Long non-coding ribonucleic acid W5 inhibits progression and predicts favorable prognosis in hepatocellular carcinoma.

BACKGROUND: Accumulating evidence has revealed that several long non-coding ribonucleic acids (lncRNAs) are crucial in the progress of hepatocellular carcinoma (HCC). AIM: To classify a long non-coding RNA, i.e., lncRNA W5, and to determine the clinical significance and potential roles of lncRNA W5 in HCC. METHODS: The results showed that lncRNA W5 expression was significantly downregulated in HCC cell lines and tissues. Analysis of the association between lncRNA W5 expression levels and clinicopathological features suggested that low lncRNA W5 expression was related to large tumor size (P < 0.01), poor histological grade (P < 0.05) and serious portal vein tumor thrombosis (P < 0.05). Furthermore, Kaplan-Meier survival analysis showed that low expression of lncRNA W5 predicts poor overall survival (P = 0.016). RESULTS: Gain-of-loss function experiments, including cell counting kit8 assays, colony formation assays, and transwell assays, were performed in vitro to investigate the biological roles of lncRNA W5. In vitro experiments showed that ectopic overexpression of lncRNA W5 suppressed HCC cell proliferation, migration and invasion; conversely, silencing of lncRNA W5 promoted cell proliferation, migration and invasion. In addition, acting as a tumor suppressor gene in HCC, lncRNA W5 inhibited the growth of HCC xenograft tumors in vivo. CONCLUSION: These results showed that lncRNA W5 is down-regulated in HCC, and it may suppress HCC progression and predict poor clinical outcomes in patients with HCC. LncRNA W5 may serve as a potential HCC prognostic biomarker in addition to a therapeutic target.

Silymarin-mediated regulation of the cell cycle and DNA damage response exerts antitumor activity in human hepatocellular carcinoma.

A novel module-search algorithm method was used to screen for potential signatures and investigate the molecular mechanisms of inhibiting hepatocellular carcinoma (HCC) growth following treatment with silymarin (SM). The modules algorithm was used to identify the modules via three major steps: i) Seed gene selection; ii) module search by seed expansion and entropy minimization; and iii) module refinement. The statistical significance of modules was computed to select the differential modules (DMs), followed by the identification of core modules using the attract method. Pathway analysis for core modules was implemented to identify the biological functions associated with the disease. Subsequently, results were verified in an independent sample set using reverse transcription polymerase chain reaction (RT-PCR). In total, 18 seed genes and 12 DMs (modules 1-12) were identified. The core modules were isolated using gene expression data. Overall, there were 4 core modules (modules 11, 5, 6 and 12). Additionally, DNA topoisomerase 2-binding protein 1 (TOPBP1), non-structural maintenance of chromosomes condensing I complex subunit H, nucleolar and spindle associated protein 1 (NUSAP1) and cell division cycle associated 3 (CDCA3) were the initial seed genes of module 11, 5, 6 and 12, respectively. Pathway results revealed that cell cycle signaling pathway was enriched by all core modules simultaneously. RT-PCR results indicated that the level of CDCA3, TOPBP1 and NUSAP1 in SM-treated HCC samples was markedly decreased compared with that in non-SM-treated HCC. No statistically significant difference between the transcriptional levels of CDCA3 in SM-treated and non-treated HCC groups was identified, although CDCA3 expression was increased in the treated group compared with the untreated group. Furthermore, although the expression level of TOPBP1 and NUSAP1 in the SM-treated group was decreased compared with that in the normal group, no significant difference was observed. From the results of the present study it can be inferred that TOPBP1, NUSAP1 and CDCA3 of the core modules may serve notable functions in SM-associated growth suppression of HCC.

Clinical analysis of bevacizumab targeting therapy in treating early colorectal carcinoma after operation.

