This study aimed to characterize the epidemiology, risk factors, and survival outcomes of differentiated thyroid carcinoma (DTC) patients aged <55 years with distant metastasis (DM). The Surveillance, Epidemiology, and End Results (SEER) database was researched retrospectively for young DTC patients. Multivariate Logistic and Cox regression analyses were used to identify DM predictors and survival determinants. The overall survival (OS) and cancer-specific survival (CSS) were evaluated and compared by Kaplan-Meier analysis and Log-Rank test, respectively. The risk and prognostic nomogram were constructed and validated using ROC curves, calibration plots, and decision curve analysis. A total of 68,373 young DTC patients were identified. The DM prevalence was 0.65% (95% CI: 0.59-0.72%), predominantly involving the lungs (35.8%). Independent risk factors for DM included male sex (OR=1.84), follicular histology (OR=2.43), advanced T4 (OR=22.3) and N1 stages (OR=4.91). The 5-year and 10-year OS rates for young DTC patients with DM were 83.6% and 73.3%, while the 5-year and 10-year CSS rates were 85.0% and 79.9%, respectively. Multivariate Cox analysis identified several independent prognostic factors for both OS and CSS including age, the presence of lung, bone, liver, and brain metastases, as well as the use of surgery and chemotherapy. Established prognostic nomograms demonstrated robust performance (C-index =0.87 for OS; C-index =0.85 for CSS), with time-dependent area under the curves (AUCs) exceeding 0.80 at 5, and 10 years. In conclusion, this study presents the first population-level analysis of young DTC patients with distant metastases and establishes SEER-derived predictive models for assessing risk and survival among these patients.

Background:Signet-ring cell carcinoma (SRCC) of the bile duct is a rare malignancy with poorly characterized clinical features and prognosis. Given its rarity and the critical need to differentiate its behavior from common cholangiocarcinoma (CCA), this population-based study aimed to characterize SRCC and compare survival outcomes with CCA, while developing prognostic nomograms for individualized prediction. Methods:We analyzed data from the Surveillance, Epidemiology, and End Results (SEER) database [2000-2021], identifying 98 SRCC and 18,979 CCA cases. Propensity score matching (PSM) analysis (1:5 ratio) was conducted to balance baseline characteristics between groups (98 SRCC vs. 490 CCA). Independent prognostic factors for overall survival (OS) and cancer-specific survival (CSS) were identified using multivariable Cox regression. These factors were incorporated into nomograms for OS and CSS prediction. Model performance was evaluated using the concordance index (C-index), calibration curves, receiver operating characteristic (ROC) curve analysis, and decision curve analysis (DCA). Results:SRCC predominantly involved the extrahepatic bile duct (86.7%) and exhibited poor survival [median OS (mOS) 7.0 months, 95% confidence interval (CI): 4.0-10.0; median CSS (mCSS) 8.0 months, 95% CI: 5.0-12.0]. After PSM, no significant survival difference was observed between SRCC and CCA (P>0.05). Older age and distant metastasis were independent factors associated with poor survival, whereas surgery and chemotherapy were independently associated with better survival. The nomograms demonstrated moderate predictive accuracy (C-index: OS 0.78, 95% CI: 0.71-0.84; CSS 0.78, 95% CI: 0.70-0.85). Calibration curves showed excellent agreement between predicted and observed survival, ROC analysis confirmed discriminative ability, and DCA indicated strong clinical utility. Conclusions:SRCC of the bile duct is a rare, aggressive malignancy with a prognosis similar to CCA. The developed nomograms, integrating readily available clinical factors, provide clinically applicable tools for individualized prognosis prediction.

