S100A6 is a positive regulator of PPP5C-FKBP51-dependent regulation of endothelial calcium signaling.

ISOC is a cation current permeating the ISOC channel. In pulmonary endothelial cells, ISOC activation leads to formation of inter-endothelial cell gaps and barrier disruption. The immunophilin FK506-binding protein 51 (FKBP51), in conjunction with the serine/threonine protein phosphatase 5C (PPP5C), inhibits ISOC . Free PPP5C assumes an autoinhibitory state, which has low "basal" catalytic activity. Several S100 protein family members bind PPP5C increasing PPP5C catalytic activity in vitro. One of these family members, S100A6, exhibits a calcium-dependent translocation to the plasma membrane. The goal of this study was to determine whether S100A6 activates PPP5C in pulmonary endothelial cells and contributes to ISOC inhibition by the PPP5C-FKBP51 axis. We observed that S100A6 activates PPP5C to dephosphorylate tau T231. Following ISOC activation, cytosolic S100A6 translocates to the plasma membrane and interacts with the TRPC4 subunit of the ISOC channel. Global calcium entry and ISOC are decreased by S100A6 in a PPP5C-dependent manner and by FKBP51 in a S100A6-dependent manner. Further, calcium entry-induced endothelial barrier disruption is decreased by S100A6 dependent upon PPP5C, and by FKBP51 dependent upon S100A6. Overall, these data reveal that S100A6 plays a key role in the PPP5C-FKBP51 axis to inhibit ISOC and protect the endothelial barrier against calcium entry-induced disruption.

Assessing interest in cardiothoracic surgery at an osteopathic medical school: Results of an institutional survey.

Objective: Cardiothoracic surgery is a surgical subspecialty that attracts few medical students. As integrated surgical residency programs continue to grow in number, there is increased interest in what factors influence specialty selection during undergraduate medical education. Previous institutional studies have studied allopathic medical schools affiliated with academic institutions. This study aimed to assess the interest and perception of cardiothoracic surgery at an osteopathic institution. Methods: Active medical students at a US osteopathic institution were invited to complete an original online survey. Means and 95% confidence intervals were calculated and graphed for questions using Likert scale responses. Comparison of mean responses for preclinical versus clinical students was assessed by a Kruskal-Wallis nonparametric analysis of variance. Results: There were 166 surveys (22%) completed, and interest in cardiothoracic surgery was indicated by 7.8% of respondents. Work/life balance, personality of cardiothoracic surgeons, and lack of family time were negative factors associated with cardiothoracic surgery. Clinical exposure, shadowing, mentorship, and significant personal/life events before medical school were strong factors in establishing students' interest in cardiothoracic surgery. Preclinical students noted exposure to cardiothoracic surgery would further increase their interest when compared with clinical students (µ = 3.39 vs µ = 2.69, P = .008). Conclusions: All factors that established interest in cardiothoracic surgery occurred before students entered medical school. Although there are negative perceptions associated with cardiothoracic surgery, these may be ameliorated with more exposure to the field. Further research is needed to explore how early exposure in preclinical years of medical school affects students' perceptions and ultimate interest in cardiothoracic surgery.

Building Interest in Cardiothoracic Surgery at an Osteopathic Medical School: Results of an Institutional Study and a Guide for Medical Schools.

Objective A previous study at this institution revealed a connection between interest group involvement and specialty interest while identifying the negative perceptions of cardiothoracic (CT) surgery. This study aimed to build interest and ameliorate the negative perceptions of CT surgery by exposing pre-clinical students to the field through engaging events. Methods Students at a US osteopathic institution who attended CT surgery committee events were invited to complete an online survey after each event. Associations between the number of events attended and ranked responses to survey questions were assessed by two-tailed Spearman correlations. Statistical comparisons in ranked responses between the events attended and the survey questions were assessed by a two-way analysis of variance (ANOVA). Pre-clinical students actively enrolled at the institution during the 2022-2023 academic year were eligible for inclusion. Results There were 83 surveys completed over seven events. There was a significant association between the number of events a student attended and their perception of CT surgeon's work/life balance with a correlation coefficient of .258 (P=0.019) and whether CT surgeons have time for their families with a correlation coefficient of .235 (P=0.035). Residents and medical student events as well as wet lab events increased interest the most and helped students feel equipped to apply for CT surgery. Conclusions While negative perceptions associated with CT surgery exist, these may be ameliorated with more exposure to the field. Unique events that expose pre-clinical students to multiple facets of CT surgery, including physicians and trainees in the field, as well as offering hands-on activities, may increase interest in the field and further pursuit of the field during clinical years.

