Triptolide inhibits proliferation and invasion of colorectal cancer cells by blocking Nrf2 expression.

Triptolide (TPL), the main active ingredient of Tripterygium wilfordii, has anti-inflammatory, immunomodulatory, and antitumor actions. It can also inhibit cell proliferation and metastasis while promoting apoptosis of several tumors, such as colorectal cancer (CRC). However, the mechanism of TPL against CRC is not clear. This study was designed to investigate the effects and molecular mechanisms of TPL on the proliferation and invasion ability of CRC cells. A human CRC cell line (HT29 cell line) cultured in vitro was treated with different concentrations of TPL (0, 25, 50, and 100 nmol/L). The proliferation of cells was detected by MTT, the invasion ability of cells by Transwell, and the apoptosis level by flow cytometry. The protein expression levels of nuclear factor-erythroid 2-related factor 2 (Nrf2), matrix metalloproteinase (MMP)-2, and MMP-9 were detected by western blotting. After transfection with sh-Nrf2, HT29 cells were divided into NC group, NC + TPL group and sh-Nrf2 + TPL group, and the above assays were repeated for each group. TPL significantly inhibited the proliferation and invasion ability of HT29 cells and promoted apoptosis (p < .05). Notably, its inhibitory or promotional effects were concentration-dependent, which were enhanced with increasing drug concentration (p < .05). After silencing Nrf2 expression, the proliferation, and invasion ability of HT29 cells were further significantly inhibited while cells apoptosis was further promoted (p < .05). Besides, the decreased Nrf2 expression reduced the protein expression levels of MMP-2 and MMP-9 (p < .05). TPL can effectively inhibit the proliferation and invasion while promoting apoptosis of HT29 cells. And its mechanism of action may be related to the inhibition of Nrf2 signaling expression.

Comparative analysis of five type II TA systems identified in Pseudomonas aeruginosa reveals their contributions to persistence and intracellular survival.

Background: Pseudomonas aeruginosa is a grave nosocomial pathogen that persistently inhabits the lungs of patients with cystic fibrosis (CF) and causes various chronic infections. The bacterial toxin-antitoxin (TA) system is associated with latent and long-term infections, but the underlying mechanisms remain to be fully characterized. Methods: We here investigated the diversity and function of five genomic type II TA systems widely distributed among P. aeruginosa clinical isolates. We also examined the distinct structural features of the toxin protein from different TA systems and characterized their contributions to persistence, invasion ability, and intracellular infection caused by P. aeruginosa. Results: ParDE, PA1030/PA1029, and HigBA could modulate persister cell formation under treatment with specific antibiotics. Furthermore, cell-based transcriptional and invasion assays revealed that PA1030/PA1029 and HigBA TA systems were critical for intracellular survival. Discussion: Our results highlight the prevalence and diverse roles of type II TA systems in P. aeruginosa and evaluate the possibility of using PA1030/PA1029 and HigBA TA pairs as targets for novel antibiotic treatments.

High Expression of PKMYT1 Predicts Poor Prognosis and Aggravates the Progression of Osteosarcoma via the NF-κB Pathway in MG63 Cells.

