Bone marrow adipocytes is a new player in supporting myeloma cells proliferation and survival in myeloma microenvironment.

Multiple myeloma (MM) is a lethal B cell neoplasm characterized by clonal expansion of malignant plasma cells in the bone marrow and remains incurable due to disease relapse and drug resistance. Bone marrow adipocytes (BMAs) are emerging as playing active functions that can support myeloma cell growth and survival. The aim of this study is to investigate myeloma-mesenchymal stem cells (MSCs) interaction and the impact of such interactions on the pathogenesis of MM using in vitro co-culture assay. Here we provide evidence that MM cell up-regulated MSCs to express PPAR-γ and pushes MSCs differentiation toward adipocytes at the expense of osteoblasts in co-culture manner. The increased BMAs can effectively enhance MM cell to proliferation, migration, and chemoresistance via cell-cell contact and/or cytokines release regulated by PPAR-γ signal pathway. This effect was partially reversed in medium containing PPAR-γ antagonist G3335 and indicated that G3335 distorts the maturation of MSC-derived adipocytes and cytokines release by adipocytes through inhibition of PPAR-γ, a key transcriptional factor for the activation of adipogenesis, or cell to cell contact, or both. In meantime, we observed higher expression of adipocyte differentiation associated genes DLK1, DGAT1, FABP4, and FASN both in MSCs and MSC derived adipocytes, but the osteoblast differentiation-associated gene ALP was down regulated in MSCs. These finding mean that direct consequence of MM/MSC interaction that play a role in MM pathogenesis. Consistent with those in vitro results, our primary clinical observation also showed that bone marrow samples from MM patients had significantly higher bone adiposity in comparison with controls and the number of adipocytes decreased in those who were response to anti-MM therapy. Our finding suggested that BMAs may have an important contribution to MM progression, particularly in drugs resistant of MM cells, and plays an important contribution in MM bone disease and treatment failure, but more clinical studies are needed to confirm its role.

[Bone Marrow Adipocytes Promote the Survival of Multiple Myeloma Cells and Up-Regulate Their Chemoresistance].

OBJECTIVE: To investigate the effect of adipocytes in the bone marrow microenvironment of patients with multiple myeloma (MM) on the pathogenesis of MM. METHODS: Bone marrow adipocytes (BMA) in bone marrow smears of health donors (HD) and newly diagnosed MM (ND-MM) patients were evaluated with oil red O staining. The mesenchymal stem cells (MSC) from HD and ND-MM patients were isolated, and in vitro co-culture assay was used to explore the effects of MM cells on the adipogenic differentiation of MSC and the role of BMA in the survival and drug resistance of MM cells. The expression of adipogenic/osteogenic differentiation-related genes PPAR-γ, DLK1, DGAT1, FABP4, FASN and ALP both in MSC and MSC-derived adipocytes was determined with real-time quantitative PCR. The Western blot was employed to detect the expression levels of IL-6, IL-10, SDF-1α, TNF-α and IGF-1 in the supernatant with or without PPAR-γ inhibitor. RESULTS: The results of oil red O staining of bone marrow smears showed that BMA increased significantly in patients of ND-MM compared with the normal control group, and the BMA content was related to the disease status. The content of BMA decreased in the patients with effective chemotherapy. MM cells up-regulated the expression of MSC adipogenic differentiation-related genes PPAR-γ, DLK1, DGAT1, FABP4 and FASN, but the expression of osteogenic differentiation-related gene ALP was significantly down-regulated. This means that the direct consequence of the interaction between MM cells and MSC in the bone marrow microenvironment is to promote the differentiation of MSC into adipocytes at the expense of osteoblasts, and the cytokines detected in supernatant changed. PPAR-γ inhibitor G3335 could partially reverse the release of cytokines by BMA. Those results confirmed that BMA regulated the release of cytokines via PPAR-γ signal, and PPAR-γ inhibitor G3335 could distort PPAR-γ mediated BMA maturation and cytokines release. The increased BMA and related cytokines effectively promoted the proliferation, migration and drug resistance of MM cells. CONCLUSION: The BMA and its associated cytokines are the promoting factors in the survival, proliferation and migration of MM cells. BMA can protect MM cells from drug-induced apoptosis and plays an important role in MM treatment failure and disease progression.

