Gankyrin inhibits ferroptosis through the p53/SLC7A11/GPX4 axis in triple-negative breast cancer cells.

Gankyrin is found in high levels in triple-negative breast cancer (TNBC) and has been established to form a complex with the E3 ubiquitin ligase MDM2 and p53, resulting in the degradation of p53 in hepatocarcinoma cells. Therefore, this study sought to determine whether gankyrin could inhibit ferroptosis through this mechanism in TNBC cells. The expression of gankyrin was investigated in relation to the prognosis of TNBC using bioinformatics. Co-immunoprecipitation and GST pull-down assays were then conducted to determine the presence of a gankyrin and MDM2 complex. RT-qPCR and immunoblotting were used to examine molecules related to ferroptosis, such as gankyrin, p53, MDM2, SLC7A11, and GPX4. Additionally, cell death was evaluated using flow cytometry detection of 7-AAD and a lactate dehydrogenase release assay, as well as lipid peroxide C11-BODIPY. Results showed that the expression of gankyrin is significantly higher in TNBC tissues and cell lines, and is associated with a poor prognosis for patients. Subsequent studies revealed that inhibiting gankyrin activity triggered ferroptosis in TNBC cells. Additionally, silencing gankyrin caused an increase in the expression of the p53 protein, without altering its mRNA expression. Co-immunoprecipitation and GST pull-down experiments indicated that gankyrin and MDM2 form a complex. In mouse embryonic fibroblasts lacking both MDM2 and p53, this gankyrin/MDM2 complex was observed to ubiquitinate p53, thus raising the expression of molecules inhibited by ferroptosis, such as SLC7A11 and GPX4. Furthermore, silencing gankyrin in TNBC cells disrupted the formation of the gankyrin/MDM2 complex, hindered the degradation of p53, increased SLC7A11 expression, impeded cysteine uptake, and decreased GPX4 production. Our findings suggest that TNBC cells are able to prevent cell ferroptosis through the gankyrin/p53/SLC7A11/GPX4 signaling pathway, indicating that gankyrin may be a useful biomarker for predicting TNBC prognosis or a potential therapeutic target.

Three-Dimensional de Sitter Holography and Bulk Correlators at Late Time.

We propose an explicitly calculable example of holography on three-dimensional de Sitter space by providing a prescription to analytic continue a higher-spin holography on three-dimensional anti-de Sitter space. Applying the de Sitter holography, we explicitly compute bulk correlation functions on three-dimensional de Sitter space at late time in a higher-spin gravity. These expressions are consistent with recent analysis based on bulk Feynman diagrams. Our explicit computations reveal how holographic computations could provide fruitful information.

A Metabolic-associated Nomogram Predicts Recurrence Survival of Thyroid Cancer.

OBJECTIVE: Various studies have suggested that metabolic genes play a significant role in papillary thyroid cancer (PTC). The current study aimed to identify a metabolic signature related biomarker to predict the prognosis of patients with PTC. METHODS: We conducted a comprehensive analysis on the data obtained from the Cancer Genome Atlas (TCGA) database. The correlation between survival result and metabolic genes was evaluated based on the univariate Cox analyses, least absolute shrinkage and selection operator (LASSO) and multivariate Cox analyses. The performance of a 7-gene signature was assessed according to Kaplan-Meier and receiver operating characteristic (ROC) analysis. Multivariate Cox regression analysis was adopted to unearth clinical factors related to the recurrence free survival (RFS) of patients with PTC. Finally, a prognostic nomogram was developed based on risk score, cancer status and cancer width to improve the prediction for RFS of PTC patients. RESULTS: Seven metabolic genes were used to establish the prognostic model. The ROC curve and C-index exhibited high value in training, testing and the whole TCGA datasets. The established nomogram, incorporating the 7-metabolic gene signature and clinical factors, was able to predict the RFS with high effectiveness. The 7-metabolic gene signature-based nomogram had a good performance to predict the RFS of patients with PTC. CONCLUSION: Our study identified a 7-metabolic gene signature and established a prognostic nomogram, which were useful in predicting the RFS of PTC.

