Clinical sequencing reveals diagnostic, therapeutic, and prognostic biomarkers for adult-type diffuse gliomas.

Diffuse gliomas in adults are highly infiltrative and largely incurable. Whole exome sequencing (WES) has been demonstrated very useful in genetic analysis. Here WES was performed to characterize genomic landscape of adult-type diffuse gliomas to discover the diagnostic, therapeutic and prognostic biomarkers. Somatic and germline variants of 66 patients with adult-type diffuse gliomas were detected by WES based on the next-generation sequencing. TCGA and CGGA datasets were included to analyze the integrated diagnosis and prognosis. Among 66 patients, the diagnosis of 9 cases was changed, in which 8 cases of astrocytoma were corrected into IDH-wildtype glioblastoma (GBM), and 1 oligodendroglioma without 1p/19q co-deletion into astrocytoma. The distribution of mutations including ATRX/TP53 differed in three cohorts. The genetic mutations in GBM mainly concentrated on the cell cycle, PI3K and RTK pathways. The mutational landscape of astrocytoma was more similar to that of GBM, with the highest frequency in germline variants. Patients with IDH-mutant astrocytoma harboring SNVs of PIK3CA and PIK3R1 showed a significantly worse overall survival (OS) than wild-type patients. AEBP1 amplification was associated with shorter OS in GBM. Our study suggests that clinical sequencing can recapitulate previous findings, which may provide a powerful approach to discover diagnostic, therapeutic and prognostic markers for precision medicine in adult-type diffuse gliomas.

Structural insights into the Caprin-2 HR1 domain in canonical Wnt signaling.

The canonical Wnt signaling pathway plays crucial roles in cell fate decisions as well as in pathogenesis of various diseases. Previously, we reported Caprin-2 as a new regulator of canonical Wnt signaling through a mechanism of facilitating LRP5/6 phosphorylation. Here, we resolved the crystal structure of the N-terminal homologous region 1 (HR1) domain of human Caprin-2. HR1 domain is so far only observed in Caprin-2 and its homologous protein Caprin-1, and the function of this domain remains largely mysterious. Here, the structure showed that HR1 domain of human Caprin-2 forms a homo-dimer and exhibits an overall structure roughly resembling the appearance of a pair of scissors. Moreover, we found that residues R200 and R201, which located at a basic cluster within the N-terminal "blades" region, are critical for Caprin-2's localization to the plasma membrane. In line with this, mutations targeting these two residues decrease Caprin-2's activity in the canonical Wnt signaling. Overall, we characterized a previously unknown "scissors"-like structure of the full-length HR1 domain and revealed its function in mediating Caprin-2's localization to the plasma membrane.

MFGE8 promotes adult hippocampal neurogenesis in rats following experimental subarachnoid hemorrhage via modifying the integrin ß3/Akt signaling pathway.

Subarachnoid hemorrhage (SAH) is one of the most severe type of cerebral strokes, which can cause multiple cellular changes in the brain leading to neuronal injury and neurological deficits. Specifically, SAH can impair adult neurogenesis in the hippocampal dentate gyrus, thus may affecting poststroke neurological and cognitive recovery. Here, we identified a non-canonical role of milk fat globule epidermal growth factor 8 (MFGE8) in rat brain after experimental SAH, involving a stimulation on adult hippocampal neurogenesis(AHN). Experimental SAH was induced in Sprague-Dawley rats via endovascular perforation, with the in vivo effect of MFGE8 evaluated via the application of recombinant human MFGE8 (rhMFGE8) along with pharmacological interventions, as determined by hemorrhagic grading, neurobehavioral test, and histological and biochemical analyses of neurogenesis related markers. Results: Levels of the endogenous hippocampal MFGE8 protein, integrin-ß3 and protein kinase B (p-Akt) were elevated in the SAH relative to control groups, while that of hippocalcin (HPCA) and cyclin D1 showed the opposite change. Intraventricular rhMGFE8 infusion reversed the decrease in doublecortin (DCX) immature neurons in the DG after SAH, along with improved the short/long term neurobehavioral scores. rhMGFE8 treatment elevated the levels of phosphatidylinositol 3-kinase (PI3K), p-Akt, mammalian target of rapamycin (mTOR), CyclinD1, HPCA and DCX in hippocampal lysates, but not that of integrin ß3 and Akt, at 24 hr after SAH. Treatment of integrin ß3 siRNA, the PI3K selective inhibitor ly294002 or Akt selective inhibitor MK2206 abolished the effects of rhMGFE8 after SAH. In conclusion, MFGE8 is upregulated in the hippocampus in adult rats with reduced granule cell genesis. rhMFGE8 administration can rescue this impaired adult neurogenesis and improve neurobehavioral recovery. Mechanistically, the effect of MFGE8 on hippocampal adult neurogenesis is mediated by the activation of integrin ß3/Akt pathway. These findings suggest that exogenous MFGE8 may be of potential therapeutic value in SAH management. Graphical abstract and proposed pathway of rhMFGE8 administration attenuate hippocampal injury by improving neurogenesis in SAH models. SAH caused hippocampal injury and neurogenesis interruption. Administered exogenous MFGE8, recombinant human MFGE8(rhMFGE8), could ameliorate hippocampal injury and improve neurological functions after SAH. Mechanistically, MFGE8 bind to the receptor integrin ß3, which activated the PI3K/Akt pathway to increase the mTOR expression, and further promote the expression of cyclin D1, HPCA and DCX. rhMFGE8 could attenuated hippocampal injury by improving neurogenesis after SAH, however, know down integrin ß3 or pharmacological inhibited PI3K/Akt by ly294002 or MK2206 reversed the neuro-protective effect of rhMFGE8.