BACKGROUND: Protein kinase, membrane-associated tyrosine/threonine 1 (PKMYT1) contributes to the proliferative, migratory, invasive and colony-forming capabilities of oncocytes. Dysregulated expression of PKMYT1 is associated with numerous malignancies. However, at present, the functional role of PKMYT1 in osteosarcoma is still not clarified. OBJECTIVE: The present study, therefore, aimed to investigate the prognostic value of PKMYT1 in osteosarcoma, and to explore the underlying molecular mechanism(s). METHODS: To meet this end, the expression level of PKMYT1 in osteosarcoma was measured by immunohistochemical analysis. The prognostic value of PKMYT1 in osteosarcoma was analyzed on the basis of R2: Genomics Analysis and Visualization Platform. The functional role of PKMYT1 was subsequently investigated in MG63 cells by knocking down PKMYT1 expression via lentivirus encoding shRNA. MTT assay, scratch-wound and Transwell assays were then used to determine whether PKMYT1 fulfills a role in the proliferative and invasive capabilities of the MG63 cells. Subsequently, the role of PKMYT1 in the apoptosis of the cells was assessed using western blot and immunofluorescence analyses. Finally, to determine whether PKMYT1 exerts its role through the NF-κB pathway, fibroblast-stimulating lipopeptide-1 (FSL-1) was used as an NF-κB activator. RESULTS: Compared with normal tissues, osteosarcoma tissues showed a significantly increased level of PKMYT1 expression. The clinical survival analysis indicated that patients with high PKMYT1 expression were associated with lower probabilities of overall survival and metastasis-free survival compared with those with low PKMYT1 expression levels. Knockdown of PKMYT1 inhibited the migratory and invasive capabilities of the MG63 cells, and also facilitated their apoptosis. Moreover, the knockdown of PKMYT1 restrained the NF-κB pathway in MG63 cells, whereas activating the NF- κB pathway ameliorated the effects of silencing PKMYT1 on MG63 cells, suggesting that PKMYT1 functions via the NF-κB pathway in MG63 cells. CONCLUSION: Taken together, the results of the present study have shown that a high expression level of PKMYT1 is associated with poor prognosis of osteosarcoma, and that PKMYT1 is able to aggravate the malignant progression of MG63 cells via negatively regulating the NF-κB pathway, suggesting that PKMYT1 may be a potential molecular therapeutic target for the treatment of osteosarcoma.

Clinical significance of RAC3 expression in glioma tissue and its effect on migration and invasion ability of glioma cells / 中华内分泌外科杂志

Objective:To investigate the expression of ras-related C3 botulinum toxin substrate 3 (RAC3) in glioma tissues and its effect on the migration and invasion of glioma cells.Methods:The expression of RAC3 in 57 glioma patients and their adjacent tissues from the First People’s Hospital of Shangqiu was detected by immunohistochemical assay. According to the experimental requirements, brain glioma cells U87MG were divided into experimental group and control group. The experimental group U87MG cells were transfected with RAC3-siRNA plasmid, and the control group U87MG cells were transfected with MOCK-siRNA plasmid. RAC3 mRNA in each group was detected by fluorescence quantitative PCR. The expressions of RAC3 and MMP2 in each group were detected by Western blot. Transwell was used to detect the migration and invasion ability of cells in each group.Results:The positive rate of RAC3 in glioma patients was 89.47% (51/57 cases) , and the expression rate in paracancer tissues was 14.04% (8/57 cases) . The expression rate of RAC3 in glioma tissues was significantly higher than that in paracancer tissues, with statistical significance ( P<0.01) . After siRNA transfection, mRNA expression of RAC3 in experimental group and control group was 1.23±0.20 and 0.43±0.12, and protein expression of RAC3 was 1.19±0.11 and 0.23±0.08, respectively. The expression of MMP2 protein was 1.19±0.11 and 0.23±0.08, respectively. The expression of MMP2 in experimental group was significantly decreased ( P<0.05) . Transwell assay showed that the number of invasive cells in experimental group and control group U87MG cells was (22±5) and (45±8) , and the number of migratory cells was (34±6) and (90±11) , respectively. In experimental group, U87MG cell migration and invasion ability decreased significantly (both P<0.05) . Conclusion:The high expression of RAC3 in glioma tissues may be related to the malignant degree of development, and affect the migration and invasion ability of glioma cells by regulating the expression of MMP2.