Clinical effects of bevacizumab target therapy in treating early colorectal carcinoma (CRC) after resection were analyzed. Ninety-two patients diagnosed with early CRC and treated with endoscopic mucosal resection for the first time were selected for the study. They were randomly divided into the control group and the observation group with 46 cases in each group. Control group was administered the chemotherapy regimen with oxaliplatin, calcium folinate and 5-fluorouracil, while bevacizumab targeting therapy was given to the observation group. The follow-up median time in these two groups was 30 months. In the observation group, objective response rate and disease control rate were higher than those in the control group, the adverse reaction rate was lower, and the differences were statistically significant (p<0.05). In the observation group, disease-free survival was prolonged (38.6 vs. 30.5 months, p<0.05); the recurrence rate was lower (13.0 vs. 30.4%, p<0.05); the survival rate was improved (91.3 vs. 76.1%, p<0.05). Vascular endothelial growth factor (VEGF) expressions of follow-up serum in these two groups were lower; VEGF expression in the observation group was lower than that in the control group, and the differences had statistical significance (p<0.05). There was no statistical significance in comparison of positive expression in tissue VEGF (p>0.05). In conclusion, after bevacizumab targeting therapy in treating early CRC, VEGF expression of serum was significantly lower; treatment effects improved; adverse drug reaction was reduced; survival time was prolonged; the recurrence rate was reduced; the survival rate improved. It has good application values.

Application and Exploration of Big Data Mining in Clinical Medicine.

OBJECTIVE: To review theories and technologies of big data mining and their application in clinical medicine. DATA SOURCES: Literatures published in English or Chinese regarding theories and technologies of big data mining and the concrete applications of data mining technology in clinical medicine were obtained from PubMed and Chinese Hospital Knowledge Database from 1975 to 2015. STUDY SELECTION: Original articles regarding big data mining theory/technology and big data mining's application in the medical field were selected. RESULTS: This review characterized the basic theories and technologies of big data mining including fuzzy theory, rough set theory, cloud theory, Dempster-Shafer theory, artificial neural network, genetic algorithm, inductive learning theory, Bayesian network, decision tree, pattern recognition, high-performance computing, and statistical analysis. The application of big data mining in clinical medicine was analyzed in the fields of disease risk assessment, clinical decision support, prediction of disease development, guidance of rational use of drugs, medical management, and evidence-based medicine. CONCLUSION: Big data mining has the potential to play an important role in clinical medicine.

Effect of immune modulation therapy on cardiac function and T-bet/GATA-3 gene expression in aging male patients with chronic cardiac insufficiency.

AIM: The aim of this study was to explore the role of immune modulation therapy in regulating the imbalance of Th1/Th2, serum IFN-γ, IL-4 and the T-cell-specific transcription factors T-bet/GATA-3 in peripheral blood in aging male patients with chronic cardiac insufficiency (CCI). PATIENTS & METHODS: In total, 156 participants were divided into three groups: the CCI intervention group, which received regular therapy and thymopetidum (20 mg intramuscular injection, once every other day for 3 months; n = 70), the CCI control group, which received regular therapy (n = 56) and 50 healthy individuals older than 57 years of age, who served as normal controls. RESULTS: Before therapy, in comparison with the control group, levels of left ventricular end diastolic diameter, NT-proBNP, C-reactive protein (CRP), Th1, Th1/Th2, IFN-γ, and T-bet mRNA and T-bet/GATA-3 mRNA all increased, and the level of left ventricular ejection fraction (LVEF), 6MWT, Th2, IL-4, and GATA-3 mRNA also decreased in both the CCI intervention and control groups. Linear correlation analysis indicated that LVEF was inversely correlated with serum NT-proBNP, CRP, Th1/Th2, IFN-γ and T-bet mRNA/GATA-3 mRNA, and was positively correlated with plasma IL-4. After 3 months of therapy, levels of left ventricular end diastolic diameter, NT-proBNP, CRP, Th1, Th1/Th2, IFN-γ, T-bet mRNA and T-bet/GATA-3 mRNA decreased in the two CCI subgroups, but levels in the CCI intervention group were lower in comparison to the control group. Levels of LVEF, 6MWT, Th2 and GATA-3 mRNA increased in the two CCI subgroups, while levels in the CCI intervention group were higher in comparison with the control group. Plasma levels of IL-4 showed no change after treatment. CONCLUSION: Immune modulation improved cardiac function of CCI patients and was associated with amelioration of T-helper superficial transcription factor polarization and its related cytokine imbalance. Immune modulation might be a new treatment strategy for aging CCI patients.

[Establishment of human cardiac C protein induced experimental autoimmune myocarditis model in rat].

