Cav1.2

In this work, we propose a simple, reliable, and versatile strategy to create 3D electroconductive scaffolds suitable for bone tissue engineering (TE) applications with electrical stimulation (ES). The proposed scaffolds are made of 3D-extruded poly(ε-caprolactone) (PCL), subjected to alkaline treatment, and of poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (PEDOT:PSS), anchored to PCL with one of two different crosslinkers: (3-glycidyloxypropyl)trimethoxysilane (GOPS) and divinyl sulfone (DVS). Both cross-linkers allowed the formation of a homogenous and continuous coating of PEDOT:PSS to PCL. We show that these PEDOT:PSS coatings are electroconductive (11.3-20.1 S cm^-1), stable (up to 21 days in saline solution), and allow the immobilization of gelatin (Gel) to further improve bioactivity. In vitro mineralization of the corresponding 3D conductive scaffolds was greatly enhanced (GOPS(NaOH)-Gel - 3.1 fold, DVS(NaOH)-Gel - 2.0 fold) and cell colonization and proliferation were the highest for the DVS(NaOH)-Gel scaffold. In silico modelling of ES application in DVS(NaOH)-Gel scaffolds indicates that the electrical field distribution is homogeneous, which reduces the probability of formation of faradaic products. Osteogenic differentiation of human bone marrow derived mesenchymal stem/stromal cells (hBM-MSCs) was performed under ES. Importantly, our results clearly demonstrated a synergistic effect of scaffold electroconductivity and ES on the enhancement of MSC osteogenic differentiation, particularly on cell-secreted calcium deposition and the upregulation of osteogenic gene markers such as COL I, OC and CACNA1C. These scaffolds hold promise for future clinical applications, including manufacturing of personalized bone TE grafts for transplantation with enhanced maturation/functionality or bioelectronic devices.

Adipocytes normally accumulate in the epicardial and pericardial layers around the human heart, but their infiltration into the myocardium can be proarrhythmic. METHODS AND RESULTS: Human adipose derived stem/stromal cells and human induced pluripotent stem cells (hiPSC) were differentiated, respectively into predominantly white fat-like adipocytes (hAdip) and ventricular cardiomyocytes (CMs). Adipocytes cultured in CM maintenance medium (CM medium) maintained their morphology, continued to express adipogenic markers, and retained clusters of intracellular lipid droplets. In contrast, hiPSC-CMs cultivated in adipogenic growth medium displayed abnormal cell morphologies and more clustering across the monolayer. Pre-plated hiPSC-CMs co-cultured in direct contact with hAdips in CM medium displayed prolonged action potential durations, increased triangulation, slowed conduction velocity, increased conduction velocity heterogeneity, and prolonged calcium transients. When hAdip-conditioned medium was added to monolayer cultures of hiPSC-CMs, results similar to those recorded with direct co-cultures were observed. Both co-culture and conditioned medium experiments resulted in increases in transcript abundance of SCN10A, CACNA1C, SLC8A1, and RYR2, with a decrease in KCNJ2. Human adipokine immunoblots revealed the presence of cytokines that were elevated in adipocyte-conditioned medium, including MCP-1, IL-6, IL-8 and CFD that could induce electrophysiological changes in cultured hiPSC-CMs. CONCLUSIONS: Co-culture of hiPSC-CMs with hAdips reveals a potentially pathogenic role of infiltrating human adipocytes on myocardial tissue. In the absence of structural changes, hAdip paracrine release alone is sufficient to cause CM electrophysiological dysfunction mirroring the co-culture conditions. These effects, mediated largely by paracrine mechanisms, could promote arrhythmias in the heart.

Brain-derived estrogen (BDE2) is gaining attention as an endogenous neurotransmitter. Recent research has revealed that selectively removing the aromatase gene, the pivotal enzyme responsible for BDE2 synthesis, in forebrain neurons or astrocytes can lead to synaptic loss and cognitive impairment. It is worth noting that remote ischemia post-conditioning (RIP), a non-invasive technique, has been shown to activate natural protective mechanisms against severe ischemic events. The aim of our study was to investigate whether RIP triggers aromatase-BDE2 signaling, shedding light on its neuroprotective mechanisms after global cerebral ischemia (GCI) in ovariectomized rats. Our findings are as follows: (1) RIP was effective in mitigating ischemic damage in hippocampal CA1 neurons and improved cognitive function after GCI. This was partially due to increased Aro-BDE2 signaling in CA1 neurons. (2) RIP intervention efficiently enhanced pro-survival kinase pathways, such as AKT, ERK1/2, CREB, and suppressed CaMKIIα signaling in CA1 astrocytes induced by GCI. Remarkably, inhibiting CaMKIIα activity led to elevated Aro-BDE2 levels and replicated the benefits of RIP. (3) We also identified the positive mediation of Cav1.2, an LVGCC calcium channel, on CaMKIIα-Aro/BDE2 pathway response to RIP intervention. (4) Significantly, either RIP or CaMKIIα inhibition was found to alleviate reactive astrogliosis, which was accompanied by increased pro-survival A2-astrocyte protein S100A10 and decreased pro-death A1-astrocyte marker C3 levels. In summary, our study provides compelling evidence that Aro-BDE2 signaling is a critical target for the reparative effects of RIP following ischemic insult. This effect may be mediated through the CaV1.2-CaMKIIα signaling pathway, in collaboration with astrocyte-neuron interactions, thereby maintaining calcium homeostasis in the neuronal microenvironment and reducing neuronal damage after ischemia.