Membrane trafficking from lysosomes to the plasma membrane regulates phosphatidylserine externalization during apoptosis

Programmed cell death is characterized by tightly controlled temporal and spatial intracellular Ca2+ responses that regulate the release of key proapoptotic proteins from mitochondria to the cytosol. Since apoptotic cells retain their ability to exclude membrane impermeable dyes, it is possible that the cells evoke repair mechanisms that, similar to those in normal cells, patch any damaged areas of the plasma membrane that preclude dye permeation. One critical distinction between plasma membrane repair in normal and apoptotic cells is the preservation of membrane lipid asymmetry. In normal cells, phosphatidylserine (PS) retains its normal asymmetric distribution in the inner membrane leaflet. In apoptotic cells, PS redistributes to the outer membrane leaflet by a Ca2+ dependent mechanism where it serves as a recognition ligand for phagocytes(1). In this study Ca 2+-specific fluorescent probes were employed to investigate the source of Ca2+ required for PS externalization. Experiments employing Rhod2-AM, calcium green 1, fura2-AM and the aqueous space marker FITC-dextran, demonstrated that exogenous Ca2+ imported with endocytotic vesicles into the cell was released into the cytosol in an apoptosis dependent manner. Labeling of the luminal side of the endocytotic vesicles with FITC-annexin 5, revealed that membrane lipid asymmetry was disrupted upon endosome formation. Specific labeling of the lysosomal luminal surface with the non-exchangeable membrane lipid probe, N-rhodamine-labeled-phosphatidylethanolamine (N-Rho-PE) and the lysosomal specific probe, lysotracker green, facilitated real-time monitoring of plasma membrane-to-endosome-to-lysosome transitions. Enforced elevation of cytosolic [Ca2+] with ionophore resulted in the redistribution of N-Rho-PE and PS from the inner membrane leaflet to the PM outer membrane leaflet. Identical results were obtained during apoptosis, however, the redistribution of both N-RhoPE and PS was dependent on the release of intra-lysosomal Ca2+ to the cytosol. Additional experiments suggested that lipid redistribution was dependent on the activity of lysosomal phospholipase A2 activity since lipid trafficking was abolished in the presence of chloroquine and lipase inhibitors. These data indicate that endosomal/lysosomal Ca2+ and the fusion of hybrid organelles to the plasma membrane regulates the externalization of PS during apoptosis. ^

A Time-Motion Study of Registered Nurses’ Workflow in Intensive Care Unit Remote Monitoring

Utilizing advanced information technology, Intensive Care Unit (ICU) remote monitoring allows highly trained specialists to oversee a large number of patients at multiple sites on a continuous basis. In the current research, we conducted a time-motion study of registered nurses’ work in an ICU remote monitoring facility. Data were collected on seven nurses through 40 hours of observation. The results showed that nurses’ essential tasks were centered on three themes: monitoring patients, maintaining patients’ health records, and managing technology use. In monitoring patients, nurses spent 52% of the time assimilating information embedded in a clinical information system and 15% on monitoring live vitals. System-generated alerts frequently interrupted nurses in their task performance and redirected them to manage suddenly appearing events. These findings provide insight into nurses’ workflow in a new, technology-driven critical care setting and have important implications for system design, work engineering, and personnel selection and training.