A Translocation Pathway for Vesicle-Mediated Unconventional Protein Secretion.

Many cytosolic proteins lacking a signal peptide, called leaderless cargoes, are secreted through unconventional secretion. Vesicle trafficking is a major pathway involved. It is unclear how leaderless cargoes enter into the vesicle. Here, we find a translocation pathway regulating vesicle entry and secretion of leaderless cargoes. We identify TMED10 as a protein channel for the vesicle entry and secretion of many leaderless cargoes. The interaction of TMED10 C-terminal region with a motif in the cargo accounts for the selective release of the cargoes. In an in vitro reconstitution assay, TMED10 directly mediates the membrane translocation of leaderless cargoes into the liposome, which is dependent on protein unfolding and enhanced by HSP90s. In the cell, TMED10 localizes on the endoplasmic reticulum (ER)-Golgi intermediate compartment and directs the entry of cargoes into this compartment. Furthermore, cargo induces the formation of TMED10 homo-oligomers which may act as a protein channel for cargo translocation.

An electric molecular motor.

Macroscopic electric motors continue to have a large impact on almost every aspect of modern society. Consequently, the effort towards developing molecular motors1-3 that can be driven by electricity could not be more timely. Here we describe an electric molecular motor based on a [3]catenane4,5, in which two cyclobis(paraquat-p-phenylene)6 (CBPQT4+) rings are powered by electricity in solution to circumrotate unidirectionally around a 50-membered loop. The constitution of the loop ensures that both rings undergo highly (85%) unidirectional movement under the guidance of a flashing energy ratchet7,8, whereas the interactions between the two rings give rise to a two-dimensional potential energy surface (PES) similar to that shown by FOF1 ATP synthase9. The unidirectionality is powered by an oscillating10 voltage11,12 or external modulation of the redox potential13. Initially, we focused our attention on the homologous [2]catenane, only to find that the kinetic asymmetry was insufficient to support unidirectional movement of the sole ring. Accordingly, we incorporated a second CBPQT4+ ring to provide further symmetry breaking by interactions between the two mobile rings. This demonstration of electrically driven continual circumrotatory motion of two rings around a loop in a [3]catenane is free from the production of waste products and represents an important step towards surface-bound14 electric molecular motors.

Correlating Transcription Initiation and Conformational Changes by a Single-Subunit RNA Polymerase with Near Base-Pair Resolution.

We provide a comprehensive analysis of transcription in real time by T7 RNA Polymerase (RNAP) using single-molecule fluorescence resonance energy transfer by monitoring the entire life history of transcription initiation, including stepwise RNA synthesis with near base-pair resolution, abortive cycling, and transition into elongation. Kinetically branching pathways were observed for abortive initiation with an RNAP either recycling on the same promoter or exchanging with another RNAP from solution. We detected fast and slow populations of RNAP in their transition into elongation, consistent with the efficient and delayed promoter release, respectively, observed in ensemble studies. Real-time monitoring of abortive cycling using three-probe analysis showed that the initiation events are stochastically branched into productive and failed transcription. The abortive products are generated primarily from initiation events that fail to progress to elongation, and a majority of the productive events transit to elongation without making abortive products.

Primordial aminoacyl-tRNA synthetases preferred minihelices to full-length tRNA.

Aminoacyl-tRNA synthetases (AARS) and tRNAs translate the genetic code in all living cells. Little is known about how their molecular ancestors began to enforce the coding rules for the expression of their own genes. Schimmel et al. proposed in 1993 that AARS catalytic domains began by reading an 'operational' code in the acceptor stems of tRNA minihelices. We show here that the enzymology of an AARS urzyme•TΨC-minihelix cognate pair is a rich in vitro realization of that idea. The TΨC-minihelixLeu is a very poor substrate for full-length Leucyl-tRNA synthetase. It is a superior RNA substrate for the corresponding urzyme, LeuAC. LeuAC active-site mutations shift the choice of both amino acid and RNA substrates. AARS urzyme•minihelix cognate pairs are thus small, pliant models for the ancestral decoding hardware. They are thus an ideal platform for detailed experimental study of the operational RNA code.

