ABSTRACT
Objective: Skin fibrosis is a lesion in the dermis causing to itching, pain, and psychological stress. The gut microbiome plays as an essential role in skin diseases developments. We conducted a Mendelian randomization study to determine the causal association between the gut microbiome and skin fibrosis. Methods: We retrieved valid instrumental variables from the genome-wide association study (GWAS) files of the gut microbiome (n = 18,340) conducted by the MiBioGen consortium. Skin fibrosis-associated data were downloaded from the GWAS Catalog. Subsequently, a two-sample Mendelian randomization (MR) analysis was performed to determine whether the gut microbiome was related to skin fibrosis. A reverse MR analysis was also performed on the bacterial traits which were causally associated with skin fibrosis in the forward MR analysis. In addition, we performed an MR-Pleiotropy Residual Sum and Outlier analysis to remove outliers and a sensitivity analysis to verify our results. Results: According to the inverse variance-weighted estimation, we identified that ten bacterial traits (Class Actinobacteria, Class Bacteroidia, family Bifidobacteriaceae, family Rikenellaceae, genus Lachnospiraceae (UCG004 group), genus Ruminococcaceae (UCG013 group), order Bacteroidales, order Bifidobacteriales, genus Peptococcus and genus Victivallis) were negatively correlated with skin fibrosis while five bacterial traits (genus Olsenella, genus Oscillospira, genus Turicibacter, genus Lachnospiraceae (NK4A136group), and genus Sellimonas) were positively correlated. No results were obtained from reverse MR analysis. No significant heterogeneity or horizontal pleiotropy was observed in MR analysis. Objective conclusion: There is a causal association between the gut microbiome and skin fibrosis, indicating the existence of a gut-skin axis. This provides a new breakthrough point for mechanistic and clinical studies of skin fibrosis.
ABSTRACT
Chronic diabetic wounds remain a globally recognized clinical challenge, which occurs mainly due to the disturbances of wound microenvironmental induced by high concentrations of reactive oxygen species (ROS). Impairments in angiogenesis and inflammation in the wound microenvironment ultimately impede the normal healing process. Therefore, targeting macrophage and vascular endothelial cell dysfunction is a promising therapeutic strategy. In our study, we fabricated artificial composite scaffolds composed of naringin/carboxymethyl chitosan/sodium hyaluronate/silk fibroin (NG/CMCS/HA/SF) to promote wound healing. The NG/CMCS/HA/SF scaffold demonstrated favorable anti-inflammatory, anti-oxidative, and pro-angiogenic properties in both in vitro and in vivo experiments, effectively promoting the healing of diabetic wounds. The positive therapeutic effects observed indicate that the composite scaffolds have great potential in clinical wound healing applications.
Subject(s)
Chitosan , Diabetes Mellitus , Fibroins , Flavanones , Humans , Fibroins/pharmacology , Chitosan/pharmacology , Hyaluronic Acid/pharmacology , Tissue Scaffolds , Reactive Oxygen Species/pharmacology , Wound Healing , Glycosaminoglycans/pharmacology , MacrophagesABSTRACT
Background: Keloids are aberrant dermal wound healing characterized by invasive growth, extracellular matrix deposition, cytokine overexpression and easy recurrence. Many factors have been implicated as pathological causes of keloids, particularly hyperactive inflammation, tension alignment and genetic predisposition. S-Nitrosylation (SNO), a unique form of protein modification, is associated with the local inflammatory response but its function in excessive fibrosis and keloid formation remains unknown. We aimed to discover the association between protein SNO and keloid formation. Methods: Normal and keloid fibroblasts were isolated from collected normal skin and keloid tissues. The obtained fibroblasts were cultured in DMEM supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. The effects of DJ-1 on cell proliferation, apoptosis, migration and invasion, and on the expression of proteins were assayed. TurboID-based