Follicular CD8+ T cells promote immunoglobulin production and demyelination in multiple sclerosis and a murine model.

Emerging evidence underscores the importance of CD8+ T cells in the pathogenesis of multiple sclerosis (MS), but the precise mechanisms remain ambiguous. This study intends to elucidate the involvement of a novel subset of follicular CD8+ T cells (CD8+CXCR5+ T) in MS and an experimental autoimmune encephalomyelitis (EAE) murine model. The expansion of CD8+CXCR5+ T cells was observed in both MS patients and EAE mice during the acute phase. In relapsing MS patients, higher frequencies of circulating CD8+CXCR5+ T cells were positively correlated with new gadolinium-enhancement lesions in the central nervous system (CNS). In EAE mice, frequencies of CD8+CXCR5+ T cells were also positively correlated with clinical scores. These cells were found to infiltrate into ectopic lymphoid-like structures in the spinal cords during the peak of the disease. Furthermore, CD8+CXCR5+ T cells, exhibiting high expression levels of ICOS, CD40L, IL-21, and IL-6, were shown to facilitate B cell activation and differentiation through a synergistic interaction between CD40L and IL-21. Transferring CD8+CXCR5+ T cells into naïve mice confirmed their ability to enhance the production of anti-MOG35-55 antibodies and contribute to the disease progression. Consequently, CD8+CXCR5+ T cells may play a role in CNS demyelination through heightening humoral immune responses.

A General Strategy for Food Traceability and Authentication Based on Assembly-Tunable Fluorescence Sensor Arrays.

Food traceability and authentication systems play an important role in ensuring food quality and safety. Current techniques mainly rely on direct measurement by instrumental analysis, which is usually designed for one or a group of specific foods, not available for various food categories. To develop a general strategy for food identification and discrimination, a novel method based on fluorescence sensor arrays is proposed, composed of supramolecular assemblies regulated by non-covalent interactions as an information conversion system. The stimuli-responsiveness and tunability of supramolecular assemblies provided an excellent platform for interacting with various molecules in different foods. In this work, five sensor arrays constructed by supramolecular assemblies composed of pyrene derivatives and perylene derivatives are designed and prepared. Assembly behavior and sensing mechanisms are investigated systematically by spectroscopy techniques. The traceability and authentication effects on several kinds of food from different origins or grades are evaluated and verified by linear discriminant analysis (LDA). It is confirmed that the cross-reactive signals from different sensor units encompassing all molecular interactions can generate a unique fingerprint pattern for each food and can be used for traceability and authentication toward universal food categories with 100% accuracy.

IGF2BP3 enhances lipid metabolism in cervical cancer by upregulating the expression of SCD.

Cervical cancer (CC) is the most common gynecologic malignancy, which seriously threatens the health of women. Lipid metabolism is necessary for tumor proliferation and metastasis. However, the molecular mechanism of the relationship between CC and lipid metabolism remains poorly defined. We revealed the expression of IGF2BP3 in CC exceeded adjacent tissues, and was positively associated with tumor stage using human CC tissue microarrays. The Cell Counting Kit-8, colony formation assay, 5-ethynyl-2'-deoxyuridine assay, transwell assays, wound-healing assays, and flow cytometry assessed the role of IGF2BP3 in proliferation and metastasis of CC cells. Besides, exploring the molecular mechanism participating in IGF2BP3-driven lipid metabolism used RNA-seq, which determined SCD as the target of IGF2BP3. Further, lipid droplets, cellular triglyceride (TG) contents, and fatty acids were accessed to discover that IGF2BP3 can enhance lipid metabolism in CC. Moreover, RIP assay and methylated RNA immunoprecipitation experiments seeked the aimed-gene-binding specificity. Lastly, the IGF2BP3 knockdown restrained CC growth and lipid metabolism, after which SCD overexpression rescued the influence in vitro and in vivo using nude mouse tumor-bearing model. Mechanistically, IGF2BP3 regulated SCD mRNA m6A modifications via IGF2BP3-METTL14 complex, thereby enhanced CC proliferation, metastasis, and lipid metabolism. Our study highlights IGF2BP3 plays a crucial role in CC progression and represents a therapeutic latent strategy. It is a potential tactic that blocks the metabolic pathway relevant to IGF2BP3 with the purpose of treating CC.

