARS-CoV-2 and its past variants (Alpha, Delta, Omicron) and how the knowledge has grown today with new emerging variants, with an emphasis on cardiovascular complexities. We focus on the possible mechanisms that cause the observations of chronic cardiac conditions seen even after patients have recovered from the infection. Further understanding of these mechanisms will help to close the gap in knowledge on post-acute sequelae of COVID-19 and the differences between the effects of variants.

Primary cardiac fibroblast cell culture: methodological considerations for physiologically relevant conditions.

Primary cardiac fibroblast (CF) tissue culture is a necessary tool for interrogating specific signaling mechanisms that dictate the phenotypic heterogeneity observed in vivo in different disease states. Traditional approaches that use tissue culture plastic and nutrient-rich medium have been shown to induce CF activation and, therefore, alter CF subpopulation composition. This shift away from in vivo phenotypes complicate the interpretation of results through the lens of the animal model. As the field works to identify CF diversity, these methodological flaws have begun to be addressed and more studies are focused on the dynamic interaction of CFs with their environment. This review focuses on the aspects of tissue culture that impact CF activation and, therefore, require consideration when designing in vitro experiments. The complexity of CF biology overlaid onto diverse model systems highlight the need for study-specific optimization and validation.

Dopamine receptor 3: A mystery at the heart of cardiac fibrosis.

Dopamine receptors have been extensively studied in the mammalian brain and spinal cord, as dopamine is a vital determinant of bodily movement, cognition, and overall behavior. Thus, dopamine receptor antagonist antipsychotic drugs are commonly used to treat multiple psychiatric disorders. Although less discussed, these receptors are also expressed in other peripheral organ systems, such as the kidneys, eyes, gastrointestinal tract, and cardiac tissue. Consequently, therapies for certain psychiatric disorders which target dopamine receptors could have unidentified consequences on certain functions of these peripheral tissues. The existence of an intrinsic dopaminergic system in the human heart remains controversial and debated within the literature. Therefore, this review focuses on literature related to dopamine receptors within cardiac tissue, specifically dopamine receptor 3 (D3R), and summarizes the current state of knowledge while highlighting areas of research which may be lacking. Additionally, recent findings regarding crosstalk between D3R and dopamine receptor 1 (D1R) are examined. This review discusses the novel concept of understanding the role of the loss of function of D3R may play in collagen accumulation and cardiac fibrosis, eventually leading to heart failure.

CVD and COVID-19: Emerging Roles of Cardiac Fibroblasts and Myofibroblasts.

Cardiovascular disease (CVD) is the leading cause of death worldwide. Current data suggest that patients with cardiovascular diseases experience more serious complications with coronavirus disease-19 (COVID-19) than those without CVD. In addition, severe COVID-19 appears to cause acute cardiac injury, as well as long-term adverse remodeling of heart tissue. Cardiac fibroblasts and myofibroblasts, being crucial in response to injury, may play a pivotal role in both contributing to and healing COVID-19-induced cardiac injury. The role of cardiac myofibroblasts in cardiac fibrosis has been well-established in the literature for decades. However, with the emergence of the novel coronavirus SARS-CoV-2, new cardiac complications are arising. Bursts of inflammatory cytokines and upregulation of TGF-ß1 and angiotensin (AngII) are common in severe COVID-19 patients. Cytokines, TGF-ß1, and Ang II can induce cardiac fibroblast differentiation, potentially leading to fibrosis. This review details the key information concerning the role of cardiac myofibroblasts in CVD and COVID-19 complications. Additionally, new factors including controlling ACE2 expression and microRNA regulation are explored as promising treatments for both COVID-19 and CVD. Further understanding of this topic may provide insight into the long-term cardiac manifestations of the COVID-19 pandemic and ways to mitigate its negative effects.

Low Intrinsic Aerobic Capacity Limits Recovery Response to Hindlimb Ischemia.

