Laparoscopic vs. open distal gastrectomy for locally advanced gastric cancer: A systematic review and meta-analysis of randomized controlled trials.

Objective: The aim of this systematic review and meta-analysis is to compare the short- and long-term outcomes of laparoscopic distal gastrectomy (LDG) with those of open distal gastrectomy (ODG) for patients with advanced gastric cancer (AGC) who exclusively underwent distal gastrectomy and D2 lymphadenectomy in randomized controlled trials (RCTs). Background: Data in published meta-analyses that included different gastrectomy types and mixed tumor stages prevented an accurate comparison between LDG and ODG. Recently, several RCTs that compared LDG with ODG included AGC patients specifically for distal gastrectomy, with D2 lymphadenectomy being reported and updated with the long-term outcomes. Methods: PubMed, Embase, and Cochrane databases were searched to identify RCTs for comparing LDG with ODG for advanced distal gastric cancer. Short-term surgical outcomes and mortality, morbidity, and long-term survival were compared. The Cochrane tool and GRADE approach were used for evaluating the quality of evidence (Prospero registration ID: CRD42022301155). Results: Five RCTs consisting of a total of 2,746 patients were included. Meta-analyses showed no significant differences in terms of intraoperative complications, overall morbidity, severe postoperative complications, R0 resection, D2 lymphadenectomy, recurrence, 3-year disease-free survival, intraoperative blood transfusion, time to first liquid diet, time to first ambulation, distal margin, reoperation, mortality, or readmission between LDG and ODG. Operative times were significantly longer for LDG [weighted mean difference (WMD) 49.2 min, p < 0.05], whereas harvested lymph nodes, intraoperative blood loss, postoperative hospital stay, time to first flatus, and proximal margin were lower for LDG (WMD -1.3, p < 0.05; WMD -33.6 mL, p < 0.05; WMD -0.7 day, p < 0.05; WMD -0.2 day, p < 0.05; WMD -0.4 mm, p < 0.05). Intra-abdominal fluid collection and bleeding were found to be less after LDG. Certainty of evidence ranged from moderate to very low. Conclusions: Data from five RCTs suggest that LDG with D2 lymphadenectomy for AGC has similar short-term surgical outcomes and long-term survival to ODG when performed by experienced surgeons in hospitals contending with high patient volumes. It can be concluded that RCTs should highlight the potential advantages of LDG for AGC. Systematic Review Registration: PROSPERO, registration number CRD42022301155.

A novel model based on necroptosis-related genes for predicting immune status and prognosis in glioma.

Background: Glioma is a highly aggressive brain cancer with a poor prognosis. Necroptosis is a form of programmed cell death occurring during tumor development and in immune microenvironments. The prognostic value of necroptosis in glioma is unclear. This study aimed to develop a prognostic glioma model based on necroptosis. Methods: A necroptosis-related risk model was constructed by Cox regression analysis based on The Cancer Genome Atlas (TCGA) training set, validated in two Chinese Glioma Genome Atlas (CGGA) validation sets. We explored the differences in immune infiltration and immune checkpoint genes between low and high risk groups and constructed a nomogram. Moreover, we compiled a third validation cohort including 43 glioma patients. The expression of necroptosis-related genes was verified in matched tissues using immunochemical staining in the third cohort, and we analyzed their relationship to clinicopathological features. Results: Three necroptosis-related differentially expressed genes (EZH2, LEF1, and CASP1) were selected to construct the prognostic model. Glioma patients with a high risk score in the TCGA and CGGA cohorts had significantly shorter overall survival. The necroptosis-related risk model and nomogram exhibited good predictive performance in the TCGA training set and the CGGA validation sets. Furthermore, patients in the high risk group had higher immune infiltration status and higher expression of immune checkpoint genes, which was positively correlated with poorer outcomes. In the third validation cohort, the expression levels of the three proteins encoded by EZH2, LEF1, and CASP1 in glioma tissues were significantly higher than those from paracancerous tissues. They were also closely associated with disease severity and prognosis. Conclusions: Our necroptosis-related risk model can be used to predict the prognosis of glioma patients and improve prognostic accuracy, which may provide potential therapeutic targets and a theoretical basis for treatment.

A modified LC-MS/MS method for the detection of whole blood tacrolimus and its clinical value in Chinese kidney transplant patients.

