MIR-448 Regulates MAGEA6/AMPK Signaling Pathway in Hepatocellular Carcinoma Tumor Stem Cells

Objective: To explore the role of miR-448 in regulating MAGEA6/AMPK signaling pathway in the biological study of hepatocellular carcinoma (HCC) tumor stem cells. Methods: Using the database, the hepatocellular carcinoma related expression chips were obtained and the regulatory mirnas of candidate genes were predicted, and the predicted results were analyzed. The effects of miR-448 and MAGEA6 on the pellet formation rate and clone formation rate of hepatocellular carcinoma stem cells were detected by immunofluorescence identification of stem cell markers and light microscope counting method. The effects of miR-448 and MAGEA6 on migration and invasion of hepatocellular carcinoma stem cells were detected by scratch and Transwell assay. Dual luciferase reporter assay to verify whether miR-448 targets MAGEA6. The expression and influence of miR-448 on MAGEA6 and AMPK pathway were detected by qRT-PCR and Western blot. Results: It was found that miR-448 may directly regulate the expression of MAGEA6. Overexpression of miR-448 inhibited the characteristics, proliferation, migration, and invasion of hepatocellular carcinoma stem cells in vitro, as well as the ability of xenograft tumor formation in vivo. However, inhibition of miR-448 showed opposite results. In addition, miR-448 directly targets MAGEA6 and regulates AMPK signaling. Silencing MAGEA6 and adding AMPK activator further verified that miR-448 activated AMPK signaling pathway by targeting MAGEA6, thus affecting characteristics, proliferation, migration and invasion of hepatoma stem cells. Conclusions: Our results reveal that miR-448 activates AMPK signaling pathway by targeting MAGEA6, thereby affecting characteristics, proliferation, migration and invasion of hepatoma stem cells. It is suggested that overexpression of miR-448 may be a new therapeutic strategy for hepatocellular carcinoma.

Similarity on neural stem cells and brain tumor stem cells in transgenic brain tumor mouse models

Although it is believed that glioma is derived from brain tumor stem cells, the source and molecular signal pathways of these cells are still unclear. In this study, we used stable doxycycline-inducible transgenic mouse brain tumor models （c-myc/SV40Tag＋/Tet-on＋） to explore the malignant trans- formation potential of neural stem cells by observing the differences of neural stem cells and brain tumor stem cells in the tumor models. Results showed that chromosome instability occurred in brain tumor stem cells. The numbers of cytolysosomes and autophagosomes in brain tumor stem cells and induced neural stem cells were lower and the proliferative activity was obviously stronger than that in normal neural stem cells. Normal neural stem cells could differentiate into glial fibrillary acidic protein-positive and microtubule associated protein-2-positive cells, which were also negative for nestin. However, glial fibrillary acidic protein/nestin, microtubule associated protein-2/nestin, and glial fibrillary acidic protein/microtubule associated protein-2 double-positive cells were found in induced neural stem cells and brain tumor stem cells. Results indicate that induced neural stem cells are similar to brain tumor stem cells, and are possibly the source of brain tumor stem cells.

MicroRNAs in tumor stem cells

MicroRNAs （miRNAs） are a class of non-coding RNAs that are believed to have a significant role in tumorigenesis and cancer metastasis. Cancer stem cells play a major role in tumor recurrence, metastasis, and drug resistance. Research has shown that miRNAs can promote or inhibit the sternness of cancer stem cells and regulate the differentiation and self-renewal of cancer stem cells. In this article, the phenotype and regulatory mechanisms of miRNAs in cancer stem cells will be described, together with an explanation of their potential role in tumor diagnosis and treatment.

Experimental study on dendritic cells pulsed with brain tumor stem cells for immunotherapy of glioma

Objective To investigate the effect of dendritic cells pulsed with brain tumor stem cells which are used to treat on intracranial glioma. Methods We obtained murine brain tumor stem cells by grow ing C6 cells in epidermal grow th factor/basic fibroblast grow th factor w ithout serum.Dendritic cells isolated from rat bone marrow w ere pulsed w ith BTSCs. Rat brain

Differentiation profile of brain tumor stem cells:a comparative study with neural stem cells