Mechanism of glycyrrhetinic acid on inhibiting malignant biological behaviors of melanoma by Wnt / β - catenin pathways / 安徽医科大学学报

Objective @#To explore the possible mechanism of glycyrrhetinic acid on inhibiting malignant biological behaviors of melanoma by Wnt / β-catenin pathways．@*Methods@#The melanoma cells B16-F10 were selected as the research objects．The concentration gradient tests (0，1，2，4 μmol /L) were conducted by MTT．The cells given cisplatin intervention was enrolled as positive controls．The cells invasion and migration were detected by Transwell chamber assay．The expression levels of Wnt / β-catenin pathway proteins，invasion and migration related proteins (MMP-2，MMP-9) in B16-F10 cells were detected by Wester blot．The xenograft models of nude mice were constructed，and they were divided into control group (without drugs treatment) and glycyrrhetinic acid group (40 mg / kg) ．The growth of tumor tissues，and expression levels of Wnt / β-catenin pathway proteins，invasion and migration related proteins were observed. @*Results @#MTT results showed that glycyrrhetic acid could inhibit the proliferation of B16-F10 cells in a concentration-dependent manner．The inhibition effect of glycyrrhetic acid ( ≥2 μmol /L) was significant on the proliferation of B16-F10 cells (P ＜0. 05) ．The results of Transwell chamber assay showed that compared with control group，invasion and migration abilities of B16-F10 cells were significantly reduced after treatment with glycyrrhetinic acid (2，4 μmol /L) (P ＜0. 05) ．Wester blot results showed that compared with those without glycyrrhetinic acid treatment，expression levels of MMP-9 ，MMP-2，Wnt1 and β-catenin protein in B16- F10 cells significantly decreased after treatment with glycyrrhetinic acid (2，4 μmol /L) (P＜0. 05) ．The results of tumor-bearing assay showed that compared with control group ，weight and volume of tumors significantly decreased in glycyrrhetinic acid group，and expression levels of Wnt1 ，β-catenin，MMP-9 and MMP-2 proteins also significantly decreased (P＜0. 05) ．@*Conclusion @#Glycyrrhetinic acid can significantly inhibit the malignant biological behaviors of melanoma in vitro and vivo．And its mechanism may be related to inhibiting the activation of Wnt / β-catenin signaling pathways.

The Mechanism of Downregulation of Twist1 Inhibiting Trophoblast Invasion and Aggravating the Development of Preeclampsia.

To study the expression of under expressed transcription factor Twist1 in preeclampsia (PE) and its effect on the invasion of placental trophoblast cells and to explore its related mechanism on the development of PE by establishing a pregnant rat model. Methods: the villi were collected from the induced abortion in the first trimester (6-8 weeks), the normal placenta (18-20 weeks) induced by the second trimester, the term placenta tissue of normal pregnancy (37-40 weeks), and the placental tissue of patients with PE, to detect the expression of Twist1. Trophoblast cells were subjected to primary culture in placental tissues of normal pregnant women and placental tissues of PE patients. The invasion ability of the two groups of trophoblasts was detected, and the primary cultured trophoblasts were divided into two groups: an experimental group and a control group. Specific Twist1 siRNA was added to the experimental group, and no reagents were added to the control group. The above-mentioned cells were given different interventions. To explore the effect of Twist1 on trophoblast cell invasion, cells were cultivated for 72 h. The SD rats were conceived. After the pregnancy was stable, the SD rats in different groups were treated with different treatments (interference with Twist1), and the average systolic blood pressure and urine protein of the gestational mothers in the different treatment groups were measured at 1 week, 2 weeks, and full-term pregnancy. The expression of Twist1 in the placenta tissue of SD rats with different interventions at full-term pregnancy was detected. The results showed that Twist1 expression is down-regulated in PE, and the invasion ability of placental trophoblast cells in PE patients is weak. After inhibiting Twist1, the mean tail artery pressure and urine protein level of SD pregnant rats increase, showing a trend of PE. The mechanism may be related to the inhibition of the placenta by Twist1 Trophoblast cell invasion.

Porphyromonas gingivalis outer membrane vesicles inhibit the invasion of Fusobacterium nucleatum into oral epithelial cells by downregulating FadA and FomA.