Renal venous sampling assisted the diagnosis of juxtaglomerular cell tumor: a case report and literature review.

Juxtaglomerular cell tumor (JCT) is an endocrine tumor marked by elevated renin levels and high blood pressure. This case report presents the clinical findings of a 47-year-old woman with a history of recurrent hypokalemia, headaches, hypertension, and increased plasma renin activity (PRA). Dynamic enhanced magnetic resonance imaging (MRI) revealed a small nodule on the upper part of the right kidney. Selective renal venous sampling indicated a higher PRA only in the right upper pole renal vein. The patient underwent surgical removal of the right kidney mass, and the pathology results confirmed the diagnosis of JCT. This case underscores the importance of conducting selective renal venous sampling for accurate JCT diagnosis.

EZH2 inhibitor DZNep blocks cell proliferation of GCB-DLBCL cells by upregulating p16.

Diffuse large B-cell lymphomas (DLBCLs) are phenotypically and genetically heterogeneous. Two main subgroups of DLBCL include germinal center B-cell-like (GCB) and activated B-cell-like (ABC). Molecular profiling can further classify DLBCL into four subtypes: MCD (both CD79B and MYD88 L265P), BN2 (NOTCH2 mutation or BCL6 fusion), N1 (NOTCH1 mutation), or EZB (EZH2 mutation or BCL2 fusion). EZH2 inhibitors were recommended for patients with the EZB subtype of DLBCLs; however, little is known about the therapeutic mechanisms. Our results showed that DZNep arrested G1/S phase of GCB-DLBCL cells and inhibited the cell proliferation in vitro through upregulation of p16 by demethylating its promoter. These results suggest that DZNep may have potential as a novel therapeutic agent for DFLBL therapy. This agent may serve as a novel molecular agent to be applied to GCB DLBCL.

[Effect of the Connexin 43 Coupling to the Biobehavior of Multiple Myeloma Cells].

OBJECTIVE: To investigate the effect of gap junction intercellular communication (GJIC) combined by connexin43 (Cx43) and its signal to the biobehavior of multiple myeloma (MM) cells, and its possible mechanism. METHODS: The mesenchymal stem cell (MSC) cells were isolated and cultured from patients with MM and normal donors. The expression of connexin43 (Cx43) in MSC cells from different sources was detected by RT-PCR and Western blot. The side population (SP) cells were sorted by flow cytometry (FCM). The effect of MSC cells from different sources to the cell cycle, Cx43 expression, colony formation in vitro, stem cell related genes expression, cytokines secretion and chemoresistance in MM SP cells as well as with or without Cx43 inhibitor 18α-glycyrrhetinic acid (18α-GA) was observed. RESULTS: There was no significantly difference between the MSC isolated from normal donor and MM patients. Western blot showed that Cx43 expression in SP cells was up-regulated when the cells were incubated with MSC, and medium containing 18α-GA could partially inhibit it, moreover, it was more significant in MSC cells of MM patients. The ability of colony formation of SP cells in vitro was higher than those of MM cells and MM-MSC could promote the colony formation in a co-culture manner. The effect of MM-MSC to SP cells was down-regulated after 18α-GA was added. RT-PCR showed that there was several important stem cell-related genes including c-myc, Oct-4 Klf-4, and Sox-2 were found in RPMI 8226 cells, but those cells were up-regulated in SP cells (P<0.001). Meanwhile, MM-MSC could up-regulate the expression of c-myc, Klf-4 and Sox-2 (P<0.001), but down-regulate Oct-4 gene in the SP cells. The expression of those genes decreased after 18α-GA was added, but showed no significant difference (P>0.05). Cytometry bead array assays showed that MM-MSCs could secrete high level of IL-6, but the levels of IL-6, IL-10 and TGF-ß increased significantly when the MM-MSCs were co-cultured with SP cells (P<0.05), especially the levels of IL-6 and IL-10 were significantly higher than cultured alone. There was no significant change in the levels of bFGF and IL-17 before and after co-cultured. The levels of IL-6, IL-10 and TGF-ß in supernatant decreased significantly after GJ inhibitor 18α-GA was added. PI/Annexin V assay showed that MM cells were sensitive to bortezomib (BTZ)-induced apoptosis, but the sensitivity for SP cells was weaker. The ratio of cell apoptosis was 75.2%±0.77% and 8.12%±0.86% (P<0.001), respectively. MM-MSC could down-regulate the cell apoptosis induced by BTZ, while the sensitivity of MM cells to BTZ could be partially recovered after GJ inhibitor was added. CONCLUSION: MSC derived from MM patients can enhance GJIC to maintain its "hematopoiesis" by up-regulating the expression of Cx43 in MM cells, and at the same time promote cell proliferation and drug recistance by secreting multiple cytokines, which finally contributes to the relapse of MM.