Efficacy, Safety, and Early Relapse After Cessation of Upadacitinib Versus Dupilumab in Adolescents with Moderate-to-Severe Atopic Dermatitis: A Real-World Study in China.

Background: The efficacy and safety of upadacitinib and dupilumab for atopic dermatitis (AD) in adolescent patients have been proven in clinical trials. However, few daily practice studies comparing agents have been conducted in this patient population. Objectives: This single-center retrospective cohort study aimed to investigate the efficacy, safety, and early relapse after cessation of upadacitinib and dupilumab for moderate-to-severe AD in Chinese adolescents. Methods: A retrospective study collected data on a cohort of 83 adolescent patients receiving upadacitinib or dupilumab from October 2021 to October 2023. This study comprised two main emphases. The first main emphasis involved the treatment phase, where the efficacy and safety of the two treatments were evaluated. Primary endpoints included the proportion of patients achieving an improvement of ≥50%, ≥75%, and ≥90% in Eczema Area and Severity Index (EASI) from baseline (EASI-50, EASI-75, EASI-90) and the proportion of patients achieving Validated Investigator Global Assessment (vIGA) 0/1 at week 4 and week 40. In the second main emphasis, AD recurrence after treatment discontinuation was assessed in the two treatment groups. The median time to relapse was calculated. Results: A total of 83 patients were enrolled. At week 4, the proportion of patients achieving the primary endpoints, including EASI-75 and EASI-90, was substantially higher with upadacitinib than with dupilumab (51.5% vs 14.0% [P < 0.001], 18.2% vs 2.0% [P < 0.05]). However, at week 40, higher proportion of patients on dupilumab were reaching EASI-50 and EASI-75 and vIGA 0/1. After the discontinuation of dupilumab or upadacitinib therapy, the Kaplan-Meier survival curve showed that the median time to relapse was 270 days in the dupilumab group and 18 days in the upadacitinib group. Conclusions: This study demonstrated that upadacitinib has superior short-term efficacy over dupilumab in adolescents with moderate-to-severe AD, whereas dupilumab trends toward better long-term remission over upadacitinib under the condition of treatment discontinuation in some patients.

Clinical characteristics and treatment strategies for pituitary adenoma associated with intracranial aneurysm.

BACKGROUND: This study aimed to investigate clinical features and treatment strategies for intracranial aneurysm (IA) associated with pituitary adenoma (PA). METHODS: We enrolled patients with lesions in the sellar region and age-matched general population who were confirmed with IA from two hospitals. Four types of treatment strategies were performed, which included Type I (both IA and PA were treated with surgery), Type II (IA was treated with surgery and PA was performed by non-surgical treatment), Type III (PA was performed with surgery and observation was available for IA) and Type IV (both IA and PA were performed with non-surgical treatment). RESULTS: The incidence of IA was 2.2% in the general population, 6.1% in patients with PA, 4.3% in patients with Rathke cleft cyst, 2.8% in patients with meningioma and none were found with IA in patients with craniopharyngioma. Age over 50 years (OR, 2.69; 95% CI, 1.20-6.04; P = 0.016), female (OR, 3.83, P = 0.003), and invasive tumor (OR, 3.26, P = 0.003) were associated with a higher incidence of IA in patients with PA. During the mean follow-up of 49.2 months, no patients experienced stroke, and recurrence of aneurysms and aneurysms treated with observation were stable. Of four patients with recurrence of PA, three patients were treated for type I and one patient for type III. CONCLUSIONS: Preoperative evaluation for aneurysm screening is necessary due to the high incidence of IA in PA patients. Our current treatment strategies may provide a benefit for these patients.