Kapß2 Inhibits Perioperative Neurocognitive Disorders in Rats with Mild Cognitive Impairment by Reversing the Nuclear-Cytoplasmic Mislocalization of hnRNPA2/B1.

Harmful stimuli trigger mutations lead to uncontrolled accumulation of hnRNPA2/B1 in the cytoplasm, exacerbating neuronal damage. Kapß2 mediates the bidirectional transport of most substances between the cytoplasm and the nucleus. Kapß2 guides hnRNPA2/B1 back into the nucleus and restores its function, alleviating related protein toxicity. Here, we aim to explore the involvement of Kapß2 in neurodegeneration in rats with MCI following sevoflurane anesthesia and surgery. Firstly, novel object recognition test and Barnes maze were conducted to assess behavioral performances, and we found Kapß2 positively regulated the recovery of memory and cognitive function. In vivo electrophysiological experiments revealed that the hippocampal theta rhythm energy distribution was disrupted, coherence was reduced, and long-term potentiation was attenuated in MCI rats. LTP was greatly improved with positive modulation of Kapß2. Next, functional MRI and BOLD imaging will be employed to examine the AFLL and FC values of dynamic connectivity between the cortex and hippocampus of the brain. The findings show that regulating Kapß2 in the hippocampus region enhances functional activity and connections between brain regions in MCI rats. WB results showed that increasing Kapß2 expression improved the expression and recovery of cognitive-related proteins in the hippocampus of MCI rats. Finally, WB and immunofluorescence were used to examine the changes in hnRNPA2/B1 expression in the nucleus and cytoplasm after overexpression of Kapß2, and it was found that nucleocytoplasmic mis location was alleviated. Overall, these data show that Kapß2 reverses the nucleoplasmic misalignment of hnRNPA2/B1, which slows neurodegeneration towards dementia in MCI after sevoflurane anesthesia and surgery. Our findings may lead to new approaches for perioperative neuroprotection of MCI patients.

Nanoassemblies Based on a Cationic Perylene Diimide Derivative and Sodium Dodecyl Sulfate: A Simple Fluorescent Platform for Efficient Analysis of Aflatoxin B1.

Aflatoxin B1 (AFB1) is a kind of potently carcinogenic fungal metabolite in food threatening human health, and it is crucial and challenging to develop advanced nonimmune approaches for AFB1 determination. Addressing this challenge, we successfully constructed a nanoassembly (PdE-PDI/SDS) by noncovalently coupling a cationic perylene diimide derivative (PdE-PDI) and sodium dodecyl sulfate (SDS), exhibiting high-density charges and a specific surface area for rapid sensing of AFB1. The large electronic conjugate structure and rigid plane of PdE-PDI enable it to form more stable σ-π, π-π coordination, and hydrogen bonds with AFB1. Additionally, the introduction of SDS significantly amplifies noncovalent interactions and enhances the quenching efficiency of PdE-PDI toward AFB1. The proposed PdE-PDI/SDS exhibited excellent specificity to AFB1 and showed dosage-sensitive detection with detection limit as low as 0.74 ng mL-1. Finally, the PdE-PDI/SDS was successfully applied in cereal samples with good recoveries from 94.61 to 109.92%. To our knowledge, this is the first time a fluorescent strategy from the point of self-assembly for AFB1 determination is reported, which holds great promise for wide applications of perylene diimide derivative in food safety.

[Knockdown of IGF2BP2 inhibits colorectal cancer cell proliferation, migration and promotes tumor immunity by down-regulating MYC expression].

