Induced pluripotent stem cells (iPSCs) can be propagated indefinitely, while maintaining the capacity to differentiate into all cell types in the body except for the extra-embryonic tissues. This iPSC technology not only represents a new way to use individual-specific stem cells for regenerative medicine but also constitutes a novel method to obtain large numbers of disease-specific cells for biomedical research. However, the low efficiency of reprogramming and genomic integration of oncogenes and viral vectors limit the potential application of iPSCs. Chemical-induced reprogramming offers a novel approach to generating iPSCs. In this study, a new combination of small-molecule compounds (SMs) (sodium butyrate, A-83-01, CHIR99021, Y-27632) under conditions of transient folate deprivation was used to generate iPSC. It was found that transient folate deprivation combined with SMs was sufficient to permit reprogramming from mouse embryonic fibroblasts (MEFs) in the presence of transcription factors, Oct4 and Klf4, within 25 days, replacing Sox2 and c-Myc, and accelerated the generation of mouse iPSCs. The resulting cell lines resembled mouse embryonic stem (ES) cells with respect to proliferation rate, morphology, pluripotency-associated markers and gene expressions. Deprivation of folic acid, combined with treating MEFs with SMs, can improve the inducing efficiency of iPSCs and reduce their carcinogenicity and the use of exogenous reprogramming factors.
Autophagy dysregulation, mitochondrial dynamic abnormality and cell cycle re-entry are implicated in the vulnerable neurons of patients with Alzheimer’s disease. This study was designed to testify the association among autophagy, mitochondrial dynamics and cell cycle in dividing neuroblastoma (N2a) cells. The N2a cells were cultured in vitro and treated with different concentrations of 3-methyladenine (3-MA). The cell viability was detected by methyl thiazolyl tetrazolium (MTT) assay. They were randomly divided into control group (cells cultured in normal culture medium) and 3-MA group (cells treated with 10 mmol/L 3-MA). The cell cycle was analyzed in the two groups 3, 6, 12, and 24 h after treatment by flow cytometry. Western blotting was used to evaluate the expression levels of mitofission 1 (Fis1), mitofusin 2 (Mfn2), microtubule-associated protein 1 light chain 3 (LC3), cell cycle-dependent kinase 4 (CDK4) and cdc2. The flow cytometry revealed that the proportion of cells in G2/M was significantly increased, and that in G0/G1 was significantly reduced in the 3-MA group as compared with the control group. Western blotting showed that the expression levels of Fis1, LC3, and CDK4 were significantly up-regulated in the 3-MA group at the four indicated time points as compared with the control group. Mfn2 was initially decreased in the 3-MA group, and then significantly increased at 6 h or 12 h. Cdc2 was significantly increased in the 3-MA group at 3 h and 6 h, and then dropped significantly at 12 h and 24 h. Our data indicated that 3-MA-induced suppressed autophagy may interfere with the cell cycle progression and mitochondrial dynamics, and cause cell death. There are interactions among cell cycle, mitochondrial dynamics and autophagy in neurons.
In order to investigate the role of Toll-like receptor 4 (TLR4) in cerebrocardiac syndrome (CCS), the partial cerebral ischemia/reperfusion (I/R) models in mice with different TLR4 genotypes were established in the present study. TLR4 wild-type (C3H/HeN) and mutant (C3H/HeJ) mice of 6–8 weeks of age were divided into 4 groups at random: C3H/HeN sham group (n=10), C3H/HeJ sham group (n=10), C3H/HeN model group (n=10) and C3H/HeJ model group (n=10). Partial cerebral I/R was caused by the middle cerebral artery occlusion (MCAO) to duplicate CCS models in mice. After the operation, the electrocardiogram (ECG), the level of tumor necrosis factor-alpha (TNF-α) in myocardial tissue and the cardiac pathological changes were observed in each group. It was shown that the brain infarct volume in C3H/HeN model group was larger than that in C3H/HeJ model group (P<0.01). The ST segment change and T wave inversion occurred frequently in model groups. Moreover, the TNF-α level in C3H/HeN model group was higher than that in C3H/HeJ model group (P<0.01). The myocardial injury was aggravated in C3H/HeN group as compared with C3H/HeJ group. It was concluded that TLR4 was implicated in the development of CCS.
