Higher TIGIT+CD226- γδ T cells in Patients with Acute Myeloid Leukemia.

The diverse structural and functional heterogeneity of γδ T cells is related to their distinct role in cancer immunity. The different phenotypes of γδ T cells in patients with acute myeloid leukemia (AML) is far from clear. In particular, the expression pattern of co-inhibitory and co-stimulatory receptors on γδ T cells remains unknown. In this study, we analyzed the distribution of γδ T cell subsets by expression of the immune checkpoint co-inhibitor TIGIT (T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain) and its competing co-stimulatory receptor CD226 in AML patients of different clinical statuses (including de novo AML, AML in non-remission (NR), and AML in complete remission (CR)). Our data demonstrated an imbalanced distribution of TIGIT and CD226 on γδ T cells with a decrease in CD226+ γδ T cells and an increase in TIGIT+ γδ T cells in de novo AML patients, while TIGIT-CD226+ γδ T cells were restored in AML patients who achieved CR after chemotherapy. Moreover, the patients who had higher TIGIT+CD226- γδ T cells showed lower overall survival rate for non-M3 AML, which may be considered a novel prognostic immune biomarker. In conclusion, our study reveals for the first time that imbalance in the TIGIT/CD226 axis might be related to different clinical outcomes for AML patients.Abbreviations: AML: acute myeloid leukemia; CR: complete remission; ICs: immune checkpoints; PD-1: programmed death-1; γδ T cells: gamma delta T cells; TCR: T cell receptor; MHC: major histocompatibility complex; TIGIT: T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain; NK: natural killer; PB: Peripheral blood; NR: non-remission; FAB: French-American-British; WHO: World Health Organization; HIs: healthy individuals; OS: overall survival.

Vasorin stimulates malignant progression and angiogenesis in glioma.

Glioma, the most common human primary brain tumor, is characterized by invasive capabilities and angiogenesis. Vasorin (VASN), a transmembrane protein, is reported to be associated with vascular injury repair and is overexpressed in some human tumors. However, its role in tumor progression and angiogenesis in glioma is unknown. In this study, VASN was shown to be overexpressed in high-grade gliomas, and the expression level correlated with tumor grade and microvessel density in glioma specimens. Glioma patients with high VASN expression had a shorter overall survival time. Knockdown of VASN in glioma cells by shRNA significantly inhibited the malignancy of glioma, including cell proliferation, colony formation, invasion, and sphere formation. Ectopic expression of VASN increased glioma progression in vitro. The expression of VASN correlated with the mesenchymal type of glioblastoma multiforme (GBM) subtyped by gene set enrichment analysis (GSEA). Our results showed that the concentration of VASN was increased in the conditioned medium (CM) from glioma cells with VASN overexpression, and the CM from glioma cells with knockdown or overexpressed VASN inhibited or promoted HUVEC migration and tubulogenesis in vitro, respectively. Glioma growth and angiogenesis were stimulated upon ectopic expression of VASN in vivo. The STAT3 and NOTCH pathways were found to be activated and inhibited by VASN overexpression. Our findings suggest that VASN stimulates tumor progression and angiogenesis in glioma, and, as such, represents a novel therapeutic target for glioma.

CD44 collaborates with ERBB2 mediate radiation resistance via p38 phosphorylation and DNA homologous recombination pathway in prostate cancer.

CD44, a glycoprotein, has been reported to have relationship with resistance to radiation in prostate cancer (Cap) cells. However, its molecular mechanism remains unknown. In this study, we demonstrated that inhibited CD44 enhanced the radiosentivity in Cap cells. It has been hypothesized that CD44 combine with ERBB2 and activate downstream phosphated protein to mediate DNA damage repair. Therefore, we conducted a detailed analysis of effects of radiation by clonogenic assay and immunofluorescence stain for p-H2AX foci. The downstream of CD44/ERBB2 and DNA damage repair proteins was detected by western blot. The results reveal that CD44 interacted with ERBB2, the downstream of CD44/ERBB2 was p-p38 when Cap cells were irradiated. Among the pathways, homologous recombination (HR) related proteins Mre11 and Rad50 were involved in CD44/ERBB2/p-p38 mediated radioresistance in Cap. In conclusion, CD44 could stabilize ERBB2 and co-activate p-p38 expression then promote the DNA damage repair by HR pathway, which finally contribute to the radioresistance of CaP.

miR-182-5p Promotes Growth in Oral Squamous Cell Carcinoma by Inhibiting CAMK2N1.