OBJECTIVE: To construct the recombinant plasmid of human cardiac C protein (CCP) peptide with immunogenicity and to express, purification and renature fusion protein. The fusion protein was injected to Lewis rats to establish experimental autoimmune myocarditis (EAM) model. METHODS: Total RNA was extracted from human heart and used as the template for reverse transcriptase-directed cDNA synthesis. The cDNA was then amplified by polymerase chain reaction (PCR) using oligonucleotide primers specific for CCP peptide with immunogenicity. Subsequently, the purified CCP peptide gene was cloned into PEASY-T1 vector and the ligated product was identified by PCR and DNA sequence analysis. Then the CCP target gene of positive clone was inserted into the pQE30, a prokaryotic expression vector, and the inserting plasmid was transformed into Escherichia coli. host M15. The positive clone extracted from the bacterium liquid was sieved by insertional inactivation sieve method and identified by PCR of bacterium liquid, CCP immunological peptide was purified and renatured in semipermeable membrane. EAM model in Lewis rats was induced by injection of mixture of 100 µg CCP fusion protein immunological peptide and 2.5 g/L completed Freund adjuvant from two double foot pad and subsequent abdominal injection of 0.5 µg pertussis toxin. Two, four, six, and eight weeks after immunization, hemodynamic evaluation was made and hearts underwent histological examination. RESULTS: The DNA sequence analysis for cloning vector extraction revealed that the CCP target gene was cloned into pQE30 exactly. The DNA of 1000 bp length was obtained by PCR examination of bacterium liquid with transformation of express recombinants which were consistent with the expected size. Purified fusion protein in vertical slab gel electrophoresis showed 35 000 as expected. The recombinant CCP fusion protein existed in inclusion bodies of E. coli and amounted to 80% - 90% of the total protein. Hemodynamic and histological evaluations showed typical acute inflammatory responses at 2 weeks, subacute inflammatory and fibrosis changes at 4 weeks after injection, and signs of chronic dilated cardiomyopathy at 6 weeks post injection. CONCLUSION: Combination of gene clone technique and histidine tag protein purification technique can be used to synthesize human cardiac C protein to induce EAM model in Lewis rat.

[Effects of matrix metalloproteinase-9 inhibitor in Lewis rats with experimental autoimmune myocarditis].

OBJECTIVE: To investigate the effects of matrix metalloproteinase-9 (MMP-9) inhibitor minocyclin hydrochloride in Lewis rats with experimental autoimmune myocarditis (EAM). METHODS: EAM was induced by injection of cardiac C protein emulsified in completed Freund adjuvant in double footpad and intraperitoneal injection of pertussis toxin on 6- to 8-week old Lewis rats. Sixty EAM Lewis rats were divided into 3 groups (early, middle and late intervention groups, n = 20 each: 10 minocyclin treated and 10 control rats). In early intervention group, rats in treatment group received intraperitoneal injection of minocyclin hydrochloride from 1(st) to 21(st) day after immunization; in middle intervention group, rats were treated from 8(th) to 28(th) day after immunization and in late intervention group, rats were treated from 15(th) to 35(th) day after immunization (50 mg/kg body weight, once daily). Control rats received intraperitoneal injection of same volumetric physiological saline at corresponding time periods. At the end of intervention, rats were euthanatized and hearts were harvested. Paraffin sections were used for hematoxylin and eosin stain to determine the inflammatory score, for picrosirius stain to determine fibrosis score and collagen content, and for immunohistological stain to determine macrophages and T lymphocytes. Real time PCR was used to detect mRNA expression of myocardial MMP-2 and MMP-9. Cryostat sections were used for in situ zymography to detect protein activity of gelatinase. RESULTS: Inflammatory score in cardiac paraffin slides, number of cardiac macrophages and T lymphocytes, cardiac interstitial fibrosis score and content, expression of MMP-2, 9 mRNA and activity of gelatinase in treatment group were all significantly lower than in control group for early and middle intervention groups (inflammatory score: early control group vs. treatment group: 3.03 ± 1.35 vs.1.51 ± 0.36, P < 0.05, middle control group vs. treatment group: 3.75 ± 0.29 vs. 2.11 ± 0.82, P < 0.01; cardiac interstitial fibrosis score, early control group vs. treatment group: 2.75 ± 0.29 vs.1.51 ± 0.35, P < 0.01, middle control group vs. treatment group: 2.50 ± 0.41 vs. 1.61 ± 0.42, P < 0.05; gelatinase, early control group vs. treatment group: 162 367 ± 5095 vs. 62 366 ± 2131, P < 0.01, middle control group vs. treatment group: 184 256 ± 5427 vs. 113 197 ± 4809, P < 0.01) while these parameters were similar between minocyclin-treated and control rats in late intervention group (all P > 0.05). CONCLUSIONS: MMP-9 plays an important role in the pathogenesis of autoimmune myocarditis. Inhibition of MMP-9 in early and middle stage could significantly attenuate inflammatory responses and myocardial fibrosis in this experimental EAM model.

