Workforce, Workload, and Burnout Among Intensivists and Advanced Practice Providers: A Narrative Review.

OBJECTIVES: To assess-by literature review and expert consensus-workforce, workload, and burnout considerations among intensivists and advanced practice providers. DESIGN: Data were synthesized from monthly expert consensus and literature review. SETTING: Workforce and Workload section workgroup of the Academic Leaders in Critical Care Medicine Task Force. MEASUREMENTS AND MAIN RESULTS: Multidisciplinary care teams led by intensivists are an essential component of critical care delivery. Advanced practice providers (nurse practitioners and physician assistants) are progressively being integrated into ICU practice models. The ever-increasing number of patients with complex, life-threatening diseases, concentration of ICU beds in few centralized hospitals, expansion of specialty ICU services, and desire for 24/7 availability have contributed to growing intensivist staffing concerns. Such staffing challenges may negatively impact practitioner wellness, team perception of care quality, time available for teaching, and length of stay when the patient to intensivist ratio is greater than or equal to 15. Enhanced team communication and reduction of practice variation are important factors for improved patient outcomes. A diverse workforce adds value and enrichment to the overall work environment. Formal succession planning for ICU leaders is crucial to the success of critical care organizations. Implementation of a continuous 24/7 ICU coverage care model in high-acuity, high-volume centers should be based on patient-centered outcomes. High levels of burnout syndrome are common among intensivists. Prospective analyses of interventions to decrease burnout within the ICU setting are limited. However, organizational interventions are felt to be more effective than those directed at individuals. CONCLUSIONS: Critical care workforce and staffing models are myriad and based on several factors including local culture and resources, ICU organization, and strategies to reduce burden on the ICU provider workforce. Prospective studies to assess and avoid the burnout syndrome among intensivists and advanced practice providers are needed.

Critical Care Organizations: Business of Critical Care and Value/Performance Building.

OBJECTIVE: New, value-based regulations and reimbursement structures are creating historic care management challenges, thinning the margins and threatening the viability of hospitals and health systems. The Society of Critical Care Medicine convened a taskforce of Academic Leaders in Critical Care Medicine on February 22, 2016, during the 45th Critical Care Congress to develop a toolkit drawing on the experience of successful leaders of critical care organizations in North America for advancing critical care organizations (Appendix 1). The goal of this article was to provide a roadmap and call attention to key factors that adult critical care medicine leadership in both academic and nonacademic setting should consider when planning for value-based care. DESIGN: Relevant medical literature was accessed through a literature search. Material published by federal health agencies and other specialty organizations was also reviewed. Collaboratively and iteratively, taskforce members corresponded by electronic mail and held monthly conference calls to finalize this report. SETTING: The business and value/performance critical care organization building section comprised of leaders of critical care organizations with expertise in critical care administration, healthcare management, and clinical practice. MEASUREMENTS AND MAIN RESULTS: Two phases of critical care organizations care integration are described: "horizontal," within the system and regionalization of care as an initial phase, and "vertical," with a post-ICU and postacute care continuum as a succeeding phase. The tools required for the clinical and financial transformation are provided, including the essential prerequisites of forming a critical care organization; the manner in which a critical care organization can help manage transformational domains is considered. Lastly, how to achieve organizational health system support for critical care organization implementation is discussed. CONCLUSIONS: A critical care organization that incorporates functional clinical horizontal and vertical integration for ICU patients and survivors, aligns strategy and operations with those of the parent health system, and encompasses knowledge on finance and risk will be better positioned to succeed in the value-based world.

Using Incentives to Improve Resource Utilization: A Quasi-Experimental Evaluation of an ICU Quality Improvement Program.