Liver ChREBP Protects Against Fructose-Induced Glycogenic Hepatotoxicity by Regulating L-Type Pyruvate Kinase.

Excessive fructose consumption is closely linked to the pathogenesis of metabolic disease. Carbohydrate response element-binding protein (ChREBP) is a transcription factor essential for fructose tolerance in mice. However, the functional significance of liver ChREBP in fructose metabolism remains unclear. Here, we show that liver ChREBP protects mice against fructose-induced hepatotoxicity by regulating liver glycogen metabolism and ATP homeostasis. Liver-specific ablation of ChREBP did not compromise fructose tolerance, but rather caused severe transaminitis and hepatomegaly with massive glycogen overload in mice fed a high-fructose diet, while no obvious inflammation, cell death, or fibrosis was detected in the liver. In addition, liver ATP contents were significantly decreased by ChREBP deficiency in the fed state, which was rendered more pronounced by fructose feeding. Mechanistically, liver contents of glucose-6-phosphate (G6P), an allosteric activator of glycogen synthase, were markedly increased in the absence of liver ChREBP, while fasting-induced glycogen breakdown was not compromised. Furthermore, hepatic overexpression of LPK, a ChREBP target gene in glycolysis, could effectively rescue glycogen overload and ATP reduction, as well as mitigate fructose-induced hepatotoxicity in ChREBP-deficient mice. Taken together, our findings establish a critical role of liver ChREBP in coping with hepatic fructose stress and protecting from hepatotoxicity by regulating LPK.

Risk Factors and Management of Postoperative Pancreatic Fistula Following Pancreaticoduodenectomy: Single-center Experience.

Pancreatic fistula (PF) remains the most frequent complication after pancreaticoduodenectomy (PD). This study was undertaken to explore the risk factors of postoperative PF following PD and discuss the management of PF in our center. A single-center respective study, involving 241 patients who underwent PD between September 2015 and June 2018, was conducted. Differences in the demographic data, preoperative, intraoperative and postoperative variables between the group with PF [International Study Group on Pancreatic Surgery (ISGPS) grade B/C] and the group without PF (no PF and ISGPS grade BL) were evaluated. The diagnosis and grading of PF were in strict accordance with ISGPS. Risk factors were analyzed by univariate analysis and multivariate logistic regression analysis. The results showed that postoperative PF occurred in 50 (20.7%) of the patients; 25 (10.4%) patients had a PF type BL, 46 (19.1%) patients developed a PF type B and 4 (1.6%) had a PF type C. Univariate analysis showed that fasting blood glucose (P=0.02), pancreatic texture (P< 0.001) and pancreatic duct diameter (P=0.01) were correlated with PF. Multivariate logistic regression analysis identified one independent risk factor for postoperative PF: soft pancreatic texture (OR=3.251, P=0.002). Among the cases, there were three postoperative deaths, giving a 60-day hospital mortality rate of 1.2% (3/241), and the mortality related to PF was 4.0% (2/50). One of the patients died from multiple organ failure caused by postoperative abdominal hemorrhage. In conclusion, soft pancreatic texture is an independent risk factor for PF. Surgeons should be well aware of this risk factor when performing a PD.

Nutrient Stress-Dysregulated Antisense lncRNA GLS-AS Impairs GLS-Mediated Metabolism and Represses Pancreatic Cancer Progression.

