Differences in cardiac adaptation to exercise in male and female athletes assessed by noninvasive techniques: a state-of-the-art review.

Athlete's heart is generally regarded as a physiological adaptation to regular training, with specific morphological and functional alterations in the cardiovascular system. Development of the noninvasive imaging techniques over the past several years enabled better assessment of cardiac remodeling in athletes, which may eventually mimic certain pathological conditions with the potential for sudden cardiac death, or disease progression. The current literature provides a compelling overview of the available methods that target the interrelation of prolonged exercise with cardiac structure and function. However, this data stems from scientific studies that included mostly male athletes. Despite the growing participation of females in competitive sport meetings, little is known about the long-term cardiac effects of repetitive training in this population. There are several factors-biochemical, physiological and psychological, that determine sex-dependent cardiac response. Herein, the aim of this review was to compare cardiac adaptation to endurance exercise in male and female athletes with the use of electrocardiographic, echocardiographic, and biochemical examination, to determine the sex-specific phenotypes, and to improve the healthcare providers' awareness of cardiac remodeling in athletes. Finally, we discuss the possible exercise-induced alternations that should arouse suspicion of pathology and be further evaluated.

The Impact of LY487379 or CDPPB on eNOS Expression in the Mouse Brain and the Effect of Joint Administration of Compounds with NO• Releasers on MK-801- or Scopolamine-Driven Cognitive Dysfunction in Mice.

The role of endothelial nitric oxide synthase (eNOS) in the regulation of a variety of biological processes is well established, and its dysfunction contributes to brain pathologies, including schizophrenia or Alzheimer's disease (AD). Positive allosteric modulators (PAMs) of metabotropic glutamate (mGlu) receptors were shown to be effective procognitive compounds, but little is known about their impact on eNOS expression and stability. Here, we investigated the influence of the acute and chronic administration of LY487379 or CDPPB (mGlu2 and mGlu5 PAMs), on eNOS expression in the mouse brain and the effect of the joint administration of the ligands with nitric oxide (NO) releasers, spermineNONOate or DETANONOate, in different combinations of doses, on MK-801- or scopolamine-induced amnesia in the novel object recognition (NOR) test. Our results indicate that both compounds provoked eNOS monomer formation, and CDPPB at a dose of 5 mg/kg exaggerated the effect of MK-801 or scopolamine. The coadministration of spermineNONOate or DETANONOate enhanced the antiamnesic effect of CDPPB or LY487379. The best activity was observed for ineffective or moderate dose combinations. The results indicate that treatment with mGluR2 and mGluR5 PAMs may be burdened with the risk of promoting eNOS uncoupling through the induction of dimer dissociation. Administration of the lowest possible doses of the compounds with NO• donors, which themselves have procognitive efficacy, may be proposed for the treatment of schizophrenia or AD.

Molecular Imaging and Nanotechnology-Emerging Tools in Diagnostics and Therapy.

Personalized medicine is emerging as a new goal in the diagnosis and treatment of diseases. This approach aims to establish differences between patients suffering from the same disease, which allows to choose the most effective treatment. Molecular imaging (MI) enables advanced insight into molecule interactions and disease pathology, improving the process of diagnosis and therapy and, for that reason, plays a crucial role in personalized medicine. Nanoparticles are widely used in MI techniques due to their size, high surface area to volume ratio, and multifunctional properties. After conjugation to specific ligands and drugs, nanoparticles can transport therapeutic compounds directly to their area of action and therefore may be used in theranostics-the simultaneous implementation of treatment and diagnostics. This review summarizes different MI techniques, including optical imaging, ultrasound imaging, magnetic resonance imaging, nuclear imaging, and computed tomography imaging with theranostics nanoparticles. Furthermore, it explores the potential use of constructs that enables multimodal imaging and track diseases in real time.

Nitric Oxide-Dependent Mechanisms Underlying MK-801- or Scopolamine-Induced Memory Dysfunction in Animals: Mechanistic Studies.

