Tumor necrosis factor-␣-induced TRPC1 expression amplifies store-operated Ca 2ϩ influx and endothelial permeability

influx in response to thrombin exposure. We observed that thrombininduced Ca 2ϩ influx in TNF-␣-stimulated HPAEC was twofold greater than in control cells. To address the relationship between store-operated Ca 2ϩ influx and TRPC1 expression, we overexpressed TRPC1 by three-to fourfold in the human dermal microvascular endothelial cell line (HMEC) using the TRPC1 cDNA. Thrombininduced store Ca 2ϩ depletion in these cells caused approximately twofold greater increase in Ca 2ϩ influx than in control cells. Furthermore, the inositol 1,4,5-trisphosphate-sensitive store-operated cationic current was increased greater than twofold in TRPC1-transfected cells compared with control. To address the role of Ca 2ϩ influx via TRPC1 in signaling endothelial permeability, we measured actinstress fiber formation and transendothelial monolayer electrical resistance (TER) in the TRPC1 cDNA-transfected HMEC and TNF-␣-challenged HPAEC. Both thrombin-induced actin-stress fiber formation and a decrease in TER were augmented in TRPC1-overexpressing HMEC compared with control cells. TNF-␣-induced increased TRPC1 expression in HPAEC also resulted in marked endothelial barrier dysfunction in response to thrombin. These findings indicate the expression level of TRPC1 in endothelial cells is a critical determinant of Ca 2ϩ influx and signaling of the increase in endothelial permeability. tumor necrosis factor-␣; store-operated calcium ion influx; transient receptor potential channel 1; endothelial barrier dysfunction MICROVASCULAR ENDOTHELIAL cells regulate the vessel wall permeability of solutes, liquid, and macromolecules and produce numerous autocrine and paracrine factors, such as NO, that modulate the contractility of the underlying vascular smooth muscl

Immunologic Recognition of a Shared p53 Mutated Neoantigen in a Patient with Metastatic Colorectal Cancer.

Adoptive cell therapy (ACT) with T cells targeting neoantigens can mediate durable responses in patients with metastatic cancer. Cell therapies targeting common shared antigens for epithelial cancers are not yet broadly available. Here, we report the identification and characterization in one patient of T-cell receptors (TCRs) recognizing mutated p53 p.R175H, which is shared among a subset of patients with cancer. Tumor-infiltrating lymphocytes were screened for recognition of mutated neoantigens in a patient with metastatic colorectal cancer. HLA-A*0201-restricted recognition of mutated p53 p.R175H was identified, and the minimal peptide epitope was HMTEVV