Gene expression and methylation profiles show the involvement of POMC in primary hyperparathyroidsm.

Primary hyperparathyroidism (PHPT) is mainly caused by parathyroid adenoma, which produces excess parathyroid hormones. Its pathogenic mechanisms have not yet been fully understood. To investigate the mechanism in the pathogenesis of PHPT, the transcriptome and genome-wide DNA methylation profiles of parathyroid adenoma were analyzed. The candidate genes that may be involved in the PHPT were verified via qRT-PCR, immunohistochemistry, western blot, and methylation-specific PCR. A total of 1650 differentially expressed genes and 2373 differentially methylated regions were identified. After the integration of its transcriptome and DNA methylation data, IL6, SYP, GNA01, and pro-opiomelanocortin (POMC) were the candidate genes that demonstrated a similar pattern between their mRNA expression and DNA methylation status. Of the 4 candidate genes, POMC, a pro-peptide which is processed to a range of bioactive peptide products like ACTH, was further confirmed to be expressed at low levels at both the mRNA and protein levels, which may be due to POMC promoter hypermethylation. Hypermethylation of the POMC promoter may contribute to its low expression, which may be involved in the pathogenesis of PHPT.

Knockdown of lncRNA ANRIL suppresses the production of inflammatory cytokines and mucin 5AC in nasal epithelial cells via the miR-15a-5p/JAK2 axis.

The incidence of allergic rhinitis (AR) is increasing worldwide. Human nasal epithelial cells (HNECs) are the key cells in the occurrence of AR. Antisense non-coding RNA in the INK4 locus (ANRIL) was discovered to be involved in the progression of AR. However, the mechanism by which ANRIL mediates the progression of AR remains to be determined. The present study aimed to further explore the mechanism by which ANRIL regulates AR. Thereby, HNECs were treated with IL-13 to mimic AR in vitro. The mRNA expression levels of ANRIL, microRNA (miR)-15a-5p, JAK2, mucin 5AC (MUC5AC), granulocyte-macrophage colony-stimulating factor (GM-CSF) and eotaxin-1, and protein expression levels of JAK2, STAT3 and phosphorylated-STAT3 in HNECs were analyzed using reverse transcription-quantitative PCR and western blotting, respectively. ELISAs were used to detect the secretory levels of inflammatory cytokines and mucin in cell supernatants. In addition, a dual luciferase reporter assay was used to confirm the downstream target of ANRIL and the target gene of miR-15a-5p. The results revealed that the secretory levels of eotaxin-1, GM-CSF and MUC5AC were significantly upregulated by IL-13 in the supernatant of HNECs. The expression levels of ANRIL and JAK2 were also upregulated in IL-13-induced HNECs, while the expression levels of miR-15a-5p were downregulated. In addition, ANRIL was identified to bind to miR-15a-5p. The IL-13-induced upregulation of eotaxin-1, GM-CSF and MUC5AC mRNA expression and secretory levels was significantly inhibited by the genetic knockdown of ANRIL, while the miR-15a-5p inhibitor effectively reversed this effect. JAK2 was also discovered to be directly targeted by miR-15a-5p. The overexpression of JAK2 significantly suppressed the therapeutic effect of miR-15a-5p mimics on IL-13-induced inflammation in vitro. In conclusion, the findings of the present study suggested that the genetic knockdown of ANRIL may suppress the production of inflammatory cytokines and mucin in IL-13-treated HNECs via regulation of the miR-15a-5p/JAK2 axis. Thus, ANRIL may serve as a novel target for AR treatment.

Fibroblasts play a potential role in bone destruction via osteopontin related caldesmon expression and polymerization in human non-functioning pituitary adenomas.