MK-801, an NMDA receptor antagonist, and scopolamine, a cholinergic receptor blocker, are widely used as tool compounds to induce learning and memory deficits in animal models to study schizophrenia or Alzheimer-type dementia (AD), respectively. Memory impairments are observed after either acute or chronic administration of either compound. The present experiments were performed to study the nitric oxide (NO)-related mechanisms underlying memory dysfunction induced by acute or chronic (14 days) administration of MK-801 (0.3 mg/kg, i.p.) or scopolamine (1 mg/kg, i.p.). The levels of L-arginine and its derivatives, L-citrulline, L-glutamate, L-glutamine and L-ornithine, were measured. The expression of constitutive nitric oxide synthases (cNOS), dimethylaminohydrolase (DDAH1) and protein arginine N-methyltransferases (PMRTs) 1 and 5 was evaluated, and the impact of the studied tool compounds on cGMP production and NMDA receptors was measured. The studies were performed in both the cortex and hippocampus of mice. S-nitrosylation of selected proteins, such as GLT-1, APP and tau, was also investigated. Our results indicate that the availability of L-arginine decreased after chronic administration of MK-801 or scopolamine, as both the amino acid itself as well as its level in proportion to its derivatives (SDMA and NMMA) were decreased. Additionally, among all three methylamines, SDMA was the most abundant in the brain (~70%). Administration of either compound impaired eNOS-derived NO production, increasing the monomer levels, and had no significant impact on nNOS. Both compounds elevated DDAH1 expression, and slight decreases in PMRT1 and PMRT5 in the cortex after scopolamine (acute) and MK-801 (chronic) administration were observed in the PFC, respectively. Administration of MK-801 induced a decrease in the cGMP level in the hippocampus, accompanied by decreased NMDA expression, while increased cGMP production and decreased NMDA receptor expression were observed after scopolamine administration. Chronic MK-801 and scopolamine administration affected S-nitrosylation of GLT-1 transport protein. Our results indicate that the analyzed tool compounds used in pharmacological models of schizophrenia or AD induce changes in NO-related pathways in the brain structures involved in cognition. To some extent, the changes resemble those observed in human samples.

Endothelial Dysfunction Driven by Hypoxia-The Influence of Oxygen Deficiency on NO Bioavailability.

Cardiovascular diseases (CVDs) are the leading cause of death worldwide. The initial stage of CVDs is characterized by endothelial dysfunction, defined as the limited bioavailability of nitric oxide (NO). Thus, any factors that interfere with the synthesis or metabolism of NO in endothelial cells are involved in CVD pathogenesis. It is well established that hypoxia is both the triggering factor as well as the accompanying factor in cardiovascular disease, and diminished tissue oxygen levels have been reported to influence endothelial NO bioavailability. In endothelial cells, NO is produced by endothelial nitric oxide synthase (eNOS) from L-Arg, with tetrahydrobiopterin (BH4) as an essential cofactor. Here, we discuss the mechanisms by which hypoxia affects NO bioavailability, including regulation of eNOS expression and activity. What is particularly important is the fact that hypoxia contributes to the depletion of cofactor BH4 and deficiency of substrate L-Arg, and thus elicits eNOS uncoupling-a state in which the enzyme produces superoxide instead of NO. eNOS uncoupling and the resulting oxidative stress is the major driver of endothelial dysfunction and atherogenesis. Moreover, hypoxia induces impairment in mitochondrial respiration and endothelial cell activation; thus, oxidative stress and inflammation, along with the hypoxic response, contribute to the development of endothelial dysfunction.

Neurochemical changes underlying cognitive impairment in olfactory bulbectomized rats and the impact of the mGlu5-positive allosteric modulator CDPPB.

The olfactory bulbectomized (OBX) rat model is a well-established model of depression in which antidepressant drugs reverse deficits in the passive avoidance test 14 days after administration. Recently, the olfactory bulbectomized rat model has been proposed to be a model of Alzheimer's disease (AD), and the available data indicate similarities between the changes that typically occur in AD and those observed in OBX animals. In the present study, the occurrence of neurochemical impairments related to AD were investigated 8 months after OB ablation. The expression of the nitric oxide synthases eNOS and nNOS, receptor for advanced glycation endproducts (RAGEs) and dimethylarginine dimethylaminohydrolase (DDAH1) in the prefrontal cortices (PFCs), hippocampi and striata of olfactory bulbectomized and sham-operated rats was evaluated. Subsequently, the impact of the administration of a positive allosteric modulator of the mGlu5 receptor, CDPPB (14 days, 2.5 or 5 mg/kg), on OBX-related changes was assessed. OB ablation induced typical deficits in passive avoidance. Significant aberrations in the expression of both isoforms of NOS were observed in the hippocampus and striatum, and the expression of DDAH1 was increased in the PFCs of OBX animals. CDPPB at a dose of 5 mg/kg ameliorated cognitive impairment in the passive avoidance test and partially reversed the changes in eNOS and nNOS expression induced by the lesion. The results of this study confirm that some of the neurochemical changes observed in OBX animals may resemble those associated with AD pathology and that activation of the mGlu5 receptor may partially counteract these pathological alterations.

Molecularly targeted nanoparticles: an emerging tool for evaluation of expression of the receptor for advanced glycation end products in a murine model of peripheral artery disease.

