Factors Influencing the Satisfaction of Second Language Learners of Chinese in Online Courses.

The study aims to investigate the relationship among the key factors affecting second language learners' satisfaction with online Chinese courses and their willingness to continue utilizing them by constructing a Model of Chinese Learners' Satisfaction in Online Courses. Additionally, the influence of participants' individual differences was also explored. A total of 203 second language learners of Chinese participated in the questionnaire survey, with 5 learners further participating in interviews. Learner expectations, learner perceived quality, and learner perceived value were identified as important factors influencing learner satisfaction and willingness to continue using the online course. The results of the questionnaire survey showed that (1) learner individual differences, such as age, Chinese proficiency, weekly study duration, and offline Chinese course experience, significantly influence learner satisfaction. (2) Learner expectations have a significant positive impact on perceived quality, while perceived quality positively affects perceived value. (3) Learner satisfaction significantly influences the willingness to continue using online courses. (4) The results of the interview revealed that most learners still prefer traditional offline courses, indicating that online teaching has several shortcomings and deficiencies. Overall, this study provides some scientific and reasonable decision-making references for improving online teaching methods, aiming to enhance learner satisfaction and promote the development of online education.

A Lysosome-Targeted Magnetic Nanotorquer Mechanically Triggers Ferroptosis for Breast Cancer Treatment.

Targeting ferroptosis has attracted exponential attention to eradicate cancer cells with high iron-dependent growth. Increasing the level of intracellular labile iron pool via small molecules and iron-containing nanomaterials is an effective approach to induce ferroptosis but often faces insufficient efficacy due to the fast drug metabolism and toxicity issues on normal tissues. Therefore, developing a long-acting and selective approach to regulate ferroptosis is highly demanded in cancer treatment. Herein, a lysosome-targeted magnetic nanotorquer (T7-MNT) is proposed as the mechanical tool to dynamically induce the endogenous Fe2+ pool outbreak for ferroptosis of breast cancer. T7-MNTs target lysosomes via the transferrin receptor-mediated endocytosis in breast cancer cells. Under the programmed rotating magnetic field, T7-MNTs generate torques to trigger endogenous Fe2+ release by disrupting the lysosomal membrane. This magneto-mechanical manipulation can induce oxidative damage and antioxidant defense imbalance to boost frequency- and time-dependent lipid peroxidization. Importantly, in vivo studies show that T7-MNTs can efficiently trigger ferroptosis under the magnetic field and play as a long-acting physical inducer to boost ferrotherapy efficacy in combination with RSL3. It is anticipated that this dynamic targeted strategy can be coupled with current ferroptosis inducers to achieve enhanced efficacy and inspire the design of mechanical-based ferroptosis inducers for cancer treatment.

[Anti-glioma mechanism of pterostilbene by regulating apoptosis and GSDME-mediated pyroptosis pathways: a study based on network pharmacology and experimental research].

