A common Shox2–Nkx2-5 antagonistic mechanism primes the pacemaker cell fate in the pulmonary vein myocardium and sinoatrial node

In humans, atrial fibrillation is often triggered by ectopic pacemaking activity in the myocardium sleeves of the pulmonary vein (PV) and systemic venous return. The genetic programs that abnormally reinforce pacemaker properties at these sites and how this relates to normal sinoatrial node (SAN) development remain uncharacterized. It was noted previously that Nkx2-5, which is expressed in the PV myocardium and reinforces a chamber-like myocardial identity in the PV, is lacking in the SAN. Here we present evidence that in mice Shox2 antagonizes the transcriptional output of Nkx2-5 in the PV myocardium and in a functional Nkx2-5(+) domain within the SAN to determine cell fate. Shox2 deletion in the Nkx2-5(+) domain of the SAN caused sick sinus syndrome, associated with the loss of the pacemaker program. Explanted Shox2(+) cells from the embryonic PV myocardium exhibited pacemaker characteristics including node-like electrophysiological properties and the capability to pace surrounding Shox2(−) cells. Shox2 deletion led to Hcn4 ablation in the developing PV myocardium. Nkx2-5 hypomorphism rescued the requirement for Shox2 for the expression of genes essential for SAN development in Shox2 mutants. Similarly, the pacemaker-like phenotype induced in the PV myocardium in Nkx2-5 hypomorphs reverted back to a working myocardial phenotype when Shox2 was simultaneously deleted. A similar mechanism is also adopted in differentiated embryoid bodies. We found that Shox2 interacts with Nkx2-5 directly, and discovered a substantial genome-wide co-occupancy of Shox2, Nkx2-5 and Tbx5, further supporting a pivotal role for Shox2 in the core myogenic program orchestrating venous pole and pacemaker development

Influence of Terrain Slope on Sub-Surface Fire Behavior in Boreal Forests of China

In recent years, the influence of extreme weather patterns has led to an alarming increase in the frequency and severity of sub-surface forest fires in boreal forests. The Ledum palustre-Larix gmelinii forests of the Daxing’an Mountains of China have emerged as a hotspot for sub-surface fires, and terrain slope has been recognized as a pivotal factor shaping forest fire behavior. The present study was conducted to (1) study the effect of terrain slope on the smoldering temperature and spread rate using simulated smoldering experiments and (2) establish occurrence probability prediction model of the sub-surface fires’ smoldering with different slopes based on the random forest model. The results showed that all the temperatures with different slopes were high, and the highest temperature was 947.91 °C. The spread rates in the horizontal direction were higher than those in the vertical direction, and the difference increased as the slope increased. The influence of slope on the peak temperature was greater than that of spread rate. The peak temperature was extremely positively correlated with the slope, horizontal distance and vertical depth. The spread rate was extremely positively correlated with the slope. The spread rate in the vertical direction was strongly positively correlated with the depth, but was strongly negatively correlated with the horizontal distance; the horizontal spread rate was opposite. The prediction equations for smoldering peak temperature and spread rate were established based on slope, horizontal distance, and vertical depth, and the model had a good fit (p 0.9) and high prediction accuracy (accuracy > 80%). The study proved the effect of slope on the characteristics of sub-surface fire smoldering, explained the variation in peak temperature and spread rate between different slopes, and established the occurrence prediction model based on the random forest model. The selected models had a good fit, and prediction accuracy met the requirement of the sub-surface fire prediction

Phospholipid‐flippase chaperone CDC50A is required for synapse maintenance by regulating phosphatidylserine exposure

Synaptic refinement is a critical physiological process that removes excess synapses to establish and maintain functional neuronal circuits. Recent studies have shown that focal exposure of phosphatidylserine (PS) on synapses acts as an "eat me" signal to mediate synaptic pruning. However, the molecular mechanism underlying PS externalization at synapses remains elusive. Here, we find that murine CDC50A, a chaperone of phospholipid flippases, localizes to synapses, and that its expression depends on neuronal activity. Cdc50a knockdown leads to phosphatidylserine exposure at synapses and subsequent erroneous synapse removal by microglia partly via the GPR56 pathway. Taken together, our data support that CDC50A safeguards synapse maintenance by regulating focal phosphatidylserine exposure at synapses

Microglial GPR56 is the molecular target of maternal immune activation-induced parvalbumin-positive interneuron deficits