BACKGROUND: Molecular imaging with molecularly targeted probes is a powerful tool for studying the spatio-temporal interactions between complex biological processes. The pivotal role of the receptor for advanced glycation end products (RAGE), and its involvement in numerous pathological processes, aroused the demand for RAGE-targeted imaging in various diseases. In the present study, we evaluated the use of a diagnostic imaging agent for RAGE quantification in an animal model of peripheral artery disease, a multimodal dual-labeled probe targeted at RAGE (MMIA-CML). METHODS: PAMAM dendrimer was conjugated with Nε-carboxymethyl-lysine (CML) modified albumin to synthesize the RAGE-targeted probe. A control untargeted agent carried native non-modified human albumin (HSA). Bifunctional p-SCN-Bn-NOTA was used to conjugate the 64Cu radioisotope. Surgical right femoral artery ligation was performed on C57BL/6 male mice. One week after femoral artery ligation, mice were injected with MMIA-CML or MMIA-HSA labeled with 64Cu radioisotope and 60 min later in vivo microPET-CT imaging was performed. Immediately after PET imaging studies, the murine hindlimb muscle tissues were excised and prepared for gene and protein expression analysis. RAGE gene and protein expression was assessed using real-time qPCR and Western blot technique respectively. To visualize RAGE expression in excised tissues, microscopic fluorescence imaging was performed using RAGE-specific antibodies and RAGE-targeted and -control MMIA. RESULTS: Animals subjected to PET imaging exhibited greater MMIA-CML uptake in ischemic hindlimbs than non-ischemic hindlimbs. We observed a high correlation between fluorescent signal detection and radioactivity measurement. Significant RAGE gene and protein overexpression were observed in ischemic hindlimbs compared to non-ischemic hindlimbs at one week after surgical ligation. Fluorescence microscopic staining revealed significantly increased uptake of RAGE-targeted nanoparticles in both ischemic and non-ischemic muscle tissues compared to the control probe but at a higher level in ischemic hindlimbs. Ischemic tissue exhibited explicit RAGE dyeing following anti-RAGE antibody and high colocalization with the MMIA-CML targeted at RAGE. CONCLUSIONS: The present results indicate increased expression of RAGE in the ischemic hindlimb and enable the use of multimodal nanoparticles in both in vitro and in vivo experimental models, creating the possibility for imaging structural and functional changes with a RAGE-targeted tracer.

Pathogenesis of psoriasis in the "omic" era. Part IV. Epidemiology, genetics, immunopathogenesis, clinical manifestation and treatment of psoriatic arthritis.

Psoriatic arthritis (PsA) is a chronic, progressive, inflammatory arthropathy associated with psoriasis as well as a complex pathogenesis. Genetic and environmental factors trigger the development of the immune-mediated auto-inflammatory response in different sites: skin, bone marrow, entheses and synovial tissues. Studies of the last two decades have changed the view of PsA from a mild, non-progressive arthritis to an inflammatory systemic disease with serious health consequences, not only associated with joint dysfunction, but also with an increased risk of cardiovascular disease and socioeconomic consequences with significantly reduced quality of life. The joint damage starts early in the course of the disease, thus early recognition and treatment with modern biological treatments, which may modify the natural history and slow down progression of this debilitating disease, is essential for the patient long-term outcome.

Genetic and Epigenetic Aspects of Atopic Dermatitis.

Atopic dermatitis is a heterogeneous disease, in which the pathogenesis is associated with mutations in genes encoding epidermal structural proteins, barrier enzymes, and their inhibitors; the role of genes regulating innate and adaptive immune responses and environmental factors inducing the disease is also noted. Recent studies point to the key role of epigenetic changes in the development of the disease. Epigenetic modifications are mainly mediated by DNA methylation, histone acetylation, and the action of specific non-coding RNAs. It has been documented that the profile of epigenetic changes in patients with atopic dermatitis (AD) differs from that observed in healthy people. This applies to the genes affecting the regulation of immune response and inflammatory processes, e.g., both affecting Th1 bias and promoting Th2 responses and the genes of innate immunity, as well as those encoding the structural proteins of the epidermis. Understanding of the epigenetic alterations is therefore pivotal to both create new molecular classifications of atopic dermatitis and to enable the development of personalized treatment strategies.

Pathogenesis of psoriasis in the "omic" era. Part I. Epidemiology, clinical manifestation, immunological and neuroendocrine disturbances.