BACKGROUND: Porphyromonas gingivalis (P. gingivalis) and Fusobacterium nucleatum (F. nucleatum) participate in the formation and progression of periodontitis. They can exert virulence by invading into host cells, but the interaction between them and their specific mechanisms remain unclear. The purpose of this study was to study the effect of P. gingivalis outer membrane vesicles (OMVs) on the ability of F. nucleatum to invade oral epithelial cells, and the reasons for the influence. METHODS: The invasion abilities of the two bacteria were detected separately after mixed infection of P. gingivalis and F. nucleatum. Next, P. gingivalis OMVs were extracted with the kit, and their influence on the invasion ability of F. nucleatum was tested. The effects of P. gingivalis OMVs on F. nucleatum were evaluated by assessment of bacterial morphology, growth curves, auto-aggregation morphology, and the expression of adhesion-related proteins FadA and FomA. RESULTS: Our results showed that P. gingivalis inhibited the invasion of F. nucleatum into oral epithelial cells but F. nucleatum promoted the invasion of P. gingivalis. In subsequent experiments, we extracted P. gingivalis OMVs successfully and revealed that proteases in P. gingivalis OMVs inhibited the invasion of F. nucleatum into oral epithelial cells. Furthermore, P. gingivalis OMVs did not affect the morphology and proliferation of F. nucleatum, but proteases inside decreased the auto-aggregation of F. nucleatum. Additionally, proteases in P. gingivalis OMVs reduced the expression levels of F. nucleatum surface adhesion-related proteins FadA and FomA. CONCLUSION: Our study demonstrated that proteases in P. gingivalis OMVs inhibited the invasion of F. nucleatum into oral epithelial cells by downregulating FadA and FomA.

Intracellular invasion ability of Streptococcus agalactiae among non-invasive isolates from human adults and companion animals in Japan.

OBJECTIVE: This study evaluated the cell invasion ability (CIA) of Streptococcus agalactiae isolates from humans and companion animals and clarified the relationship between CIA populations and their microbiological features. METHODS: Human-origin and companion animal-origin isolates were collected along with host information. We measured CIA using human-lineage colon cancer epithelium (Caco-2) and keratinocyte (HaCaT) cell lines, via virulence-associated gene profiling (bca-rib-bac-lmb-cylE-hylB-pavA-pilB-spb1-srtC1-brpA), capsular genotyping, multilocus sequence typing, and antimicrobial resistance (AMR) phenotyping/genotyping. Significant differences in data regarding CIA into epithelium and keratinocytes and those of isolates from different hosts were assessed. We analyzed the association of CIA populations with the virulence genotypes, capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes. RESULTS: A comparative analysis was performed between human (n = 15) and canine (n = 17) non-invasive isolates. There was a difference in CIA data between Caco-2 and HaCaT cells using human and animal isolates. For percent invasion ability into Caco-2 cells, we designated values ≥ 0.1 as high-frequency CIA and values < 0.1 as low-frequency CIA. Fourteen isolates harbored high-frequency and 18 isolates harbored low-frequency strains. There was no association between the high-frequency population and the virulence genotypes, capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes. CONCLUSION: This is the first report assessing the invasion ability of S. agalactiae into HaCaT and Caco-2 cells. Our observations suggest that S. agalactiae is more capable of entering Caco-2 rather than HaCaT.

MiR-122 Promotes the Development of Colon Cancer by Targeting ALDOA In Vitro.

Non-coding RNAs, originally considered junk gene products, have taken center stage in view of their significant involvement in a spectrum of biological processes during human development, thereby offering novel therapeutic targets for improvement of treatment options. Accumulating evidence has demonstrated non-coding RNA dysfunction across various human cancers. In particular, microRNAs have emerged as key regulatory molecules in cancer biology. MicroRNAs are noninvasive, readily accessible biomarkers that can be effectively applied for diagnosis and prognosis of different tumor types, including colon cancer. In this study, we reanalyzed the available data with bioinformatics tools to identify differentially expressed microRNAs in colon cancer cells. The top 3 upregulated microRNAs (miR-10, miR-199, and miR-122) in colon cancer cells were further validated in tissues of clinical patients via reverse transcription-quantitative polymerase chain reaction. Our results showed that miR-122 significantly promotes the proliferation and invasion ability of SW480 and SW620 cells through inhibition of Aldolase, Fructose-Bisphosphate A (ALDOA) expression. We further summarized recent advances in our understanding of the functional relevance of microRNAs in cancer development and discussed the possible implications of specific microRNAs in colon cancer. This study extends our knowledge of microRNA involvement in colon cancer biology and presents novel candidates for the development of attractive therapeutic strategies.

