Controlling Nutritional Status score predict the individualized survival of patients with gastric cancer.

BACKGROUND AND OBJECTIVES: The present study aimed to determine the correlation between Controlling Nutritional Status (CONUT) score and prognosis in gastric cancer patients undergoing total gastrectomy. METHODS AND STUDY DESIGN: The clinical data of 245 gastric cancer patients who underwent total gastrectomy in Peking University, First Hospital between January1st 2005 and December 30th 2015 were retrospectively collected. According to the CONUT level, they were divided into high CONUT (>3) group and low CONUT (≤3) group. The relationship between CONUT and the disease-free survival (DFS) and overall survival (OS) were analyzed by statistical analysis. RESULTS: The results showed that the optimal cutoff value for CONUT to predict the 5-year survival was 3 and CONUT had a higher area under the ROC curve (AUC) for 5-year disease free survival (DFS) and overall survival (OS) prediction. Additionally, when age was considered as a stratified factor, univariate analyses demonstrated that high CONUT correlated with shorter DFS in non-elderly (<65) patients and shorter DFS and OS in elderly (≥65) patients. CONCLUSIONS: High CONUT was significantly correlated with older age, advanced TNM-stage, higher Ki-67 and pathological subtype. Patients with high pre-operative high CONUT levels should be given more observation and constant follow-up after surgery.

Transformation of alkylating regimen of thiotepa into tepa determines the disease progression through GSTP1 gene polymorphism for metastatic breast cancer patients receiving thiotepa containing salvage chemotherapy.

BACKGROUND: The shifts to second-line chemotherapy for metastatic breast cancer (MBC) were widely required based on pharmaceutical molecular profiles to reach out precision medicine. The emerging precise treatment of cancer requires the implementation of clarified pharmacogenetic profiles which are capable of elucidating the predictive responses to cancer chemotherapy. Therefore we were interested in the analysis of the roles of single nucleotide polymorphism (SNP) of GSTP1 (glutathione S-transferase pi 1 gene) alleles to identify pharmacological links with predictors of clinical responses and toxicities. METHODS: 93 MBC patients receiving thiotepa plus docetaxel chemotherapy were enrolled in this study. Optimized CYP3A5, CYP2B6, and GSTP1 were predominantly selected as candidate genes and their three SNPs (CYP2B6 G516T, CYP3A5 A6986G, and GSTP1 A313G) were genotyped by matrix-assisted laser desorption ionization/time of flight (MALDI-TOF) mass spectrometry. Progression-free survival (PFS), disease control rate, and chemo-related toxicities were recorded. RESULTS: GSTP1 A313G (rs1695) was identified to be related with disease progression. In particular, patients harboring AG/GG genotype demonstrated a statistically longer PFS than those with AA. Multivariate analysis confirmed that AG/GG genotype was associated with both clinical responses and liver-localized metastatic lesions. No correlation was found between these three SNPs and chemotherapy-induced toxicity. CONCLUSIONS: These results suggest that the GSTP1 polymorphism is a novel prognostic marker for clinical response to thiotepa-containing chemotherapy regimens. Such evidence could provide insight into the role of pharmacogenetics to deprive of biases in shifting regimens solely by empirical choices.

Studies on microRNAs that are correlated with the cancer stem cells in chronic myeloid leukemia.

Accumulating data indicate that cancer stem cells play an important role in tumorigenesis and are underlying cause of tumor recurrence and metastasis, specifically in chronic myeloid leukemia (CML). We aim to detect the miRNAs that are correlated with the cancer stem cells in CML to provide theoretical basis for clinical application. We first analyzed microRNA expression profiles of CML leukemia patients compared with normal controls by microarray analysis and validated the results by real-time PCR. A single microRNA signature classified CML from normal was detected. We also determined the absolute copy numbers of these three microRNAs in normal adults. The results showed that three microRNAs (miR-150, miR-23a, and miR-130a) were identified to significantly decrease in expanded 38 CML patients compared with 90 normal controls. Molecular and statistical analysis showed that the decreased microRNAs were significant in clinical analysis. All these results indicated that those three microRNAs could act as a tumor suppressor and their decreased expression might be one of the causes of leukemia. Accordingly, clarifying their regulatory mechanisms might delineate their potentials as drug targets of gene therapy for CML.

