Oncolytic adenovirus-mediated expression of CCL5 and IL12 facilitates CA9-targeting CAR-T therapy against renal cell carcinoma.

Chimeric antigen receptor T-cell (CAR-T) is particularly prominent in hematological but not in solid tumors, mainly based on the complex tumor immune microenvironment. Oncolytic virus (OVs) is an emerging adjuvant therapy method. OVs may prime tumor lesions to induce anti-tumor immune response, thereby enhancing CAR-T cells functionality and possibly increasing response rates. Here, we combined CAR-T cells targeting carbonic anhydrase 9 (CA9) and an oncolytic adenovirus (OAV) carrying chemokine (C-C motif) ligand 5 (CCL5), cytokine interleukin-12 (IL12) to explore the anti-tumor effects of this combination strategy. The data showed that Ad5-ZD55-hCCL5-hIL12 could infect and replicate in renal cancer cell lines and induced a moderate inhibition of xenografted tumor in nude mice. IL12 mediated by Ad5-ZD55-hCCL5-hIL12 promoted the phosphorylation of Stat4 in CAR-T cells, induced CAR-T cells to secrete more IFN-γ. We also found that Ad5-ZD55-hCCL5-hIL-12 combined with CA9-CAR-T cells significantly increased the infiltration of CAR-T cells in tumor mass, prolonged the survival of the mice and restrained tumor growth in immunodeficient mice. Ad5-ZD55-mCCL5-mIL-12 could also increase CD45+CD3+T cell infiltration and prolong mice survival in immunocompetent mice. These results provided feasibility for the combination of oncolytic adenovirus and CAR-T cells, which demonstrated the sufficient potential and prospects of CAR-T for the treatment of solid tumors.

Temperature impacts fate of antibiotic resistance genes during vermicomposting of domestic excess activated sludge.

Effect of temperature on antibiotic resistance genes (ARGs) during vermicomposting of domestic excess sludge remains poorly understood. Vermicomposting experiment with excess sludge was conducted at three different temperatures (15 °C, 20 °C, and 25 °C) to investigate the fate of ARGs, bacterial community and their relationship in the process. The vermicomposting at 25 °C did not significantly attenuate the targeted ARGs relative to that at 15 °C and 20 °C. The dynamics of qnrA, qnrS, and tetM genes during vermicomposting at 15 °C and 20 °C followed the first-order kinetic model. Temperature remarkably impacted bacterial diversity of the final products with the lowest Shannon index at 25 °C. The presence of the genus (Aeromonas and Chitinophagaceae) at 25 °C may contribute to the rebound of the genes (qnrA, qnrS and tetM). The study indicates that 20 °C is a suitable vermicomposting temperature to simultaneously reach the highest removal efficiency of the ARGs and the good biostability of the final product.

Paracrine and epigenetic control of CAF-induced metastasis: the role of HOTAIR stimulated by TGF-ß1 secretion.

BACKGROUND: The communication between carcinoma associated fibroblasts (CAFs) and cancer cells facilitate tumor metastasis. In this study, we further underlying the epigenetic mechanisms of CAFs feed the cancer cells and the molecular mediators involved in these processes. METHODS: MCF-7 and MDA-MB-231 cells were treated with CAFs culture conditioned medium, respectively. Cytokine antibody array, enzyme-linked immunosorbent assay, western blotting and immunofluorescence were used to identify the key chemokines. Chromatin immunoprecipitation and luciferase reporter assay were performed to explore the transactivation of target LncRNA by CAFs. A series of in vitro assays was performed with RNAi-mediated knockdown to elucidate the function of LncRNA. An orthotopic mouse model of MDA-MB-231 was conducted to confirm the mechanism in vivo. RESULTS: Here we reported that TGF-ß1 was top one highest level of cytokine secreted by CAFs as revealed by cytokine antibody array. Paracrine TGF-ß1 was essential for CAFs induced EMT and metastasis in breast cancer cells, which is a crucial mediator of the interaction between stromal and cancer cells. CAF-CM significantly enhanced the HOTAIR expression to promote EMT, whereas treatment with small-molecule inhibitors of TGF-ß1 attenuated the activation of HOTAIR. Most importantly, SMAD2/3/4 directly bound the promoter site of HOTAIR, located between nucleotides -386 and -398, -440 and -452, suggesting that HOTAIR was a directly transcriptional target of SMAD2/3/4. Additionally, CAFs mediated EMT by targeting CDK5 signaling through H3K27 tri-methylation. Depletion of HOTAIR inhibited CAFs-induced tumor growth and lung metastasis in MDA-MB-231 orthotopic animal model. CONCLUSIONS: Our findings demonstrated that CAFs promoted the metastatic activity of breast cancer cells by activating the transcription of HOTAIR via TGF-ß1 secretion, supporting the pursuit of the TGF-ß1/HOTAIR axis as a target in breast cancer treatment.