proximity labelling and liquid chromatography-mass spectrometry were conducted to explore the potential targets of DJ-1. Biotin-switch assays and transnitrosylation reactions were used to detect protein SNO. Quantitative data were compared by two-tailed Student's t test. Results: We found that DJ-1 served as an essential positive modulator to facilitate keloid cell proliferation, migration and invasion. A higher S-nitrosylated DJ-1 (SNO-DJ-1) level was observed in keloids, and the effect of DJ-1 on keloids was dependent on SNO of the Cys106 residue of the DJ-1 protein. SNO-DJ-1 was found to increase the level of phosphatase and tensin homolog (PTEN) S-nitrosylated at its Cys136 residue via transnitrosylation in keloids, thus diminishing the phosphatase activity of PTEN and activating the PI3K/AKT/mTOR pathway. Furthermore, Cys106-mutant DJ-1 is refractory to SNO and abrogates DJ-1-PTEN coupling and the SNO of the PTEN protein, thus repressing the PI3K/AKT/mTOR pathway and alleviating keloid formation. Importantly, the biological effect of DJ-1 in keloids is dependent on the SNO-DJ-1/SNO-PTEN/PI3K/AKT/mTOR axis. Conclusions: For the first time, this study demonstrated the effect of transnitrosylation from DJ-1 to PTEN on promoting keloid formation via the PI3K/AKT/mTOR signaling pathway, suggesting that SNO of DJ-1 may be a novel therapeutic target for keloid treatment.
ABSTRACT
The dysfunction of endothelial cells caused by hyperglycemia is observed as a decrease in neovascularization in diabetic wound healing. Studies have found that epidermal stem cells (EpiSCs) can promote the angiogenesis of full-thickness wounds. To further explain the therapeutic effect of EpiSCs, EpiSC-derived exosomes (EpiSC-EXOs) are considered the main substance contributing to stem cell effectivity. In our study, EpiSCs and EpiSC-EXOs were supplied to the dorsal wounds of db/db mice. Results showed that EpiSCs could colonize in the wound area and both EpiSCs and EpiSC-EXOs could accelerate diabetic wound healing by promoting angiogenesis. In vitro, persistent high glucose led to the malfunction and apoptosis of endothelial cells. The apoptosis induced by high glucose is due to excessive autophagy and was alleviated by EpiSC-EXOs. RNA sequencing of EpiSC-EXOs showed that miR200b-3p was enriched in EpiSC-EXOs and alleviated the apoptosis of endothelial cells. Synapse defective rho GTPase homolog 1 was identified the target of miR200b-3p and affected the phosphorylation of ERK to regulate intracellular autophagy and apoptosis. Furthermore, animal experiments validated the angiogenic effect of miR200b-3p. Collectively, our results verified the effect of EpiSC-EXOs on apoptosis caused by hyperglycemia in endothelial cells through the miR200b-3p/synapse defective rho GTPase homolog 1 /RAS/ERK/autophagy pathway, providing a theoretical basis for EpiSC in treating diabetic wounds.
ABSTRACT
Moderate inflammation is essential for standard wound healing. In pathological conditions, such as diabetes, protracted and refractory wounds are associated with excessive inflammation, manifested by persistent proinflammatory macrophage states. However, the mechanisms are still unclear. Herein, we perform a metabolomic profile and find a significant phenylpyruvate accumulation in diabetic foot ulcers. Increased phenylpyruvate impairs wound healing and augments inflammatory responses, whereas reducing phenylpyruvate via dietary phenylalanine restriction relieves uncontrolled inflammation and benefits diabetic wounds. Mechanistically, phenylpyruvate is ingested into macrophages in a scavenger receptor CD36-dependent manner, binds to PPT1, and inhibits depalmitoylase activity, thus increasing palmitoylation of the NLRP3 protein. Increased NLRP3 palmitoylation is found to enhance NLRP3 protein stability, decrease lysosome degradation, and promote NLRP3 inflammasome activation and the release of inflammatory factors, such as interleukin (IL)-1ß, finally triggering the proinflammatory macrophage phenotype. Our study suggests a potential strategy of targeting phenylpyruvate to prevent excessive inflammation in diabetic wounds.
