Localization of the sodium-potassium-chloride cotransporter (SLc12A2) during zebrafish embryogenesis and myogenesis and a screen for additional antibodies to study zebrafish myogenesis

The objective of the current study was to screen a series of commercially avaliable antibodies in order to determine if they would bind specifically to target proteins in zebrafish musculature for use in fluorscence confocal microscopy. Many of these antibodies were not developed specifically for use in zebrafish; but because of the near universality of many basic muscle proteins among species these antibodies could potentially bind to homologus proteins in zebrafish. Of these, T4, for the protein Slc12a, has not been well described. We reviewed antibody expression in the muscle and evaluated the protein localization of Slc12a in zebrafish development. T4 was visualized within the skeletal muscle, where it may play an important role in ion regulation during muscle activity. Overall, the use of these antibodies will allow researchers access to tools known to work within the zebrafish and enable more studies using this organism as a model system for muscle development

Intracellular Cryptococcus neoformans disrupts the transcriptome profile of M1- and M2-polarized host macrophages.

Macrophages serve as a first line of defense against infection with the facultative intracellular pathogen, Cryptococcus neoformans (Cn). However, the ability of these innate phagocytic cells to destroy ingested Cn is strongly influenced by polarization state with classically (M1) activated macrophages better able to control cryptococcal infections than alternatively (M2) activated cells. While earlier studies have demonstrated that intracellular Cn minimally affects the expression of M1 and M2 markers, the impact on the broader transcriptome associated with these states remains unclear. To investigate this, an in vitro cell culture model of intracellular infection together with RNA sequencing-based transcriptome profiling was used to measure the impact of Cn infection on gene expression in both polarization states. The gene expression profile of both M1 and M2 cells was extensively altered to become more like naive (M0) macrophages. Gene ontology analysis suggested that this involved changes in the activity of the Janus kinase-signal transducers and activators of transcription (JAK-STAT), p53, and nuclear factor-κB (NF-κB) pathways. Analyses of the principle polarization markers at the protein-level also revealed discrepancies between the RNA- and protein-level responses. In contrast to earlier studies, intracellular Cn was found to increase protein levels of the M1 marker iNos. In addition, common gene expression changes were identified that occurred post-Cn infection, independent of polarization state. This included upregulation of the transcriptional co-regulator Cited1, which was also apparent at the protein level in M1-polarized macrophages. These changes constitute a transcriptional signature of macrophage Cn infection and provide new insights into how Cn impacts gene expression and the phenotype of host phagocytes