Introduction: In this study, we determined the influence of intrinsic exercise capacity on the vascular adaptive responses to hind limb ischemia. High Capacity Running, HCR; Low Capacity Running, LCR, rats were used to assess intrinsic aerobic capacity effects on adaptive responses to ischemia. Methods: Muscle samples from both ischemic and non-ischemic limb in both strains were compared, histologically for the muscle-capillary relationship, and functionally using microspheres to track blood flow and muscle stimulation to test fatigability. PCR was used to identify the differences in gene expression between the phenotypes following occlusive ischemia. Results: Prior to ligation, there were not significant differences between the phenotypes in the exhaustion time with high frequency pacing. Following ligation, LCR decreased significantly in the exhaustion time compare with HCRs (437 ± 47 vs. 824 ± 56, p < 0.001). The immediate decrease in flow was significantly more severe in LCRs than HCRs (52.5 vs. 37.8%, p < 0.001). VEGF, eNOS, and ANG2 (but not ANG1) gene expression were decreased in LCRs vs. HCRs before occlusion, and increased significantly in LCRs 14D after occlusion, but not in HCRs. LCR capillary density (CD) was significantly lower at all time points after occlusion (LCR 7D = 564.76 ± 40.5, LCR 14D = 507.48 ± 54.2, both p < 0.05 vs. HCR for respective time point). NCAF increased significantly in HCR and LCR in response to ischemia. Summary: These results suggest that LCR confers increased risk for ischemic injury and is subject to delayed and less effective adaptive response to ischemic stress.

Influence of Intrinsic Aerobic Exercise Capacity and Sex on Cardiac Injury Following Acute Myocardial Ischemia and Reperfusion.

Purpose: Previous reports have suggested that active exercise aside, intrinsic aerobic running capacity (Low = LCR, high = HCR) in otherwise sedentary animals may influence several cardiovascular health-related indicators. Relative to the HCR phenotype, the LCR phenotype is characterized by decreased endothelial reactivity, increased susceptibility to reperfusion-induced arrhythmias following short, non-infarction ischemia, and increased diet-induced insulin resistance. More broadly, the LCR phenotype has come to be characterized as a "disease prone" model, with the HCRs as "disease resistant." Whether these effects extend to injury outcomes in an overt infarction or whether the effects are gender specific is not known. This study was designed to determine whether HCR/LCR phenotypic differences would be evident in injury responses to acute myocardial ischemia-reperfusion injury (AIR), measured as infarct size and to determine whether sex differences in infarction size were preserved with phenotypic selection. Methods: Regional myocardial AIR was induced in vivo by either 15 or 30 min ligation of the left anterior descending coronary artery, followed by 2 h of reperfusion. Global ischemia was induced in isolated hearts ex vivo using a Langendorff perfusion system and cessation of perfusion for either 15 or 30 min followed by 2 h of reperfusion. Infarct size was determined using 2, 3, 5-triphenyltetrazolium chloride (TTC) staining, and normalized to area at risk in the regional model, or whole heart in the global model. Portions of the tissue were paraffin embedded for H&E staining and histology analysis. Results: Phenotype dependent differences in infarct size were seen with 15 min occlusion/2 h reperfusion (LCR > HCR, p < 0.05) in both regional and global models. In both models, longer occlusion times (30 min/2 h) produced significantly larger infarctions in both phenotypes, but phenotypic differences were no longer present (LCR vs. HCR, p = n.s.). Sex differences in infarct size were present in each phenotype (LCR male > LCR female, p < 0.05; HCR male > HCR female, p < 0.05 regardless of length of occlusion, or ischemia model. Conclusions: There is cardioprotection afforded by high intrinsic aerobic capacity, but it is not infinite/continuous, and may be overcome with sufficient injury burden. Phenotypic selection based on endurance running capacity preserved sex differences in response to both short and longer term coronary occlusive challenges. Outcomes could not be associated with differences in system characteristics such as circulating inflammatory mediators or autonomic nervous system influences, as similar phenotypic injury patterns were seen in vivo, and in isolated crystalloid perfused heart ex vivo.

Augmented Cardiac Mitochondrial Capacity in High Capacity Aerobic Running "Disease-Resistant" Phenotype at Rest Is Lost Following Ischemia Reperfusion.