Simultaneous removal of fluoride, manganese and iron by manganese oxide supported activated alumina: characterization and optimization via response surface methodology.

Fluoride, iron and manganese simultaneous exceedance of standard can be observed in groundwater in northeastern China. This work aims to apply a highly efficient method combining adsorption and oxidation for the synchronous removal of the inorganic ions. An innovative adsorbent (manganese-supported activated alumina) was synthesized by the impregnation method and showed a significant adsorption capacity better than that of fresh activated alumina. The characterization (scanning electron microscope; Brunauer, Emmett and Teller; X-ray diffraction and Fourier transform infrared spectroscopy) results verified the successful introduction of MnOOH and MnO2, and the improvement of surface microstructure enhanced the removal ability. The effect of single factors, such as pH value, reaction time or dosage on the removal performance has been verified. The maximum removal efficiencies of fluoride, iron and manganese were optimized via Response surface methodology considering the independent factors in the range of MO@AA dosage (5-9 g/L), pH (4-6) and contact time (4-12 h). Noted that compared with control, MO@AA exhibited 59.4% of improved fluoride performance. At pH of 5.79, contacting time of 12 h and 8.21 g/L of MO@AA, fluoride, iron and manganese removal were found to be 91, 100 and 23%, respectively. Herein, MO@AA was distinguished as good applicability for the treatment of fluoride-, iron- and manganese-containing groundwater.

Phase 1 studies comparing safety, tolerability, pharmacokinetics and pharmacodynamics of HLX01 (a rituximab biosimilar) to reference rituximab in Chinese patients with CD20-positive B-cell lymphoma.

OBJECTIVE: This study aimed to compare the pharmacokinetic, pharmacodynamic and safety profiles of HLX01 (a rituximab biosimilar) and reference rituximab sourced from China (MabThera®; rituximab-CN). METHODS: Here we report the results of two phase 1 studies. In the phase 1a, open-label, dose-escalation study (NCT03218072, CTR20140400), eligible patients received 250, 375 and 500 mg/m2 HLX01 sequentially at 7-day intervals, after confirming no dose-limiting toxicity (DLT). In the phase 1b, double-blind study (NCT02584920, CTR20140764), eligible patients were given a single dose of 375 mg/m2 HLX01 or rituximab-CN. The primary endpoints included safety and tolerability parameters for the phase 1a and the area under the plasma concentration-time curve from time zero to day 91 (AUC0-91 d) for the phase 1b study. Equivalence was concluded if 90% confidence interval (90% CI) for the geometric least squares mean ratio (GLSMR) fell in the pre-specified equivalence criteria (80%-125%). RESULTS: Between June 20, 2014 and January 5, 2015, 12 patients were enrolled in the phase 1a study. The pharmacokinetics of HLX01 showed dose proportionality and accumulation to steady state. HLX01 was well tolerated, with no serious adverse events (AEs), discontinuations or DLTs. Between November 8, 2014 and August 13, 2015, 87 eligible patients were enrolled in the phase 1b study, including 43 who received HLX01 and 44 who were treated with rituximab-CN. The equivalence endpoint was met with GLSMR for AUC0-91 d being 89.6% (90% CI: 80.4%-99.8%). AEs, anti-drug antibodies, and CD19+ and CD20+ B lymphocyte counts were similar between the HLX01 and rituximab-CN treatment groups. CONCLUSIONS: Treatment with HLX01 was safe and well tolerated in Chinese patients with B-cell lymphoma. HLX01 and rituximab-CN have similar pharmacokinetic, pharmacodynamic and safety profiles.

Anti-TWEAK Antibody Alleviates Renal Interstitial Fibrosis by Increasing PGC-1α Expression in Lupus Nephritis.