Background: For patients who treated with tacrolimus after kidney transplant, therapeutic drug monitoring is essential to improve their prognosis. However, previous detection methods have limitations, such as the overestimation and unacceptable bias in the immunoassays. Precision medicine has been challenged. The liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is recognized as the gold standard due to its accuracy and specificity, but lack of throughput and complex process limits its clinical application. Therefore, an accurate, simple and high throughput method for tacrolimus monitoring is needed for clinical practice. Methods: A modified LC-MS/MS method was introduced and validated. Whole blood samples were prepared by a one-step protein precipitation method. Chromatographic separation was achieved using a Phenomenex Kinetex 2.6 µm XB-C18 2.1 × 50 mm column with a total run time of 3.5 min to avoid matrix effect. An electrospray ionization source (ESI) was used in positive ion multiple reaction monitoring (MRM) mode for mass spectrometric detection. In order to protect the mass spectrometer, only part of the sample after LC separation was allowed to enter the mass spectrum, through a two HPLC systems coupled one mass spectrometry design. In this way, the instrument throughput is also improved and realizing the detection of 2 samples within 3.5 min and carried out a shorter analyzing time for each sample of 1.75 min. Additionally, we calculated tacrolimus-intrapatient variant (Tac-IPV) based on this modified method and assessed the prognostic value of Tac-IPV in Chinese kidney transplant patients. Results: The LC-MS/MS was modified by streamlining the procedure and increasing the throughput. The method proved to be accurate and reproducible with all performance parameters suitably meeting the clinical requirements over a calibration ranged from 0.37 to 42.90 ng/mL. Parameters such as linearity, limit of quantification (LoQ) and dilution integrity were validated with a clinical reportable range from 0.37 to 343.20 ng/mL, which was particularly useful for high drug concentrations patients (rare but very serious). Both cross-contamination and matrix effects were negligible. Clinical data of 83 patients showed that Tac-IPV was associated with poor kidney transplant outcome in Chinese (Hazard Ratio (HR) = 3.96, 4.75; 95% Cl: 1.10-14.21, 1.23-18.36; P < 0.05). Conclusions: This modified LC-MS/MS method possessed high throughput and simple sample preparation, allowing it to meet daily clinical needs. At the same time, Tac-IPV based on this modified LC-MS/MS had excellent prognostic value in kidney transplantation. These advantages have great significance for the individualized treatment of Chinese kidney transplant patients and broad application of Tac-IPV.

Long Noncoding RNA RP11-732M18.3 Promotes Glioma Angiogenesis by Upregulating VEGFA.

Gliomas are the most aggressive and common type of malignant brain tumor, with limited treatment options and a dismal prognosis. Angiogenesis, a hallmarks of cancer, is one of two critical events in the progression of gliomas. Accumulating evidence has demonstrated that in glioma dysregulated molecules like long noncoding RNAs (lncRNAs), are closely linked to tumorigenesis and prognosis. However, the effects of and mechanisms of action of lncRNAs during tumor angiogenesis are poorly understood. The effect of lncRNA RP11-732M18.3 on angiogenesis was elucidated through an intracranial orthotopic glioma model, immunohistochemistry, and an in vitro angiogenesis assay. Co-culture experiments and cell migration assays were performed to investigate the function of lncRNA RP11-732M18.3 in vitro. lncRNA RP11-732M18.3 increased CD31+ microvessel density, and overexpression of lncRNA RP11-732M18.3 resulted in poor mouse survival. lncRNA RP11-732M18.3 promoted endothelial cell migration and tube formation. Nomogram and Kaplan-Meier survival analyses indicated that higher VEGFA is correlated with a poor prognosis. Mechanistically, lncRNA RP11-732M18.3 promotes angiogenesis by increasing the nuclear level of EP300 and facilitating the transcription and secretion of VEGFA. Our study contributes to the latest understanding of glioma angiogenesis and prognosis. lncRNA RP11-732M18.3 may be a potential treatment target in glioma.

Patient-based real-time quality control for quantitative hepatitis B virus DNA test using moving rate of positive and negative patient results.

OBJECTIVES: Patient-based real-time quality control (PBRTQC) has gained increasing attention in the field of laboratory quality management in recent years. However, PBRTQC has not been reported for use in molecular diagnostics. This study introduces PBRTQC to quantitative hepatitis B virus (HBV) DNA test using moving rate (MR) of positive and negative patient results. METHODS: In contrast to the MR protocols described in other literature, MR protocol for HBV-DNA test has an additional logarithmic transformation and binary conversion steps before using a common statistical process control algorithm, such as the MR. We used all patient test results of HBV-DNA assay from August 2018 to August 2021 at the Second Affiliated Hospital of Guangzhou University of Chinese Medicine, for parameters setting, optimization, and performance validation. The false rejection rate, error detection curves and validation charts were used to assess the MR protocols. RESULTS: The false rejection rates of two MR protocols were both <0.7%. The optimal block sizes for positive and negative errors in each cut-off value were not the same, so we first proposed a combined protocol that used different block size to detect negative and positive errors. It turned out that the combined protocols outperformed the simple protocols for each cut-off value, especially detecting positive errors. CONCLUSIONS: The performances of MR protocols using positive or negative patient results to detect constant errors of HBV-DNA test could meet laboratory requirements. Therefore, we have provided an effective alternative tool for internal quality control in the field of molecular diagnostics.

