A plant-derived natural photosynthetic system for improving cell anabolism.

Insufficient intracellular anabolism is a crucial factor involved in many pathological processes in the body1,2. The anabolism of intracellular substances requires the consumption of sufficient intracellular energy and the production of reducing equivalents. ATP acts as an 'energy currency' for biological processes in cells3,4, and the reduced form of NADPH is a key electron donor that provides reducing power for anabolism5. Under pathological conditions, it is difficult to correct impaired anabolism and to increase insufficient levels of ATP and NADPH to optimum concentrations1,4,6-8. Here we develop an independent and controllable nanosized plant-derived photosynthetic system based on nanothylakoid units (NTUs). To enable cross-species applications, we use a specific mature cell membrane (the chondrocyte membrane (CM)) for camouflage encapsulation. As proof of concept, we demonstrate that these CM-NTUs enter chondrocytes through membrane fusion, avoid lysosome degradation and achieve rapid penetration. Moreover, the CM-NTUs increase intracellular ATP and NADPH levels in situ following exposure to light and improve anabolism in degenerated chondrocytes. They can also systemically correct energy imbalance and restore cellular metabolism to improve cartilage homeostasis and protect against pathological progression of osteoarthritis. Our therapeutic strategy for degenerative diseases is based on a natural photosynthetic system that can controllably enhance cell anabolism by independently providing key energy and metabolic carriers. This study also provides an enhanced understanding of the preparation and application of bioorganisms and composite biomaterials for the treatment of disease.

Histone Deacetylase 3 Couples Mitochondria to Drive IL-1ß-Dependent Inflammation by Configuring Fatty Acid Oxidation.

Immune cell function depends on specific metabolic programs dictated by mitochondria, including nutrient oxidation, macromolecule synthesis, and post-translational modifications. Mitochondrial adaptations have been linked to acute and chronic inflammation, but the metabolic cues and precise mechanisms remain unclear. Here we reveal that histone deacetylase 3 (HDAC3) is essential for shaping mitochondrial adaptations for IL-1ß production in macrophages through non-histone deacetylation. In vivo, HDAC3 promoted lipopolysaccharide-induced acute inflammation and high-fat diet-induced chronic inflammation by enhancing NLRP3-dependent caspase-1 activation. HDAC3 configured the lipid profile in stimulated macrophages and restricted fatty acid oxidation (FAO) supported by exogenous fatty acids for mitochondria to acquire their adaptations and depolarization. Rather than affecting nuclear gene expression, HDAC3 translocated to mitochondria to deacetylate and inactivate an FAO enzyme, mitochondrial trifunctional enzyme subunit α. HDAC3 may serve as a controlling node that balances between acquiring mitochondrial adaptations and sustaining their fitness for IL-1ß-dependent inflammation.

A circular RNA, circPTPN14, increases MYC transcription by interacting with FUBP1 and exacerbates renal fibrosis.

Fibrosis is a relentlessly progressive and irreversible cause of organ damage, as in chronic kidney disease (CKD), but its underlying mechanisms remain elusive. We found that a circular RNA, circPTPN14, is highly expressed in human kidneys with biopsy-proved chronic interstitial fibrosis, mouse kidneys subjected to ischemia/reperfusion (IR) or unilateral ureteral obstruction (UUO), and TGFß1-stimulated renal tubule epithelial cells (TECs). The intrarenal injection of circPTPN14 shRNA alleviated the progression of fibrosis in kidneys subjected to IR or UUO. Knockdown of circPTPN14 in TECs inhibited TGFß1-induced expression of profibrotic genes, whereas overexpressing circPTPN14 increased the profibrotic effect of TGFß1. The profibrotic action of circPTPN14 was ascribed to an increase in MYC transcription. The binding of circPTPN14 to the KH3 and KH4 domains of far upstream element (FUSE) binding protein 1 (FUBP1) enhanced the interaction between FUBP1 and FUSE domain, which was required for the initiation of MYC transcription. In human kidneys (n = 30) with biopsy-proved chronic interstitial fibrosis, the expression of circPTPN14 positively correlated with MYC expression. Taken together these studies show a novel mechanism in the pathogenesis of renal fibrosis, mediated by circPTPN14, which can be a target in the diagnosis and treatment of CKD.