PURPOSE: Current studies on the mechanism of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) in lupus nephritis (LN) mainly focus on the inflammatory pathway. Herein, we aimed to determine whether TWEAK could promote the progression of renal interstitial fibrosis by regulating peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α) expression and intervening in lipid metabolism in LN. MATERIALS AND METHODS: MRL/lpr mice, an animal model of lupus, were treated with the anti-TWEAK antibody or co-treated with adeno-associated virus-mediated PGC-1α short hairpin RNA (shRNA). In addition, human proximal tubular epithelial cells (HK2 cells) were treated with recombinant human TWEAK (rhTWEAK) or ammonium pyrrolidine dithiocarbamate (PDTC) in vitro. RESULTS: The renal contents of free fatty acids and triglycerides were higher in MRL/lpr mice than in MRL/MpJ mice; however, these contents were decreased by treatment with the anti-TWEAK antibody. Based on immunofluorescence staining, the expression of PGC-1α was markedly more in the renal tubules of MRL/MpJ mice than in the glomeruli. However, treatment with anti-TWEAK antibody increased the levels of PGC-1α and its downstream target genes, which were remarkably lower in MRL/lpr mice than in MRL/MpJ mice. Anti-TWEAK antibody effectively eased renal interstitial fibrosis, which manifested as a decrease in the deposition of collagen fibers and the inhibition of type I collagen and fibronectin expression. However, the therapeutic effects of the anti-TWEAK antibody were abolished by PGC-1α shRNA. Treatment with rhTWEAK decreased PGC-1α expression in both dose- and time-dependent manners in HK2 cells in vitro. PDTC, an inhibitor of IκBα phosphorylation, suppressed the decrease in the PGC-1α protein level induced by rhTWEAK treatment. CONCLUSION: Our results suggest that TWEAK prevents renal tubular PGC-1α expression by promoting NF-κB activation, resulting in a deficiency in lipid metabolism and the progress of renal interstitial fibrosis. The upregulation of renal tubular PGC-1α expression to improve lipid metabolism is one of the mechanisms employed by the anti-TWEAK antibody to treat renal interstitial fibrosis.

Exosomal miR-1910-3p promotes proliferation, metastasis, and autophagy of breast cancer cells by targeting MTMR3 and activating the NF-κB signaling pathway.

Exosomes are key mediators of intercellular communication and play a role in the pathogenesis and progression of cancer. Exosomes in circulating body fluids serve as molecular markers for cancer diagnosis. This study aimed to investigate the role of exosomal microRNA (miR)-1910-3p in breast cancer and determine its clinical diagnostic value. MiR-1910-3p promoted proliferation and migration of breast cancer cells in vitro and in vivo. In vitro, exosomes enriched in miR-1910-3p transferred miR-1910-3p to mammary epithelial cells and breast cancer cells, promoting proliferation and migration, inhibiting apoptosis, and inducing autophagy. In vivo, exosomes enriched in miR-1910-3p promoted the proliferation and migration of breast cancer cells. MiR-1910-3p downregulated myotubularin-related protein 3, activated the NF-κB and wnt/ß-catenin signaling pathway, and promoted breast cancer progression. Serum miR-1910-3p in exosomes was an effective diagnostic marker that improved the sensitivity of breast cancer diagnosis when used in combination with the traditional tumor marker CA153. In conclusion, breast cancer cell-derived exosomes promoted the growth, metastasis, and autophagy of breast cancer cells by transferring miR-1910-3p. MiR-1910-3p in serum exosomes may serve as a novel molecular marker for breast cancer diagnosis.

Inverse Relationship between Plasma Tumor Necrosis Factor-Like Weak Inducer of Apoptosis and Carotid Intima-Media Thickness among Patients Undergoing Hemodialysis and Peritoneal Dialysis.