BACKGROUND/AIMS: Emerging evidence suggests that the propagation of oral squamous cell carcinoma (OSCC) is influenced by the abnormal expression of microRNAs (miRNAs). This study aimed to characterize the involvement of miR-182-5p in OSCC by targeting the calcium/ calmodulin-dependent protein kinase II inhibitor CAMK2N1. METHODS: miR-182-5p expression was quantified in OSCC tissues and cell lines with reverse transcription polymerase chain reaction (RT-PCR). Cell colony formation, Cell Counting Kit-8 (CCK-8), Ki-67, and nude mouse xenograft assays were used to characterize the role of miR-182-5p in the proliferation of OSCC. A miR-182-5p target gene was identified with western blotting, RT-PCR, and luciferase activity assays. OSCC patient survival based on CAMK2N1 expression was also analyzed. RESULTS: miR-182-5p was up-regulated in in vitro cell lines and in vivo clinical OSCC samples. CCK-8, colony formation, and Ki-67 assays revealed that miR-182-5p promoted the growth and proliferation of OSCC cells. miR-182-5p directly targeted CAMK2N1, as evidenced by luciferase assays and target prediction algorithms. CAMK2N1 operated as a tumor suppressor gene in patients with OSCC. Down-regulating miR-182-5p expression in the CAL-27 cell line restored CAMK2N1-mediated OSCC cell proliferation. miR-182-5p expression inhibited the activation of AKT, ERK1/2, and NF-κB. Mice injected with CAL-27 cells transfected with miR-182-5p-inhibitor demonstrated a significant increase in tumor size and weight and increased CAMK2N1 mRNA and protein expression compared with the miR-negative control group. CONCLUSION: The miR-182-5p-CAMK2N1 pathway can be potentially targeted to regulate the proliferation of OSCC cells.

Reciprocal androgen receptor/interleukin-6 crosstalk drives oesophageal carcinoma progression and contributes to patient prognosis.

Oesophageal squamous cell carcinoma (ESCC), a leading lethal malignancy of the digestive tract, is characterized by marked gender disparity. Clarifying the roles of the function and regulatory pathway of the androgen receptor (AR) will improve our understanding of oesophageal cancer progression, thereby facilitating the personalized management of ESCC. Here we report evidence to show that AR is a key mediator of inflammatory signals in ESCC cancer progression. High AR expression was associated with poor overall survival in tobacco-using ESCC patients but not in ESCC patients not using tobacco. A gain and loss of AR function enhanced and repressed ESCC cell growth, respectively, by altering cell cycle progression. In mice bearing human ESCC xenografts, silencing AR expression attenuated tumour growth, whereas AR overexpression promoted tumour growth in mice of different androgen statuses (male, female, and castrated male). Array assays revealed that the inflammatory cytokine interleukin-6 (IL6) is a prominent AR target gene in ESCC. By directly binding to the IL6 promoter, AR enhances IL6 transcription, and IL6 can in turn activate AR expression, thus forming a reciprocal regulatory circuit to sustain STAT3 oncogenic signalling in ESCC. Moreover, high expression levels of both AR and IL6 in human ESCC predict poor clinical outcome in tobacco users. Together, these data establish that AR promotes ESCC growth and is associated with poor patient prognosis. The discovery of a positive feedback loop between IL6 and AR bridges the knowledge gaps among lifestyle factor-associated inflammation, gender disparity, and oesophageal carcinoma. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Mitochondrial estrogen receptor ß inhibits cell apoptosis via interaction with Bad in a ligand-independent manner.