The relationship between methylenetetrahydrofolate reductase polymorphism and hematological malignancy.

BACKGROUND: Methylenetetrahydrofolate reductase (MTHFR) is the key enzyme for folate metabolism. Previous studies suggest a relationship between its single nucleotide polymorphisms (SNP) of C677T and A1298C with a variety of tumor susceptibility including hematological malignancy. SNP frequency distribution in different ethnic populations might lead to differences in disease susceptibility. There has been little research in Chinese people on the MTHFR SNP with the susceptibility of the hematological malignancy. Therefore, this study investigated the relationship between MTHFR SNPs and hematological malignancy in Jiangsu province in China. METHODS: Gene microarray was used to detect MTHFR C677T and A1298C single nucleotide polymorphism loci on 157 healthy controls and 127 patients from Jiangsu province with hematological malignancies (30 with multiple myeloma, 28 with non-Hodgkin's lymphoma, 22 with acute lymphoblastic leukemia, 40 with acute myeloid leukemia, and seven with chronic myeloid leukemia). RESULTS: The allele frequency of 677T was 41.3% in patients and 33.1% in controls, showed significant difference (chi2 = 4.08, p = 0.043); 677TT genotype with a high susceptibility to hematological malignancy (OR 1.96, 95% CI 1.01 - 4.45, p = 0.041). In subgroup analyses, the genotypes 677TT and 1298CC were associated with significantly increased multiple myeloma risk (TT vs. CC: OR 8.92, 95% CI 1.06 - 75.24, p = 0.006; CC vs. AA: OR = 4.80, 95% CI 1.56 - 14.73, p = 0.044). No associations were found between polymorphisms and susceptibilities to acute lymphoblastic leukemia, acute myeloid leukemia, or non-Hodgkin's lymphoma. CONCLUSIONS: MTHFRC677T polymorphisms influence the risk of hematological malignancy among the population in Jiangsu province. Both MTHFR 677TT and MTHFR 1298CC genotypes increase susceptibility to myeloid leukemia.

Elevator block brake structural optimization design based on an approximate model.

An Aquila optimizer-back propagation (AO-BP) neural network was used to establish an approximate model of the relationship between the design variables and the optimization objective to improve elevator block brake capabilities and achieve a lightweight brake design. Subsequently, the constraint conditions and objective functions were determined. Moreover, the multi-objective genetic algorithm optimized the structural block brake design. Finally, the effectiveness of the optimization results was verified using simulation experiments. The results demonstrate that the maximum temperature of the optimized brake wheel during emergency braking was 222.09°C, which is 36.71°C lower than that of 258.8°C before optimization, with a change rate of 14.2%. The maximum equivalent stress after optimization was 246.89 MPa, 28.87 MPa lower than that of 275.66 MPa before optimization, with a change rate of 10.5%. In addition, the brake wheel mass was reduced from 58.85 kg to 52.40 kg, and the thermal fatigue life at the maximum equivalent stress increased from 64 times before optimization to 94 times after optimization.

Effects of transplanted bone-marrow-derived mesenchymal stem cells in animal models of acute hepatitis.

Our aim was to evaluate the therapeutic effects of bone-marrow-derived mesenchymal stem cells (BMMSCs) on ConA-induced hepatitis and to elucidate the possible mechanism involved. MSCs were isolated from bone marrow and their characteristics and anti-apoptotic effects on the L02 cell line were analyzed. The effect of intravenous infusion of BMMSCs on liver damage was also tested. Furthermore, the recruitment of donor BMMSCs to the liver of recipient animals and their effects on the activity of intrahepatic natural killer T (NKT) cells were investigated. BMMSCs ameliorated liver damage in a time- and dose-dependent manner. Donor BMMSCs were detected in the livers of recipient animals, suggesting that tissue damage stimulated the migration of BMMSCs. Transplanted BMMSCs also suppressed the activity of intrahepatic NKT cells, not only in the liver but throughout the body. The general infusion of BMMSCS ameliorated immunoregulatory activities by the suppression of intrahepatic NKT cells.