GLP-1 in the Hypothalamic Paraventricular Nucleus Promotes Sympathetic Activation and Hypertension.

Glucagon-like peptide-1 (GLP-1) and its analogs are widely used for diabetes treatment. The paraventricular nucleus (PVN) is crucial for regulating cardiovascular activity. This study aims to determine the roles of GLP-1 and its receptors (GLP-1R) in the PVN in regulating sympathetic outflow and blood pressure. Experiments were carried out in male normotensive rats and spontaneously hypertensive rats (SHR). Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded. GLP-1 and GLP-1R expressions were present in the PVN. PVN microinjection of GLP-1R agonist recombinant human GLP-1 (rhGLP-1) or EX-4 increased RSNA and MAP, which were prevented by GLP-1R antagonist exendin 9-39 (EX9-39) or GLP-1R antagonist 1, superoxide scavenger tempol, antioxidant N-acetylcysteine, NADPH oxidase (NOX) inhibitor apocynin, adenylyl cyclase (AC) inhibitor SQ22536 or protein kinase A (PKA) inhibitor H89. PVN microinjection of rhGLP-1 increased superoxide production, NADPH oxidase activity, cAMP level, AC, and PKA activity, which were prevented by SQ22536 or H89. GLP-1 and GLP-1R were upregulated in the PVN of SHR. PVN microinjection of GLP-1 agonist increased RSNA and MAP in both WKY and SHR, but GLP-1 antagonists caused greater effects in reducing RSNA and MAP in SHR than in WKY. The increased superoxide production and NADPH oxidase activity in the PVN of SHR were augmented by GLP-1R agonists but attenuated by GLP-1R antagonists. These results indicate that activation of GLP-1R in the PVN increased sympathetic outflow and blood pressure via cAMP-PKA-mediated NADPH oxidase activation and subsequent superoxide production. GLP-1 and GLP-1R upregulation in the PVN partially contributes to sympathetic overactivity and hypertension.

Self-assembled soft alloy with Frank-Kasper phases beyond metals.

Soft building blocks, such as micelles, cells or soap bubbles, tend to adopt near-spherical geometry when densely packed together. As a result, their packing structures do not extend beyond those discovered in metallic glasses, quasicrystals and crystals. Here we report the emergence of two Frank-Kasper phases from the self-assembly of five-fold symmetric molecular pentagons. The µ phase, an important intermediate in superalloys, is indexed in soft matter, whereas the ϕ phase exhibits a structure distinct from known Frank-Kasper phases in metallic systems. We find a broad size and shape distribution of self-assembled mesoatoms formed by molecular pentagons while approaching equilibrium that contribute to the unique packing structures. This work provides insight into the manipulation of soft building blocks that deviate from the typical spherical geometry and opens avenues for the fabrication of 'soft alloy' structures that were previously unattainable in metal alloys.

IFIH1 variants are associated with generalised epilepsy preceded by febrile seizures.

BACKGROUND: IFIH1 variants have been reported to be associated with immune-related disorders with/without seizures. It is unknown whether IFIH1 variants are associated with common epilepsy without acquired causes and the mechanism underlying phenotypic variation remains elusive. METHODS: Trio-based whole-exome sequencing was performed on patients with febrile seizures or epilepsy with antecedent febrile seizures. Previously reported variants were systematically reviewed to investigate genotype-phenotype associations. RESULTS: Two de novo heterozygous and three biallelic missense variants were identified in five patients with generalised epilepsy with antecedent febrile seizures. The variants were predicted to be damaging by in silico tools and were associated with hydrogen bonding changes to neighbouring amino acids or decreased protein stability. Patients exhibited an early onset age and became seizure-free with favourable outcome. Further analysis revealed that de novo missense variants located in the Hel region resulted in seizures with multiple neurological abnormalities, while those in the pincer domain or C-terminal domain led to seizures with normal neurodevelopment, suggesting a sub-molecular effect. Biallelic missense variants, which were inherited from unaffected parents and presented low allele frequencies in general populations, were associated with seizures without neurological abnormalities. Truncation variants were related to refractory epilepsy and severe developmental delay, suggesting a genotype-phenotype correlation. IFIH1 is predominantly expressed in the neonatal stage and decreases dramatically in the adulthood, which is consistent with the early onset age and favourable outcome of the patients. CONCLUSIONS: IFIH1 variants are potentially associated with generalised epilepsy with antecedent febrile seizures. The sub-molecular implication and genotype-phenotype association help explain phenotype variations of IFIH1 variants.