Background Although expression of MTA1 inversely correlates with the nuclear localization of ERα, the effect and molecular mechanism of ERα regulation of MTA1 remain unknown. Methods Quantitative real-time PCR and western blot analyses were used to measure levels of MTA1. The effect on HCC cell proliferation and invasion was assessed by EdU incorporation assays and Transwell, respectively. ShRNA and dual-luciferase assays were used to investigate the regulatory relationship between MTA1 and ERα in cell lines. Results We found that MTA1 gene regulation by ERα may be influenced by nuclear corepressors. The MTA1 promoter has three functional ER-element half-sites that lead to decreased MTA1 transcription and expression. ERα overexpression suppressed the proliferation and invasion of hepatocellular carcinoma cells (HCC). In addition, overexpression of MTA1 attenuated ERα-mediated suppression of the proliferation and invasion of HCC cells and tumor formation in vivo. These results suggested feedback regulation between ERα and MTA1. In summary, our results demonstrated that ERα suppressed proliferation and invasion of human HCC cells through downregulation of MTA1 transcription. Conclusions Our study is an improved description of the mechanisms of the suppressive effect of ERα on HCCs, adding understanding to the gender disparity of HCC progression.

Objective The purpose of this study was to describe the clinicopathological characteristics and prognosis of primary small cell carcinoma of the breast (PSCCB) and compare PSCCB with breast invasive ductal carcinoma (IDC).Design A retrospective cohort study.Setting Data of patients with PSCCB and breast IDC were identified from the Surveillance, Epidemiology, and End Results (SEER) database between 2004 and 2016.Participants Eighty-three patients with PSCCB and 410 699 patients with breast IDC were enrolled in the present cohort study.Materials and methods Patients with PSCCB and breast IDC were identified from the SEER database between 2004 and 2016. The clinicopathological characteristics and survival of patients with PSCCB and IDC were compared. Propensity score matching (PSM) analysis was performed to adjust for differences in baseline characteristics when comparing overall survival (OS) and cancer-specific survival (CSS). Moreover, OS-/CSS-specific nomograms were established to predict the prognosis of PSCCB.Results Compared with IDC, PSCCB was significantly correlated with older age, male, higher pathological grade, higher TNM (tumour, node, metastases) stage, a higher proportion of triple-negative breast cancer, a lower proportion of ER/PR positivity and significantly worse clinical outcome. The median OS and CSS of patients with PSCCB were 23.0 m (95%CI 13.0 to 56.0) and 28.0 m (95%CI 18.0 to 66.0), respectively. The 5-year OS and CSS rates in the PSCCB group were 36.1% and 42.4%, respectively. In the matched cohort after PSM analysis, patients with PSCCB had significantly worse OS and CSS than IDC patients. Multivariate Cox regression analysis demonstrated that T stage and administration of chemotherapy were independent prognostic factors for both OS and CSS in patients with PSCCB. The C-index for OS-/CSS-specific nomogram was 0.75 (95%CI 0.66 to 0.85)/0.79 (95%CI 0.69 to 0.89), respectively. The calibration curve in the ROC analysis indicated that the predicted value was consistent with the actual observation value. Decision curve analysis suggested that the nomogram model has a significant positive net benefit from the risk of death and are better than the traditional TNM staging system.Conclusion PSCCB has distinct clinicopathological characteristics, and patients with PSCCB have significantly worse clinical outcomes than those with IDC.