Previous studies reported that estrogen receptor ß (ERß) is localized to mitochondria, whereas little is known about the physiological functions of mitochondrial ERß. In the present study, we explored the role of mitochondrial ERß in regulating apoptosis using stable ERß-expressing and ERß knockdown cells lines. We found that exogenous ERß was mainly expressed in mitochondrial but not in nuclear after ERß overexpression and protected cells from apoptosis induced by hydrogen peroxide (H2O2), ultraviolet (UV), and staurosporine (STS). Moreover, overexpression of ERß prevented Bax activation, cytochrome c release, caspase-3 activation, and PARP cleavage during apoptosis. Furthermore, knockdown of ERß significantly suppressed the expression of ERß in mitochondrial and promoted cell apoptosis induced by H2O2, UV, and STS. Downregulation of ERß also enhanced Bax activation, cytochrome c release, caspase-3 activation and PARP cleavage. In addition, our study discovered that mitochondrial ERß interacted with proapoptotic protein Bad in a ligand-independent manner, which suggests that mitochondrial ERß inhibits Bad, and prevents Bax activation and cytochrome c release. Collectively, the results of this study support that mitochondrial ERß prevents cell apoptosis via the mitochondrial apoptotic pathway in a ligand-independent manner.

EGCG inhibits properties of glioma stem-like cells and synergizes with temozolomide through downregulation of P-glycoprotein inhibition.

RATIONAL: Combination therapy to inhibit cancer stem cells may have important clinical implications. Here, we examine the molecular mechanisms by which epigallocatechin gallate (EGCG), a bioactive polyphenol in green tea, inhibits the stem cell characteristics of glioma stem-like cells (GSLCs) and synergizes with temozolomide (TMZ), a DNA-methylating agent commonly used as first-line chemotherapy in gliomas. GSLCs were enriched from the human glioblastoma cell line U87 using neurosphere culture. Cells were analyzed using flow cytometry, quantitative PCR, and western blotting. Compared to U87 cells, a higher percentage of U87 GSLCs remained in the G0/G1 phase, with downregulation of the cell-cycle protein CylinD1 and overexpression of stem cell markers CD133 and ALDH1. The drug-resistance gene ABCB1 (but not ABCG2 or MGMT) also showed high mRNA and protein expression. The resistance index of U87 GSLCs against TMZ and carmustine (BCNU) was 3.0 and 16.8, respectively. These results indicate that U87 GSLCs possess neural stem cell and drug-resistance properties. Interestingly, EGCG treatment inhibited cell viability, neurosphere formation, and migration in this cell model. EGCG also induced apoptosis, downregulation of p-Akt and Bcl-2, and cleaving PARP in a dose-dependent manner. Importantly, EGCG treatment significantly downregulated P-glycoprotein expression but not that of ABCG2 or MGMT and simultaneously enhanced sensitivity to TMZ. Our study demonstrates that the use of EGCG alone or in combination with TMZ may be an effective therapeutic strategy for glioma.

SNX-25a, a novel Hsp90 inhibitor, inhibited human cancer growth more potently than 17-AAG.

17-Allylamino-17-demethoxygeldanamycin (17-AAG), a typical Hsp90 inhibitor derived from geldanamycin (GA), has entered Phase III clinical trials for cancer therapy. However, it has several significant limitations such as poor solubility, limited bioavailability and unacceptable hepatotoxicity. In this study, the anticancer activity and mechanism of SNX-25a, a novel Hsp90 inhibitor, was investigated comparing with that of 17-AAG. We showed that SNX-25a triggered growth inhibition more sensitively than 17-AAG against many human cancer cells, including K562, SW-620, A375, Hep-2, MCF-7, HepG2, HeLa, and A549 cell lines, especially at low concentrations (<1 µM). It showed low cytotoxicity in L-02, HDF and MRC5 normal human cells. Compared with 17-AAG, SNX-25a was more potent in arresting the cell cycle at G2 phase, and displayed more potent effects on human cancer cell apoptosis and Hsp90 client proteins. It also exhibited a stronger binding affinity to Hsp90 than 17-AAG using molecular docking. Considering the superiority effects on Hsp90 affinity, cell growth, cell cycle, apoptosis, and Hsp90 client proteins, SNX-25a is supposed as a potential anticancer agent that needs to be explored in detail.

