Desulfovibrio vulgaris bacterioferritin uses H(2)O(2) as a co-substrate for iron oxidation and reveals DPS-like DNA protection and binding activities

β?-GPI (β?-glycoprotein I) is a plasma glycoprotein ascribed with an anti-angiogenic function; however, the biological role and molecular basis of its action in cell migration remain unknown. The aim of the present study was to assess the contribution of β?-GPI to HAEC (human aortic endothelial cell) migration and the details of its underlying mechanism. Using wound healing and Boyden chamber assays, we found that β?-GPI inhibited endothelial cell migration, which was restored by its neutralizing antibody. NF-κB (nuclear factor κB) inhibitors and lentiviral siRNA (small interfering RNA) silencing of NF-κB significantly attenuated the inhibitory effect of β?-GPI on cell migration. Moreover, β?-GPI was found to induce IκBα (inhibitor of NF-κB) phosphorylation and translocation of p65 and p50. We further demonstrated that mRNA and protein levels of eNOS [endothelial NO (nitric oxide) synthase] and NO production were all increased by β?-GPI and these effects were remarkably inhibited by NF-κB inhibitors and siRNAs of p65 and p50. Furthermore, β?-GPI-mediated inhibition of cell migration was reversed by eNOS inhibitors and eNOS siRNAs. The findings of the present study provide novel insight into the ability of β?-GPI to inhibit endothelial cell migration predominantly through the NF-κB/eNOS/NO signalling pathway, which indicates a potential direction for clinical therapy in vascular diseases.     

Auxotrophic recombinant Mycobacterium bovis BCG overexpressing Ag85B enhances cytotoxicity on superficial bladder cancer cells in vitro

BCG therapy remains at the forefront of immunotherapy for treating patients with superficial bladder cancer. The high incidence of local side effects and the presence of non-responder diseases have led to efforts to improve the therapy. Hence, we proposed that an auxotrophic recombinant BCG strain overexpressing Ag85B (BCG ?leuD/Ag85B), could enhance the cytotoxicity to the human bladder carcinoma cell line 5637. The rBCG was generated using an expression plasmid encoding the mycobacterial antigen Ag85B to transform a BCG ?leuD strain. The inhibitory effect of BCG ?leuD/Ag85B on 5637 cells was determined by the MTT method, morphology observation and a LIVE/DEAD assay. Gene expression profiles for apoptotic, cell cycle-related and oxidative stress-related genes were investigated by qRT-PCR. Bax, bcl-2 and p53 induction by BCG ?leuD/Ag85B treatment was evaluated by Western blotting. BCG ?leuD/Ag85B revealed a superior cytotoxicity effect compared to the control strains used in this study. The results showed that the expression level of pro-apoptotic and cell cycle-related genes increased after BCG ?leuD/Ag85B treatment, whereas the mRNA levels of anti-apoptotic genes decreased. Interestingly, BCG ?leuD/Ag85B also increased the mRNA level of antioxidant enzymes in the bladder cancer cell line. Bax and p53 proteins levels increased following treatment. In conclusion, these results suggest that treatment with BCG ?leuD/Ag85B enhances cytotoxicity for superficial bladder cancer cells in vitro. Therefore, rBCG therapy may have potential benefits in the treatment of bladder cancer.     

Fibroblasts enhance the in vitro survival of human memory and naïve B cells by maintaining intracellular levels of glutathione

To gain insight into the mechanism of memory B cell survival, we cultured highly purified subpopulations of tonsillar B cells with tonsillar fibroblasts. The fibroblasts greatly enhanced the survival of memory and na?ve B cells but did not delay the rapid apoptosis of germinal center B cells. B cell activation was not observed during the period of culture, as shown by the absence of activation markers and of cycling cells. These findings were reproduced when the B cells were physically separated from the fibroblasts by a semi-permeable transwell-membrane, indicating that the survival factor(s) were diffusible. Several cytokines including IL-6, IL-15, and VEGF were tested for survival activity but none could replace the fibroblasts. However, the addition of reduced glutathione (GSH) to the na?ve and memory B cells significantly enhanced their survival, and depletion of GSH resulted in rapid loss of B cell viability. Furthermore, intracellular glutathione levels were maintained when the B cells were co-cultured with fibroblasts. Our results suggest that glutathione plays an important role in the survival of memory and na?ve B cells in the presence of stromal cells.     

