A novel entire traction method with clip-anchored nylon ring facilitating endoscopic submucosal dissection (ESD) of early gastric cancer.

A 48-year-old woman with a high-grade intraepithelial neoplasia of the gastric antrum was referred for ESD treatment.The difficulty of ESD is to expose the submucosa,especially in difficult sites and lesions with severe fibrosis.Adequate submucosal exposure is the most critical technology to reduce complications and improving efficiency.Here we report a novel entire traction method to facilitate safe and efficient ESD.

[Adsorption Characteristics and Mechanism of Ammonia Nitrogen in Water by Nano Zero-valent Iron-modified Biochar].

The adsorption performances of ammonia nitrogen (NH+4-N) in water by unmodified biochar are ineffective. In this study, nano zero-valent iron-modified biochar (nZVI@BC) was prepared to remove NH+4-N from water. The NH+4-N adsorption characteristics of nZVI@BC were investigated through adsorption batch experiments. The composition and structure characteristics of nZVI@BC were analyzed using scanning electron microscopy, energy spectrum analysis, BET-N2 surface area (SSA), X-ray diffraction, and FTIR spectra to explore the main adsorption mechanism of NH+4-N by nZVI@BC. The results showed that the composite synthesized at the iron to biochar mass ratio of 1:30 (nZVI@BC1/30) performed well in NH+4-N adsorption at 298 K. The maximum adsorption amount of nZVI@BC1/30 at 298 K was remarkably increased by 45.96% and reached 16.60 mg·g-1. The pseudo-second-order model and Langmuir model fitted well with the adsorption process of NH+4-N by nZVI@BC1/30. There was competitive adsorption between coexisting cations and NH+4-N, and the sequence of coexisting cations to the adsorption of NH+4-N by nZVI@BC1/30 was Ca2+> Mg2+> K+> Na+. The adsorption mechanism of NH+4-N by nZVI@BC1/30 could be mainly attributed to ion exchange and hydrogen bonding. In conclusion, nano zero-valent iron-modified biochar can improve the adsorption performance of NH+4-N and enhance the application potential of biochar in the field of nitrogen removal from water.

Single-Cell Sequencing Analysis and Multiple Machine Learning Methods Identified G0S2 and HPSE as Novel Biomarkers for Abdominal Aortic Aneurysm.

Identifying biomarkers for abdominal aortic aneurysms (AAA) is key to understanding their pathogenesis, developing novel targeted therapeutics, and possibly improving patients outcomes and risk of rupture. Here, we identified AAA biomarkers from public databases using single-cell RNA-sequencing, weighted co-expression network (WGCNA), and differential expression analyses. Additionally, we used the multiple machine learning methods to identify biomarkers that differentiated large AAA from small AAA. Biomarkers were validated using GEO datasets. CIBERSORT was used to assess immune cell infiltration into AAA tissues and investigate the relationship between biomarkers and infiltrating immune cells. Therefore, 288 differentially expressed genes (DEGs) were screened for AAA and normal samples. The identified DEGs were mostly related to inflammatory responses, lipids, and atherosclerosis. For the large and small AAA samples, 17 DEGs, mostly related to necroptosis, were screened. As biomarkers for AAA, G0/G1 switch 2 (G0S2) (Area under the curve [AUC] = 0.861, 0.875, and 0.911, in GSE57691, GSE47472, and GSE7284, respectively) and for large AAA, heparinase (HPSE) (AUC = 0.669 and 0.754, in GSE57691 and GSE98278, respectively) were identified and further verified by qRT-PCR. Immune cell infiltration analysis revealed that the AAA process may be mediated by T follicular helper (Tfh) cells and the large AAA process may also be mediated by Tfh cells, M1, and M2 macrophages. Additionally, G0S2 expression was associated with neutrophils, activated and resting mast cells, M0 and M1 macrophages, regulatory T cells (Tregs), resting dendritic cells, and resting CD4 memory T cells. Moreover, HPSE expression was associated with M0 and M1 macrophages, activated and resting mast cells, Tregs, and resting CD4 memory T cells. Additional, G0S2 may be an effective diagnostic biomarker for AAA, whereas HPSE may be used to confer risk of rupture in large AAAs. Immune cells play a role in the onset and progression of AAA, which may improve its diagnosis and treatment.

Contrastive study for coadsorption of copper and two dihydroxybenzene isomers by a multi-amine modified resin.