Long non­coding RNA AL355711 promotes smooth muscle cell migration through the ABCG1/MMP3 pathway.

Atherosclerosis and related cardiovascular diseases pose severe threats to human health worldwide. There is evidence to suggest that at least 50% of foam cells in atheromas are derived from vascular smooth muscle cells (VSMCs); the first step in this process involves migration to human atherosclerotic lesions. Long non­coding RNAs (lncRNAs) have been found to play significant roles in diverse biological processes. The present study aimed to investigate the role of lncRNAs in VSMCs. The expression of lncRNAs or mRNAs was detected using reverse transcription­quantitative polymerase chain reaction. The Gene Expression Omnibus datasets in the NCBI portal were searched using the key words 'Atherosclerosis AND tissue AND Homo sapiens' and the GSE12288 dataset. Gene expression in circulating leukocytes was measured to identify patients with coronary artery disease (CAD) or controls, and used to analyze the correlation coefficient and expression profiles. The protein level of ATP­binding cassette sub­family G member 1 (ABCG1) and matrix metalloproteinase (MMP)3 was determined using immunohistochemistry and western blot analysis. The analysis of mouse aortic roots was performed using Masson's and Oil Red O staining. The expression of lncRNA AL355711, ABCG1 and MMP3 was found to be higher in human atherosclerotic plaques or in patients with atherosclerotic CAD. The correlation analysis revealed that ABCG1 may be involved in the regulation between lncRNA AL355711 and MMP3 in atherosclerotic CAD. The knockdown of lncRNA AL355711 inhibited ABCG1 transcription and smooth muscle cell migration. In addition, lncRNA AL355711 was found to regulate MMP3 expression through the ABCG1 pathway. The expression of ABCG1 and MMP3 was found to be high in an animal model of atherosclerosis. The results indicated that lncRNA AL355711 promoted VSMC migration and atherosclerosis partly via the ABCG1/MMP3 pathway. On the whole, the present study demonstrates that the inhibition of lncRNA AL355711 may serve as a novel therapeutic target for atherosclerosis. lncRNA AL355711 in circulating leukocytes may be a novel biomarker for atherosclerotic CAD.

Work-Related Musculoskeletal Disorders Among Hospital Midwives in Chenzhou, Hunan Province, China and Associations with Job Stress and Working Conditions.

BACKGROUND: Midwives have a high risk of work-related musculoskeletal disorders (WMSDs), which are the leading cause of pain and disability. However, the association between job stress and WMSDs among midwives has not been studied. The aim of this project was to specifically describe relationships between the job stress and WMSDs among a sample of hospital midwives in Chenzhou, Hunan Province, China. METHODS: We conducted a cross-sectional study among a sample of 769 eligible midwives in the city of Chenzhou in Hunan Province, China from May 2018 to January 2019. All participants completed a questionnaire regarding the presence and severity of different pain symptoms and job stress assessed by the Korean occupational stress scale. RESULTS: A total of 597 participants responded (77.6%), and 491 (82.2%) reported that they had experienced WMSDs at some time over a 12-month period. The most common pain site was low back (72.7%), followed by the neck (52.8%) and shoulders (42.7%). We revealed that various aspects of job stress were associated with WMSDs in the current study, namely "Owing to my workload, I always feel time pressure" (OR, 2.05; 95% CI: 1.28-3.28), "My job has become increasingly overbearing" (OR, 2.34; 95% CI: 1.46-3.77), "My work requires long lasting concentration" (OR, 3.50; 95% CI: 2.13-5.74), "I have to do various jobs simultaneously", (OR, 3.15; 95% CI: 1.93-5.14)), "My work requires creativity" (OR, 2.15; 95% CI: 1.31-3.54), "My work requires a high level of skill or knowledge" (OR, 2.83; 95% CI: 1.67-4.80), "My supervisor is helpful in getting the job done" (OR, 0.53; 95% CI: 0.33-0.84), "I have someone who understands my difficulties at work" (OR, 0.53; 95% CI: 0.34-0.85), "Undesirable changes (ie, downsizing) will come to my job" (OR, 3.28; 95% CI: 2.01-5.77), "My company provides me with sufficient organizational support" (OR, 0.47; 95% CI: 0.29-0.74), "Departments cooperate each other without conflicts" (OR, 0.50; 95% CI: 0.32-0.80), "I am provided with the opportunity to develop my capacity" (OR, 0.57; 95% CI: 0.36-0.91) and "My company climate is authoritative and hierarchical" (OR: 3.21; 95% CI: 1.97-5.23). CONCLUSION: Overall, this study suggests that job stress has an important influence on WMSDs among a sample of hospital midwives in Chenzhou, Hunan Province, China. Given the multifaceted nature of identified risk, a comprehensive approach to reduce this risk is clearly required and a job stress management program will be essential.