Understanding of the differentiation profile of brain tumor stem cells （BTSCs）, the key ones among tumor cell population, through comparison with neural stem cells （NSCs） would lend insight into the origin of glioma and ultimately yield new approaches to fight this intractable disease. Here, we cultured and purified BTSCs from surgical glioma specimens and NSCs from human fetal brain tissue, and further analyzed their cellular biological behaviors, especially their differentiation property. As expected, NSCs differentiated into mature neural phenotypes. In the same differentiation condition, however, BTSCs exhibited distinguished differences. Morphologically, cells grew flattened and attached for the first week, but gradually aggregated and reformed floating tumor sphere thereafter. During the corresponding period, the expression rate of undifferentiated cell marker CD 133 and nestin in BTSCs kept decreasing, but 1 week later, they regained ascending tendency. Interestingly, the differentiated cell markers GFAP and β-tubulinlII showed an expression change inverse to that of undifferentiated cell markers. Taken together, BTSCs were revealed to possess a capacity to resist differentiation, which actually represents the malignant behaviors of glioma.

Research Progress in the Regulation of Tumor Cells and Tumor Stem Cells at Multiple Targets by Antrodia camphorata

Extensive in vitro and in vivo research reveals multiple intracellular molecular targets of Antrodia camphorata,and these targets affect growth,apoptosis,angiogenesis,invasion and metastasis of cells.These targets include tumor suppressor,cell cycle regulator,transcription factor,angiogenesis and metastasis factor,apoptosis and survival regulator,etc.Additionally,more and more attention has been paid to the molecular mechanism of A.camphorata on the regulation of tumor stem cells.Meanwhile,there is evidence that the immunoregulation of A.camphorata is enhanced,which may lead cell cycle arrest or apoptosis.In this paper,molecular mechanism of tumor cells and tumor stem cells regulated at multiple targets by A.camphorata in vitro and in vivo in the past decade is summarized.

IL-13Ra2-and glioma stem cell-pulsed dendritic cells induce glioma cell death in vitro

Objective Gliomas are the most common malignant tumors in the central nervous system.Despite multiple therapies including surgery,chemotherapy,and radiotherapy,the prognosis of patients remains poor.Immunotherapy is an alternative method of treating glioma,and the use of dendritic cell vaccines is one of the promising treatment options.However,there is no specific tumor cell antigen that can trigger dendritic cells(DCs).IL-13Ra2 is a specific antigen expressed in glioma cells;in the current study,we have attempted to explore whether IL-13Ra2 could be the antigen that triggers DCs and to envisage its application as potential therapy for glioma.Methods The expression of IL-13Ra2 was detected in U251 glioma cell lines and primary glioma tissues using different methods.DCs from human blood were isolated and pulsed with recombinant IL-13Ra2,following which the cytotoxicity of these DCs on glioma cells was detected and analyzed.Results About 55.9% human glioma tissue cells expressed IL-13Ra2,while normal brain tissue cells did not show any expression.DC vaccines loaded with IL-13Ra2,glioma cell antigen,and brain tumor stem cell(BTSC) antigen could significantly stimulate the proliferation of T lymphocytes and induce cell death in the glioma tissue.Compared to other groups,DC vaccines loaded with BTSC antigen showed the strongest ability to activate cytotoxic T lymphocytes(CTLs),while the glioma cell antigen group showed no significant difference.Conclusion IL-13Ra2,which is expressed in gliomas and by glioma stem cells,as well as IL-13Ra2 could prove to be potential antigens for DC vaccine-based immunotherapy.

Meningioma originating from the superior petrosal vein without dural attachment:A case report

BACKGROUND Meningioma in the cerebellopontine angle(CPA)without dural attachment is extremely rare.We report a unique case of meningioma derived from the superior petrosal vein without dural attachment.CASE SUMMARY A 44-year-old right-handed woman presented with a two-month history of headache and tinnitus.Brain magnetic resonance imaging showed a well-defined contrast-enhancing lesion in the right CPA without a dural tail sign.Tumor resection was performed using a right retro sigmoid approach.A dural attachment was not seen at the tentorium or posterior surface of the petrous pyramid.The tumor was firmly adherent to the superior petrosal vein.The origin site was cauterized and resected with the preservation of the superior petrosal vein.A diagnosis of meningothelial meningioma was made.The patient’s headache and tinnitus gradually disappeared,and a recurrence was not observed five years after the surgery.CONCLUSION The rare occurrence of meningioma without dural attachment makes it difficult to determine dural attachment before surgery.The absence of dural attachment makes it easy to completely resect such tumors.Vessels related to tumors should be removed carefully,considering the possible presence of tumor stem cells in the microvessels.

