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Possible role of VEGF/Akt transduction pathway after beta-2 adrenergic receptors overexpression-promoted angiogenesis.

Exercise upregulates beta-adrenergic receptor expression and this, in turn, stimulates angiogenesis.

Angiogenesis in the failing heart is stimulated beta-1 AR blockade and cardiac beta-2 AR overexpression.

We review the literature on the little-known roles of specific CaMKs in regulating endocrine functions of the pineal gland, the pituitary gland, and the hypothalamus. Melatonin activates hippocampal CaMKII, which then influences dendritogenesis. In the pituitary gland, the signal pathways activated by the CaMK in lower vertebrates, such as fishes, differ from those of mammals. In the teleost anterior pituitary, the activation of CaMKII induces the expression of somatolactin by glucagon b. In rats and humans, CaMKIVs have been associated with gonadotropes and thyrotropes and CaMKII with several types of human tumor cells and with a specific signaling pathway. Neuropeptides such as vasopressin and endothelin are also involved in the CaMKII signaling chain, as is the CaMKIIδ isoform which participates in generating the circadian rhythms of the suprachiasmatic nucleus. What arises from this review is that most of the hypothalamic CaMKs are involved in activities of the endocrine brain. Furthermore, among the CaMKs, type II occurs with the highest frequency followed by CaMKIV and CaMKI.

Human induced pluripotent stem cells (hiPSCs) are emerging as a useful tool for modelling in vitro early brain development and neurological disorders. Molecular mechanisms and cell interactions that regulate the neurodevelopment at early stages remain unclear because of human brain’s complexity and limitations of functional studies. Two major culture methodologies are used to differentiate in vitro hiPSCs into neurons: monolayer (2D) and organoid (3D) cultures. Here we investigate the effect of cell dissociation and the loss of 3D organization during the early differentiation process of neuronal progenitors. Using the same culture media, we first differentiated hiPSCs into neural progenitor cells (NPCs) and then induced their differentiation into neurons in 3 different modalities: 3D undissociated organoids, dissociated NPCs followed by immediate re-aggregation into an organoid, and dissociated NPCs cultured as monolayer. We assessed neuronal differentiation efficiency of each method by immunocytochemistry, qPCR, western blot, and RNA-Seq analysis over a time course. Our data revealed substantial differences in gene and protein expression among the three systems, including genes of the Notch pathway (e.g. NEUROD1, NEUROG2), earliest determinants of cortical region differentiation (e.g. SOX1, FEZF1) as well as later transcriptional regulators that specify cortical neuron subtypes (e.g. TBR1, CTIP2), which were all downregulated in monolayer. Moreover, we found that genes and pathways mediating cell-to-cell interactions (e.g. CNTNs, CAMs) were mostly upregulated in the 3D culture systems, whereas cell-extracellular matrix interaction molecules (e.g. ITG, LAM) were mostly upregulated in 2D, indicating that cell surface molecules may be involved in specification of neuronal cell types. Our results address the methodological question of the appropriateness of a differentiation method for a particular experimental goal, and, beyond that, reveal important early determinants that exert a decisive influence on neuronal differentiation and regional specification of human neural stem cells., Italian Journal of Anatomy and Embryology, Vol. 122, No. 1 (Supplement) 2017

It has already been reported that cannabinoids are neuroprotective agents against excitotoxicity in vitro and increase after acute brain damage in vivo. This background prompted us to study the localization and expression of the calcium -binding protein calretinin in a condition similar to Alzheimer disease and its possible relationship with cannabinoids and their supposed protective role. We carried out quantitative analysis of the transient changes in calretinin expression shown by hybridochemistry within neuronal cell populations in the hippocampus of a beta amyloid-treated rat model of Alzheimer׳s disease and their correlation with endocannabinoid increase. Calretinin expression increases throughout the first week after cortical amyloid-beta peptide injection, and then decreases towards normal levels in the rat hippocampus during the following weeks, indicating that decreased calretinin gene expression may be associated with either increase of endocannabinoids or VDM11-induced accumulation of endocannabinoids. In contrast, SR1, an antagonist, which limits the cannabinoid effect by selective binding to the cannabinoid receptor CB1, up-regulates calretinin expression with respect to non-treated rats. This could mean that the SR1 endocannabinoid-blocking action through CB1 receptors, that are normally stimulated by endocannabinoids to inhibit calcium increase, might cause a higher calretinin expression. This would allow us to speculate on a possible reverse relationship between endocannabinoid and calretinin levels in the hippocampal calcium-homeostasis balance.

Background Uncoupling protein 3 (ucp3) is a member of the mitochondrial anion carrier superfamily of proteins uncoupling mitochondrial respiration. In this study, we investigated the effects of ucp3 genetic deletion on mitochondrial function and cell survival under low oxygen conditions in vitro and in vivo. Methods and Results To test the effects of ucp3 deletion in vitro, murine embryonic fibroblasts and adult cardiomyocytes were isolated from wild‐type ( WT , n=67) and ucp3 knockout mice (ucp3 −/− , n=70). To test the effects of ucp3 genetic deletion in vivo, myocardial infarction ( MI ) was induced by permanent coronary artery ligation in WT and ucp3 −/− mice. Compared with WT , ucp3 −/− murine embryonic fibroblasts and cardiomyocytes exhibited mitochondrial dysfunction and increased mitochondrial reactive oxygen species generation and apoptotic cell death under hypoxic conditions in vitro (terminal deoxynucleotidyl transferase‐dUTP nick end labeling–positive nuclei: WT hypoxia, 70.3±1.2%; ucp3 −/− hypoxia, 85.3±0.9%; P MI , despite similar areas at risk in the 2 groups, ucp3 −/− hearts demonstrated a significantly larger infarct size compared with WT (infarct area/area at risk: WT , 48.2±3.7%; ucp3 −/− , 65.0±2.9%; P MI , cardiac function was significantly decreased in ucp3 −/− mice compared with WT (fractional shortening: WT MI , 42.7±3.1%; ucp3 −/− MI , 24.4±2.9; P WT MI , 0.7±0.04%; ucp3 −/− MI , 1.1±0.09%, P Conclusions Our data indicate that ucp3 levels regulate reactive oxygen species levels and cell survival during hypoxia, modulating infarct size in the ischemic heart.