Cancer cells are known to undergo metabolic reprogramming, such as glycolysis and glutamine addiction, to sustain rapid proliferation and metastasis. It remains undefined whether long noncoding RNAs (lncRNA) coordinate the metabolic switch in pancreatic cancer. Here we identify a nuclear-enriched antisense lncRNA of glutaminase (GLS-AS) as a critical regulator involved in pancreatic cancer metabolism. GLS-AS was downregulated in pancreatic cancer tissues compared with noncancerous peritumor tissues. Depletion of GLS-AS promoted proliferation and invasion of pancreatic cancer cells both in vitro and in xenograft tumors of nude mice. GLS-AS inhibited GLS expression at the posttranscriptional level via formation of double stranded RNA with GLS pre-mRNA through ADAR/Dicer-dependent RNA interference. GLS-AS expression was transcriptionally downregulated by nutrient stress-induced Myc. Conversely, GLS-AS decreased Myc expression by impairing the GLS-mediated stability of Myc protein. These results imply a reciprocal feedback loop wherein Myc and GLS-AS regulate GLS overexpression during nutrient stress. Ectopic overexpression of GLS-AS inhibited proliferation and invasion of pancreatic cancer cells by repressing the Myc/GLS pathway. Moreover, expression of GLS-AS and GLS was inversely correlated in clinical samples of pancreatic cancer, while low expression of GLS-AS was associated with poor clinical outcomes. Collectively, our study implicates a novel lncRNA-mediated Myc/GLS pathway, which may serve as a metabolic target for pancreatic cancer therapy, and advances our understanding of the coupling role of lncRNA in nutrition stress and tumorigenesis.Significance: These findings show that lncRNA GLS-AS mediates a feedback loop of Myc and GLS, providing a potential therapeutic target for metabolic reprogramming in pancreatic cancer.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/7/1398/F1.large.jpg.See related commentary by Mafra and Dias, p. 1302.

Hypoxia-induced LncRNA-BX111 promotes metastasis and progression of pancreatic cancer through regulating ZEB1 transcription.

The contribution of long noncoding RNAs (lncRNAs) to pancreatic cancer progression and the regulatory mechanisms of their expression are attractive areas. In the present study, the overexpression of lncRNA-BX111887 (BX111) in pancreatic cancer tissues was detected by microarray and further validated in a cohort of pancreatic cancer tissues. We further demonstrated that knockdown or overexpression of BX111 dramatically repressed or enhanced proliferation and invasion of pancreatic cancer cells. Mechanically, BX111 activated transcription of ZEB1, a key regulator for epithelia-mesenchymal transition (EMT), via recruiting transcriptional factor Y-box protein (YB1) to its promoter region. Moreover, we revealed that BX111 transcription was induced by hypoxia-inducible factor (HIF-1α) in response to hypoxia. In addition, BX111 contributed to the hypoxia-induced EMT of pancreatic cells by regulating expression of ZEB1 and its downstream proteins E-cadherin and MMP2. Coincidence with in vitro results, BX111 depletion effectively inhibited growth and metastasis of xenograft tumor in vivo. The clinical samples of pancreatic cancer further confirmed a positive association between BX111 and ZEB1. Moreover, high BX111 expression was correlated with late TNM stage, lymphatic invasion and distant metastasis, as well as short overall survival time in patients. Taken together, our findings implicate a hypoxia-induced lncRNA contributes to metastasis and progression of pancreatic cancer, and suggest BX111 might be applied as a potential biomarker and therapeutic target for pancreatic cancer.

MiRNA-646-mediated reciprocal repression between HIF-1α and MIIP contributes to tumorigenesis of pancreatic cancer.