The study aimed to investigate the impact of intraclot recombinant tissue-type plasminogen activator (rt-PA) on perihematomal edema (PHE) development in patients with intracerebral hemorrhage (ICH) treated with minimally invasive surgery (MIS) and the effects of intraclot rt-PA on the 30-day survival. We reviewed the medical records of ICH patients undergoing MIS between October 2011 and July 2013. A volumetric analysis was done to assess the change in PHE and ICH volumes at pre-MIS (T1), post-MIS (T2) and day 10–16 (T3) following diagnostic computed tomographic scans (T0). Forty-three patients aged 52.8±11.1 years with (n=30) or without rt-PA (n=13) were enrolled from our institutional ICH database. The median rt-PA dose was 1.5 (1) mg, with a maximum dose of 4.0 mg. The ratio of clot evacuation was significantly increased by intraclot rt-PA as compared with controls (77.9%±20.4% vs. 64%±15%; P=0.046). From T1 to T2, reduction in PHE volume was strongly associated with the percentage of clot evacuation (ρ=0.34; P=0.027). In addition, PHE volume was positively correlated with residual ICH volume at the same day (ρ ranging from 0.39–0.56, P<0.01). There was no correlation between the cumulative dose of rt-PA and early (T2) PHE volume (ρ=0.24; P=0.12) or delayed (T3) PHE volume (ρ=0.19; P=0.16). The 30-day mortality was zero in this cohort. In the selected cohort of ICH patients treated with MIS, intraclot rt-PA accelerated clot removal and had no effects on PHE formation. MIS aspiration and low dose of rt-PA seemed to be feasible to reduce the 30-day mortality in patients with severe ICH. A large, randomized study addressing dose titration and long-term outcome is needed.
Aminoacyl-tRNA syntheses (AARS) can catalyze the adenosine triphosphate (ATP)-dependent acylation of their cognate tRNA(s) with a specific amino acid. They can be seen as an index to reflect the energy metabolic rate of ischemic brain cells in ischemic penumbra. This study examined the relationship between arginyl-tRNA synthetase (ArgRS), one of the AARS, and cerebral ischemia in rats. The model of middle cerebral artery occlusion (MCAO) was established in rats. The expression levels of ArgRS protein and mRNA were detected in rat brain tissues at different time points following MCAO by Western blotting and RT-PCR, respectively. The results showed that the MCAO model was successfully established. Western blotting and RT-PCR analysis revealed that the ArgRS protein and mRNA were expressed in brain cells in both ischemic and normal penumbra tissues. The expression levels of ArgRS protein and mRNA peaked at 6 h after MCAO and decreased gradually. At 24 h, the expression levels of ArgRs protein and mRNA in ischemic penumbral tissues were lower than those in normal tissues. The expression levels of ArgRS mRNA and protein in ischemic penumbra varied with ischemic time, suggesting that the energy metabolism of brain cells in penumbra changed dynamically after ischemia to ensure the endogenous self-protection of the body. The brain oxygen supply should be improved as soon as possible, especially within 6–12 h after ischemia, so as to meet the demand for energy metabolism in ischemic penumbra and make sure the cell structure remains stable.
The cytokines of acute leukemia (AL) patients have certain expression patterns, forming a complex network involved in diagnosis, progression, and prognosis. We collected the serum of different AL patients before and after complete remission (CR) for detection of cytokines by using an antibody chip. The expression patterns of cytokines were determined by using bioinformatics computational analysis. The results showed that there were significant differences in the cytokine expression patterns between AL patients and normal controls, as well as between acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). In confirmatory test, ELISA revealed the expression of uPAR in AL. Moreover, the bioinformatic analysis showed that the differentially expressed cytokines among the AL groups were involved in different biological behaviors and were closely related with the development of the disease. It was concluded that the cytokine expression pattern of AL patients is significantly different from that of healthy volunteers. Also, differences of cytokine expression patterns exist between AML and ALL, and between before and after CR in the same subtype of AL, which holds important clinical significance for revealing disease progression.