Apoptotic body-inspired nanotherapeutics efficiently attenuate osteoarthritis by targeting BRD4-regulated synovial macrophage polarization.

Bromodomain-containing protein 4 (BRD4) is the most well-studied BET protein that is important for the innate immune response. We recently revealed that targeting BRD4 triggers apoptosis in tumor-associated macrophages, but its role in synovial macrophages and joint inflammation is largely unknown. Herein, we demonstrated that BRD4 was highly expressed in the iNOS-positive M1 macrophages in the human and mouse osteoarthritis (OA) synovium, and conditional knockout of BRD4 in the myeloid lineage using Lyz2-cre; BRD4flox/flox mice significantly abolished anterior cruciate ligament transection (ACLT)-induced M1 macrophage accumulation and synovial inflammation. Accordingly, we successfully constructed apoptotic body-inspired phosphatidylserine-containing nanoliposomes (PSLs) loaded with the BRD4 inhibitor JQ1 to regulate inflammatory macrophages. JQ1-loaded PSLs (JQ1@PSLs) exhibited a higher cellular uptake by macrophages than fibroblast-like synoviocytes (FLSs) in vitro and in vivo, as well as the reduction in proinflammatory M1 macrophage polarization. Intra-articular injections of JQ1@PSLs showed prolonged retention within the joint, and remarkably reduced synovial inflammation and joint pain via suppressing M1 polarization accompanied by reduced TRPA1 expression by targeted inhibition of BRD4 in the macrophages, thus attenuating cartilage degradation during OA development. The results show that BRD4-inhibiting JQ1@PSLs can targeted-modulate macrophage polarization, which opens a new avenue for efficient OA therapy via a "Trojan horse".

Multi-Omics Analysis Reveals the Role of Changes in Platelet Activation Pathways in the Survival Outcome of Patients with Esophageal Squamous Cell Carcinoma.

Objective: The objective of this study was to integrate metabolomics and transcriptomics data to identify key diagnostic and prognostic markers for esophageal squamous cell carcinoma (ESCC). Plasma samples were collected from 85 ESCC patients at different stages and 50 healthy volunteers for non-targeted metabolomic analysis. Methods: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed for non-targeted metabolomic analysis. Subsequently, we integrated the metabolomic data with transcriptomic data from the Gene Expression Omnibus (GEO) and prognosis data from The Cancer Genome Atlas Program (TCGA) to perform pathway analysis. Our focus was on pathways that involve both metabolites and upstream genes, as they often exhibit higher accuracy. Results: Through the integration of metabolomics and transcriptomics, we identified significant alterations in the platelet activation pathway in ESCC. This pathway involves the participation of both metabolites and genes, making it a more accurate reflection of pathological changes associated with the disease. Notably, metabolite arachidonic acid (AA) and chemokine receptor type 2(CXCR2) were significantly downregulated in ESCC, while genes collagen type I alpha 1(COL1A1), collagen type I alpha 2(COL1A2), collagen type III alpha 1(COL3A1), type 3 inositol 1,4,5-trisphosphate receptor (ITPR3), and insulin-like growth factor II mRNA binding protein 3(IGF2BP3) were significantly upregulated, indicating the presence of tumor-induced platelet activation in ESCC. Further analysis of prognosis data revealed that high expression of COL1A1, IGF2BP3, and ITPR3 was associated with a favorable prognosis for ESCC, while high CXCR2 expression was linked to an adverse prognosis. In addition, we combined COL1A1, ITPR3, IGF2BP3, CXCR2, and AA to form a diagnostic biomarker panel. The receiver operating characteristic curve (ROC) demonstrated excellent diagnostic capability (AUC=0.987). Conclusion: Our study underscores the significant role of platelet activation pathways and related genes in the diagnosis and prognosis of ESCC patients. These findings offer promising insights for improving the clinical management of ESCC.

