SUMMARYAdult zebrafish possess a significant ability to regenerate injured heart tissue through proliferation of pre-existing cardiomyocytes, which contrasts with the inability of mammals to do so after the immediate postnatal period. Zebrafish therefore provide a model system in which to study how an injured heart can be repaired. However, it remains unknown what important processes cardiomyocytes are involved in other than partial de-differentiation and proliferation. Here we show that migration of cardiomyocytes to the injury site is essential for heart regeneration. Ventricular amputation induced expression of cxcl12a and cxcr4b, genes encoding a chemokine ligand and its receptor. We found that cxcl12a was expressed in the epicardial tissue and that Cxcr4 was expressed in cardiomyocytes. We show that pharmacological blocking of Cxcr4 function as well as genetic loss of cxcr4b function causes failure to regenerate the heart after ventricular resection. Cardiomyocyte proliferation was not affected but a large portion of proliferating cardiomyocytes remained localized outside the injury site. A photoconvertible fluorescent reporter-based cardiomyocyte-tracing assay demonstrates that cardiomyocytes migrated into the injury site in control hearts but that migration was inhibited in the Cxcr4-blocked hearts. By contrast, the epicardial cells and vascular endothelial cells were not affected by blocking Cxcr4 function. Our data show that the migration of cardiomyocytes into the injury site is regulated independently of proliferation, and that coordination of both processes is necessary for heart regeneration.

SUMMARYThe transcriptional basis of vertebrate limb initiation, which is a well-studied system for the initiation of organogenesis, remains elusive. Specifically, involvement of the -catenin pathway in limb initiation, as well as its role in hindlimb-specific transcriptional regulation, are under debate. Here, we show that the -catenin pathway is active in the limb-forming area in mouse embryos. Furthermore, conditional inactivation of -catenin as well as Islet1, a hindlimb-specific factor, in the lateral plate mesoderm results in a failure to induce hindlimb outgrowth. We further show that Islet1 is required for the nuclear accumulation of -catenin and hence for activation of the -catenin pathway, and that the -catenin pathway maintains Islet1 expression. These two factors influence each other and function upstream of active proliferation of hindlimb progenitors in the lateral plate mesoderm and the expression of a common factor, Fgf10. Our data demonstrate that Islet1 and -catenin regulate outgrowth and Fgf10-Fgf8 feedback loop formation during vertebrate hindlimb initiation. Our study identifies Islet1 as a hindlimb-specific transcriptional regulator of initiation, and clarifies the controversy regarding the requirement of -catenin for limb initiation.

Classical approaches to immunotherapy that show promise in some malignancies have generally been disappointing when applied to high-grade brain tumors such as glioblastoma multiforme (GBM). We recently showed that ex vivo expanded/activated γδ T cells recognize NKG2D ligands expressed on malignant glioma and are cytotoxic to glioma cell lines and primary GBM explants. In addition, γδ T cells extend survival and slow tumor progression when administered to immunodeficient mice with intracranial human glioma xenografts. We now show that temozolomide (TMZ), a principal chemotherapeutic agent used to treat GBM, increases the expression of stress-associated NKG2D ligands on TMZ-resistant glioma cells, potentially rendering them vulnerable to γδ T cell recognition and lysis. TMZ is also highly toxic to γδ T cells, however, and to overcome this cytotoxic effect γδ T cells were genetically modified using a lentiviral vector encoding the DNA repair enzyme O(6)-alkylguanine DNA alkyltransferase (AGT) from the O(6)-methylguanine methyltransferase (MGMT) cDNA, which confers resistance to TMZ. Genetic modification of γδ T cells did not alter their phenotype or their cytotoxicity against GBM target cells. Importantly, gene modified γδ T cells showed greater cytotoxicity to two TMZ resistant GBM cell lines, U373TMZ-R and SNB-19TMZ-R cells, in the presence of TMZ than unmodified cells, suggesting that TMZ exposed more receptors for γδ T cell-targeted lysis. Therefore, TMZ resistant γδ T cells can be generated without impairing their anti-tumor functions in the presence of high concentrations of TMZ. These results provide a mechanistic basis for combining chemotherapy and γδ T cell-based drug resistant cellular immunotherapy to treat GBM.

This article examines whether the public holds biased perceptions of public organizations (in this case, hospitals) in the United States and whether organizations get credit for positive results from program evaluations. Using an experimental design that replicates Hvidman and Andersen's 2016 Danish study, the study finds no negative public sector biases in the United States, but organizations are not given any credit for positive program evaluations. These results hold in two experimental replications. The implications of the findings for the measurement of public perceptions of government programs and for effective democratic governance are discussed. Evidence for Practice• Performance appraisal results generated by an agency do not appear to affect public evaluations of that agency. • U.S. public hospitals are not perceived more negatively than private hospitals in terms of efficiency, effectiveness, red tape, or benevolence. • Experiments on citizen evaluation need to be replicated in specific agency and national contexts.• Practitioners need to be especially careful about arguments related to efficiency and whether reliable measures of efficiency are possible.

Background: Phototherapy (PT) is widely used to prevent and treat severe hyperbilirubinemia and its associated risks for both acute and chronic bilirubin encephalopathy. Intensive PT, recommended for inpatient treatment of hyperbilirubinemia in term and near-term infants, is defined as having a spectral irradiance of ≥30 μW/cm²/nm. Objectives: We aimed to assess local PT practices by measuring the irradiance of PT devices in local neonatal intensive care units and newborn nurseries. Methods: The irradiance footprint, including maximum irradiance at the center of the footprint, of 39 PT devices in 7 area hospitals was measured according to current practice in these facilities. Results: The mean ± SD (range) footprint irradiance was 20.7 ± 5.8 (8.8-29.4) μW/cm²/nm. The mean ± SD maximum irradiance at the footprint center for all devices at a mean clinically used treatment distance of 33.1 ± 9.3 (25.5-60.0) cm was 27.8 ± 7.0 (14.7-42.0) μW/cm²/nm. Sixty-two percent of the devices did not meet the minimum recommended spectral irradiance for intensive PT. For the sites without irradiance-based protocols, the maximum irradiance of the devices (n = 33) at the treatment distances was 25.8 ± 6.1 μW/cm²/nm. Conclusions: Despite established PT guidelines, local protocols and practices vary. Based on an assessment of 7 local hospitals, intensive PT was suboptimal for 62% of devices. Straightforward changes, such as decreasing the distance between an infant and the light source and establishing a consistent irradiance-based protocol, could substantially improve the quality of the intervention.