Vascular smooth muscle cells exhibit elevated hypoxia-inducible Factor-1α expression in human blood vessel organoids, influencing osteogenic performance.

Considering the importance of alternative methodologies to animal experimentation, we propose an organoid-based biological model for in vitro blood vessel generation, achieved through co-culturing endothelial and vascular smooth muscle cells (VSMCs). Initially, the organoids underwent comprehensive characterization, revealing VSMCs (α-SMA + cells) at the periphery and endothelial cells (CD31+ cells) at the core. Additionally, ephrin B2 and ephrin B4, genes implicated in arterial and venous formation respectively, were used to validate the obtained organoid. Moreover, the data indicates exclusive HIF-1α expression in VSMCs, identified through various methodologies. Subsequently, we tested the hypothesis that the generated blood vessels have the capacity to modulate the osteogenic phenotype, demonstrating the ability of HIF-1α to promote osteogenic signals, primarily by influencing Runx2 expression. Overall, this study underscores that the methodology employed to create blood vessel organoids establishes an experimental framework capable of producing a 3D culture model of both venous and arterial endothelial tissues. This model effectively guides morphogenesis from mesenchymal stem cells through paracrine signaling, ultimately leading to an osteogenic acquisition phenotype, with the dynamic involvement of HIF-1α.

Trifluoroacetic acid as excipient destabilizes melittin causing the selective aggregation of melittin within the centrin-melittin-trifluoroacetic acid complex.

Trifluoroacetic acid (TFA) may be the cause of the bottleneck in high resolution structure determination for protein-peptide complexes. Fragment based drug design often involves the use of synthetic peptides which contain TFA (excipient). Our goal was to explore the effects of this excipient on a model complex: centrin-melittin-TFA. We performed Fourier transform infrared, two-dimensional infrared correlation spectroscopies and spectral simulations to analyze the amide I'/I'* band for the components and the ternary complex. Melittin (MLT) was observed to have increased helicity upon its interaction with centrin, followed by the thermally induced aggregation of MLT within the ternary complex in the TFA presence.

Estradiol interacts with insulin through membrane receptors to induce an antimitogenic effect on lactotroph cells.

The signaling mechanisms of estrogens interact with those of growth factors to control the pituitary gland functions. The contribution of the membrane bound estrogen receptor in these actions is not fully understood. In this study, we focused on the regulatory action of estradiol in interaction with insulin on the secretory and proliferative lactotroph cell activities from primary pituitary cell cultures. Furthermore, we studied the involvement of ERK1/2, PKC epsilon and Pit-1 in these actions. In serum free conditions, estradiol and estradiol-BSA promoted a differential secretory activity on PRL cells but were unable to induce lactotroph cell proliferation. However, both free and conjugated estradiol were competent arresting the mitogenic activity promoted by insulin. Estradiol, estradiol-BSA and insulin stimuli increased the PKC epsilon, phosphorylated ERK 1/2 and Pit-1 expression, although combined treatments with estradiol/insulin or estradiol-BSA/insulin induced a significant reduction in these levels, in close correlation with the decrease of lactotroph cell proliferation. The pre-treatment with PKC inhibitor BIM significantly inhibited the ERK activation promoted by insulin without modifying the ERK expression levels induced by estradiol or estradiol-BSA. By immuno-electron-microscopy the alpha nuclear estrogen receptor was localized in the plasma membrane of lactotroph cells. These findings suggest that the membrane bound ER participates modulating lactotroph cells proliferation via PKC epsilon, ERK1/2 and Pit-1. The interactions between estradiol and growth factors, inducing both mitogenic and antimitogenic effects, could provide glandular plasticity preventing an over-proliferation induced by growth factors.