Objective To investigate the effect of insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) on the proliferation, migration and tumor immune microenvironment of colorectal cancer cells and its possible molecular mechanism. Methods The Cancer Genome Atlas (TCGA) database was used to analyze the expression levels of IGF2BP2 and MYC in colorectal cancer and adjacent tissues. The expression of IGF2BP2 in HCT-116 and SW480 human colorectal cancer cells was silenced by RNA interference (RNAi), and the silencing effect was detected by quantitative real-time PCR. After knocking down IGF2BP2, colony formation assay, CCK-8 assay and 5-ethynyl-2'-deoxyuridine (EdU) assay were employed to detect cell colony formation and proliferation ability. TranswellTM assay was used to detect cell migration ability. Quantitative real-time PCR was used to detect the mRNA expression of IGF2BP2, MYC, tumor necrosis factor-α (TNF-α), transforming growth factor-ß (TGF-ß) and interleukin-10 (IL-10). The protein expression of IGF2BP2 and MYC was detected by western blot. The binding ability of IGF2BP2 and MYC in HCT-116 cells was detected by quantitative real-time PCR after RNA immunoprecipitation. Results The results of TCGA database showed that the expression of IGF2BP2 and MYC in colorectal cancer tissues was significantly higher than that in adjacent tissues, and the survival time of colorectal cancer patients with high expression of IGF2BP2 was shorter. After silencing IGF2BP2, the viability, proliferation and migration of HCT-116 and SW480 cells were decreased. The mRNA expression of MYC, TGF-ß and IL-10 in IGF2BP2 knockdown group was significantly decreased, while the expression of TNF-α mRNA was increased. The expression of MYC protein and the stability of MYC mRNA were significantly decreased. RIP-qPCR results showed that IGF2BP2 could bind to MYC mRNA. Conclusion Knockdown of IGF2BP2 inhibits colorectal cancer cell proliferation, migration and promotes tumor immunity by down-regulating MYC expression.

IGF2BP2 promotes colorectal cancer progression by upregulating the expression of TFRC and enhancing iron metabolism.

BACKGROUND: Colorectal cancer (CRC) is one of the most common malignant tumors of the digestive system, ranking third for morbidity and mortality worldwide. At present, no effective control method is available for this cancer type. In tumor cells, especially iron metabolization, is necessary for its growth and proliferation. High levels of iron are an important feature to maintain tumor growth; however, the overall mechanism remains unclear. METHODS: We used western blotting, immunohistochemistry (IHC) and real-time quantitative PCR to analyze the expression of IGF2BP2 in cell lines and tissues. Further, RNA-sequencing, RNA immunoprecipitation and methylated RNA immunoprecipitation experiments explored the specific binding of target genes. Moreover, the RNA stability assay was performed to determine the half-life of genes downstream of IGF2BP2. In addition, the Cell Counting Kit-8, colony formation assay, 5-ethynyl-2'-deoxyuridine assay and flow cytometry were used to evaluate the effects of IGF2BP2 on proliferation and iron metabolism. Lastly, the role of IGF2BP2 in promoting CRC growth was demonstrated in animal models. RESULTS: We observed that IGF2BP2 is associated with iron homeostasis and that TFRC is a downstream target of IGF2BP2. Further, overexpression of TFRC can rescue the growth of IGF2BP2-knockdown CRC cells. Mechanistically, we determined that IGF2BP2 regulates TFRC methylation via METTL4, thereby regulating iron metabolism and promoting CRC growth. Furthermore, using animal models, we observed that IGF2BP2 promotes CRC growth. CONCLUSION: IGF2BP2 regulates TFRC mRNA methylation via METTL4, thereby regulating iron metabolism and promoting CRC growth. Our study highlights the key roles of IGF2BP2 in CRC carcinogenesis and the iron transport pathways.

miR-145 inhibits aerobic glycolysis and cell proliferation of cervical cancer by acting on MYC.