This study aimed to explore the anti-glioma effect of natural compound pterostilbene(PTE) through regulating pyroptosis and apoptosis pathways, and to analyze the possible anti-glioma pathways and targets of PTE by network pharmacology and molecular docking. In this study, the action targets of PTE and the glioma targets were obtained by network pharmacology to construct a target network and a protein-protein interaction(PPI) network to predict the possible action targets of PTE against glioma. Molecular docking was performed on the core targets by AutoDock and the action pathways of PTE against glioma were predicted by enrichment analysis. In addition, the effect of PTE on the viability of U87MG and GL261 glioma cells was detected by CCK-8 assay. Clone formation assay and cell scratching assay were used to explore the effect of different concentrations of PTE on the proliferation and migration, respectively of glioma cells. Hoechst staining was used to observe PTE-induced apoptosis in glioma cells. The changes in mitochondrial membrane potential were detected by JC-1 staining. The pyroptosis-inducing effect of PTE on glioma cells was observed by inverted microscopy and lactate dehydrogenase(LDH) assay. Hoechst 33342/PI dual staining assay was performed to detect the integrity of glioma cell membranes. The expressions of pyroptosis and apoptosis-related proteins in glioma cells after PTE induction were determined by Western blot. In this study, 37 anti-glioma targets of PTE were obtained, and enrichment analysis suggested that PTE exerted anti-glioma effects through various signaling pathways including cancer pathway, proteoglycan in cancer, PI3K/AKT pathway, and apoptosis regulatory pathway. Molecular docking revealed that PTE had good binding activity with the main targets. Compared with the control group, PTE significantly reduced the viability as well as the proliferation, migration and adhesion abilities of U87MG and GL261 cells; it induced the apoptosis of the two glioma cells and the decrease of mitochondrial membrane potential in U87MG cells, and the effects increased with the increase of drug concentration. Compared with the conditions in the control group, glioma cells in the PTE group had increased pyroptosis-specific appearance and gradually increased LDH release; the number of PI positive cells was significantly elevated with the increase of PTE concentration as revealed by Hoechst 33342/PI staining; the expression levels of apoptosis-related factors cleaved PARP1 and B-cell lymphoma-2(Bcl-2) associated X(BAX) in the PTE group were markedly up-regulated, while the expression level of Bcl-2 was markedly down-regulated; the activation levels of pyroptosis-related proteins cleaved caspase-3 and gasdermin E-N(GSDME-N) had a remarkable rise in the PTE group, while no significant changes were found in the activation levels of gasdermin D-N(GSDMD-N) and cleaved caspase-1. In summary, PTE plays an anti-glioma role by inhibiting cell viability, proliferation, and migration and activating the caspase-3/GSDME-mediated pyroptosis pathway and mitochondrial apoptosis pathway.

Jinyinqingre Oral Liquid alleviates LPS-induced acute lung injury by inhibiting the NF-κB/NLRP3/GSDMD pathway.

Acute lung injury (ALI) is a prevalent and severe clinical condition characterized by inflammatory damage to the lung endothelial and epithelial barriers, resulting in high incidence and mortality rates. Currently, there is a lack of safe and effective drugs for the treatment of ALI. In a previous clinical study, we observed that Jinyinqingre oral liquid (JYQR), a Traditional Chinese Medicine formulation prepared by the Taihe Hospital, Affiliated Hospital of Hubei University of Medicine, exhibited notable efficacy in treating inflammation-related hepatitis and cholecystitis in clinical settings. However, the potential role of JYQR in ALI/acute respiratory distress syndrome (ARDS) and its anti-inflammatory mechanism remains unexplored. Thus, the present study aimed to investigate the therapeutic effects and underlying molecular mechanisms of JYQR in ALI using a mouse model of lipopolysaccharide (LPS)-induced ALI and an in vitro RAW264.7 cell model. JYQR yielded substantial improvements in LPS-induced histological alterations in lung tissues. Additionally, JYQR administration led to a noteworthy reduction in total protein levels within the BALF, a decrease in MPAP, and attenuation of pleural thickness. These findings collectively highlight the remarkable efficacy of JYQR in mitigating the deleterious effects of LPS-induced ALI. Mechanistic investigations revealed that JYQR pretreatment significantly inhibited NF-κB activation and downregulated the expressions of the downstream proteins, namely NLRP3 and GSDMD, as well as proinflammatory cytokine levels in mice and RAW2647 cells. Consequently, JYQR alleviated LPS-induced ALI by inhibiting the NF-κB/NLRP3/GSDMD pathway. JYQR exerts a protective effect against LPS-induced ALI in mice, and its mechanism of action involves the downregulation of the NF-κB/NLRP3/GSDMD inflammatory pathway.

Correlation between plasma long non-coding RNA MEG-3 and inflammatory cytokines in patients with diabetic nephropathy.