The comparition of biological characteristics and multilineage differentiation of bone marrow and adipose derived Mesenchymal stem cells.

We compared the two sources of adipose and bone marrow-derived mesenchymal stem cells (BMSCs and AMSCs ) in multiple differentiation capacity and biological characteristics to provide a theoretical basis for stem cells transplantation. We isolated bone marrow- and adipose-derived mesenchymal stem cells and compared their phenotype,cell doubling time, the secretion of factors and their ability of multi-differentiation. We also compared their differences in T lymphocyte activation, proliferation and suppression. BMSCs and AMSCs were similar in cell phenotype and the differences existed only in the expression of CD106. On the proliferation rate, AMSCs were faster than BMSCs (doubling time 28 vs. 39 h). In addition, both of these two sources of cells were able to differentiate into bone, fat and cartilage that proved their stem cells properties and the number of stem cell progenitors (CFU-F) from adipose tissue were 10 times larger than those from bone marrow. But AMSCs showed a diminished capacity for suppressing T lymphocyte proliferation and activation compared to BMSCs. Cell origin and abundance were decisive factors in stem cells applications and, in the same volume, with the same premise of AMSCs and BMSCs, adipose tissue is a more promising source of stem cells.

The comparison of multilineage differentiation of bone marrow and adipose-derived mesenchymal stem cells.

BACKGROUND: To compare the roles of adipose and bone marrow derived mesenchymal stem cells (AMSCs and BMSCs) in multiple differentiation capacity to provide a theoretical basis for stem cell transplantation. METHODS: We isolated bone marrow and adipose derived mesenchymal stem cells and compared their phenotype, cell doubling time, the secretion of factors, and the ability of multi-differentiation. RESULTS: BMSCs and AMSCs were similar in cell phenotype and the differences existed only between the expression of CD106. The proliferation rate of AMSCs was faster than of BMSCs (doubling time 28h vs. 39h) and the capacity to suppress T cell proliferation and activation was weakened in AMSCs. In addition, both sources of cells were able to differentiate into bone, fat, and cartilage which proved their stem cell properties. CONCLUSIONS: Cell origin and abundance were decisive factors in stem cell applications and with the same premise as for AMSCs and BMSCs, adipose tissue is a more promising source of stem cells.

Matrix metalloproteinase-9 silencing by RNA interference promotes the adhesive-invasive switch in HT1080 human fibrosarcoma cells.

BACKGROUND: A high level of matrix metalloproteinase-9 (MMP-9) is associated with human tumor invasion and/or metastasis. The HT1080 human fibrosarcoma cell line is highly invasive and metastatic which constitutively express MMP-9. METHODS: HT1080 cells transfected with a double stranded RNA that targeted the MMP-9 mRNA and the cellular characteristics were examined before and after interference. The inhibition effects of MMP-9 interference on the tumor growth of HT1080 cells in nude mice was also tested by xenograft assay. RESULTS: MMP-9 extinction in HT1080 resulted in the following: (1) inhibited cell mobility; (2) increased cell adhesion, and (3) attenuated tumor cell migration. In addition, MMP-9 knockdown concomitantly resulted in decreased levels of soluble ICAM-1, leading to an adhesion defect and tumor metastasis. Moreover, in vivo assay further demonstrated MMP-9 interference affecting the tumorigenesis of HT1080 cells in mice as follows (1) inhibition of tumor growth; (2) reduced tumor volume, and (3) prolonged survival time. CONCLUSIONS: Our observations defined a novel critical role for MMP-9 in the progression of HT1080 fibrosarcoma by changing the inter-cellular adhesion molecular-1 from membrane-anchored state to a soluble one which provides a target for promising tumor therapy in clinics.