Phosphatidylethanolamine methylase and cyclic nucleotide phosphodiesterase activities in human B lymphoid hemopathies

Phospholipid methylase and cyclic nucleotide phosphodiesterase activities were studied in human B lympho?d hemopathies (51 patients: acute lymphoblastic leukemia, B lymphoma, chronic lymphocytic leukemia, hairy cell leukemia) and compared with activities in lymphoblast?d and Burkitt lymphoma cell lines and with normal B lymphocytes: methylase activity proved to be lower in ALL and high grade lymphoma and inversely related to the percent of cells in S phase state; the A/G ratio of phosphodiesterases was low in ALL and CLL and high in hairy cell leukemia and it was related to the percent of cells in S phase state.     

Root hair development involves asymmetric cell division in Brachypodium distachyon and symmetric division in Oryza sativa

? The root epidermis of most angiosperms comprises hair (H) cells and nonhair (N) cells. H cells are shorter than N cells in grasses (Poaceae). ? The aim of this study was to determine the developmental basis for differences in H and N cell size in the grasses Brachypodium distachyon and Oryza sativa. ? We show that cytokinesis in the last cell division in each epidermal file is asymmetric in B. distachyon. The smaller daughter cell becomes an H cell and the larger cell forms an N cell. By contrast, asymmetric cytokinesis does not occur during H cell and N cell development in O. sativa and the differences in size arise because there is more cell expansion in N cells than in H cells after root hair initiation. ? The different sizes of mature H and N cells result from cell division asymmetry in B. distachyon but different rates of cell expansion in O. sativa. We hypothesize that the mechanism that includes asymmetric cytokinesis during the development of H and N cells evolved among the Pooideae or ancestors of this subfamily.     

Improved accumulation of betulin and betulinic acid in cell suspension culture of Betula pendula roth by abiotic and biotic elicitors

Abstract

Betulin (B) and betulinic acid (BA) are two triterpenes with diverse pharmacological and physiological actions. Elicitation of Betula pendula Roth cell cultures by elicitors is an excellent strategy to increase B and BA levels. Six abiotic and biotic elicitors were studied to improve accumulation of B and BA in the cell culture of B. pendula. The B and BA production in treated cells was verified by HPLC. The results showed the maximum growth index (7) on day 3 in cells treated with 0.5?mg L^?1 chlorocholine chloride (CCC). The increased accumulation of BA in the cells treated with 200?mg L^?1 of chitosan was found to be 5.9?×?(6.5?mg g^?1 DW) higher over control cells. Treating the cells with 2?mg L^?1 of CCC, after 7?days, led to 149.3× enhancement of B content (19.4?mg g^?1 DW) over the controls. Production of this triterpenoid at a much shorter time with a much higher growth rate can be economic and lead to producing large amounts of B and BA for anti-cancer and HIV drugs preparation.     

Antiproliferative activity of Humulus lupulus extracts on human hepatoma (Hep3B), colon (HT-29) cancer cells and proteases,tyrosinase, β-lactamase enzyme inhibition studies

The aims of this study were to examine the antiproliferation of Humulus lupulus extracts on human hepatoma carcinoma (Hep3B) and human colon carcinoma (HT-29) cell lines along with enzyme inhibitory effects of the crude extracts. Potential cell cytotoxicity of six different H. lupulus extracts were assayed on various cancer cells using MTT assay at 24, 48 and 72?h intervals. Methanol-1 extract has inhibited the cell proliferation with doses of 0.6–1?mg/mL in a time dependent (48 and 72 hours) manner in Hep3B cells with 70% inhibition, while inhibitory effect was not seen in colon cancer cells. Acetone extract has increased the cell proliferation at low doses of 0.1?mg/mL for 72?h in Hep3B cells and 0.1–0.2?mg/mL for 48 and 72?h in HT29 cells. The inhibitory effects of the extracts were compared by relative maximum activity values (V_max) using proteases such as α-chymotrypsin, trypsin and papain, tyrosinase and β-lactamase (penicillinase).     

Effect of Boron on Osteogenic Differentiation of Human Bone Marrow Stromal Cells

Bone marrow stromal cells (BMSCs) have been well established as an ideal source of cell-based therapy for bone tissue engineering applications. Boron (B) is a notable trace element in humans; so far, the effects of boron on the osteogenic differentiation of BMSCs have not been reported. The aim of this study was to evaluate the effects of boron (0, 1, 10,100, and 1,000?ng/ml) on osteogenic differentiation of human BMSCs. In this study, BMSCs proliferation was analyzed by cell counting kit-8 (CCK8) assay, and cell osteogenic differentiation was evaluated by alkaline phosphatase (ALP) activity assay, Von Kossa staining, and real-time PCR. The results indicated that the proliferation of BMSCs was no different from the control group when added with B at the concentration of 1, 10, and 100?ng/ml respectively (P?>?0.05); in contrast, 1,000?ng/ml B inhibited the proliferation of BMSCs at days?4, 7, and 14 (P?相似文献   