Maresin 1 Activates LGR6 to Alleviate Neuroinflammation via the CREB/JMJD3/IRF4 Pathway in a Rat Model of Subarachnoid Hemorrhage.

Neuroinflammation is an early event of brain injury after subarachnoid hemorrhage (SAH). Whether the macrophage mediators in resolving inflammation 1 (MaR1) is involved in SAH pathogenesis is unknown. In this study, 205 male Sprague-Dawley rats were subjected to SAH via endovascular perforation in the experimental and control groups. MaR1 was dosed intranasally at 1 h after SAH, with LGR6 siRNA and KG-501, GSK-J4 administered to determine the signaling pathway. Neurobehavioral, histological and biochemical data were obtained from the animal groups with designated treatments. The results showed: (i) The leucine-rich repeat containing G protein-coupled receptor 6 (LGR6) was decreased after SAH and reached to the lowest level at 24 h after SAH. Jumonji d3 (JMJD3) protein levels tended to increase and peaked at 24 h after SAH. LGR6 and JMJD3 expression were co-localized with microglia. (ii) MaR1 administration mitigated short-term neurological deficits, brain edema and long-term neurobehavioral performance after SAH, and attenuated microglial activation and neutrophil infiltration. (iii) Knockdown of LGR6, inhibition of CREB phosphorylation or JMJD3 activity abolished the anti-neuroinflammatory effect of MaR1 on the expression of CREB, CBP, JMJD3, IRF4, IRF5, IL-1ß, IL-6 and IL-10, thus prevented microglial activation and neutrophil infiltration. Together, the results show that MaR1 can activate LGR6 and affect CREB/JMJD3/IRF4 signaling to attenuate neuroinflammation after SAH, pointing to a potential pharmacological utility in this disorder.

The mir-199b-5p encapsulated in adipocyte-derived exosomes mediates radioresistance of colorectal cancer cells by targeting JAG1.

Radiotherapy is a key treatment option for colorectal cancer, but its efficacy varies among patients. Our previous studies suggested that adipose tissue may confer the radioresistance of several abdominal tumors, such as pancreatic cancer, biliary cancer, and others. In the present work, the effects of adipocytes in regulating the radiosensitivity of colorectal cancer are explored for the first time. It was found that colony formation was increased and radiation-induced apoptosis decreased in colorectal cancer cells HCT8 and HCT116 co-cultured with adipocytes, which verified the mediation of adipocyte-driven radioresistance in colorectal cancer in vitro. Next, the colorectal cancer cells were incubated with adipocyte-derived exosomes, and a perceptible reduction in radiosensitivity was detected. Furthermore, to investigate the possible mechanisms involved, the exosomes were isolated, the encapsulated microRNAs were extracted and analyzed by small RNA sequencing. Based on bioinformatics analysis and qRT-PCR verification, miR-199b-5p was chosen for functional annotation. It was shown that miR-199b-5p expression was significantly upregulated after 6 Gy irradiation, and overexpressed miR-199b-5p significantly suppressed the radiosensitivity of HCT8 and HCT116 cells. In addition, jagged canonical Notch ligand 1(JAG1) was identified as the target gene of miR-199b-5p by using bioinformatics prediction and dual luciferase reporter gene assay. It was demonstrated that JAG1 conferred the radioresistance of colorectal cancer cells both in vivo and in vitro. Taken together, the present study demonstrates that adipocytes trigger the radioresistance of colorectal cancer cells, probably by targeting JAG1 through an adipocyte-derived exosomal miR-199b-5p.

Trim21 Regulates the Postnatal Development and Thermogenesis of Brown Adipose Tissue.

Brown adipose tissue undergoes rapid postnatal development to mature and plays a crucial role in thermoregulation and energy expenditure, which protects against cold and obesity. Herein, it is shown that the expression of Trim21 mRNA level of interscapular brown adipose tissue elevates after birth, and peaks at P14 (postnatal day 14). Trim21 depletion severely impairs the maturation of interscapular brown adipose tissue, decreases the expression of a series of thermogenic genes, and reduces energy expenditure. Consistently, the loss of Trim21 also leads to a suppression of white adipose tissue "browning", in response to cold exposure and a ß-adrenergic agonist, CL316,243. In addition, Trim21-/- mice are more prone to high-fat diet-induced obesity compared with the control littermates. Taken together, the study for the first time reveals a critical role of Trim21 in regulating iBAT postnatal development and thermogenesis.