Subject(s)
Diabetes Mellitus , NLR Family, Pyrin Domain-Containing 3 Protein , Humans , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Inflammasomes/metabolism , Wound Healing/physiology , InflammationABSTRACT
BACKGROUND: The composite transplantation of a split-thickness skin graft (STSG) combined with an acellular dermal matrix (ADM) is a promising repair method for full-thickness skin defects. Due to delayed vascularization of the ADM, no currently available engineered skin tissue is able to permanently cover full-thickness skin defects via a single-stage procedure. Epidermal stem cells (EpSCs) have been found to promote angiogenesis in the wound bed. Whether EpSCs can induce early angiogenesis of dermal substitutes and promote the survival of single-stage tissue-engineered skin transplantation needs to be further studied. METHODS: In vitro, rat vascular endothelial cells (RVECs) were treated with the supernatant of EpSCs cultured in ADM and stimulated for 48 h. RVECs were analysed by RNA sequencing and tube formation assays. For the in vivo experiment, 75 rats were randomly divided into five groups: ADM, ADM + EpSCs (AE), STSG, ADM + STSG (AS), and ADM + STSG + EpSCs (ASE) groups. The quality of wound healing was estimated by general observation and H&E and Masson staining. The blood perfusion volume was evaluated using the LDPI system, and the expression of vascular markers was determined by immunohistochemistry (IHC). RESULTS: The active substances secreted by EpSCs cultured in ADM promoted angiogenesis, as shown by tube formation experiments and RNA-seq. EpSCs promoted epithelialization of the ADM and vascularization of the ADM implant. The ASE group showed significantly increased skin graft survival, reduced skin contraction, and an improved cosmetic appearance compared with the AS group and the STSG control group. CONCLUSIONS: In summary, our findings suggest that EpSCs promote the formation of new blood vessels in dermal substitutes and support one-step transplantation of tissue-engineered skin, and thereby provide new ideas for clinical application.
Subject(s)
Skin, Artificial , Wound Healing , Rats , Animals , Endothelial Cells , Skin , Stem CellsABSTRACT
BACKGROUND: The development of therapeutic strategies to improve wound healing in individual diabetic patients remains challenging. Stem cell-derived exosomes represent a promising nanomaterial, and microRNAs (miRNAs) can be isolated from them. It is important to identify the potential therapeutic role of specific miRNAs, given that miRNAs can play a therapeutic role. METHODS: qPCR, flow cytometry, and western blotting were used to verify the effect of epidermal stem cell-derived exosomes (EpiSC-EXOs) on M2 macrophage polarization and SOCS3 expression. By screening key miRNAs targeting SOCS3 in EpiSC-EXOs by high-throughput sequencing, we verified the mechanism in vitro. Finally, an animal model was used to verify the effect of promoting healing. RESULTS: The use of EpiSC-EXOs reduced SOCS3 expression and promoted M2 macrophage polarization. The abundant miR-203a-3p present in the EpiSC-EXOs specifically bound to SOCS3 and activated the JAK2/STAT3 signaling pathway to induce M2 macrophage polarization. Treatment of the db/db mouse wound model with miR-203a-3p agomir exerted a pro-healing effect. CONCLUSIONS: Our results demonstrated that the abundant miR-203a-3p present in EpiSC-EXOs can promote M2 macrophage polarization by downregulating SOCS3 and suggested that diabetic wounds can obtain better healing effects through this mechanism.