To identify the correlation between the plasma long non-coding RNA maternally expressed gene 3 (lncRNA MEG-3) and inflammatory cytokines in patients with diabetic nephropathy (DN) and search for a potential index for the diagnosis of DN. Quantitative real-time PCR (qPCR) was used to assess lncRNA MEG-3 expression. The levels of plasma cytokines were detected via enzyme-linked immunosorbent assay (-ELISA). 20 patients with type 2 diabetes (T2DM) and DN (DM+DN+ group), 19 patients with T2DM (DM+DN- group), and 17 healthy subjects (DM-DN- group) were finally enrolled. The expression of lncRNA MEG-3 was significantly upregulated in the DM+DN+ group compared to the DM+DN- group (p < 0.05) and the DM-DN- group (p < 0.001). Pearson's correlation analysis showed a positive correlation of lncRNA MEG-3 levels with cystatin C (Cys-C) (r = 0.468, p < 0.05), albumin-creatinine ratio (ACR) (r = 0.532, p < 0.05), and creatinine (Cr) (r = 0.468, p < 0.05), and a negative correlation with estimated glomerular filtration rate (eGFR) (r = -0.674, p < 0.01). Furthermore, the expression level of plasma lncRNA MEG-3 had a significantly positive correlation with the level of interleukin 1ß (IL-1ß) (r = 0.524, p < 0.05) and interleukin 18 (IL-18) (r = 0.230, p < 0.05). Binary regression analysis showed that lncRNA MEG-3 was a risk factor for DN with odds ration (OR) value of 1.71 (p < 0.05). The area under receiver operation characteristic (ROC) curve (AUC) of DN identified by lncRNA MEG-3 was 0.724. LncRNA MEG-3 was highly expressed in DN patients and showed a positive correlation with IL-1ß, IL-18, ACR, Cys-C, and Cr.

The role of vascular endothelial cells in tumor metastasis.

Vascular endothelial cells (VECs) are an integral component of the inner lining of blood vessels, and their functions are essential for the proper functioning of the vascular system. The tight junctions formed by VECs act as a significant barrier to the intravasation and extravasation of tumor cells (TCs). In addition to that, the proliferation, activation, and migration of VECs play a vital role in the growth of new blood vessels, a process known as tumor angiogenesis, which is closely related to the malignant progression of tumors. However, during tumor progression, VECs undergo endothelial-to-mesenchymal transition (EndMT), which further promotes tumor progression. Furthermore, VECs act as the first line of defense against effector immune cells and help prevent immune cells from infiltrating into tumor tissues. VECs also secrete various cytokines that can contribute to regulating the stemness of tumor stem cells. Thus, it has been increasingly recognized that dysfunction of VECs is one of the key driving forces behind tumor metastasis, and therapeutic strategies targeting VECs have the potential to be an effective means of antitumor therapy. This review aims to present a comprehensive overview of the role and mechanisms of VECs in regulating tumor progression and metastasis, providing insights into the possibilities for the development of novel antitumor therapies that target VECs.

Dual role of ANGPTL8 in promoting tumor cell proliferation and immune escape during hepatocarcinogenesis.

The interplay between hepatocellular carcinoma (HCC) cells and the tumor microenvironment is essential for hepatocarcinogenesis, but their contributions to HCC development are incompletely understood. We assessed the role of ANGPTL8, a protein secreted by HCC cells, in hepatocarcinogenesis and the mechanisms through which ANGPTL8 mediates crosstalk between HCC cells and tumor-associated macrophages. Immunohistochemical, Western blotting, RNA-Seq, and flow cytometry analyses of ANGPTL8 were performed. A series of in vitro and in vivo experiments were conducted to reveal the role of ANGPTL8 in the progression of HCC. ANGPTL8 expression was positively correlated with tumor malignancy in HCC, and high ANGPTL8 expression was associated with poor overall survival (OS) and disease-free survival (DFS). ANGPTL8 promoted HCC cell proliferation in vitro and in vivo, and ANGPTL8 KO inhibited the development of HCC in both DEN-induced and DEN-plus-CCL4-induced mouse HCC tumors. Mechanistically, the ANGPTL8-LILRB2/PIRB interaction promoted polarization of macrophages to the immunosuppressive M2 phenotype in macrophages and recruited immunosuppressive T cells. In hepatocytes, ANGPTL8-mediated stimulation of LILRB2/PIRB regulated the ROS/ERK pathway and upregulated autophagy, leading to the proliferation of HCC cells. Our data support the notion that ANGPTL8 has a dual role in promoting tumor cell proliferation and immune escape during hepatocarcinogenesis.