MicroRNA-221 targeting PI3-K/Akt signaling axis induces cell proliferation and BCNU resistance in human glioblastoma.

MicroRNAs (miRNAs) are short regulatory RNAs that negatively regulate protein biosynthesis at the post-transcriptional level and participate in the pathogenesis of different types of human cancers, including glioblastoma. In particular, the levels of miRNA-221 are overexpressed in many cancers and miRNA-221 exerts its functions as an oncogene. Nevertheless, the roles of miRNA-221 in carmustine (BCNU)-resistant glioma cells have not been totally elucidated. In the present study, we explored the effects of miRNA-221 on BCNU-resistant glioma cells and the possible molecular mechanisms by which miRNA-221 mediated the cell proliferation, survival, apoptosis and BCNU resistance were investigated. We found that miR-221 was overexpressed in glioma cells, including BCNU-resistant cells. Moreover, we found that miR-221 regulated cell proliferation and BCNU resistance in glioma cells. Overexpression of miR-221 led to cell survival and BCNU resistance and reduced cell apoptosis induced by BCNU, whereas knockdown of miR-221 inhibited cell proliferation and prompted BCNU sensitivity and cell apoptosis. Further investigation revealed that miR-221 down-regulated PTEN and activated Akt, which resulted in cell survival and BCNU resistance. Overexpression of PTEN lacking 3'UTR or PI3-K/Akt specific inhibitor wortmannin attenuated miR-221-mediated BCNU resistance and prompted cell apoptosis. We propose that miR-221 regulated cell proliferation and BCNU resistance in glioma cells by targeting PI3-K/PTEN/Akt signaling axis. Our findings may provide a new potential therapeutic target for treatment of glioblastoma.

4E-BP1 regulates the sensitivity of human glioma cells to chemotherapy through PI3K/Akt/mTOR-independent pathway.

Drug resistance is one of the most formidable obstacles for treatment of glioma. Eukaryotic initiation factor 4E-binding protein (4E-BP1), a key component in the rate-limiting step of protein translation initiation, is closely associated with poor prognosis in multiple tumor types. However, it is unclear whether 4E-BP1 is involved in the drug resistance of human glioma. Herein we show that the expression of 4E-BP1 in human SWOZ2-BCNU drug-resistant glioma cells is significantly lower than that of the parent SWOZ2 cell line. Moreover, down-regulation of 4E-BP1 by short interfering RNA significantly impaired the sensitivity of SWOZ2 and U251 cells to carmustine (BCNU). Furthermore, overexpression of 4E-BP1 with plasmid transfection regained this sensitivity. Clinical studies showed that the expression levels of 4E-BP1 in primary glioma tissues were markedly higher than those of recrudescent glioma tissues. Taken together, our results suggest that 4E-BP1 is a novel protein that contributes to acquired drug resistance and it may be a potential target for reversing drug resistance in human glioma.

Epithelial-to-mesenchymal transition is involved in BCNU resistance in human glioma cells.