[Experimental study of MMP-2 inhibitor treatment of experimental autoimmune myocarditis in Lewis rats].

OBJECTIVE: To investigate the inhibitor of matrix metalloproteinase-2 (MMP-2) (2R)-2-[5-[4-[ ethyl-methylamino] phenyl [thiophene-2-sulfonylamino]-3-methylbutyric acid (TISAM) therapeutic effect on experimental autoimmune myocarditis (EAM) in Lewis rats. METHODS: Treatment protocol of oral administration of 5 mg/kg TISAM once a day for 14 days was performed on EAM Lewis rats. EAM Lewis rats were divided into 3 groups: treatment in early, middle and later stage respectively (n = 20). After experiment at the designate time point, the rats were euthanatized and hearts were harvested. Cardiac inflammatory score, fibrosis score and content, and infiltration of macrophages and T lyminflammatory score, fibrosis score and content, and infiltration of macrophages and T lymphocytes, message RNA (mRNA) expression of matrix metalloproteinase (MMP)-2 and MMP-9 and protein activity of gelatinase were determined. RESULTS: TISAM treatment in early phase was invalid (treatment started from the creation of the model), treatment in middle and later phase was effective (treatment started from 7 and 14 day after the creation of the model). CONCLUSION: Inhibitor of MMP-2 can block ventricular remodeling in middle stage in EAM Lewis rats. The mechanism maybe alleviate the inflammatory cell cardiac infiltration, decrease the mRNA expression of MMP-2 at transcript level and downregulate gelatinase activity at protein level.

[Immune state of Th1, Th2 and Th17 subpopulation in experimental autoimmune myocarditis].

OBJECTIVE: To shed light on changes in the gene expression of T helper lymphocyte (Th) subpopulation, Th1, Th2 and Th17 in autoimmune myocarditis and to gain insight into the immunological mechanisms underlying the essence of myocarditis. METHODS: An experimental Lewis rat autoimmune myocarditis model was induced by immunization with cardiac C protein and completed Freund adjuvant in double foot pads and subsequent intraperitoneal injection of pertussis toxin. Two groups of normal rats without immunological injection acted as control group. Transthoracic echocardiography was performed and subsequently hearts and spleens were obtained from EAM rats at 1 week, 2 weeks, 4 weeks, 6 weeks and 8 weeks after immunization. The pathological sections of heart samples were prepared, the inflammatory score was determined by hematoxylin and eosin stain, the fibrosis score was determined by picrosirius red stain. The ratio of Th1, Th2 and Th17 subpopulation in spleen cells were measured by flow cytometry, and enzyme linked immunoabsorption assay (ELISA) was used to determine the serum level of Th1 related cytokine interferon-gamma (IFN-gamma), Th2 related cytokine interleukin (IL)-4 and Th17 related cytokine IL-17. RESULTS: In EAM rats, cardiac ejection fraction remained normal until 4th week, and left ventricular end systolic diameter and left ventricular end diastolic diameter decreased. However, cardiac ejection fraction decreased obviously and left ventricular end systolic diameter and left ventricular end diastolic diameter rose until 8th week. Inflammatory score increased rapidly at 2nd week after immunization and remained peek level until 4th week and then gradually decreased at 6th and 8th week. Fibrosis score and fibrosis content increased from 4th week and maintained the peek level until 8th week. Ratio of Th1 in spleen cells of EAM rats and its related cytokine in serum, IFN-gamma, increased at 1st week, arrived at the peek level until 4th week and gradually decreased at 6th week. Ratio of Th17 and IL-17 rose from 2nd-4th week and remained until 6th to 8th week. Ratio of Th2 showed no change in the previous four weeks, ratio of IL-4 increased from 4th week, and both rose at 6th week rapidly and remained until 8th week. CONCLUSIONS: In EAM Lewis rats, The time duration from 2nd to 4th week was inflammatory stage of myocarditis while during the period of 4th week to 8th week myocarditis develops into fibrotic stage. Imbalance of Th1/Th2 takes part in the occurrence of ventricular remodeling, cellular immunity mediated by Th1 and Th17 being preponderant at inflammatory myocarditis stage while humoral immunity mediated by Th2 and Th17 being preponderant at fibrotic carditis stage.