Targeting F-actin stress fibers to suppress the dedifferentiated phenotype in chondrocytes.

Actin is a central mediator of the chondrocyte phenotype. Monolayer expansion of articular chondrocytes on tissue culture polystyrene, for cell-based repair therapies, leads to chondrocyte dedifferentiation. During dedifferentiation, chondrocytes spread and filamentous (F-)actin reorganizes from a cortical to a stress fiber arrangement causing a reduction in cartilage matrix expression and an increase in fibroblastic matrix and contractile molecule expression. While the downstream mechanisms regulating chondrocyte molecular expression by alterations in F-actin organization have become elucidated, the critical upstream regulators of F-actin networks in chondrocytes are not completely known. Tropomyosin (TPM) and the RhoGTPases are known regulators of F-actin networks. The main purpose of this study is to elucidate the regulation of passaged chondrocyte F-actin stress fiber networks and cell phenotype by the specific TPM, TPM3.1, and the RhoGTPase, CDC42. Our results demonstrated that TPM3.1 associates with cortical F-actin and stress fiber F-actin in primary and passaged chondrocytes, respectively. In passaged cells, we found that pharmacological TPM3.1 inhibition or siRNA knockdown causes F-actin reorganization from stress fibers back to cortical F-actin and causes an increase in G/F-actin. CDC42 inhibition also causes formation of cortical F-actin. However, pharmacological CDC42 inhibition, but not TPM3.1 inhibition, leads to the re-association of TPM3.1 with cortical F-actin. Both TPM3.1 and CDC42 inhibition, as well as TPM3.1 knockdown, reduces nuclear localization of myocardin related transcription factor, which suppresses dedifferentiated molecule expression. We confirmed that TPM3.1 or CDC42 inhibition partially redifferentiates passaged cells by reducing fibroblast matrix and contractile expression, and increasing chondrogenic SOX9 expression. A further understanding on the regulation of F-actin in passaged cells may lead into new insights to stimulate cartilage matrix expression in cells for regenerative therapies.

Accessing Ambiphilic Phosphine Boronates through C-H Borylation by an Unforeseen Cationic Iridium Complex.

Ambiphilic molecules, which contain a Lewis base and Lewis acid, are of great interest based on their unique ability to activate small molecules. Phosphine boronates are one class of these substrates that have interesting catalytic activity. Direct access to these phosphine boronates is described through the iridium-catalyzed C-H borylation of phosphines. An unconventional cationic iridium catalyst was identified as optimal for a range of phosphines, providing good yields and selectivity across a diverse class of phosphine boronates (isolated as the borane-protected phosphine). A complimentary catalyst system (quinoline-based silane ligand with [(COD)IrOMe]2 ) was optimal for biphenyl-based phosphines. Selective polyborylation was also shown providing bis- and tris-borylated phosphines. Deprotection of the phosphine boronate provided free ambiphilic phosphine boronates, which do not have detectable interactions between the phosphorus and boron atoms in solution or the solid state.