Ubiquitin-specific peptidase 14 maintains estrogen receptor α stability via its deubiquitination activity in endometrial cancer.

USP14 deubiquitinates ERα to maintain its stability in ECEndometrial cancer (EC) is one of the common gynecological malignancies of which the incidence has been rising for decades. It is considered that continuously unopposed estrogen exposure is the main risk factor for EC initiation. Thus, exploring the modulation of estrogen/estrogen receptor α (ERα) signaling pathway in EC would be helpful to well understand the mechanism of EC development and find the potential target for EC therapy. Ubiquitin-specific peptidase 14 (USP14), a member of the proteasome-associated deubiquitinating enzyme family, plays a crucial role in a series of tumors. However, the function of USP14 in EC is still elusive. Here, our results have demonstrated that USP14 is highly expressed in EC tissues compared with that in normal endometrial tissues, and higher expression of USP14 is positively correlated with poor prognosis. Moreover, USP14 maintains ERα stability through its deubiquitination activity. Our results further demonstrate that USP14 depletion decreases the expression of ERα-regulated genes in EC-derived cell lines. Moreover, knockdown of USP14 or USP14-specific inhibitor treatment significantly suppresses cell growth and migration in EC cell lines or in mice. We further provide the evidence to show that the effect of USP14 on EC cell growth, if not all, at least is partially related to ERα pathway. Our study provides new sights for USP14 to be a potential therapeutic target for the treatment of EC, especially for EC patients with fertility preservation needs.

MOF upregulates the estrogen receptor α signaling pathway by its acetylase activity in hepatocellular carcinoma.

The histone acetyltransferase MOF (KAT8) is mainly involved in the acetylation of histone H4 at lysine 16 (H4K16) and some non-histone proteins. The MOF expression level is significantly reduced in many cancers, however the biological function of MOF and its underlying mechanism are still elusive in hepatocellular carcinoma (HCC). Estrogen receptor α (ERα) has been considered as a tumor suppressor in HCC. Here, we demonstrated that MOF expression is significantly reduced in HCC samples, and is positively correlated with that of ERα. MOF interacts with ERα, and participates in acetylation of ERα at K266, K268, K299, thereby inhibiting ERα ubiquitination to maintain the stability of ERα. In addition, MOF participates in the upregulation of ERα-mediated transactivation. Depletion of MOF significantly promotes cell growth, migration, and invasion in HCC cell lines. Taken together, our results provide new insights to understand the mechanism underlying the modulation function of MOF on ERα action in HCC, suggesting that MOF might be a potential therapeutic target for HCC.

Splicing factor PRPF6 upregulates oncogenic androgen receptor signaling pathway in hepatocellular carcinoma.

Androgen receptor (AR) signaling is considered to be crucial for the pathogenesis of hepatocellular carcinoma (HCC) with obvious sexual dimorphism. Pre-mRNA processing factor 6 (PRPF6) was identified as a coactivator of AR. However, the molecular mechanism underlying the modulation function of PRPF6 on AR-mediated transcriptional activity in HCC needs to be further clarified. In this study, we analyzed data from The Cancer Genome Atlas to show that PRPF6 is highly expressed in HCC. . Our data indicated that PRPF6 interacts with AR/AR splice variants (AR-Vs) and upregulates AR/AR splice variant 7-mediated transcriptional activity even without dihydrotestosterone treatment. We observed that AR is obviously induced by androgen treatment and is mainly expressed in the nucleus in HCC-derived cell lines. Moreover, overexpression of PRPF6 enhances AR expression accompanied with the increase of AR-Vs expression. We provided evidence that PRPF6 participates in upregulating AR self-transcription. PRPF6 facilitates the recruitment of AR to the androgen responsive element region of the AR gene. Finally, PRPF6 depletion inhibits cell proliferation in HCC cells and mouse xenografts. Taken together, our results suggest that PRPF6 as a splicing factor enhances AR self-transcription, thereby coactivating oncogenic AR/AR-Vs actions in HCC.

A new tumor suppressor LncRNA ADAMTS9-AS2 is regulated by DNMT1 and inhibits migration of glioma cells.