Recently, the immunotherapy has been highlighted among cancer treatments. Cancer-testis antigen (CTA) has been studied in a variety of solid tumors because of its specific expression in tumors, and testis, ovary and placenta tissues, but not in other normal tissues. In order to provide a new approach for multiple myeloma (MM) immunotherapy, we examined the CTA expression in MM cell lines, and primary myeloma cells in patients with MM. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the mRNA expression of MAGE-C1/CT7, SSX1, SSX2 and SSX4 in MM cell lines of RPMI-8226 and U266, and bone marrow (BM) cells of 25 MM patients and 18 healthy volunteers. The results showed that the 4 CTAs were expressed in RPMI-8226 and U266 cell lines. The positive expression rate of MAGE-C1/CT7, SSX1, SSX2 and SSX4 in the BM cells of 25 MM patients was 28% (7/25), 80% (20/25), 40% (10/25) and 68% (17/25), respectively. In contrast, the expression of any member of the CTAs was not detected in BM cells of 18 healthy volunteers. The expression of two or more CTAs was detected in 80% (20/25) MM patients, and that of at least one CTA in 88% (22/25). The mRNA expression levels of SSX1 and SSX4 were significantly higher in patients with MM at stage III than in those at stage I and II (P<0.05). No statistically significant differences were observed in the mRNA expression levels of MAGE-C1/CT7 and SSX2 in further stratified analyses by age, gender, MM types and percentage of MM cells in BM (P>0.05). In conclusion, our present study showed that MAGE-C1/CT7, SSX1, SSX2 and SSX4 were co-expressed in MM cell lines and the primary myeloma cells in MM patients, but not expressed in BM cells of healthy subjects. The mRNA levels of SSX1 and SSX4 are associated with MM clinical stage. This work may provide a new insight into MM immunotherapy in the future.
The intestinal absorption of berberine (Ber) and its structural modified compound 8-hydroxy dihydroberberine (Hdber) was compared, and their effects on the intestinal absorption of sugar by perfusion experiment were investigated in order to reveal the mechanism of low dose and high activity of Hdber in the treatment of hyperglycemia. The absorption of Hdber and Ber in rat small intestine was measured by in situ perfusion. High performance liquid chromatography (HPLC) was used to determine the concentrations of Hdber and Ber. In situ perfusion method was also used to study the effects of Hdber and Ber on sugar intestinal absorption. Glucose oxidase method and UV spectrophotometry were applied to examine the concentrations of glucose and sucrose in the perfusion fluid. The results showed that the absorption rate of Ber in the small intestine was lower than 10%, but that of Hdber was larger than 70%. Both Hdber and Ber inhibited the absorption of glucose and sucrose at the doses of 10 and 20 μg/mL. However, Hdber presented stronger activity than Ber (P<0.01). It is suggested that Hdber is absorbed easily in rat small intestine and that its inhibitory effect on the absorption of sugar is better than Ber.
Over-expression of Fas ligand (FasL) on tumor cell surface can induce the apoptosis of specific activated tumor infiltrating lymphocytes (TILs) via the Fas/FasL pathway, leading to the formation of a site of immune privilege surrounding the tumor mass for escaping immune surveillance and promoting tumor proliferation, invasion and metastasis. The blocking effect of miR-21 on FasL-mediated apoptosis in breast cancers was investigated in this study. The expression levels of miR-21 and FasL in human breast carcinoma cell lines were detected by using RT-PCR and Western blotting. FasL as a target gene of miR-21 was identified by Luciferase assay. The apoptosis of Jurkat T lymphocytes induced by MCF-7 cells was determined by flow cytometry. It was found that in four human breast cancer cell lines, FasL expression level in MCF-7 cells was the highest, while miR-21 was down-regulated the most notably. After miR-21 expression in MCF-7 cells was up-regulated, FasL was identified as a target gene of miR-21. When the effector/target (E/T) ratio of MCF-7 cells and Jurkat cells was 10:1, 5:1 and 1:1, the inhibitory rate of apoptosis of Jurkat T lymphocytes induced by MCF-7 cells was 95.81%, 93.16% and 91.94%, respectively. It is suggested that in breast cancers miR-21 expression is negatively associated with FasL expression, and FasL is a target gene of miR-21. miR-21 targeting and regulating FasL-mediated apoptosis will bring us the possibility of a new tumor immunotherapy via breaking tumor immune privilege.
This study examined the effect of Notch-1 signaling on malignant behaviors of breast cancer cells by regulating breast cancer stem cells (BCSCs). BCSCs were enriched by using serum-free medium and knocked out of Notch-1 by using a lentiviral vector. Real-time polymerase chain reaction (RT-PCR) and Western blotting were used to detect the Notch-1 expression levels in breast cancer cell lines and BCSCs, and flow cytometry to detect the proportion of BCSCs in BCSC spheres. The BCSC self-renewal, migration, invasion, and tumorigenicity were examined by the tumor microsphere-forming assay and transwell assay and after xenotransplantation. The results showed that the Notch-1 silencing reduced the number of BCSC spheres, the proportion of BCSCs, and the number of cells penetrating through the transwell membrane. It also decreased the size of tumors that were implanted in the nude mice. These results suggest that Notch-1 signaling is intimately linked to the behaviors of BCSCs. Blocking Notch-1 signaling can inhibit the malignant behaviors of BCSCs, which may provide a promising therapeutical approach for breast cancer.
