Integrative mRNA-miRNA interaction analysis associated with the immune response in the head kidney of rainbow trout (Oncorhynchus mykiss) after infectious hematopoietic necrosis virus infection.

Rainbow trout (Oncorhynchus mykiss) is an important cold-water fish widely cultivated in China. The frequent occurrence of viral diseases caused by infectious hematopoietic necrosis virus (IHNV) seriously restricted the healthy development of the rainbow trout farming industry. However, the immune defense mechanism induced by IHNV in rainbow trout has not been fully elucidated. In the present study, we detected mRNA and miRNA expression profiles in rainbow trout head kidney after IHNV infection using RNA-seq and identified key immune-related genes and miRNAs. The results showed that a total of 7486 genes and 277 miRNAs were differentially expressed, and numerous differentially expressed genes (DEGs) enriched in the immune-related pathways such as Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, NOD-like receptor signaling pathway, cytokine-cytokine receptor interaction, and JAK-STAT signaling pathway were significantly up-regulated, including LGP2, MDA5, TRIM25, IRF3, IRF7, TLR3, TLR7, TLR8, MYD88, and IFN1. Integration analysis identified six miRNAs (miR-141-y, miR-200-y, miR-144-y, miR-2188-y, miR-725-y, and miR-203-y) that target at least six key immune-related genes (TRIM25, LGP2, TLR3, TLR7, IRF3, and IRF7). Further, we verified selected immune-related mRNAs and miRNAs through qRT-PCR and confirmed the reliability of the RNA-seq results. These findings improve our understanding of the immune mechanism of rainbow trout infected with IHNV and provide basic data for future breeding for disease resistance in rainbow trout.

Effects of Astaxanthin on Growth Performance, Gut Structure, and Intestinal Microorganisms of Penaeus vannamei under Microcystin-LR Stress.

Microcystin-LR (MC-LR) are biologically active cycloheptapeptide compounds that are released by cyanobacteria during water blooms and are extensively found in aquatic ecosystems. The Penaeus vannamei is a significant species in global aquaculture. However, the high level of eutrophication in aquaculture water frequently leads to outbreaks of cyanobacterial blooms, posing a significant threat to its sustainable cultivation. Astaxanthin (AX) is commonly utilized in aquaculture for its physiological benefits, including promoting growth and enhancing immune function in cultured organisms. This study aimed to examine the protective effect of astaxanthin on P. vannamei exposed to microcystin-induced stress. The experiment consisted of three groups: one group was fed formulated feed containing MC (100 µg/kg), another group was fed formulated feed containing MC (100 µg/kg) + AX (100 mg/kg), and the third group was fed basic feed (control group). After 15 days of feeding, the specific growth rate (SGR) was significantly higher in the MCAX group (2.21% day-1) compared to the MC group (0.77% day-1), and there was no significant difference between the MCAX group (2.21% day-1) and the control group (2.24% day-1). Similarly, the percent of weight gain (PWG) was also significantly higher in the MCAX group (14.61%) compared to the MC group (13.44%) and the control group (16.64%). Compared to the control group, the epithelial cells in the MC group suffered severe damage and detachment from the basement membrane. However, in the MCAX group, although there was still a gap between the intestinal epithelial cells and the basement membrane, the overall intestinal morphology was slightly less impaired than it was in the MC group. The analysis of the intestinal microbiota revealed a significant disparity in the community composition (chao 1 and ACE) between the MC and MCAX groups. When comparing the various bacterial genera, the MC group exhibited an increase in Vibrio abundance, whereas the MCAX group showed a decrease in both Shewanella and Vibrio abundance. The results indicate that AX has a positive impact on the growth performance and resistance of P. vannamei against MC by regulating the composition of the intestinal microbiota. AX can be utilized to mitigate the detrimental effects of MC in aquaculture practices. This function could be attributed to the role of AX in preserving the structural integrity of the intestinal mucosa and regulating the composition of the intestinal microbiota.

Increased BBB Permeability Enhances Activation of Microglia and Exacerbates Loss of Dendritic Spines After Transient Global Cerebral Ischemia.

Ischemic stroke can induce rapid disruption of blood-brain barrier (BBB). It has been suggested that increased BBB permeability can affect the pathological progression of ischemic tissue. However, the impact of increased BBB permeability on microglial activation and synaptic structures following reperfusion after ischemia remains unclear. In this study, we investigated microglial activation, dendritic damage and plasticity of dendritic spines after increasing BBB permeability following transient global cerebral ischemia in the somatosensory cortices in mice. Bilateral common carotid artery ligation (BCAL) was used to induce transient global cerebral ischemia. Mannitol was used to increase the BBB permeability. Intravital two-photon imaging was performed to image the dendritic structures and BBB extravasation. Microglial morphology was quantitated using a skeletonization analysis method. To evaluate inflammation of cerebral cortex, the mRNA expression levels of integrin alpha M (CD11b), CD68, chemokine (C-X-C motif) ligand 10 (IP10) and tumor necrosis factor alpha (TNF-α) were measured by fluorescent quantitative PCR. Intravital two-photon imaging revealed that mannitol caused a drastic increase in BBB extravasation during reperfusion after transient global ischemia. Increased BBB permeability induced by mannitol had no significant effect on inflammation and dendritic spines in healthy mice but triggered a marked de-ramification of microglia; importantly, in ischemic animals, mannitol accelerated de-ramification of microglia and aggravated inflammation at 3 h but not at 3 days following reperfusion after ischemia. Although mannitol did not cause significant change in the percentage of blebbed dendrites and did not affect the reversible recovery of the dendritic structures, excessive extravasation was accompanied with significant decrease in spine formation and increase in spine elimination during reperfusion in ischemic mice. These findings suggest that increased BBB permeability induced by mannitol can lead to acute activation of microglia and cause excessive loss of dendritic spines after transient global cerebral ischemia.