Promoter DNA methylation is associated with KLF11 expression in epithelial ovarian cancer.

As a transforming growth factor-ß (TGF-ß)-inducible gene, the expression of Krüppel-like transcription factor 11 (KLF11) is altered in several types of cancer. In the current study, through using human 9K CpG island array, KLF11 was identified as one of hypermethylated genes in RAS-transformed ovarian T29H cells. Methylation of the KLF11 promoter was also observed in ovarian cancer tissue samples accompanied by significantly reduced KLF11 gene expression. Interestingly, the expression of SMAD2, SMAD3, and SMAD7 genes was reduced in the tumour, whilst no change was found in TGF-ß expression. Our data suggest a relationship between promoter DNA methylation and KLF11 gene expression in ovarian cancer tumorigenesis.

Experimental study on the failure process and modes of loess spoil slope induced by rainfall and engineering disturbance.

In this paper, indoor model tests were conducted using image analysis, pore pressure, and displacement measurement methods to investigate the failure evolution process and modes of loess spoil slopes with various components under the influence of rainfall and artificial excavation. The results of the experiments reveal that, under the action of rainfall, there are two types of cracks-to-failure modes for pure loess spoil slopes. One involves the formation of a large gully through the dominant channel, while the other is characterized by step-by-step retreating soil damage between cracks. The failure exhibits three distinct stages, and after failure, the slope angle is relatively large (>45°). The process of rainfall-induced destruction affecting loess spoil containing 25% coarse-grained content similarly unfolds in three stages, ultimately resulting in the formation of a regional landslide. This landslide typically encompasses a broader damage range compared to pure loess spoil, albeit with a shallower depth of damage. After the landslide stops and stabilizes, a tiny slope (45°) is created (<45°). The excavation at the toe of the slope induces loess spoil damage in a progressive multi-stage receding manner. This study provides a reference and basis for disaster prevention and warning of spoiled ground in loess areas.

Characteristics and mechanism of local scour reduction around spur dike using the collar in clear water.

To reduce the local scour around the spur dike, the U-shaped collar is proposed in this study. The influence of the collar's length, width, and porosity on the local scour reduction in clear water is studied by model tests and numerical simulations. Experimental studies show that the collar has a significant effect on reducing the local scour. The location of the maximum scour depth of the spur dike moves downstream. The width of the collar has the greatest impact on the reduction effect among the three selected factors, followed by the porosity and the length. Local scour reduction efficiency of the collar can reach 56.9%. Based on the regression analysis of the range and variety, a formula for predicting the reduction effect around the spur dike is put forward, and the deviation between the values by formula and that in experiments are within ± 4%. The characteristics of the flow field around the spur dike under constant conditions with a collar are studied via numerical simulation. The numerical simulation results show that compared to the case without collar, the flow velocities around the spur dike in cases with permeable collar and solid collar reduced by 45% and 25%, respectively, and the shear stresses reduced by 20% and 28.6%, respectively. The results of this study can provide a reference for local scour reduction using the solid collar or collar made of permeable materials such as gabions.

Analysis of the response subsidence and grouting treatment in the goaf of multi-layer inclined coal seam.

Based on existing researches, field drillings and numerical simulations are carried out in this paper to analyze the problems of subsidence control in the goaf of multi-layer inclined coal seam. Midas/GTS NX is used to build a three-dimensional calculation model of the goaf. A new method of using borehole data to check simulation parameters is proposed. The whole process of goaf excavation, construction of roadbed (pile foundation) and grouting treatment is analyzed. Analysis theory of different subgrade construction schemes and grouting treatment process on goaf is established. Response characteristics of displacement and equivalent stress and strain of goaf in multilayer inclined coal seam are obtained. A new method for analyzing the characteristics of the stress and deformation of the rock strata before and after grouting in the goaf under the conditions of different foundation schemes on the surface is provided in this research.

Oncolytic adenovirus-mediated expression of decorin facilitates CAIX-targeting CAR-T therapy against renal cell carcinoma.