Subject(s)
Diabetes Mellitus , Exosomes , MicroRNAs , Mice , Animals , Exosomes/genetics , Exosomes/metabolism , MicroRNAs/genetics , MicroRNAs/metabolism , Wound Healing/genetics , Stem Cells/metabolism , Diabetes Mellitus/metabolism , Macrophages/metabolism , Suppressor of Cytokine Signaling 3 Protein/genetics , Suppressor of Cytokine Signaling 3 Protein/metabolismABSTRACT
Background: Immunotherapy with checkpoint inhibitors usually has a low response rate in some cutaneous melanoma (CM) cases due to its cold nature. Hence, identification of hot tumors is important to improve the immunotherapeutic efficacy and prognoses of CMs. Methods: Fatty acid (FA) metabolism-related genes were extracted from the Gene Set Enrichment Analysis and used in the non-negative matrix factorization (NMF), copy number variation frequency, tumor mutation burden (TMB), and immune-related analyses, such as immunophenoscore (IPS). We generate a risk model and a nomogram for predicting patient prognoses and predicted the potential drugs for therapies using the Connectivity Map. Moreover, the NMF and the risk model were validated in a cohort of cases in the GSE65904 and GSE54467. At last, immunohistochemistry (IHC) was used for further validation. Results: Based on the NMF of 11 FA metabolism-related DEGs, CM cases were stratified into two clusters. Cluster 2 cases had the characteristics of a hot tumor with higher immune infiltration levels, higher immune checkpoint (IC) molecules expression levels, higher TMB, and more sensitivity to immunotherapy and more potential immunotherapeutic drugs and were identified as hot tumors for immunotherapy. The risk model and nomogram displayed excellent predictor values. In addition, there were more small potential molecule drugs for therapies of CM patients, such as ambroxol. In immunohistochemistry (IHC), we could find that expression of PLA2G2D, ACOXL, and KMO was upregulated in CM tissues, while the expression of IL4I1, BBOX1, and CIDEA was reversed or not detected. Conclusion: The transcriptome profiles of FA metabolism-related genes were effective for distinguishing CM into hot-cold tumors. Our findings may be valuable for development of effective immunotherapy for CM patients and for proposing new therapy strategies.
ABSTRACT
Background: Hypertrophic scar (HS) is a fibrotic cutaneous disease with few effective therapies. Lycorine is a drug with pro-apoptotic ability and anti-fibrosis potential. This study aimed to test whether lycorine could trigger the apoptosis of hypertrophic scar fibroblasts (HSFs) to inhibit HS formation. Methods: The proapoptotic and anti-fibrosis effects of lycorine on the viability and apoptosis of human primary HSFs and their reactive oxygen species (ROS) production as well as a rabbit ear model of HS were determined by CCK-8, flow cytometry, Western blot, immunofluorescence, transwell migration, collagen gel contraction assays. Results: Lycorine treatment selectively decreased the viability of HSFs, and induced their apoptosis, but not normal fibroblasts (NFs). Lycorine treatment increased the relative levels of Bax and cleaved PARP expression, cytochrome C cytoplasm translocation, but decreased Bcl-2, caspase-3 and caspase-9 expression, the mitochondrial membrane potential (MMP) in HSFs. Lycorine inhibited the migration and contraction of HSFs, and reduced the expression of collagen I, collagen III and α-SMA. Mechanistically, lycorine treatment stimulated high levels of ROS production, leading to apoptosis of HSFs while treatment with NAC, a ROS inhibitor, significantly mitigated or abrogated the pro-apoptotic and antifibrotic activity of lycorine in HSFs. Moreover, lycorine treatment mitigated the severity of HS in rabbit ears by inducing fibroblast apoptosis. Conclusion: These results indicate that lycorine has a potent anti-fibrotic activity and is a potential drug for intervention of HS.