OBJECTIVES: Healthcare systems strive to provide quality care at lower cost. Arterial blood gas testing, chest radiographs, and RBC transfusions provide an important example of opportunities to reduce excess resource utilization within the ICU. We describe the effect of a multifaceted quality improvement program designed to decrease the avoidable arterial blood gases, chest radiographs, and RBC utilization on utilization of these resources and patient outcomes. DESIGN: Prospective pre-post cohort study. SETTING: Seven ICUs in an academic healthcare system. PATIENTS: All adult ICU patients admitted to study ICUs during consecutive baseline (n = 7,357), intervention (n = 7,553), and follow-up (n = 7,657) years between September 2010 and August 2013. INTERVENTIONS: A multifaceted quality improvement program including provider education, audit and feedback, and unit-based provider financial incentives targeting arterial blood gas, chest radiograph, and RBC utilization. MEASUREMENTS AND MAIN RESULTS: The primary outcome was the number of orders for arterial blood gases, chest radiographs, and RBCs per patient. Compared with the baseline period, unadjusted arterial blood gas, chest radiograph, and RBC utilization in the intervention period was reduced by 42%, 26%, and 17%, respectively (p < 0.01). After adjusting for potentially relevant patient factors, the intervention was associated with 128 fewer arterial blood gases, 73 fewer chest radiographs, and 16 fewer RBCs per 100 patients (p < 0.01). This effect was durable during the follow-up year. This reduction yielded an approximate net savings of $1.5 M in direct costs over the intervention and follow-up years after accounting for the direct costs of the program. Unadjusted hospital mortality decreased from 7% in the baseline period to 5.2% in the intervention period (p < 0.01). This reduction remained significant after adjusting for patient factors (odds ratio = 0.43; p < 0.01). CONCLUSIONS: Implementation of a multifaceted quality improvement program including financial incentives was associated with significant improvements in resource utilization. Our findings provide evidence supporting the safety, effectiveness, and sustainability of incentive-based quality improvement interventions.

Prevalence and Impact of Unknown Diabetes in the ICU.

OBJECTIVES: Many patients with diabetes and their care providers are unaware of the presence of the disease. Dysglycemia encompassing hyperglycemia, hypoglycemia, and glucose variability is common in the ICU in patients with and without diabetes. The purpose of this study was to determine the impact of unknown diabetes on glycemic control in the ICU. DESIGN: Prospective observational study. SETTING: Nine ICUs in an academic, tertiary hospital and a hybrid academic/community hospital. PATIENTS: Hemoglobin A1c levels were ordered at all ICU admissions from March 1, 2011 to September 30, 2013. Electronic medical records were examined for a history of antihyperglycemic medications or International Classification of Diseases, 9th Edition diagnosis of diabetes. Patients were categorized as having unknown diabetes (hemoglobin A1c > 6.5%, without history of diabetes), no diabetes (hemoglobin A1c < 6.5%, without history of diabetes), controlled known diabetes (hemoglobin A1c < 6.5%, with documented history of diabetes), and uncontrolled known diabetes (hemoglobin A1c > 6.5%, with documented history of diabetes). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: A total of 15,737 patients had an hemoglobin A1c and medical record evaluable for the history of diabetes, and 5,635 patients had diabetes diagnosed by either medical history or an elevated hemoglobin A1c in the ICU. Of these, 1,460 patients had unknown diabetes, accounting for 26.0% of all patients with diabetes. This represented 41.0% of patients with an hemoglobin A1c > 6.5% and 9.3% of all ICU patients. Compared with patients without diabetes, patients with unknown diabetes had a higher likelihood of requiring an insulin infusion (44.3% vs 29.3%; p < 0.0001), a higher average blood glucose (172 vs 126 mg/dL; p < 0.0001), an increased percentage of hyperglycemia (19.7% vs 7.0%; blood glucose > 180 mg/dL; p < 0.0001) and hypoglycemia (8.9% vs 2.5%; blood glucose < 70 mg/dL; p < 0.0001), higher glycemic variability (55.6 vs 28.8, average of patient SD of glucose; p < 0.0001), and increased mortality (13.8% vs 11.4%; p = 0.01). CONCLUSIONS: Patients with unknown diabetes represent a significant percentage of ICU admissions. Measurement of hemoglobin A1c at admission can prospectively identify a population that are not known to have diabetes but have significant challenges in glycemic control in the ICU.