Value of Preoperative Hematological Parameters in the Prognosis of Gastric Cancer Patients Undergoing a Total Gastrectomy.

OBJECTIVE: This study aimed to evaluate the relationships between the albumin/globulin ratio (AGR), neutrophil/lymphocyte ratio (NLR), and platelet/lymphocyte ratio (PLR) and clinicopathological information for gastric cancer patients. In addition, the prognostic values of these hematological parameters for resectable gastric cancer patients undergoing a total gastrectomy were determined. METHODS: A total of 245 patients with gastric cancer who underwent a total gastrectomy at our hospital between January 1, 2005, and December 30, 2015, were enrolled into this study. The preoperative AGR, NLR, and PLR in the serum samples of the patients were measured. The relationships between the hematological parameters and the disease-free survival (DFS) as well as overall survival (OS) were analyzed by statistical analysis. RESULTS: The cutoff values of AGR, NLR, and PLR were 1.57, 3.5, and 193, respectively. Univariate analyses demonstrated that a low AGR, a high NLR, and a high PLR were significant risk factors for a poor prognosis. According to multivariate analysis, a high PLR was found to be independently associated with a poor survival. Additionally, when age was considered as a stratified factor, univariate analyses demonstrated that a low AGR had the tendency to be correlated with a shorter DFS in nonelderly patients (<65 years old). A low AGR was significantly correlated with a shorter DFS and OS in elderly patients (≥65 years old). CONCLUSION: AGR, NLR, and PLR are independent risk factors associated with a poor gastric cancer survival by univariate analysis, and AGR is an independent risk factor for predicting DFS and OS in elderly patients (≥65 years old) with gastric cancer after total gastrectomy.

Mesenchymal stem cells induce mature dendritic cells into a novel Jagged-2-dependent regulatory dendritic cell population.

Mesenchymal stem cells (MSCs), in addition to their multilineage differentiation, exert immunomodulatory effects on immune cells, even dendritic cells (DCs). However, whether they influence the destiny of full mature DCs (maDCs) remains controversial. Here we report that MSCs vigorously promote proliferation of maDCs, significantly reduce their expression of Ia, CD11c, CD80, CD86, and CD40 while increasing CD11b expression. Interestingly, though these phenotypes clearly suggest their skew to immature status, bacterial lipopolysaccharide (LPS) stimulation could not reverse this trend. Moreover, high endocytosic capacity, low immunogenicity, and strong immunoregulatory function of MSC-treated maDCs (MSC-DCs) were also observed. Furthermore we found that MSCs, partly via cell-cell contact, drive maDCs to differentiate into a novel Jagged-2-dependent regulatory DC population and escape their apoptotic fate. These results further support the role of MSCs in preventing rejection in organ transplantation and treatment of autoimmune disease.

Effect of interleukin-31 on septic shock through regulating inflammasomes and interleukin-1ß.

Sepsis with severe systemic inflammation remains a great challenge for the intensive care unit in clinics. Although biomarkers have been identified to diagnose, monitor and predict these syndromes, novel therapeutic approaches are required for the amelioration of symptoms of sepsis and septic shock. The present study demonstrated that interleukin (IL)-31 was able reduce the mortality rate of lipopolysaccharide (LPS)-induced sepsis with the reduction of inflammatory cytokines in the sera. IL-31 also inhibited IL-1ß production in the peritoneal lavage fluid in LPS-induced or cecal ligation and puncture-induced sepsis. The in vitro mechanism responsible for IL-31 regulation on peritoneal IL-1ß activation following LPS challenge was explored. It was demonstrated that IL-1ß secretion was suppressed by IL-31 treatment from LPS-challenged peritoneal macrophages following adenosine triphosphate stimulation, which is an activator of NLR family, pyrin domain-containing 3 (NLRP3). Furthermore, IL-31 inhibited the expression of NLRP3 at the transcriptional level. In human THP-1 cells, anti-IL-31/anti-IL-31 receptor (R) neutralizing antibody enhanced NLRP3 expression as well as IL-1ß activation, suggesting a role of the IL-31-IL-31R-NLRP3-IL-1ß signaling axis in the physiological status of sepsis. On the other hand, IL-31 displayed a negative effect on the NLRP1 inflammasome, but not on NLRP3 on the LPS-primed human peripheral blood monocytes, resulting in reduction of the inflammatory cytokine, tumor necrosis factor (TNF)-α, in the supernatant. Taken together, the present data implied that T helper 2-type cytokine, IL-31, may be a promising therapeutic option for treatment of sepsis and septic shock in clinics.