BACKGROUND/AIMS: This study aimed to investigate the level of soluble tumor necrosis factor-like weak inducer of apoptosis (sTWEAK) and its correlation with micro-inflammation and atherosclerosis in continuous ambulatory peritoneal dialysis (PD) patients. METHODS: This retrospective study involved 23 healthy subjects (control group), 23 hemodialysis (HD) patients (HD group) and 26 PD patients (PD group). Serum biochemical measurements and sTWEAK assessments were tested. The association between intima-media thickness (IMT) and sTWEAK concentrations was evaluated. RESULTS: The TWEAK level was lower in PD (155.16 ± 3.69 pg/mL, p < 0.001) and the HD group (150.16 ± 7.23 pg/mL, p < 0.001) than that in the control group (193.05 ± 5.36 pg/mL), with no significant difference between the PD group and the HD group. In the PD and HD groups, sTWEAK was significant negatively correlated with CPR, fibrinogen, and white blood cell (p < 0.05). Besides, compared to lower sTWEAK concentration end-stage renal disease (ESRD) patients (no >161.9 pg/mL), patients who had a higher level of sTWEAK (>161.9 pg/mL) had a lower IMT (0.97 ± 0.04 vs. 0.84 ± 0.03 cm, p = 0.029). After adjusted for sex, age, hypertension, diabetes, duration of dialysis, triglyceride, total cholesterol, low-density lipoprotein, and serum glucose, sTWEAK (B = -0.002, r = 0.015) and CRP (B = 0.022, r = 0.015) were independent risk factors for the IMT of ESRD patients. CONCLUSION: Plasma TWEAK is inversely associated with carotid IMT among patients undergoing HD and PD.

Shenqi Fuzheng Injection (SFI) Enhances IFN-α Inhibitory Effect on Hepatocellular Carcinoma Cells by Reducing VEGF Expression: Validation by Gene Silencing Technique.

Shenqi Fuzheng Injection (SFI) is a traditional Chinese medicine injection with anticancer properties and is mainly composed of ginseng and astragalus. Its efficacy has been confirmed in clinical trials, but the mechanism remains unclear. We investigated the effect of SFI on vascular endothelial growth factor (VEGF) gene expression in hepatocellular carcinoma (HCC) cells and identified its possible mechanism of synergistic effects when combined with the chemotherapeutic drug interferon (IFN-) α. An MTT assay was used to measure the inhibition effects of low-dose IFN-α (6000 IU) with or without SFI (0.5 g/L) on the HCC cell line MHCC97. VEGF-silenced MHCC97L-mir200 cell lines were prepared using lentiviral vectors and evaluated by real-time PCR to determine the inhibition effect. We examined MHCC97L-mir200 and MHCC97L cells by MTT assay, using IFN-α alone or in combination with SFI. The inhibition ratio of IFN-α (6000 IU) was -29.5%, while that for IFN-α (6000 IU) + SFI (0.5 g/L) was 17.0%, which was significantly higher than that for the IFN-α group (P < 0.01). The VEGF gene was silenced successfully in MHCC97-L cells. After interference of VEGF, the inhibition by SFI and IFN-α in MHCC97L-mir200 did not differ from that in MHCC97-L cells (P > 0.05). SFI can reduce the expression of VEGF in HCC, which can increase the efficacy of IFN-α, providing a theoretical basis for clinical application.

Impact of cell fusion in myeloma marrow microenvironment on tumor progression.

BACKGROUND: Mesenchymal stem cells (MSCs) represent a subset of non-hematopoietic adult stem cells, which can also fuse with other cells spontaneously in bone marrow and capable of adopting the phenotype of other cells. The fusion of somatic cells with stem cells can reprogram somatic cells to a pluripotent state. Our research on the fusion of bone marrow mesenchymal stem cells(BM-MSCs) and MM cells demonstrate that the fused cells can exhibit stemness and cancer cell-like characteristics. RESULTS: We successfully produced a hybrid cells that acquired larger size and multinucleation, in which partial chromatin condensation, a visible nucleolus, and one or more round or oval nucleus. Experiments results showed that the stemness markers highly expressed in these fused cells and there were much more chromosomes in fused cells than those in parental cells as well as exhibited increased resistance to drug treatment. CONCLUSIONS: Our results suggest that cell fusion between BM-MSCs and MM cells could contribute it genomic heterogeneity and play a role on disease progression. METHODS: We fused human BM-MSCs with MM cells lines RPMI 8226 or XG1 in vitro by polyethylene glycol (PEG), and the hybrid cells were sorted by sedimentation assays. The growth, migration, cell cycle, chromosome and drug sensitive of hybrids were assessed by cell counting, cell colony formation, transwell assays, cytogenetic assay and flow cytometry (FCM). The proteins and genes related to stemness and cytokines were tested by western blot and/or real-time quantitative RT-PCR.