Rationale: Regular active exercise is considered therapeutic for cardiovascular disease, in part by increasing mitochondrial respiratory capacity, but a significant amount of exercise capacity is determined genetically. Animal models, demonstrating either high capacity aerobic running (HCR) or low capacity aerobic running (LCR) phenotypes, have been developed to study the intrinsic contribution, with HCR rats subsequently characterized as "disease resistant" and the LCRs as "disease prone." Enhanced cardioprotection in HCRs has been variable and mutifactoral, but likely includes a metabolic component. These studies were conducted to determine the influence of intrinsic aerobic phenotype on cardiac mitochondrial function before and after ischemia and reperfusion. Methods: A total of 34 HCR and LCR rats were obtained from the parent colony at the University of Toledo, housed under sedentary conditions, and fed normal chow. LCR and HCR animals were randomly assigned to either control or ischemia-reperfusion (IR). On each study day, one HCR/LCR pair was anesthetized, and hearts were rapidly excised. In IR animals, the hearts were immediately flushed with iced hyperkalemic, hyperosmotic, cardioplegia solution, and subjected to global hypothermic ischemic arrest (80 min). Following the arrest, the hearts underwent warm reperfusion (120 min) using a Langendorff perfusion system. Following reperfusion, the heart was weighed and the left ventricle (LV) was isolated. A midventricular ring was obtained to estimate infarction size [triphenyltetrazolium chloride (TTC)] and part of the remaining tissue (~150 mg) was transferred to a homogenation buffer on ice. Isolated mitochondria (MITO) samples were prepared and used to determine respiratory capacity under different metabolic conditions. In control animals, MITO were obtained and prepared similarly immediately following anesthesia and heart removal, but without IR. Results: In the control rats, both resting and maximally stimulated respiratory rates were higher (32 and 40%, respectively; p < 0.05) in HCR mitochondria compared to LCR. After IR, resting MITO respiratory rates were decreased to about 10% of control in both strains, and the augmented capacity in HCRs was absent. Maximally stimulated rates also were decreased more than 50% from control and were no longer different between phenotypes. Ca++ retention capacity and infarct size were not significantly different between HCR and LCR (49.2 ± 5.6 vs. 53.7 ± 4.9%), nor was average coronary flow during reperfusion or arrhythmogenesis. There was a significant loss of mitochondria following IR, which was coupled with decreased function in the remaining mitochondria in both strains. Conclusion: Cardiac mitochondrial capacity from HCR was significantly higher than LCR in the controls under each condition. After IR insult, the cardiac mitochondrial respiratory rates were similar between phenotypes, as was Ca++ retention capacity, infarct size, and arrhythmogenicity, despite the increased mitochondrial capacity in the HCRs before ischemia. Relatively, the loss of respiratory capacity was actually greater in HCR than LCR. These data could suggest limits in the extent to which the HCR phenotype might be "protective" against acute tissue stressors. The extent to which any of these deficits could be "rescued" by adding an active exercise component to the intrinsic phenotype is unknown.

Loss of Function in Dopamine D3 Receptor Attenuates Left Ventricular Cardiac Fibroblast Migration and Proliferation in vitro.

Evidence suggests the existence of an intracardiac dopaminergic system that plays a pivotal role in regulating cardiac function and fibrosis through G-protein coupled receptors, particularly mediated by dopamine receptor 3 (D3R). However, the expression of dopamine receptors in cardiac tissue and their role in cardiac fibroblast function is unclear. In this brief report, first we determined expression of D1R and D3R both in left ventricle (LV) tissue and fibroblasts. Then, we explored the role of D3R in the proliferation and migration of fibroblast cell cultures using both genetic and pharmaceutical approaches; specifically, we compared cardiac fibroblasts isolated from LV of wild type (WT) and D3R knockout (D3KO) mice in response to D3R-specific pharmacological agents. Finally, we determined if loss of D3R function could significantly alter LV fibroblast expression of collagen types I (Col1a1) and III (Col3a1). Cardiac fibroblast proliferation was attenuated in D3KO cells, mimicking the behavior of WT cardiac fibroblasts treated with D3R antagonist. In response to scratch injury, WT cardiac fibroblasts treated with the D3R agonist, pramipexole, displayed enhanced migration compared to control WT and D3KO cells. Loss of function in D3R resulted in attenuation of both proliferation and migration in response to scratch injury, and significantly increased the expression of Col3a1 in LV fibroblasts. These findings suggest that D3R may mediate cardiac fibroblast function during the wound healing response. To our knowledge this is the first report of D3R's expression and functional significance directly in mouse cardiac fibroblasts.

Pro-remodeling peptides modulate collagen α1(I) promoter activity in rat cardiac myofibroblasts.

