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Pavlik, V.

Publications and source records attributed to Pavlik, V..

2 recordsLinked to original sources
bioRxiv2026

Macrophage-based assays for the in vitro testing of the anti-inflammatory activity of mesenchymal stem cell-based products

Therapies based on mesenchymal stromal cells (MSCs) have high potential in the field of regenerative medicine due mainly to their immunomodulatory properties. However, their clinical translation is hampered by a lack of sufficiently standardised potency tests. Since macrophages comprise key mediators of the effects of MSCs, macrophage-based assays potentially provide a relevant in vitro tool for the evaluation of the activity of MSC products. This study involved the coculturing of canine adipose-derived mesenchymal stem cells (ASCs) with macrophages derived from human THP-1 and U937 monocyte cell lines, murine RAW264.7 macrophages and primary human macrophages. The M2 polarisation was assessed following stimulation with IL-4/IL-13. The mRNA expression of the pro- and anti-inflammatory markers was analysed applying qPCR. The ASC secretome acted to reduce the pro-inflammatory mRNA expression across all the macrophage models, albeit with a certain degree of model-dependent variability. Only the U937 macrophages responded consistently to the M2-polarising stimuli, while the RAW264.7 cells provided practical advantages in terms of routine screening. The results thus provided support for the application of macrophage-based potency assays as a suitable platform for the testing of MSC products; the U937 cells were found to be particularly suitable for the study of polarisation and the RAW264.7 cells for standardised screening.

cell biology↗
bioRxiv2022

Phosphorylation of tyrosine 90 in SH3 domain is a new regulatory switch controlling Src kinase

The activation of Src kinase in cells is strictly controlled by intramolecular inhibitory interactions mediated by SH3 and SH2 domains. They impose structural constraints on the kinase domain holding it in a catalytically non-permissive state. The transition between inactive and active conformation is known to be largely regulated by the phosphorylation state of key tyrosines 416 and 527. Here we identified that phosphorylation of tyrosine 90 reduces binding affinity of the SH3 domain to its interacting partners, opens the Src structure, and renders Src catalytically active. This is accompanied by an increased affinity to the plasma membrane, decreased membrane motility and slower diffusion from focal adhesions. Phosphorylation of tyrosine 90 controlling SH3-medited intramolecular inhibitory interaction, analogical to tyrosine 527 regulating SH2-C-terminus bond, enables SH3 and SH2 domains to serve as cooperative but independent regulatory elements. This mechanism allows Src to adopt several distinct conformations of varying catalytic activities and interacting properties, enabling it to operate not as a simple switch but as a tunable regulator functioning as a signaling hub in a variety of cellular processes.

cell biology↗