Radiomics model based on contrast-enhanced computed tomography to predict early recurrence in patients with hepatocellular carcinoma after radical resection.

BACKGROUND: Radical resection remains an effective strategy for patients with hepatocellular carcinoma (HCC). Unfortunately, the postoperative early recurrence (recurrence within 2 years) rate is still high. AIM: To develop a radiomics model based on preoperative contrast-enhanced computed tomography (CECT) to evaluate early recurrence in HCC patients with a single tumour. METHODS: We enrolled a total of 402 HCC patients from two centres who were diagnosed with a single tumour and underwent radical resection. First, the features from the portal venous and arterial phases of CECT were extracted based on the region of interest, and the early recurrence-related radiomics features were selected via the least absolute shrinkage and selection operator proportional hazards model (LASSO Cox) to determine radiomics scores for each patient. Then, the clinicopathologic data were combined to develop a model to predict early recurrence by Cox regression. Finally, we evaluated the prediction performance of this model by multiple methods. RESULTS: A total of 1915 radiomics features were extracted from CECT images, and 31 of them were used to determine the radiomics scores, which showed a significant difference between the early recurrence and nonearly recurrence groups. Univariate and multivariate Cox regression analyses showed that radiomics scores and serum alpha-fetoprotein were independent indicators, and they were used to develop a combined model to predict early recurrence. The area under the receiver operating characteristic curve values for the training and validation cohorts were 0.77 and 0.74, respectively, while the C-indices were 0.712 and 0.674, respectively. The calibration curves and decision curve analysis showed satisfactory accuracy and clinical utilities. Kaplan-Meier curves based on recurrence-free survival and overall survival showed significant differences. CONCLUSION: The preoperative radiomics model was shown to be effective for predicting early recurrence among HCC patients with a single tumour.

Novel index for the prediction of significant liver fibrosis and cirrhosis in chronic hepatitis B patients in China.

BACKGROUND: Noninvasive, practical, and convenient means of detection for the prediction of liver fibrosis and cirrhosis in China are greatly needed. AIM: To develop a precise noninvasive test to stage liver fibrosis and cirrhosis. METHODS: With liver biopsy as the gold standard, we established a new index, [alkaline phosphatase (U/L) + gamma-glutamyl transpeptidase (U/L)/platelet (109/L) (AGPR)], to predict liver fibrosis and cirrhosis. In addition, we compared the area under the receiver operating characteristic curve (AUROC) of AGPR, gamma-glutamyl transpeptidase to platelet ratio, aspartate transaminase to platelet ratio index, and FIB-4 and evaluated the accuracy of these routine laboratory indices in predicting liver fibrosis and cirrhosis. RESULTS: Correlation analysis revealed a significant positive correlation between AGPR and liver fibrosis stage (P < 0.001). In the training cohort, the AUROC of AGPR was 0.83 (95%CI: 0.78-0.87) for predicting fibrosis (≥ F2), 0.84 (95%CI: 0.79-0.88) for predicting extensive fibrosis (≥ F3), and 0.87 (95%CI: 0.83-0.91) for predicting cirrhosis (F4). In the validation cohort, the AUROCs of AGPR to predict ≥ F2, ≥ F3 and F4 were 0.83 (95%CI: 0.77-0.88), 0.83 (95%CI: 0.77-0.89), and 0.84 (95%CI: 0.78-0.89), respectively. CONCLUSION: The AGPR index should become a new, simple, accurate, and noninvasive marker to predict liver fibrosis and cirrhosis in chronic hepatitis B patients.

The BcLAE1 is involved in the regulation of ABA biosynthesis in Botrytis cinerea TB-31.