Coadsorption of Cu(II) and two dihydroxybenzene isomers (hydroquinone, HQ and catechol, CAT) onto a multi-amine modified resin (CEAD) were comparatively studied. The presence of Cu(II) promoted adsorption of both HQ and CAT by a maximum of 25.8% and 41.6%, respectively. However, two diphenols exerted a very different influence on Cu(II) uptake. Higher concentrations of HQ consistently suppressed Cu(II) adsorption while the coexistence of CAT facilitated it, especially at lower CAT concentrations. The interactions among solutes and adsorbents were revealed by means of kinetic tracking, sequential adsorption experiments, and characterizations/calculations (FTIR, XPS, MINTEQ and DFT). Cu(II) and HQ/CAT competed for amine sites with the order of adsorption affinity as HQ > Cu(II) > CAT. The bridging effect of Cu(II) forming ternary complexes (amine-Cu-CAT/HQ) on the resin phase was the dominant mechanism for the enhanced adsorption of diphenols. The [Cu-CAT] complex species showed a lower affinity to bind directly to amine sites compared with free Cu2+. Instead, the complex could be attracted by the polyphenyl matrix of CEAD, contributing to the increase of Cu(II) adsorption. Additionally, Cu(II) and diphenols were successively recovered, and CEAD could be stably reused. The findings will guide adsorbent applications and the environmental fate of concurrent heavy metals and phenolic compounds.

Multifold enhanced synergistic removal of nickel and phosphate by a (N,Fe)-dual-functional bio-sorbent: Mechanism and application.

A novel (N,Fe)-dual-functional biosorbent (N/Fe-DB) capable of efficient synergistic removal of Ni(II) and H2PO4- from aqueous solution was synthesized. The adsorption capacities of Ni(II) and H2PO4- were both remarkably enhanced over 3 times compared with those in single systems. Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy confirmed that complexation of amino groups and ligand exchange of hydrous ferric oxide in N/Fe-DB played dominant roles. The electric double layer compressing and chelating ligand of deprotonated H2PO4- accounted for the enhanced removal of Ni(II) in binary system, while cation bridge interaction promoted uptake of H2PO4-. Furthermore, the coadsorbates were sequentially recovered, with the ratios of more than 99.0%. Besides, the recovered N/Fe-DB remained stable and applicable to the treatment of real electroplating wastewater even after six adsorption-regeneration cycles. Since the electroplating industry is springing up, effective control of heavy metals and phosphate has attracted global concerns. Based on the enhanced coremoval properties and superb regenerability, N/Fe-DB is potentially applicable to practical production.

Association between neutrophilic granulocyte percentage and depression in hospitalized patients with heart failure.

BACKGROUND: Previous researches reveal that depression is associated with increased inflammatory markers. As a simple and cheap inflammatory marker, we hypothesize that neutrophilic granulocyte percentage is associated with depression in hospitalized heart failure patients, whose prevalence of depression is at a very high level. METHODS: Three hundred sixty-six cases of hospitalized heart failure patients with left ventricular ejection fraction (LVEF) ≤45% and New York Heart Association (NYHA) class II-IV were enrolled. All the enrolled patients received Hamilton Rating Scale for Depression (24-items) (HAM-D24). The demographic, clinical data, blood samples and echocardiography were documented. The Pearson simple linear correlation was performed to evaluate the confounding factors correlated with HAM-D24 depression index. The significantly correlated factors were enrolled as independent variables in Logistic regression to determine the risk or protective factors for depression, which was taken as dependent variable. RESULTS: Two hundred ten cases of hospitalized heart failure patients (57.4%) had depression. Among them, 134 patients (63.8%) had mild depression, 58 patients (27.6%) had moderate depression and 18 patients (8.6%) had severe depression. Pearson simple linear correlation revealed that in hospitalized patients with heart failure, the neutrophils granulocyte percentage was positively correlated with the HAM-D24 depression index (r = .435, p < .001). After the adjustment of age, BMI, number of members of the household, smoking index, New York Heart Association (NYHA) classification, hemoglobin, TC, LDL-C, creatinine, cystatin-C, TBIL and albumin, the neutrophils granulocyte percentage is still significantly associated with depression in hospitalized heart failure patients (OR = 1.046, p < .001). CONCLUSIONS: The neutrophils granulocyte percentage may be used as a new marker for depression in hospitalized heart failure patients.