Domain acquisition by class I aminoacyl-tRNA synthetase urzymes coordinated the catalytic functions of HVGH and KMSKS motifs.

Leucyl-tRNA synthetase (LeuRS) is a Class I aminoacyl-tRNA synthetase (aaRS) that synthesizes leucyl-tRNAleu for codon-directed protein synthesis. Two signature sequences, HxGH and KMSKS help stabilize transition-states for amino acid activation and tRNA aminoacylation by all Class I aaRS. Separate alanine mutants of each signature, together with the double mutant, behave in opposite ways in Pyrococcus horikoshii LeuRS and the 129-residue urzyme ancestral model generated from it (LeuAC). Free energy coupling terms, Δ(ΔG‡), for both reactions are large and favourable for LeuRS, but unfavourable for LeuAC. Single turnover assays with 32Pα-ATP show correspondingly different internal products. These results implicate domain motion in catalysis by full-length LeuRS. The distributed thermodynamic cycle of mutational changes authenticates LeuAC urzyme catalysis far more convincingly than do single point mutations. Most importantly, the evolutionary gain of function induced by acquiring the anticodon-binding (ABD) and multiple insertion modules in the catalytic domain appears to be to coordinate the catalytic function of the HxGH and KMSKS signature sequences. The implication that backbone elements of secondary structures achieve a major portion of the overall transition-state stabilization by LeuAC is also consistent with coevolution of the genetic code and metabolic pathways necessary to produce histidine and lysine sidechains.

Expanding quasiperiodicity in soft matter: Supramolecular decagonal quasicrystals by binary giant molecule blends.

The quasiperiodic structures in metal alloys have been known to depend on the existence of icosahedral order in the melt. Among different phases observed in intermetallics, decagonal quasicrystal (DQC) structures have been identified in many glass-forming alloys yet remain inaccessible in bulk-state condensed soft matters. Via annealing the mixture of two giant molecules, the binary system assemblies into an axial DQC superlattice, which is identified comprehensively with meso-atomic accuracy. Analysis indicates that the DQC superlattice is composed of mesoatoms with an unusually broad volume distribution. The interplays of submesoatomic (molecular) and mesoatomic (supramolecular) local packings are found to play a crucial role in not only the formation of the metastable DQC superlattice but also its transition to dodecagonal quasicrystal and Frank-Kasper σ superlattices.

Syntheses of three-dimensional catenanes under kinetic control.

Although catenanes comprising two ring-shaped components can be made in large quantities by templation, the preparation of three-dimensional (3D) catenanes with cage-shaped components is still in its infancy. Here, we report the design and syntheses of two 3D catenanes by a sequence of SN2 reactions in one pot. The resulting triply mechanically interlocked molecules were fully characterized in both the solution and solid states. Mechanistic studies have revealed that a suit[3]ane, which contains a threefold symmetric cage component as the suit and a tribromide component as the body, is formed at elevated temperatures. This suit[3]ane was identified as the key reactive intermediate for the selective formation of the two 3D catenanes which do not represent thermodynamic minima. We foresee a future in which this particular synthetic strategy guides the rational design and production of mechanically interlocked molecules under kinetic control.