Previous studies have extensively demonstrated the effect of endothelin-1 (ET-1), angiotensin II (Ang II), and TGF-ß1 on the stimulation of collagen type I expression in cardiac myofibroblasts. However, the role of pro-remodeling peptides in the transcriptional regulation of the collagen promoter remains unclear. Thus, the purpose of this study was to investigate the net regulatory effects of pro-remodeling peptides on collagen type I promoter activity. Constructs of various lengths (300 bp, 1.1 kbp, 1.7 kbp, 2.3 kbp and 3.5 kbp) of the rat collagen α1(I) promoter were transfected into cardiac myofibroblasts in vitro and promoter activity was measured using chloramphenicol acetyl transferase (CAT) assays. Reduced promoter activity occurred across all treatments in myofibroblasts transfected with the 1.7 kbp construct. ET-1 was unable to increase promoter activity with constructs 300, 1.1, and 1.7 kbp, but induced promoter activity in cells with the 2.3 kbp construct. Additionally, while a combination of pro-remodeling peptides induced promoter activity across constructs, the resultant increase in the 2.3 and 3.5 kbp constructs were comparable to that observed from ET-1 treatment alone. Lastly, cells transfected with the entire promoter sequence had the lowest promoter activity. This data suggests that the collagen promoter is tightly regulated and that pro-remodeling factors produce an overall net effect on collagen expression, rather than additive.

What is the role of peptide fragments of collagen I and IV in health and disease?

Collagen is the most abundant protein in mammalian systems; it can be found in organs such as bones, the liver, kidney, heart, teeth, and skin. Collagen provides the necessary structural framework for tissues in which it is found. However, if there are any alterations in the delicate balance of collagen types in the extracellular matrix (ECM), then problems arise. For example, increasing collagen I:III ratio would provide additional rigidity to tissue structure, whereas decreasing this ratio would provide elasticity and flexibility to the tissue. The proper function of tissues is reliant on this scale not tipping too far in either direction. Major players in the process of ECM remodeling, both normal and adverse, are the fibroblast cells via the secretion of collagen precursors and matrix metalloproteinases, with the latter responsible for ECM degradation. The collagen peptides created by the proteolytic cleavage of these collagen fibrils, while once thought to have an absence of function, have been shown over recent years to potentiate and regulate a variety of cellular processes acting through integrin receptors. Many collagen peptides have been identified from many different collagen types and have been shown to regulate processes such as cell proliferation, migration, apoptosis, and reduce angiogenesis. The collagen peptides of interest are those generated from the primary collagen type of tissue interstitial matrix, collagen type I, and the basement membrane, collagen type IV. Thus, this review looks to highlight some examples of unorthodox functional roles of collagen and its peptides in regulating physiological health and disease.

Are There Limitations to Exercise Benefits in Peripheral Arterial Disease?

Substantial evidence exists indicating that inactivity contributes to the progression of chronic disease, and conversely, that regular physical activity can both prevent the onset of disease as well as delay the progression of existing disease. To that end "exercise as medicine" has been advocated in the broad context as general medical care, but also in the specific context as a therapeutic, to be considered in much the same way as other drugs. As there are non-responders to many medications, there also are non-responders to exercise; individual who participate but do not demonstrate appreciable improvement/benefit. In some settings, the stress induced by exercise may aggravate an underlying condition, rather than attenuate chronic disease. As personalized medicine evolves with ready access to genetic information, so too will the incorporation of exercise in the context of those individual genetics. The focus of this brief review is to distinguish between the inherent capacity to perform, as compared to adaptive response to active exercise training in relation to cardiovascular health and peripheral arterial disease.

Dietary bitter melon seed increases peroxisome proliferator-activated receptor-γ gene expression in adipose tissue, down-regulates the nuclear factor-κB expression, and alleviates the symptoms associated with metabolic syndrome.