Targeting YAP1-regulated Glycolysis in Fibroblast-Like Synoviocytes Impairs Macrophage Infiltration to Ameliorate Diabetic Osteoarthritis Progression.

The interplay between immune cells/macrophages and fibroblast-like synoviocytes (FLSs) plays a pivotal role in initiating synovitis; however, their involvement in metabolic disorders, including diabetic osteoarthritis (DOA), is largely unknown. In this study, single-cell RNA sequencing (scRNA-seq) is employed to investigate the synovial cell composition of DOA. A significant enrichment of activated macrophages within eight distinct synovial cell clusters is found in DOA synovium. Moreover, it is demonstrated that increased glycolysis in FLSs is a key driver for DOA patients' synovial macrophage infiltration and polarization. In addition, the yes-associated protein 1 (YAP1)/thioredoxin-interacting protein (TXNIP) signaling axis is demonstrated to play a crucial role in regulating glucose transporter 1 (GLUT1)-dependent glycolysis in FLSs, thereby controlling the expression of a series of adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1) which may subsequently fine-tune the infiltration of M1-polarized synovial macrophages in DOA patients and db/db diabetic OA mice. For treatment, M1 macrophage membrane-camouflaged Verteporfin (Vt)-loaded PLGA nanoparticles (MVPs) are developed to ameliorate DOA progression by regulating the YAP1/TXNIP signaling axis, thus suppressing the synovial glycolysis and the infiltration of M1-polarized macrophages. The results provide several novel insights into the pathogenesis of DOA and offer a promising treatment approach for DOA.

Trim21 depletion alleviates bone loss in osteoporosis via activation of YAP1/ß-catenin signaling.

Despite the diverse roles of tripartite motif (Trim)-containing proteins in the regulation of autophagy, the innate immune response, and cell differentiation, their roles in skeletal diseases are largely unknown. We recently demonstrated that Trim21 plays a crucial role in regulating osteoblast (OB) differentiation in osteosarcoma. However, how Trim21 contributes to skeletal degenerative disorders, including osteoporosis, remains unknown. First, human and mouse bone specimens were evaluated, and the results showed that Trim21 expression was significantly elevated in bone tissues obtained from osteoporosis patients. Next, we found that global knockout of the Trim21 gene (KO, Trim21-/-) resulted in higher bone mass compared to that of the control littermates. We further demonstrated that loss of Trim21 promoted bone formation by enhancing the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and elevating the activity of OBs; moreover, Trim21 depletion suppressed osteoclast (OC) formation of RAW264.7 cells. In addition, the differentiation of OCs from bone marrow-derived macrophages (BMMs) isolated from Trim21-/- and Ctsk-cre; Trim21f/f mice was largely compromised compared to that of the littermate control mice. Mechanistically, YAP1/ß-catenin signaling was identified and demonstrated to be required for the Trim21-mediated osteogenic differentiation of BMSCs. More importantly, the loss of Trim21 prevented ovariectomy (OVX)- and lipopolysaccharide (LPS)-induced bone loss in vivo by orchestrating the coupling of OBs and OCs through YAP1 signaling. Our current study demonstrated that Trim21 is crucial for regulating OB-mediated bone formation and OC-mediated bone resorption, thereby providing a basis for exploring Trim21 as a novel dual-targeting approach for treating osteoporosis and pathological bone loss.

p55γ degrades RIP3 via MG53 to suppress ischaemia-induced myocardial necroptosis and mediates cardioprotection of preconditioning.