Notch4 inhibition reduces migration and invasion and enhances sensitivity to docetaxel by inhibiting Akt/fascin in pancreatic cancer cells.

Overexpression of Notch4 is associated with a variety of tumor types. Only sparse information exists on Notch4 expression in pancreatic cancer (PC). The present study demonstrated that Notch4 expression was significantly upregulated in PC cell lines compared with a non-transformed pancreatic epithelial cell line, HPDE6c-7. To investigate the possible role of Notch4 in PC cells, an RNA interference approach was used to silence Notch4 expression. The results revealed that small interfering RNA (siRNA) targeting Notch4 significantly impeded the viability, migration and invasion abilities of PC cells in vitro. Downregulation of Notch4 with siRNA sensitized cells to the action of docetaxel. Furthermore, Notch4 downregulation enhanced the inhibition of Akt activation and the fascin expression induced by docetaxel in PC cells. Together, these data provide insight into the function of Notch4 and suggest that Notch4 may represent a new potential target for gene therapy in PC.

Effects of Salt Loading on Plasma Osteoprotegerin Levels and Protective Role of Potassium Supplement in Normotensive Subjects.

BACKGROUND: Excess dietary salt is strongly correlated with cardiovascular disease, morbidity, and mortality. Conversely, potassium likely elicits favorable effects on cardiovascular disorders. In epidemiological studies, increased plasma osteoprotegerin (OPG) concentrations are associated with atherosclerosis and vascular deaths. Our study was designed to examine the effects of salt intake and potassium supplementation on plasma OPG levels in normotensive subjects.Methods and Results:The 18 normotensive subjects were selected from a rural community in China. They were sequentially maintained on low-salt diet for 7 days (3 g/day, NaCl), high-salt diet for 7 days (18 g/day), and high-salt diet with potassium supplementation for 7 days (18 g/day of NaCl+4.5 g/day of KCl). High-salt intake enhanced plasma OPG levels (252.7±13.9 vs. 293.4±16.1 pg/mL). This phenomenon was abolished through potassium supplementation (293.4±16.1 vs. 235.1±11.3 pg/mL). Further analyses revealed that the OPG concentration positively correlated with 24-h urinary sodium excretion (r=0.497, P<0.01). By contrast, OPG concentration negatively correlated with 24-h urinary potassium excretion (r=0.594, P<0.01). CONCLUSIONS: Salt loading can enhance the production of circulating OPG. Potassium supplementation can reverse the effects of excessive OPG. Our study results may improve our understanding of the roles of salt and potassium in the risk of cardiovascular disorders.

Elevation of Fasting Ghrelin in Healthy Human Subjects Consuming a High-Salt Diet: A Novel Mechanism of Obesity?

Overweight/obesity is a chronic disease that carries an increased risk of hypertension, diabetes mellitus, and premature death. Several epidemiological studies have demonstrated a clear relationship between salt intake and obesity, but the pathophysiologic mechanisms remain unknown. We hypothesized that ghrelin, which regulates appetite, food intake, and fat deposition, becomes elevated when one consumes a high-salt diet, contributing to the progression of obesity. We, therefore, investigated fasting ghrelin concentrations during a high-salt diet. Thirty-eight non-obese and normotensive subjects (aged 25 to 50 years) were selected from a rural community in Northern China. They were sequentially maintained on a normal diet for three days at baseline, a low-salt diet for seven days (3 g/day, NaCl), then a high-salt diet for seven days (18 g/day). The concentration of plasma ghrelin was measured using an immunoenzyme method (ELISA). High-salt intake significantly increased fasting ghrelin levels, which were higher during the high-salt diet (320.7 ± 30.6 pg/mL) than during the low-salt diet (172.9 ± 8.9 pg/mL). The comparison of ghrelin levels between the different salt diets was statistically-significantly different (p < 0.01). A positive correlation between 24-h urinary sodium excretion and fasting ghrelin levels was demonstrated. Our data indicate that a high-salt diet elevates fasting ghrelin in healthy human subjects, which may be a novel underlying mechanism of obesity.

High Salt Diet Affects Renal Sodium Excretion and ERRα Expression.