ABSTRACT
Bladder cancer (BC) is one of the most common malignant urinary system tumors, and its prognosis is poor. In recent years, autophagy has been closely linked to the development of BC. Therefore, we investigated the potential prognostic role of autophagy-related long non-coding RNA (lncRNA) in patients with BC. We obtained the lncRNA information and autophagy genes, respectively, from The Cancer Genome Atlas (TCGA) data set and the human autophagy database (HADb) and performed a co-expression analysis to identify autophagy gene-associated lncRNAs. Then, we divided the data into training group and testing group. In the training group, 15 autophagy-related lncRNAs were found to have a prognostic value (AC026369.3, USP30-as1, AC007991.2, AC104785.1, AC010503.4, AC037198.1, AC010331.1, AF131215.6, AC084357.2, THUMPD3-AS1, U62317.4, MAN1B1-DTt, AC024060.1, AL662844.4, and AC005229.4). The patients were divided into low-risk group and high-risk group based on the prognostic lncRNAs. The overall survival (OS) time for the high-risk group was shorter than that for the low-risk group [risk ratio (hazard ratio, HR) = 1.08, 95% CI: 1.06-1.10; p < 0.0001]. Using our model, the defined risk value can predict the prognosis of a patient. Next, the model was assessed in the TCGA testing group to further validate these results. A total of 203 patients with BC were recruited to verify the lncRNA characteristics. We divided these patients into high-risk group and low-risk group. The results of testing data set show that the survival time of high-risk patients is shorter than that of low-risk patients. In the training group, the area under the curve (AUC) was more than 0.7, indicating a high level of accuracy. The AUC for a risk model was greater than that for each clinical feature alone, indicating that the risk value of a model was the best indicator for predicting the prognosis. Further training data analysis showed that the gene set was significantly enriched in cancer-related pathways, including actin cytoskeleton regulation and gap junctions. In conclusion, our 15 autophagy-related lncRNAs have a prognostic potential for BC, and may play key roles in the biology of BC.
ABSTRACT
BACKGROUND: Deep brain stimulation (DBS) of the subthalamic nucleus (STN) and the globus pallidus internus (GPi) are currently the most common and effective surgical targets for advanced Parkinson's disease (APD). Herein, we conducted a meta-analysis to evaluate the comprehensive efficacy of STN-DBS and GPi-DBS in patients with APD. METHODS: We conducted a systematic search for relevant articles written in English in the Cochrane Library, PubMed, and EMBASE databases through January 2020. Studies comparing the efficacy and clinical outcomes of GPi-DBS and STN-DBS for APD were included and analyzed. RESULTS: Ten eligible trials with a total of 857 patients were included in this meta-analysis. The results showed no significant difference between the STN-DBS and GPi-DBS groups in Unified Parkinson's Disease Rating Scale (UPDRS) III scores during the on and off-medication phases(SMD, 0.1; 95 % CI, -0.04 to 0.25; p = 0.17, on-med), (SMD,-0.12;95 % CI -0.37 to 0.13, p = 0.33,off-med). Dyskinesia scores and the activities of daily living (ADLs) scores during the on-medication phase showed significant differences in favor of GPi stimulation (SMD, 0.16; 95 % CI, 0.01-0.32; P < 0.05)/(SMD, 0.18; 95 % CI, 0.01-0.34; P < 0.05). The ADLs score during the off-medication phase showed no significant difference between the STN-DBS and GPi-DBS groups (SMD, -0.11; 95 % CI, -0.32-0.11; P = 0.33). The LED showed significant differences in favor of STN stimulation (SMD, -0.57; 95 % CI, -0.74-0.40; P < 0.00001). CONCLUSIONS: Both STN and GPi-DBS were equally effective in improving motor dysfunction. STN-DBS was superior for medication reduction, whereas GPi-DBS perhaps led to less dyskinesia and improved the postoperative ADLs (on-medication) in APD patients. Hence, the goals of DBS can be important in the target selection. More studies comparing the adverse events and quality of life between the two targets are needed.