Background: Intrahepatic cholangiocarcinoma (iCCA) is a highly malignant subtype of cholangiocarcinoma (CCA) with poor prognosis. In iCCA, the interplay between the stroma and tumor cells results in resistance to adjuvant chemotherapy. Increasing evidence indicates that miR-206 participates in tumor progression, but its role in iCCA is still unclear. The aim of this study was to identify dysregulated miR-206 expression in iCCA and to further explore the underlying mechanism. Methods: MiR-206 expression was proven to be downregulated in iCCA tissues by qPCR, and its correlation with clinical characteristics and prognosis was investigated. iCCA-derived cancer-associated fibroblast cells (CAFs) and normal fibroblast cells (NFs) were isolated and identified. MiR-206 was knocked in or down in CAFs and CCA cells, respectively, to explore the role of miR-206, and coculture of these treated CCAs and CAFs was conducted to explore the effects of miR-206 on their mutual promoting effects. Exosomes carrying miR-206 and an orthotopic mouse model were used to determine the inhibitory effects of miR-206 on iCCA deterioration in vivo. Results: We confirmed that miR-206 is a suppressor of iCCA. Overexpressing miR-206 in CCA cells inhibited cell proliferation, migration and invasion. When cocultured with CCA cells, NFs downregulated miR-206 expression, and NFs were susceptible to transforming into CAFs. Moreover, CAFs promoted CCA cell malignant behaviors and gemcitabine resistance. Overexpressing miR-206 in CAFs or CCA cells inhibited this mutual promoting effect. Additionally, when delivered by exosomes, miR-206 suppressed tumor deterioration. And combined with gemcitabine, this treatment resulted in a longer survival time. Conclusion: Our study explained that the interaction between CCA cells and CAFs promoted iCCA deterioration. As a suppressive factor, miR-206 inhibited aggressive characteristics and gemcitabine resistance by interfering with this mutual promoting effect. This research elucidated the molecular mechanism underlying the unfavorable chemotherapeutic response of patients with iCCA, which provided a promising target for iCCA treatment.

目的 探讨环状 RNA-MYC-overlap.2(circ-MYC-overlap.2)/微小 RNA-19a-3p(miR-19a-3p)在胃癌的表达及其临床意义.方法 采用RT-PCR检测56例胃癌组织及其癌旁正常组织以及56例胃癌患者和50例健康人群血清中circ-MYC-overlap.2、miR-19a-3p的表达,并分析circ-MYC-overlap.2表达与胃癌患者临床特征的相关性.采用Kaplan-Meier生存曲线分析胃癌患者血清中circ-MYC-overlap.2表达与术后5年生存期的关系.观察人胃癌细胞株SGC-7901中信号传导与转录激活因子3(STAT3)信号通路对circ-MYC-overlap.2表达的调控作用.结果 与癌旁正常组织相比,胃癌组织中circ-MYC-overlap.2表达上调,miR-19a-3p表达下调(P＜0.01);胃癌组织中circ-MYC-overlap.2与miR-19a-3p表达呈负相关(r=-0.503,P＜0.01).与健康人群相比,胃癌患者血清中circ-MYC-overlap.2表达上调,miR-19a-3p表达下调(P＜0.01);胃癌患者血清中circ-MYC-overlap.2 与 miR-19a-3p表达呈负相关(r=-0.658,P＜0.01).TNM分期Ⅲ和Ⅳ期、伴有淋巴结或远处转移患者的胃癌组织和血清中circ-MYC-overlap.2表达分别高于TNM分期Ⅰ和Ⅱ期、未转移患者(P＜0.05).circ-MYC-overlap.2低表达患者5年生存率高于circ-MYC-overlap.2高表达患者(P＜0.01).在 SGC-7901 细胞中,IL-6 可上调 circ-MYC-overlap.2 表达(P＜0.01),而加入STAT3抑制剂硝呋齐特后circ-MYC-overlap.2表达下调(P＜0.01).结论 circ-MYC-overlap.2可作为判断胃癌患者临床分级、转移及预后的潜在标志物.circ-MYC-overlap.2可能通过吸附效应下调miR-19a-3p的作用.

Objective:To study gene therapy for Wilson′s disease (WD) with adeno-associated virus (AAV)-clustered regularly interspaced short palindromic repeats (CRISPR) system in vitro. Methods:Three small-guide RNAs (sgRNAs) were designed and AAV particles containing CRISPR-sgRNA1, 2, 3 or W/O homologous template (HT) were transduced into Toxic milk (TX) mouse hepatocytes, respectively. Gene editing efficiency and HT repair efficiency were evaluated by sequencing, and repair template-specific primers were used for PCR to verify HT incorporation.Results:Sanger sequencing data showed that the editing efficiency of sgRNA1 reached (20.2±2.3)%, which was significantly higher than others ( P<0.05). When AAV-HT treatment was combined, the corrected ATP7B gene band could be amplified by PCR, and NGS showed that repair efficiency was (7.9±2.7)%. Conclusion:AAV transduction can achieve pretty high editing efficiency. We selected sgRNA1 and verified its editing efficiency and the repair efficiency of HT, providing a theoretical and experimental basis for further in vivo experiments.