Parvalbumin-positive (PV+) interneurons play a critical role in maintaining circuit rhythm in the brain, and their reduction is implicated in autism spectrum disorders. Animal studies demonstrate that maternal immune activation (MIA) leads to reduced PV+ interneurons in the somatosensory cortex and autism-like behaviors. However, the underlying molecular mechanisms remain largely unknown. Here, we show that MIA down-regulates microglial Gpr56 expression in fetal brains in an interleukin-17a-dependent manner and that conditional deletion of microglial Gpr56 [Gpr56 conditional knockout (cKO)] mimics MIA-induced PV+ interneuron defects and autism-like behaviors in offspring. We further demonstrate that elevated microglial tumor necrosis factor-α expression is the underlying mechanism by which MIA and Gpr56 cKO impair interneuron generation. Genetically restoring Gpr56 expression in microglia ameliorates PV+ interneuron deficits and autism-like behaviors in MIA offspring. Together, our study demonstrates that microglial GPR56 plays an important role in PV+ interneuron development and serves as a salient target of MIA-induced neurodevelopmental disorders

In vivo self-assembly and delivery of VEGFR2 siRNA-encapsulated small extracellular vesicles for lung metastatic osteosarcoma therapy

Abstract The prognosis of lung metastatic osteosarcoma (OS) remains disappointing. siRNA-based gene silencing of VEGFR2 is a promising treatment strategy for lung metastatic OS, but there is a lack of safe and efficient delivery systems to encapsulate siRNAs for in vivo administration. This study presented a synthetic biological strategy that remolds the host liver with synthesized genetic circuits for efficient in vivo VEGFR2 siRNA delivery. After being taken-up by hepatocytes, the genetic circuit (in the form of a DNA plasmid) reprogrammed the liver to drive the autonomous intrahepatic assembly and encapsulation of VEGFR2 siRNAs into secretory small extracellular vesicles (sEVs), thus allowing for the transport of self-assembled VEGFR2 siRNAs towards the lung. The results showed that our strategy was superior to the positive medicine (Apatinib) for OS lung metastasis in terms of therapeutic efficacy and toxic adverse effects and may provide a feasible and viable therapeutic solution for lung metastatic OS

A splicing isoform of GPR56 mediates microglial synaptic refinement via phosphatidylserine binding

Developmental synaptic remodeling is important for the formation of precise neural circuitry, and its disruption has been linked to neurodevelopmental disorders such as autism and schizophrenia. Microglia prune synapses, but integration of this synapse pruning with overlapping and concurrent neurodevelopmental processes, remains elusive. Adhesion G protein‐coupled receptor ADGRG1/GPR56 controls multiple aspects of brain development in a cell type‐specific manner: In neural progenitor cells, GPR56 regulates cortical lamination, whereas in oligodendrocyte progenitor cells, GPR56 controls developmental myelination and myelin repair. Here, we show that microglial GPR56 maintains appropriate synaptic numbers in several brain regions in a time‐ and circuit‐dependent fashion. Phosphatidylserine (PS) on presynaptic elements binds GPR56 in a domain‐specific manner, and microglia‐specific deletion of Gpr56 leads to increased synapses as a result of reduced microglial engulfment of PS (+) presynaptic inputs. Remarkably, a particular alternatively spliced isoform of GPR56 is selectively required for microglia‐mediated synaptic pruning. Our present data provide a ligand‐ and isoform‐specific mechanism underlying microglial GPR56‐mediated synapse pruning in the context of complex neurodevelopmental processes

A unique stylopod patterning mechanism by Shox2-controlled osteogenesis.

Vertebrate appendage patterning is programmed by Hox-TALE factor-bound regulatory elements. However, it remains unclear which cell lineages are commissioned by Hox-TALE factors to generate regional specific patterns and whether other Hox-TALE co-factors exist. In this study, we investigated the transcriptional mechanisms controlled by the Shox2 transcriptional regulator in limb patterning. Harnessing an osteogenic lineage-specific Shox2 inactivation approach we show that despite widespread Shox2 expression in multiple cell lineages, lack of the stylopod observed upon Shox2 deficiency is a specific result of Shox2 loss of function in the osteogenic lineage. ChIP-Seq revealed robust interaction of Shox2 with cis-regulatory enhancers clustering around skeletogenic genes that are also bound by Hox-TALE factors, supporting a lineage autonomous function of Shox2 in osteogenic lineage fate determination and skeleton patterning. Pbx ChIP-Seq further allowed the genome-wide identification of cis-regulatory modules exhibiting co-occupancy of Pbx, Meis and Shox2 transcriptional regulators. Integrative analysis of ChIP-Seq and RNA-Seq data and transgenic enhancer assays indicate that Shox2 patterns the stylopod as a repressor via interaction with enhancers active in the proximal limb mesenchyme and antagonizes the repressive function of TALE factors in osteogenesis