Up regulation of miR-96-5p is responsible for TiO2 NPs induced invasion dysfunction of human trophoblastic cells via disturbing Ezrin mediated cytoskeletons arrangement.

Titanium dioxide nanoparticles (TiO2 NPs) are used extensively in our daily lives, and their toxic effects on the placenta have been reported. Animal studies indicated that placental development is impaired after maternal exposure of TiO2 NPs, but the underlying mechanisms remain largely unknown. In the present study, we used a human trophoblast-derived cell, HTR8-SVneo, to determine how TiO2 NPs affected placental functions, and found out potential reversal targets. TEM was employed for TiO2 NPs morphology observation and uptake assessment. RT-PCR was used to detect the expression of both mRNA and miRNA, and western blotting was used for protein examination. Cell invasion ability was evaluated by Transwell assay, and cytoskeletons were observed by immunofluorescence combined with confocal microscope examination. We found that TiO2 NPs disrupted cytoskeletons and impaired cell invasion ability. Further investigations showed that TiO2 NPs increased the expression of a microRNA (miR-96-5p), which targeted and down-regulated the translation of EZR mRNA, a gene that encodes ezrin protein, and affected the cell cytoskeletons and ultimately cell invasion ability. When the expression of miR-96-5p was down-regulated, the expression level of ezrin protein was also reversed, and cell invasion ability was partially restored. Collectively, we determined how miR-96-5p mediates TiO2 NP-induced placental dysfunction, and provided a potential rescue target for future therapy.

Adeno-associated virus type 2-mediated gene transfer of a short hairpin-RNA targeting human IGFBP-2 suppresses the proliferation and invasion of MDA-MB-468 cells.

Adeno-associated virus 2 (AAV2) is prepotent in the biological treatment of breast tumor because of its low pathogenicity and immunogenicity. Our previous study demonstrated that insulin­like growth factor­binding protein 2 (IGFBP­2) was highly expressed in patients with breast metastasis. In the present study, the effects of recombinant AAV2 on the growth and metastasis of breast cancer cells were determined in vitro, and in vivo. rAAV2-ZsGreen-shRNA-scramble and rAAV2­ZsGreen­shRNA­hIGFBP­2 were used to transfect MDA­MB­468, and MCF­10A cells respectively, and observed that these virus could not penetrate the normal human breast epithelia MCF­10A cell line. To investigate the effect of the recombinant virus on chemotherapeutics, paclitaxel was added to MDA­MB­468 cells and it was demonstrated that rAAV2­ZsGreen­shRNA­hIGFBP-2-infected MDA-MB-468 cells were highly chemosensitive to paclitaxel compared with rAAV2­ZsGreen­shRNA­scramble­injected cells. In addition, it was demonstrated that the invasive ability of rAAV2­ZsGreen­shRNA­hIGFBP­2­infected MDA-MB-468 cells was highly impaired compared with the rAAV2­ZsGreen­shRNA­scramble group. In the nude mice xenografts, the rAAV2­ZsGreen­shRNA­hIGFBP­2 injection inhibited tumor growth and Ki­67 expression was significantly downregulated compared with the scramble group. Following IGFBP­2 knockdown using rAAV2-ZsGreen-shRNA-hIGFBP­2, matrix metalloproteinase­2 expression was significantly reduced in tumor tissues compared with that in rAAV2­ZsGreen­shRNA­scramble treated tumor tissues. These findings have provided a direction for the application of novel AAV2­based therapeutics for treating aggressive triple­negative breast cancer types.