miR-200c has been shown to regulate the epithelial-mesenchymal transition (EMT) by inhibiting ZEB1 and ZEB2 expression in breast cancer cells. This study further examined the role of miR-200c in the invasion and metastasis of breast cancer that goes beyond the regulation on ZEB1 and ZEB2 expression. In this study, the bioinformatics software (miRanda) was used to predict the target gene of miR-200c and Renilla luciferase assay to verify the result. The metastatic breast cancer cells MDA-MB-231 were cultured and transfected with the miR-200c mimic or inhibitor. The expressions of miR-200c and HMGB1 were detected by RT-PCR and Western blotting, respectively. Transwell assay and wound healing assay were employed to examine the invasive and migrating ability of transfected cells. Target prediction and Renilla luciferase analysis revealed that HMGB1 was a putative target gene of miR-200c. After transfection of MDA-MB-231 cells with the miR-200c mimic or inhibitor, the expression of miR-200c was significantly increased or decreased when compared with cells transfected with the miR-200c mimic NC or inhibitor NC. Moreover, the expression of HMGB1 was reversely correlated with that of miR-200c in transfected cells. Tranwell assay showed that the number of invasive cells was significantly reduced in miR-200c mimic group when compared with miR-200c inhibitor group. It was also found that the migrating ability of cells transfected with miR-200c mimics was much lower than that of cells transfected with miR-200c inhibitors. It was suggested that miR-200c can suppress the invasion and migration of breast cancer cells by regulating the expression of HMGB1. miR-200c and HMGB1 may become useful biomarkers for progression of breast cancer and targets of gene therapy.
Recently, microRNAs (miRNAs) have been shown to be involved in multiple biological pathways that can influence tumor progression and metastasis and they can serve as prognostic biomarkers in many cancers. The present study examined the prognostic significance of miR-215 in cervical cancer. The paraffin-embedded paired cervical scrape samples and tumor tissue samples from 302 patients with stage II cervical cancer were detected for the expression of miR-215 by using qRT-PCR. A miR-215-based classifier was established by using the Cox regression model. The prognostic and predictive accuracy of this classifier was determined in both the internal testing group of 138 patients, and the external independent group of 280 patients. Moreover, cervical cancer HeLa cells overexpressing miR-215 (HeLa-miR-215) were constructed and subcutaneously injected into the nude mice to examine the effect of miR-215 on tumor growth and metastasis in vivo. The results showed that the expression level of miR-215 was significantly higher in cervical cancer tissues than in paired normal tissues (P<0.0001). When patients were classified into high- and low-risk cancer progression groups according to miR-215 level, the 5-year disease-free survival in high- and low-risk groups were 43% (95% CI: 32.1–51.6) and 67% (95% CI: 48.6–77.3) (hazard ratio [HR] 2.02, 95% CI: 1.16–3.52; P=0.013) respectively. Moreover, the expression level of miR-215 was negatively associated with survival rate in patients at TNM stage T3 (HR: 3.317; 95% CI: 1.18–5.14, P=0.017) and TNM stage T4 (HR: 3.48; 95% CI: 1.49–4.45, P=0.008). Tumor volume in nude mice injected with HeLa-miR-215 cells was significantly larger than that in mice injected with control HeLa cells. It was concluded that the expression level of miR-215 is associated with cervical tumor progression and worse survival rate, suggesting that it may serve as a potential prognostic marker to identify patients at higher risk of recurrence.
Ovarian endometrioma is a common form of endometriosis, which may cause infertility, dysmenorrhea and pelvic pain in women of reproductive age. Although surgery is the treatment of choice for endometriomas, recurrence poses a formidable frustration. This study investigated potential risk factors of endometriomas recurrence, aiming to better understand its pathogenesis. A total of 307 patients with endometriomas were followed up for an average of 28.6 months and the 1-, 2- and 3-year cumulative recurrence rate was 9.5%, 21.9%, and 29.2%, respectively. Twenty-one potential risk factors for endometriomas recurrence were evaluated using Cox’s proportional hazards models. Total revised American Fertility Society (rAFS) score was significantly associated with higher recurrence (OR=1.858, 95% CI=1.122–3.075, P=0.016), as well as younger age at surgery (OR=0.953, 95% CI=0.915–0.992, P=0.020). Semiradical surgical treatment was defined as surgical removal of cyst plus hysterectomy with preservation of bilateral or unilateral ovary, and was a significant factor that was associated with lower recurrence than the conservative surgery (OR=0.318, 95% CI=0.107–0.951, P=0.040). Postoperative pregnancy was favorable factors for disease recurrence (OR=0.217, 95% CI=0.102–0.460, P=0.000). The results suggest that endometrioma recurrence is inversely associated with age at surgery and postoperative pregnancy, and may correlate with total rAFS score and conservative surgery method.