Psoriasis is a common, chronic, inflammatory, immune-mediated skin disease affecting about 2% of the world's population. According to current knowledge, psoriasis is a complex disease that involves various genes and environmental factors, such as stress, injuries, infections and certain medications. The chronic inflammation of psoriasis lesions develops upon epidermal infiltration, activation, and expansion of type 1 and type 17 Th cells. Despite the enormous progress in understanding the mechanisms that cause psoriasis, the target cells and antigens that drive pathogenic T cell responses in psoriatic lesions are still unproven and the autoimmune basis of psoriasis still remains hypothetical. However, since the identification of the Th17 cell subset, the IL-23/Th17 immune axis has been considered a key driver of psoriatic inflammation, which has led to the development of biologic agents that target crucial elements of this pathway. Here we present the current understanding of various aspects in psoriasis pathogenesis.

Quantitative imaging of the receptor for advanced glycation end-products in prostate cancer.

PURPOSE: Current screening and monitoring of prostate cancer (PCa) is insufficient, producing inaccurate diagnoses. Presence of the receptor for advanced glycation end-products (RAGE) is associated with signature characteristics of PCa development such as cell proliferation, anchorage-independent growth, angiogenesis, migration, invasion, and poor patient survival. Therefore, we developed a preclinical multimodal imaging strategy targeted at RAGE to diagnose and monitor PCa. METHODS: In this work, RAGE-targeted multimodal nanoparticles (64Cu-Cy5-G4-CML) were synthesized and rendered functional for nuclear and optical imaging using previously established methods. The probe's binding affinity and targeting specificity was assessed in androgen-dependent (LNCaP) and androgen-independent (DU145) prostate cancer cells using flow cytometry and confocal microscopy. In vivo PET-CT imaging was used to evaluate RAGE levels in DU145 and LNCaP xenograft models in mice. Then, tumors were excised post-imaging for histological staining and autoradiography to further assess RAGE levels and targeting efficiency of the tracer. Finally, RAGE levels from human PCa samples of varying Gleason Scores were evaluated using Western blot and immunohistochemical staining. RESULTS: PCa cell culture studies confirmed adequate RAGE-targeting with 64Cu-Cy5-G4-CML with KD between 360 and 540 nM as measured by flow cytometry. In vivo PET-CT images of PCa xenografts revealed favorable kinetics, rapid blood clearance, and a non-homogenous, enhanced uptake in tumors, which varied based on cell type and tumor size with mean uptake between 0.5 and 1.4%ID/g. RAGE quantification of human samples confirmed increased RAGE uptake corresponding to increased Gleason scoring. CONCLUSIONS: Our study has shown that RAGE-targeted cancer imaging is feasible and could significantly impact PCa management.

Multimodal imaging of the receptor for advanced glycation end-products with molecularly targeted nanoparticles.

The receptor for advanced glycation end-products (RAGE) is central to multiple disease states, including diabetes-related conditions such as peripheral arterial disease (PAD). Despite RAGE's importance in these pathologies, there remains a need for a molecular imaging agent that can accurately assess RAGE levels in vivo. Therefore, we have developed a multimodal nanoparticle-based imaging agent targeted at RAGE with the well-characterized RAGE ligand, carboxymethyllysine (CML)-modified human serum albumin (HSA). Methods: A multimodal tracer (64Cu-Rho-G4-CML) was developed using a generation-4 (G4) polyamidoamine (PAMAM) dendrimer, conjugated with both rhodamine and copper-64 (64Cu) chelator (NOTA) for optical and PET imaging, respectively. First, 64Cu-Rho-G4-CML and its non-targeted analogue (64Cu-Rho-G4-HSA) were evaluated chemically using techniques such as dynamic light scattering (DLS), electron microscopy and nuclear magnetic resonance (NMR). The tracers' binding capabilities were examined at the cellular level and optimized using live and fixed HUVEC cells grown in 5.5-30 mM glucose, followed by in vivo PET-CT imaging, where the probes' kinetics, biodistribution, and RAGE targeting properties were examined in a murine model of hindlimb ischemia. Finally, histological assessment of RAGE levels in both ischemic and non-ischemic tissues was performed. Conclusions: Our RAGE-targeted probe demonstrated an average size of 450 nm, a Kd of 340-390 nM, rapid blood clearance, and a 3.4 times greater PET uptake in ischemic RAGE-expressing hindlimbs than their non-ischemic counterpart. We successfully demonstrated increased RAGE expression in a murine model of hindlimb ischemia and the feasibility for non-invasive examination of cellular, tissue, and whole-body RAGE levels with a molecularly targeted tracer.

Altered circadian genes expression in breast cancer tissue according to the clinical characteristics.