Massive Open Online Courses-based blended versus face-to-face classroom teaching methods for fundamental nursing course.

ABSTRACT: An increasing number of studies focus on the effectiveness of Massive Open Online Courses (MOOC)-based blended learning, whereas none have yet studied using it for teaching fundamental nursing skills at an undergraduate level.To evaluate the effectiveness of MOOC-based blended learning versus face-to-face classroom teaching techniques within the fundamental nursing course at the Faculty of Nursing, University of Xiang Nan, China.This cluster randomized controlled trial enrolled 181 students and assigned them into either an MOOC-based blended or a face-to-face classroom teaching group, both involving the Fundamental Nursing course for undergraduate nursing students. The analyzed outcomes included test scores, critical thinking ability, and feedback received from the students on the Fundamental Nursing course.MOOC-based blended techniques versus face-to-face classroom teaching methods demonstrated higher daily performance (P = .014), operational performance (P = .001), theoretical achievements (P < .001), and final grades (P < .001) in Fundamental Nursing.Moreover, the mean change in the participants' critical thinking ability items between groups were, mostly, statistically significant. The items focusing on the feedback from the students demonstrated significant differences between the groups in terms of their satisfaction with the teaching they received (P < .001) and the overall learning effects (P = .030).This study confirmed that receiving MOOC-based blended learning was superior when compared against face-to-face classroom teaching techniques for learning within the Fundamental Nursing course.

Moving Rate of Positive Patient Results as a Quality Control Tool for High-Sensitivity Cardiac Troponin T Assays.

BACKGROUND: A small shift in high-sensitivity cardiac troponin T (hs-cTnT) assays can lead to different result interpretation and consequent patient management. We explored whether a small bias could be detected using conventional internal quality control (QC) procedures, evaluated the performance of moving average (MA)-based QC procedures, and proposed a new QC procedure based on the moving rate (MR) of positive patient results of hs-cTnT assays. METHODS: The ability of conventional QC to detect a 5 ng/L bias was examined using the13s/ 22s/R4s multi-rule procedure as deviation rules.We developed MA and MR procedures for the hs-cTnT assay using eight months of patient data. The performance of different MA or MR procedures was investigated by calculating the median number of patient samples affected until a bias introduced into the dataset was detected (MNPed). After comparing the MNPed across different procedures, we selected an optimal MA or MR procedure for validation. Validation graphs were plotted using the minimum, median, and maximum number of results affected until bias detection. RESULTS: Our conventional QC procedures could not detect a positive bias of 5 ng/L. When a positive bias was introduced, MNPed was much higher using MA than using MR, with cut-off values of 5 ng/L and 14 ng/L, respectively. MR validation charts for optimal procedures provided insight into the MR performance. CONCLUSIONS: The MR procedure could detect different errors with few false alarms. In the hs-cTnT assay, the MR procedure with a smaller cut-off value outperformed MA and conventional QC procedures for small bias detection.

Prevalence of Percutaneous Injuries and Associated Factors Among a Sample of Midwives in Hunan Province, China.

Background: Percutaneous injuries and blood-borne-related infections pose occupational hazards to healthcare professionals. However, the prevalence and associated factors for these hazards among midwives in Hunan Province, China are poorly documented. Methods: A cross-sectional study was conducted among a sample of 1,282 eligible midwives in the cities of Yongzhou, Chenzhou, Hengyang, and Changsha in Hunan Province, China, from January 2017 to July 2017. The association of selected independent variables with percutaneous injuries was investigated using binary logistic regression. Results: 992 participants responded (77.3%), and within the previous 12 months, 15.7% experienced percutaneous injuries. In multivariate analysis, hospital size, age, length of employment as a midwife, weekly working hours, and three aspects of Hospital Safety Climate Scale were associated with percutaneous injuries. The risk of percutaneous injuries among the midwives working in hospitals with ≤399 beds was higher than that among those working in hospitals with ≥400 beds by nearly 3 times. Furthermore, the percutaneous injury prevalence of midwives decreased as age increased. Moreover, the probability of percutaneous injuries among the midwives with weekly working hours of >40 was 4.35 times higher compared with that among midwives with weekly working hours of ≤40. Conclusion/Application to practice: The prevalence of percutaneous injuries among midwives in the study hospitals was substantial. Our results further proved that risk mitigation strategies tailored to midwives are needed to reduce this risk. These strategies include ensuring a positive organizational climate, providing highly safe devices, and reducing the workload.