Although chimeric antigen receptor T cell (CAR-T) therapy has been successful for hematological malignancies, it is less effective for solid tumors. The primary reason is that the immune microenvironment restricts CAR-T cells from infiltrating and proliferating in tumors. Oncolytic virotherapy has emerged as a novel immunogenic therapy to augment antitumor immune response. Here we combined an oncolytic adenovirus carrying decorin with a CAR-T targeting carbonic anhydrase IX (CAIX) to perform the antitumor activity for renal cancer cells. We found that OAV-Decorin combined with CAIX-CAR-T exhibited significantly reduced tumor burden, altered the composition of extracellular matrix (ECM) by inhibiting the distribution of collagen fibers, decreased the expression of TGF-ß in tumor cells, enhanced IFN-γ secretion, and obtained higher numbers of CAR-T cells. The combination treatment modality showed prolonged mice survival. The intratumoral injection of OAV-Decorin into tumor-bearing immunocompetent mice activated the inflammatory immune status and resulted in tumor regression. These data supported further investigation of the combination of OAV-Decorin and CAIX-CAR-T cells in solid tumors.

Functional expression of TRPA1 channel, TRPV1 channel and TMEM100 in human odontoblasts.

TRPA1 and TRPV1 channels respond to external stimulation as pain mediators and form a complex with a transmembrane protein TMEM100 in some tissues. However, their expression and interaction in dental pulp is unclear. To investigate the functional co-expression of TRPA1 channel, TRPV1 channel and TMEM100 in human odontoblasts (HODs), immunohistochemistry, immunofluorescence staining and Western blot were used to study their co-localization and expression in both native HODs and cultured HOD-like cells. Calcium imaging was used to detect the functional interaction between TRPA1 and TRPV1 channels. Immunohistochemistry and multiple immunofluorescence staining of tooth slices showed positive expression of TRPA1 channel, TRPV1 channel and TMEM100 mainly in the cell bodies of HODs, and TRPA1 channel presented more obvious immunofluorescence in the cell processes than TRPV1 channel and TMEM100. HALO software analysis showed that TRPA1 and TRPV1 channels were positively expressed in most TMEM100+ HODs and these three proteins were strongly correlated in HODs (P < 0.01). The protein expression levels of TRPA1 channel, TRPV1 channel and TMEM100 in HODs showed no significant difference (P > 0.05). Double immunofluorescence staining of cultured HOD-like cells visually demonstrated that TRPA1 and TRPV1 channel were both highly co-localized with TMEM100 with similar expressive intensity. Calcium imaging showed that there was a functional interaction between TRPA1 and TRPV1 channels in HOD-like cells, and TRPA1 channel might play a greater role in this interaction. Overall, we concluded that TRPA1 channel, TRPV1 channel and TMEM100 could be functionally co-expressed in HODs.

Activation of Transient Receptor Potential Ankyrin 1 and Vanilloid 1 Channels Promotes Odontogenic Differentiation of Human Dental Pulp Cells.

INTRODUCTION: Transient receptor potential ankyrin 1 (TRPA1) and vanilloid 1 (TRPV1) are thermosensitive channels that play an important role in thermal sensation or tooth pain by regulating intracellular Ca2+ concentration that is essential for pulp tissue repair. The aim of this study was to evaluate the role of TRPA1 and TRPV1 channels in the odontogenic differentiation of human dental pulp cells (HDPCs). METHODS: HDPCs were isolated from healthy human intact third molars and cultured in odontogenic differentiation medium. Gene and protein expression levels of TRPA1 and TRPV1 channels during the odontogenic differentiation of HDPCs were evaluated by real-time quantitative polymerase chain reaction and Western blot analysis. HDPCs were then treated with channel agonists or antagonists, and the expression levels of odontogenic markers dentin sialophosphoprotein (DSPP) and osteopontin (OPN) were examined. Alkaline phosphatase activity and alizarin red staining were also conducted to detect mineralization levels. RESULTS: Consistent with the mineralization degree and DSPP and OPN expression, messenger RNA and protein expression of TRPA1 and TRPV1 channels was up-regulated during the odontogenic differentiation of HDPCs. The application of TRPA1 or TRPV1 agonists increased the mineralized nodules of alizarin red staining and alkaline phosphatase activity and up-regulated the messenger RNA and protein expression of DSPP and OPN, respectively, with the highest values reached on the seventh day (P < .05). On the contrary, the mineralization level and DSPP and OPN expression could be suppressed by using the antagonists of these 2 channels. CONCLUSIONS: TRPA1 and TRPV1 channels not only showed up-regulated expression along with the odontogenic differentiation of HDPCs but also could affect the odontogenic differentiation by regulating intracellular Ca2+ concentration.

miR-595 suppresses cell proliferation and metastasis in hepatocellular carcinoma by inhibiting NF-κB signalling pathway.