The objective of this study was to examine the extent to which bitter melon seed (BMS) alleviates the symptoms associated with metabolic syndrome and elucidate the mechanism by which BMS exerts beneficial effects. Three-month-old female Zucker rats were assigned to following groups: lean control (L-Ctrl), obese control (O-Ctrl), and obese + BMS (O-BMS). The control groups were fed AIN-93M purified rodent diet, and the O-BMS group was fed AIN-93M diet modified to contain 3.0% (wt/wt) ground BMS for 100 days. After 100 days of treatment, BMS supplementation in the obese rats lowered the total serum cholesterol by 38% and low-density lipoprotein-cholesterol levels by about 52% and increased the ratio of serum high-density lipoprotein-cholesterol to total cholesterol compared to the O-Ctrl group. The percentage of total liver lipids was about 32% lower and serum triglyceride levels were 71% higher in the O-BMS group compared to the O-Ctrl group. Serum glucose levels were significantly lowered partly because of the increase in the serum insulin levels in the BMS-based diet groups. BMS supplementation increased the expression of peroxisome proliferator-activated receptor-γ (PPAR-γ) in the white adipose tissue of the obese rats significantly (P < .05) and down-regulated the expression of PPAR-γ, nuclear factor-κB (NF-κB), and interferon-γ mRNA in heart tissue of the obese rats. The findings of this study suggest that BMS improves the serum and liver lipid profiles and serum glucose levels by modulating PPAR-γ gene expression. To our knowledge, this study for the first time shows that BMS exerts cardioprotective effects by down-regulating the NF-κB inflammatory pathway.

Urotensin II alters vascular reactivity in animals subjected to volume overload.

Congestive heart failure (CHF) alters vascular reactivity and up regulates in urotensin II (UTII), a potent vasoactive peptide. The aim of this study was to investigate the interaction between CHF and UTII in altering vascular reactivity in a rat model of volume overload heart failure. Animals were divided into 4 groups: control, UTII infused (UTII), volume overload only (VO) or volume overload+UTII (VO+UTII). Volume overload was established by the formation of an aortocaval fistula. Following fistula formation animals were administered UTII at a rate of 300 pmol/kg/h for 4 weeks subcutaneously with mini-osmotic pumps. Thoracic aorta rings, with/without endothelium, were subjected to cumulative dose-responses to phenylephrine, sodium nitroprusside (SNP), acetylcholine (ACH), UTII, and the Rho-kinase inhibitor HA-1077. Aortas from VO animals exhibited increased sensitivity to phenylephrine and UTII with a decreased relaxation response to ACH and HA-1077. Aortas from animals subjected to chronic UTII with volume overload (VO + UTII) retained their sensitivity to phenylephrine and UTII while they improved their relaxation to HA-1077 but not ACH. The constrictive response to UTII was dose-dependent and augmented at concentrations <0.01 µM in VO animals. The changes in vascular reactivity paralleled an elevation of both the UTII and α(1A)-adrenergic receptor while the Rho and Rho-kinase signalling proteins were diminished. We found that volume overload increased sensitivity to the vasoconstrictor agents that was inversely related to changes in the Rho-kinase expression. The addition of UTII with VO reversed the constrictive vascular response through alterations in the Rho-kinase signalling pathway.

Role of protein kinase C-delta in angiotensin II induced cardiac fibrosis.

Previous studies have demonstrated a role for angiotensin II (AngII) and myofibroblasts (myoFb) in cardiac fibrosis. However, the role of PKC-delta in AngII mediated cardiac fibrosis is unclear. Therefore, the present study was designed to investigate the role of PKC-delta in AngII induced cardiac collagen expression and fibrosis. AngII treatment significantly (p<0.05) increased myoFb collagen expression, whereas PKC-delta siRNA treatment or rottlerin, a PKC-delta inhibitor abrogated (p<0.05) AngII induced collagen expression. MyoFb transfected with PKC-delta over expression vector showed significant increase (p<0.05) in the collagen expression as compared to control. Two weeks of chronic AngII infused rats showed significant (p<0.05) increase in collagen expression compared to sham operated rats. This increase in cardiac collagen expression was abrogated by rottlerin treatment. In conclusion, both in vitro and in vivo data strongly suggest a role for PKC-delta in AngII induced cardiac fibrosis.

Visfatin in pregnancy: proposed mechanism of peptide delivery.