Non-functioning pituitary adenomas (NFPAs) are the most frequent pituitary tumors. The elucidation of the mechanisms of aggressive NFPAs in bone destruction is required in order to guide the clinical diagnosis and treatment of NFPAs. In the present study, we investigated the differential proteomics of fibroblasts isolated from clinical specimens of NFPAs with or without bone destruction. Proteomic analysis revealed a group of molecules associated with cytoskeleton organization, including caldesmon, were differentially expressed between fibroblasts isolated from bone destruction NFPAs (BD-NFPAs) and fibroblasts isolated from non-bone destruction NFPAs (NBD-NFPAs). The secreted proteins analysis found that osteopontin was significantly upregulated in BD-NFPAs fibroblasts. Furthermore, immunohistochemical staining of the NFPAs clinical samples showed that the expression of caldesmon in stromal cells and the expression of osteopontin in both tumor cells and stroma were significantly increased in BD-NFPAs. Taken together, our results indicate a possible way that osteopontin secreted from both NFPA cells and surrounding fibroblasts modify caldesmon expression and polymerization in fibroblasts, which may contribute to bone destruction in NFPA patients.

Large-scale synthesis of tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate by a stereoselective carbonyl reductase with high substrate concentration and product yield.

To biosynthesize the (3R,5S)-CDHH in an industrial scale, a newly synthesized stereoselective short chain carbonyl reductase (SCR) was successfully cloned and expressed in Escherichia coli. The fermentation of recombinant E. coli harboring SCR was carried out in 500 L and 5000 L fermenters, with biomass and specific activity of 9.7 g DCW/L, 15749.95 U/g DCW, and 10.97 g DCW/L, 19210.12 U/g DCW, respectively. The recombinant SCR was successfully applied for efficient production of (3R,5S)-CDHH. The scale-up synthesis of (3R,5S)-CDHH was performed in 5000 L bioreactor with 400 g/L of (S)-CHOH at 30°C, resulting in a space-time yield of 13.7 mM/h/g DCW, which was the highest ever reported. After isolation and purification, the yield and d.e. of (3R,5S)-CDHH reached 97.5% and 99.5%, respectively. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:612-620, 2017.

NK/T cell lymphoma involving mediastinum: report of a case and review of literature.

Extranodal natural killer (NK)/T-cell lymphoma is a very aggressive malignant neoplasia with a poor prognosis. Herein we reported a case of NK/T cell lymphoma involving mediastinum. It was a 28-year-old Chinese male patient. The tumor cells were medium-sized, had irregularly folded nuclei, and inconspicuous or small nucleoli with coagulative necrosis. The tumor cells were positive for CD3ε, TIA-1, but negative for CD56. In situ hybridization revealed that tumor cells also expressed Epstein-Barr virus encoded RNA. To our knowledge, this is the first case of NK/T cell lymphoma involving mediastinum.

Discovery of N-(3,5-bis(1-pyrrolidylmethyl)-4-hydroxybenzyl)-4-methoxybenzenesulfamide (sulcardine) as a novel anti-arrhythmic agent.

AIM: To investigate the anti-arrhythmic effects of sulfamide analogues of changrolin and to characterize the sulfate of compound 6f (sulcardine sulfate, Sul) as a novel anti-arrhythmic agent. METHODS: The anti-arrhythmic effects of compounds were studied against aconitine-induced arrhythmias in rats and ouabain-induced arrhythmias in guinea pigs. The effects of Sul on transmembrane action potentials were investigated in isolated rabbit sinoatrial nodes and guinea-pig papillary muscles using intracellular recording. With a whole-cell recording technique, the effects of Sul on sodium current, calcium current, and potassium currents were examined in isolated single guinea-pig ventricular myocytes. RESULTS: In aconitine-induced arrhythmias of rats, sulfamide analogues of changrolin (4, 5, and 6a-6p) exhibited various anti-arrhythmic activities. The sulfate of compound 6f (Sul) increased the amount of aconitine required to induce arrhythmias in each treated animal. The ED50 value of Sul in rats was 196 mg/kg. In ouabain-induced arrhythmias of guinea pigs, 25, 50, and 100 mg/kg doses of Sul increased the dose of ouabain required to induce VP, VT, and VF in a dose-dependent manner. In papillary preparations, Sul produced a concentration-dependent decrease in APA and V(max), prolonged APD(90) and ERP, whereas RP was unaffected. In the spontaneously beating sinus nodes, Sul reduced APA and V(max) in a concentration-dependent manner. The whole-cell recording studies revealed that Sul produced a reversible reduction in I(Na) (IC50=26.9 µmol/L) and I(Ca,L)(IC50=69.2 µmol/L), whereas the inward rectifier (I(K1)) and the delayed rectifier potassium currents (I(K)) were unaffected. CONCLUSION: As a multi-ion channel blocker, Sul may have potent efficacy in anti-atrial and ventricular arrhythmias.