Chemotherapy has been considered as an effective treatment for malignant glioma; however, it becomes increasingly ineffective with tumor progression. Epithelial-to-mesenchymal transition (EMT) is a process whereby cells acquire morphologic and molecular alterations that facilitate tumor metastasis and progression. Emerging evidence associates chemoresistance with the acquisition of EMT in cancer. However, it is not clear whether this phenomenon is involved in glioma. We used the previously established human glioma cell lines SWOZ1, SWOZ2 and SWOZ2-BCNU to assess cellular morphology, molecular changes, migration and invasion. We found that BCNU-resistant cells showed multiple drug resistance and phenotypic changes consistent with EMT, including spindle-shaped morphology and enhanced pseudopodia formation. Decreased expression of the epithelial adhesion molecule E-cadherin and increased expression of the mesenchymal marker vimentin were observed in BCNU-resistant SWOZ1 and SWOZ2-BCNU cells compared to SWOZ2 cells. Migratory and metastatic potentials were markedly enhanced in SWOZ1 and SWOZ2-BCNU cells compared to SWOZ2 cells. These data suggest that there is a possible link between drug resistance and EMT induction in glioma cells. Gaining further insight into the mechanisms underlying chemoresistance and EMT may enable the restoration of chemosensitivity or suppression of metastasis.

A novel selenadiazole derivative induces apoptosis in human glioma cells by dephosphorylation of AKT.

Selenadiazole derivatives are synthetic organoselenium compounds with improved anticancer activity and greater selectivity than inorganic selenium. In this study, 4-(benzo[c][1,2,5]selenadiazol-6-yl)-benzene-1,2-diamine (BSBD) was shown to induce time- and dose-dependent apoptosis in SWO-38 human glioma cells by accumulation of a sub-G1 cell population, DNA fragmentation, nuclear condensation, caspase activation and poly(ADP-ribose) polymerase (PARP) cleavage. Further mechanistic investigation showed that BSBD treatment induced dephosphorylation of AKT and DNA damage-mediated activation of p53, leading to extensive apoptosis through the mitochondrial pathway. Our findings suggest that BSBD represents a potential human glioma therapeutic.

The positive correlation between DJ-1 and ß-catenin expression shows prognostic value for patients with glioma.

The relationship between DJ-1 and ß-catenin, and its impact on the prognosis for glioma patients has not been fully understood. This study determined the effect of DJ-1 on ß-catenin and the prognostic significance of this interaction in glioma patients. We collected tumor specimens from 88 glioma patients and determined the expression of DJ-1, ß-catenin and PTEN by using immunohistochemical staining. The involvement of DJ-1 and ß-catenin in glioma cell lines was evaluated by immunohistochemistry and Western blotting. High DJ-1 expression (37.5%) and high ß-catenin expression (34.1%) in glioma specimens were significantly associated with high grade and poor prognosis in glioma patients. However, only high levels of DJ-1 (P = 0.014) was a strong independent prognostic factor, correlated with a reduced overall survival time. In vitro DJ-1 expression was positively correlated with the expression levels of ß-catenin and p-Akt, and negatively correlated with PTEN expression in U87, U251 MG, SWO-38 and SHG44 human glioma cell lines. After the knockdown of DJ-1, Akt, p-Akt or ß-catenin expression levels were not affected in the PTEN-null cell lines (U87 and U251 MG). However, in the SWO-38 cell line, which has wild-type PTEN protein, the level of PTEN increased while Akt/p-Akt and ß-catenin levels were reduced. Furthermore, ß-catenin staining weakened in SWO-38 cells after DJ-1 levels decreased according to immunocytochemical analysis. In conclusion, DJ-1 and ß-catenin may contribute to the development and recurrence of glioma and are valuable prognostic factors for glioma patients. DJ-1 may regulate ß-catenin expression via PTEN and p-Akt.

Conditioned serum-free medium from umbilical cord mesenchymal stem cells has anti-photoaging properties.