Abscisic acid (ABA), as a classic plant hormone, is a key factor in balancing the metabolism of endogenous plant hormones, and plays an important role in regulating the activation of mammalian innate immune cells and glucose homeostasis. Currently, Botrytis cinerea has been used for fermentation to produce ABA. However, the mechanism of the regulation of ABA biosynthesis in B. cinerea is still not fully understood. The putative methyltransferase LaeA/LAE1 is a global regulator involved in the biosynthesis of a variety of secondary metabolites in filamentous fungi. In this study, we demonstrated that BcLAE1 plays an important role in the regulation of ABA biosynthesis in B. cinerea TB-31 by knockout experiment. The deletion of Bclae1 caused a 95% reduction in ABA yields, accompanied by a decrease of the transcriptional level of the ABA synthesis gene cluster Bcaba1-4. Further RNA-seq analysis indicated that deletion of Bclae1 also affected the expression level of key enzymes of BOA and BOT in secondary metabolism, and accompanied by clustering regulatory features. Meanwhile, we found that BcLAE1 is involved in epigenetic regulation as a methyltransferase, with enhanced H3K9me3 modification and attenuated H3K4me2 modification in ΔBclae1 mutant, and this may be a strategy for BcLAE1 to regulate ABA synthesis.

Neoantigen vaccine: An emerging immunotherapy for hepatocellular carcinoma.

Tumor-specific neoantigens, which are expressed on tumor cells, can induce an effective antitumor cytotoxic T-cell response and mediate tumor regression. Among tumor immunotherapies, neoantigen vaccines are in early human clinical trials and have demonstrated substantial efficiency. Compared with more neoantigens in melanoma, the paucity and inefficient identification of effective neoantigens in hepatocellular carcinoma (HCC) remain enormous challenges in effectively treating this malignancy. In this review, we highlight the current development of HCC neoantigens in its generation, screening, and identification. We also discuss the possibility that there are more effective neoantigens in hepatitis B virus (HBV)-related HCC than in non-HBV-related HCC. In addition, since HCC is an immunosuppressive tumor, strategies that reverse immunosuppression and enhance the immune response should be considered for the practical exploitation of HCC neoantigens. In summary, this review offers some strategies to solve existing problems in HCC neoantigen research and provide further insights for immunotherapy.

RFX5 promotes the progression of hepatocellular carcinoma through transcriptional activation of KDM4A.

Regulatory factor X-5 (RFX5) represents a key transcription regulator of MHCII gene expression in the immune system. This study aims to explore the molecular mechanisms and biological significance of RFX5. Firstly, by analyzing ENCODE chromatin immunoprecipitation (ChIP)-seq in HepG2 and TCGA RNA-seq data, we discovered lysine-specific demethylase 4A (KDM4A), also named JMJD2A, to be a major downstream target gene of RFX5. Moreover, RFX5 was verified to bind directly to the KDM4A's promoter region and sequentially promoted its transcription determined by the ChIP-PCR assay and luciferase assay. In addition, RFX5-dependent regulation of KDM4A was demonstrated in HCC. Compared with adjacent non-tumor tissues, the expression levels of KDM4A were significantly raised in HCC tumor tissues. Notably, elevated levels of KDM4A were strongly correlated with HCC patient prognosis. Functionally, KDM4A overexpression largely rescued the growth inhibitory effects of RFX5 deletion, highlighting KDM4A as a downstream effector of RFX5. Mechanistically, the RFX5-KDM4A pathway promoted the progression of the cell cycle from G0/G1 to S phase and was protective against cell apoptosis through regulation of p53 and its downstream genes in HCC. In conclusion, RFX5 could promote HCC progression via transcriptionally activating KDM4A expression.

Genome-scale CRISPR activation screening identifies a role of ELAVL2-CDKN1A axis in paclitaxel resistance in esophageal squamous cell carcinoma.

Neoadjuvant chemotherapy (NAC) may provide survival benefits for patients with advanced esophageal squamous cell carcinoma. However, tumor cells can display primary or secondary resistance to paclitaxel (PTX), a primary component of induction chemotherapy regimen. To identify genes capable of conveying PTX resistance, we performed a genome-wide CRISPR transcriptional activation library in human KYSE-180 cells. High throughput next generation sequencing was further applied to establish the phenotype-to-genotype relationship. Our highest-ranking hits are CDKN1A, TSPAN4, ELAVL2, JUNB and PAAF1. We generated evidence that esophageal tumors with high CDKN1A, ELAVL2 and TSPAN4 levels, quantified using qRT-PCR and Western blot assays, showed poorer chemotherapy response. Higher expression levels of TSPAN4 and ELAVL2 protein are independent risk factors for poor chemotherapy response in ESCC patients. We then found that overexpression of CDKN1A, ELAVL2 or TSPAN4 in ESCC cell lines significantly promoted the resistance to PTX by inhibiting cell apoptosis. Interestingly, ESCC cells overexpressed CDKN1A, ELAVL2 or TSPAN4 also acquired resistance to cisplatin (DDP). This phenomenon may be explained by cross-resistance of chemotherapy. We additionally found an association between ELAVL2 and CDKN1A, which may be regarded as the upstream and downstream factors that synergistically involved in the regulation of chemo-resistance in ESCC. Therefore, our study demonstrated that the genome-wide CRISPR activation library is a powerful strategy for the discovery of chemo-resistant genes critical for ESCC and we reported the first evidence that the ELAVL2-CDKN1A axis may be an important mechanism involved in chemo-resistance in ESCC.