Serine/threonine phosphatase 5 (PP5C/PPP5C) regulates the ISOC channel through a PP5C-FKBP51 axis.

Pulmonary endothelial cells express a store-operated calcium entry current ( Isoc), which contributes to inter-endothelial cell gap formation. Isoc is regulated by a heterocomplex of proteins that includes the immunophilin FKBP51. FKBP51 inhibits Isoc by mechanisms that are not fully understood. In pulmonary artery endothelial cells (PAECs) we have shown that FKBP51 increases microtubule polymerization, an event that is critical for Isoc inhibition by FKBP51. In neurons, FKBP51 promotes microtubule stability through facilitation of tau dephosphorylation. However, FKBP51 does not possess phosphatase activity. Protein phosphatase 5 (PP5C/PPP5C) can dephosphorylate tau, and similar to FKBP51, PP5C possesses tetratricopeptide repeats (TPR) that mediate interaction with heat shock protein-90 (HSP90) chaperone/scaffolding complexes. We therefore tested whether PP5C contributes to FKBP51-mediated inhibition of Isoc. Both siRNA-mediated suppression of PP5C expression in PAECs and genetic disruption of PP5C in HEK293 cells attenuate FKBP51-mediated inhibition of Isoc. Reintroduction of catalytically competent, but not catalytically inactive PP5C, restored FKBP51-mediated inhibition of Isoc. PAEC cell fractionation studies identified both PP5C and the ISOC heterocomplex in the same membrane fractions. Further, PP5C co-precipitates with TRPC4, an essential subunit of ISOC channel. Finally, to determine if PP5C is required for FKBP51-mediated inhibition of calcium entry-induced inter-endothelial cell gap formation, we measured gap area by wide-field microscopy and performed biotin gap quantification assay and electric cell-substrate impedance sensing (ECIS®). Collectively, the data presented indicate that suppression of PP5C expression negates the protective effect of FKBP51. These observations identify PP5C as a novel member of the ISOC heterocomplex that is required for FKBP51-mediated inhibition of Isoc.

Protective role of FKBP51 in calcium entry-induced endothelial barrier disruption.

Pulmonary artery endothelial cells (PAECs) express a cation current, ISOC (store-operated calcium entry current), which when activated permits calcium entry leading to inter-endothelial cell gap formation. The large molecular weight immunophilin FKBP51 inhibits ISOC but not other calcium entry pathways in PAECs. However, it is unknown whether FKBP51-mediated inhibition of ISOC is sufficient to protect the endothelial barrier from calcium entry-induced disruption. The major objective of this study was to determine whether FKBP51-mediated inhibition of ISOC leads to decreased calcium entry-induced inter-endothelial gap formation and thus preservation of the endothelial barrier. Here, we measured the effects of thapsigargin-induced ISOC on the endothelial barrier in control and FKBP51 overexpressing PAECs. FKBP51 overexpression decreased actin stress fiber and inter-endothelial cell gap formation in addition to attenuating the decrease in resistance observed with control cells using electric cell-substrate impedance sensing. Finally, the thapsigargin-induced increase in dextran flux was abolished in FKBP51 overexpressing PAECs. We then measured endothelial permeability in perfused lungs of FKBP51 knockout (FKBP51-/-) mice and observed increased calcium entry-induced permeability compared to wild-type mice. To begin to dissect the mechanism underlying the FKBP51-mediated inhibition of ISOC, a second goal of this study was to determine the role of the microtubule network. We observed that FKBP51 overexpressing PAECs exhibited increased microtubule polymerization that is critical for inhibition of ISOC by FKBP51. Overall, we have identified FKBP51 as a novel regulator of endothelial barrier integrity, and these findings are significant as they reveal a protective mechanism for endothelium against calcium entry-induced disruption.