The purpose of this study was to determine the combined effect of transmyocardial laser revascularization (TMLR) and the implantation of endothelial progenitor cells (EPCs) on cardiac function of ischemic hearts in canines. The left anterior descending artery (LAD) was occluded to establish the canine model of acute myocardial infarct (AMI). Four weeks later, the animals were randomly divided into four groups: TMLR group, in which transmyocardial laser-induced channels were established at the ischemic region; EPCs+TMLR group, in which EPCs were locally transplanted into laser-induced channels at the ischemic region; EPCs group, in which the EPCs were injected into the ischemic region; control group, in which the AMI animals received neither TMLR nor EPCs. The peripheral blood (50 mL) was sampled in all groups. Mononuclear cells from the peripheral blood were separated and cultured to obtain spindle-shaped attaching (AT) cells in vitro. AT cells were labeled with 1, 1′-dioctadecyl-1 to 3,3, 3′,3′-tetramethyl-indocarbocyanine perchlorate (DiI) before injecting into the laser-induced channels or ischemic region. Four weeks after the first operation, TMLR was performed in the TMLR group and EPCs+TMLR group, and at the same time, the EPCs originating from the AT cells were mixed with calcium alginate (CA). Then the EPCs-CA composites were implanted into myocardial channels induced by laser in the EPCs+TMLR group, and into the myocardial infarct area in the EPCs group. All dogs underwent echocardiography at second month after LAD occlusion. Finally the samples of myocardium around the LAD were subjected to histochemical and immunohistologic examinations. The results showed there was no significant difference in the diameter of left atrium and ventricle before treatment among all groups (P>0.05). Eight weeks after modeling, the regional contractility in the LAD territory in the EPCs+TMLR group was increased as compared with control group and TMLR group, but there was no significant difference between control group and TMLR group. Neoangiogenesis was observed in the EPCs+TMLR group, and the fibrosis was seen in the TMLR group. There was no significant difference in neoangiogenesis around the channels induced by laser among EPCs+TMLR, EPCs and TMLR groups. It was concluded that TMLR combined with EPCs could improve the regional and global cardiac function in AMI, and augment neovascularizaiton in channels of ischemic myocardium induced by laser.
This study examined the expressions of miR-22 and miR-135a in rats with acute edematous pancreatitis (AEP) and their target genes in order to shed light on the involvement of miR-22 and miR-135a in the pathogenesis of acute pancreatitis (AP). The in vivo model of AEP was established by introperitoneal injection of L-arginine (150 mg/kg) in rats. The miRNA microarray analysis was used to detect the differential expression of miRNAs in pancreatic tissue in AEP and normal rats. The in vitro AEP model was established by inducing the rat pancreatic acinar cell line (AR42J) with 50 ng/mL recombinant rat TNF-α. Real-time quantitative RT-PCR was employed to detect the expression of miR-22 and miR-135a in AR42J cells. Lentiviruses carrying the miRNA mimic and anti-miRNA oligonucleotide (AMO) of miR-22 and miR-135a were transfected into the AR42J cells. The AR42J cells transfected with vehicle served as control. Western blotting was used to measure the expression of activated caspase3 and flow cytometry analysis to detect the apoptosis of AR42J cells. Targets of miR-22 and miR-135a were predicted by using TargetScan, miRanda, and TarBase. Luciferase reporter assay and quantitative real-time RT-PCR were performed to confirm whether ErbB3 and Ptk2 were the target gene of miR-22 and miR-135a, respectively. The results showed that the expression levels of miR-22 and miR-135a were obviously increased in AEP group compared with the control group in in-vivo and in-vitro models. The expression levels of miR-22 and miR-135a were elevated conspicuously and the expression levels of their target genes were reduced significantly in AR42J cells transfected with lentiviruses carrying the miRNA mimic. The apoptosis rate was much higher in the TNF-α-induced cells than in non-treated cells. The AR42J cells transfected with miRNA AMOs expressed lower level of miR-22 and miR-135a and had lower apoptosis rate, but the expression levels of ErbB3 and Ptk2 were increased obviously. It was concluded that the expression levels of miR-22 and miR-135a were elevated in AEP. Up-regulating the expression of miR-22 and miR-135a may promote the apoptosis of pancreatic acinar cells by repressing ErbB3 and Ptk2 expression in AEP.