Deafness Gene Mutations in Newborns in the Foshan Area of South China With Bloodspot-Based Genetic Screening Tests.

Purpose The aim of this study was to determine the rate of deafness gene mutations in the Foshan area of South China. Method We enrolled the infants delivered in Foshan Maternity and Children's Healthcare Hospital. Deafness gene mutation was detected by HibriMax method. Our study tested 47,538 newborns within 3 days after birth, including 13 sites in four genes: GJB2 (c.35 del G, c.176 del 16, c.235 del C, c.299 del AT, c.155 del TCTG), GJB3 (c.583 C>T), SLC26A4 (c.2168 A>G, c.919-2 A>G, c.1299 C>T), and mtDNA 12S rRNA (m.1555 A>G, m.1494 C>T, m.12201 T>C, m.7445 A>G). The birth condition of infants was collected, including sex, low or high birth weight, twins, and premature delivery. Results In a total of 47,538 newborns, 1,415 were positively identified with deafness gene mutations. The total rate of the deafness gene mutation was 2.976%. The carrier rates of GJB2 (c.35 del G, c.176 del 16, c.235 del C, c.299 del AT, c.155 del TCTG), GJB3 (c.583 C>T), SLC26A4 (c.2168 A>G, c.919-2 A>G, c.1299 C>T), and mtDNA 12S rRNA (m.1555 A>G, m.1494 C>T, m.12201 T>C, m.7445 A>G) mutations were 0.000%, 0.048%, 1.422%, 0.185%, 0.000%, 0.076%, 0.116%, 0.755%, 0.160%, 0.187%, 0.021%, 0.000%, and 0.006%, respectively. Conclusions Our study showed that the c.235 del C GJB2 mutation was the leading deafness-related mutation in the Foshan area of South China. Deafness gene mutations screening in newborns detected by bloodspot-based genetic screening tests can help the diagnosis of newborn congenital hearing loss.

Metabolic classification of circulating tumor cells as a biomarker for metastasis and prognosis in breast cancer.

BACKGROUND: Circulating tumor cells (CTCs) has been demonstrated as a promising liquid biopsy marker for breast cancer (BC). However, the intra-patient heterogeneity of CTCs remains a challenge to clinical application. We aim at profiling aggressive CTCs subpopulation in BC utilizing the distinctive metabolic reprogramming which is a hallmark of metastatic tumor cells. METHODS: Oncomine, TCGA and Kaplan-Meier plotter databases were utilized to analyze expression and survival relevance of the previously screened metastasis-promoting metabolic markers (PGK1/G6PD) in BC patients. CTCs detection and metabolic classification were performed through micro-filtration and multiple RNA in situ hybridization using CD45 and PGK1/G6PD probes. Blood samples were collected from 64 BC patients before treatment for CTCs analysis. Patient characteristics were recorded to evaluate clinical applications of CTCs metabolic subtypes, as well as morphological EMT subtypes classified by epithelial (EpCAM/CKs) and mesenchymal (Vimentin/Twist) markers. RESULTS: PGK1 and G6PD expressions were up-regulated in invasive BC tissues compared with normal mammary tissues. Increased tissue expressions of PGK1 or G6PD indicated shortened overall and relapse-free survival of BC patients (P < 0.001). Blood GM+CTCs (DAPI+CD45-PGK1/G6PD+) was detectable (range 0-54 cells/5 mL) in 61.8% of tCTCs > 0 patients. Increased GM+CTCs number and positive rate were correlated with tumor metastasis and progression (P < 0.05). The GM+CTCs ≥ 2/5 mL level presented superior AUC of ROC at 0.854 (95% CI 0.741-0.968) in the diagnosis of BC metastasis (sensitivity/specificity: 66.7%/91.3%), compared with that of tCTCs (0.779) and CTCs-EMT subtypes (E-CTCs 0.645, H-CTCs 0.727 and M-CTCs 0.697). Moreover, GM+CTCs+ group had inferior survival with decreased 2 years-PFS proportion (18.5%) than GM+CTCs- group (87.9%; P = 0.001). CONCLUSIONS: This work establishes a PGK1/G6PD-based method for CTCs metabolic classification to identify the aggressive CTCs subpopulation. Metabolically active GM+CTCs subtype is suggested a favorable biomarker of distant metastasis and prognosis in BC patients.