Efficacy of long-term nonsteroidal antiinflammatory drug treatment on magnetic resonance imaging-determined bone marrow oedema in early, active axial spondyloarthritis patients.

To assess the efficacy of long-term treatment with nonsteroidal antiinflammatory drugs (NSAIDs) on bone marrow oedema (BMO) of the sacroiliac joint in newly diagnosed axial spondyloarthritis (SpA) with a symptom duration of less than 4 years, a single-center, open-label study in a cohort of consecutive patients with newly diagnosed axial SpA was conducted. Eligible patients had magnetic resonance imaging (MRI)-determined BMO of the sacroiliac joint at baseline, had a symptom duration of less than 4 years, and were naïve to NSAIDs. After the baseline MRI, an optimal dose of NSAID was administered for 24 or 48 weeks. BMO of sacroiliac joint was quantified by applying the Spondyloarthritis Research Consortium of Canada (SPARCC) system. Disease activity was expressed using the Ankylosing Spondylitis Disease Activity Score (ASDAS). Primary end points were improvement in BMO of sacroiliac joint at week 24 or week 48. Forty-three patients were recruited, and 33 patients eventually completed the study, including 10 patients having follow-up MRI at week 24 and 23 patients having follow-up MRI at week 48. Overall, the mean of SPARCC score decreased from 21.8 ± 16.1 at baseline to 10.2 ± 12.8 at follow-up (p < 0.001). 75.8% of the patients displayed a minimally important change, and 30.3% became free of BMO. The mean of ASDAS-CRP decreased from 3.1 ± 1.0 at baseline to 2.1 ± 1.0 at follow-up (p < 0.001). Long-term treatment with optimal dose NSAIDs could significantly alleviate BMO of sacroiliac joint in early and active axial SpA.

Impact of connexin 43 coupling on survival and migration of multiple myeloma cells.

INTRODUCTION: Gap junctions (GJs) represent the best known intercellular communication (IC) system and are membrane-spanning channels that facilitate intercellular communication by allowing small signaling molecules to pass from cell to cell. In this study, we constructed an amino terminus of human Cx43 (Cx43NT-GFP), verified the overexpression of Cx43-NT in HUVEC cells and explored the impact of gap junctions (GJs) on multiple myeloma (MM). MATERIAL AND METHODS: The levels of phosphorylated Cx43(s368) and the change of MAPK pathway associated molecules (ERK1/2, JNK, p38, NFκB) were also investigated in our cell models. Cx43 mRNA and proteins were detected in both MM cell lines and mesenchymal stem cells (MSCs). Dye transfer assays demonstrated that gap junction intercellular communication (GJIC) occurring via Cx43 situated between MM and MSCs or MM and HUVECCx43NT is functional. RESULTS: Our results present evidence for a channel-dependent modulator action of connexin 43 on the migratory activity of MM cells toward MSCs or HUVECCx43-N was higher than those of spontaneous migration (p < 0.05) and protection them from apoptosis in the presence of dexamethasone via cytokines secretion. In the meantime, the migration of MM cells involves an augmented response of p38 and JNK signaling pathway of carboxyl tail of the protein. CONCLUSIONS: Our data suggest that GJIC between MM and MSCs is one of the essential factors in tumor cell proliferation and drug sensitivity, and is implicated in MM pathogenesis.

MiR-155 promotes cell proliferation and inhibits apoptosis by PTEN signaling pathway in the psoriasis.