Myeloid-derived suppressor cells (MDSCs) play a crucial role in T cell dysfunction, which is related to poor outcome in patients with severe trauma. Cyclooxygenase-2 (Cox-2) contributes to immune disorder in trauma and infection via production of prostaglandin E2. However, the role of Cox-2 in the accumulation and function of MDSCs after traumatic stress has not been fully elucidated. In the present study, we treated murine trauma model with NS398, a selective Cox-2 inhibitor. Then the percentages of CD11b+/Gr-1+ cells, proliferation and apoptosis of CD4+ T cells were determined. Arginase activity and arginase-1 (Arg-1) protein expression of splenic CD11b+/Gr-1+ cells, and delayed-type hypersensitivity (DTH) response were analyzed. The results showed that Cox-2 blockade significantly decreased the percentages of CD11b+/Gr-1+ cells in the spleen and bone marrow 48 and 72 h after traumatic stress. NS398 inhibited arginase activity and down-regulated the Arg-1 expression of splenic CD11b+/Gr-1+ cells. Moreover, NS398 could promote proliferation and inhibit apoptosis of CD4+ T cells. It also restored DTH response of traumatic mice. Taken together, our data revealed that Cox-2 might play a pivotal role in the accumulation and function of MDSC after traumatic stress.
Effect of interleukin-6 receptor (IL-6R) antibody on polymethyl methacrylate (PMMA) bone cement-mediated expression of osteoprotegerin (OPG) and receptor activator of nuclear factor-kappaB ligand (RANKL) in synovial fibroblasts was investigated. Synovial tissue obtained from total knee arthroplasty was digested and cultured. Inverted microscope was employed to observe the synovial cells and immunocytochemistry (SABC method) staining was used to identify synovial fibroblasts. This experiment was divided into three groups according to different culture media: PMMA group (75 μg/mL PMMA bone cement particles), IL-6R antibody group (10 ng/mL IL-6R antibody+75 μg/mL PMMA bone cement particles), and control group (no IL-6R antibody or PMMA bone cement particles). Influence of IL-6R antibody and PMMA on proliferation of synovial fibroblasts was measured by cell counting kit-8 (CCK-8). ELISA method was used to measure OPG and RANKL levels in culture solution. Fluorescence quantitative real-time PCR (FQ-PCR) was used to detect the expression of OPG and RANKL mRNA. After three consecutive passages, more than 95% of the primary synovial cells became long spindle fibroblast-like cells. SABC staining results showed that the fibroblast-like cells were negative for anti-CD68 antibody and positive for anti-vimentin antibody, with brown madder stained. CCK-8 test demonstrated that the absorbance (A) value at 450 nm was significantly lower in IL-6R antibody group than in PMMA group and control group (P<0.01), but there was no statistically significant difference in A value at 450 nm between the control group and PMMA group (P>0.05). Results of ELISA indicated that the expression of OPG was significantly higher in IL-6R antibody group than in PMMA group and control group (P<0.01). The expression of RANKL was inhibited (P<0.05), and the ratio of OPG/RANKL was significantly increased in IL-6R antibody group as compared with PMMA group and control group. There was no significant difference in the expression of OPG between control group and PMMA group (P>0.05), but the expression of RANKL was higher in PMMA group than in control group (P<0.05), and there was a significant difference in the ratio of OPG/RANKL between them (P<0.05). Results of FQ-PCR revealed the expression of RANKL mRNA was significantly inhibited (P<0.01) and the expression of OPG mRNA was significantly increased (P<0.01) in IL-6R antibody group as compared with PMMA group and control group. The expression of RANKL mRNA was higher in PMMA group than in control group (P<0.05), but the expression of OPG mRNA had no significant difference between them (P>0.05). IL-6R antibody could significantly increase the expression of OPG, but inhibit the expression of RANKL, which might provide a theoretical basis of molecular biology for the prevention and treatment of aseptic loosening of prosthesis.