MicroRNAs (miRNAs) have been proven to be critical regulators of cancer development. To date, many of them are still in urgent need of characterisation, and role of miR-595 in hepatocellular carcinoma (HCC) remains unknown. To better understand the mechanism of miR-595 in HCC development, a series of experiments were carried out to explore the effects of miR-595 on malignant behaviour in HCC. First, we found that miR-595 was downregulated in HCC tissues and cells and tightly associated with poor overall survival in HCC patients. Then, we further demonstrated that miR-595 inhibited cell proliferation, migration and invasion in vitro. Additionally, animal experimental results demonstrated that miR-595 inhibited HCC carcinogenesis in vivo. Moreover, we demonstrated that upregulation of miR-595 expression inhibited the NF-κB signalling pathway in HCC cells. To further uncover the molecular mechanism of miR-595 action on the NF-κB signalling pathway, we identified ABCB1 as a direct target of miR-595 through bioinformatics prediction and supported our results with luciferase assays. Finally, we showed that miR-595 inhibited the NF-κB pathway by suppressing ABCB1 expression in HCC cells. Taken together, our findings uncover a pivotal role for the miR-595/ABCB1/NF-κB axis in HCC development, and this novel axis may be a suitable target for diagnostic or therapeutic interventions in HCC.

Study on Properties Prediction and Braiding Optimization of Axial Braided Carbon/Carbon Composite.

It is well established that the microstructure has significant effects on the properties of axial braided C/C composites. In this study, a method coupling the homogenization method and finite element method (FEM) was proposed to predict the relationship between the microstructure characteristics and macroscopic properties. Based on the representative volume element (RVE) model, the periodic displacement boundary condition was introduced to predict the equivalent elastic properties of the RVE and component of C/C composite material, and the coefficient of thermal expansion (CTE) of the material was predicted by the energy prediction method. The predicted results were in good agreement with experimental results. By predicting the thermal and mechanical properties of the materials with different braiding spacing and fiber rod diameter, the variation of the properties with braiding spacing and fiber rod diameter was obtained. The research methods and results in this paper could provide important references for the optimization and rational application of composite materials.

Upregulation of Prickle2 Ameliorates Alzheimer's Disease-Like Pathology in a Transgenic Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder that has no effective therapies. Prickle planar cell polarity protein 2 (Prickle2), is an important cytoplasmic regulator of Wnt/PCP signaling. It has been reported that Prickle2 deficiency reduced neurite outgrowth levels in mouse N2a cells and led to autism-like behaviors and hippocampal synaptic dysfunction in mice. However, much less is known about the relationship of Prickle2 to AD pathogenesis. RT-qPCR, Western blot and IHC results showed that the mRNA and protein levels of Prickle2 were reduced in APP/PS1/Tau transgenic (3xTg) mice. Intravenous injection of Prickle2-overexpressing AAV-PHP.eB vectors improved the cognitive deficits in 3xTg mice. We also demonstrated that Prickle2 could repress oxidative stress and neuroinflammation, ameliorate the amyloid ß (Aß) plaque pathology and reduce Tau hyperphosphorylation in APP/PS1 mice. Further investigation of the mechanism of Prickle2 in AD revealed that Prickle2 inhibited Wnt/PCP/JNK pathway in vivo and in vitro. Our results suggest that Prickle2 might be a potential candidate for the diagnosis and treatment of AD.

Efficacy of an Oncolytic Adenovirus Driven by a Chimeric Promoter and Armed with Decorin Against Renal Cell Carcinoma.

Virus-targeted therapy for tumors can effectively prolong the survival rate of patients and has become a new trend for cancer biotherapy. Oncolytic adenovirus (OAd) can specifically replicate in tumor cells, allowing the therapeutic genes carried to be rapidly copied. As known, solid tumors are always hypoxic, and researchers often overlook a key point, the replication of OAd depends not only on its own activity but also on the cellular hypoxic environment in which the virus replicates. In this study, we constructed an OAd carrying Decorin, HRE-Ki67-Decorin, combining the Ki67 promoter upstreamed with hypoxia-response element (HRE) sequences to drive adenoviral E1A. The OAd HRE-Ki67-Decorin had better replication ability under hypoxic conditions, downregulated cellular immunosuppressed growth factor TGF-ß. In addition, HRE-Ki67-Decorin was potent in suppressing tumor growth and participated in the assembly of tumor extracellular matrix by expressing Decorin in subcutaneous renal cancer cell tumor models. Tumor sections from HRE-Ki67-Decorin-treated tissues had less collagen fibers and more spread of virus among tumor tissues. These results indicated that chimeric HRE-Ki67 promoter-regulated OAd carrying Decorin might be an effective anticancer treatment strategy.