Chronic exposure to solar radiation is the primary cause of photoaging and benign and malignant skin tumors. A conditioned serum-free medium (SFM) was prepared from umbilical cord mesenchymal stem cells (UC-MSCs) and its anti-photoaging effect, following chronic UV irradiation in vitro and in vivo, was evaluated. UC-MSC SFM had a stimulatory effect on human dermal fibroblast proliferation and reduced UVA-induced cell death. In addition, UC-MSC SFM blocked UVA inhibition of superoxide dismutase activity. Topical application of UC-MSC SFM to mouse skin prior to UV irradiation blocked the inhibition of superoxide dismutase and glutathione peroxidase activities, and prevented the upregulation of malonaldehyde. UC-MSC SFM thus protects against photoaging induced by UVA and UVB radiation and is a promising candidate for skin anti-photoaging treatments.

Transplantation of adipose-derived mesenchymal stem cells into a murine model of passive chronic immune thrombocytopenia.

BACKGROUND: Immune thrombocytopenia (ITP) is a bleeding disorder characterized by antibody-opsonized platelets (PLTs) being prematurely destroyed by macrophages in the reticuloendothelial system. T helper (Th) cells and different Th cytokines play an important role in the pathophysiology of ITP. As immunomodulators, adipose-derived mesenchymal stem cells (ADSCs) regulate Th cells and show therapeutic effects in autoimmune diseases. However, it is not clear how ADSCs affect ITP. In this study, we explored the specific effects of ADSCs on ITP in mice. STUDY DESIGN AND METHODS: BALB/c mice were randomly divided into three groups: normal controls, ITP controls, and ITP with ADSC transplantation. PLT levels were monitored by an automatic blood cell counter, and the cytokines interferon-γ (IFN-γ); interleukin (IL)-2, -4, -10, and -17; and transforming growth factor-ß1 (TGF-ß1) were analyzed by enzyme-linked immunosorbent assays. RESULTS: Compared to the untreated ITP mice, the PLT level of the ITP mice significantly increased after ADSC treatment. In the ADSC group, IFN-γ, IL-2, and IL-17 significantly decreased, while IL-4, IL-10, and TGF-ß1 increased. CONCLUSION: These findings constitute the first experimental evidence that ADSCs are efficacious in improving PLT levels and reducing the related Th cytokines mediating proinflammatory response in ITP mice, which may provide a scientific basis for using ADSCs as a new therapy for ITP.

Glycogen Synthase Kinase 3ß Inhibitor (2'Z,3'E)-6-Bromo-indirubin- 3'-Oxime Enhances Drug Resistance to 5-Fluorouracil Chemotherapy in Colon Cancer Cells.

OBJECTIVE: To explore the effects and mechanism of glycogen synthase kinase 3ß (GSK-3ß) inhibitor (2'Z,3'E)-6-bromo-indirubin-3'-oxime (BIO) on drug resistance in colon cancer cells. METHODS: The colon cancer SW480 and SW620 cells were treated with BIO, 5-fluorouracil (5-FU) and BIO/5-FU, separately. Cell cycle distribution, apoptosis level and efflux ability of rhodamine 123 (Rh123) were detected by flow cytometry. The protein expressions of P-glycoprotein (P-gp), multidrug resistance protein 2 (MRP2), thymidylate synthase (TS), ß-catenin, E2F-1 and Bcl-2 were detected by Western blot. ß-catenin and P-gp were stained with double immunofluorescence and observed under a confocal microscope. RESULTS: BIO up-regulated ß-catenin, P-gp, MRP2 and TS, enhanced the efflux ability of Rh123, decreased Bcl-2 protein and gave the opposite effect to E2F-1 protein in SW480 and SW620 cells. Furthermore, BIO significantly inhibited cell apoptosis, increased S and G(2)/M phase cells, and reduced the cell apoptosis induced by 5-FU in SW480 cells, whereas the effects were slight or not obvious in SW620 cells. CONCLUSION: GSK-3ß was involved in drug resistance regulation, and activation of ß-catenin and inhibition of E2F-1 may be the most responsible for the enhancement of 5-FU chemotherapy resistance induced by GSK-3ß inhibitor BIO in colon cancer.

[Effects of human umbilical cord mesenchymal stem cells in the treatment of paraquat-induced lung injury].