AIMS: Regulated necrosis (necroptosis) and apoptosis are important biological features of myocardial infarction, ischaemia-reperfusion (I/R) injury, and heart failure. However, the molecular mechanisms underlying myocardial necroptosis remain elusive. Ischaemic preconditioning (IPC) is the most powerful intrinsic cardioprotection against myocardial I/R injury. In this study, we aimed to determine whether IPC suppresses I/R-induced necroptosis and the underlying molecular mechanisms. METHODS AND RESULTS: We generated p55γ transgenic and knockout mice and used ligation of left anterior descending coronary artery to produce an in vivo I/R model. The effects of p55γ and its downstream molecules were subsequently identified using mass spectroscopy and co-immunoprecipitation and pulldown assays. We found that p55γ expression was down-regulated in failing human myocardium caused by coronary heart disease as well as in I/R mouse hearts. Cardiac-specific p55γ overexpression ameliorated the I/R-induced necroptosis. In striking contrast, p55γ deficiency (p55γ-/-) and cardiac-specific deletion of p55γ (p55γc-KO) worsened I/R-induced injury. IPC up-regulated p55γ expression in vitro and in vivo. Using reporter and chromatin immunoprecipitation assays, we found that Hif1α transcriptionally regulated p55γ expression and mediated the cardioprotection of IPC. IPC-mediated suppression of necroptosis was attenuated in p55γ-/- and p55γc-KO hearts. Mechanistically, p55γ overexpression decreased the protein levels of RIP3 rather than the mRNA levels, while p55γ deficiency increased the protein abundance of RIP3. IPC attenuated the I/R-induced up-regulation of RIP3, which was abolished in p55γ-deficient mice. Up-regulation of RIP3 attenuated the p55γ- or IPC-induced inhibition of necroptosis in vivo. Importantly, p55γ directly bound and degraded RIP3 in a ubiquitin-dependent manner. We identified MG53 as the E3 ligase that mediated the p55γ-induced degradation of RIP3. In addition, we also found that p55γ activated the RISK pathway during IPC. CONCLUSIONS: Our findings reveal that activation of the MG53-RIP3 signal pathway by p55γ protects the heart against I/R-induced necroptosis and underlies IPC-induced cardioprotection.

Early osteoarthritis diagnosis based on near-infrared spectroscopy combined with aquaphotomics.

Osteoarthritis (OA) is the most common joint disease and the leading cause of disability in elderly individuals. Despite rapid advances in imaging techniques, early OA diagnosis remains a clinical challenge. In the present study, the feasibility of early OA diagnosis was explored via near-infrared spectroscopy (NIRS) combined with aquaphotomics. Synovial fluid samples from 65 cases of OA categorized as mild, moderate, and severe according to theKellgrenandLawrence classification criteria were analyzed via NIRS. The 1st overtone of water (1300-1600 nm) was considered as the research object for an aquaphotomics model, and aquagrams of the mild, moderate, and severe OA cases were generated using 12 water absorption patterns for early OA diagnosis.The aquaphotomics results exhibited clear differences in the region of 1300-1500 nm, and the number of hydrogen bonds of different water species (1412,1424, 1482, and 1496 nm) evidently correlated with OA occurrence and development. With OA progression, the absorption intensity of water molecules without hydrogen bonds (1412 nm/1424 nm) became stronger, while the absorption intensity of water molecules with four hydrogen bonds (1482 nm/1496 nm) decreased.These results together reveal that the established accurate and rapid early OA diagnosis model based on NIRS combined with aquaphotomics is effective and feasible, and that the number of hydrogen bonds can be used as a biomarker for early OA diagnosis.

HSP90-regulated CHIP/TRIM21/p21 Axis Involves in the Senescence of Osteosarcoma Cells.

BACKGROUND: OS is the most frequent malignant bone tumor with a poor prognosis. TRIM21 has been reported to play a critical role in OS by regulating the expression of the TXNIP/p21 axis and inhibiting the senescence of OS cells. AIM: Investigation of the molecular mechanism of tripartite motif 21 (TRIM21) in osteosarcoma (OS) would shed light on the understanding of the pathogenesis of OS. OBJECTIVE: This study aimed to explore the mechanism regulating the protein stability of TRIM21 in the process of OS senescence. METHODS: Human U2 OS cells were used to establish stable cells overexpressing TRIM21 (induced by Dox) or knocking down TRIM21. The co-immunoprecipitation (co-IP) assay was used to examine the interaction between TRIM21 and HSP90. Immunofluorescence (IF) assay was used to observe colocalization in OS cells. Western blot analysis was applied to detect the protein expression, and quantitative real-time PCR (qRT-PCR) assay was used to test the mRNA expression of corresponding genes. SA-ß-gal staining was used to evaluate OS senescence. RESULTS: In this study, we verified the interaction between HSP90 and TRIM21 using a co-IP assay. Knockdown or inhibition of HSP90 with its inhibitor 17-AAG accelerated the degradation of TRIM21 by the proteasome in OS cells. CHIP E3 ligase mediated this degradation of TRIM21, with the knockdown of CHIP rescuing the downregulation of TRIM21 induced by 17-AAG. TRIM21 inhibited OS senescence and downregulated the expression of senescence marker p21, while CHIP exhibited an opposite regulatory role on p21 expression. CONCLUSION: Taken together, our results demonstrated that HSP90 is responsible for the stabilization of TRIM21 in OS and that the CHIP/TRIM21/p21 axis controlled by HSP90 affects the senescence of OS cells.