Prognostic and clinicopathological significance of NRF2 expression in non-small cell lung cancer: A meta-analysis.

Nuclear factor erythroid 2-related factor 2 (NRF2) functions as a transcription factor and regulates a wide array of antioxidant and stress-responsive genes. NRF2 has been widely implicated in different types of cancers, but only limited studies concerning the relationship between NRF2 expression and tumour invasion or prognosis in lung cancer. Therefore, we conducted a meta-analysis to determine the prognostic value of NRF2 in patients with non-small cell lung cancer (NSCLC). The relationship between NRF2 expression in NSCLC patients and clinicopathological features was also investigated. Overall survival (OS) and treatment response rate were evaluated using STATA software. Twenty eligible articles with 2530 lung cancer patients were included in this meta-analysis. The results revealed that high expression level of NRF2 was associated with pathologic distant metastasis (odds ratio (OR) = 2.64, 95% confidence interval (CI) 1.62-4.31; P < 0.001), lymph node metastasis (OR = 2.14, 95% CI: 1.53-3.00; P < 0.001), and tumour node metastasis (TNM) stage (OR = 1.95, 95% CI: 1.52-2.49, P < 0.001). High NRF2 expression was associated with low treatment response rate in platinum-based chemotherapy (HR = 0.11, 95% CI 0.02-0.51; P = 0.005). High expression level of NRF2 is predictive for poor overall survival rate (HR = 1.86, 95% CI 1.44-2.41, P < 0.001) and poor progression-free survival (PFS) (HR = 2.27, 95% CI 1.26-4.09, P = 0.006). Compared to patients with a low level of NRF2 expression, patients with high NRF2 expression levels were associated with worse OS and PFS when given the chemotherapy or EGFR-TKI. Together, our meta-analysis results suggest that NRF2 can act as a potential indicator of NSCLC tumour aggressiveness and help the prognosis and design of a better treatment strategy for NSCLC patients.

MicroRNA-621 inhibits cell proliferation and metastasis in bladder cancer by suppressing Wnt/ß-catenin signaling.

Increasing evidence has shown that dysregulation of microRNA-621 (miR-621) is demonstrated to be associated with several cancers. However, the role of miR-621 in bladder cancer (BCa) remains unclear. Herein, we aimed to study the expression pattern, biological function, and molecular mechanism of miR-621 in BCa. First, we demonstrated that miR-621 was frequently downregulated in BCa tissues and cell lines compared with the adjacent normal BCa tissues and non-cancerous immortalized urothelial cell line. In addition, the expression of miR-621 was negatively correlated with overall survival of BCa patients. Functional experiments suggessted that miR-621 inhibited the proliferation and metastasis of BCa cells. Notably, dual-luciferase assay showed that miR-621 directly targeted the 3' UTR of TRIM29, which was frequently upregulated in BCa tissues and displayed inverse correlation with miR-621 expression. Furthermore, we demonstrated that miR-621 inhibited the proliferation and metastasis of BCa cells via Wnt/ß-catenin signaling pathway by targeting TRIM29. Our study suggested that the miR-621/TRIM29 axis inhibits the proliferation and metastasis of BCa cells via Wnt/ß-catenin signaling pathway and may have potential applications for development of BCa diagnosis or treatment.

A novel role mediated by adenoviral E1A in suppressing cancer through modulating decorin.