Kidneys regulate the balance of water and sodium and therefore are related to blood pressure. It is unclear whether estrogen-related receptor α (ERRα), an orphan nuclear receptor and transcription factor highly expressed in kidneys, affects the reabsorption of water and sodium. The aim of this study was to determine whether changes in the expressions of ERRα, Na⁺/K⁺-ATPase and epithelial sodium channel (ENaC) proteins affected the reabsorption of water and sodium in kidneys of Dahl salt-sensitive (DS) rats. SS.13BN rats, 98% homologous to the DS rats, were used as a normotensive control group. The 24 h urinary sodium excretion of the DS and SS.13BN rats increased after the 6-week high salt diet intervention, while sodium excretion was increased in DS rats with daidzein (agonist of ERRα) treatment. ERRα expression was decreased, while ß- and γ-ENaC mRNA expressions were increased upon high sodium diet treatment in the DS rats. In the chromatin immunoprecipitation (CHIP) assay, positive PCR signals were obtained in samples treated with anti-ERRα antibody. The transcriptional activity of ERRα was decreased upon high salt diet intervention. ERRα reduced the expressions of ß- and γ-ENaC by binding to the ENaC promoter, thereby increased Na+ reabsorption. Therefore, ERRα might be one of the factors causing salt-sensitive hypertension.

Poly (ADP-ribose) transferase/polymerase-1-deficient mice resistant to age-dependent decrease in ß-cell proliferation.

Basal and adaptive ß-cell regeneration capacity declines with old age, but the underlying molecular mechanisms remain incompletely understood. Poly (adenosine diphosphate [ADP]-ribose) polymerase 1 (PARP-1) is considered a multifunctional enzyme and transcription factor that regulates pancreatic ß-cell death, regeneration and insulin secretion. We analyzed the capacity of ß-cell regeneration in 2-month-old (young) and 12-month-old (old) wild-type (WT) and PARP-1⁻/⁻ mice before and after low-dose streptozotocin (STZ), a stimulus of ß-cell regeneration and the underlying mechanism. Before STZ administration, young WT and PARP-1⁻/⁻ mice showed similar ß-cell proliferation. By contrast, old WT but not old PARP-1⁻/⁻ mice showed severely restricted ß-cell proliferation. In further assessment of the adaptive ß-cell regeneration capacity with age, we observed that with a single low dose of STZ, young WT and PARP-1⁻/⁻ mice showed a similar increase in ß-cell proliferation, with few changes in old WT mice. Surprisingly, adaptive ß-cell proliferation capacity was significantly higher in old PARP-1⁻/⁻ mice than old WT mice after STZ administration. The ability of ß-cell mass to expand was associated with increased levels of the regenerating (Reg) genes RegI and RegII but not RegIV. Therefore, PARP-1 is a key regulator in ß-cell regeneration with advancing age in mice.

Arginase I attenuates inflammatory cytokine secretion induced by lipopolysaccharide in vascular smooth muscle cells.

OBJECTIVE: Inflammation plays an important role in atherosclerosis. Arginase I (Arg I) promotes the proliferation of vascular smooth muscle cells; however, the effect of Arg I on inflammation remains unknown. The present study investigated the role of Arg I in inflammation in vitro and in vivo. METHODS AND RESULTS: Quantitative reverse transcription-polymerase chain reaction and Western blot analysis demonstrated that Arg I inhibited tumor necrosis factor-α production induced by lipopolysaccharide in human aortic smooth muscle cells. Inducible nitric oxide synthase substrate competition and nuclear factor-κB activation were main contributors to lipopolysaccharide-mediated inflammatory cytokine generation. However, Arg I could attenuate the function of inducible nitric oxide synthase and inhibit the subsequent nuclear factor-κB activation, leading to inhibition of tumor necrosis factor-α generation. Furthermore, upregulation of Arg I significantly decreased macrophage infiltration and inflammation in atherosclerotic plaque of rabbits, whereas downregulation of Arg I aggravated these adverse effects. CONCLUSIONS: The results indicate the antiinflammatory effects of Arg I and suggest an unexpected beneficial role of Arg I in inflammatory disease.

Hypertrophy of ligamentum flavum in lumbar spine stenosis associated with the increased expression of connective tissue growth factor.