We investigated whether β(2) -adrenoceptor overexpression could promote angiogenesis and improve blood perfusion and left ventricular (LV) remodeling of the failing heart.We explored the angiogenic effects of β(2) -adrenoceptor overexpression in a rat model of post-myocardial infarction (MI) heart failure (HF). Cardiac adenoviral-mediated β(2) -adrenoceptor overexpression was obtained via direct intramyocardial injection 4-weeks post-MI. Adenovirus(Ad)-GFP and saline injected rats served as controls. Furthermore, we extended our observation to β(2) -adrenoceptor -/- mice undergoing MI.Transgenes were robustly expressed in the LV at 2 weeks post-gene therapy, whereas their expression was minimal at 4-weeks post-gene delivery. In HF rats, cardiac β(2) -adrenoceptor overexpression resulted in enhanced basal and isoprenaline-stimulated cardiac contractility at 2-weeks post-gene delivery. At 4 weeks post-gene transfer, Ad-β(2) -adrenoceptor HF rats showed improved LV remodeling and cardiac function. Importantly, β(2) -adrenoceptor overexpression was associated with a markedly increased capillary and arteriolar length density and enhanced in vivo myocardial blood flow and coronary reserve. At the molecular level, cardiac β(2) -adrenoceptor gene transfer induced the activation of the VEGF/PKB/eNOS pro-angiogenic pathway. In β(2) -adrenoceptor-/- mice, we found a ~25% reduction in cardiac capillary density compared with β(2) -adrenoceptor+/+ mice. The lack of β(2) -adrenoceptors was associated with a higher mortality rate at 30 days and LV dilatation, and a worse global cardiac contractility compared with controls.β(2) -Adrenoceptors play an important role in the regulation of the angiogenic response in HF. The activation of VEGF/PKB/eNOS pathway seems to be strongly involved in this mechanism.

Background LMO2 is highly expressed at the most immature stages of lymphopoiesis. In T-lymphocytes, aberrant LMO2 expression beyond those stages leads to T-cell acute lymphoblastic leukemia, while in B cells LMO2 is also expressed in germinal center lymphocytes and diffuse large B-cell lymphomas, where it predicts better clinical outcome. The implication of LMO2 in B-cell acute lymphoblastic leukemia must still be explored. Design and Methods We measured LMO2 expression by real time RT-PCR in 247 acute lymphoblastic leukemia patient samples with cytogenetic data (144 of them also with survival and immunophenotypical data) and in normal hematopoietic and lymphoid cells. Results B-cell acute lymphoblastic leukemia cases expressed variable levels of LMO2 depending on immunophenotypical and cytogenetic features. Thus, the most immature subtype, pro-B cells, displayed three-fold higher LMO2 expression than pre-B cells, common-CD10+ or mature subtypes. Additionally, cases with TEL-AML1 or MLL rearrangements exhibited two-fold higher LMO2 expression compared to cases with BCR-ABL rearrangements or hyperdyploid karyotype. Clinically, high LMO2 expression correlated with better overall survival in adult patients (5-year survival rate 64.8% (42.5%–87.1%) vs . 25.8% (10.9%–40.7%), P = 0.001) and constituted a favorable independent prognostic factor in B-ALL with normal karyotype: 5-year survival rate 80.3% (66.4%–94.2%) vs. 63.0% (46.1%–79.9%) ( P = 0.043). Conclusions Our data indicate that LMO2 expression depends on the molecular features and the differentiation stage of B-cell acute lymphoblastic leukemia cells. Furthermore, assessment of LMO2 expression in adult patients with a normal karyotype, a group which lacks molecular prognostic factors, could be of clinical relevance.

β 2 -Adrenergic receptors (β 2 ARs) are widely expressed, although their physiological relevance in many tissues is not yet fully understood. In vascular endothelial cells, they regulate NO release and vessel tone. Here we provide novel evidence that β 2 ARs can regulate neoangiogenesis in response to chronic ischemia. We used in vivo adenoviral-mediated gene transfer of the human β 2 AR to the endothelium of the rat femoral artery and increased β 2 AR signaling resulting in ameliorated angiographic blood flow and hindlimb perfusion after chronic ischemia. Histological analysis confirmed that β 2 AR overexpression also produced benefits on capillary density. The same maneuver partially rescued impaired angiogenesis in spontaneously hypertensive rats (SHR), whereas gene delivery of the G-protein–coupling defective mutant Ile164 β 2 AR failed to provide ameliorations. Stimulation of endogenous and overexpressed β 2 AR on endothelial cells in vitro was found to regulate cell number by inducing proliferation and [ 3 H]-thymidine incorporation through means of extracellular receptor-activated kinase and vascular endothelial growth factor. The β 2 AR also has novel effects on endothelial cell number through stimulation of proapoptosis and antiapoptosis pathways involving p38 mitogen-activated protein kinase and PI3-kinase/Akt activation. Therefore, β 2 ARs play a critical role in endothelial cell proliferation and function including revascularization, suggesting a novel and physiologically relevant role in neoangiogenesis in response to ischemia.