Metabolic reprogramming-based characterization of circulating tumor cells in prostate cancer.

BACKGROUND: Circulating tumor cells (CTCs), an advantageous target of liquid biopsy, is an important biomarker for the prognosis and monitoring of cancer. Currently, detection techniques for CTCs are mainly based on the physical and/or epithelial characteristics of tumor cells. However, biofunctional activity markers that can indicate the high metastatic capacity of CTCs are lacking. METHODS: Functional microarray, quantitative real-time polymerase chain reaction, and Western blot were used on five prostate cancer cell lines with different metastatic capacities to identify the metastasis-related metabolic genes. The identified genes were detected in the CTCs of 64 clinical samples using the RNA in situ hybridization. A multi-criteria weighted model was used to determine the optimal metabolic markers for the CTCs test. Based on five fluorescent signals targeting DAPI, CD45, metabolic, epithelial (EpCAM/CKs), and mesenchymal (Vimentin/Twist) markers, the filtration-enriched CTCs were classified as GM+CTCs/GM-CTCs (metabolic types) or E-CTCs/H-CTCs/M-CTCs (EMT types). Correlation analysis and ROC curve were conducted on 54 prostate cancer samples to evaluate the clinical significance of CTCs subtypes. RESULTS: Eight metastasis-related metabolic genes were identified, including HK2, PDP2, G6PD, PGK1, PHKA1, PYGL, PDK1, and PKM2. Among them, PGK1 and G6PD were determined as optimal glucose metabolic (GM) markers for CTCs. GM+CTCs (marked by PGK1/G6PD) were detectable in 64.8% (35/54) of prostate cancer patients, accounting for 46.5% (134/288) of total CTCs. An increased GM+CTCs level was associated with advanced tumor stage and metastasis (P <  0.05). In the discrimination of cancer metastasis from non-metastasis, GM+CTCs presented a higher AUC of the ROC curve (0.780) compared with the EMT CTCs subtypes (E-CTCs 0.729, H-CTCs 0.741, and M-CTCs 0.648). A triple tPSA-Gleason-GM+CTCs marker increased the AUC to 0.904, which was better than that of the tPSA-Gleason-H-CTCs marker (0.874). CONCLUSIONS: The metabolic marker (PGK1/G6PD) is determined as the indicator for the biofunctional activity analysis of CTCs, compared with the existing morphological (EMT) classification on CTCs. The metabolic characterization of CTCs demonstrates that hypermetabolic GM+CTCs are promising biomarkers for prostate cancer metastasis.

Long non-coding RNA UBE2CP3 enhances HCC cell secretion of VEGFA and promotes angiogenesis by activating ERK1/2/HIF-1α/VEGFA signalling in hepatocellular carcinoma.

BACKGROUND: Angiogenesis is considered as an important process in the development of malignancies and is associated with cancer progression and metastasis. Hepatocellular carcinoma (HCC) is the most common primary tumor of the liver and is recognized as a typical angiogenic tumor. Thus, it is of great importance to study the underlying mechanism of angiogenesis in HCC. The long non-coding RNA (lncRNA) ubiquitin conjugating enzyme E2C pseudogene 3 (UBE2CP3) has been reported as an oncogene that promotes tumor metastasis in HCC. However, the role and underlying mechanisms of UBE2CP3 in HCC angiogenesis are still unclear. METHODS: We measured the expression levels of UBE2CP3 by in situ hybridization (ISH) and quantitative real-time polymerase chain reaction (qRT-PCR) in HCC patient samples. We also concomitantly used CD31/PAS double-staining to measure endothelial vessel (EV) density and used qRT-PCR to measure the CD31 mRNA level. HepG2 and SMMC-7721 cells were transfected with Lv-UBE2CP3 or Sh-UBE2CP3 virus to obtain stably over-expressing or knocking-down UBE2CP3 cell lines. The indirect effects of UBE2CP3 on ECs were studied by establishing a co-culture system using Transwell chambers with a 0.4-µm pore size. HCC cells and ECs in the co-culture system were separated, but the cytokines and growth factors were able to communicate with each other. Following exposed to HCC cells, ECs were collected for functional studies. Finally, we studied the function of UBE2CP3 in vivo by chick embryo chorioallantoic membrane (CAM) angiogenesis assays and nude mouse tumorigenicity assays. RESULTS: In this study, we found that UBE2CP3 expression was higher in HCC tissues than in para-tumor tissues and was up-regulated in tissues with high EV density. Functionally, we found that in the co-culture systems, HCC cells overexpressing UBE2CP3 promoted HUVEC proliferation, migration and tube formation via the activation of ERK/HIF-1α/p70S6K/VEGFA signalling, increasing the level of VEGFA in HCC cell supernatant. In addition, the opposite results appeared when the expression of UBE2CP3 in HCC cells was knocked down. Consistent with these results, CAM angiogenesis assays and nude mouse tumorigenicity assays showed that UBE2CP3 expression up-regulated EV density in vivo. CONCLUSION: Our study suggests that UBE2CP3 can enhance the interaction between HCC tumor cells and HUVECs and promote HCC tumorigenicity by facilitating angiogenesis.