Growing number of long noncoding RNAs (lncRNAs) are emerging as new modulators in cancer origination and progression. A lncRNA, ADAM metallopeptidase with thrombospondin type 1 motif, 9 (ADAMTS9) antisense RNA 2 (ADAMTS9-AS2), with unknown function, is the antisense transcript of tumor suppressor ADAMTS9. In the present study, we investigated the expression pattern and functional role of ADAMTS9-AS2 in glioma by using real-time PCR and gain-/loss-of-function studies. The results showed that the ADAMTS9-AS2 expression was significantly downregulated in tumor tissues compared with normal tissues and reversely associated with tumor grade and prognosis. Multivariate analysis of the prognosis factors showed that low ADAMTS9-AS2 expression was a significant independent predictor of poor survival in glioma. Overexpression of ADAMTS9-AS2 resulted in significant inhibition of cell migration in glioma, whereas knockdown of ADAMTS9-AS2 showed the opposite effect. We also found that ADAMTS9-AS2 expression was negatively correlated with DNA methyltransferase-1 (DNMT1). In addition, DNMT1 knockdown led to remarkable enhancement of ADAMTS9-AS2 expression. By 5-aza-dC treatment, the ADAMTS9-AS2 expression was also reactivated. The results suggested that ADAMTS9-AS2 is a novel tumor suppressor modulated by DNMT1 in glioma. LncRNA ADAMTS9-AS2 may serve as a potential biomarker and therapeutic target for glioma.

Tagging SNPs in the MTHFR gene and risk of ischemic stroke in a Chinese population.

Stroke is currently the leading cause of functional impairments worldwide. Folate supplementation is inversely associated with risk of ischemic stroke. Methylenetetrahydrofolate reductase (MTHFR) is an important enzyme involved in folate metabolism. The aim of this study is to examine whether genetic variants in MTHFR gene are associated with the risk of ischemic stroke and fasting total serum homocysteine (tHcy) level. We genotyped nine tag SNPs in the MTHFR gene in a case-control study, including 543 ischemic stroke cases and 655 healthy controls in China. We found that subjects with the rs1801133 TT genotype and rs1801131 CC genotype had significant increased risks of ischemic stroke (adjusted odds ratio (OR)=1.82, 95% confidence interval (CI): 1.27-2.61, p=0.004; adjusted OR=1.99, 95% CI: 1.12-3.56, p=0.01) compared with subjects with the major alleles. Haplotype analysis also found that carriers of the MTHFR CTTCGA haplotype (rs12121543-rs13306553-rs9651118-rs1801133-rs2274976-rs1801131) had a significant reduced risk of ischemic stroke (adjusted OR=0.53, 95% CI: 0.35-0.82) compared with those with the CTTTGA haplotype. Besides, the MTHFR rs1801133 and rs9651118 were significantly associated with serum levels of tHcy in healthy controls (p<0.0001 and p=0.02). These findings suggest that variants in the MTHFR gene may influence the risk of ischemic stroke and serum tHcy.

MYSM1 acts as a novel co-activator of ERα to confer antiestrogen resistance in breast cancer.

Endocrine resistance is a crucial challenge in estrogen receptor alpha (ERα)-positive breast cancer (BCa). Aberrant alteration in modulation of E2/ERα signaling pathway has emerged as the putative contributor for endocrine resistance in BCa. Herein, we demonstrate that MYSM1 as a deubiquitinase participates in modulating ERα action via histone and non-histone deubiquitination. MYSM1 is involved in maintenance of ERα stability via ERα deubiquitination. MYSM1 regulates relevant histone modifications on cis regulatory elements of ERα-regulated genes, facilitating chromatin decondensation. MYSM1 is highly expressed in clinical BCa samples. MYSM1 depletion attenuates BCa-derived cell growth in xenograft models and increases the sensitivity of antiestrogen agents in BCa cells. A virtual screen shows that the small molecule Imatinib could potentially interact with catalytic MPN domain of MYSM1 to inhibit BCa cell growth via MYSM1-ERα axis. These findings clarify the molecular mechanism of MYSM1 as an epigenetic modifier in regulation of ERα action and provide a potential therapeutic target for endocrine resistance in BCa.