Reduced red blood cell transfusion in cardiothoracic surgery after implementation of a novel clinical decision support tool.

BACKGROUND: Packed red blood cell (PRBC) transfusion can increase short- and long-term adverse outcomes and health care costs. We compared the transfusion practices in cardiothoracic surgery before and after implementation of a novel clinical decision support (CDS) tool. STUDY DESIGN: The transfusion CDS tool was implemented within computerized provider order entry of a multi-institutional urban hospital system in September 2012. Data were queried for 12 months pre-intervention and for another 12 months post-intervention to compare transfusion practices for all adult patients having isolated coronary artery bypass grafting (CABG) or isolated surgical aortic valve replacement (SAVR). RESULTS: The total number of patients undergoing either isolated CABG or isolated SAVR was 744 pre-intervention and 765 post-intervention (p = 0.84). There was no significant difference in age (64 ± 11.4 years vs 64.5 ± 11.2 years, p = 0.37) or sex (30.2% vs 32.2% female, p = 0.42) between the 2 groups. The number of postoperative transfusions (374 [50.3%] vs 312 [40.8%], p < 0.001), postoperative PRBC units given (1.59 ± 2.9 vs 1.25 ± 2.5, p = 0.01), pre-transfusion hemoglobin level (8.09 ± 1.5 g/dL vs 7.65 ± 1.4 g/dL, p < 0.001), and incidence of surgical site infection (3.1% vs 1.1%; p = 0.005) were significantly reduced after implementation of the transfusion CDS tool. There were no significant differences in intraoperative transfusions (206 [27.7%] vs 180 [23.5%], p = 0.06), intraoperative PRBC units given (0.73 ± 1.5 vs 0.65 ± 1.4, p = 0.28), ICU length of stay (3.29 ± 3.9 days vs 3.37 ± 4.8 days, p = 0.74), or in-hospital mortality (1.3% vs 1.4%, p = 0.87). CONCLUSIONS: Implementation of a transfusion CDS tool was associated with lower pre-transfusion hemoglobin levels, fewer transfusions, decreased infection rates, and decreased health care costs, without an increase in short-term mortality.

Prescribing patterns of hydrocortisone in septic shock: a single-center experience of how surviving sepsis guidelines are interpreted and translated into bedside practice.

OBJECTIVES: The Surviving Sepsis Campaign suggests giving hydrocortisone to septic patients only if their "blood pressure is poorly responsive to fluid resuscitation and vasopressor therapy." Because the definition of "poorly responsive" is not provided, the purpose of this study was to identify prescribing triggers for hydrocortisone in septic shock. DESIGN: Retrospective chart review of patients with septic shock over 17 months, who received hydrocortisone, followed by a survey of all intensivists who attended in the study ICUs to determine whether provider attitudes matched clinical practice. SETTING: Eight ICUs in an academic hospital and a hybrid academic/community hospital. PATIENTS: A total of 155 patients with septic shock in whom vasopressors were initiated and hydrocortisone was prescribed. MEASUREMENTS AND MAIN RESULTS: Ninety-nine patients (64%) were already receiving two vasopressors before hydrocortisone was prescribed. An additional 22 patients were on a single high-dose vasopressor prior to corticosteroid initiation. Of patients who survived to have their hydrocortisone dose changed, 57% had their corticosteroids tapered, whereas 43% were abruptly discontinued. Seventy-six percent of patients were no longer on vasopressors when the first dosing change was made. Twenty-seven out of 36 intensivists (75%) completed the survey. The majority (72%) defined "poorly responsive to vasopressors" as the presence of two vasopressors, and 70% stated that they required patients to be off vasopressors prior to altering the corticosteroid dose. CONCLUSIONS: Significant variability exists when corticosteroids are prescribed for septic shock, with the most common interpretation in our institution of "poorly responsive to fluid resuscitation and vasopressor therapy" being the presence of two vasopressors. The method and timing of corticosteroid discontinuation also differed among providers. Self-described prescribing patterns from intensivists closely matched their actual behavior, suggesting variability is due to differing interpretations of the guidelines themselves, rather than a deficit in knowledge translation.