MicroRNAs (miRNAs) have been demonstrated to contribute to malignant progression in psoriasis development. The purposes of the study was to evaluated the effects of miRNA-155 on cell proliferation, migration and apoptosis in psoriasis development via PTEN singaling pathway and identify its direct target protein. Quantitative real-time RT-PCR (qRT-PCR) was performed to examine the level of miR-155 in psoriasis cells, miR-155 was downregulated in a psoriasis cell line Hacat by transfected with small interfering RNA (siRNA), respectively. Cell survival was detected by the MTT assay and colony formation assay. Cell migration and invasion were measured via wound-healing assayand transwell assay. In addition, cell cycle and apoptosis about psoriasis cells was measured by flow cytometry. In this study, qRT-PCR assay showed that the expressions of miR-155 mRNA in psoriasis tissues were significantly higher than that in normal tissues. The assays about cell growth and proliferation showed that miR-155 knockdown led to a significant decrease in cell proliferation which was determined by MTT assay and colony formation assay compared to those of Lv-NC cells. Flow cytometry analysis showed that depletion of miR-155 could cause cell cycle change and the number of apoptotic cells was significantly increased in Lv-miR155 cells compared with control cells. In addition, the expression of several apoptosis-related factors were dramatically changed, such as PTEN, PIP3, AKT, p-AKT, Bax and Bcl-2. Our findings indicate that down-regulation of miR-155 significantly inhibits proliferation, migration, invasion and promotes apoptosis through PTEN singaling pathway in psoriasis cells. miR-155 might function as an oncogene miRNA in the progress of psoriasis.

The role of follicular T helper cells in patients with malignant lymphoid disease.

OBJECTIVES: To investigate the dynamic change of follicular T helper cells (TFH) in patients with malignant lymphoid disease (MLD) and to explore its clinical significance. METHODS: The dynamic change of TFH cells, ICOS+- and PD-1+ TFH cells at pretreatment and different treatment periods was determined by flow cytometry in 85 MLD patients. Concentration of interleukin 21 (IL-21) was evaluated by ELISA, and the correlation between clinical prognosis and the ratio of TFH cells was analyzed. RESULTS: Significantly increased ICOS+- and PD-1+ TFH cells were found in MLD patients at pretreatment compared to healthy controls. Decreased or even close to normal levels of ICOS+- and PD-1+ TFH cells were found at the end of treatment. However, in the patients with progressive disease, high levels of ICOS+- and PD-1+ TFH cells were found. Moreover, a significantly increased plasma IL-21 level was found in MLD patients. Negative correlation was found between the level of ICOS+-, PD-1+ TFH cells, as well as IL-21 and the prognosis of MLD. CONCLUSIONS: Significantly increased TFH cell ratios were found in patients with MLD, and decreased TFH cells ratios could be expected in those treatment-effective patients, which could be used as the therapeutic efficacy index.

Cx43 expressed on bone marrow stromal cells plays an essential role in multiple myeloma cell survival and drug resistance.

INTRODUCTION: Connexin-43 (Cx43), a connexin constituent of gap junctions (GJs) is mainly expressed in bone marrow stromal cells (BMSCs) and played a important role on hematopoiesis. In this study, we explored the role of gap junctions (GJs) formed by Cx43 between BMSCs and multiple myeloma (MM) cells. MATERIAL AND METHODS: qPCR and western blot assays were employed to assay Cx43 expression in three MM cell lines (RPMI 8266, U266, and XG7), freshly isolated MM cells, and bone marrow stromal cells (BMSCs). Cx43 mRNA and proteins were detected in all three MM cell lines and six out of seven freshly isolated MM cells. RESUTHS: The BMSCs from MM patients expressed Cx43 at higher levels than of normal donor (ND-BMSCs). Dye transfer assays demonstrated that gap junction intercellular communication (GJIC) occurring via Cx43 situated between MM and BMSCs is functional. Cytometry beads array (CBA) assays showed that cytokines production changed when the ND-BMSCs were co-cultured with MM cells, especially the levels of IL-6, SDF-1α and IL-10 were higher than those the cells cultured alone and decreased significantly in the presence of GJ inhibitor heptanol. Our results demonstrated that the cytotoxicity of BTZ to MM cells decreased significantly in the presence of BMSCs, an effect that was partially recovered in the presence of GJ inhibitor. CONCLUSIONS: Our data suggest that GJIC between MM and BMSCs is a critical factor in tumor cell proliferation and drug sensitivity, and is implicated in MM pathogenesis.

Increased autophagy in fibroblast-like synoviocytes leads to immune enhancement potential in rheumatoid arthritis.