[Fine root biomass and its nutrient storage in karst ecosystems under different vegetations in Central Guizhou, China].

The degeneration of karst ecosystem is closely associated with the reduction of soil nutrients and fine root biomass, and the retention of soil nutrients is of significance in sustaining ecosystem functioning. To understand the changes in the fine root biomass and soil nutrient retention in degenerated karst ecosystems, a comparative study was conducted with three typical vegetation stands (forest, shrub, and shrub-grass) in Central Guizhou of Southwest China. Soil samples with fine roots were collected from the depths 0-5 cm, 5-10 cm, and 10-15 cm, with the related indices of fine roots and nutrients measured. In the three stands, fine roots dominantly distributed in 0 -10 cm soil layer, and decreased sharply with soil depth. The living fine root biomass in 0-10 cm soil layer under forest, shrub, and shrub-grass occupied 83.36%, 86.91%, and 93.79% of the total fine root biomass, and 42.78%, 56.75%, and 53.38% of the total living fine root biomass within the 0-15 cm soil layer, respectively. The fine root biomass of predominant plant species varied with vegetation types. The N and P storage of the living fine roots in 0-5 cm soil layer under forest stand was significantly higher than those under shrub stand and shrub-grass stand, and no significant differences were observed between the latter two stands. However, the N and P storage of the living fine roots in 5-10 cm soil layer under different stands decreased in the order of forest > shrub > shrub-grass (P < 0.05). There was a significant positive correlation between the plant aboveground biomass and the living fine root biomass in 0-10 cm soil layer, and a significant negative correlation between the N and P contents of plant leaves and the specific length of living fine roots, illustrating that the nutrient uptake and retention by the living fine roots could have particular importance in the aboveground biomass establishment and ecosystem functioning.

A case of mesothelial/monocytic incidental cardiac excrescence and literature review.

Mesothelial/monocytic incidental cardiac excrescence (MICE) is a rare entity which is an amalgam of mesothelial cells, histiocytes, and fibrin, often found occasionally during cardiac valve replacement. We report a case in a 25-year-old Chinese female with serous mitral stenosis and patent foramen ovale. Routine and immunohistochemical stains and ultrastructure examination revealed the vegetation was predominantly composed of histocytes with scattered mesothelial cells. In fact nodular histiocytic/mesothelial hyperplasia (NHMH) is a similar lesion to MICE. MICE and NHMH could be unified, and NHMH may be a better choice.

Synthesis and characteristics of graphene oxide-derived carbon nanosheet-Pd nanosized particle composites.

Carbon nanosheet (CNS)-Pd nanosized particle (NP) composites were synthesized by using graphite oxide (GO) and bis(ethylenediamine)palladium(II) (Pd(en)(2)(2+)) as the precursors, and their structure and adsorption properties were examined. It was found that the Pd(en)(2)(2+) complex ions can be intercalated into GO layers highly efficiently to form a layered structure containing a large amount of Pd (approximately 12 wt %). By the subsequent chemical reduction, Pd NPs (2-6 nm in size) are well dispersed between CNS to form a CNS-Pd NP composite and serve as spacers to increase the porosity of the composite. Hydrogen adsorption results demonstrate that both Pd NPs and CNS play important roles in hydrogen adsorption, particularly at a lower temperature and for CNS with deficient sites, which bring about a H(2) adsorption greater than those on other Pd-loaded nanocarbon materials reported so far. The unique composite nanostructure having large contents of Pd NPs (20-25 wt %) stabilized by CNSs is hopeful to be applied to the fields of H(2)-related catalysis, sensing, and so forth.