Interferon Regulatory Factor 5 Inhibits Invasion Ability of Nasopharyngeal Carcinoma by Reducing PARP-1 / 中山大学学报(医学科学版)

[Objective]To investigate whether IRF5 can inhibit invasion ability of nasopharyngeal carcinoma by re-ducing PARP-1(poly(ADP-ribose)polymerase-1).[Methods]Forty-six specimens of nasopharyngeal carcinoma and 51 specimens of normal tissue were confirmed by pathologically in this study.The expression of IRF5 and PARP-1 in naso-pharyngeal carcinoma tissues and normal tissues was detected by immunohistochemistry.The IFR5 overexpression plasmid was transfected into the nasopharyngeal carcinoma cell line CNE-2,quantitative PCR and immunoblotting was used to value the expression of IRF5 after transfection.The wound healing and transwell assay was used to investigate the invasion ability. The expression of PARP-1 was valued by quantitative PCR and immunoblotting after over-expression of PFR5.[Results]The results showed that the expression of IRF5 in cancer tissues was lower than that in normal tissues,but the PARP-1 expression was opposite. The IRF5 overexpressing cell line CNE-2/IFR5 was established. The healing rate of CNE-2/IFR5 cells was lower than that of the control cells(P<0.01). Transwell experiments revealed that the number of CNE-2/IFR5 cells passing through the basement membrane was smaller than that of the control group(P<0.01),suggest-ing that up-regulation of IFR5 could inhibit the invasiveness of nasopharyngeal carcinoma cells.Over-expression of IFR5 led to reduced PARP-1 mRNA and protein(P<0.01).Besides,elevation of PARP-1 can prevent IRF5-induced changes of invasion ability.[Conclusion]Therefore,we speculated that IRF5 can inhibit invasion ability of nasopharyngeal carci-noma by reducing the expression of PARP-1.This study provided a new target for inhibiting the invasion ability of naso-pharyngeal carcinoma based on IRF5.

miRNA-125b over-expression for promoting proliferation and invasion ability of lung cancer A549 cells and its mechanism / 重庆医学

Objective To study the influence of miRNA-125b over-expression on proliferation and invasion ability of lung cancer A549 cells and its mechanism.Methods A549 cells were divided into 3 groups:miRNA-125b group(transfected with miRNA-125b mimics),NC group(transfected with NC mimics) and blank group(same volume of GIBCO serum mixed with transfection agent).The transfection and expression efficiency of miRNA-125b was detected with Q-PCR,the proliferation ability was detected with MTT,and the invasion ability was detected with the transwell chamber test.The expression level of BMF in A549 cells was detected with Western blot.Results Compared with the blank group,the expression level of miRNA-125b,proliferation ability and invasion ability in the miRNA-125b group were increased(P＜0.05);while the above indexes in the NC group demonstrated no significant change(P＞0.05).Compared with the blank group,the expression level of BMF in the miRNA-125b group was decreased (P＜0.05);while which in the NC group had no significant change(P＞0.05).Conclusion miRNA-125b can promote the proliferation and invasion ability of A549 cells via inhibiting the expression of BMF.

Relationship between DLK1 gene promoter region DNA methylation and non-small cell lung cancer biological behavior.

We investigated the possible association between DLKI gene promoter region methylation and the increased invasion capacity of non-small cell lung cancer (NSCLC). Lung cancer cell line H1299, as well as the gene transfection and RNA interference technology were used to build DLK gene overexpression and knockdown cells. An in vitro invasion assay was performed to observe the changes in the invasion ability of lung cancer cells. Western blot analysis was used to verify Notchl and matrix metalloproteinase-9 (MMP-9) expression levels and a sulfurous acid sequencing technique was used to test the DNA methylation level in the promoter region. Our results showed that the invasion ability of cells in the overexpression group was significantly enhanced. This ability was considerably reduced in the knockdown group. The Notchl and MMP-9 expression level increased significantly in the overexpression group, while it was reduced considerably in the knockdown group. We detected significantly lower levels of DNA methylation in the promoter region in the overexpression group. It was concluded that methylation of the DLK1 gene promoter region increased the invasion ability of NSCLC. Furthermore, it is possible that this process is related to the Notch signaling pathway.