Breast cancer has a multifactorial etiology. One of the supposed and novel mechanisms is an alteration of circadian gene expression. Circadian genes play a crucial role in many physiological processes. These processes, such as genomic stability, DNA repair mechanism and apoptosis, are frequently disrupted in breast tumors. To assess the significance of circadian gene expression in breast cancer, we carried out an analysis of CLOCK, BMAL1, NPAS2, PER1, PER2, PER3 and CRY1, CRY2, TIMELESS, CSNK1E expression by the use of the quantitative Real-Time PCR technique in tumor tissue and non-tumor adjacent normal tissue sampled from 107 women with a newly diagnosed disease. The obtained data were compared to the clinical and histopathological features. PER1, PER2, PER3, CRY2 were found to be significantly down-expressed, while CLOCK, TIMELESS were over-expressed in the studied tumor samples compared to the non-tumor samples. Only gene expression of CRY1 was significantly down-regulated with progression according to the TNM classification. We found significantly decreased expression of CRY2, PER1, PER2 genes in the ER/PR negative breast tumors compared to the ER/PR positive tumors. Additionally, expression of CRY2, NPAS2 genes had a decreased level in the poorly differentiated tumors in comparison with the well and moderately differentiated ones. Our results indicate that circadian gene expression is altered in breast cancer tissue, which confirms previous observations from various animal and in vitro studies.

eNOS expression and NO release during hypoxia is inhibited by miR-200b in human endothelial cells.

The nitric oxide (NO) secreted by vascular endothelium is required for the maintenance of cardiovascular homeostasis. Diminished release of NO generated by endothelial NO synthase contributes to endothelial dysfunction. Hypoxia and ischemia reduce endothelial eNOS expression via posttranscriptional mechanisms that result in NOS3 transcript destabilization. Here, we examine whether microRNAs contribute to this mechanism. We followed the kinetics of hypoxia-induced changes in NOS3 mRNA and eNOS protein levels in primary human umbilical vein endothelial cells (HUVECs). Utilizing in silico predictive protocols to identify potential miRNAs that regulate eNOS expression, we identified miR-200b as a candidate. We established the functional miR-200b target sequence within the NOS3 3'UTR, and demonstrated that manipulation of the miRNA levels during hypoxia using miR-200b mimics and antagomirs regulates eNOS levels, and established that miR-200b physiologically limits eNOS expression during hypoxia. Furthermore, we demonstrated that the specific ablation of the hypoxic induction of miR-200b in HUVECs restored eNOS-driven hypoxic NO release to the normoxic levels. To determine whether miR-200b might be the only miRNA that had this effect, we utilized Next Generation Sequencing (NGS) to follow hypoxia-induced changes in the miRNA levels in HUVECS and found 83 novel hypoxamiRs, with two candidate miRNAs besides miR-200b that could potentially influence eNOS levels. Taken together, the data establish miR-200b-eNOS regulation as a first hypoxamiR-based mechanism that limits NO bioavailability during hypoxia in endothelial cells, and show that hypoxamiRs could become useful therapeutic targets for cardiovascular diseases and other hypoxic-related diseases including various types of cancer.

Posttranscriptional and transcriptional regulation of endothelial nitric-oxide synthase during hypoxia: the role of microRNAs.

Understanding the cellular pathways that regulate endothelial nitric oxide (eNOS, NOS3) expression and consequently nitric oxide (NO) bioavailability during hypoxia is a necessary aspect in the development of novel treatments for cardiovascular disorders. eNOS expression and eNOS-dependent NO cellular signaling during hypoxia promote an equilibrium of transcriptional and posttranscriptional molecular mechanisms that belong to both proapoptotic and survival pathways. Furthermore, NO bioavailability results not only from eNOS levels, but also relies on the presence of eNOS substrate and cofactors, the phosphorylation status of eNOS, and the presence of reactive oxygen species (ROS) that can inactivate eNOS. Since both NOS3 levels and these signaling pathways can also be a subject of posttranscriptional modulation by microRNAs (miRNAs), this class of short noncoding RNAs contribute another level of regulation for NO bioavailability. As miRNA antagomirs or specific target protectors could be used in therapeutic approaches to regulate NO levels, either by changing NOS3 mRNA stability or through factors governing eNOS activity, it is critical to understand their role in governing eNOS activity during hypoxa. In contrast to a large number of miRNAs reported to the change eNOS expression during hypoxia, only a few miRNAs modulate eNOS activity. Furthermore, impaired miRNA biogenesis leads to NOS3 mRNA stabilization under hypoxia. Here we discuss the recent studies that define miRNAs' role in maintaining endothelial NO bioavailability emphasizing those miRNAs that directly modulate NOS3 expression or eNOS activity.