Regulatory factor X5 promotes hepatocellular carcinoma progression by transactivating tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein theta and suppressing apoptosis.

BACKGROUND: Our previous studies have shown that regulatory factor X5 (RFX5), a classical transcription regulator of MHCII genes, was obviously overexpressed in hepatocellular carcinoma (HCC) tumors. However, the role of RFX5 in the carcinogenesis and progress of HCC remains unknown. This study aimed to reveal its biological significance and the underlying mechanism in HCC. METHODS: RFX5 mRNA expression level and copy number variation in HCC tumors and cell lines were determined by analyzing deposited data sets in the Cancer Genome Atlas and Gene Expression Omnibus database. The biological significance of RFX5 in HCC was investigated by monitoring the colony formation and subcutaneous tumor growth capacity when RFX5 was silenced with lentiviral short hairpin RNA and CRISPR/Cas9 system in HCC cell lines. The downstream gene transcriptionally activated by RFX5 in HCC cells was determined by chromatin immunoprecipitation and luciferase reporter assay. The involvement of tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein theta (YWHAQ) in HCC development was further determined by performing colony formation rescue assay and subcutaneous tumor growth rescue experiment. The association of YWHAQ with recurrence-free survival of patients with HCC was assessed by Kaplan-Meier analysis. Moreover, apoptosis level and the protein level of p53 pathway were determined to reveal the mechanism of RFX5 in driving HCC development. RESULTS: RFX5 was amplified and highly overexpressed in HCC tumor tissues compared with the corresponding non-tumor tissues. The mRNA expression level of RFX5 was significantly correlated with its DNA copy number (r = 0.4, P < 0.001). Functional study demonstrated that RFX5 was required for both clonogenic forming in vitro and subcutaneous tumor growth in vivo of HCC cells. Further study identified YWHAQ, namely 14-3-3 tau, as a key downstream transcriptional target gene of RFX5, which was tightly regulated by RFX5 in HCC. Moreover, overexpression of YWHAQ largely rescued the clonogenic growth of HCC cells that was suppressed by RFX5 knockdown. In addition, overexpression of YWHAQ in primary tumor was linked to poor prognosis of patients with HCC. These results demonstrated that YWHAQ was a downstream effector of RFX5 in HCC. Notably, RFX5-YWHAQ pathway could protect cells from apoptosis by suppressing the p53 and Bax in HCC. CONCLUSION: RFX5 is a putative HCC driver gene that plays an important role in the development and progression of HCC by transactivating YWHAQ and suppressing apoptosis.

Interaction between Oc-1 and Lmx1a promotes ventral midbrain dopamine neural stem cells differentiation into dopamine neurons.

Recent studies have shown that Onecut (Oc) transcription factors may be involved in the early development of midbrain dopaminergic neurons (mdDA). The expression profile of Oc factors matches that of Lmx1a, an important intrinsic transcription factor in the development of mDA neuron. Moreover, the Wnt1-Lmx1a pathway controls the mdDA differentiation. However, their expression dynamics and molecular mechanisms remain to be determined. To address these issues, we hypothesize that cross-talk between Oc-1 and Lmx1a regulates the mdDA specification and differentiation through the canonical Wnt-ß-catenin pathway. We found that Oc-1 and Lmx1a displayed a very similar expression profile from embryonic to adult ventral midbrain (VM) tissues. Oc-1 regulated the proliferation and differentiation of ventral midbrain neural stem cells (vmNSCs). Downregulation of Oc-1 decreased both transcript and protein level of Lmx1a. Oc-1 interacted with lmx1a in vmNSCs in vitro and in VM tissues in vivo. Knockdown of Lmx1a reduced the expression of Oc-1 and Wnt1 in vmNSCs. Inhibiting Wnt1 signaling in vmNSCs provoked similar responses. Our data suggested that Oc-1 interacts with Lmx1a to promote vmNSCs differentiation into dopamine neuron through Wnt1-Lmx1a pathway.