ADAMTS8 inhibits glioma development in vitro and in vivo.

INTRODUCTION: In recent years, novel RNAs have been revealed to be regulators in glioma. ADAMTS8 has been reported to be reduced in brain tumours. In this study, we aimed to explore the role of ADAMTS8 in glioma. MATERIAL AND METHODS: Online bioinformatic tools, Gepia and Chinese Glioma Genome Atlas database (CGGA) were used to analyse the differential expression of ADAMTS8, overall survival and disease-free survival rates and the correlations between ADAMTS8 and matrix metallopeptidases (MMP2 and MMP9) in glioma. RT-qPCR and western blot experiments were performed to measure the mRNA and protein expression. ADAMTS8 expression was regulated in cells through transfection. Thereafter, the effect of ADAMTS8 on cells was investigated through the cell viability, apoptosis and transwell experiments. The epithelial-mesenchymal transition (EMT)-related proteins and also MMP2 and MMP9 were examined. The subcutaneous tumour model was established to validate the suppressive role of ADAMTS8 in tumour growth. RESULTS: ADAMTS8 expression was reduced in glioma tissues and cells. Higher expression of ADAMTS8 was correlated with higher survival rates. ADAMTS8 was correlated with MMP2 and MMP9 in glioma tissues. In glioma cells, overexpression of ADAMTS8 could inhibit the viability, invasion, migration and EMT, and MMP2 and MMP9, but promote the apoptosis of cells. The upregulation of ADAMTS8 could inhibit the tumour growth in vivo. CONCLUSIONS: ADAMTS8 was inhibited in glioma and the higher expression of ADAMTS8 might be related to better prognosis among glioma patients. Overexpression of ADAMTS8 inhibited the development of glioma in vitro and in vivo.

BAP18 facilitates CTCF-mediated chromatin accessible to regulate enhancer activity in breast cancer.

The estrogen receptor alpha (ERα) signaling pathway is a crucial target for ERα-positive breast cancer therapeutic strategies. Co-regulators and other transcription factors cooperate for effective ERα-related enhancer activation. Recent studies demonstrate that the transcription factor CTCF is essential to participate in ERα/E2-induced enhancer transactivation. However, the mechanism of how CTCF is achieved remains unknown. Here, we provided evidence that BAP18 is required for CTCF recruitment on ERα-enriched enhancers, facilitating CTCF-mediated chromatin accessibility to promote enhancer RNAs transcription. Consistently, GRO-seq demonstrates that the enhancer activity is positively correlated with BAP18 enrichment. Furthermore, BAP18 interacts with SMARCA1/BPTF to accelerate the recruitment of CTCF to ERα-related enhancers. Interestingly, BAP18 is involved in chromatin accessibility within enhancer regions, thereby increasing enhancer transactivation and enhancer-promoter looping. BAP18 depletion increases the sensitivity of anti-estrogen and anti-enhancer treatment in MCF7 cells. Collectively, our study indicates that BAP18 coordinates with CTCF to enlarge the transactivation of ERα-related enhancers, providing a better understanding of BAP18/CTCF coupling chromatin remodeling and E-P looping in the regulation of enhancer transcription.

USP22 upregulates ZEB1-mediated VEGFA transcription in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a common solid tumor with high rate of recurrence and mortality. Anti-angiogenesis drugs have been used for the therapy of HCC. However, anti-angiogenic drug resistance commonly occurs during HCC treatment. Thus, identification of a novel VEGFA regulator would be better understanding for HCC progression and anti-angiogenic therapy resistance. Ubiquitin specific protease 22 (USP22) as a deubiquitinating enzyme, participates in a variety of biological processes in numerous tumors. While the molecular mechanism underlying the effects of USP22 on angiogenesis is still needed to be clarified. Here, our results demonstrated that USP22 acts as a co-activator of VEGFA transcription. Importantly, USP22 is involved in maintenance of ZEB1 stability via its deubiquitinase activity. USP22 was recruited to ZEB1-binding elements on the promoter of VEGFA, thereby altering histone H2Bub levels, to enhance ZEB1-mediated VEGFA transcription. USP22 depletion decreased cell proliferation, migration, Vascular Mimicry (VM) formation, and angiogenesis. Furthermore, we provided the evidence to show that knockdown of USP22 inhibited HCC growth in tumor-bearing nude mice. In addition, the expression of USP22 is positively correlated with that of ZEB1 in clinical HCC samples. Our findings suggest that USP22 participates in the promotion of HCC progression, if not all, at least partially via up-regulation of VEGFA transcription, providing a novel therapeutic target for anti-angiogenic drug resistance in HCC.