Clinical safety and pharmacological profile of the HLA-DR antibody 1D09C3 in patients with B cell chronic lymphocytic leukemia and lymphoma: results from a phase I study

1D09C3 is a human monoclonal IgG4-type antibody against human leukocyte antigen-DR (HLA-DR) which has demonstrated pro-apoptotic activity against lymphoid tumors in vitro and in vivo. We report results from a phase I dose-escalation study which aimed to identify tolerated dosing, and the pharmacokinetic and pharmacodynamic profile of 1D09C3. Fourteen patients with relapsed/refractory B cell type leukemia/lymphoma were treated and followed after up to 4 weekly infusions of 1D09C3, administered in 6 dose levels at 0.25?C8?mg/kg/day. Treatment was tolerated well with mostly mild side effects. The most common grade III?CIV toxicities were hematological events observed in 4 patients. In one patient, treated at 8.0?mg/kg/day, a dose limiting toxicity occurred, identified as an invasive catheter-related infection. Adverse events resolved completely without long-term sequelae. 1D09C3 reduced peripheral blood B cells and monocytes by a median of 73?C81?% in all patients, with a nadir reached 30?C60?min after infusion and sustained for <96?h. Granulocytes and natural killer cells predominantly increased with variable time courses. Pharmacokinetic assessments showed detectable drug concentrations at doses 4?C8?mg/kg/day and a terminal half-life of 0.7?C7.9?h. Effective saturation of HLA-DR on peripheral blood B cells/monocytes was achieved, varying consistently with available serum concentrations and the cell-reducing activity of 1D09C3. In summary, 1D09C3 could be administered safely in patients with advanced B cell malignancies. Pharmacodynamic studies demonstrated a strong dose dependent but transient reduction of peripheral blood B cells and monocytes, consistent with a short drug serum availability.     

Immunomodulatory activity of Lactobacillus strains isolated from fermented vegetables and infant stool

Four Lactobacillus strains?- Lactobacillus plantarum CJLP133, L. plantarum CJLP243, L. plantarum CJNR26, and Lactobacillus gasseri CJMF3?- were isolated from Korean fermented food or healthy infant feces, and their capacity to modulate cellular and humoral immune responses was studied. Feeding of the tested lactobacilli for 8?weeks did not alter the weight of and cell numbers in the spleen of mice. However, CJLP133 and CJLP243 strains increased the T lymphocyte population in the spleen of mice, while CJNR26 and CJMF3 increased the B lymphocyte population. In splenocytes treated with concanavalin A, ingestion of CJLP133 and CJLP243 promoted T lymphocyte proliferation and secretion of T cell cytokines, whereas feeding of the CJNR26 and CJMF3 strains enhanced B lymphocyte proliferation in splenocytes treated with lipopolysaccharide and plaque formation. These results suggest that CJLP133 and CJLP243 have immunostimulating activity through the enhancement of T cell activation, while CJNR26 and CJMF3 exhibit immunopotentiation through the increment of B cell activation.     

HD39 (B3), a B lineage-restricted antigen whose cell surface expression is limited to resting and activated human B lymphocytes

The B cell-restricted antigen HD39, whose cell surface expression is limited to resting and activated human B lymphocytes, is described in this report. The monoclonal antibody HD39 detects a two-chain glycoprotein with apparent molecular weights of 130,000 and 140,000. During B cell ontogeny, HD39 is first expressed in the cytoplasm of bone marrow derived pre-B cells, then appears on the cell surface of sIgM+ B cells, and finally on the majority of sIgM+ sIgD+ resting B cells. After activation in vitro, the expression of HD39 on the cell surface first increases, and then the antigen is lost as cells begin to differentiate. HD39 is weakly expressed on very few non-T cell ALL and B cell CLL, on approximately 50% of B cell lymphomas, and not on Waldenstr?m's macroglobulinemias and myelomas. In contrast, it is strongly expressed on all hairy cell leukemias. Its limited cell surface expression in B cell ontogeny suggests that HD39 may be important in the events that regulate the activation of the human resting B lymphocytes.     