Identification of label-retaining cells in human gastric cancer xenograft in nude mice

Objective:This study has investigated the existence of label-retaining cell and its distribution in gastric cancer,in the hope that this information will assist investigations on gastric cancer stem cells.Methods:The gastric carcinoma cell line BGC-823 was labeled with BrdU in vitro and then engrafted into the right axilla of nude mice,which developed tumors.Label-retaining cells were quantified by immunohistochemical methods.Results:BrdU positive cells constituted about 96%of the cells in xenograft tumors after 10 days.Subsequently,BrdU positive cells gradually decreased,at the 80th day,labelretaining cells steadily occupied about 0.5%.This set of population cell localized in the margin of cancer nests,which had no difference in cellular morpha.Conclusion:The study demonstrates the presence of label-retaining cells in human gastric cancer xenografts in nude mice and the label-retaining cells may be related with cancer stem cells,which are most likely the cause for spread,metastasis and recurrence.

Pancreas duodenal homeobox-1 expression and significance in pancreatic cancer

AIM： To study the correlations of Pancreas duodenal homeobox-1 with pancreatic cancer characteristics, including pathological grading, TNM grading, tumor metastasis and tumor cell proliferation. METHODS： Reverse transcriptase-polymerase chain reaction （RT-PCR） was used to detect PDX-1 mRNA expression in pancreatic cancer tissue and normal pancreatic tissue. The expression of PDX-1 protein was measured by Western blot and immunohistochemistry. Immunohistochemistry was also used to detect proliferative cell nuclear antigen （PCNA）. Correlations of PDX-1 with pancreatic cancer characteristics, including pathological grading, TNM grading, tumor metastasis and tumor cell proliferation, were analyzed by using χ^2 test. RESULTS： Immunohistochemistry showed that 41.1% of pancreatic cancers were positive for PDX-1 expression, but normal pancreatic tissue except islets showed no staining for PDX-1. In consistent with the result of imunohistochemistry, Western blot showed that 37.5% of pancreatic cancers were positive for PDX-1. RT-PCR showed that PDX-1 expression was significantly higher in pancreatic cancer tissues than normal pancreatic tissues （2^-3.56±0.35 vs 2^-8.76±0.14, p 〈 0.01）. Lymph node metastasis （P 〈 0.01）, TNM grading （P 〈 0.05）, pathological grading （P 〈 0.05） and tumor cell proliferation （P 〈 0.01） were significantly correlated with PDX-1 expression levels. CONCLUSION： PDX-1 is re-expressed in pancreatic cancer, and PDX-l-positive pancreatic cancer cells show more malignant potential compared to PDX-l-negative cells. Therefore, PDX-l-positive cells may be tumor stemcells and PDX-1 may act as alternate surface marker of pancreatic cancer stem cells.

Expression of CD44,CD24 and ESA in pancreatic adenocarcinoma cell lines varies with local microenvironment

BACKGROUND:Emerging evidence suggests that pancreatic adenocarcinoma is hierarchically organized and sustained by pancreatic cancer stem cells.Furthermore,elimination of these cells is possible and therapeutically relevant.This study aimed to investigate the expression patterns of pancreatic cancer stem cell surface markers CD44,CD24 and ESA in pancreatic adenocarcinoma cell lines and explore the influence of their local microenvironment.METHODS:Flow cytometry was used to analyze the expression patterns of CD44,CD24 and ESA in five pancreatic adenocarcinoma cell lines (PANC-1,PC-2,MIA-Paca-2,AsPC-1 and BxPC-3).In addition,the capacity for sphereformation in serum-free medium of four cell lines (PANC-1,PC-2,MIA-Paca-2 and BxPC-3) was assessed.Then,the same assays were performed when tumor cell spheres were developed.The role of sonic hedgehog (SHH) in cell spheres from PANC-1 and MIA-Paca-2 were also assessed by RT-PCR.RESULTS:CD44 and CD24 were detected in PANC-1.Only CD44 expression was detected in PC-2,MIA-Paca-2 and AsPC-1.CD44,CD24 and ESA were all detected in BxPC-3.Tumor cell spheres developed in PANC-1 and MIA-Paca-2 in serumfree medium.This was accompanied by an increase in CD24 expression and a decrease in CD44 expression in PANC-1.Interestingly,the expression of CD44 and CD24 returned to initial levels once the medium was changed back from serumfree to serum-containing medium.No significant change in the expression of CD44 was detected in MIA-Paca-2.Furthermore,the relative quantification of SHH mRNA in PANC-1 cell spheres was significantly higher than that in cells cultured in the serum-containing medium.CONCLUSION:The expression patterns of the pancreatic cancer stem cell surface markers CD44,CD24 and ESA were diverse in different pancreatic adenocarcinoma cell lines and changed with their local microenvironment.