Expression of the pluripotency markers Oct3/4, Nanog and Sox2 in human breast cancer cell lines.

Previous studies have demonstrated that pluripotency-associated transcription factors, such as Oct3/4, Nanog and Sox2, play a crucial role in the development and malignant progression of various types of tumors. Breast cancer is the most frequent cancer among females, being a heterogeneous disease, with distinct morphologies, metastatic behavior and therapeutic responses. The expression of Oct3/4, Nanog and Sox2 in 3 human breast cancer cell lines, MCF7, T-47D and MDA-MB-231, was determined. The expression of Oct3/4, Nanog and Sox2 mRNA was determined by reverse transcription polymerase chain reaction (RT-PCR) and protein expression was detected by immunocytohistochemistry. RT-PCR revealed that all three human breast cancer cell lines tested expressed evident Oct3/4, Nanog and Sox-2 mRNA at various levels. Higher levels of Oct3/4 were identified in MCF7 and MDA-MB-231 cells compared with T-47D cells. Higher levels of Nanog were observed in MCF7 and T-47D cells compared with MDA-MB-231 cells and the highest expression of Sox-2 was detected in MCF7 cells. The nuclear localization of Oct3/4, Nanog and Sox-2 was confirmed by immunostaining. Oct3/4, Nanog and Sox2 were expressed in human breast cancer cell lines. Further studies are required to characterize the role of Oct3/4, Nanog and Sox2 in human breast cancer.

[Analyzing hematopoietic development of mouse embryonic AGM region by tissue culture].

To analyze hematopoietic kinetics of mouse embryonic aorta-gonad-mesonephros (AGM) region, an in vitro tissue culture method was developed in this study, partly based on previous reports. After 2 days of tissue culture, a significant number of erythro myeloid progenitors, as quantitated by colony forming assay was detected in the AGM region. Moreover, the cells from cultured E10.5 AGM region could generate 10.8 ± 3.5 colony-forming unit in spleen (CFU-S) per tissue on average. Transplantation of cultured E10.5-E11.0 AGM cells resulted in efficient (85.7% repopulated) and long-term (> 4 months) reconstitution of lethally irradiated adult recipients with remarkable chimerism [(51.12 ± 21.17)%]. The multilineage contribution of donor cells was validated by significant engraftment of myeloid and/or lymphoid cells in peripheral blood, bone marrow, spleen and thymus of recipients. Taken together, the tissue culture method can enable us to manipulate the AGM region in vitro, fulfilling a systematic evaluation of developmental kinetics of various hematopoietic precursor cells, particularly HSC, in normal and mutant mid-gestation mouse embryos.

Migration of dorsal aorta mesenchymal stem cells induced by mouse embryonic circulation.

Mesenchymal stem cells (MSCs) represent powerful tools for regenerative medicine for their differentiation and migration capacity. However, ontogeny and migration of MSCs in mammalian mid-gestation conceptus is poorly understood. We identified canonical MSCs in the mouse embryonic day (E) 11.5 dorsal aorta (DA). They possessed homogenous immunophenotype (CD45(-)CD31(-)Flk-1(-)CD44(+)CD29(+)), expressed perivascular markers (α-SMA(+)NG2(+)PDGFRß(+)PDGFRα(+)), and had tri-lineage differentiation potential (osteoblasts, adipocytes, and chondrocytes). Of interest, MSCs were also detected in E12.5-E13.5 embryonic circulation, 24 hr later than in DA, suggesting migration like hematopoietic stem cells. Functionally, E12.5 embryonic blood could trigger efficient migration of DA-MSCs through platelet-derived growth factor (PDGF) receptor-, transforming growth factor-beta receptor-, but not basic fibroblast growth factor receptor-mediated signaling. Moreover, downstream JNK and AKT signaling pathway played important roles in embryonic blood- or PDGF-mediated migration of DA-derived MSCs. Taken together, these results revealed that clonal MSCs developed in the mouse DA. More importantly, the embryonic circulation, in addition to its conventional transporting roles, could modulate migration of MSC during early embryogenesis.