Smart Nanosacrificial Layer on the Bone Surface Prevents Osteoporosis through Acid-Base Neutralization Regulated Biocascade Effects.

Osteoporosis is a global chronic disease characterized by severe bone loss and high susceptibility to fragile fracture. It is widely accepted that the origin acidified microenvironment created by excessive osteoclasts causes irreversible bone mineral dissolution and organic degradation during osteoclastic resorption. However, current clinically available approaches are mainly developed from the perspective of osteoclast biology rather than the critical acidified niche. Here, we developed a smart "nanosacrificial layer" consisting of sodium bicarbonate (NaHCO3)-containing and tetracycline-functionalized nanoliposomes (NaHCO3-TNLs) that can target bone surfaces and respond to external secreted acidification from osteoclasts, preventing osteoporosis. In vitro and in vivo results prove that this nanosacrificial layer precisely inhibits the initial acidification of osteoclasts and initiates a chemically regulated biocascade to remodel the bone microenvironment and realize bone protection: extracellular acid-base neutralization first inhibits osteoclast function and also promotes its apoptosis, in which the apoptosis-derived extracellular vesicles containing RANK (receptor activator of nuclear factor-κ B) further consume RANKL (RANK ligand) in serum, achieving comprehensive osteoclast inhibition. Our therapeutic strategy for osteoporosis is based on original and precise acid-base neutralization, aiming to reestablish bone homeostasis by using a smart nanosacrificial layer that is able to induce chemically regulated biocascade effects. This study also provides a novel understanding of osteoporosis therapy in biomedicine and clinical treatments.

Anti-fatigue effect of phlorizin on exhaustive exercise-induced oxidative injury mediated by Nrf2/ARE signaling pathway in mice.

Oxidative stress plays a crucial role in fatigue, thus it is of significance to develop safe and efficient antioxidant to prevent fatigue. Phlorizin (PHZ) is a major active ingredient of dihydrochalcone from Lithocarpus polystachyus Rehd., which has already been approved as a new food material in China since 2017. The current study was designed to investigate the effect of PHZ on fatigue, and further to elucidate its possible underlying mechanism. Our results revealed that PHZ exerted beneficial effect on exhaustive exercise-induced fatigue in mice, as reflected by rotarod test and exhaustive swimming test. Moreover, PHZ also effectively decreased the levels of blood urea nitrogen, creatine kinase and plasma lactic acid, increased the liver glycogen and skeletal muscle glycogen of fatigued mice, as evidenced by enzyme linked immunosorbent assay. PHZ balanced the redox status through reducing generation of reactive oxygen species, enhancing the activities of antioxidative enzymes. Furthermore, PHZ not only increased the ratio of Bcl2/Bax, but also decreased the level of cleaved-caspase 3. Notably, PHZ facilitated nuclear factor erythroid 2-related factor 2 (Nrf2) translocated from cytoplasm to nucleus, and up-regulated its downstream antioxidant response element including heme oxygenase-1 and NADPH quinone oxidoreductase-1. Intriguingly, PHZ directly bound to Nrf2, as evidenced by molecular docking, and the anti-fatigue effects of PHZ were almost abolished in Nrf2 deficient mice. In summary, our findings suggest that PHZ might be a natural occurring antioxidant with safety profile to relieve fatigue via targeting Nrf2 to inhibit apoptosis.

Epithelial Gasdermin D shapes the host-microbial interface by driving mucus layer formation.