Octreotide-mediated neurofunctional recovery in rats following traumatic brain injury. Role of H2S, Nrf2 and TNF-α.

PURPOSE: To explore the role and mechanisms of octreotide in neurofunctional recovery in the traumatic brain injury (TBI) model. METHODS: Rats were subjected to midline incision followed by TBI in the prefrontal cortex region. After 72 hours, the behavioural and neurological deficits tests were performed, which included memory testing on Morris water maze for 5 days. Octreotide (15 and 30 mg/kg i.p.) was administered 30 minutes before subjecting to TBI, and its administration was continued for three days. RESULTS: In TBI-subjected rats, administration of octreotide restored on day 4 escape latency time (ELT) and increased the time spent in the target quadrant (TSTQ) on day 5, suggesting the improvement in learning and memory. It also increased the expression of H2S, Nrf2, and cystathionine-γ-lyase (CSE) in the prefrontal cortex, without any significant effect on cystathionine-ß-synthase. Octreotide also decreased the TNF-α levels and neurological severity score. However, co-administration of CSE inhibitor (D,L-propargylglycine) abolished octreotide-mediated neurofunctional recovery, decreased the levels of H2S and Nrf2 and increased the levels of TNF-α. CONCLUSIONS: Octreotide improved the neurological functions in TBI-subjected rats, which may be due to up-regulation of H2S biosynthetic enzyme (CSE), levels of H2S and Nrf2 and down-regulation of neuroinflammation.

Licochalcone B attenuates neuronal injury through anti-oxidant effect and enhancement of Nrf2 pathway in MCAO rat model of stroke.

BACKGROUND: Investigating anti-oxidant therapies that lead to the diminution of oxidative injury is priority in clinical. We herein aimed to explore whether and how Licochalcone B (Lico B) act as an anti-oxidant in the stroke model. METHODS: Middle cerebral artery occlusion (MCAO) was constructed as stroke model and exposed to various doses of Lico B. Behavioral tests and neurological behavior status were detected for neurological function examination. Histological staining was used for evaluating cerebral injury, and neuronal apoptosis or damage. Levels of oxidative stress and inflammation were also assessed by biochemical analysis and expression analysis. Nrf2 knockdown induced by lentiviral vector was used for the research on mechanism. RESULTS: Lico B had improvement effects on cerebral infarction size, memory impairments, and neurological deficits after MCAO. Histological evaluation also revealed the amelioration of neuronal injury and apoptosis by Lico B, along with down-regulation of apoptosis-related proteins. Additionally, Lico B rescued the down-regulation of BDNF and NGF after MCAO. Moreover, Lico B suppressed the oxidative stress and inflammation, manifesting as the enhancement of SOD, GSH and IL-4, but the decline of MDA, iNOS, and TNF-α. Finally, Nrf2 knockdown reversed the Lico B-caused improvement in neuronal injury, apoptosis and oxidative stress levels. CONCLUSIONS: The present study revealed the neuroprotective effects of Lico B in MCAO rats. Importantly, we proposed a potential mechanism that Lico B activated the Nrf2 pathway, thereby acting as anti-oxidant to attenuate neuronal injury and apoptosis after stroke. The proposed mechanism provided an encouraging possibility for anti-oxidant therapy of stroke.

Corrosion Resistance of 2060 Aluminum⁻Lithium Alloy LBW Welds Filled with Al-5.6Cu Wire.