Subject(s)
Deep Brain Stimulation/methods , Parkinson Disease/therapy , Dyskinesias/etiology , Dyskinesias/therapy , Globus Pallidus/physiology , Humans , Parkinson Disease/complications , Subthalamic Nucleus/physiologyABSTRACT
Tumor-immune crosstalk within the tumor microenvironment (TME) occurs at all stages of tumorigenesis. Tumor-associated M2 macrophages play a central role in tumor development, but the molecular underpinnings have not been fully elucidated. We demonstrated that M2 macrophages produce interleukin 1ß (IL-1ß), which activates phosphorylation of the glycolytic enzyme glycerol-3-phosphate dehydrogenase (GPD2) at threonine 10 (GPD2 pT10) through phosphatidylinositol-3-kinase-mediated activation of protein kinase-delta (PKCδ) in glioma cells. GPD2 pT10 enhanced its substrate affinity and increased the catalytic rate of glycolysis in glioma cells. Inhibiting PKCδ or GPD2 pT10 in glioma cells or blocking IL-1ß generated by macrophages attenuated the glycolytic rate and proliferation of glioma cells. Furthermore, human glioblastoma tumor GPD2 pT10 levels were positively correlated with tumor p-PKCδ and IL-1ß levels as well as intratumoral macrophage recruitment, tumor grade and human glioblastoma patient survival. These results reveal a novel tumorigenic role for M2 macrophages in the TME. In addition, these findings suggest possible treatment strategies for glioma patients through blockade of cytokine crosstalk between M2 macrophages and glioma cells.
Subject(s)
Brain Neoplasms/metabolism , Glioma/metabolism , Glycerolphosphate Dehydrogenase/metabolism , Macrophages/metabolism , Tumor Microenvironment/physiology , Animals , Brain Neoplasms/pathology , Carcinogenesis/metabolism , Cell Line, Tumor , Glioma/pathology , Glycolysis/physiology , Heterografts , Humans , Interleukin-1beta/metabolism , Mice , Mice, Nude , Receptor Cross-Talk/physiology , Signal Transduction/physiologyABSTRACT
INTRODUCTION: Calcium supplementation is one of the most important factors in maintaining the safety and efficacy of regional citrate anticoagulation (RCA) during continuous renal replacement therapy (CRRT). The aims of this study were to assess the determinants of calcium requirements in RCA-CVVH and to simplify the calcium supplementation approach. METHODS: Our study consisted of two parts. The first part was a discovery phase to determine the key factors of calcium supplementation. Twenty critically ill patients who required RCA-CVVH were enrolled in this part. Systemic citrate, total calcium, protein-bound calcium, and ionized calcium concentrations were serially measured using the traditional RCA protocol. A two-phase calcium supplementation protocol was then proposed, and algorithms were developed for calcium supplementation. The second part of the study was the validation phase. Another 97 critically ill patients were enrolled and were treated with RCA-CVVH using the new version of the calcium supplementation protocol. FINDINGS: The loss of calcium flux in the extracorporeal circuit and the increase in citrate-calcium complexes in vivo were the main determinants of the required calcium supplementation. In the CVVH mode, the rate of calcium infusion had to be reduced after systemic citrate level reached a steady state. With the aid of mathematical models, systemic calcium levels could be stably maintained in the normal range, and the frequencies of calcium monitoring were reduced. DISCUSSION: Calcium supplementation during RCA-CVVH undergoes two phases. We propose mathematical models to quantify the need for calcium supplementation, which enable individualization of the RCA prescription and simplify the management of RCA in the CVVH mode.