Goblet cells and their main secretory product, mucus, play crucial roles in orchestrating the colonic host-microbe interactions that help maintain gut homeostasis. However, the precise intracellular machinery underlying this goblet cell-induced mucus secretion remains poorly understood. Gasdermin D (GSDMD) is a recently identified pore-forming effector protein that causes pyroptosis, a lytic proinflammatory type of cell death occurring during various pathophysiological conditions. Here, we reveal an unexpected function of GSDMD in goblet cell mucin secretion and mucus layer formation. Specific deletion of Gsdmd in intestinal epithelial cells (ΔIEC) led to abrogated mucus secretion with a concomitant loss of the mucus layer. This impaired colonic mucus layer in GsdmdΔIEC mice featured a disturbed host-microbial interface and inefficient clearance of enteric pathogens from the mucosal surface. Mechanistically, stimulation of goblet cells activates caspases to process GSDMD via reactive oxygen species production; in turn, this activated GSDMD drives mucin secretion through calcium ion-dependent scinderin-mediated cortical F-actin disassembly, which is a key step in granule exocytosis. This study links epithelial GSDMD to the secretory granule exocytotic pathway and highlights its physiological nonpyroptotic role in shaping mucosal homeostasis in the gut.

Gasdermin D maintains bone mass by rewiring the endo-lysosomal pathway of osteoclastic bone resorption.

Gasdermin D (GSDMD)-mediated pyroptosis induces immunogenic cell death and promotes inflammation. However, the functions of GSDMD in tissue homeostasis remain unclear. Here, we identify a physiological function of GSDMD in osteoclasts via a non-lytic p20-generated protein, which prevents bone loss to maintain bone homeostasis. In the late stage of RANKL-induced osteoclastogenesis, GSDMD underwent cleavage, which is dependent on RIPK1 and caspase-8/-3, to yield this p20 product. Gsdmd-deficient osteoclasts showed normal differentiation but enhanced bone resorption with excessive lysosomal activity. Mice with complete or myeloid-specific Gsdmd deletion exhibited increased trabecular bone loss and more severe aging/ovariectomy-induced osteoporosis. GSDMD p20 was preferentially localized to early endosomes and limited endo-lysosomal trafficking and maturation, relying on its oligomerization and control of phosphoinositide conversion by binding to phosphatidylinositol 3-phosphate (PI(3)P). We have thus identified an anti-osteoclastic function of GSDMD as a checkpoint for lysosomal maturation and secretion and linked this to bone homeostasis and endosome-lysosome biology.

Ubiquitination of NLRP3 by gp78/Insig-1 restrains NLRP3 inflammasome activation.

The NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome plays a pivotal role in defending the host against infection as well as sterile inflammation. Activation of the NLRP3 inflammasome is critically regulated by a de-ubiquitination mechanism, but little is known about how ubiquitination restrains NLRP3 activity. Here, we showed that the membrane-bound E3 ubiquitin ligase gp78 mediated mixed ubiquitination of NLRP3, which inhibited NLRP3 inflammasome activation by suppressing the oligomerization and subcellular translocation of NLRP3. In addition, the endoplasmic reticulum membrane protein insulin-induced gene 1 (Insig-1) was required for this gp78-NLRP3 interaction and gp78-mediated NLRP3 ubiquitination. gp78 or Insig-1 deficiency in myeloid cells led to exacerbated NLRP3 inflammasome-dependent inflammation in vivo, including lipopolysaccharide-induced systemic inflammation and alum-induced peritonitis. Taken together, our study identifies gp78-mediated NLRP3 ubiquitination as a regulatory mechanism that restrains inflammasome activation and highlights NLRP3 ubiquitination as a potential therapeutic target for inflammatory diseases.

Kir2.1-mediated membrane potential promotes nutrient acquisition and inflammation through regulation of nutrient transporters.