Visfatin is a newly identified 52 kD adipocytokine that appears to have insulinomimetic properties. We examined visfatin expression in visceral fat from lean and pregnant women. Visfatin gene expression was seven times higher in omental fat of pregnant women than in lean women. Both immunohistochemistry and immunoblot confirmed that visfatin protein was much higher in pregnant women than in nonpregnant women. However, serum visfatin was 20.8 +/- 7.7 ng/ml (n = 7) in lean women as compared to only a slight increase to 40.3 ng/ml in pregnant women (n = 4). We measured visfatin mRNA content of human placenta and found that placenta expresses high levels of visfatin mRNA and protein. At a concentration of 2 nM, visfatin and insulin produced nearly identical increase in glucose transport. The discrepancy between the elevated visfatin expression and tissue visfatin compared to only a small increase in serum visfatin is a matter of controversy. The data on serum visfatin concentrations are replete with contradictory data. Taken together, we suggest that visfatin is not a hormone. Instead, we propose that visfatin acts in either a paracrine or autocrine mode. This hypothesis would explain what various laboratories have found widely discrepant values for serum visfatin. Since visfatin potently and efficaciously induced glucose transport in a cell culture model, any hypothetical role for visfatin in pregnancy should include the possibility that it may play a role in maternal/fetal glucose metabolism or distribution and that it may do so by acting locally.

PPAR-gamma expression in animals subjected to volume overload and chronic Urotensin II administration.

Activation of PPAR-gamma through the administration of glitazones has shown promise in preserving function following cardiac injury, although recent evidence has suggested their use may be contraindicated in the case of severe heart failure. This study tested the hypothesis that PPAR-gamma expression increases in a time dependent manner in response to chronic volume overload (VO) induced heart failure. Additionally, we attempted to determine what effect 4 week administration of Urotensin II (UTII) may have on PPAR-gamma expression. VO induced heart failure was produced in Sprague-Dawley rats (n=32) by aorta-caval fistula. Animals were sacrificed at 1, 4, and 14 weeks following shunt creation. In a separate set of experiments, animals were administered 300 pmol/kg/h of UTII for 4 weeks, subjected to 4 weeks of volume overload, or given UTII+VO. Densitometric analysis of left ventricular (LV) protein demonstrated PPAR-gamma expression was significantly ((*)p<0.05) upregulated at 4 and 14 weeks (31.5% and 37%, respectively) post-fistula formation compared to control values. PPAR-gamma activation was decreased in the 4 and 14 week (39.16% and 42.4%, respectively), but not in the 1-week animals, and these changes did not correlate with NF-kappaB activity. Animals given UTII either with or without VO demonstrated increased expression of PPAR-gamma as did animals subjected to 4 week VO alone. Animals given UTII either with or without VO had decreased activity vs. control. These data suggest PPAR-gamma may play a role in the progression of heart failure, however, the exact nature has yet to be determined.

Hemodynamic effects of chronic urotensin II administration in animals with and without aorto-caval fistula.

Urotensin II (UTII) is a potent vasoactive peptide. Recent studies have demonstrated increased expression of both UTII and its receptor (UTR) expression in end-stage congestive heart failure (CHF), but it is unclear whether UTII and UTR are late stage markers of decompensation, or earlier adaptive responses. The purpose of this study was to measure the effects of chronic UTII administration in normal and volume overloaded animals. Chronic 4 weeks administration of UTII produced decreases in hemodynamic function in animals not subjected to volume overload while returning function to control levels in animals with overload. Expression levels of calcium regulatory proteins phospholamban (PLN), sarcoplasmic reticulum Ca(2+) ATPase (SERCA2), and Na(+)/Ca(2+) exchanger (NCX) were measured to determine if administration of UTII resulted in aberrant Ca(2+) handling. Changes in protein expression revealed that UTII influenced Ca(2+) handling proteins in normal animals although these changes are not seen in the volume overload.

PPAR-gamma agonists induce the expression of VEGF and its receptors in cultured cardiac myofibroblasts.