Hypertrophy of the ligamentum flavum (LF) contributes to lumbar spinal stenosis (LSS), and results mainly from fibrosis. Connective tissue growth factor (CTGF) is a profibrotic factor involved in the fibrotic process. This study aimed to evaluate CTGF expression in hypertrophied lumbar LF and the involvement of CTGF in LF hypertrophy. Ten patients with LSS were enrolled in this study. The control group included 10 patients with lumbar disc herniation. LF thickness was measured on the preoperative axial T1-weighted MRI. LF samples were collected during surgery. LF fibrosis was scored by Masson's trichrome staining. CTGF expression was determined by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry. Correlation between LF thickness and CTGF expression was analyzed. Human LF cells were cultured and treated with recombinant human (rh) CTGF. Expression of types I and III collagen was determined by real-time PCR and ELISA. The thickness and fibrosis scores of LF in the LSS group were higher than that in the control group (all P < 0.001). CTGF was expressed in the extracellular matrix of all ligament samples, and was significantly higher in the LSS group than that in the control group (P < 0.001). The increase of CTGF expression was positive correlation with the LF thickness (r = 0.969, P = 0.000). rhCTGF treatment increased the mRNA expression and protein synthesis of types I and III collagen of the LF cells (all P < 0.001). Our results suggest that the increased expression of CTGF is associated with hypertrophy of the LF in patients with LSS.

Glucagon-like peptide 1 protects microvascular endothelial cells by inactivating the PARP-1/iNOS/NO pathway.

Increasing studies suggest that the activity of GLP-1 might be of significant importance in the development of type 2 diabetes beyond its serum glucose-lowering effects. However, to date, the anti-apoptosis mechanism by which GLP-1 acts on MILE SVEN 1 (MS-1) cells has not been fully explored with regard to the intracellular signaling pathway. Increasing evidence shows that apoptosis of islet microvascular endothelial cells (IMECs) participates in the pathogenesis of diabetes. We wondered whether GLP-1 exerts its anti-apoptosis effects by inactivating the PARP-1/iNOS/NO pathway in oxidized low-density-lipoprotein (oxLDL)-induced apoptosis in mouse IMECs (MS-1 cells), which may linked to GLP-1R/cAMP levels. MTT assay revealed that 2-h pre-incubation with GLP-1 markedly restored the oxLDL-induced loss of MS-1 viability in a concentration-dependent manner. This effect was accompanied by a significant decrease in intracellular nitric oxide (NO) activity. Moreover, GLP-1 suppressed lipid peroxidation, restored the activities of endogenous antioxidants, and decreased the level of NO. Pre-incubating MS-1 cells with GLP-1 reduced cell apoptosis. Finally, GLP-1 could efficiently prevent the upregulation of poly(ADP-ribose) polymerase-1/nitrotyrosine and inducible NO synthase protein. Simultaneously, the expression of GLP-1 receptor and the level of cAMP was consistent with the administration of GLP-1. Our findings suggest that GLP-1 can effectively protect MS-1 cells against oxLDL-induced apoptosis, which may be important in preventing the pathogenesis of diabetes mellitus.

Amide-linkage formed between ammonia plasma treated poly(D,L-lactide acid) scaffolds and bio-peptides: enhancement of cell adhesion and osteogenic differentiation in vitro.

The surface characteristics of scaffolds for bone tissue engineering must support cell adhesion, migration, proliferation, and osteogenic differentiation. In the study, poly(D,L-lactide acid) (PDLLA) scaffolds were modified by combing ammonia (NH(3) ) plasma pretreatment with Gly-Arg-Gly-Asp-Ser (GRGDS)-peptides coupling technologies. The x-ray photoelectron spectroscopy (XPS) survey spectra showed the peak of N1s at the surface of NH(3) plasma pretreated PDLLA, which was further raised after GRGDS conjugation. Furthermore, N1s and C1s in the high-resolution XPS spectra revealed the presence of -C=N(imine), -C-NH-(amine), and -C=O-NH- (amide) groups. The GRGDS conjugation increased amide groups and decreased amine groups in the plasma-treated PDLLA. Confocal microscope and high performance liquid chromatography verified the anchored peptides after the conjugation process. Bone marrow mesenchymal stem cells were co-cultured with scaffolds. Fluorescent microscope and scanning electron microscope photographs revealed the best cell adhesion in NH(3) plasma pretreated and GRGDS conjugated scaffolds, and the least attachment in unmodified scaffolds. Real-time PCR demonstrated that expression of osteogenesis-related genes, such as osteocalcin, alkaline phosphatase, type I collagen, bone morphogenetic protein-2 and osteopontin, was upregulated in the single NH(3) plasma treated and NH(3) plasma pretreated scaffolds following GRGDS conjugation. The results show that NH(3) plasma treatment promotes the conjugation of GRGDS peptides to the PDLLA scaffolds via the formation of amide linkage, and combination of NH(3) plasma treatment and peptides conjugation may enhance the cell adhesion and osteogenic differentiation in the PDLLA scaffolds. © 2011 Wiley Periodicals, Inc. Biopolymers 95: 682-694, 2011.