Intravenous transplantation of mesenchymal stem cells attenuates oleic acid induced acute lung injury in rats

Background Acute lung injury （ALl） and end-stage acute respiratory distress syndrome （ARDS） were among the most common causes of death in intensive care units. The activation of an inflammatory response and the damage of pulmonary epithelium and endotheliumwerethe hallmark of ALI/ARDS. Recent studies had demonstrated the importance of mesenchymal stem cells （MSCs） in maintaining the normal pulmonary endothelial and epithelial function as well as participating in modulating the inflammatory response and they are involved in epithelial and endothelial repair after injury. Here, our study demonstrates MSCs therapeutic potential in a rat model of ALI/ARDS. Methods Bone marrow derived MSCs were obtained from Sprague-Dawley （SD） rats and their differential potential was verified. ALl was induced in rats byoleic acid （OA）, and MSCs were transplanted intravenously. The lung injury and the concentration of cytokines in plasma and lung tissue extracts were assessed at 8 hours, 24 hours and 48 hours after OA-injection. Results The histological appearance and water content in rat lung tissue were significantly improved at different time points in rats treated with MSCs. The concentration of tumor necrosis factor-a and intercellular adhesion molecular-1 in rats plasma and lung tissue extracts were significantly inhibited after intravenous transplantation of MSCs, whereas interleukin-10 was significantly higher after MSCs transplantation at 8 hours, 24 hours and 48 hours after OA-challenge. Conclusions Intravenous transplantation of MSCs could maintain the integrity of the pulmonary alveolar-capillary barrier and modulate the inflammatory response to attenuate the experimental ALI/ARDS. Transplantation of MSCs could be a novel cell-based therapeutic strategy for prevention and treatment of ALI/ARDS.

Clinical outcomes of ATP-tumor chemosensitivity assay directed chemotherapy in hepatocellular carcinoma

This study aims to investigate the clinical response of ATP tumor chemosensitivity assay(ATP-TCA)directed chemotherapy regimens delivered via hepatic artery infusion in 104 cases of primary liver carcinoma(PLC).Tumor tissue was obtained via laparotomy and cultivated in vitro.This tissue was put through the assay to determine chemosensitivity.A single drug regimen of either 5-FU,MMC and ADM and a combination drug regimen were used.The treatment assigned was dependent on the result of the ATP-TCA.In the control group,30 cases of diagnosed PLC were given the conventional three-combination drug.The two groups were evaluated after three courses of chemotherapy.The results are as follows.The overall response rate of sensitivity test ranged from 36%to 44%in the single drug therapy groups and 81%in the combination drug group.The clinical overall response rate was 75%in the treatment group and 56%in control group.The treatment group had better results than the control group as survival period over six months was 80% and over one year 44%.In the control group,survival period over six months was 60% and 30% over one year.In short,ATP-TCA directed chemotherapy shows better results for terminal stages of PLC in that you can decrease the dose of drugs thereby reducing the side-effects with possible improvements in therapeutic effects.

Effect of Acanthopanax giraldii Harms Var. Hispidus Hoo polysaccharides on the human gastric cancer cell line SGC-7901 and its possible mechanism

OBJECTIVE: To study the inhibitory effect of Acanthopanax giraldii Harms Var. Hispidus Hoo polysaccharides (AGP) on SGC-7901 gastric cancer cells and its possible mechanism. METHODS: Cell doubling time analysis, colony forming assay and MTT assay were adopted to study the inhibitory effect and its characteristics. We also analyzed the amount of protein expressed by oncogenes, antioncogenes and cell factors using flow cytometric analysis. RESULTS: AGP inhibited the proliferation of SGC-7901 cells and cell colony forming ability. AGP did not inhibit the viability and function of lymphocytes of peripheral blood in healthy subjects and human embryonic tenocytes, except for the highest dosage of AGP (P