Knockdown of fascin1 expression suppresses the proliferation and metastasis of gastric cancer cells.

AIMS: Our current investigation attempts to study the role of the fascin1 gene in growth and metastasis of gastric cancer cell line MKN45. METHODS: Small interfering RNA (siRNA) was used to inhibit fascin1 expression in the human gastric cancer cell line MKN45. Expression of fascin1 in fascin1 siRNA transfected cells (sifascin1), non-transfected cells (NT) and non-specific fascin1 siRNA cells (CON) were examined by Western blotting and reverse transcription polymerase chain reaction (RT-PCR). Cell growth ability in vitro was evaluated by MTT and clone formation assays. Cell mobility in vitro was examined by the Boyden chamber assay. Nude mice metastasis models were established by abdominal cavity transfer method. Tumour growth was evaluated by immunohistochemistry with proliferating cell nuclear antigen (PCNA). RESULTS: Knockdown of fascin1 expression in MKN45 cells resulted in decreased cellular proliferative and migratory abilities. In vitro, the cloning efficiency of siFascin1 cells (34.2%) was significantly lower compared to that in NT (78.5%) (p < 0.05). The migration rate in siFascin1 cells was significantly decreased (33.7%) compared with NT cells (89.4%) (p < 0.05). In vivo, the cell proliferation rate was lower in siFascin1 cells (25.8%) compared to that in NT (75.0%) (p < 0.05). The number of tumour clones in the liver was significantly lower in siFascin1 cells (2.0 +/- 1.1) compared to that in NT (5.1 +/- 1.6) (p < 0.05). CONCLUSIONS: Our study demonstrates that down-regulation of fascin1 suppresses the proliferation and migration of gastric cancer cells MKN45, suggesting that fascin1 siRNA may offer a novel potential gene therapy approach for human gastric cancer with fascin1 over-expression.

[Relationship between p53 gene and chromosome 13q14 variations and prognosis in primary intestinal lymphoma].

OBJECTIVE: Some research has shown that primary intestinal lymphoma with the same immunophenotype has different prognosis. It suggests that the prognosis of this disease is not determined by a single factor but may be related to genetic or chromosomal variations. The p53 gene is an important tumor suppressor gene, and 13q14 deletion is a common chromosomal abnormality of lymphocyte proliferation diseases. This study aimed to explore the role of the p53 gene and chromosome 13q14 variations in the assessment of prognosis in primary intestinal lymphoma. METHODS: p53 gene and chromosome 13q14 expression in paraffin sections of 30 cases of primary intestinal lymphoma and 10 cases of lymph node reactive hyperplasia were ascertained using an improved FISH technique. RESULTS: p53 gene deletion was found in 22.7% of patients with primary intestinal lymphoma at stage I-II and in 75.0% of patients at stage III-IV (x2=6.903, p<0.01). The 30 patients with primary intestinal lymphoma were pathologically classified into-mucosa-associated lymphoid tissue (MALT) (n=14) and non-MALT types (n=16). The MALT lymphoma group had significantly lower incidence of p53 gene deletion (14.3% vs 56.3%; x2=5.662, p<0.05). Average survival time in patients with p53 gene deletion was 13.41 months, being shorter than the patients with normal p53 gene (36.1 months) (t=2.637, p<0.05). 13q14 deletion was found in 40.0% of patients with primary intestinal lymphoma, but none of patients with lymph node reactive hyperplasia showed 13q14 deletion. 13q14 deletion was not significantly related to the pathological type and the clinical stage of primary intestinal lymphoma as well as the survival time. There was no significant correlation between p53 gene and 13q14 deletions. CONCLUSIONS: There was a high incidence of p53 gene deletion in patients with non-MALT lymphoma or at stage III-IV. p53 gene deletion is related to a high tumor malignant degree and a poor prognosis, while-chromosome 13q14 variation is not associated with the prognosis in patients with primary intestinal lymphoma.