Comparison of Drug-Eluting and Bare Metal Stents in Patients With Chronic Kidney Disease: An Updated Systematic Review and Meta-Analysis.

BACKGROUND: Drug-eluting stents (DESs) and bare metal stents (BMSs) are both recommended to improve coronary revascularization and to treat coronary artery disease in patients with chronic kidney disease (CKD). However, the potential superiority of DESs over BMSs for reducing the incidence of long-term major adverse cardiovascular events and mortality in CKD patients has not been established, and the results remain controversial. We aimed to systematically assess and quantify the total weight of evidence regarding the use of DESs versus BMSs in CKD patients. METHODS AND RESULTS: In this systematic review and conventional meta-analysis, electronic studies published in any language until May 20, 2016, were systematically searched through PubMed, Embase, Web of Science, and the Cochrane Central Register of Controlled Trials. We included randomized controlled trials and observational studies comparing outcomes in CKD patients with DESs versus BMSs and extracted data in a standard form. Pooled odd ratios and 95% CIs were calculated using random- and fixed-effects models. Finally, 38 studies involving 123 396 patients were included. The use of DESs versus BMSs was associated with significant reductions in major adverse cardiovascular events (pooled odds ratio 0.75; 95% CI, 0.64-0.88; P<0.001), all-cause mortality (odds ratio 0.81; 95% CI, 0.73-0.90; P<0.001), myocardial infarction, target-lesion revascularization, and target-vessel revascularization. The superiority of DESs over BMSs for improving clinical outcomes was attenuated in randomized controlled trials. CONCLUSIONS: The use of DESs significantly improves the above outcomes in CKD patients. Nevertheless, large-sized randomized controlled trials are necessary to determine the real effect on CKD patients and whether efficacy differs by type of DES.

Effects of mineralocorticoid receptor antagonists on left ventricular mass in chronic kidney disease patients: a systematic review and meta-analysis.

BACKGROUND: Mineralocorticoid receptor antagonists (MRAs) are used widely in treatment of heart failure, but their effects on cardiovascular complications and mortality of chronic kidney disease (CKD) are not well known. Thus, we aim to assess such therapeutic effects of MRAs on CKD. METHODS: Electronic literature published in any language until Dec 31, 2015, was systematically searched on PubMed, Embase, and Cochrane Central Register of Controlled Trials. Primary outcome was left ventricular mass (LVM) or LVM index (LVMI), and secondary outcome was all-cause mortality and major adverse cardiovascular events (MACEs). Results of continuous outcomes were pooled using mean difference (MD) and standard mean difference (SMD). Risk ratios (RRs) with 95 % confidence intervals (CIs) were pooled using a random- or fixed-effects model. RESULTS: Totally 12 studies (6 randomized controlled trials with 1003 participants) involving 4935 patients were included. MRA treatment versus non-MRA treatment resulted in a significant change of 0.93 SMD (standard mean difference) in LVM (LVMI), a significant reduction of 22 % in all-cause mortality, a significant reduction of incidence of MACEs (RR 0.65, P = 0.001), significantly higher prevalence rates of hyperkalemia (>5.5 mmol/L), but no significant change in prevalence rates of severe hyperkalemia (>6.0 mmol/L). CONCLUSION: MRA benefits CKD patients in terms of LVMI, all-cause mortality, and MACEs with no incidence of severe hyperkalemia. Nevertheless, the real effects of MRAs on cardiovascular events and mortality as well as their safety in CKD patients should be identified by further studies with prospective and large-sample clinical trials.