Migration and invasion inhibitory protein (MIIP) is recently identified as an inhibitor in tumor development. However, the regulatory mechanism and biological contributions of MIIP in pancreatic cancer (PC) have been not elucidated. In this study, we demonstrated a negative feedback of MIIP and hypoxia-induced factor-1α (HIF-1α), which was mediated by a hypoxia-induced microRNA. Compared with paracarcinoma tissues, MIIP was downregulated in PC tissues. Overexpression of MIIP significantly impeded the proliferation and invasion of PC cells both in vitro and in mouse xenograft models. We further verified MIIP was downregulated under hypoxia in a HIF-1α-mediated manner. Interestingly, although MIIP promoter containing two putative hypoxia response elements (HREs), the chromatin immunoprecipitation (ChIP) and luciferase reporter assays did not support an active interaction between HIF-1α and MIIP promoter. Meanwhile, microRNA array revealed a hypoxia-induced microRNA, miR-646, impaired stability of MIIP mRNA and consequently inhibited its expression by targeting the coding sequence (CDS). Coincidently, knockdown of miR-646 significantly repressed proliferation and invasion ability of PC cells both in vitro and in vivo by upregulating MIIP expression. Besides, ChIP and luciferase reporter assays further validated that HIF-1α activated transcription of miR-646 in hypoxia condition. Therefore, these results suggested HIF-1α indirectly regulated MIIP expression in post-transcriptional level through upregulating miR-646 transcription. Conversely, our results further revealed that MIIP suppressed deacetylase ability of histone deacetylase 6 (HDAC6) to promote the acetylation and degradation of HIF-1α, by which impairing HIF-1α accumulation. What is more, a specific relationship between downregulated MIIP and upregulated miR-646 expression was validated in PC samples. Moreover, the dysregulated miR-646 and MIIP expression was correlated with advanced tumor stage, lymphatic invasion, metastasis and shorter overall survival in PC patients. Together, our results highlight that the reciprocal loop of HIF-1α/miR-646/MIIP might be implemented as an applicable target for pancreatic cancer therapy.

Hypoxia-induced lncRNA-NUTF2P3-001 contributes to tumorigenesis of pancreatic cancer by derepressing the miR-3923/KRAS pathway.

Recent studies indicate that long non-coding RNAs (lncRNAs) play crucial roles in numerous cancers, while their function in pancreatic cancer is rarely elucidated. The present study identifies a functional lncRNA and its potential role in tumorigenesis of pancreatic cancer. Microarray co-assay for lncRNAs and mRNAs demonstrates that lncRNA-NUTF2P3-001 is remarkably overexpressed in pancreatic cancer and chronic pancreatitis tissues, which positively correlates with KRAS mRNA expression. After downregulating lncRNA-NUTF2P3-001, the proliferation and invasion of pancreatic cancer cell are significantly inhibited both in vitro and vivo, accompanying with decreased KRAS expression. The dual-luciferase reporter assay further validates that lncRNA-NUTF2P3-001 and 3'UTR of KRAS mRNA competitively bind with miR-3923. Furthermore, miR-3923 overexpression simulates the inhibiting effects of lncRNA-NUTF2P3-001-siRNA on pancreatic cancer cell, which is rescued by miR-3923 inhibitor. Specifically, the present study further reveals that lncRNA-NUTF2P3-001 is upregulated in pancreatic cancer cells under hypoxia and CoCl2 treatment, which is attributed to the binding of hypoxia-inducible factor-1α (HIF-1α) to hypoxia response elements (HREs) in the upstream of KRAS promoter. Data from pancreatic cancer patients show a positive correlation between lncRNA-NUTF2P3-001 and KRAS, which is associated with advanced tumor stage and worse prognosis. Hence, our data provide a new lncRNA-mediated regulatory mechanism for the tumor oncogene KRAS and implicate that lncRNA-NUTF2P3-001 and miR-3923 can be applied as novel predictors and therapeutic targets for pancreatic cancer.

[Zebrafish as a model animal for the study of blood-brain barrier permeability by biomolecules].

Blood-brain barrier (BBB) is the major obstacle for drug delivery into the central nervous system (CNS). However, there is no ideal model animal for the study of BBB permeability till now. Currently zebrafish (Danio rerio) has emerged as a powerful model organism for the study of vertebrate biology. In this study, the feasibility of using zebrafish as model animal was investigated for BBB permeability by comparing the results of administration of BBB-penetrating peptide and protein to mouse and zebrafish. The results showed that the BBBs of mouse and zebrafish were similar in molecular permeability. Additionally, zebrafish has advantageous features as a model animal, such as small size, fertile and easy to breed. Therefore, it is suggested that zebrafish may be a favored model for the study of BBB permeability.