The long non-coding RNA PTTG3P promotes cell growth and metastasis via up-regulating PTTG1 and activating PI3K/AKT signaling in hepatocellular carcinoma.

BACKGROUND: Dysfunctions of long non-coding RNA (lncRNAs) have been associated with the initiation and progression of hepatocellular carcinoma (HCC), but the clinicopathologic significance and potential role of lncRNA PTTG3P (pituitary tumor-transforming 3, pseudogene) in HCC remains largely unknown. METHODS: We compared the expression profiles of lncRNAs in 3 HCC tumor tissues and adjacent non-tumor tissues by microarrays. In situ hybridization (ISH) and quantitative real-time polymerase chain reaction (qRT-PCR) were applied to assess the level of PTTG3P and prognostic values of PTTG3P were assayed in two HCC cohorts (n = 46 and 90). Artificial modulation of PTTG3P (down- and over-expression) was performed to explore the role of PTTG3P in tumor growth and metastasis in vitro and in vivo. Involvement of PTTG1 (pituitary tumor-transforming 1), PI3K/AKT signaling and its downstream signals were validated by qRT-PCR and western blot. RESULTS: We found that PTTG3P was frequently up-regulated in HCC and its level was positively correlated to tumor size, TNM stage and poor survival of patients with HCC. Enforced expression of PTTG3P significantly promoted cell proliferation, migration, and invasion in vitro, as well as tumorigenesis and metastasis in vivo. Conversely, PTTG3P knockdown had opposite effects. Mechanistically, over-expression of PTTG3P up-regulated PTTG1, activated PI3K/AKT signaling and its downstream signals including cell cycle progression, cell apoptosis and epithelial-mesenchymal transition (EMT)-associated genes. CONCLUSIONS: Our findings suggest that PTTG3P, a valuable marker of HCC prognosis, promotes tumor growth and metastasis via up-regulating PTTG1 and activating PI3K/AKT signaling in HCC and might represent a potential target for gene-based therapy.

Long non-coding RNA UBE2CP3 promotes tumor metastasis by inducing epithelial-mesenchymal transition in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a highly aggressive, solid malignancy that has a poor prognosis. Long non-coding RNAs (lncRNAs) have been found to be dysregulated in various cancers, including HCC. However, the molecular mechanism involving lncRNAs in HCC remains largely unknown. In this study, lncRNAs differentially expressed between HCC and corresponding non-cancerous tissue were identified by microarray analysis. A specific differentially expressed lncRNA UBE2CP3 (ubiquitin conjugating enzyme E2 C pseudogene 3) was identified. LncRNA UBE2CP3 was frequently up-regulated in HCC samples as assessed by quantitative real-time polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH) experiments. Clinical data showed that high levels of lncRNA UBE2CP3 were correlated with poor prognosis in HCC patients. Functional studies demonstrated that over-expression of lncRNA UBE2CP3 promoted cell invasion and migration in vitro and in vivo. Mechanistically, enhanced expression of lncRNA UBE2CP3 increased the expression of Snail1 and N-cadherin, but decreased the expression of E-cadherin, thus promoting the process of epithelial to mesenchymal transition (EMT) and finally inducing cell invasion and migration. Furthermore, serum levels of lncRNA UBE2CP3 were increased in HCC patients and decreased after surgery. Our results suggest that lncRNA UBE2CP3 promotes the metastasis of HCC and that serum lncRNA UBE2CP3 may be a new biomarker for the diagnosis of HCC.

Epithelial-mesenchymal transition phenotypes of circulating tumor cells correlate with the clinical stages and cancer metastasis in hepatocellular carcinoma patients.