BCR/ABL mRNA targeting small interfering RNA effects on proliferation and apoptosis in chronic myeloid leukemia.

BACKGROUND: To investigate the effects of small interference RNA (siRNA) targeting BCR/ABL mRNA on proliferation and apoptosis in the K562 human chronic myeloid leukemia (CML) cell line and to provide a theoretical rationale and experimental evidence for its potential clinical application for anti-CML treatment. MATERIALS AND METHODS: The gene sequence for BCR/ABL mRNA was found from the GeneBank. The target gene site on the BCR/ABL mRNA were selected according to Max-Planck-Institute (MPI) and rational siRNA design rules, the secondary structure of the candidate targeted mRNA was predicted, the relevant thermodynamic parameters were analyzed, and the targeted gene sequences were compared with BLAST to eliminate any sequences with significant homology. Inhibition of proliferation was evaluated by MTT assay and colony-formation inhibiting test. Apoptosis was determined by flow cytometry (FCM) and the morphology of apoptotic cells was identified by Giemsa-Wright staining. Western blotting was used to analyze the expression of BCR/ABL fusion protein in K562 cells after siRNA treatment. RESULTS: The mRNA local secondary structure calculated by RNA structure software, and the optimal design of specific siRNA were contributed by bioinformatics rules. Five sequences of BCR/ABL siRNAs were designed and synthesized in vitro. Three sequences, siRNA1384, siRNA1276 and siRNA1786, which showed the most effective inhibition of K562 cell growth, were identified among the five candidate siRNAs, with a cell proliferative inhibitory rate nearly 50% after exposure to 12.5 nmol/L~50 nmol/L siRNA1384 for 24,48 and 72 hours. The 50% inhibitory concentrations (IC50) of siRNA1384, siRNA1276 and siRNA1786 for 24 hours were 46.6 nmol/L, 59.3 nmol/L and 62.6 nmol/L, respectively, and 65.668 nmol/L, 76.6 nmol/L, 74.4 nmol/L for 72 hours. The colony-formation inhibiting test also indicated that, compared with control, cell growth of siRNA treated group was inhibited. FCM results showed that the rate of cell apoptosis increased 24 hours after transfecting siRNA. The results of annexinV/PI staining indicated that the rate of apoptosis imcreased (1.53%, 15.3%, 64.5%, 57.5% and 21.5%) following treamtne with siRNAs (siRNA34, siRNA372, siRNA1384, siRNA1276 and siRNA1786). Morphological analysis showed td typical morphologic changes of apoptosis such as shrunken, fragmentation nucleus as well as "apoptotic bodies" after K562 cell exposure to siRNA. Western blot analysis showed that BCR/ABL protein was reduced sharply after a single dose of 50 nmol/L siRNA transfection. CONCLUSIONS: Proliferation of K562 cells was remarkbly inhibited by siRNAs (siRNA1384, siRNA1276 and siRNA1786) in a concentration-dependent manner in vitro, with effective induction of apoptosis at a concentration of 50 nmol/L. One anti-leukemia mechanism in K562 cells appeared that BCR/ABL targeted protein was highly down-regulated. The siRNAs (siRNA1384, siRNA1276 and siRNA1786) may prove valuable in the treatment of CML.

Antitumor effects of 32P-chromic-poly (L-lactide) brachytherapy in nude mice with human prostate cancer.