Identification of the Acid-Sensitive Site Critical for Chloral Hydrate (CH) Activation of the Proton-Activated Chloride Channel.

The transmembrane protein TMEM206 was recently identified as the molecular basis of the extracellular proton-activated Cl- channel (PAC), which plays an essential role in neuronal death in ischemia-reperfusion. The PAC channel is activated by extracellular acid, but the proton-sensitive mechanism remains unclear, although different acid-sensitive pockets have been suggested based on the cryo-EM structure of the human PAC (hPAC) channel. In the present study, we firstly identified two acidic amino acid residues that removed the pH-dependent activation of the hPAC channel by neutralization all the conservative negative charged residues located in the extracellular domain of the hPAC channel and some positively charged residues at the hotspot combined with two-electrode voltage-clamp (TEVC) recording in the Xenopus oocytes system. Double-mutant cycle analysis and double cysteine mutant of these two residues proved that these two residues cooperatively form a proton-sensitive site. In addition, we found that chloral hydrate activates the hPAC channel depending on the normal pH sensitivity of the hPAC channel. Furthermore, the PAC channel knock-out (KO) male mice (C57BL/6J) resist chloral hydrate-induced sedation and hypnosis. Our study provides a molecular basis for understanding the proton-dependent activation mechanism of the hPAC channel and a novel drug target of chloral hydrate.SIGNIFICANCE STATEMENT Proton-activated Cl- channel (PAC) channels are widely distributed in the nervous system and play a vital pathophysiological role in ischemia and endosomal acidification. The main discovery of this paper is that we identified the proton activation mechanism of the human proton-activated chloride channel (hPAC). Intriguingly, we also found that anesthetic chloral hydrate can activate the hPAC channel in a pH-dependent manner. We found that the chloral hydrate activates the hPAC channel and needs the integrity of the pH-sensitive site. In addition, the PAC channel knock-out (KO) mice are resistant to chloral hydrate-induced anesthesia. The study on PAC channels' pH activation mechanism enables us to better understand PAC's biophysical mechanism and provides a novel target of chloral hydrate.

FNDC5 inhibits malignant growth of human cervical cancer cells via restraining PI3K/AKT pathway.

Cervical cancer (CxCa) is the fourth most frequent cancer in women. This study aimed to determine the role and underlying mechanism of fibronectin type III domain-containing protein 5 (FNDC5) in inhibiting CxCa growth. Experiments were performed in human CxCa tissues, human CxCa cell lines (HeLa and SiHa), and xenograft mouse model established by subcutaneous injection of SiHa cells in nude mice. Bioinformatics analysis showed that CxCa patients with high FNDC5 levels have a longer overall survival period. FNDC5 expression was increased in human CxCa tissues, HeLa and SiHa cells. FNDC5 overexpression or FNDC5 protein not only inhibited proliferation, but also restrained invasion and migration of HeLa and SiHa cells. The effects of FNDC5 were prevented by inhibiting integrin with cilengitide, activating PI3K with recilisib or activating Akt with SC79. FNDC5 inhibited the phosphorylation of PI3K and Akt, which was attenuated by recilisib. PI3K inhibitor LY294002 showed similar effects to FNDC5 in HeLa and SiHa cells. Intravenous injection of FNDC5 (20 µg/day) for 14 days inhibited the tumor growth, and reduced the proliferation marker Ki67 expression and the Akt phosphorylation in the CxCa xenograft mouse model. These results indicate that FNDC5 inhibits the malignant phenotype of CxCa cells through restraining PI3K/Akt signaling. Upregulation of FNDC5 may play a beneficial role in retarding the tumor growth of CxCa.

Visualizing the Promoting Role of Interfacial Water in the Deprotonation of Formic Acid on Cu(111).