OBJECTIVES: Myofibroblasts (myoFb) are the major cell types that appear at the site of myocardial infarction (MI) in response to injury and play a vital role in tissue repair/remodeling. Since vascular endothelial growth factor (VEGF) plays a crucial role in the infarcted/ischemic heart, we hypothesized that activation of the peroxisome proliferator-activated receptor (PPAR)-gamma by its agonists induces VEGF expression while simultaneously decreasing inflammation (NF-kappaB). Such an increase in myoFb VEGF expression by PPAR-gamma agonists may play a role in angiogenesis. METHODS: Rat myoFb were treated with PPAR-gamma agonists and VEGF expression was measured by ELISA. The effect of these agonists on VEGF receptors was determined by qRT-PCR and flow-cytometric analysis. VEGF produced by these cells was also used for analysis of in vitro tubule formation (Matrigel assay). RESULTS: The PPAR-gamma activators troglitazone (TZ) and 15-deoxy-prostaglandin J2 (15J2) induced the expression of VEGF and its receptors (Flt-1 and KDR) in myoFb. TZ and 15J2 elicited a significant increase in the expression of KDR (14.7+/-1.0% and 9.6+/-2.1% respectively) and Flt-1 (24.5+/-2.0%, and 14.0+/-2.2% respectively) when compared to untreated myoFb. MyoFb treated with PPAR-gamma agonists increased extracellular VEGF, augmenting tubule formation on a Matrigel. The PPAR-gamma activator 15J2 significantly decreased the NF-kappaB activity in myoFb. CONCLUSION: This study demonstrates the induction of the VEGF accompanied by a reduction of NF-kappaB activity (inflammatory signaling) by PPAR-gamma agonists in cardiac myoFb. These results may further the understanding of the beneficial effects of PPAR-gamma agonists on infarcted tissue repair and angiogenesis.

RNAi pathway is functional in peripheral nerve axons.

Recent observations demonstrated that translation of mRNAs may occur in axonal processes at sites that are long distances away from the neuronal perikaria. While axonal protein synthesis has been documented in several studies, the mechanism of its regulation remains unclear. The aim of this study was to investigate whether RNA interference (RNAi) may be one of the pathways that control local protein synthesis in axons. Here we show that sciatic nerve contains Argonaute2 nuclease, fragile X mental retardation protein, p100 nuclease, and Gemin3 helicase-components of the RNA-induced silencing complex (RISC). Application of short-interfering RNAs against neuronal beta-tubulin to the sciatic nerve initiated RISC formation, causing a decrease in levels of neuronal beta-tubulin III mRNA and corresponding protein, as well as a significant reduction in retrograde labeling of lumbar motor neurons. Our observations indicate that RNAi is functional in peripheral mammalian axons and is independent from the neuronal cell body or Schwann cells. We introduce a concept of local regulation of axonal translation via RNAi.

Effects of targeted deletion of A1 adenosine receptors on postischemic cardiac function and expression of adenosine receptor subtypes.

To examine ischemic tolerance in the absence of A(1) adenosine receptors (A(1)ARs), isolated wild-type (WT) and A(1)AR knockout (A(1)KO) murine hearts underwent global ischemia-reperfusion, and injury was measured in terms of functional recovery and efflux of lactate dehydrogenase (LDH). Hearts were analyzed by real-time RT-PCR both at baseline and at intervals during ischemia-reperfusion to determine whether compensatory expression of other adenosine receptor subtypes occurs with either A(1)AR deletion and/or ischemia-reperfusion. A(1)KO hearts had higher baseline coronary flow (CF) and left ventricular developed pressure (LVDP) than WT hearts, whereas heart rate was unchanged by A(1)AR deletion. After 20 min of ischemia, CF was attenuated in A(1)KO compared with WT hearts, and this reduction persisted throughout reperfusion. Final recovery of LVDP was decreased in A(1)KO hearts (54.4 +/- 5.1 vs. WT 81.1 +/- 3.4% preischemic baseline) and correlated with higher diastolic pressure during reperfusion. Postischemic efflux of LDH was greater in A(1)KO compared with WT hearts. Real-time RT-PCR demonstrated the absence of A(1)AR transcript in A(1)KO hearts, and the message for A(2A), A(2B), and A(3) adenosine receptors was similar in uninstrumented A(1)KO and WT hearts. Ischemia-reperfusion increased A(2B) mRNA expression 2.5-fold in both WT and A(1)KO hearts without changing A(1) or A(3) expression. In WT hearts, ischemia transiently doubled A(2A) mRNA, which returned to preischemic level upon reperfusion, a pattern not observed in A(1)KO hearts. Together, these data affirm the cardioprotective role of A(1)ARs and suggest that induced expression of other adenosine receptor subtypes may participate in the response to ischemia-reperfusion in isolated murine hearts.