Subject(s)
Anticoagulants/therapeutic use , Calcium/therapeutic use , Citric Acid/therapeutic use , Continuous Renal Replacement Therapy/methods , Renal Dialysis/methods , Aged , Anticoagulants/administration & dosage , Anticoagulants/pharmacology , Calcium/pharmacology , Citric Acid/administration & dosage , Citric Acid/pharmacology , Female , Hemofiltration , Humans , Male , Middle AgedSubject(s)
Alzheimer Disease/complications , Cognition/drug effects , Cognitive Dysfunction/drug therapy , Insulin/administration & dosage , Administration, Intranasal , Cognitive Dysfunction/diagnosis , Cognitive Dysfunction/etiology , Humans , Memory and Learning Tests , Randomized Controlled Trials as Topic , Treatment OutcomeABSTRACT
Enhancer of Zeste 2 (EZH2) is the key enzymatic factor in Polycomb Repressive Complex 2 (PRC2), a transcriptional repressor which contributes to oncogenesis. Recent research has revealed the key role of aberrant EZH2 hyper-activity in human gliomas. Here, we examined the role of the lesser-known PRC2-associated PHD Finger Protein 19 (PHF19) in human glioma. We found that PHF19 transcript and protein levels were significantly elevated in human glioma tumors, which was negatively associated with expression of the anti-PHF19 microRNA miR-124a. miR-124a over-expression in the A172 and U251MG glioma cell lines and patient glioma cells suppressed PHF19 expression, EZH2 activation, and cell proliferation. However, miR-124a did not suppress cell proliferation with PHF19 silencing or mutation. Knockdown of PHF19 suppressed EZH2 phosphorylation and proliferation of glioma cells. Co-immunoprecipitation confirmed that PHF19 forms the PRC2 with EZH2, EED, and SUZ12. In a nude murine model, subcutaneous and orthotopic xenograft tumor growth was significantly inhibited by miRNA-124a or PHF19 shRNA. In conclusion, miR-124a suppresses PHF19 over-expression, EZH2 hyper-activation, and aberrant glioma cell proliferation. Targeting PHF19 via miR-124a agomir therapy may block aberrant EZH2 hyper-activity in these tumors.
Subject(s)
Enhancer of Zeste Homolog 2 Protein/metabolism , Glioma/metabolism , Glioma/pathology , MicroRNAs/genetics , Nuclear Proteins/genetics , Animals , Cell Proliferation/drug effects , Cell Proliferation/genetics , DNA-Binding Proteins , Glioma/genetics , Humans , Mice , Nuclear Proteins/biosynthesis , Nuclear Proteins/metabolism , Transcription Factors , Xenograft Model Antitumor AssaysABSTRACT
Patient-derived xenografts (PDX) provide in vivo glioblastoma (GBM) models that recapitulate actual tumors. Orthotopic tumor xenografts within the mouse brain are obtained by injection of GBM stem-like cells derived from fresh surgical specimens. These xenografts reproduce GBM's histologic complexity and hallmark biological behaviors, such as brain invasion, angiogenesis, and resistance to therapy. This method has become essential for analyzing mechanisms of tumorigenesis and testing the therapeutic effect of candidate agents in the preclinical setting. Here, we describe a protocol for establishing orthotopic tumor xenografts in the mouse brain with human GBM cells.
Subject(s)
Disease Models, Animal , Glioblastoma/pathology , Heterografts , Animals , Brain Neoplasms/diagnostic imaging , Brain Neoplasms/pathology , Fluorescent Antibody Technique , Glioblastoma/diagnostic imaging , Humans , Magnetic Resonance Imaging/methods , Mice , Microscopy, FluorescenceABSTRACT
Delivery of imaging agents to brain glioma is challenging because the blood-brain barrier (BBB) functions as a physiological checkpoint guarding the central nervous system from circulating large molecules. Moreover, the ability of existing probes to target glioma has been insufficient and needs to be improved. In present study, PEG-based long circulation, CdSe/ZnS quantum dots (QDs)-based nanoscale and fluorescence, asparagines-glycine-arginine peptides (NGR)-based specific CD13 recognition were integrated to design and synthesize a novel nanoprobe by conjugating biotinylated NGR peptides to avidin-PEG-coated QDs. Our data showed that the NGR-PEG-QDs were nanoscale with less than 100 nm and were stable in various pH (4.0~8.0). These nanomaterials with non-toxic concentrations could cross the BBB and target CD13-overexpressing glioma and tumor vasculature in vitro and in vivo, contributing to fluorescence imaging of this brain malignancy. These achievements allowed groundbreaking technological advances in targeted fluorescence imaging for the diagnosis and surgical removal of glioma, facilitating potential transformation toward clinical nanomedicine.