Patient-care time allocation by nurse practitioners and physician assistants in the intensive care unit.

INTRODUCTION: Use of nurse practitioners and physician assistants ("affiliates") is increasing significantly in the intensive care unit (ICU). Despite this, few data exist on how affiliates allocate their time in the ICU. The purpose of this study was to understand the allocation of affiliate time into patient-care and non-patient-care activity, further dividing the time devoted to patient care into billable service and equally important but nonbillable care. METHODS: We conducted a quasi experimental study in seven ICUs in an academic hospital and a hybrid academic/community hospital. After a period of self-reporting, a one-time monetary incentive of $2,500 was offered to 39 affiliates in each ICU in which every affiliate documented greater than 75% of their time devoted to patient care over a 6-month period in an effort to understand how affiliates allocated their time throughout a shift. Documentation included billable time (critical care, evaluation and management, procedures) and a new category ("zero charge time"), which facilitated record keeping of other patient-care activities. RESULTS: At baseline, no ICUs had documentation of 75% patient-care time by all of its affiliates. In the 6 months in which reporting was tied to a group incentive, six of seven ICUs had every affiliate document greater than 75% of their time. Individual time documentation increased from 53% to 84%. Zero-charge time accounted for an average of 21% of each shift. The most common reason was rounding, which accounted for nearly half of all zero-charge time. Sign out, chart review, and teaching were the next most common zero-charge activities. Documentation of time spent on billable activities also increased from 53% of an affiliate's shift to 63%. Time documentation was similar regardless of during which shift an affiliate worked. CONCLUSIONS: Approximately two thirds of an affiliate's shift is spent providing billable services to patients. Greater than 20% of each shift is spent providing equally important but not reimbursable patient care. Understanding how affiliates spend their time and what proportion of time is spent in billable activities can be used to plan the financial impact of staffing ICUs with affiliates.

Patterns and dynamics of subventricular zone neuroblast migration in the ischemic striatum of the adult mouse.

The migratory behavior of neuroblasts after a stroke is poorly understood. Using time-lapse microscopy, we imaged migration of neuroblasts and cerebral vessels in living brain slices of adult doublecortin (DCX, a marker of neuroblasts) enhanced green fluorescent protein (eGFP) transgenic mice that were subjected to 7 days of stroke. Our results show that neuroblasts originating in the subventricular zone (SVZ) of adult mouse brain laterally migrated in chains or individually to reach the ischemic striatum. The chains were initially formed at the border between the SVZ and the striatum by neuroblasts in the SVZ and then extended to the striatum. The average speed of DCX-eGFP-expressing cells within chains was 28.67+/-1.04 microm/h, which was significantly faster (P<0.01) than the speed of the cells in the SVZ (17.98+/-0.57 microm/h). Within the ischemic striatum, individual neuroblasts actively extended or retracted their processes, suggestive of probing the immediate microenvironment. The neuroblasts close to cerebral blood vessels exhibited multiple processes. Our data suggest that neuroblasts actively interact with the microenvironment to reach the ischemic striatum by multiple migratory routes.

Gene profiles and electrophysiology of doublecortin-expressing cells in the subventricular zone after ischemic stroke.