This study examined the osteogenic effect of electromagnetic fields (EMF) under the simulated in vivo conditions. Rat bone marrow mesenchymal stem cells (BMSCs) and rat osteoblasts were co-cultured and exposed to 50 Hz, 1.0 mT EMF for different terms. Unexposed single-cultured BMSCs and osteoblasts were set as controls. Cell proliferation features of single-cultured BMSCs and osteoblasts were studied by using a cell counting kit (CCK-8). For the co-culture system, cells in each group were randomly chosen for alkaline phosphatase (ALP) staining on the day 7. When EMF exposure lasted for 14 days, dishes in each group were randomly chosen for total RNA extraction and von Kossa staining. The mRNA expression of osteogenic markers was detected by using real-time PCR. Our study showed that short-term EMF exposure (2 h/day) could obviously promote proliferation of BMSCs and osteoblasts, while long-term EMF (8 h/day) could promote osteogenic differentiation significantly under co-cultured conditions. Under EMF exposure, osteogenesis-related mRNA expression changed obviously in co-cultured and single-cultured cells. It was noteworthy that most osteogenic indices in osteoblasts were increased markedly after co-culture except Bmp2, which was increased gradually when cells were exposed to EMF. Compared to other indices, the expression of Bmp2 in BMSCs was increased sharply in both single-cultured and co-cultured groups when they were exposed to EMF. The mRNA expression of Bmp2 in BMSCs was approximately four times higher in 8-h EMF group than that in the unexposed group. Our results suggest that Bmp2-mediated cellular interaction induced by EMF exposure might play an important role in the osteogenic differentiation of BMSCs.
The abnormal expression of MUC15, a novel cell membrane-associated mucin, has been reported to predict poor survival in several cancers. The aim of the present study was to examine the expression of MUC15 in glioma and its correlation with clinicopathological features, including the survival of patients with glioma. The mRNA expression level of MUC15 was determined by RT-PCR, quantitative RT-PCR (RT-qPCR) and Western blotting in seven normal brain tissues and seven glioma tissues, respectively. The protein expression level of MUC15 was immunohistochemically detected in paraffin-embedded samples of 317 glioma tissues and 115 noncancerous brain tissues. The association of MUC15 expression levels with the clinicopathologic features and the prognosis was analyzed. The results showed that both mRNA and protein levels of MUC15 were significantly increased in glioma as compared with those in noncancerous brain tissue. Moreover, MUC15 overexpression was positively correlated with the advanced clinical stages of glioam patients (P<0.01). Furthermore, MUC15 expression levels were significantly correlated with the progression of glioma (P<0.001). Survival analysis indicated that glioma patients with higher MUC15 expression had a significantly shorter overall and 5-year survival time than those with low MUC15 expression. Multivariate analysis suggested that MUC15 overexpression was an independent factor for prognosis (hazard risk: 3.216; P=0.009). It was concluded that MUC15 is overexpressed in glioma tissues. Its overexpression correlates with tumor progression and it is a potentially unfavorable prognostic factor for patients with glioma.
This retrospective analysis compared standard regimen of doxorubicin, bleomycin, vinblastine, and dacarbazine (ABVD) with the dose-dense ABVD regimen (ABVD-21) in terms of efficacy and toxicity. Patients who had early-stage unfavorable or advanced Hodgkin’s lymphoma (HL) according to German Hodgkin Study Group criteria from March 1999 to February 2011 were analyzed for treatment response, long-term survival and hematological toxicity. There were 85 patients in the ABVD-21 group and 118 patients in the ABVD group respectively. The complete remission rates after completion of treatment were 92.9% and 90.7% for ABVD-21 and ABVD, respectively. During a median follow-up period of 62 months, no significant difference was found in projected 10-year progression-free survival (PFS) and overall survival (OS) rates (84.7% and 94.1% respectively for ABVD-21; 81.4% and 91.5% for ABVD). Subgroup analyses showed that ABVD-21 was significantly better than ABVD for patients with IPS≥3 in terms of PFS and OS rates. Grade 3 to 4 leukopenia (51.8% vs. 28.8%, P=0.001) and neutropenia (57.6% vs. 39.0%, P=0.009) were more common with ABVD-21. We were led to conclude that dose-dense ABVD did not result in better tumor control and overall survival than did ABVD for early-stage unfavorable HL. However, patients at high risk, for example, with IPS≥3, may benefit from dose-dense ABVD.