Acute rejection is commonly encountered for long-term survival in liver transplant (LT) recipients and may impact their long-term survival if rejection is severe or recurrent. The aim of this study is to examine the therapeutic potential of transforming growth factor (TGF-β)-overexpressing mesenchymal stem cells (MSCs) in inducing a local immunosuppression in liver grafts after transplantation. MSCs were transduced with a lentiviral vector expressing the human TGF-β1 gene; TGF-β1-overexpressing MSCs (designated as TGF/MSCs) were then transfused into the liver grafts via the portal vein of a rat LT model of acute rejection. Rejection severity was assessed by clinical and histologic analysis. The immunity suppression effects and mechanism of TGF/MSCs were tested, focusing on their ability to induce generation of regulatory T cells (Tregs) in the liver grafts. Our findings demonstrate that transfusion of TGF/MSCs prevented rejection, reduced mortality, and improved survival of rats after LT. The therapeutic effects were associated with the immunosuppressive effects of MSCs and TGF-β1. Their reciprocal effects on Tregs induction and function resulted in more CD4 + Foxp3 + Helios- induced Tregs, fewer Th17 cells, and improved immunosuppressive effects in local liver grafts. Thus, TGF/MSCs can induce a local immunosuppressive effect in liver grafts after transplantation. The immunomodulatory activity of TGF-β1 modified MSCs may be a gateway to new therapeutic approaches to prevent organ rejection in clinical transplantation. Stem Cells 2016;34:2681-2692.

Objectives Costimulatory signals are essential for T-cell activation and hence play a very important role in antitumor immunity. B7 and 4-1BBL which belongs to tumor necrosis factor (TNF) family provide costimulatory interaction for T-cell activation and function. This study investigated the role of B7 and 4-1BBL in the amplification of tumor immunity by transduction of the B7-1, B7-2 and 4-1BBL into mouse hepatocellular carcinoma cell line H22. Methods The tumorigenicity of H22 variants expressing either B7-1, B7-2 (H22/B7-1/B7-2) or 4-1BBL was compared with an H22 variant expressing B7-1, B7-2 and 4-1BBL (H22/B7-1/B7-2/4-1BBL). The study next investigated whether the combination of B7-1/B7-2 and 4-1BBL cell injection induced cytotoxic T lymphocyte (CTL) response and IL-2/IFN-γ secretion. The immune mechanisms underlying this combination treatment were then analyzed. Results Syngeneic BALB/c mice injected with H22/B7-1/B7-2/4-1BBL cells that expressed elevated levels of B7-1, B7-2 and 4-1BBL showed a tumor development frequency of 50% compared with 100% in mice injected with the H22 parental line, H22/neo, H22/B7-1/B7-2 and H22/4-1BBL. Mice inoculated with H22 tumor cells expressing B7-1, B7-2 and 4-1BBL developed a strong cytotoxic T lymphocyte response and long-term immunity against wild-type tumor, suggesting a synergistic effect between the B7 and 4-1BBL costimulatory pathways. Results showed that H22/B7-1/B7-2/4-1BBL tumor vaccines probably protect the infiltrating lymphocytes from apoptosis and induce NF-κB activation to improve T-cell-mediated antitumor response. Conclusions In this study, the antitumor consequences of using B7-1, B7-2 and 4-1BBL gene transfer have demonstrated the therapeutic potential of gene therapy approach for hepatocellular carcinoma.