Stroke increases neuroblasts in the subventricular zone (SVZ) of the lateral ventricle and these neuroblasts migrate toward the ischemic boundary to replace damaged neurons. Using brain slices from the nonischemic adult rat and transgenic mice that expressed enhanced green fluorescent protein (EGFP) concomitantly with doublecortin (DCX), a marker for migrating neuroblasts, we recorded electrophysiological characteristics while simultaneously analyzing the gene expression in single SVZ cells. We found that SVZ cells expressing the DCX gene from the nonischemic rat had a mean resting membrane potential (RMP) of -30 mV. DCX-EGFP-positive cells in the nonischemic SVZ of the transgenic mouse had a mean RMP of -25+/-7 mV and did not exhibit Na(+) currents, characteristic of immature neurons. However, DCX-EGFP-positive cells in the ischemic SVZ exhibited a hyperpolarized mean RMP of -54+/-18 mV and displayed Na(+) currents, indicative of more mature neurons. Single-cell multiplex RT-PCR analysis revealed that DCX-EGFP-positive cells in the nonischemic SVZ of the transgenic mouse expressed high neural progenitor marker genes, Sox2 and nestin, but not mature neuronal marker genes. In contrast, DCX-EGFP-positive cells in the ischemic SVZ expressed tyrosine hydroxylase, a mature neuronal marker gene. Together, these data indicate that stroke changes gene profiles and the electrophysiology of migrating neuroblasts.

Neural progenitor cells treated with EPO induce angiogenesis through the production of VEGF.

Recombinant human erythropoietin (rhEPO) induces neurogenesis and angiogenesis. Using a coculture system of mouse brain endothelial cells (MBECs) and neural progenitor cells derived from the subventricular zone of adult mouse, we investigated the hypothesis that neural progenitor cells treated with rhEPO promote angiogenesis. Treatment of neural progenitor cells with rhEPO significantly increased their expression and secretion of vascular endothelial growth factor (VEGF) and activated phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and extracellular signal-regulated kinase (ERK1/2). Selective inhibition of the Akt and ERK1/2 signaling pathways significantly attenuated the rhEPO-induced VEGF expression in neural progenitor cells. The supernatant harvested from neural progenitor cells treated with rhEPO significantly increased the capillary-like tube formation of MBECs. SU1498, a specific VEGF type-2 receptor (VEGFR2) antagonist, abolished the supernatant-enhanced angiogenesis. In addition, coculture of MBECs with neural progenitor cells treated with rhEPO substantially increased VEGFR2 mRNA and protein levels in MBECs. These in vitro results suggest that EPO enhances VEGF secretion in neural progenitor cells through activation of the PI3K/Akt and ERK1/2 signaling pathways and that neural progenitor cells treated with rhEPO upregulate VEGFR2 expression in cerebral endothelial cells, which along with VEGF secreted by neural progenitor cells promotes angiogenesis.

Coupling of angiogenesis and neurogenesis in cultured endothelial cells and neural progenitor cells after stroke.

Angiogenesis and neurogenesis are coupled processes. Using a coculture system, we tested the hypothesis that cerebral endothelial cells activated by ischemia enhance neural progenitor cell proliferation and differentiation, while neural progenitor cells isolated from the ischemic subventricular zone promote angiogenesis. Coculture of neural progenitor cells isolated from the subventricular zone of the adult normal rat with cerebral endothelial cells isolated from the stroke boundary substantially increased neural progenitor cell proliferation and neuronal differentiation and reduced astrocytic differentiation. Conditioned medium harvested from the stroke neural progenitor cells promoted capillary tube formation of normal cerebral endothelial cells. Blockage of vascular endothelial growth factor receptor 2 suppressed the effect of the endothelial cells activated by stroke on neurogenesis as well as the effect of the supernatant obtained from stroke neural progenitor cells on angiogenesis. These data suggest that angiogenesis couples to neurogenesis after stroke and vascular endothelial growth factor likely mediates this coupling.

The Sonic hedgehog pathway mediates carbamylated erythropoietin-enhanced proliferation and differentiation of adult neural progenitor cells.