[Clarification of lymphoid potential in mouse E8.5 embryos by OP9/OP9-DL1 coculture].

The anatomical location of lymphocyte ontogeny in the developing mouse embryo remains controversial. To define the site that can generate lymphocytes de novo, the intraembryonic splanchnopleura (SP) and extraembryonic yolk sac (YS) at 8.5 days postcoitum, when systemic circulation is not established, were investigated. The results indicated that in standard colony forming assay, the cells from both splanchnopleura and yolk sac formed typical myeloerythroid colonies, but their types were distinct. When cocultured with the OP9, the splanchnopleura produced B cells expressing B220, CD19 and surface IgM. Using a three-step culture protocols with the OP9 expressing Delta-like 1 as feeders, the splanchnopleura produced immature T precursor cells (CD44-/CD25+) and more mature single positive T cells (CD4+/CD8-) after 16 days of incubation. However, the yolk sac failed to generate B and T lymphocytes under identical conditions. It is concluded that prior to linked embryonic circulation, the splanchnopleura other than the yolk sac had robust lymphoid potential in vitro. In the future, more reliable evidence from novel model animals will ultimately delineate the embryonic origin of lymphocytes in vivo.

Self-assembly of luminescent Sn(IV)/Cu/S clusters using metal thiolates as metalloligands.

Through the use of (Bu4N)2[Sn3S4(edt)3] (edt=SCH2CH2S(2-)) and Sn(SPh)4 as metalloligands, three neutral compounds have been obtained: [(Ph3P) 2Cu] 2SnS(edt)(2).2CH2Cl2.H2O (1a), [(Ph3P) 2Cu]2SnS(edt)2.2DMF.H2O (1b), and [(Ph3P)Cu] 2Sn(SPh)(6).3H 2O (2). Single-crystal X-ray diffraction studies revealed that compounds 1a and 1b contain the same neutral butterfly-like [(Ph3P)2Cu]2SnS(edt)2 cluster, which consists of one central SnS 5 dreich trigonal bipyramid sharing one vertex and two sides with two slightly distorted CuS 2P2 tetrahedrons. Compound 2 has a linear [(Ph3P)Cu]2Sn(SPh)6 cluster that is composed of a central distorted SnS 6 octahedron sharing two opposite planes with two slightly distorted CuS 3P tetrahedrons. Compound 1a exhibited an emission at 568 nm (tau=12.86 micros) in the solid state, while in CH 2Cl 2 solution, 1a exhibited a green emission at 534 nm (tau=4.75 micros). Compound 2 showed an intense red emission at 696 nm (tau=3.64 micros) upon excitation at 307 nm in the solid state.

[Radiosensitivity of nasopharyngeal carcinoma in relation to cell cycle synchronization effect of tumor necrosis factor alpha].

OBJECTIVE: To investigate the effect of tumor necrosis factor alpha (TNFalpha) on radiosensitivity of nasopharyngeal carcinoma (NPC) in relation to TNFalpha-induced cell cycle synchronization. METHODS: The radio-resistance of a NPC cell line subclone CNE-2Z-S1 was verified by in vivo experiments and flow cytometry was performed to evaluate cell cycle synchronization in TNFalpha-treated CNE-2Z-S1 cells. The radiosensitivity of the cell synchronized CNE-2Z-S1 cells was determined by clone formation in vitro and in vivo experiment in nude mice. RESULTS: TNFalpha was capable of inducing cell cycle arrest and synchronization of CNE-2Z-S1 cells. Pretreatment with TNFalpha remarkably enhanced the radiosensitivity of CNE-2Z-S1 in vitro, and in vivo experiments with nude mice also suggested the role of TNFalpha in enhancing the radiosensitivity of NPC. CONCLUSION: TNFalpha can enhance the radiosensitivity of NPC cells by inducing cell cycle synchronization.