Objective:To investigate the expression of metastasis associated 1(MTA1) in human hepatocellular carcinoma(HCC) and adjacent tissues,and its correlation with survival and recurrence,to consequenthy determin whether MTA1 is a suitable prognostic index for prognosis or not.Methods:The transcription and translation of MTA1 gene in HCC tissue,corresponding adjacent tissue,and normal tissue samples were determined by quantity real-time PCR and immunohistochemistry(IHC),and the correlation between MTA1 and survival and recurrence of HCC patients was analyzed by Kaplan-Meier curve followed by log-rank test.Results:The transcription and translation of MTA1 gene were significantly higher in the HCC tissues and the adjacent cirrhotic tissues compared to the normal liver tissues(P 0.01).Expression level of MTA1 was related to survival and recurrence rates of the HCC patients.Conclusion:MTA1 was a suitable prognostic index predicting survival and recurrence after surgical resection of HCC.

Objective To determine whether interleukin(IL)-1α regulates hepatocellular carcinoma(HCC) development in humans.Methods HBV-associated HCC tissues,the tumor-adjacent tissues and normal liver tissues were obtained from 80 male and 36 female patients.IL-1α,estrin receptorα(ERα),IL-6,and MyD88 were quantified by RT-PCR.Dynamic expressions of IL-1α and IL-6 were detected in the DEN-induced mice HCC models as well.Results Compared with normal liver tissues,IL-1α was highly expressed in the tumor-adjacent tissues of male HCC cases(P<0.01),which was not in the female HCC subjects.Compared with normal liver tissues,the expression of ERα was dramatically decreased in HCC and the tumor-adjacent tissues of male HCC cases(P<0.01).A linear relationship was observed between increased IL-1α and decreased ERα expression in the tumor-adjacent tissues of male HCC cases(r=-0.61,P<0.01).In the DEN-induced HCC mice models,IL-1α was highly expressed in the xenografts,which was gradually decreased as experimental HCC grew.Conclusion IL-1α is an essential factor in the development of human HCC in male and may be a valuable index for clinical diagnosis and prediction of reoccurrence of HCC.

Objective:To investigate the inhibitory effect on the orthotopic transplantation tumor model of human hepatocellular carcinomain using hydrodynamics-based transfection of recombinant retrovirus vector containg IκBα super repressor gene and its possible mechanism.Methods:The animal model of orthotopic transplantation tumor in nude mice was established with MHCC97-H and MHCC97-L cell lines.Tumor-bearing nude mice were divided into 4 groups:MHCC97-H group(control group),MHCC97-H+IκBαSR group(transfection group),MHCC97-L group(control group)and MHCC97-L+IκBαSR group(transfection group).After treatment for 7 days,plasmid pBABE-puro-IκBαSR and plasmid pBABE-puro in PBS solution were diluted according to the mouse weight.Plasmid pBABE-puro-IκBαSR and plasmid pBABE-puro were respectively transferred into transfection group and control group via the tail vein by hydrodynamic injection.The survival rate of nude mice bearing the tumor was observed.The size,weight and volume of tumor in situ and the number of metastatic tumor in liver were examined,and the inhibitory rates of tumor growth were calculated.ALT and AST in sera were observed.Immunohistochemistry was used to detect the expression of NF-κB in liver cancer tissue.IκBα was determined with western blot.Results:Plasmid pBABE-puro-IκBαSR significantly promoted the survival rate and inhibited the level of ALT and AST in transfection groups than in control groups.In transfection groups,the tumors were significantly smaller and lighter than those in control groups,and the inhibition rates were 29.3% and 33.0% respectively.In cellular nucleus of the transfection groups,the ex pression levels of NF-κB p65 and IκBα were higher than the control groups.Conclusion:Hydrodynamics-based transfection of plasimd pBABE-puro-IκBαSR gene to the orthotopic implant model of human hepatocellular carcinoma can inhibit the tumor.Its mechanism may be that the transfected IκBαSR gene suppress the activation of NF-κB signaling in cancer cells.

Objective: To investigate the efficacy of decision tree incorporated with indocyanine green clearance test on hepatic reserve function in patients with hepatocellular carcinoma.Methods:Eighty-two patients with hepatocellular carcinoma confirmed by pathology were enrolled.The presence of ascites,serum bilirubin level and indocyanine green retention rate at 15 minutes were elevated.Results:During strict application of this decision tree to 82 hepatectomies,no postoperative mortality(overall morbidity of 0%) and no major complications were observed.Conclusion:The decision tree can provide more precise information of liver function so as to avoid liver failure of patients with hepatocellular carcinoma.