Boosting Glioblastoma Therapy with Targeted Pyroptosis Induction.

Glioblastoma (GBM) is a highly aggressive cancer that currently lacks effective treatments. Pyroptosis has emerged as a promising therapeutic approach for cancer, but there is still a need for new pyroptosis boosters to target cancer cells. In this study, it is reported that Aloe-emodin (AE), a natural compound derived from plants, can inhibit GBM cells by inducing pyroptosis, making it a potential booster for pyroptosis-mediated GBM therapy. However, administering AE is challenging due to the blood-brain barrier (BBB) and its non-selectivity. To overcome this obstacle, AE@ZIF-8 NPs are developed, a biomineralized nanocarrier that releases AE in response to the tumor's acidic microenvironment (TAM). Further modification of the nanocarrier with transferrin (Tf) and polyethylene glycol-poly (lactic-co-glycolic acid) (PEG-PLGA) improves its penetration through the BBB and tumor targeting, respectively. The results show that AE-NPs (Tf-PEG-PLGA modified AE@ZIF-8 NPs) significantly increase the intracranial distribution and tumor tissue accumulation, enhancing GBM pyroptosis. Additionally, AE-NPs activate antitumor immunity and reduce AE-related toxicity. Overall, this study provides a new approach for GBM therapy and offers a nanocarrier that is capable of penetrating the BBB, targeting tumors, and attenuating toxicity.

ANGPTL8 accelerates liver fibrosis mediated by HFD-induced inflammatory activity via LILRB2/ERK signaling pathways.

INTRODUCTION: High calorie intake is known to induce nonalcoholic fatty liver disease (NAFLD) by promoting chronic inflammation. However, the mechanisms are poorly understood. OBJECTIVES: This study examined the roles of ANGPTL8 in the regulation of NAFLD-associated liver fibrosis progression induced by high fat diet (HFD)-mediated inflammation. METHODS: The ANGPTL8 concentration was measured in serum samples from liver cancer and liver cirrhosis patients. ANGPTL8 knockout(KO) mice were used to induce disease models (HFD, HFHC and CCL4) followed by pathological staining, western blot and immunohistochemistry. Hydrodynamic injection of an adeno-associated virus 8 (AAV8) was used to establish a model for restoring ANGPTL8 expression specifically in ANGPTL8 KO mice livers. RNA-sequencing, protein array, Co-IP, etc. were used to study ANGPTL8's mechanisms in regulating liver fibrosis progression, and drug screening was used to identify an effective inhibitor of ANGPTL8 expression. RESULTS: ANGPTL8 level is associated with liver fibrogenesis in both cirrhosis and hepatocellular carcinoma patients. Mouse studies demonstrated that ANGPTL8 deficiency suppresses HFD-stimulated inflammatory activity, hepatic steatosis and liver fibrosis. The AAV-mediated restoration of liver ANGPTL8 expression indicated that liver-derived ANGPTL8 accelerates HFD-induced liver fibrosis. Liver-derived ANGPTL8, as a proinflammatory factor, activates HSCs (hepatic stellate cells) by interacting with the LILRB2 receptor to induce ERK signaling and increase the expression of genes that promote liver fibrosis. The FDA-approved anti-diabetic drug metformin, an ANGPTL8 inhibitor, inhibited HFD-induced liver fibrosis in vivo. CONCLUSIONS: Our data support that ANGPTL8 is a proinflammatory factor that accelerates NAFLD-associated liver fibrosis induced by HFD. The serum ANGPTL8 level may be a potential and specific diagnostic marker for liver fibrosis, and targeting ANGPTL8 holds great promise for developing innovative therapies to treat NAFLD-associated liver fibrosis.

Expression and Clinical Significance of Plasma miR-223 in Patients with Diabetic Nephropathy.