Nearly half a million women are diagnosed with cervical cancer (CC) each year, with the incidence of CC stabilizing or rising in low-income and middle-income countries. Cancer cells use metabolic reprogramming to meet the needs of rapid proliferation, known as the Warburg effect, but the mechanism of the Warburg effect in CC remains unclear. microRNAs (miRNAs) have a wide range of effects on gene expression and diverse modes of action, and they regulate genes for metabolic reprogramming. Dysregulation of miRNA expression leads to metabolic abnormalities in tumor cells and promotes tumorigenesis and tumor progression. In this study, we found that miR-145 was negatively correlated with metabolic reprogramming-related genes and prevented the proliferation and metastasis of CC cell lines by impeding aerobic glycolysis. A dual-luciferase reporter assay showed that miR-145 can bind to the 3'-untranslated region (3'-UTR) of MYC. Chromatin Immunoprecipitation-quantitative real-time PCR indicated that MYC was involved in the regulation of glycolysis-related genes. In addition, miR-145 mimics significantly suppressed the growth of CC cell xenograft tumor, prolonged the survival time of mice, and dramatically silenced the expression of tumor proliferation marker Ki-67. Therefore, the results suggested that miR-145 affects aerobic glycolysis through MYC, which may be a potential target for the treatment of CC.

Convolution neural network with batch normalization and inception-residual modules for Android malware classification.

Deep learning technology is changing the landscape of cybersecurity research, especially the study of large amounts of data. With the rapid growth in the number of malware, developing of an efficient and reliable method for classifying malware has become one of the research priorities. In this paper, a new method, BIR-CNN, is proposed to classify of Android malware. It combines convolution neural network (CNN) with batch normalization and inception-residual (BIR) network modules by using 347-dim network traffic features. CNN combines inception-residual modules with a convolution layer that can enhance the learning ability of the model. Batch Normalization can speed up the training process and avoid over-fitting of the model. Finally, experiments are conducted on the publicly available network traffic dataset CICAndMal2017 and compared with three traditional machine learning algorithms and CNN. The accuracy of BIR-CNN is 99.73% in binary classification (2-classifier). Moreover, the BIR-CNN can classify malware by its category (4-classifier) and malicious family (35-classifier), with a classification accuracy of 99.53% and 94.38%, respectively. The experimental results show that the proposed model is an effective method for Android malware classification, especially in malware category and family classifier.

Analysis of the function and mechanism of DIRAS1 in osteosarcoma.

BACKGROUND: Osteosarcoma is a prevalent malignant bone tumor with a tendency to metastasize to the lungs. In this study, we intend to detect the function and mechanism of DIRAS family GTPase 1 (DIRAS1) in osteosarcoma cells. METHODS: Expression level of DIRAS1 in osteosarcoma cells was analyzed by western blot. Cell location of DIRAS1 in osteosarcoma cells was detected by immunofluorescence. Small interfering RNAs (siRNA)-DIRAS1 and pcDNA3.1-DIRAS1 were employed to regulate DIRAS1 expression. The malignant behaviors of osteosarcoma cells were examined by cell counting kit-8, colony formation, transwell, and wound healing assays. The expression of related proteins was measured by western blot. ELISA and dot blot assays were used to detect the methylation level of m6A. Rescue assays were performed to detect the function of METTL3/METTL14 and DIRASI on osteosarcoma cells. RESULTS: DIRAS1 was located in the nucleus of osteosarcoma cells. Silencing of DIRAS1 in MG63 cells strengthened the proliferation, invasion and migration abilities, as well as blocked the apoptosis ability. Also, p-ERK expression was regulated by DIRAS1 expression, while p-AKT was not affected. Furthermore, DIRAS1 expression was suppressed by METTL3 or/and METTL14 treatment. Moreover, the inhibitory effect of DIRAS1 overexpression on HOS cells malignant behaviors can be reversed by METTL3 and METTL14 joint treatment. The reduced expression of p-ERK induced by DIRAS1 overexpression can be inversed by METTL3 and METTL14 co-treatment. CONCLUSIONS: Taken together, our findings illustrated that DIRAS1 regulated by METTL3 and METTL14 can obviously modulate the malignant behaviors of osteosarcoma cells by inactivating ERK pathway.

