Neutrophil cathepsin G proteolysis of protease-activated receptor 4 generates a novel, functional tethered ligand.

Platelet-neutrophil interactions regulate ischemic vascular injury. Platelets are activated by serine proteases that cleave protease-activated receptor (PAR) amino termini, resulting in an activating tethered ligand. Neutrophils release cathepsin G (CatG) at sites of injury and inflammation, which activates PAR4 but not PAR1, although the molecular mechanism of CatG-induced PAR4 activation is unknown. We show that blockade of the canonical PAR4 thrombin cleavage site did not alter CatG-induced platelet aggregation, suggesting CatG cleaves a different site than thrombin. Mass spectrometry analysis using PAR4 N-terminus peptides revealed CatG cleavage at Ser67-Arg68. A synthetic peptide, RALLLGWVPTR, representing the tethered ligand resulting from CatG proteolyzed PAR4, induced PAR4-dependent calcium flux and greater platelet aggregation than the thrombin-generated GYPGQV peptide. Mutating PAR4 Ser67or Arg68 reduced CatG-induced calcium flux without affecting thrombin-induced calcium flux. Dog platelets, which contain a conserved CatG PAR4 Ser-Arg cleavage site, aggregated in response to human CatG and RALLLGWVPTR, while mouse (Ser-Gln) and rat (Ser-Glu) platelets were unresponsive. Thus, CatG amputates the PAR4 thrombin cleavage site by cleavage at Ser67-Arg68 and activates PAR4 by generating a new functional tethered ligand. These findings support PAR4 as an important CatG signaling receptor and suggest a novel therapeutic approach for blocking platelet-neutrophil-mediated pathophysiologies.

Transfusion with Cryoprecipitate for Very Low Fibrinogen Levels Does Not Affect Bleeding or Survival in Critically Ill Cirrhosis Patients.

BACKGROUND: Fibrinogen (FIB) levels less than 150 mg/dL have been associated with increased rates of bleeding and lower survival in critically ill cirrhosis patients. OBJECTIVE: We aimed to determine if treatment with cryoprecipitate (CRYO) for low FIB levels is associated with bleeding outcomes or survival. METHODS: A total of 237 cirrhosis patients admitted to an intensive care unit at a tertiary care liver transplant center with initial FIB levels less than 150 mg/dL were retrospectively assessed for CRYO transfusion, bleeding events, and survival outcomes. RESULTS: The mean MELD score was 27.2 (95% confidence interval [CI]: 26.0-28.3) and CLIF-C acute on chronic liver failure score was 53.4 (51.9-54.8). Ninety-nine (41.8%) were admitted for acute bleeding and the remainder were admitted for nonbleeding illnesses. FIB level on admission correlated strongly with disease severity. After adjusting for disease severity, FIB on admission was not an independent predictor of 30-day survival (hazard ratio [HR]: 0.99, 95% CI: 0.99-1.01, p = 0.68). CRYO transfusion increased FIB levels but had no independent effect on mortality or bleeding complications (HR: 1.10, 95% CI: 0.72-1.70, p = 0.65). CONCLUSION: In cirrhosis patients with critical illness, low FIB levels on presentation reflect severity of illness but are not independently associated with 30-day mortality. Treatment of low FIB with CRYO also does not affect survival or bleeding complications, suggesting FIB is an additional marker of severity of illness but is not itself a direct factor in the pathophysiology of bleeding in critically ill cirrhosis patients.

The Effect of Radiation on Quality of Life throughout the Breast Reconstruction Process: A Prospective, Longitudinal Pilot Study of 200 Patients with Long-Term Follow-Up.

BACKGROUND: Despite well-established correlation of postmastectomy radiotherapy and surgical complications in breast reconstruction, its impact on patient reported outcomes is less clear. We sought to determine the effect of postmastectomy radiotherapy on patient reported outcomes throughout the breast reconstruction process. METHODS: Patients undergoing prosthetic and autologous breast reconstruction from November 2010 to June 2013 were prospectively followed with BREAST-Q surveys (preoperatively, after expander placement, and 6 and 12 months after final reconstruction). Paired t test, Wilcoxon rank sum test, and multiple linear regression were used to determine the effect of radiation on patient reported outcomes. RESULTS: Two hundred patients were included in the study, of which 51 (25.5 percent) received postmastectomy radiotherapy. Prosthetic reconstruction was performed in 75 patients (37.5 percent), autologous reconstruction was performed in 118 (59 percent), and pure fat grafting was performed in seven (3.5 percent). At one-year follow-up, the nonirradiated group reported higher BREAST-Q scores when compared with the irradiated group, in Satisfaction with Breasts (p = 0.003), Psychosocial Well-being (p = 0.003), Sexual Well-being (p < 0.001), Physical Well-being of Chest (p = 0.024), and Satisfaction with Outcome (p = 0.03). When accounting for baseline values, Satisfaction with Breasts and Physical Well-being of Chest significantly worsened in irradiated patients undergoing prosthetic reconstruction, an effect not seen with autologous reconstructions. All irradiated patients significantly worsened in Psychosocial Well-being and Sexual Well-being scores. CONCLUSIONS: Postmastectomy radiotherapy is associated with worse patient reported outcomes following breast reconstruction. Autologous reconstruction can mitigate patient dissatisfaction in some domains. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, II.