Single-Cell RNA Sequencing Reveals Transcriptional Changes in the Cartilage of Subchondral Insufficiency Fracture of the Knee.

Purpose: Subchondral insufficiency fracture of the knee (SIFK) is a common cause of knee joint pain that mainly afflicts the elderly. Until now, how a sudden insufficiency fracture of subchondral bone affects the transcriptomic profiles of cartilage in SIFK and OA patients are largely unknown. Methods: Single-cell RNA sequencing (scRNA-seq) was used to identify various cell subsets and evaluate transcriptomic differences in cartilage of SIFK and OA patients. In addition, the above findings were confirmed by histological evaluation and immunohistochemical (IHC) staining. Results: We found that the transcriptomic profiles of cartilage in the SIFK patient was completely different from those of normal and OA patients. Accordingly, several novel cell clusters with activation of hypoxia and endochondral ossification signaling were identified in the SIFK cartilage. Chondrocyte trajectories analysis and IHC staining revealed that transcription factors including TCF4 were found to be highly up-regulated during the occurrence of SIFK, which might drive the reactive formation of cartilage and fibrous tissue and the activation of endochondral ossification. Conclusion: This is the first report to elucidate the transcriptomic alterations and distinct cell type subpopulations in the cartilage of SIFK and OA by the use of scRNA-seq, which provides a new insight in the understanding of the initiation and progression of SIFK.

Versatile Nano-PROTAC-Induced Epigenetic Reader Degradation for Efficient Lung Cancer Therapy.

Recent evidence has indicated that overexpression of the epigenetic reader bromodomain-containing protein 4 (BRD4) contributes to a poor prognosis of lung cancers, and the suppression of its expression promotes cell apoptosis and leads to tumor shrinkage. Proteolysis targeting chimera (PROTAC) has recently emerged as a promising therapeutic strategy with the capability to precisely degrade targeted proteins. Herein, a novel style of versatile nano-PROTAC (CREATE (CRV-LLC membrane/DS-PLGA/dBET6)) is developed, which is constructed by using a pH/GSH (glutathione)-responsive polymer (disulfide bond-linked poly(lactic-co-glycolic acid), DS-PLGA) to load BRD4-targeted PROTAC (dBET6), followed by the camouflage with engineered lung cancer cell membranes with dual targeting capability. Notably, CREATE remarkably confers simultaneous targeting ability to lung cancer cells and tumor-associated macrophages (TAMs). The pH/GSH-responsive design improves the release of dBET6 payload from nanoparticles to induce pronounced apoptosis of both cells, which synergistically inhibits tumor growth in both subcutaneous and orthotopic tumor-bearing mouse model. Furthermore, the efficient tumor inhibition is due to the direct elimination of lung cancer cells and TAMs, which remodels the tumor microenvironment. Taken together, the results elucidate the construction of a versatile nano-PROTAC enables to eliminate both lung cancer cells and TAMs, which opens a new avenue for efficient lung cancer therapy via PROTAC.

Intraneuronal sortilin aggregation relative to granulovacuolar degeneration, tau pathogenesis and sorfra plaque formation in human hippocampal formation.

Extracellular ß-amyloid (Aß) deposition and intraneuronal phosphorylated-tau (pTau) accumulation are the hallmark lesions of Alzheimer's disease (AD). Recently, "sorfra" plaques, named for the extracellular deposition of sortilin c-terminal fragments, are reported as a new AD-related proteopathy, which develop in the human cerebrum resembling the spatiotemporal trajectory of tauopathy. Here, we identified intraneuronal sortilin aggregation as a change related to the development of granulovacuolar degeneration (GVD), tauopathy, and sorfra plaques in the human hippocampal formation. Intraneuronal sortilin aggregation occurred as cytoplasmic inclusions among the pyramidal neurons, co-labeled by antibodies to the extracellular domain and intracellular C-terminal of sortilin. They existed infrequently in the brains of adults, while their density as quantified in the subiculum/CA1 areas increased in the brains from elderly lacking Aß/pTau, with pTau (i.e., primary age-related tauopathy, PART cases), and with Aß/pTau (probably/definitive AD, pAD/AD cases) pathologies. In PART and pAD/AD cases, the intraneuronal sortilin aggregates colocalized partially with various GVD markers including casein kinase 1 delta (Ck1δ) and charged multivesicular body protein 2B (CHMP2B). Single-cell densitometry established an inverse correlation between sortilin immunoreactivity and that of Ck1δ, CHMP2B, p62, and pTau among pyramidal neurons. In pAD/AD cases, the sortilin aggregates were reduced in density as moving from the subiculum to CA subregions, wherein sorfra plaques became fewer and absent. Taken together, we consider intraneuronal sortilin aggregation an aging/stress-related change implicating protein sorting deficit, which can activate protein clearance responses including via enhanced phosphorylation and hydrolysis, thereby promoting GVD, sorfra, and Tau pathogenesis, and ultimately, neuronal destruction and death.