Background: MicroRNA-223 (miR-223) is associated with diabetes and kidney diseases and serves as a novel marker for diagnosing diabetic kidney disease (DKD). This study was conducted to investigate the plasma expression of miR-223 and its clinical significance in type 2 diabetes (T2DM) and diabetic nephropathy (DN) patients. Methods: In this research, 20 patients with T2DM and DN, 19 patients with T2DM, and 17 healthy volunteers were finally enrolled. miR-223 expression was detected by quantitative real-time PCR (qPCR), and the diagnostic value of miR-223 in DN was further analyzed. Results: miR-223 was downregulated in the DN group compared to that in the T2DM group (P=0.031) and the control group (P < 0.001). Pearson's correlation analysis showed a negative correlation of miR-223 levels with an albumin-creatinine ratio (ACR) (r = -0.481; P=0.044), urine ß2-microglobulin (ß2-MG) (r = -0.494; P=0.037), urine α1-microglobulin (α1-MG) (r = -0.537; P=0.022), creatinine (Cr) (r = -0.664; P < 0.01), cystatin C (Cyc-C) (r = -0.553; P=0.017), and glycosylated hemoglobin (HbA1c) (r = -0.761; P < 0.01). The findings of a binary regression analysis indicated that miR-223, ACR, Cr, and α1-MG were the risk factors for DN (OR: 2.019, 1.166, 1.031, and 1.031; all P < 0.05). Furthermore, miR-223 had a favorable diagnostic value for DN (AUC: 0.752; sensitivity: 0.722; specificity: 0.842) (2.5 was utilized as the diagnostic cutoff point). Conclusion: miR-223 was lowly expressed in DN patients, and the evaluation of miR-223 may be a good approach for diagnosing DN.

ANGPTL8 promotes adipogenic differentiation of mesenchymal stem cells: potential role in ectopic lipid deposition.

Background: Ectopic lipid deposition plays a promoting role in many chronic metabolic diseases. Abnormal adipogenic differentiation of mesenchymal stem cells (MSCs) is an important cause of lipid deposition in organs. Studies have shown that serum angiopoietin-like protein 8 (ANGPTL8) levels are increased in patients with many chronic metabolic diseases (such as type 2 diabetes, obesity, and hepatic steatosis), while the role of ANGPTL8 in ectopic lipid accumulation has not been reported. Methods: We used the Gene Expression Omnibus (GEO) database to analyze the expression of ANGPTL8 in subcutaneous adipose tissue of obese patients and qPCR to analyze the expression of ANGPTL8 in the liver of high-fat diet (HFD)-induced obese mice. To explore the potential roles of ANGPTL8 in the progression of ectopic lipid deposition, ANGPTL8 knockout (KO) mice were constructed, and obesity models were induced by diet and ovariectomy (OVX). We analyzed lipid deposition (TG) in the liver, kidney, and heart tissues of different groups of mice by Oil Red O, Sudan black B staining, and the single reagent GPO-PAP method. We isolated and characterized MSCs to analyze the regulatory effect of ANGPTL8 on Wnt/ß-Catenin, a key pathway in adipogenic differentiation. Finally, we used the pathway activator LiCl and a GSK3ß inhibitor (i.e., CHIR99021) to analyze the regulatory mechanism of this pathway by ANGPTL8. Results: ANGPTL8 is highly expressed in the subcutaneous adipose tissue of obese patients and the liver of HFD-induced obese mice. Both normal chow diet (NCD)- and HFD-treated ANGPTL8 KO male mice gained significantly less weight than wild-type (WT) male mice and reduced ectopic lipid deposition in organs. However, the female mice of ANGPTL8 KO, especially the HFD group, did not show differences in body weight or ectopic lipid deposition because HFD could induce estrogen overexpression and then downregulate ANGPTL8 expression, thereby counteracting the reduction in HFD-induced ectopic lipid deposition by ANGPTL8 deletion, and this result was also further proven by the OVX model. Mechanistic studies demonstrated that ANGPTL8 could promote the differentiation of MSCs into adipocytes by inhibiting the Wnt/ß-Catenin pathway and upregulating PPARγ and c/EBPα mRNA expression. Conclusions: ANGPTL8 promotes the differentiation of MSCs into adipocytes, suggesting that ANGPTL8 may be a new target for the prevention and treatment of ectopic lipid deposition in males.