VCAM-1 improves migration and invasion of human glioma cell lines / 基础医学与临床

Objective To investigate the effects of VCAM-1 on migration and invasion of glioma cell lines . Methods The techniques of lentivirus pSGU6/GFP/Neo-based VCAM-1 shRNA and EF1 a-GFP/puro-based VCAM-1 expression vector, the scratch wound healing migration and transwell invasion assays , and the Western blot and cell staining were applied to observe the effects of VCAM-1 expression levels on migration and invasion of glioma cell line cells.There are four groups in T98G cells including control, vector, scramble and shRNA-VCAM-1 groups and three groups in U251 cells covering control, vector and VCAM-1 overexpressed groups ( n=6 per group) .Results The stabled glioma cell lines of T98 G cells with down-regulated VCAM-1 and U251 cells with VCAM-1 overexpression were established by using lentivirus-based VCAM-1 shRNA and expression vector.The ability of scratch wound healing (migration activity) decreased significantly (P<0.01) in T98G cells with lower VCAM-1 expression levels, while the migration activity was obviously improved in U251 cells with overexpressed VCAM-1 ( P <0.05 ) .Similarly, the invasion ability was significantly inhibited ( P <0.05) in T98G cells with silenced VCAM-1, as well as VCAM-1 overexpression could enhance the invasion ability of U251 cells ( P<0.01 ) .Conclusions VCAM-1 improves the migration activity and invasion ability of human glioma cell line cells.

Legionella dumoffii Tex-KL Mutated in an Operon Homologous to traC-traD is Defective in Epithelial Cell Invasion.

OBJECTIVE: To understand the mechanism of invasion by Legionella dumoffii. METHODS: The L. dumoffii strain Tex-KL was mutated using the Tn903 derivative, Tn903dIIlacZ. After screening 799 transposon insertion mutants, we isolated one defective mutant. We then constructed the gene-disrupted mutant, KL16, and studied its invasion of and intracellular growth in HeLa and A549 cells, and in A/J mice survival experiments. The structure of traC-traD operon was analyzed by RT-PCR. RESULTS: The transposon insertion was in a gene homologous to Salmonella typhi traC, which is required for the assembly of F pilin into the mature F pilus structure and for conjugal DNA transmission. Results from RT-PCR suggested that the traC-traD region formed an operon. We found that when the traC gene was disrupted, invasion and intracellular growth of L. dumoffii Tex-KL were impaired in human epithelial cells. When mice were infected by intranasal inoculation with a traC deficient mutant, their survival significantly increased when compared to mice infected with the wild-type strain.. CONCLUSION: Our results indicated that the traC-traD operon is required for the invasion and intracellular growth abilities of L. dumoffii Tex-KL in epithelial cells.

Effect of overexpression of HOX genes on its invasive tendency in cerebral glioma.

Transcription factors encoded by HOX genes are vital in the determination of cell fate and identity during embryonic development. In certain malignancies, HOX genes also behave as oncogenes. The present study demonstrated suppression of the invasive tendency of glioblastoma multiforme U-118 and U-138 cells by the introduction of the antisense fragments of HOXA6 and B13 genes using electroporation. The invasion index indicated 79 and 72% reductions in the invasive ability of antisense HOXA6 and B13, respectively. No significant differences in the invasive index of the parental and mock cells of each HOX gene were observed (invasive index, 0.75-0.91; P=0.05). A reduction in invasion tendency was also observed following betulinic acid (BA) treatment: The results from the matrigel assay analysis clearly demonstrated a significant inhibition in the invasive behaviour of U-118 and U-138 cell lines from day 15 following BA treatment, with a maximum effect on day 30. The invasion index demonstrated 62 and 65% reductions in invasion ability in the U-118 and U-138 cell lines, respectively. The suppression of HOXC6 and B13 expression by the introduction of the corresponding antisense fragments in addition to BA reduced invasion tendency in U-118 and U-138 cell lines. The mechanism underlying the association between the HOX gene and invasive behavior in glioma cells is yet to be understood. However, the anti-invasive behavior of BA may aid understanding of the mechanism in future studies.