[Biological implication of PTEN gene expression in human gastric cancer and related molecular mechanisms].

OBJECTIVE: To study the role of PTEN gene involved in the biological behavior of human gastric carcinoma cells and underlying molecular mechanisms. METHODS: Gastric carcinoma cell line, SGC7901, was transfected with plasmid PBP-PTEN and stable high PTEN expression clones were selected by Western blot and cell immunohistochemistry screening. Cell proliferation rate and apoptosis index of transfected cells were investigated by growth curve analysis, colony-formation assay and flow cytometry (FCM). Expressions of vascular endothelial growth factor (VEGF), matrix metalloprotease-2 (MMP-2) and MMP-9 proteins in cell culture supernatant and cytoplasm were determined by ELISA, gelatin zymogram, Western blot and cell immunohistochemistry. RESULTS: Stable clone with high level expression of PTEN was successfully established (PTEN-SGC7901). Cell doubling time of PTEN-SGC7901 was longer than that of the control cells (P < 0.05). The size and colony-forming efficiency of PTEN-SGC7901 cells deceased compared with those of the control. The relative colony-inhibition efficiency of PTEN-SGC7901 to SGC7901 (naïve untransfected) and PBP-SGC7901 (control vector transfected) cells were 69.8% and 64.8%, respectively. PTEN-SGC7901 clone had more cells at G1 phase (P < 0.05) compared with that of the control. However, the apoptosis index did not show significant differences among the three groups (P > 0.05). There were significantly less VEGF and MMP-9 protein expressions in the PTEN-SGC7901 culture supernatant and cytoplasm (P < 0.05). In contrast, the MMP-2 expression among three cell groups had no significant difference (P > 0.05). CONCLUSIONS: PTEN expression suppresses the growth and proliferation of gastric carcinoma cell SGC7901, possibly through an inhibition of the expressions of VEGF and MMP-9.

Effect of DPC4 gene on invasion and metastasis of colorectal carcinoma cells.

To investigate the effect of DPC4 gene on invasion and metastasis of colorectal carcinoma cells, the expression of DPC4 was detected in sixty-three samples of colorectal tumors and seven cases of colorectal mucosa. The biological behavior of tumors expressing DPC4 was evaluated (including tumor staging, differentiation degree and metastasis). pcDNA3.1-DPC4 plasmid was constructed and transferred into HCT116 cells not expressing DPC4. The cell models (DPC4(+)-HCT116) steadily expressing DPC4 were obtained. Compared with HCT116 and pcDNA3.1-HCT116 cells, the doubling time of DPC4(+)-HCT116 cells was lengthened obviously (P<0.01), the apoptosis rate of DPC4(+)-HCT116 cells was significantly increased (PP<0.01), the cloning efficiency, cell adherency, migration and invasion ability of DPC4+-HCT116 cells were dropped obviously (P<0.01). The number of cancer nodules was decreased significantly in abdominal cavity and liver of the nude mice inoculated with DPC4(+)-HCT116 cells. The activity of MMP-9 and MMP-2 was detected by gelatin zymography. In comparison with HCT116 and pcDNA3.1-HCT116 cells, the activity of MMP-9 was decreased in DPC4(+)-HCT116 cells. Therefore, the down-regulation of DPC4 expression may be associated with the carcinogenesis of colorectal carcinoma. DPC4 may inhibit the proliferation of colon cancer cell by restraining growth and inducing apoptosis, and the invasion and metastasis of colorectal carcinoma cells. MMP-9 may be one of the downstream target genes regulated by DPC4.