Carbamylated erythropoietin (CEPO), a well characterized erythropoietin (EPO) derivative, does not bind to the classical EPO receptor and does not stimulate erythropoiesis. Using neural progenitor cells derived from the subventricular zone of the adult mouse, we investigated the effect of CEPO on neurogenesis and the associated signaling pathways in vitro. We found that CEPO significantly increased neural progenitor cell proliferation and promoted neural progenitor cell differentiation into neurons, which was associated with up-regulation of Sonic hedgehog (Shh), its receptor ptc, and mammalian achaete-scute homolog 1 (Mash1), a pro-neuron basic helix-loop-helix protein transcription factor. Blockage of the Shh signaling pathway with a pharmacological inhibitor, cyclopamine, abolished the CEPO-induced neurogenesis. Attenuation of endogenous Mash1 expression by short-interfering RNA blocked CEPO-promoted neuronal differentiation. In addition, recombinant mouse Shh up-regulated Mash1 expression in neural progenitor cells. These results demonstrate that the Shh signaling pathway mediates CEPO-enhanced neurogenesis and Mash1 is a downstream target of the Shh signaling pathway that regulates CEPO-enhanced neuronal differentiation.

Chemokine ligand 2 (CCL2) induces migration and differentiation of subventricular zone cells after stroke.

Ischemic stroke stimulates neurogenesis in the adult rodent brain. The molecules that mediate stroke-induced neurogenesis have not been fully investigated. Using a microarray containing 113 known genes associated with angiogenesis, we analyzed transcriptional profiles in subventricular zone (SVZ) tissue and in cultured neural progenitor cells isolated from the SVZ of adult mice subjected to middle cerebral artery occlusion (MCAo). Among the genes most robustly up-regulated by MCAo were chemokine ligand 2 (CCL2) and chemokine ligand 10 (CXCL10). Consistent with the mRNA data, immunofluorescent staining revealed that MCAo substantially increased the number of CCL2-positive cells in the ipsilateral SVZ and that CCL2-positive cells were positive for both glial fibrillary acidic protein (GFAP) and nestin. In vitro studies showed that incubation of neural progenitor cells with recombinant human CCL2 substantially increased the number of Tuj1-positive cells dose dependently compared with the number in the control group, indicating that CCL2 promotes neuronal differentiation. Blockage of CCL2 with a neutralized antibody against CCL2 abolished the effects of CCL2 on neural progenitor cell migration and differentiation. Treatment of neural progenitor cells with CCL2 did not alter the number of BrdU cells and the number of apoptotic cells compared with those in the control group, suggesting that CCL2 does not affect neural progenitor cell proliferation and cell survival. These data demonstrate that in addition to its role in cell motility, CCL2 plays an important role in neuronal differentiation.

Neuroblast division during migration toward the ischemic striatum: a study of dynamic migratory and proliferative characteristics of neuroblasts from the subventricular zone.

Ischemic stroke induces neurogenesis in the subventricular zone (SVZ), and newly generated neurons in the SVZ migrate toward the ischemic boundary. However, the characteristics of migrating SVZ cells have not been investigated after stroke. Using time-lapse imaging in both SVZ cells and organotypic brain slice cultures, we measured the dynamics of SVZ cell division and migration of adult rats subjected to stroke. In normal brain slices, SVZ cells primarily migrated dorsally and ventrally along the lateral ventricular surface. However, in stroke brain slices, SVZ cells migrated laterally toward the striatal ischemic boundary. Cultured stroke-derived SVZ cells exhibited a significant (p < 0.01) increase in the migration distance (212 +/- 21 microm) compared with the nonstroke-derived SVZ cells (97 +/- 12 microm). Migrating stroke-derived SVZ cells spent significantly (p = 0.01) less time in cytokinesis (0.63 +/- 0.04 h) compared with the time (1.09 +/- 0.09 h) for nonstroke-derived SVZ cells. Newborn cells with a single leading process exhibited fast migration (7.2 +/- 0.8 microm/h), and cells with multiple processes showed stationary migration (3.6 +/- 0.8 microm/h). Stroke SVZ daughter cells further divided during their migration. The morphology of doublecortin (DCX)-positive cells in fixed brain sections resembled those observed in cultured newborn cells, and the DCX-positive cells proliferated in the ischemic striatum. Collectively, the present study suggests that stroke promotes cytokinesis of migrating neuroblasts, and these cells migrate toward the ischemic striatum with distinct migratory behaviors and retain the capacity for cell division during migration.

Stroke induces ependymal cell transformation into radial glia in the subventricular zone of the adult rodent brain.