Immunometabolism contributes to inflammation, but how activated macrophages acquire extracellular nutrients to fuel inflammation is largely unknown. Here, we show that the plasma membrane potential (Vm) of macrophages mediated by Kir2.1, an inwardly-rectifying K+ channel, is an important determinant of nutrient acquisition and subsequent metabolic reprogramming promoting inflammation. In the absence of Kir2.1 activity, depolarized macrophage Vm lead to a caloric restriction state by limiting nutrient uptake and concomitant adaptations in nutrient conservation inducing autophagy, AMPK (Adenosine 5'-monophosphate-activated protein kinase), and GCN2 (General control nonderepressible 2), which subsequently depletes epigenetic substrates feeding histone methylation at loci of a cluster of metabolism-responsive inflammatory genes, thereby suppressing their transcription. Kir2.1-mediated Vm supports nutrient uptake by facilitating cell-surface retention of nutrient transporters such as 4F2hc and GLUT1 by its modulation of plasma membrane phospholipid dynamics. Pharmacological targeting of Kir2.1 alleviated inflammation triggered by LPS or bacterial infection in a sepsis model and sterile inflammation in human samples. These findings identify an ionic control of macrophage activation and advance our understanding of the immunomodulatory properties of Vm that links nutrient inputs to inflammatory diseases.

Tumor-associated macrophages are shaped by intratumoral high potassium via Kir2.1.

The tumor microenvironment (TME) is a unique niche governed by constant crosstalk within and across all intratumoral cellular compartments. In particular, intratumoral high potassium (K+) has shown immune-suppressive potency on T cells. However, as a pan-cancer characteristic associated with local necrosis, the impact of this ionic disturbance on innate immunity is unknown. Here, we reveal that intratumoral high K+ suppresses the anti-tumor capacity of tumor-associated macrophages (TAMs). We identify the inwardly rectifying K+ channel Kir2.1 as a central modulator of TAM functional polarization in high K+ TME, and its conditional depletion repolarizes TAMs toward an anti-tumor state, sequentially boosting local anti-tumor immunity. Kir2.1 deficiency disturbs the electrochemically dependent glutamine uptake, engendering TAM metabolic reprogramming from oxidative phosphorylation toward glycolysis. Kir2.1 blockade attenuates both murine tumor- and patient-derived xenograft growth. Collectively, our findings reveal Kir2.1 as a determinant and potential therapeutic target for regaining the anti-tumor capacity of TAMs within ionic-imbalanced TME.

Predictive model for the 5-year survival status of osteosarcoma patients based on the SEER database and XGBoost algorithm.

Osteosarcoma is the most common bone malignancy, with the highest incidence in children and adolescents. Survival rate prediction is important for improving prognosis and planning therapy. However, there is still no prediction model with a high accuracy rate for osteosarcoma. Therefore, we aimed to construct an artificial intelligence (AI) model for predicting the 5-year survival of osteosarcoma patients by using extreme gradient boosting (XGBoost), a large-scale machine-learning algorithm. We identified cases of osteosarcoma in the Surveillance, Epidemiology, and End Results (SEER) Research Database and excluded substandard samples. The study population was 835 and was divided into the training set (n = 668) and validation set (n = 167). Characteristics selected via survival analyses were used to construct the model. Receiver operating characteristic (ROC) curve and decision curve analyses were performed to evaluate the prediction. The accuracy of the prediction model was excellent both in the training set (area under the ROC curve [AUC] = 0.977) and the validation set (AUC = 0.911). Decision curve analyses proved the model could be used to support clinical decisions. XGBoost is an effective algorithm for predicting 5-year survival of osteosarcoma patients. Our prediction model had excellent accuracy and is therefore useful in clinical settings.

AKT controls NLRP3 inflammasome activation by inducing DDX3X phosphorylation.