Legionella dumoffii Tex-KL Mutated in an Operon Homologous to traC-traD is Defective in Epithelial Cell Invasion / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To understand the mechanism of invasion by Legionella dumoffii.</p><p><b>METHODS</b>The L. dumoffii strain Tex-KL was mutated using the Tn903 derivative, Tn903dIIlacZ. After screening 799 transposon insertion mutants, we isolated one defective mutant. We then constructed the gene-disrupted mutant, KL16, and studied its invasion of and intracellular growth in HeLa and A549 cells, and in A/J mice survival experiments. The structure of traC-traD operon was analyzed by RT-PCR.</p><p><b>RESULTS</b>The transposon insertion was in a gene homologous to Salmonella typhi traC, which is required for the assembly of F pilin into the mature F pilus structure and for conjugal DNA transmission. Results from RT-PCR suggested that the traC-traD region formed an operon. We found that when the traC gene was disrupted, invasion and intracellular growth of L. dumoffii Tex-KL were impaired in human epithelial cells. When mice were infected by intranasal inoculation with a traC deficient mutant, their survival significantly increased when compared to mice infected with the wild-type strain..</p><p><b>CONCLUSION</b>Our results indicated that the traC-traD operon is required for the invasion and intracellular growth abilities of L. dumoffii Tex-KL in epithelial cells.</p>

Hypoxia Promotes Invasion of Endometrial Stromal Cells via Hypoxia-Inducible Factor 1α Upregulation-Mediated ß-Catenin Activation in Endometriosis.

Endometriosis is a common benign gynecological disease defined as the presence of endometrial tissue outside the uterine cavity. The aim of this study was to identify the molecular mechanism underlying hypoxia-induced increases in invasive ability of human endometrial stromal cells (HESCs). Herein, we show that the expression levels of hypoxia-inducible factor lα (HIF-1α) and ß-catenin were greater in ectopic endometriotic tissue compared with eutopic tissue from controls. Exposure of eutopic endometrial stromal cells under hypoxic conditions or treated with desferrioxamine (DFO, chemical hypoxia) resulted in a time-dependent increase in ß-catenin expression and its dephosphorylation. Hypoxia/HIF-1α also activated the ß-catenin/T-cell factor (TCF) signaling pathway and the expression of target genes, vascular endothelial growth factor and matrix metalloproteinase 9, and knockdown of HIF-1α or ß-catenin abrogated hypoxia-induced increases in HESC invasiveness. These results suggest that HIF-1α interacting with ß-catenin/TCF signaling pathway, which is activated by hypoxia, may provide new insights into the etiology of endometriosis.

Changes of TIZ expression in epithelial ovarian cancer cells.

OBJECTIVE: To study the change of TIZ expression in epithelial ovarian cancer cells. METHODS: HO8910 cells were transinfected with siRNA to inhibit the expression of TIZ. pcDNA3.1-TIZ vectors were combined to increase the TIZ expression level. The cell viability, colony forming efficiency and cycle distribution of HO8910, HO8910/NC, HO8910/pcDNA3.1-NC, HO8910/TIZ-573 and H08910/pcDNA3.1-TIZ were compared, and the invasion rate, migration rate and adhesion rate between 5 groups of cells were compared. RESULTS: Compared with those of HO8910, HO8910/NC and HO8910/pcDNA3.1-NC, the cell viability, colony forming efficiency and cell cycle distribution of HO8910/TIZ-573 were increased, while the indexes of H08910/pcDNA3.1-NC were decreased with statistical significant difference (P<0.05). There was no statistical significant difference in the invasion rate, migration rate and adhesion rate between 5 groups of cells (P>0.05). CONCLUSIONS: The expression of TIZ can inhibit the proliferation of epithelial ovarian cancer cells.