Exposure to Pulsed Electromagnetic Field (PEMF) has been shown to affect proliferation and differentiation of human mesenchymal stem cells derived from bone marrow stroma (BM-hMSC). These cells offer considerable promise in the field of regenerative medicine, but their clinical application is hampered by major limitations such as poor availability and the time required to differentiate up to a stage suitable for implantation. For this reason, several research efforts are focusing on identifying strategies to speed up the differentiation process. In this work we investigated thein vitroeffect of PEMF on Ca²⁺-related mechanisms promoting the osteogenic differentiation of BM-hMSC. Cells were daily exposed to PEMF while subjected to osteogenic differentiation and various Ca²⁺-related mechanisms were monitored using multiple approaches for identifying functional and structural modifications related to this process. The results indicate that PEMF exposure promotes chemically induced osteogenesis by mechanisms that mainly interfere with some of the calcium-related osteogenic pathways, such as permeation and regulation of cytosolic concentration, leaving others, such as extracellular deposition, unaffected. The PEMF effect is primarily associated to early enhancement of intracellular calcium concentration, which is proposed here as a reliable hallmark of the osteogenic developmental stage.

Cortical spreading depression (CSD), a wave of depolarization followed by depression of cortical activity, is a pathophysiological process implicated in migraine with aura and various other brain pathologies, such as ischemic stroke and traumatic brain injury. To gain insight into the pathophysiology of CSD, we generated a mouse model for a severe monogenic subtype of migraine with aura, familial hemiplegic migraine type 3 (FHM3). FHM3 is caused by mutations in SCN1A, encoding the voltage-gated Na⁺ channel Na_V1.1 predominantly expressed in inhibitory interneurons. Homozygous Scn1a^L1649Q knock-in mice died prematurely, whereas heterozygous mice had a normal lifespan. Heterozygous Scn1a^L1649Q knock-in mice compared with WT mice displayed a significantly enhanced susceptibility to CSD. We found L1649Q to cause a gain-of-function effect with an impaired Na⁺-channel inactivation and increased ramp Na⁺ currents leading to hyperactivity of fast-spiking inhibitory interneurons. Brain slice recordings using K⁺-sensitive electrodes revealed an increase in extracellular K⁺ in the early phase of CSD in heterozygous mice, likely representing the mechanistic link between interneuron hyperactivity and CSD initiation. The neuronal phenotype and premature death of homozygous Scn1a^L1649Q knock-in mice was partially rescued by GS967, a blocker of persistent Na⁺ currents. Collectively, our findings identify interneuron hyperactivity as a mechanism to trigger CSD.

The pore-forming subunits (α subunits) of voltage-gated sodium channels (VGSC) are encoded in humans by a family of nine highly conserved genes. Among them, SCN1A, SCN2A, SCN3A, and SCN8A are primarily expressed in the central nervous system. The encoded proteins Nav1.1, Nav1.2, Nav1.3, and Nav1.6, respectively, are important players in the initiation and propagation of action potentials and in turn of the neural network activity. In the context of neurological diseases, mutations in the genes encoding Nav1.1, 1.2, 1.3 and 1.6 are responsible for many forms of genetic epilepsy and for Nav1.1 also of hemiplegic migraine. Several pharmacological therapeutic approaches targeting these channels are used or are under study. Mutations of genes encoding VGSCs are also involved in autism and in different types of even severe intellectual disability (ID). It is conceivable that in these conditions their dysfunction could indirectly cause a certain level of neurodegenerative processes; however, so far, these mechanisms have not been deeply investigated. Conversely, VGSCs seem to have a modulatory role in the most common neurodegenerative diseases such as Alzheimer’s, where SCN8A expression has been shown to be negatively correlated with disease severity.

The observation that vitamin D-mediated enhancement of osteocalcin (OC) gene expression is dependent on and reciprocally related to the level of basal gene expression suggests that an interaction of the vitamin D responsive element (VDRE) with basal regulatory elements of the OC gene promoter contributes to both basal and vitamin D-enhanced transcription. Protein-DNA interactions at the VDRE of the rat OC gene (nucleotides -466 to -437) are reflected by direct sequence-specific and antibody-sensitive binding of the endogenous vitamin D receptor present in ROS 17/2.8 osteosarcoma nuclear protein extracts. In addition, a vitamin D-responsive increase in OC gene transcription is accompanied by enhanced non-vitamin D receptor-mediated protein-DNA interactions in the "TATA" box region (nucleotides -44 to +23), which also contains a potential glucocorticoid responsive element. Evidence for proximity of the VDRE with the basal regulatory elements is provided by two features of nuclear architecture. (i) Nuclear matrix attachment elements in the rat OC gene promoter that bind nuclear matrix proteins with sequence specificity may impose structural constraints on promoter conformation. (ii) Limited micrococcal nuclease digestion and Southern blot analysis indicate that three nucleosomes can be accommodated in the sequence spanning the OC gene VDRE, the OC/CCAAT box (nucleotides -99 to -76), and the TATA/glucocorticoid responsive element, and thereby the potential distance between the VDRE and the basal regulatory elements can be reduced. A model is presented for the contribution of both the VDRE and proximal promoter elements to the enhancement of OC gene transcription in response to vitamin D. The vitamin D receptor plus accessory proteins may function cooperatively with basal regulatory factors to modulate the extent to which the OC gene is transcribed.