Adult ependymal cells are postmitotic and highly differentiated. Radial glial cells are neurogenic precursors. Here, we show that stroke acutely stimulated adult ependymal cell proliferation, and dividing ependymal cells of the lateral ventricle had genotype, phenotype, and morphology of radial glial cells in the rat. The majority of radial glial cells exhibited symmetrical division about the cell cleavage plane, and a radial fiber was maintained throughout each stage of cell mitosis. Increases of radial glial cells parallel expansion of neural progenitors in the subventricular zone (SVZ). Furthermore, after stroke radial glial cells derived from the SVZ supported neuron migration. These results indicate that adult ependymal cells divide and transform into radial glial cells after stroke, which could function as neural progenitor cells to generate new neurons and act as scaffolds to support neuroblast migration towards the ischemic boundary region.

Matrix metalloproteinase 2 (MMP2) and MMP9 secreted by erythropoietin-activated endothelial cells promote neural progenitor cell migration.

We investigated the hypothesis that endothelial cells activated by erythropoietin (EPO) promote the migration of neuroblasts. This hypothesis is based on observations in vivo that treatment of focal cerebral ischemia with EPO enhances the migration of neuroblasts to the ischemic boundary, a site containing activated endothelial cells and angiogenic microvasculature. To model the microenvironment within the ischemic boundary zone, we used a coculture system of mouse brain endothelial cells (MBECs) and neural progenitor cells derived from the subventricular zone of the adult mouse. Treatment of MBECs with recombinant human EPO (rhEPO) significantly increased secretion of matrix metalloproteinase 2 (MMP2) and MMP9. rhEPO-treated MBEC supernatant as conditioned medium significantly increased the migration of neural progenitor cells. Application of an MMP inhibitor abolished the supernatant-enhanced migration. Incubation of neurospheres alone with rhEPO failed to increase progenitor cell migration. rhEPO activated phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and extracellular signal-regulated kinase (ERK1/2) in MBECs. Selective inhibition of the PI3K/Akt and ERK1/2 pathways significantly attenuated the rhEPO-induced MMP2 and MMP9, which suppressed neural progenitor cell migration promoted by the rhEPO-activated MBECs. Collectively, our data show that rhEPO-activated endothelial cells enhance neural progenitor cell migration by secreting MMP2 and MMP9 via the PI3K/Akt and ERK1/2 signaling pathways. These data demonstrate that activated endothelial cells can promote neural progenitor cell migration, and provide insight into the molecular mechanisms underlying the attraction of newly generated neurons to injured areas in brain.

Neurogenin 1 mediates erythropoietin enhanced differentiation of adult neural progenitor cells.

Proneuronal basic helix-loop-helix (bHLH) transcription factor, neurogenin 1 (Ngn1), regulates neuronal differentiation during development of the cerebral cortex. Akt mediates proneuronal bHLH protein function to promote neuronal differentiation. Here, we show that recombinant human erythropoietin (rhEPO) significantly increased Akt activity and Ngn1 mRNA levels in neural progenitor cells derived from the subventricular zone (SVZ) of adult rat, which was coincident with increases of neural progenitor cell proliferation, differentiation, and neurite outgrowth. Inhibition of Akt activity by the phosphatidylinositol 3-kinase/Akt (PI3K/Akt) inhibitor, LY294002, abolished rhEPO-increased Ngn1 mRNA levels and the effects of rhEPO on neural progenitor cells. In addition, reducing expression of endogenous Ngn1 by means of short-interfering RNA (siRNA) blocked rhEPO-enhanced neuronal differentiation and neurite outgrowth but not rhEPO-increased proliferation. Furthermore, treatment of stroke rat with rhEPO significantly increased Ngn1 mRNA levels in SVZ cells. These data suggest that rhEPO acts as an extracellular molecule that activates the PI3K/Akt pathway, which enhances adult neural progenitor cell proliferation, differentiation, and neurite outgrowth, and Ngn1 is required for Akt-mediated neuronal differentiation and neurite outgrowth.