Water plays a crucial role in various heterogeneous catalytic reactions, but the atomic-scale characterization of how water participates in these chemical processes remains a significant challenge. Here we directly visualize the promoting role of interfacial water in the deprotonation of formic acid (FA) on a metal surface, using combined scanning tunneling microscopy and qPlus-based noncontact atomic force microscopy. We find the dissociation of FA when coadsorbed with water on the Cu(111) surface, resulting in the formation of hydronium and formate ions. Interestingly, most of the hydrated proton and formate ions exhibit a phase-separated behavior on Cu(111), in which Eigen and Zundel cations assemble into a monolayer hexagonal hydrogen-bonding (H-bonding) network, and bidentate formate ions are solvated with water and aggregate into one-dimensional chains or two-dimensional H-bonding networks. This phase-separated behavior is essential for preventing the proton transfer back from hydronium to formate and the reformation of FA. Density functional theory calculations reveal that the participation of water significantly reduces the deprotonation barrier of FA on Cu(111), in which water catalyzes the decomposition of FA through the Grotthuss proton transfer mechanism. In addition, the separate solvation of hydronium and bidentate formate ions is energetically preferred due to the enhanced interaction with the copper substrate. The promoting role of water in the deprotonation of FA is further confirmed by the temperature-programmed desorption experiment, which shows that the intensity of the H2 desorption peak significantly increases and the desorption of FA declines when water and FA coadsorbed on the Cu(111) surface.

Iron-Catalyzed Primary Amination of C(sp3)-H Bonds.

Primary amines are privileged molecules in drug development. Yet, there is a noticeable scarcity of methods for directly introducing a primary amine group into the ubiquitous C(sp3)-H bonds within organic compounds. Here, we report an iron-based catalytic system that enables direct primary amination of C(sp3)-H bonds under aqueous conditions and air. Various types of C(sp3)-H bonds, including benzylic, allylic, and aliphatic ones, can be readily functionalized with high selectivity and efficiency. The broad utility of this method has been further verified by late-stage amination of 11 complex bioactive molecules. Mechanistic studies unveil a protonated iron-nitrene complex as the key intermediate for the C-H bond activation. This work extends the toolbox for direct C(sp3)-H functionalizations, opening up new opportunities for late-stage modifications of organic molecules.

A Geometrically Flexible Three-Dimensional Nanocarbon.

The development of architecturally unique molecular nanocarbons by bottom-up organic synthesis is essential for accessing functional organic materials awaiting technological developments in fields such as energy, electronics, and biomedicine. Herein, we describe the design and synthesis of a triptycene-based three-dimensional (3D) nanocarbon, GFN-1, with geometrical flexibility on account of its three peripheral π-panels being capable of interconverting between two curved conformations. An effective through-space electronic communication among the three π-panels of GFN-1 has been observed in its monocationic radical form, which exhibits an extensively delocalized spin density over the entire 3D π-system as revealed by electron paramagnetic resonance and UV-vis-NIR spectroscopies. The flexible 3D molecular architecture of GFN-1, along with its densely packed superstructures in the presence of fullerenes, is revealed by microcrystal electron diffraction and single-crystal X-ray diffraction, which establish the coexistence of both propeller and tweezer conformations in the solid state. GFN-1 exhibits strong binding affinities for fullerenes, leading to host-guest complexes that display rapid photoinduced electron transfer within a picosecond. The outcomes of this research could pave the way for the utilization of shape and electronically complementary nanocarbons in the construction of functional coassemblies.

Anatomical and functional evidence for renal autonomic innervation in normotensive and hypertensive rats.