Alloy sheets of type 2060 aluminum⁻lithium were welded by laser beam welding (LBW) filled with ER2319 Al-5.6Cu wire. Microstructural observations showed the uneven distribution of columnar grains, equiaxed grains and equiaxed dendrite grains in the weld. The θ'(Al2Cu) phase and other phases precipitated in the weld. The θ'(Al2Cu) phase centrally distributed at the grain boundaries. During the immersion corrosion, the pitting corrosion first occurred and then gradually expanded and transformed to intergranular corrosion and exfoliation corrosion. The electrochemical corrosion test showed a higher corrosion tendency of the base metal and heat-affected zone for the lower corrosion potential, but the corrosion current density of the weld was relatively larger. The segregation of Cu, Mg and other elements at the grain boundary aggravated the occurrence of intergranular corrosion.

Octreotide-mediated neurofunctional recovery in rats following traumatic brain injury. Role of H2S, Nrf2 and TNF-α

ABSTRACT Purpose: To explore the role and mechanisms of octreotide in neurofunctional recovery in the traumatic brain injury (TBI) model. Methods: Rats were subjected to midline incision followed by TBI in the prefrontal cortex region. After 72 hours, the behavioural and neurological deficits tests were performed, which included memory testing on Morris water maze for 5 days. Octreotide (15 and 30 mg/kg i.p.) was administered 30 minutes before subjecting to TBI, and its administration was continued for three days. Results: In TBI-subjected rats, administration of octreotide restored on day 4 escape latency time (ELT) and increased the time spent in the target quadrant (TSTQ) on day 5, suggesting the improvement in learning and memory. It also increased the expression of H2S, Nrf2, and cystathionine-γ-lyase (CSE) in the prefrontal cortex, without any significant effect on cystathionine-β-synthase. Octreotide also decreased the TNF-α levels and neurological severity score. However, co-administration of CSE inhibitor (D,L-propargylglycine) abolished octreotide-mediated neurofunctional recovery, decreased the levels of H2S and Nrf2 and increased the levels of TNF-α. Conclusions: Octreotide improved the neurological functions in TBI-subjected rats, which may be due to up-regulation of H2S biosynthetic enzyme (CSE), levels of H2S and Nrf2 and down-regulation of neuroinflammation.

Knockdown of CDC2 expression inhibits proliferation, enhances apoptosis, and increases chemosensitivity to temozolomide in glioblastoma cells.

Cell division cycle 2 (CDC2) is always overexpressed in malignant tumor cells and is correlated with chemosensitivity, but it is unclear whether CDC2 overexpression contributes to the chemoresistance potential of glioma cells. The aim of study was to determine the relationship of CDC2 expression with the prognosis and chemoresistance of glioblastoma. In this study, the glioblastoma U87 and U251 cell lines were steadily transfected with a lentivirus vector expressing a short hairpin RNA-targeting CDC2. Expression of CDC2 was evaluated in glioblastoma and cell lines by immunohistochemistry and Western blot analysis. The relationship between CDC2 expression and clinicopathological characteristics was analyzed. Using RNA interference, the effects of CDC2 on chemosensitivity to temozolomide (TMZ) were investigated in U87 and U251 cell lines in vitro. Combined CDC2 knockdown and TMZ treatment inhibited cell proliferation and induced apoptosis in vitro more effectively than either treatment alone. qRT-PCR and Western blot analysis showed that cells underexpressing CDC2 revealed lower expression of the anti-apoptotic protein B cell lymphoma-2 and increased expression of the apoptosis effector caspase-3 compared to U87 and U251 cells transfected with a control vector. Furthermore, expression levels of CDC2 in U87 and U251 cells were related to the IC50 of the antitumor drug TMZ. Knockdown of CDC2 expression was associated with decreased expression of Ral-binding protein 1, a classical chemotherapy drugs transporter. These results indicate that the ability to suppress the malignant phenotype by down-regulating CDC2 expression may provide a new gene therapy approach for overcoming CDC2-associated chemoresistance in patients with malignant glioma.