ANGPTL8 is a negative regulator in pathological cardiac hypertrophy.

Pathological cardiac hypertrophy is an independent risk factor for heart failure and is considered a target for the treatment of heart failure. However, the mechanisms underlying pathological cardiac hypertrophy remain largely unknown. We aimed to investigate the role of angiopoietin-like protein 8 (ANGPTL8) in pathological cardiac hypertrophy. We found that serum ANGPTL8 levels were significantly increased in hypertensive patients with cardiac hypertrophy and in mice with cardiac hypertrophy induced by Ang II or TAC. Furthermore, the secretion of ANGPTL8 from the liver was increased during hypertrophic processes, which were triggered by Ang II. In the Ang II- and transverse aortic constriction (TAC)-induced mouse cardiac hypertrophy model, ANGPTL8 deficiency remarkably accelerated cardiac hypertrophy and fibrosis with deteriorating cardiac dysfunction. Accordingly, both recombinant human full-length ANGPTL8 (rANGPTL8) protein and ANGPTL8 overexpression significantly mitigated Ang II-induced cell enlargement in primary neonatal rat cardiomyocytes (NRCMs) and H9c2 cells. Mechanistically, the antihypertrophic effects of ANGPTL8 depended on inhibiting Akt and GSK-3ß activation, and the Akt activator SC-79 abolished the antihypertrophic effects of rANGPTL8 in vitro. Moreover, we demonstrated that ANGPTL8 directly bound to the paired Ig-like receptor PIRB (LILRB3) by RNA-seq and immunoprecipitation-mass screening. Remarkably, the antihypertrophic effects of ANGPTL8 were largely blocked by anti-LILRB3 and siRNA-LILRB3. Our study indicated that ANGPTL8 served as a novel negative regulator of pathological cardiac hypertrophy by binding to LILRB3 (PIRB) and inhibiting Akt/GSK3ß activation, suggesting that ANGPTL8 may provide synergistic effects in combination with AT1 blockers and become a therapeutic target for cardiac hypertrophy and heart failure.

A Bibliometric Analysis of Child Language During 1900-2021.

This study purposed to provide a bibliometric overview of child language (CL) research from 1900 to 2021 and identify major trends in CL. A total of 48,453 research articles related to the CL were identified from the Web of Science. Co-authorship, co-word, and co-citation analysis was conducted by using VOSviewer and CiteSpace. The following was analyzed: annual distribution of related papers; related disciplines; mainstream journals; geographical and institutional distribution; hot topics; keyword burst detection; and co-citation analysis of journals, authors, and references. Results showed that, under the impact of new empirical methods and new theories, the field of CL is undergoing great changes. Research hotspot and the research trends mainly concentrated on autism spectrum disorder, school readiness, oral language, reading comprehension, exposure, bilingualism, vocabulary, input, skills, kindergarten, cochlear implants, and intervention. More and more pieces of research focus on the individual difference in CL development and the importance of intervention in language education by typically developing children and some children with disabilities or language disorders. Besides, child second language acquisition also attracted a lot of attention. This bibliometric analysis is of great reference significance for researchers to understand the progress and discipline development trend in this field.

Perception of English Stress of Synthesized Words by Three Chinese Dialect Groups.

This study investigated the possible prosodic transfer influences native regional dialects may have in the perception of English lexical stress by speakers of three Chinese dialects [Beijing (BJ), Changsha (CS), and Guangzhou (GZ)] compared to 20 American English (AE) speakers. F0, duration, intensity, and vowel reduction were manipulated in nonce disyllabic words. Participants performed four-word sequence recall tasks to identify lexical stress location. They performed better with natural sounds than with manipulated words. This study focused on the performance differences in manipulating words. The results showed that all four-group members performed similarly processing F0 condition nonce words. BJ and CS participants were more accurate than GZ participants in duration and vowel reduction cues. Reaction time (RT) suggested that the processing time of acoustic cues differed significantly across language groups. The findings indicate that first language (L1) dialect effect is robust in second language (L2) stress perception tasks.