The aim of the present study was to investigate the antitumor effects and tissue distribution of 32P-chromic-poly (L-lactide) (32P-CP-PLLA) in nude mice with human prostate cancer. Tumor models were obtained by transplantation of PC-3M tumor cells into male BALB/c nude mice. Animals were randomly divided into control, 32P-chromic phosphate (32P-CP) colloid and 32P-CP-PLLA groups (all n=20). A series of indices were investigated, including apoptosis of tumor cells, rate of apoptosis, expression of caspase 3 and 8, biodistribution and intratumoral concentration of 32P-CP-PLLA, intensity of radioactivity, tumor volume and microvessel density (MVD). Highly concentrated radioactivity of 32P-CP-PLLA in the tumor mass was detected by single photon emission computed tomography (SPECT) scanning. The residual activities of the 32P-CP-PLLA and 32P-CP colloid groups were 3.02±0.32 and 1.76±0.31 MBq, respectively, on day 14 following treatment. The tumor inhibition rates were 67.24±3.55 and 55.92±7.65%, respectively (P<0.01). Necrotic changes, in conjunction with apoptosis, were observed in the treatment group. MVD values for the 32P-CP-PLLA and 32P-CP colloid groups were 28.24±10.07 and 36.15±11.06, respectively. 32P-CP-PLLA showed an excellent capacity for killing tumor cells, inducing apoptosis and inhibiting angiogenesis.

Expanding Sca-1(+) mammary stem cell in the presence of oestrogen and growth hormone.

BACKGROUND: Sca-1 is controversial as a mammary stem cell marker in the literature, which may be due to the different isolation protocols and culture media used in different laboratories. The object of our study is to establish the Medium to promote the proliferation of mammary stem cell and explore the possibility of Sca-1 as mammary stem cell marker. METHODS: We used BM medium supplemented with different concentration of 17Β-oestradiol and GH to find out MaECM medium which promoted the proliferation of mouse mammary epithelial cells and inhibited the growth of fibroblasts. Flow cytometry was used to isolate Sca-1(+) and Sca-1(-) cell populations from cultured mammary epithelial cells. Mammary fat pad transplantation and Mammosphere- forming assay were done to confirm the stem cell potential of Sca-1(+) cells. Differentiating culture was used to detect the differentiation potential of Sca-1(+) cells. Real-time PCR was carried out to analyse the expression of mammary stem cell-related genes in Sca-1(+) cells. RESULTS: We first selected the medium suitable for mammary stem cell growth. Stem cell medium BM was used to culture mammary organoids, which generated many fibroblasts. We established MaECM medium supplemented with oestrogen and growth hormone (GH), in which oestrogen promoted mammary epithelial cell proliferation and inhibited fibroblast growth, and GH obviously enhanced the effect of oestrogen on mammary epithelial cell proliferation. Flow cytometry showed that 50% of cells were Sca-1(+) under the culture of MaECM medium. We confirmed that Sca-1(+) cells regenerated mammary outgrowths when transplanted in vivo, formed mammospheres in vitro and differentiated into luminal epithelial cells with milk-secreting function and myoepithelial cells under Matrigel culture. Furthermore, gene expression analysis by Real-time PCR revealed that Sca-1(+) cells expressed markedly higher levels of mammary stem cell-related genes in comparison to Sca-1(-) cells. CONCLUSION: Our research demonstrates that Sca-1(+) mammary stem cells can be more easily isolated when cultured in the presence of oestrogen and GH.

TGF-beta1-induced PI3K/Akt/NF-kappaB/MMP9 signalling pathway is activated in Philadelphia chromosome-positive chronic myeloid leukaemia hemangioblasts.

Overwhelming evidence from chronic myeloid leukaemia (CML) research indicates that patients harbour quiescent CML stem cells that are responsible for blast crisis. While the haematopoietic stem cell (HSC) origin of CML was first suggested over 30 years ago, recently CML-initiating cells beyond HSCs are also being investigated. We have previously isolated fetal liver kinase-1-positive (Flk1(+)) cells carrying the BCR/ABL fusion gene from the bone marrow of Philadelphia chromosome-positive (Ph(+)) patients with hemangioblast property. Here, we show that these cells behave abnormally comparing with the hemangioblasts in healthy donors. These Ph(+) putative CML hemangioblast up-regulated TGF-ß1 and result in activating matrix metalloproteinase-9 to enhance s-KitL and s-ICAM-1 secretion. Further studies showed that phosphatidylinositol-3 kinase (PI3K)/Akt/nuclear factor-κB signalling pathway was involved in CML pathogenesis. These findings provide direct evidence for the first time that hemangioblasts beyond HSCs play a critical role in the progression of CML.