Renal denervation (RDN) has been used for treating resistant hypertension. A few recent studies show vagal innervation of kidneys causing confusion. This study aimed to provide anatomical and functional evidence for renal autonomic innervation. Experiments were performed in male Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Pseudorabies virus (PRV) in paraventricular nucleus and rostral ventrolateral medulla was prevented by bilateral RDN, but not subdiaphragmatic vagotomy. PRV did not appear in dorsal motor nucleus of vagus and nucleus tractus solitarii 72 h after renal injection of PRV. Adrenergic fibers were approximately 7 times more than cholinergic fibers in main renal artery (MRA) and its first (1RA) and second grade (2RA) branches. Adrenergic fibers in 1RA were more than these in MRA and 2RA. Tyrosine hydroxylase immunoreactivity in these arteries was higher in SHR than WKY. Norepinephrine (NE) increased, and α-receptor antagonist reduced vascular ring tension of renal arteries. The effect of NE was greater in 1RA and 2RA than MRA, which was prevented by α-receptor antagonist. Acetylcholine (ACh) or blockage of ß-receptors, M- or N-receptors had no significant effects on vascular ring tension and the effect of NE. Renal blood flow was reduced by electrical stimulation of renal nerves, but not affected by stimulation of subdiaphragmatic vagus. These results provide anatomical and functional evidence that kidneys are innervated and renal blood flow is regulated by renal sympathetic nerves rather than vagus. Renal vasoconstriction is regulated by NE and adrenergic fibers rather than ACh or cholinergic fibers in WKY and SHR.

Clock knockout in inhibitory neurons reduces predisposition to epilepsy and influences anxiety-like behaviors in mice.

Epilepsy is a brain disorder affecting up to 1 in 26 individuals. Despite its clinical importance, the molecular mechanisms of epileptogenesis are still far from clarified. Our previous study showed that disruption of Clock in excitatory neurons alters cortical circuits and leads to generation of focal epilepsy. In this study, a GAD-Cre;Clockflox/flox mouse line with conditional Clock gene knockout in inhibitory neurons was established. We observed that seizure latency was prolonged, the severity and mortality of pilocarpine-induced seizure were significantly reduced, and memory was improved in GAD-Cre;Clockflox/flox mice. We hypothesize that mice with CLOCK knockout in inhibitory neurons have increased threshold for seizure, opposite from mice with CLOCK knockout in excitatory neurons. Further investigation showed Clock knockout in inhibitory neurons upregulated the basal protein level of ARC, a synaptic plasticity-associated immediate-early gene product, likely through the BDNF-ERK pathway. Altered basal levels of ARC may play an important role in epileptogenesis after Clock deletion in inhibitory neurons. Although sEPSCs and intrinsic properties of layer 5 pyramidal neurons in the somatosensory cortex exhibit no changes, the spine density increased in apical dendrite of pyramidal neurons in CLOCK knockout group. Our results suggest an underlying mechanism by which the circadian protein CLOCK in inhibitory neurons participates in neuronal activity and regulates the predisposition to epilepsy.

HER2-low heterogeneity between primary and paired recurrent/metastatic breast cancer: Implications in treatment and prognosis.

BACKGROUND: With the largest sample size to date, the authors' objective was to investigate the incidence of primary-to-metastatic human epidermal growth factor 2 (HER2) conversion and the predictors for such conversion. Moreover, no previous studies have evaluated the prognosis of patients who have negative HER2 expression (HER2-0) versus low HER2 expression (HER2-low) when HER2 status was assessed based on all recurrent/metastatic lesions. METHODS: The authors included 1299 patients who had available HER2 status of primary breast tumors and paired recurrent/metastatic lesions at Fudan University Shanghai Cancer Center and West China Hospital. RESULTS: In total, 370 patients (28.5%) experienced primary-to-metastatic HER2 conversion. Intrapatient intermetastasis spatial heterogeneity and temporal heterogeneity of HER2 were detected. When assessing HER2 based on recurrent/metastatic tumors, patients who had HER2-0 tumors had significantly shorter overall survival than those who had HER2-low tumors in the overall population and in the estrogen receptor (ER)-negative subgroup. However, when assessing HER2 based on primary tumors, there was no difference in overall survival between patients who had HER2-0 versus HER2-low tumors. Moreover, patients who had tumors that converted from HER2-0 to HER2-low had longer overall survival than those who had consistent HER2-0 status in the ER-negative subgroup. By combining four predictors (ER status, Ki67 index, biopsy site, and disease-free interval), the authors established the first prediction tool to estimate the probability of HER2-0 tumors converting to HER2-low/positive tumors. CONCLUSIONS: Intrapatient primary-to-metastatic and intermetastatic HER2 heterogeneity were observed in this large-scale cohort study. When evaluating HER2 based on recurrent/metastatic tumors, an overall survival difference was observed between patients who had HER2-0 versus HER2-low, recurrent/metastatic breast tumors. The developed prediction tool might help clinicians screen out patients with primary HER2-0 tumors that have a high probability of HER2 status conversion and recommend them for re-biopsy, thus helping to screen out candidate patients for trastuzumab deruxtecan treatment.