Enhancement of metastatic and invasive capacity of gastric cancer cells by transforming growth factor-beta1.

Transforming growth factor-beta (TGF-beta), a multifunctional cytokine, exerts contradictory roles in different kinds of cells. A number of studies have revealed its involvement in the progression of many types of tumors. To investigate the effect of TGF-beta on gastric carcinoma, SGC7901, BGC823 and MKN28 (a TGF-beta-resistant cell line) adenocarcinoma clones were used. After pretreatment in serum-free medium with or without 10 ng/ml TGF-beta1, their experimental metastatic potential, chemotaxis, and invasive and adhesive ability were measured. Furthermore, zymography for gelatinase was processed. Liver colonies were also measured 4 weeks after inoculation of SGC7901, BGC823 and MKN28 in Balb/c nude mice, and an increase in the number of surface liver metastases was seen in SGC7901 (from 11.0+/-3.0 to 53.3+/-3.3) and BGC823 (from 9.3+/-2.5 to 60.0+/-2.8) groups, whereas there was no difference between MKN28 groups (from 35.2+/-3.8 to 38.5+/-2.7). In vitro experiments showed that TGF-beta1 increased the adhesion capacity of SGC7901 and BGC823 cells to immobilized reconstituted basement membrane/fibronectin matrices and promoted their penetration through reconstituted basement membrane barriers. Zymography demonstrated that enhanced invasive potential was partly due to the increased type IV collagenolytic (gelatinolytic) activity, but there was no difference in type IV collagenolytic activity and other biological behaviors between MKN28 groups. These results suggested that TGF-beta1 might modulate the metastatic potential of gastric cancer cells by promoting their ability to break down and penetrate basement membrane barriers and their adhesive and motile activities. We speculated that TGF-beta1 might act as a progression-enhancing factor in gastric cancer. Therefore blockage of TGF-beta or TGF-beta signaling might prevent gastric cancer cells from invading and metastasizing.

[Expressions of TGIF, MMP9 and VEGF proteins and their clinicopathological relationship in gastric cancer].

OBJECTIVE: To detect the expressions of TG-interacting factor (TGIF), matrix metalloproteinase 9 (MMP9) and vascular endothelial growth factor (VEGF) proteins, and to analyze their clinicopathological relationship in gastric cancer. METHODS: Vascular invasion and remote metastasis were examined in nude mice inoculated with SGC-7901 cells, vector control cells or TGIF transfected cells via serial sections, and sequentially the expressions of MMP2, MMP9 and VEGF proteins in tumor tissues from nude mice were detected by immunohistochemical staining. At the same time, TGIF, MMP9 and VEGF proteins were examined in 76 patients with gastric carcinoma, and the relationships between the expressions of the three proteins and clinicopathological features were analyzed. RESULTS: Metastasis was not observed in nude mice inoculated with any cells. There were cancerous embolisms in nude mice tumor tissue inoculated with SGC-7901 cells and vector control cells, but not in TGIF transfected cells. The expressions of MMP9 and VEGF proteins were higher in the tumor tissues originated from SGC-7901 cells and vector control cells than that from TGIF transfected cells. The expression of MMP2 had no distinct difference among the tumor tissues originated from the three cells. The positive rate of TGIF protein in gastric carcinoma was 46.1% (35/76), obviously lower than that in surgical marginal mucosa (78.1%) (P < 0.05). Low expression of TGIF protein correlated significantly with the metastasis of lymph node. The positive rates of MMP9 and VEGF proteins in gastric carcinoma were 59.2% and 56.6%, respectively, obviously higher than that in surgical marginal mucosa (31.3%) (P < 0.05). High expressions of both MMP9 and VEGF also correlated significantly with the metastasis of lymph node. The expression level of VEGF protein was also associated with the invasion of gastric carcinoma (P < 0.05). The expression of TGIF protein reversely correlated with that of MMP9 and VEGF proteins (P < 0.05). CONCLUSION: TGIF may inhibit the invasion and metastasis of gastric carcinoma via the downregulation of MMP9 and VEGF proteins.