OBJECTIVE: To evaluate the clinical effect and safety of human umbilical cord mesenchymal stem cells (HUCMSCs) in the treatment of lung injury caused by acute paraquat poisoning. METHODS: Thirteen patients with lung injury caused by acute paraquat poisoning, who were admitted to Guangzhou No. 12 People's Hospital from December 2008 to December 2012, were divided into HUCMSC group (n = 5) and control group (n = 8). All patients received conventional treatment, while the HUCMSC group was treated with HUCMSCs as an addition. Sequential Organ Failure Assessment (SOFA) system, which was created by the Infection Section of European Society of Intensive Care Medicine, and Acute Physiology and Chronic Health Evaluation II were used to acquire the SOFA scores of patients. The lung injury was evaluated with lung injury score (LIS). The two groups were compared with respect to maximum SOFA scores at 1, 3, 5, 7, 14, and 15 days after paraquat poisoning. RESULTS: The HUCMSC group showed significantly lower maximum SOFA scores than the control group at 15d after poisoning (1.80 ± 2.05 vs 13.50 ± 7.59, P < 0.05). The LISs of the HUCMSC group after treatment (0.45 ± 0.27) were significantly lower than those of the HUCMSC group before treatment (1.15 ± 0.34) and those of the control group after treatment (2.94 ± 1.20) (P < 0.01). In the HUCMSC group, all patients survived, and they complained no discomfort and showed normal liver, kidney, and lung functions in reexamination; one patient showed incompletely absorbed shadow in the posterior segment of the left lower lobe of the lung during lung CT scan, and no abnormal findings were seen in other patients. In the control group, one patient survived, and others died. No adverse reactions, such as chill and fever, were presented in the HUCMSC group. CONCLUSION: HUCMSCs show promise for clinical application in the treatment of lung injury caused by acute paraquat poisoning.

MELK predicts poor prognosis and promotes metastasis in esophageal squamous cell carcinoma via activating the NF­κB pathway.

Esophageal squamous cell carcinoma (ESCC) is one of the most common malignancies worldwide with a low 5­year survival rate due to the lack of effective therapeutic strategies. Accumulating evidence has indicated that maternal embryonic leucine zipper kinase (MELK) is highly expressed in several tumors and associated with tumor development. However, the biological effects of MELK in ESCC remain unknown. In the present study, cell phenotypical experiments and animal metastasis assays were performed to detect the influence of MELK knockdown in vitro and in vivo. The potential molecular mechanism of MELK­mediated ESCC metastasis was further investigated by western blotting and immunofluorescence staining. The results revealed that the expression of MELK in human ESCC tissues was higher than that in adjacent normal tissues and was positively associated with the poor prognosis of patients. Reducing MELK expression resulted in growth inhibition and suppression of the invasive ability of ESCC cells in vitro and in vivo. MELK inhibition induced alterations of epithelial­mesenchymal transition­associated proteins. Mechanistically, MELK interacted with IκB kinase (IKK) and promoted the phosphorylation of IKK, by which MELK regulated activation of the NF­κB pathway. Collectively, the present study revealed the function and mechanism of MELK in the cell metastasis of ESCC, which may be a potential therapeutic target for ESCC.

[Hepatocyte growth factor combined with autologous bone marrow mesenchymal stem cell transplantation for treatment of silicosis].