Hyperglycemia induces apoptosis of pancreatic islet endothelial cells via reactive nitrogen species-mediated Jun N-terminal kinase activation.

Hyperglycemia significantly stimulates pancreatic islet endothelial cell apoptosis; however, the precise mechanisms are not fully understood. In the present study, treating pancreatic islet endothelial (MS-1) cells with high glucose (30mmol/l) but not mannitol significantly increased the number of apoptotic cells as compared with a physiological glucose concentration (5.5mmol/l). Hyperglycemia significantly stimulated the expression of inducible nitric oxide synthase (iNOS) and production of NO and peroxynitrite (ONOO(-)), relevant to MS-1 cell apoptosis. Moreover, induced reactive nitrogen species (RNS) significantly increased the expression of bax, cleaved caspase-3 and poly adenosine diphosphate (ADP)-ribose polymerase (PARP) via JNK activation, but the expression of bcl-2 was not altered. Furthermore, SP600125 (a specific inhibitor of JNK) and 1400W (a specific inhibitor of iNOS) significantly attenuated cell apoptosis induced by high glucose. Therefore, hyperglycemia triggers MS-1 cell apoptosis by activating an intrinsic-dependent apoptotic pathway via RNS-mediated JNK activation.

[Relationships of serum selenium, VEGF and sFas levels in leukemia patients].

In order to investigate the relationships of cancer chemopreventive trace element-selenium, vascular endothelial growth factor (VEGF) and soluble Fas (sFas) in leukemia patients, serum selenium concentration was measured by atomic spectrometry and levels of VEGF and sFas were simultaneously detected by enzyme linked immunosorbent assay (ELISA). Relationships of selenium, VEGF and sFas were analyzed by linear correlation. The results showed that serum selenium concentration in newly-diagnosed patients and relapsed patients was significantly lower than that in the control group (p < 0.05), especially in relapsed group. VEGF and sFas concentrations of refractory/relapsed group were significantly higher than those in the control group and the remission group (p < 0.05). But no significant difference was found between the remission group and the control group (p > 0.05). In leukemia patients, negative correlation was observed between selenium and VEGF (r = -0.529, p < 0.01), so did between selenium and sFas (r = -0.432, p < 0.01). Positive correlation was found between VEGF and sFas (r = 0.663, p < 0.01). It is concluded that the selenium, VEGF and sFas may take part in the occurrence of MDR in leukemia patients. Selenium has negative correlation with VEGF and sFas, which means that it may be used as an effective assistant agent to improve therapeutic effect of chemotherapy. However, further study in more cases is needed to reach a definite conclusion.

[Anti-leukemia activity of T cells impacted by dendritic cells added with sodium selenite].

