NOX4 links metabolic regulation in pancreatic cancer to endoplasmic reticulum redox vulnerability and dependence on PRDX4.

There is an urgent need to identify vulnerabilities in pancreatic ductal adenocarcinoma (PDAC). PDAC cells acquire metabolic changes that augment NADPH production and cytosolic redox homeostasis. Here, we show that high NADPH levels drive activity of NADPH oxidase 4 (NOX4) expressed in the endoplasmic reticulum (ER) membrane. NOX4 produces H2O2 metabolized by peroxiredoxin 4 (PRDX4) in the ER lumen. Using functional genomics and subsequent in vitro and in vivo validations, we find that PDAC cell lines with high NADPH levels are dependent on PRDX4 for their growth and survival. PRDX4 addiction is associated with increased reactive oxygen species, a DNA-PKcs-governed DNA damage response and radiosensitivity, which can be rescued by depletion of NOX4 or NADPH. Hence, this study has identified NOX4 as a protein that paradoxically converts the reducing power of the cytosol to an ER-specific oxidative stress vulnerability in PDAC that may be therapeutically exploited by targeting PRDX4.

DEVELOPMENT OF A PORTABLE BLOOD POTASSIUM MONITORING DEVICE FOR DIALYSIS PATIENTS.

Approximately 500,000 dialysis patients in America are at a high risk of hyperkalemia, a condition where blood potassium becomes elevated above normal levels. Hyperkalemia is extremely dangerous, as it can result in severe cardiac complications if untreated. Hyperkalemia may be silent or present vague symptoms until those complications develop, at which point patients require emergency medical care. However, if patients have the ability to measure their potassium levels at home, they could detect hyperkalemia before it reaches a dangerous stage, and seek preventative medical care to avoid severe complications. Therefore, we have designed a novel device allowing patients to measure their blood potassium levels at home. The workflow of our solution is as follows: (1) patients obtain a blood sample from a finger prick, (2) potassium concentration is measured with an ion specific electrode (ISE), and (3) the device displays their potassium levels and a recommended course of action based on their hyperkalemic risk. We validate our solution with three major tests. First, our portable ISE technology must accurately measure potassium concentration in blood samples. Second, appropriate lancet parameters (gauge and depth) to minimize hemolysis in capillary blood samples must be found to minimize falsely elevated readings. Third, device portability and ease of use must be evaluated using patient input, as these factors will affect patient compliance. We have validated the use of portable ISE technology to feasibly measure potassium, and we continue to collect data for our second and third tests.

Comparison of Tri-folded and Scroll-based Graft Viability in Preloaded Descemet Membrane Endothelial Keratoplasty.

PURPOSE: To compare corneal endothelial damage associated with 2 techniques for preloaded Descemet membrane endothelial keratoplasty (DMEK): a tri-folded graft stored in a plastic cartridge designed for DMEK and a scrolled graft stored in a modified Jones Tube, at the time of preparation and after shipping. METHODS: DMEK grafts were prepared at the Rocky Mountain Lions Eye Bank. The grafts were either tri-folded and loaded in a plastic cartridge or scrolled and loaded into a modified Jones Tube. In each group, the grafts were then either immediately removed from the cartridges or shipped for 48 hours. The grafts were then stained with Calcein AM and imaged using a fluorescent microscope. Endothelial cell loss (ECL) was determined using trainable segmentation in Fiji by 2 graders. At each time point, rates of ECL loss were compared across the 2 groups. To explore the role of donor characteristics, a multivariable regression model was produced to account for method (tri-folding vs. scroll), donor age, donor gender, death-to-preservation time, death-to-preparation time, and shipping. RESULTS: A total of 40 grafts were prepared, processed, imaged, and analyzed. No significant difference in cell loss was seen between groups at either time point alone. In the multivariate model, no significant increase in cell loss was associated with either tri-folding (3.7% less ECL; P = 0.051) or shipping (4.3% less ECL; P = 0.049). CONCLUSIONS: All techniques used resulted in clinically acceptable levels of ECL. Tri-folded tissue in a plastic cartridge did not result in ECL inferior to a scroll when prepared either immediately or preloaded for 48 hours.

Viability of Descemet Membrane Endothelial Keratoplasty Grafts Folded in the Eye Bank.

PURPOSE: Preloaded, trifolded grafts in Descemet membrane endothelial keratoplasty require transfer of the trifolding process from the corneal transplant surgeon to the eye bank technician. We sought to assess whether trifolding may be safely conducted by an eye bank technician with cell loss comparable to standard peeling and lifting. METHODS: A total of 10 grafts were stained, peeled, and transferred directly onto a bed of Calcein-AM and Amvisc Plus by an eye bank technician. Five grafts were removed and stained as a scroll, and 5 grafts were trifolded with the endothelium in before transfer. Photographs were acquired with an inverted fluorescence microscope, and image segmentation was performed. A t test was conducted to compare differences in endothelial cell loss across groups. RESULTS: Mean cell loss in the scroll group was 18.5% [95% confidence interval (CI): 15.2%-21.9%] compared with 7.6% of the trifolded group (95% CI: 1.7%-13.5%). A 2-tailed t test indicated decreased cell loss in the trifolded group (P = 0.013). CONCLUSIONS: Despite additional manipulation of the graft, trifolding of Descemet membrane and endothelium may be performed by an eye bank technician without significantly increased cell loss relative to graft preparation as a scroll.