Epstein-Barr virus induces normal B cell responses but defective suppressor T cell responses in patients with systemic lupus erythematosus

Epstein-Barr virus (EBV) is a polyclonal B cell activator, independent of helper T cells, which induces the generation of suppressor T cells in vivo and in vitro. Given the complexity of the immunologic abnormalities in patients with systemic lupus erythematosus (SLE), we used EBV as a tool to examine the following questions: a). Are SLE B cells primarily defective? and b). Does EBV stimulate the generation of suppressor activity from SLE T cells? It was found that B cells from SLE patients infected with EBV in vitro generate plaque-forming cell (PFC) responses that are similar to those raised by normal B cells infected with EBV within the first 14 days of culture. T cells from SLE patients, in contrast to T cells from normal individuals, cultured with autologous B cells plus EBV fail to develop the expected normal decrement of PFC during the late phase of the in vitro culture (day 14). However, B cells from SLE patients are susceptible to suppression as mixed cultures of SLE B cells and normal allogeneic T cells showed a pattern of PFC response to EBV similar to that of the co-culture of normal B cells with normal T cells. T cells from SLE patients, in analogous mixed cell cultures, failed to suppress either normal B cells or allogeneic SLE B cells. The above experiments indicate that the B cells are not intrinsically defective in SLE patients; rather, a specific T cell abnormality contributes to the lack of normal immunoregulation of certain B cell responses in SLE.     

HBx transfection limits proliferative capacity of podocytes through cell cycle regulation

Our previous studies have shown that podocyte number is significantly decreased in glomeruli of children with hepa- titis B virus （HBV）-associated glomerulonephritis. In this study, we aimed to explore whether exogenous expression of HBx protein could directly inhibit podocyte proliferation in vitro, and to investigate its role in cell cycle regulation. HBx gene was delivered into cultured mouse podocytes through an adenovirus-based vector. Cell morphology was evaluated with Wright-Giemsa staining. Cell growth and proliferation were measured by 3-（4,5-dimethylthiazol-2-yl）- 2,5-diphenyltetrazolium bromide （MTT） and 5,6-carboxy- fluorescein diacetate, succinimidyl ester （CFSE）-based pro- liferation assays. Cell cycle phase was analyzed by flow cytometry, and the expression of cell cycle regulatory pro- teins was examined by western blot analysis. It was found that the aberrant nuclear changes like double and multiple micronuclei, which reflect mitotic catastrophe, accumulated in podocytes after 5 days post-infection. MTT assay showed that Ad.HBx-infected podocytes grew much more slowly than controls at day 4 post-infection and thereafter. Furthermore, CFSE-based proliferation assay also showed that the prolifer- ation of HBx-expressing podocytes was significantly inhibited than that of controls at 3-day post-infection, and that the dif- ference became much more obvious at day 5 post-infection. Cell cycle analysis showed that the transfection of HBx resulted in significant up-regulation of both cyclin B1 and CDK-inhibitor p21 expression and G2/M phase arrest, and slight down-regulation of cyclin A expression. These results demonstrated that exogenous expression of HBx might limit the proliferative capacity of podocytes through cell cycle regu- lation, thus suggesting that HBx may play a role in podocyte injuries in HBV-associated glomerulonephritis.     

A2B5+/GFAP+ Cells of Rat Spinal Cord Share a Similar Lipid Profile with Progenitor Cells: A Comparative Lipidomic Study

The central nervous system (CNS) harbors multiple glial fibrillary acidic protein (GFAP) expressing cell types. In addition to the most abundant cell type of the CNS, the astrocytes, various stem cells and progenitor cells also contain GFAP+ populations. Here, in order to distinguish between two types of GFAP expressing cells with or without the expression of the A2B5 antigens, we performed lipidomic analyses on A2B5+/GFAP+ and A2B5?/GFAP+ cells from rat spinal cord. First, A2B5+/GFAP? progenitors were exposed to the leukemia inhibitory factor (LIF) or bone morphogenetic protein (BMP) to induce their differentiation to A2B5+/GFAP+ cells or A2B5?/GFAP+ astrocytes, respectively. The cells were then analyzed for changes in their phospholipid, sphingolipid or acyl chain profiles by mass spectrometry and gas chromatography. Compared to A2B5+/GFAP? progenitors, A2B5?/GFAP+ astrocytes contained higher amounts of ether phospholipids (especially the species containing arachidonic acid) and sphingomyelin, which may indicate characteristics of cellular differentiation and inability for multipotency. In comparison, principal component analyses revealed that the lipid composition of A2B5+/GFAP+ cells retained many of the characteristics of A2B5+/GFAP? progenitors, but their lipid profile was different from that of A2B5?/GFAP+ astrocytes. Thus, our study demonstrated that two GFAP+ cell populations have distinct lipid profiles with the A2B5+/GFAP+ cells sharing a phospholipid profile with progenitors rather than astrocytes. The progenitor cells may require regulated low levels of lipids known to mediate signaling functions in differentiated cells, and the precursor lipid profiles may serve as one measure of the differentiation capacity of a cell population.