Objective To evaluate the inhibitory effects of anti-telomerase siRNA in hepatocellular carcinoma cells.Methods The lentiviral expression vector of RNA interference(RNAi) targeting human telomerase reverse transcriptase(hTERT) was constructed.The plasmid pLenti-hTERT-siRNA and two packaging plasmids were cotransfected to 293T cells by calcium phosphate method.The hTERT mRNA was detected by RT-PCR.HepG2 cell proliferation in vitro was determined by MTT assay.Telomerase activity was detected by telomeric repeat amplification protocol(TRAP) assay.Results Restriction enzyme digestion demonstrated that the recombinant plasmid pLenti-hTERT-siRNA was constructed successfully.The recombinant lentivirus infected HepG2 efficiently.The effect of RNAi could be observed in siRNA group.The expression of hTERT mRNA was significantly down-regulated.Meanwhile,cell proliferation and tumor growth were significantly inhibited in vitro.Conclusion Growth of hepatocellular carcinoma cells is effectively inhibited after transfection with anti-telomerase siRNA in vitro.

Background Chronic hepatitis B virus (HBV) infection is an important cause of cirrhosis and hepatocellular carcinoma. The major challenges for current therapies are the low efficacy of current drugs and the occurrence of drug resistant HBV mutations. RNA interference (RNAi) of virus-specific genes offers the possibility of developing a new anti-HBV therapy. Recent reports have shown that lentiviral vectors based on HIV-1 are promising gene delivery vehicles due to their ability to integrate transgenes into non-dividing cells. Herein, a lentivirus-based RNAi system was developed to drive expression and delivery of HBV-specific short hairpin RNA (shRNA) in a mouse model for HBV replication. Methods Hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in the sera of the mice were analyzed by quantitative sandwich enzyme linked immunosorbent assay (ELISA) technique, hepatitis B core antigen (HBcAg) and HBsAg in the livers of the mice were detected by immunohistochemical assay, HBV DNA and HBV mRNA were measured by fluorogenic quantitative polymerase chain reaction (FQ-PCR) and quantitative real-time PCR respectively. Results Co-injection of HBV plasmids together with the lentivirus targeting HBV shRNA induced an RNAi response. Secreted HBsAg was reduced by 89% in mouse serum, and HBeAg was also significantly inhibited, immunohistochemical detection of HBcAg or HBsAg in the liver tissues also revealed substantial reduction. Lentiviral mediated shRNA caused a significant suppression in the levels of viral mRNA and DNA synthesis compared to the control group. Conclusion Lentivirus-based RNAi can be used to suppress HBV replication in vivo, it might become a potential therapeutic strategy for treating HBV and other viral infections.

OBJECTIVE:To study the efficacy of anti-telomerase siRNA in hepatocellular carcinoma both in vitro and in vivo.METHODS:Lentvirus vectors contained anti-telomerase siRNA were conducted with a high performance homologous recombination system, and then were transduced into human hepatocellular carcinoma HepG2 cells. The telomerase activity was detected by RT-PCR, HepG2 cell proliferation was determined by MTT assay, and apoptosis was detected by TUNEL assay. The in vivo experiment was carried out by inoculation of HepG2 cells into nude mice and the tumor growth was measured and analyzed.RESULTS:The growth of transfected HepG2 cells was significantly inhibited and the inhibition rate was 57.5% at the 8th day after transfection. The telomerase activity was significantly suppressed in vitro. The growth of transfected human hepatocellular HepG2 tumor in the nude mice was also significantly inhibited.CONCLUSION:The results of this study demonstrate that the growth of hepatocellular carcinoma cells is effectively inhibited by transfection of anti-telomerase siRNA both in vitro and in vivo.