Outcome of Endoscopic Transsphenoidal Surgery for Recurrent or Residual Pituitary Adenomas and Comparison to Non-Recurrent or Residual Cohort by Propensity Score Analysis.

Objective: To evaluate the long-term outcomes and safety of endoscopic transsphenoidal surgery (ETS) in recurrent and residual pituitary adenomas (rrPAs), as well as the predictors of gross total resection (GTR) and intraoperative CSF leakage. Furthermore, to compare outcomes and complications with non-rrPAs cohort. Methods: Clinical and radiological characteristics of patients with rrPAs who underwent ETS were collected between 2017 and 2020. Data of patients with non-rrPAs were collected from 2019 to 2020. Logistic regression analyses were performed to investigate the factors influencing gross total resection (GTR) and intraoperative CSF leakage. Between-group comparisons of outcomes and complications were performed through propensity score analysis. Results: We enrolled 73 patients with rrPAs. GTR was achieved in 41 (56.1%) cases; further, GTR or near-total resection was achieved in 93.2% of patients. The mean tumor volumes for GTR and non-GTR cases were 6.2 ±7.2 cm3 and 11.1 ±9.1 cm3, respectively. Multivariate regression analysis of the GTR rate in patients with rrPAs revealed that Knosp grade was an independent factor (odds ratio [OR] = 0.324; p=0.005). Moreover, previous transcranial surgery and non-functional pituitary adenomas were risk factors for intraoperative CSF leakage in patients with rrPAs (OR=6.450, p=0.019 and OR=7.472, p=0.012, respectively). After propensity score matching, There was no significant difference in the GTR rate between patients with rrPAs and patients with non-rrPAs. Contrastingly, patients with rrPAs had a higher rate of intraoperative CSF leakage and longer postoperative hospital stay than patients with non-rrPAs. During the follow-up, vision improved in 9 (22.0%) and 24 (62.5%) patients with rrPAs and non-rrPAs, respectively. Although there was a trend that reoperation of rrPAs involved a lower hypopituitarism recovery rate and biochemical remission rate, as well as a higher hypopituitarism rate, there was no statistically significant between-group difference. Conclusions: Knosp grade was an independent factor for GTR in endoscopic transsphenoidal surgery in patients with rrPAs. Previous transcranial surgery and non-functional PAs were risk factors for intraoperative CSF leakage. Although associated with longer hospital stay, rrPAs did not associate with lower GTR rate or more frequent postoperative complications than non-rrPAs cohort.

Senescent skeletal cells cross-talk with synovial cells plays a key role in the pathogenesis of osteoarthritis.

Osteoarthritis (OA) has been recognized as an age-related degenerative disease commonly seen in the elderly that affects the whole "organ" including cartilage, subchondral bone, synovium, and muscles. An increasing number of studies have suggested that the accumulation of senescent cells triggering by various stresses in the local joint contributes to the pathogenesis of age-related diseases including OA. In this review, we mainly focus on the role of the senescent skeletal cells (chondrocytes, osteoblasts, osteoclasts, osteocyte, and muscle cells) in initiating the development and progression of OA alone or through cross-talk with the macrophages/synovial cells. Accordingly, we summarize the current OA-targeted therapies based on the abovementioned theory, e.g., by eliminating senescent skeletal cells and/or inhibiting the senescence-associated secretory phenotype (SASP) that drives senescence. Furthermore, the existing animal models for the study of OA from the perspective of senescence are highlighted to fill the gap between basic research and clinical applications. Overall, in this review, we systematically assess the current understanding of cellular senescence in OA, which in turn might shed light on the stratified OA treatments.