The NLRP3 inflammasome, a critical component of the innate immune system, induces caspase-1 activation and interleukin-1ß maturation and drives cell fate toward pyroptosis. However, the mechanism of NLRP3 inflammasome activation still remains elusive. Here we provide evidence that AKT regulates NLRP3 inflammasome activation. Upon NLRP3 activation, AKT activity is inhibited by second stimulus-induced reactive oxygen species. In contrast, AKT activation leads to NLRP3 inhibition and improved mitochondrial fitness. Mechanistically, AKT induces the phosphorylation of the DDX3X (DEAD-box helicase 3, X-linked), a recently identified NLRP3 inflammasome component, and impairs the interaction between DDX3X and NLRP3. Furthermore, an AKT agonist reduces NLRP3-dependent inflammation in two in vivo models of LPS-induced sepsis and Alum-induced peritonitis. Altogether, our study highlights an important role of AKT in controlling NLRP3 inflammasome activation.

Periosteal matrix-derived hydrogel promotes bone repair through an early immune regulation coupled with enhanced angio- and osteogenesis.

Bone healing is a complex physiological process initiated by early regulation of the inflammatory immunity and entails multiple events including angiogenesis, osteogenic differentiation, and biomineralization. Here, we fabricated an injectable periosteal extracellular matrix (PEM) hydrogel that dynamically integrates multiple biological functions and, therefore, acts at different stages of the fracture healing process. PEM hydrogels were fully characterized compared with a collagen I hydrogel. The effects of PEM hydrogels on the different phases of the healing process were assessed in vitro. PEM hydrogels induced the recruitment and M2-polarization of macrophages, promoted the differentiation of MSCs into endothelial-like cells, HUVEC tube formation, osteogenic differentiation of primary calvarial osteoblasts and MSCs, and mineralization after being immersed in simulated body fluid. The dynamic and multiphase effects of the hydrogels were evaluated using a rat critical-sized calvarial defect model in vivo. During the early phase of repair, PEM hydrogels facilitated the M1-to-M2 transition of macrophages. As bone repair progressed, PEM hydrogels promoted blood vessel migration, the development of relative larger blood vessels, and functional vascularization. These effects were also verified in a subcutaneous embedding model. Eventually, PEM hydrogels promoted mature bone formation in large bone defects to a greater extent than collagen I hydrogels. These biological effects coordinated well with the natural process of bone regeneration. Thus, PEM hydrogels may serve as promising biomaterials in bone tissue engineering.

A cancellous bone matrix system with specific mineralisation degrees for mesenchymal stem cell differentiation and bone regeneration.

Bone regenerative therapies have been explored using various biomaterial systems. Notably, collagen biomineralisation is believed to be essential for promoting bone regeneration. However, ideal bone repair materials with an appropriate mineralised matrix, superior osteogenic activity with early vascularisation, and recellularisation properties are still needed. This study aimed to develop a method to subject the decellularised cancellous bone matrix (DCBM) to ultrasound to obtain specific demineralisation to investigate the effects of DCBM with different degrees of mineralisation on proliferation and osteogenic differentiation in bone marrow-derived mesenchymal stem cells (BMSCs) and in repairing femoral bone defects in rabbits. We established an optimised native DCBM mineralisation ECM scaffold for bone regeneration. Upon complete decellularisation of the cancellous bone matrix, DCBMs with specific degrees of mineralisation were obtained. We comprehensively evaluated their bioactive components, minimal immunogenicity, ultra-micro-structural mechanical properties, and degree of mineralisation. Furthermore, specific mineralised DCBMs (obtained by low-temperature rapid ultrasound for 4 and 8 h) had prominent effects in promoting the osteogenic differentiation of BMSCs in vitro. Moreover, more newly formed trabeculae, vessels, and endochondral bone were also detected in the aforementioned groups during early-stage bone repair in vivo. The underlying mechanism might be mineralisation-related regulation and ultra-micro-structural mechanical properties. Thus, the present study shows that specific demineralised DCBM obtained under optimal conditions had superior properties to those of unmineralised or completely demineralised DCBM by promoting MSC osteogenic differentiation and initiating endochondral bone formation and de novo osteogenesis.