Acoustic Correlates of English Lexical Stress Produced by Chinese Dialect Speakers Compared to Native English Speakers.

English second language learners often experience difficulties in producing native-like English lexical stress. It is unknown which acoustic correlates, such as fundamental frequency (F0), duration, and intensity, are the most problematic for Chinese dialect speakers. The present study investigated the prosodic transfer effects of first language (L1) regional dialects on the production of English stress contrasts. Native English speakers (N = 20) and Chinese learners (N = 60) with different dialect backgrounds (Beijing, Changsha, and Guangzhou dialects) produced the same stimulus including both trochaic and iambic patterns. Results showed that (a) all participants produced the stressed syllable with greater values of F0, duration, and intensity; (b) Native speakers of English employed an exquisite combination of F0, duration, and intensity, while the dialect groups transfer their native prosody into their production of English lexical stress, resulting in the deviation or abnormality of acoustic cues. Results suggest that L1 native dialect background is considered as a potentially influential factor which may transfer in L2 speech encoding and decoding process.

Synthetic mRNA-based gene therapy for glioblastoma: TRAIL-mRNA synergistically enhances PTEN-mRNA-based therapy.

Glioblastoma (GBM) is characterized as having high molecular heterogeneity and complexity, which can be well revealed by genomic study. A truly effective treatment for GBM should flexibly address its heterogeneities, complexity, and strong drug resistance. This study was performed to explore the effectiveness of an mRNA-based therapeutic strategy using in vitro synthesized PTEN-mRNA and TRAIL-mRNA in tumor cells derived from PTEN-deletion patients. The PTEN gene alterations were revealed by whole-exome sequencing of three paired clinical GBMs and selected as the therapy target. Patient-derived primary glioblastoma stem cells (GBM2) and a DBTRG-cell-derived xenograft were used to detect mRNA's cytotoxicity in vitro and tumor suppression in vivo. Following the successful in vitro synthesis of PTEN-mRNA and TRAIL-mRNA, the combinational treatment of PTEN-mRNA and TRAIL-mRNA significantly suppressed tumor growth compared with treatment with PBS (96.4%), PTEN-mRNA (89.7%), and TRAIL-mRNA (84.5%). The combinational application of PTEN-mRNA and TRAIL-mRNA showed synergistic inhibition of tumor growth, and the JNK pathway might be the major mechanism involved. This study provided a basis for an mRNA-based therapeutic strategy to be developed into an effective patient-tailored treatment for GBM.

Intracranial delivery of synthetic mRNA to suppress glioblastoma.

Owing to messenger RNA's unique biological advantages, it has received increasing attention to be used as a therapeutic, known as mRNA-based gene therapy. It is critical to have an ideal strategy of mRNA gene therapy for glioma, which grows in a special environment. In the present study, we screened out a safe and efficient transfection reagent for intracranial delivery of synthetic mRNA in mouse brain. First, in order to analyze the effect of different transfection reagents on the intracranial delivery of mRNA, the synthetic luciferase mRNA was wrapped with two different transfection reagents and microinjected into the brain at the fixed point. The expression status of delivered mRNA was monitored by a small animal imaging system. The possible reagent-induced biological toxicity was evaluated by behavioral and blood biochemical measurements. Then, to test the therapeutic effect of our intracranial delivery mRNA model on glioma, synthetic modified tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) mRNA was used as an example of therapeutic application. This model demonstrated that synthetic mRNA could be successfully delivered into the brain using commercially available transfection reagents, and TransIT-mRNA showed better results than in vivo-jetPEI kit. This model can be applied in precise targeting and personalized gene therapy of glioma.

Palmitate impairs the autophagic flux to induce p62-dependent apoptosis through the upregulation of CYLD in NRCMs.