[The effects of hepatitis C virus core protein on biological behaviors of human hepatocytes].

OBJECTIVE: To investigate the effects of hepatitis C virus (HCV) core protein on the biological behaviors of human hepatocytes and their underlying mechanism. METHODS: A cell line expressing stably HCV core protein-QSG7701/core was constructed by transfecting the plasmid pcDNA3.1-core (expressing HCV core protein) into the human immortalized hepatocytes of the line QSG7701. The biological behaviors of these transfected cells were observed through plating-efficiency test, growth curve and flow cytometry (FCM). The association between HCV core protein and the expression of activated caspase-3 protein was evaluated by immunocytochemistry. The phosphorylation of mitogen-activate protein kinases (MAPKs) was detected with Western blotting. The activation of nuclear transcriptors AP-1, important effector molecule of MAPKs, and nuclear factor-kappa binding (NF-kappaB) were evaluated with luciferase assays and electrophoretic mobility shift assay (EMSA). RESULTS: HCV core protein was expressed in the QSG7701/core cells and not in the QSG7701/pcDNA3.1 cells and untransfected QSG7701 cells. There were no significant differences in the expression levels of total P44/42(MAPK), p38(MAPK) and JNK among the QSG7701/core cells, QSG7701/pcDNA3.1 cells and untransfected QSG7701 cells. The expression levels of phophorylated P44/42(MAPK), p38(MAPK) and JNK in the QSG7701/core cells were significantly weaker than those in the QSG7701/pcDNA3.1 cells and untransfected QSG7701 cells. Plating efficiency test showed that the clone formation rate of the QSG7701/core cells was 32.25%, significantly lower than those of QSG7701/pcDNA3.1 and untransfected QSG7701 cells (47.5% and 42.5% respectively, both P < 0.01). The growth curve showed that the multiplication time of the QSG7701/core cells was 36 hours, significantly longer than those of the QSG7701/pcDNA3.1 and untransfected QSG7701 cells (27 and 28 hours respectively). FCM showed that the apoptotic rate of the QSG770/1core was 1.04%, lower than those of the QSG7701/pcDNA3.1 and untransfected QSG7701 cells (1.68% and 3.7% respectively), and that the percentage of theQSG770/1core cells at the G(0)/G(1) stage increased and those in the S stage decreased. Immunocytochemistry showed that the expression intensity of caspase-3 in the QSG7701/core cells was significantly weaker than those of the QSG7701/pcDNA3.1 cells and untransfected QSG7701 cells. CONCLUSION: HCV core protein suppresses cell proliferation and apoptosis by downregmicrolating the phosphorylation of MAPKs and activating the transcriptors AP-1 and NF-kappaB, thus promoting the persistency of HCV infection which leads to chronic hepatitis C and hepatocellular cancer.

Effect of deleted pancreatic cancer locus 4 gene transfection on biological behaviors of human colorectal carcinoma cells.