Oncolytic adenovirus is an emerging alternative to current therapeutics. The adenoviral E1A, the first protein expressed upon oncolytic adenoviral infection, has been identified as an antitumor agent, but the mechanisms of its tumor inhibition ability are unclear enough. Decorin is ubiquitous in the extracellular matrix (ECM), which regulates multiple functions through interaction with ECM. Here, we intended to explore the effects of adenoviral E1A on the tumor extracellular matrix during gene therapy. We demonstrated that reduced decorin expression was found in patients with lung cancer. The adenoviral E1A or a mutant adenoviral E1A with Rb-binding ability absent (E1A 30-60aa, 120-127aa deletion) could increase the expression of decorin and down-regulate VEGF, two members of tumor ECM, involved in both vasculogenesis and angiogenesis. E1A/mE1A-mediated suppressing the migration and invasion ability of tumor cells was depended on decorin. E1A interacted with decorin directly and induced the proteasomal degradation of VEGF. In addition, E1A or mE1A can inhibit tumor growth in a subcutaneous lung cancer xenograft model. It suggested that decorin might be a crucial mediator among ECM components for adenoviral E1A-mediated antitumor activities. These studies on adenovirus E1A provide a new mechanism for the emerging therapies of tumor gene therapy.

[Study on estimation of postmortem interval using FTIR in rat lung].

Fourier transform infrared spectroscopy (FTIR) was applied to study the decomposition process postmortem in rat lung tissue and provided a new method for the estimation of postmortem interval (PMI). The results demonstrated that, with the PMI increasing, the peak position of main absorbance bands showed no significant difference, but there was obvious variance of intensity: (1) The intensity of 1080 and 1241 cm(-1) related to nucleic acid tended to decrease. (2) The intensity ratio at Amide I (I1647) and II (I1541) decreased since death. The intensity of 1338 and 1313 cm(-1) increased lightly. (3) The intensity at 1460, 1400 and 1170 cm(-1) showed no significant difference. (4) A new band at 1120 cm(-1) appeared since 72 h after death and its intensity increased from 72 to 144 h postmortem. The authors' preliminary results suggest that it may be possible for FTIR to become an effective method to estimate the PMI in forensic practice.

Dynamic Expression Changes in the Transcriptome of the Prefrontal Cortex after Repeated Exposure to Cocaine in Mice.

Prefrontal cortex (PFC)-dependent functions, such as executive function, explicit learning, and memory, are negatively affected in cocaine abusers and experimental animal models of cocaine treatment. However, its molecular mechanisms are less understood. In the present study, we performed transcriptome profiling of the dynamic changes in the PFC after repeated cocaine administration in mice. We found 463, 14, and 535 differentially expressed genes (DEGs) at 2 h, 24 h, and 7 days, respectively, after the withdrawal of chronic cocaine treatment. Time-series correlation analysis identified 5 clusters of statistically significant expression patterns. The expression levels of DEGs in Clusters 1 and 5 exhibited a gradual or fluctuant decrease, Cluster 2 exhibited an initial increase followed by a decrease or return to the baseline level, and Clusters 3 and 4 exhibited a fluctuant increase in the expression of DEGs. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that genes related to oxidative phosphorylation, ribosomes, and neurodegenerative disorder were enriched in Cluster 1; genes related to the mitogen activated protein kinase (MAPK), transforming growth factor (TGF)-ß, insulin signaling, and circadian pathways were enriched in Cluster 2; genes related to plasticity-related pathways were enriched in Clusters 3 and 4; and genes related to the proteasome were enriched in Cluster 5. Our results suggest that maladaptive neural plasticity associated with psychostimulant dependence may be an ongoing degenerative process with dynamic changes in the gene network at different stages of withdrawal. Furthermore, it could be helpful to develop new therapeutic approaches according to different periods of abstinence.

Demethylation of c-MYB binding site mediates upregulation of Bdnf IV in cocaine-conditioned place preference.

Abnormal BDNF signaling contributes to the structural and behavioral plasticity induced by drugs of abuse. However, the mechanisms regulating expression of Bdnf in drug addiction remain elusive. In the present study, using the conditioned place preference (CPP) model, we showed that expression of Bdnf IV is upregulated in the nucleus accumbens (NAc) of conditioned animals while Bdnf I is upregulated in cocaine-treated mice irrespective of conditioning. The methylation level of a putative c-MYB binding site in the promoter region of Bdnf IV was significantly decreased in the NAc under cocaine CPP conditioning but remained unchanged without conditioning, concurrently with increased binding of c-MYB to this site. Exon IV promoter/luciferase reporter assays revealed that transactivation of Bdnf by c-MYB was blocked by methylation of this c-MYB binding site. Administration of methionine, a precursor of SAM, inhibited cocaine CPP, reversed demethylation of c-MYB binding site and induction of Bdnf IV expression by cocaine CPP. Our results imply that Bdnf IV demethylation at c-MYB binding site is involved in cocaine-triggered seeking behavior, whereas Bdnf I responds to the immediate pharmacological effects of cocaine.