[METTL14 promotes the proliferation and migration of cervical cancer cells by up-regulating m6A Myc expression].

Objective To investigate the effect of methyltransferase-like 14 (METTL14) on the proliferation and metastasis of cervical cancer cells and its possible molecular mechanism. Methods The expression of METTL14 and Myc in cervical cancer tissues and normal tissues were analyzed using Gene Expression Omnibus (GEO) database and cervical cancer tissue microarray. The expression of METTL14 in HeLa and SiHa cells was silenced by small interfering RNA. After silencing the expression of METTL14 in cervical cancer HeLa and SiHa cells by RNA interference (RNAi), real-time quantitative PCR (qPCR) was used to verify the effect. CCK-8 assay, colony formation assay, 5-ethynyl-2'-deoxyuridine (EdU) assay were adopted to detect cell proliferation and colony forming ability. TranswellTM assay was employed to evaluate cell migration ability. After knocking out METTL14, Western blot was used to detect the protein expression of METTL14 and Myc. Methylated RNA immunoprecipitation-qPCR (MeRIP-qPCR) was applied to observe the expression of m6A Myc in HeLa cells in each group. Results GEO database analysis and cervical cancer tissue microarray staining showed that the expression of METTL14 and Myc in cervical cancer tissues was significantly higher than that in adjacent tissues, and the survival time of cervical cancer patients with high expression of METTL14 was shorter. Silencing METTL14 can significantly inhibit the cell viability, proliferation and migration of cervical cancer HeLa and SiHa cells, and its mechanism of action may be related to the up-regulation of the expression of m6A Myc by METTL14. Conclusion METTL14 promotes the proliferation and migration of cervical cancer cells by up-regulating the expression of m6A Myc.

HPV E6/E7 promotes aerobic glycolysis in cervical cancer by regulating IGF2BP2 to stabilize m6A-MYC expression.

Enhanced aerobic glycolysis constitutes an additional source of energy for tumor proliferation and metastasis. Human papillomavirus (HPV) infection is the main cause of cervical cancer (CC); however, the associated molecular mechanisms remain poorly defined, as does the relationship between CC and aerobic glycolysis. To investigate whether HPV 16/18 E6/E7 can enhance aerobic glycolysis in CC, E6/E7 expression was knocked down in SiHa and HeLa cells using small interfering RNA (siRNA). Then, glucose uptake, lactate production, ATP levels, reactive oxygen species (ROS) content, extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were evaluated. RNA-seq was used to probe the molecular mechanism involved in E6/E7-driven aerobic glycolysis, and identified IGF2BP2 as a target of E6/E7. The regulatory effect of IGF2BP2 was confirmed by qRT-PCR, western blot, and RIP assay. The biological roles and mechanisms underlying how HPV E6/E7 and IGF2BP2 promote CC progression were confirmed in vitro and in vivo. Human CC tissue microarrays were used to analyze IGF2BP2 expression in CC. The knockdown of E6/E7 and IGF2BP2 attenuated the aerobic glycolytic capacity and growth of CC cells, while IGF2BP2 overexpression rescued this effect in vitro and in vivo. IGF2BP2 expression was higher in CC tissues than in adjacent tissues and was positively correlated with tumor stage. Mechanistically, E6/E7 proteins promoted aerobic glycolysis, proliferation, and metastasis in CC cells by regulating MYC mRNA m6A modifications through IGF2BP2. We found that E6/E7 promote CC by regulating MYC methylation sites via activating IGF2BP2 and established a link between E6/E7 and the promotion of aerobic glycolysis and CC progression. Blocking the HPV E6/E7-related metabolic pathway represents a potential strategy for the treatment of CC.

Preparation of antibodies against TXR1 and construction of a new DNA tumor vaccine.