OBJECTIVE: To evaluate the potential role of hepatocyte growth factor (HGF) combined with bone marrow mesenchymal stem cells (BMSC) autograft for the treatment of silicosis. METHODS: Bone marrow (100 ml) was aspirated from a severe silicosis patient. BMSCs isolated, purified and cultured in vitro. When BMSC came to 70% confluence at passage 3, the culture medium was added liposomes (lipo2000) and plasmid-HGF (p-HGF) and cultured for 2 d. HGF-MSCSs (5 × 10(7) cells) were resuspended in 50 ml 0.9% sodium chloride (NS) and infused Intravenous drip at 3 consecutive times (once a week). Clinical follow-up were performed before and after treatment: (1) pulmonary high-kV X-ray, chest CT examination; (2) pulmonary function test; (3) determination of serum ceruloplasmin. RESULTS: The symptoms such as coughing, chest tightness disappeared at 12 months after treatment. Pulmonary function tests showed significant changes after treatment: forced vital capacity (FVC) increased from 64.6% to 81.0%, forced expiratory volume in one second (FEV(1.0)) increased from 68.7% to 90.1%, 1 second rate (FEV(1.0)/FVC%) reduced from 111.6% to 107.1%, the maximum mid-expiratory flow (FEF(25%∼75%) decreased from 100.2% to 94.6%, forced expiratory vital capacity 75% of the moment bit of gas flow (MEF(75%)) increased from 99.2% to 113.5%, forced expiratory vital capacity 50% of the moment bit of gas flow (MEF(50%)) increased from 125.3% to 130.2%, forced expiratory vital capacity 25% of the moment bit of gas flow (MEF(25%)) reduced from 86.9% to 71.7%; serum ceruloplasmin levels decreased from 690 mg/L to 180.6 mg/L; lung high-kV X-ray at 1st review showed that diffuse lung nodules had been absorbed and getting smaller than before treatment; chest CT showed that the distribution and number of small nodules at double lung fields decreased than before treatment. CONCLUSION: HGF combined with BMSC transplantation may have some potential role for the treatment of silicosis patients.

[The comparative study of quartz dust and bleomycin-induced pulmonary fibrosis in rats].

OBJECTIVE: To compare the pulmonary alveolitis and the early fibrosis of pulmonary fibrosis induced by quartz dust and bleomycin in rats, and investigate their mechanism. METHODS: The female rats were divided into three groups: control group exposed to normal saline by the trachea; SiO2 group exposed to SiO2 by the trachea; BLM group exposed to BLM A5 by the trachea. Each half of the animals were sacrificed on the 7th and 14th day after exposure. The lungs of rats were collected to observe pulmonary alveolitis by HE staining and to observe fibrosis by saturated picric acid sirius red staining. The expression of tumor necrosis factor-alpha (TNF-alpha) and CD68 in pulmonary tissues were analyzed quantitatively by immunohistochemistry and image analysis system. RESULTS: (1) The alveolitis and pulmonary fibrosis of rats in both SiO2 group and BLM group were became more serious gradually over time, HE staining under light microscope showed that BLM group on the 7th day had the most obvious alveolitis (2.814 +/- 0.832), the saturated picric acid sirius red staining under polarized light showed that BLM group on the 14th day had the worst pulmonary fibrosis (1284.57 +/- 554.72), which were significantly higher than those (103.69 +/- 18.29 and 111.78 +/- 37.45) in control group and SiO2 group on the 7th day (P < 0.05). (2) The results of immunohistochemistry examination indicated that the expression (17.100 +/- 1.831) of TNF-alpha in the BLM group on the 7th day was significantly higher than those (0.451 +/- 0.441, 7.909 +/- 1.275 and 13.506 +/- 1.454) in control group, SiO2 group on 7th day and BLM group on 14th day (P < 0.05). The expression (22.778 +/- 2.512) of TNF-alpha in the SiO2 group on the 14th day was significantly higher than those in control group, SiO2 group on 7th day and BLM group on 14th day (P < 0.05). The expression (134.941 +/- 35.951) of CD68 in the SiO2 group on the 14th day was significantly higher than those in control group, SiO2 group on 7th day and BLM group on 14th day (P < 0.05). CONCLUSION: The early alveolitis of BLM-induced lung injury model was more serious than that of SiO2-induced lung injury model, and the fibrosis process of BLM-induced lung injury model was earlier than that of SiO2-induced lung injury model. TNF-alpha plays an important role in the course of both models, but macrophages is involved in SiO2-induced pulmonary in a more continuous way than in BLM-induced pulmonary fibrosis.