The most abundant saturated free fatty acid such as palmitate (PA), can accumulate in cardiomyocytes and induce lipotoxicity. CYLD is a known regulator in the development of cardiovascular disease and an important mediator of apoptosis. The role of CYLD in PA-induced cardiomyocyte apoptosis is not completely known. Here, we showed that PA treatment resulted in a concentration- and time-dependent effect on neonatal rat cardiomyocytes (NRCMs) apoptosis. PA impaired autophagy by significantly increasing the expression levels of LC3-II, Beclin 1, and also p62 in NRCMs. The autophagy flux was measured by detecting the fluorescence in the cells with Ad-mCherry-GFP-LC3B, a decrease in red puncta and a significant increase in yellow puncta in response to PA stimulation indicated that PA impairs the autophagic flux at the late stage of autophagosome-lysosome fusion. We further found knocked down of p62 by siRNA significantly decreased the expression level of cleaved caspase-3, decreased the apoptosis rate, also alleviated the loss of mitochondrial membrane potential, and decreased AIF and Cyt C releasing from mitochondria into the cytoplasm in the PA-treated NRCMs. From this, we considered that p62 accumulation was responsible for mitochondria-mediated apoptosis in PA-treated NRCMs. In addition, PA-induced a strong elevation of CYLD, siRNA-mediated knockdown of CYLD significantly antagonized PA-induced apoptosis and restored the autophagic flux in NRCMs. Knockdown of CYLD activation of the Wnt/ß-catenin pathway to restore the autophagic flux and reduce the accumulation of p62 in PA- stimulated NRCMs, while an inhibitor of the Wnt/ß-catenin pathway reversed this effect. Thus, our findings provide new insight into the molecular mechanism of PA toxicity in myocardial cells and suggest that CYLD may be a new therapeutic target for lipotoxic cardiomyopathy.

Human umbilical cord-derived mesenchymal stem cells improve the function of liver in rats with acute-on-chronic liver failure via downregulating Notch and Stat1/Stat3 signaling.

BACKGROUND: Effective treatments for acute-on-chronic liver failure (ACLF) are lacking. Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) have been applied in tissue regeneration and repair, acting through paracrine effects, cell fusion, and actual transdifferentiation. The present study was designed to investigate the therapeutic potential of hUC-MSCs in acute-on-chronic liver injury (ACLI) and ACLF rat models. METHODS: Wistar rats aged 6 weeks were intraperitoneally administered porcine serum (PS) at a dose of 0.5 mL twice per week for 11 weeks to generate an immune liver fibrosis model. After 11 weeks, rats with immune liver fibrosis were injected intravenously with lipopolysaccharide (LPS) to induce an ACLI model or combined LPS and D-galactosamine (D-GalN) to induce an ACLF model. The rats with ACLI or ACLF were injected intravenously with 2×106 hUC-MSCs, 4×106 hUC-MSCs, or 0.9% sodium chloride as a control. The rats were sacrificed at 1, 2, 4, and 6 weeks (ACLI rats) or 4, 12, and 24 h (ACLF rats). The blood and liver tissues were collected for biochemical and histological investigation. RESULTS: The application of hUC-MSCs in rats with ACLI and ACLF led to a significant decrease in the serum levels of ALT, AST, TBil, DBil, ALP, ammonia, and PT, with ALB gradually returned to normal levels. Inflammatory cell infiltration and collagen fiber deposition in liver tissues were significantly attenuated in ACLI rats that received hUC-MSCs. Inflammatory cell infiltration and apoptosis in liver tissues of ACLF rats that received hUC-MSCs were significantly attenuated. Compared with those in the rats that received 0.9% sodium chloride, a significant reduction in proinflammatory cytokine levels and elevated serum levels of hepatocyte growth factor (HGF) were found in ACLF rats that received hUC-MSCs. Furthermore, Notch, IFN-γ/Stat1, and IL-6/Stat3 signaling were inhibited in ACLI/ACLF rats that received hUC-MSCs. CONCLUSIONS: hUC-MSC transplantation can improve liver function, the degree of fibrosis, and liver damage and promote liver repair in rats with ACLI or ACLF, mediated most likely by inhibiting Notch signaling and reversing the imbalance of the Stat1/Stat3 pathway.