The study was purposed to explore the quantity, morphology and immunophenotype of dendritic cells (DC) acquired by co-cultivated system with 3 types of cytokines and sodium selenite (Se) from peripheral blood mononuclear cells (PBMNCs), and to investigate the effects of Se on inducing the cytotoxic T lymphocyte (CTL) to get specific anti-leukemic activity in vitro by DC pulsed with K562 cell frozen-thawed antigen (antigen cell loading). PBMNCs isolated from healthy donors were cultured in RPMI 1640 medium contained 10% FBS supplied with 3 cytokines (rhGM-CSF, rhIL-4, TNF-alpha) for 4 days, DCs harvested were divided into 4 groups, DCI: DC alone; DCII: DC + Se (adding 0.5 micromol/L of Se); DCIII: DC + K562 (pulsed with lysed K562 cells); DCIV: DC + Se + K562. Morphology of DCs was observed under microscope at day 7. The CD1a, CD40, CD83, and CD86 were detected by FCM. Cytotoxicity of T cells induced by DC were measured with LDH release test at day 12. The level of IL-12 in supernatant of cultured DCs were determined with ELISA. The results indicated that at 7th day DC in 4 groups showed characteristic morphology, the colony numbers of 4 groups were all higher than those before cultivation. There were no obvious differences of morphology and colony counts between DCI group and DCII group. The colony numbers of DCIII group and DCIV group increased, as well as the ratio of suspended cells enhanced. The expressions of CD1a, CD40, CD83 and CD86 in 4 groups of DC were significantly higher than those in PBMNC group (p < 0.01), the expressions of CD1a and CD40 in 4 groups of DC did not display significant difference (p > 0.05), the expressions of CD83 and CD86 in both DCIII group and DCIV group were all higher than those in DCI group and DCII group (p < 0.01), but their expressions of CD83 and CD86 in DCI and DCII were not significantly different (p > 0.05), as well as those in DCIII group and DCIV group. With the ratio of 25:1 between E:T, killing rate of CTL on K562 cells in 4 DC groups were 15.3 +/- 2.3%, 26.3 +/- 3.7%, 28.2 +/- 4.5% and 36.2 +/- 3.7% respectively, all obviously higher than those of T cell group without being sensitized by DCs (5.9 +/- 2.4%) (p < 0.01), The CTL effect in DCIV group was the highest, which was higher than those in other 3 DC groups (p < 0.01); the effects in both DCII and DCIII group were also higher than that in DCI group (p < 0.01), but their difference between DCII and DCIII groups did not show significance (p > 0.05). The levels of IL-12 in supernatant of DCI, DCII, DCIII and DCIV groups were 257.0 +/- 64.2, 328.1 +/- 43.9, 323.0 +/- 53.5 and 353.9 +/- 46.2 pg/ml respectively, all significantly higher than that in supernatant of T cell alone group without being sensitized by DCs (35.27 +/- 27.1 pg/ml) (p < 0.01), The levels in DCII, DCIII and DCIV groups were all higher than that in DCI group (p < 0.01), but their levels between DCII, DCIII and DCIV groups were not of significant difference (p > 0.05). It is concluded that matured DCs can be successfully obtained from PBMNCs by a culture system contained rhGM-CSF, rhIL-4 and TNF-alpha with or without low-dose of Se (0.5 micromol/L) in vitro. Using K562 cell frozen-thawed antigen, DC express more adhesive molecules and co-stimulating molecules (CD83, CD86), and increase the secretion of IL-12, as well as the killing effects of CTL on special target cells. Low dose of Se did not showed effects on quantity and morphology of matured DC harvested, as well as their expression of mature phenotypes, it raised levels of IL-12 secreted by DCs, reaching the same level as using K562 cell frozen-thawed antigen, and it showed synergistic effect on induction of CTL with K562 cell frozen-thawed antigen.

[Reversal effect of sodium selenite on multidrug resistance in K562/ADR cell line and its mechanisms].

This study was purposed to investigate the reversal effect of sodium selenite on multidrug resistance in adriamycin-resistant leukemic cell line K562/ADR and its mechanisms. The cytotoxicity and the reversal effect of sodium selenite on K562/ADR cells were assayed by MTT method; the apoptosis rate of K562 and K562/ADR cells were detected by flow cytometery, the mRNA expressions of mdr1 and bcl-2 were measured by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that 10 micromol/L sodium selenite significantly increased the cytotoxicity of adriamycin to K562/ADR cell and the reverse index (RI) was 2.31; the early apoptosis rate of K562 cells was elevated after treatment with 5 micromol/L Na(2)SeO(3) for 48 hours; and the medium-term and late apoptosis rate was elevated after treatment with both 5 and 10 micromol/L Na(2)SeO(3) for 48 and 72 hours. Both doses of 5 and 10 micromol/L Na(2)SeO(3) increased the early apoptosis rate of K562/ADR at 48 hours, and also increased the medium-term and late apoptosis rate after treating for 48 and 72 hours. The apoptosis rate was higher at dose of 10 micromol/L than that at 5 micromol/L, the apoptosis rate at 72 hours also was higher than that at 48 hours. The expressions of mdr1 mRNA and bcl-2 mRNA were decreased significantly by 10 micromol/L sodium selenite. It is concluded that sodium selenite can reverse the multidrug resistance in K562/ADR partially by down-regulating the expressions of mdr1 mRNA and bcl-2 mRNA, and increasing apoptosis rate of K562/ADR cells.