BACKGROUND: Epithelial-mesenchymal transition (EMT) plays a crucial role in circulating tumor cells (CTCs) dissemination and cancer metastasis. OBJECTIVE: To investigate the EMT phenotypes of CTCs in hepatocellular carcinoma (HCC) patients and the clinical utility in the early diagnosis of HCC metastasis and progression. METHODS: We retrospectively analyzed the count and EMT classification of CTCs detected by the CanPatrol® platform in 195 HCC patients. The clinical relevance with other pathological features was statistically evaluated. RESULTS: CTCs were detected in 95% of the 195 HCC patients with a range of 0-86 CTCs. Total CTCs numbers were correlated with BCLC stages, metastasis and serum AFP levels. The AUC of the ROC curve was 0.861 (95% CI: 0.782-0.940) in discriminating metastatic HCC patients with non-metastatic patients. Epithelial, hybrid and mesenchymal CTCs were found in about 53%, 83% and 57% patients, respectively. The proportion of hybrid and mesenchymal CTCs was associated with ages, BCLC stages, metastasis and AFP levels. Besides, recurrent HCC patients presented higher CTCs count and increased hybrid and mesenchymal CTCs. CONCLUSIONS: CTCs count and EMT classification are correlated with clinical stages and metastasis of HCC, suggesting that they may be potential markers for the early diagnosis of HCC metastasis and progression.

miRNA-548p suppresses hepatitis B virus X protein associated hepatocellular carcinoma by downregulating oncoprotein hepatitis B x-interacting protein.

AIM: miR-548p is a recently identified and poorly characterized miRNA. However, its role of miR-548p in tumorigenesis and progression remains poorly understood. Here, we aimed to investigate the biofunction of miR-548p in hepatocellular carcinogenesis. METHODS: The expression levels of miR-548p were detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). The role of miR-548p in hepatocellular carcinoma (HCC) was determined by colony formation, flow cytometry assay and nude mice xenograft experiments. miR-548p target genes were analyzed by miRNA target predication programs and verified by qRT-PCR, western blotting assay and dual-luciferase reporter assay. RESULTS: miR-548p is repressed by hepatitis B virus X protein (HBx) in HCC tumor tissues and hepatoma cells, and inhibited cell growth by inhibiting cell proliferation and promoting cell apoptosis. miR-548p directly downregulated the expression of hepatitis B x-interacting protein (HBXIP) by binding to the 3'-untranslated region of HBXIP mRNA. Further study showed that hepatocyte nuclear factor-4a (HNF4A) promoted the expression of miR-548p and inhibited the transcription of HBXIP. HNF4A is a dominant transcriptional regulator of hepatocyte differentiation and hepatocellular carcinogenesis, and is shown to be repressed by HBx. CONCLUSION: We proposed the model for HBx/HNF4A/miR-548p/HBXIP pathway that controls hepatoma cell growth and tumorigenesis of HCC. miR-548p was identified as a tumor-suppressor in HBx-associated hepatocellular carcinogenesis.

HBx-related long non-coding RNA DBH-AS1 promotes cell proliferation and survival by activating MAPK signaling in hepatocellular carcinoma.

Accumulating evidence supports an important role for the hepatitis B virus x protein (HBx) in the pathogenesis of hepatitis B virus (HBV)-induced hepatocellular carcinoma (HCC), but the underlying mechanisms are not entirely clear. Here, we identified a novel long noncoding RNA (lncRNA) DBH-AS1 involved in the HBx-mediated hepatocarcinogenesis. The levels of DBH-AS1 were positively correlated with hepatitis B surface antigen (HBsAg) and tumor size in HCC tissues. Functionally, transgenic expression of DBH-AS1 significantly enhanced cell proliferation and tumorigenesis, whereas short hairpin RNA knockdown of DBH-AS1 caused an inhibition of cell proliferation. Mechanistically, overexpression of DBH-AS1 induced cell cycle progression by accelerating G1/S and G2/M transition concomitantly with upregulation of CDK6, CCND1, CCNE1 and downregulation of p16, p21 and p27. We also found that enhanced DBH-AS1 expression inhibited serum starvation-induced apoptosis of HCC cells. In contrast, suppressed DBH-AS1 expression had opposite effects. Furthermore, DBH-AS1 was shown to activate MAPK pathway. We also provide evidence that DBH-AS1 could be significantly induced by HBx protein and markedly down-regulated by p53. Thus, we concluded that DBH-AS1 can be induced by HBx and inactivated by p53, and consequently promote cell proliferation and cell survival through activation of MAPK signaling in HCC. Our study suggests that DBH-AS1 acts as an oncogene for HCC.