Subject(s)
Blood-Brain Barrier , Brain Neoplasms/diagnostic imaging , Glioma/diagnostic imaging , Oligopeptides/chemistry , Quantum Dots/chemistry , Animals , Brain Neoplasms/blood supply , Fluorescence , Glioma/blood supply , Male , Neoplasms, Experimental/blood supply , Neoplasms, Experimental/diagnostic imaging , Optical Imaging , Rats , Rats, Sprague-DawleyABSTRACT
Heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) plays an important role in influence of pre-messenger RNA (pre-mRNA) processing and mRNA metabolism and transportation in cells. Increasing evidence indicates that hnRNP A2/B1 played an important role in development and progression of various human cancers. Forty cases of normal and human glioma tissue samples were analyzed using immunohistochemistry to reveal the expression of hnRNP A2/B1 protein in the samples. Then, knockdown of hnRNP A2/B1 expression induced by RNA interference (RNAi) method was used to analyze the role of hnRNP A2/B1 in glioblastoma cell viability, adhesion, migration, invasion, and chemoresistance for temozolomide (TMZ). The data showed that hnRNP A2/B1 protein was overexpressed in glioma tissue specimens and associated with advanced glioma grades. Knockdown of hnRNP A2/B1 could reduce glioblastoma cell viability, adhesion, migration, invasion, and chemoresistance for TMZ capacity, but induced tumor cells to apoptosis and reactive oxygen species (ROS) generation in glioma U251 and SHG44 cells. Molecularly, hnRNP A2/B1 knockdown reduced expression of phospho-STAT3 and MMP-2. Detection of hnRNP A2/B1 expression may be useful as a biomarker for prediction of glioma progression and knockdown of hnRNP A2/B1 expression as a novel strategy in future control of glioblastoma in clinic.
Subject(s)
Brain Neoplasms/pathology , Gene Knockdown Techniques , Glioblastoma/pathology , Heterogeneous-Nuclear Ribonucleoprotein Group A-B/metabolism , Brain Neoplasms/metabolism , Cell Adhesion/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Dacarbazine/analogs & derivatives , Dacarbazine/pharmacology , Drug Resistance, Neoplasm/drug effects , Female , Glioblastoma/metabolism , Humans , Immunohistochemistry , Male , Matrix Metalloproteinases/metabolism , Middle Aged , Neoplasm Invasiveness , STAT3 Transcription Factor/metabolism , Temozolomide , Tumor Stem Cell AssayABSTRACT
Previous evidence suggests that the humanized anti-VEGF antibody bevacizumab increases thrombosis risk in glioma patients. Here, we comprehensively assessed the risk of adverse vascular events in adult glioma patients receiving bevacizumab therapy. Systematic searches of MEDLINE, EMBASE, and the Cochrane Library were conducted to find prospective phase II/III clinical trials on adult bevacizumab-treated glioma patients and non-bevacizumab-treated controls that reported data on adverse vascular events. Four high-quality trials were finally included in the systematic review, scoring greater than or equal to 7/8 on the Newcastle-Ottawa Scale. Three trials provided sufficient data for four meta-analytical comparisons between bevacizumab-treated and control groups of newly diagnosed glioblastoma multiforme (GBM) patients: all-cause discontinuation, thrombocytopenia, deep vein thrombosis (DVT), and pulmonary embolism. None of these adverse outcomes were found to be significantly different between bevacizumab-treated and control groups (P > 0.05); however, there was a trend toward significance with regard to bevacizumab therapy and the risk of pulmonary embolism (P = 0.07). As there was a trend toward significance with regard to bevacizumab therapy and the risk of pulmonary embolism, anticoagulation may be advisable in certain newly diagnosed adult GBM patients who display a history of thromboembolism and/or more serious risk factors for thromboembolic events.