The incidence of rheumatoid arthritis (RA) has been reported to be correlated with a disorder of immunregulation. Rheumatoid arthritis fibroblast-like synoviocytes (RA-FLSs) play an important role in regulating the local immune microenvironment. However, the potential mechanism of RA-FLS in regulating the immnue response is not clearly understood. In this study, we demonstrated that the expression of HIF-1α was significantly up-regulated in rheumatoid arthritis tissue which indicated that the hypoxia condition in the microenvironment. We also observed that RA-FLSs demonstrated the potential to up-regulate immune activation. Meanwhile, the level of autophagy increased in RA-FLSs compared with control group. Besides that, the expression of IL-6 was up-regulated not only in RA-FLSs but also in the fibroblasts that treated with hypoxia condition. Accordingly, we found that autophagy inhibitiors could effectively inhibit the immune activation function of RA-FLSs medicated by IL-6. Taken together, the results we demonstrated above indicated that the hypoxia microenvironment could effectively induce the incidence of autophagy and then lead to the immune activation function of RA-FLSs medicated by IL-6.

Rs56288038 (C/G) in 3'UTR of IRF-1 Regulated by MiR-502-5p Promotes Gastric Cancer Development.

BACKGROUND/AIMS: Interferon regulatory factor 1 (IRF-1) has been shown to function as a transcriptional activator or repressor of a variety of target genes. However, its upstream, non-coding RNA-related regulatory capacity remains unknown. In this study, we focus on the miRNA-associated single nucleotide polymorphisms (SNPs) in the 3'untranslated region (UTR) of IRF-1 to further investigate the functional relationship and potential diagnostic value of the SNPs and miRNAs among Chinese gastric cancer (GC) patients. METHODS: We performed a case-control study with 819 GC patients and 756 cancer-free controls. Genotyping by realtime PCR assay, cell transfection, and the dual luciferase reporter assay were used in our study, and the 5-year overall survival rate and relapse-free survival rate in different groups were investigated. RESULTS: We found that patients suffering from Helicobacter pylori (Hp) infection were the susceptible population compared to controls. SNP rs56288038 (C/G) in IRF-1 3'UTR was involved in the occurrence of GC by acting as a tumor promoter factor. SNP rs56288038 (C/G) could be up-regulated by miR-502-5p, which caused a down-regulation of IRF-1 in cell lines and decreased apoptosis induced by IFN-γ. Carrying the G genotype was related to significantly low expression of IRF-1 and Hp infection, poor differentiation, big tumor size, invasion depth, as well as the high probability of metastasis, and moreover, the C/G SNP was associated with shorter survival of GC patients with five years of follow-up study. CONCLUSIONS: our findings have shown that the SNP rs56288038 (C/G) in IRF-1 3'UTR acted as a promotion factor in GC development through enhancing the regulatory role of miR-502-5p in IRF-1 expression.

[Effects of Bone Marrow Mesenchymal Stem Cells on the Biological Characteristics of Migrating and Homing in Multiple Myeloma Cells].

OBJECTIVE: To construct a co-culture system for bone marrow mesenchymal stem cells (BMMSC) and multiple myeloma (MM) cells, and to investigate the effects of co-cultured BMMSC on the migrating and homing of multiple myeloma cells. METHODS: The BMMSC from the transgenic mice with green fluorescent protein (GFP) fetal bone were cultured by adherent screening. A co-culture system of BMMSC and MM cell line XG-7 cells was constracted, the proliferation and apoptosis of cells were determined by trypan blue exclusion and Annexin V/PI, respectively, MDC staining was employed to detect the autophagy. The moving direction distribution of molecule in BMMSC and XG-7 cells labeled with PE-CD138 in co-culture process were observed dinamically by confocal microscopy. RESULTS: After co-culture with GFP-BMMSC, the resistance of XG-7 cells to apoptosis and autophagy were enhanced; at the same time, their proliferation increased. Apoptosis rates of XG-7 cells directly and indirectly co-cultured with BMMSC were (6.23 ± 0.12)% and (6.97 ± 0.03)% respectively, which were lower than that of XG-7 cells cultured alone (17.90 ± 1.46)% (P < 0. 01). There was low level of autophagy in XG-7 cells co-cultured with BMMSC. XG-7 cells are highly polarized and contained a specialized membrane domain with specific protein and lipid components to contact with BMMSC under confocal microscope. After methyl-ß-cyclodextrin treatment, the molecules were normally enriched in the specialized domain. CONCLUSION: BMMSC can protect XG-7 cells from apoptosis and autophagy, and obviously promote the proliferation of XG-7 cells, and can influence the migrating and homing of multiple myeloma cells.