Matrix metalloproteinase-9 was involved in the immuno-modulatory defect of mesenchymal stem cell from chronic myeloid leukemia patients.

BACKGROUND: Overwhelming evidences on chronic myeloid leukemia (CML) indicate that patients harbor quiescent CML stem cells that are responsible for blast crisis. While the hematopoietic stem cell (HSC) origin of CML was first suggested over 30 years ago, recently CML-initiating cells beyond HSCs are also being investigated. METHODS: We have previously isolated fetal liver kinase-1-positive (Flk1(+)) cells carrying the BCR/ABL fusion gene from the bone marrow of Ph(+) patients with hemangioblast property. In this study, we isolated CML patient-derived Flk1(+)CD31(-)CD34(-) mesenchymal stem cells (MSCs) and detected their biological characteristics and immunological regulation using fluorescence in situ hybridization (FISH) analysis, fluorescence activated cell sorting (FACS), enzyme-linked immunoadsorbent assay, mixed lymphocyte reaction assays; then we compared these characters with those of the healthy donors. RESULTS: CML patient-derived Flk1(+)CD31(-)CD34(-) MSCs had normal morphology, phenotype and karyotype while appeared impaired in immuno-modulatory function. The capacity of patient Flk1(+)CD31(-)CD34(-) MSCs to inhibit T lymphocyte activation and proliferation was impaired in vitro. CONCLUSIONS: CML patient-derived MSCs have impaired immuno-modulatory functions, suggesting that the dysregulation of hematopoiesis and immune response may originate from MSCs rather than hematopoietic stem cells (HSCs). MSCs might be a potential target for developing efficacious treatment for CML.

Human adipose tissue-derived mesenchymal stem cells facilitate the immunosuppressive effect of cyclosporin A on T lymphocytes through Jagged-1-mediated inhibition of NF-κB signaling.

OBJECTIVE: Cyclosporine A (CsA), known as an effective immunosuppressive agent, is widely used in clinical fields. Mesenchymal stem cells may exert immunomodulatory effects on the immune system, but the exact mechanisms underlying them remain controversial. Here we investigated whether human adipose tissue-derived mesenchymal stem cells (AMSCs) facilitate in vitro the immunomodulatory effects of CsA and we explored the molecule mechanisms that may be involved. MATERIALS AND METHODS: Proliferation of T lymphocytes was measured by uptake of (3)H-thymidine. Transcription and production of interleukin-2 and interferon-γ were evaluated by real-time quantitative polymerase chain reaction, reverse transcription polymerase chain reaction, and enzyme-linked immunosorbent assay. Nuclear factor-κB (NF-κB) was assayed by Western blotting and electrophoretic mobility shift assay. Expression of Jagged-1, Jagged-2, and Delta-1 of AMSCs were surveyed by flow cytometric analysis and Western blotting. RESULTS: The combination of moderate-dose AMSCs and low-dose CsA was significantly more powerful than moderate-dose AMSCs or large-dose CsA alone in suppressing transcription and production of interleukin-2 and interferon-γ, activation of NF-κB, and proliferation of T lymphocytes. In addition, AMSCs expressed a high level of Jagged-1, which induced activation of Notch signaling in T lymphocytes, thus reducing NF-κB activity. Anti-Jagged-1 neutralizing antibody and N [N-(3, 5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester could reverse this trend. CONCLUSIONS: Human AMSCs facilitate the immunosuppressive effect of CsA on T lymphocytes through Jagged-1/Notch-related inhibition of NF-κB signaling. The combination of AMSCs and CsA represents a rationale therapeutic approach aimed to prevent adverse effects of CsA while maintaining its adequate immunosuppressive effect. Expression of Jagged-1 on AMSCs may provide an effective mechanism for the immunomodulatory activity of AMSCs via direct cell-cell interaction.