BACKGROUND: Taxol-resistance gene 1 (TXR1) is closely correlated with the paclitaxel resistance in the cancer chemotherapy. However, due to the lack of monoclonal antibodies (mAbs) with strong specificity and high sensitivity, little information is found about TXR1 target-related tumor therapy. METHODS: We developed an TXR1 recombinant DNA vaccine by inserting TXR1 DNA sequence into lysosome-associated membrane protein 1 (LAMP1). Adaptive immune responses were assessed by indirect enzyme-linked immunosorbent assay (ELISA), Enzyme-linked immunospot test (ELISpot), and cytotoxic T-lymphocyte (CTL) cytotoxicity. RESULTS: The pGEX4T-1-TXR1 reconstructed prokaryotic expression plasmid was constructed for producing high-purity TXR1 protein. Subsequently, a total of four mAbs for TXR1 and two PcAbs were successfully constructed and identified. We further found that TXR1 was highly expressed in breast cancer tissue than normal controls. Therefore, we constructed four tumor vectors, pVAX1-LAMP/TXR1, pVAX1-LAMP, pVAX1/TXR1 and pVAX1, for immunization. After three times of immunization, ELISpot data showed that single peptide 6,9,11 could stimulate T cells secreting IFN-γ in pVAX1-LAMP/TXR1 group. Moreover, the number of specific T cells and immune response effects significantly increased comparing to the pVAX1-LAMP control group. In addition, cytotoxicity showed that when the effect to target ratio was 40:l the killing effect of pVAX1-LAMP/TXR1 group was significantly higher than the pVAX1-TXR1 group. CONCLUSION: Our results provides new evidence for the TXR1 related tumor immunology and aids the early prevention of cancer.

BAP31 Promotes Tumor Cell Proliferation by Stabilizing SERPINE2 in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) patients are mostly diagnosed at an advanced stage, resulting in systemic therapy and poor prognosis. Therefore, the identification of a novel treatment target for HCC is important. B-cell receptor-associated protein 31 (BAP31) has been identified as a cancer/testis antigen; however, BAP31 function and mechanism of action in HCC remain unclear. In this study, BAP31 was demonstrated to be upregulated in HCC and correlated with the clinical stage. BAP31 overexpression promoted HCC cell proliferation and colony formation in vitro and tumor growth in vivo. RNA-sequence (RNA-seq) analysis demonstrated that serpin family E member 2 (SERPINE2) was downregulated in BAP31-knockdown HCC cells. Coimmunoprecipitation and immunofluorescence assays demonstrated that BAP31 directly binds to SERPINE2. The inhibition of SERPINE2 significantly decreased the BAP31-induced cell proliferation and colony formation of HCC cells and phosphorylation of Erk1/2 and p38. Moreover, multiplex immunohistochemistry staining of the HCC tissue microarray showed positive associations between the expression levels of BAP31, SERPINE2, its downstream gene LRP1, and a tumor proliferation marker, Ki-67. The administration of anti-BAP31 antibody significantly inhibited HCC cell xenograft tumor growth in vivo. Thus, these findings suggest that BAP31 promotes tumor cell proliferation by stabilizing SERPINE2 and can serve as a promising candidate therapeutic target for HCC.

Effects of extremely low-frequency electromagnetic fields on morphine-induced conditioned place preferences in rats.

In the present study, we examined the effects of extremely low-frequency (ELF) electromagnetic fields on morphine-induced conditioned place preferences in rats. During the conditioning phase (12 days), three groups of rats were placed in a sensory cue-defined environment paired with morphine (10mg/kg, i.p.) following exposure to either 20 Hz (1.80 mT) or 50 Hz (2.20 mT) or sham electromagnetic fields for 60 min/day, respectively, and were placed in another sensory cue-defined environment paired with physiological saline (1 ml/kg, i.p.) without exposure to electromagnetic fields. After finishing 12 days of conditioning, preference tests for the morphine-paired place were performed during a 10-day withdrawal period. The exposure to electromagnetic fields substantially potentiated morphine-induced place preferences in rodents, suggesting that ELF electromagnetic fields can increase the propensity for morphine-induced conditioned behaviors.