There is controversy regarding the roles of Ureaplasma urealyticum (U. urealyticum) colonization in the development of bronchopulmonary dysplasia (BPD). This study explored the association between U. urealyticum and bronchopulmonary dysplasia at 36 weeks post-menstrual age (BPD36). Studies published before December 31, 2013 were searched from Medline, Embase, Ovid, Web of Science, and Cochrane databases, with the terms “Ureaplasma urealyticum”, “chronic lung disease”, or “BPD36” used, and English language as a limit. The association between U. urealyticum colonization and BPD36 was analyzed with RevMan 4.2.10 software, using the odds ratio (OR) and relative risk (RR) for dichotomous variables. Out of the enrolled 81 studies, 11 investigated the BPD36 in total 1193 infants. Pooled studies showed no association between U. urealyticum colonization and subsequent development of BPD36, with the OR and RR being 1.03 (95% CI=0.78–1.37; P=0.84) and 1.01 (95% CI= 0.88–1.16, P=0.84), respectively. These findings indicated no association between U. urealyticum colonization and the development of BPD36.
Aqueous dispersion and stability of Fe3O4 nanoparticles remain an issue unresolved since aggregation of naked iron nanoparticles in water. In this study, we successfully synthesized different Fe3O4 super-paramagnetic nanoparticles which were modified by three kinds of materials [DSPE-MPEG2000, TiO2 and poly acrylic acid (PAA)] and further detected their characteristics. Transmission electron microscopy (TEM) clearly showed sizes and morphology of the four kinds of nanoparticles. X-ray diffraction (XRD) proved successfully coating of the three kinds of nanoparticles and their structures were maintained. Vibrating sample magnetometer (VSM) verified that their magnetic properties fitted for the super-paramagnetic function. More importantly, the particle size analysis indicated that Fe3O4@PAA had a better size distribution, biocompatibility, stability and dispersion than the other two kinds of nanoparticles. In addition, using CNE2 cells as a model, we found that all nanoparticles were nontoxic. Taken together, our data suggest that Fe3O4@PAA nanoaparticles are superior in the application of biomedical field among the four kinds of Fe3O4 nanoparticles in the future.
Metabolic syndrome (MetS) is a serious threat to public health worldwide with an increased risk of developing type 2 diabetes, cardiovascular diseases and all-cause morbidity and mortality. In this study, a urinary metabolomic approach was performed on high performance liquid chromatography quadrupole time-of-flight mass spectrometry to discriminate 36 male MetS patients and 36 sex and age matched healthy controls. Pattern recognition analyses (principal component analysis and orthogonal projections to latent structures discriminate analysis) commonly demonstrated the difference between MetS patients and no-MetS subjects. This study found 8 metabolites that showed significant changes in patients with MetS, including branch-chain and aromatic amino acids (leucine, tyrosine, phenylalanine and tryptophan), short-chain acylcanitine (tiglylcarnitine), tricarboxylic acid (TCA) cycle intermediate (cis-aconitic acid) and glucuronidated products (cortolone-3-glucuronide and tetrahydroaldosterone-3-glucuronide). The candidate biomarkers revealed in this study could be useful in providing clues for further research focusing on the in-depth investigation of the cause of and cure for MetS.
The development and influencing factors of compliance behavior of investigators in clinical trials were explored. According to literature review, a hypothetical model of development of compliance behavior of investigators in clinical trials was established, and the influencing factors of compliance behavior of investigators and their interrelationships were studied based on questionnaire survey of five hundred investigators sampled randomly from one hundred clinical trial institutions in China. Cronbach’s alpha coefficient and structural equation modeling were adopted to empirically analyze the results. Six variables in the hypothetical model were included: compliance behavior of investigators, credibility of clinical trial, capability of government regulation, quality control of sponsor, quality control of clinical institution and compliance intention of investigators. Empirical analysis showed that the compliance behavior of investigators in clinical trial was directly affected by compliance intention of investigators, quality control of sponsor and quality control of clinical institution. In addition, credibility of clinical trial and capability of government regulation indirectly affected the compliance behavior of investigators in clinical trial through influencing the compliance intention of investigators, quality control of sponsor and quality control of clinical institution. Quality control of sponsor was affected by credibility of clinical trial and capability of government regulation while quality control of clinical institution was only influenced by capability of government regulation.
An evaluation scale of medical services quality was developed on the basis of “patients’ experience”. The questionnaires were developed by, among others, searching relevant literature, collecting well-established assessment scales, measuring patients’ experience and satisfaction, brain-storming, literature analysis. Delphi method was adopted for expert consultation. Scale items were screened and revised. The key indexes were converted. Field surveys were conducted for testing the reliability and validity of the scale. Our modified evaluation scale for measuring medical services quality based on “patients’ experience” included 6 dimensions (tangibility, reliability, responsiveness, assurance, empathy and continuity), and 50 items. The novel scale based on “patients’ experience” may better serve the purpose of assessing medical services quality.