Chemerin in caudal division of nucleus tractus solitarius increases sympathetic activity and blood pressure.

Chemerin is an adipokine that contributes to metabolism regulation. Nucleus tractus solitarius (NTS) is the first relay station in the brain for accepting various visceral afferent activities for regulating cardiovascular activity. However, the roles of chemerin in the NTS in regulating sympathetic activity and blood pressure are almost unknown. This study aimed to determine the role and potential mechanism of chemerin in the NTS in modulating sympathetic outflow and blood pressure. Bilateral NTS microinjections were performed in anaesthetized adult male Sprague-Dawley rats. Renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and heart rate (HR) were continuously recorded. Chemerin and its receptor chemokine-like receptor 1 (CMKLR1) were highly expressed in caudal NTS (cNTS). Microinjection of chemerin-9 to the cNTS increased RSNA, MAP and HR, which were prevented by CMKLR1 antagonist α-NETA, superoxide scavenger tempol or N-acetyl cysteine, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitors diphenyleneiodonium or apocynin. Chemerin-9 increased superoxide production and NADPH oxidase activity in the cNTS. The increased superoxide production induced by chemerin-9 was inhibited by α-NETA. The effects of cNTS microinjection of chemerin-9 on the RSNA, MAP and HR were attenuated by the pretreatment with paraventricular nucleus (PVN) microinjection of NMDA receptor antagonist MK-801 rather than AMPA/kainate receptor antagonist CNQX. These results indicate that chemerin-9 in the NTS increases sympathetic outflow, blood pressure and HR via CMKLR1-mediated NADPH oxidase activation and subsequent superoxide production in anaesthetized normotensive rats. Glutamatergic inputs in the PVN are needed for the chemerin-9-induced responses.

"Multi-in-One" Yolk-Shell Structured Nanoplatform Inducing Pyroptosis and Antitumor Immune Response Through Cascade Reactions.

Pyroptosis, a new mode of regulatory cell death, holds a promising prospect in tumor therapy. The occurrence of pyroptosis can trigger the release of damage-associated molecular patterns (DAMPs) and activate the antitumor immune response. Moreover, enhancing intracellular reactive oxygen species (ROS) generation can effectively induce pyroptosis. Herein, an integrated nanoplatform (hCZAG) based on zeolitic imidazolate framework-8 (ZIF-8) with Cu2+ and Zn2+ as active nodes and glucose oxidase (GOx) loading is constructed to evoke pyroptosis. GOx can effectively elevate intracellular hydrogen peroxide (H2O2) levels to regulate the unfavorable tumor microenvironment (TME). Cu2+ can be reduced to Cu+ by endogenous overexpressed GSH and both Cu2+ and Cu+ can exert Fenton-like activity to promote ROS generation and amplify oxidative stress. In addition, the accumulation of Cu2+ leads to the aggregation of lipoylated dihydrolipoamide S-acetyltransferase (DLAT), thus resulting in cuproptosis. Notably, the outburst of ROS induced by hCZAG activates Caspase-1 proteins, leads to the cleavage of gasdermin D (GSDMD), and induces pyroptosis. Pyroptosis further elicits an adaptive immune response, leading to immunogenic cell death (ICD). This study provides effective strategies for triggering pyroptosis-mediated immunotherapy and achieving improved therapeutic effects.