AIM: To investigate the effect of deleted pancreatic cancer locus 4 (DPC4) gene transfection on biological behaviors of human colorectal carcinoma cells and the role of DPC4 gene in colorectal carcinogenesis. METHODS: PcDNA3.1-DPC4 plasmid was re-constructed by gene-recombination technology. SW620 cells, a human colorectal carcinoma cell line, were transfected with PcDNA3.1-DPC4 plasmid using lipofectamine transfecting technique. Transfected cells were selected with G418. Expression of Smad4 protein was detected in cells transfected with DPC4 gene by immunohistochemistry and Western blot. Biological characterristics of transfected cells were evaluated by population-doubling time and cloning efficiency. Alterations of percentage of S phage cells (S%) and apoptosis rate were determined by flow- cytometry. RESULTS: PcDNA3.1-DPC4 plasmid was constructed successfully. SW620 cells transfected with PcDNA3.1-DPC4 plasmid (DPC4+-SW620 cells) showed a strong intracellular expression of Smad4 protein, and the positive signal was localized in cytoplasm and nuclei, mainly in cytoplasm, where the expressions of Smad4 protein in SW620 cells transfected with PcDNA3.1 plasmid (PcDNA3.1-SW620 cells) and non-transfected SW620 cells (SW620 cells) were weaker than those in DPC4+-SW620 cells. The population- doubling time in DPC4+-SW620 cells (116 h) was significantly longer than that in SW620 cells (31 h) and PcDNA3.1-Sw620 cells (29 h) (P<0.01). The cloning efficiencies of DPC4+-SW620 cells (12%) were markedly lower than those of SW620 cells (69%) and PcDNA3.1-Sw620 cells (67%) (P<0.01). Compared with SW620 cells and PcDNA3.1-Sw620 cells, the G0-G1% of DPC4+-SW620 cells was obviously higher and the S% was markedly lower (P<0.05). Apoptosis rate of DPC4+-SW620 cells was significantly higher than that of SW620 cells and PcDNA3.1-SW620 cells. CONCLUSION: PcDNA3.1-DPC4 plasmid can be successfully re-constructed and stably transfected into human SW620 cells, thereby the cells can steadily express Smad4. DPC4 protein may regulate proliferation of colorectal carcinoma cells by inhibiting cell growth and inducing cell apoptosis.

Effects of transforming growth interacting factor on biological behaviors of gastric carcinoma cells.

AIM: Transforming growth interacting factor (TGIF) is an inhibitor of both transforming growth factor beta (TGF-beta) and retinoid signaling pathways. Moreover, the activation of MAPK pathway can prolong its half-life. However, its role in carcinogenesis is still unknown. Thus we attempted to investigate the effect of TGIF on biologic behaviors of gastric carcinoma cells. METHODS: Gastric carcinoma cell line, SGC-7901, was stably transfected with plasmid PcDNA3.1-TGIF. Western blotting and cell immunohistochemistry screening for the highly expressing clone of TGIF were employed. The growth of transfected cells was investigated by MTT and colony-formation assays, and apoptosis was measured by flow cytometry (FCM) and transmission electron microscopy. Tumorigenicity of the transfectant cells was also analyzed. RESULTS: TGIF had no effect on the proliferation, cell cycle and apoptosis of SGC-7901 cells, but cellular organelles of cells transfected with TGIF were richer than those of vector control or parental cells. Its clones were smaller than the control ones in plate efficiency, and its tumor tissues also had no obvious necrosis compared with the vector control or parental cells. Moreover, TGIF could resist TGF-beta mediated growth inhibition. CONCLUSION: TGIF may induce differentiation of stomach neoplastic cells. In addition, TGIF can counteract the growth inhibition induced by TGF-beta.

[Growth suppression of colon cancer cells in vitro by DPC4 gene expression and its mechanism].

OBJECTIVE: To study the effect of DPC4 gene expression on the growth of colon cancer cells and its mechanism. METHODS: Expression plasmid pcDNA3.1-DPC4 was constructed and transfected into the colon cancer cell line SW620 by use of lipofectamine gene transfer technique. DPC4 protein expression was detected by Western blot and immunocytochemistry. The effect of DPC4 gene on the growth of SW620 cells was monitored by population doubling time (PDT) and cloning efficiency. The influence of DPC4 expression on p21WAF1 transcription was investigated by RT-PCR to detect p21WAF1 mRNA. RESULTS: Successful expression of DPC4 protein was detected in the transfected SW620 cells. Compared with the control cells, PDT (74 h) of the DPC4 expressing cells was prolonged and the cloning efficiency (21%) decreased. In addition, the mRNA level of p21(WAF1) in DPC4 transfected cells was increased. CONCLUSIONS: Overexpression of DPC4 gene inhibits the growth of colon cancer in vitro, and induction of p21(WAF1) expression may be an important functional aspect of DPC4.