Transcriptomic analysis reveals differential activation of microglial genes after ischemic stroke in mice.

Microglia are immune cells in the brain and play a pivotal role in the progression of ischemic injury, but the gene expression and signaling pathways related to the activation of microglia following ischemia remain unclear. In our experiment, we used digital gene expression (DGE) analysis to profile the transcriptome of ischemic tissue in a photothrombosis model. DGE analysis identified that a total of 749 genes were differentially regulated (643 up-regulated and 106 down-regulated) after 2days and 7days following stroke. We found 74.5% of these differentially expressed genes were microglial genes. Gene ontology (GO) analysis categorizes these differentially expressed genes at 2days and 7days to specific biological processes such as inflammatory response, cell activation, cell proliferation, and chemokine and cytokine production. Our data demonstrated that a large number of microglial genes were highly regulated at 2days after stroke, but the number of differentially expressed genes had reduced drastically by 7days. Importantly, some of the differentially expressed microglial genes at 7days did not show differential expression at 2days after stroke. DGE analysis indicated that specific genes related to microgliosis were regulated after ischemia. Consistent with the changes in transcriptome, the results from histological analysis of transgenic mice revealed that the microglia proliferated and aggregated surrounding the ischemic core during the period from 2days to 7days following photothrombosis. Together, these results suggested that transcriptomic changes in microglial genes after stroke may have a profound implication for pathophysiology and treatment of stroke.

Reversible recovery of neuronal structures depends on the degree of neuronal damage after global cerebral ischemia in mice.

It has been observed by in vivo imaging that damaged neuronal structures can be reversibly restored after ischemic insults with the application of timely therapeutic interventions. However, what degree of neuronal damage can be restored and the time frame for reversible recovery of neuronal structures remain unclear. Here, transcranial two-photon imaging, histological staining and electron microscopy were used to investigate the reversible recovery of neuronal structures from dendrites to soma after different durations of global cerebral ischemia in mice. Intravital imaging revealed that the damage to dendritic structures was reversible when ischemia time was <1h, but they became difficult to restore after >3h of ischemia. Data from fixed YFP brain slice and Golgi staining indicated that the damage of dendritic structures progressively extended to deeper dendritic shafts with the extension of ischemia time. Furthermore, longer duration of ischemia caused an increasing number of degenerating neurons. Importantly, significant chromatin margination and karyopyknosis of neuron were observed after 6h of ischemia. These data suggested that neuronal structures could be reversibly restored when ischemia time was <1h, but irreversible and progressive damage to neurons occurred with longer duration of ischemia. Consistently, behavioral performance of post-ischemic animals experienced an ischemia time-dependent recovery. Taken together, our data suggested that recovery of neuronal structures following ischemia was dependent on the duration of ischemia, and prevention of neuronal loss is a key target for therapeutic interventions in ischemic stroke.

Transient global cerebral ischemia induces rapid and sustained reorganization of synaptic structures.

Ischemia can cause rapid neuronal damage. Previous studies have suggested that synaptic structures and cortical functions can be rescued if therapeutic interventions are applied in time, but the structural basis for this resilience remains incompletely understood. Here, we investigated the restoration of synaptic structures and postischemic plasticity of dendritic spines in the somatosensory cortices of mice by taking advantage of a reversible global cerebral ischemia model. Intravital two-photon imaging revealed that although dendritic structures were rapidly distorted after global ischemia, only a small percentage of spines were actually lost after transient ischemia. Electron microscopy indicated that most presynaptic electron-dense structures were still apposed to postsynaptic densities, and that the majority of disrupted synaptic structures were rapidly reinstated following reperfusion after transient ischemia. Repeated imaging suggested that restored dendrites survived the initial ischemia -reperfusion challenge. Importantly, spines on the restored dendrites underwent a rapid and sustained structural reorganization following transient ischemia. These findings suggested that disrupted synapses during transient ischemia could be rapidly restored after ischemia/reperfusion, and that restored dendritic structures remained plastic to rebuild the cortical network.

Infiltrating cells from host brain restore the microglial population in grafted cortical tissue.

Transplantation of embryonic cortical tissue is considered as a promising therapy for brain injury. Grafted neurons can reestablish neuronal network and improve cortical function of the host brain. Microglia is a key player in regulating neuronal survival and plasticity, but its activation and dynamics in grafted cortical tissue remain unknown. Using two-photon intravital imaging and parabiotic model, here we investigated the proliferation and source of microglia in the donor region by transplanting embryonic cortical tissue into adult cortex. Live imaging showed that the endogenous microglia of the grafted tissue were rapidly lost after transplantation. Instead, host-derived microglia infiltrated and colonized the graft. Parabiotic model suggested that the main source of infiltrating cells is the parenchyma of the host brain. Colonized microglia proliferated and experienced an extensive morphological transition and eventually differentiated into resting ramified morphology. Collectively, these results demonstrated that donor tissue has little contribution to the activated microglia and host brain controls the microglial population in the graft.