BackgroundChloroquine (CQ), the worldwide used anti-malarial drug, has recently being focused as a potential anti-cancer agent as well as a chemosensitizer when used in combination with anti-cancer drugs. It has been shown to inhibit cell growth and/or to induce cell death in various types of cancer. 5-Fluorouracil (5-FU) is the chemotherapeutic agent of first choice in colorectal cancer, but in most cases, resistance to 5-FU develops through various mechanisms. Here, we focused on the combination of CQ as a mechanism to potentiate the inhibitory effect of 5-FU on human colon cancer cells.MethodsHT-29 cells were treated with CQ and/or 5-FU, and their proliferative ability, apoptosis and autophagy induction effects, and the affection of the cell cycle were evaluated. The proliferative ability of HT-29 was analyzed by the MTS assay. Apoptosis was quantified by flow-cytometry after double-staining of the cells with AnnexinV/PI. The cell cycle was evaluated by flow-cytometry after staining of cells with PI. Autophagy was quantified by flow-cytometry and Western blot analysis. Finally, to evaluate the fate of the cells treated with CQ and/or 5-FU, the colony formation assay was performed.Results5-FU inhibited the proliferative activity of HT-29 cells, which was mostly dependent on the arrest of the cells to the G0/G1-phase but also partially on apoptosis induction, and the effect was potentiated by CQ pre-treatment. The potentiation of the inhibitory effect of 5-FU by CQ was dependent on the increase of p21Cip1 and p27Kip1 and the decrease of CDK2. Since CQ is reported to inhibit autophagy, the catabolic process necessary for cell survival under conditions of cell starvation or stress, which is induced by cancer cells as a protective mechanism against chemotherapeutic agents, we also analyzed the induction of autophagy in HT-29. HT-29 induced autophagy in response to 5-FU, and CQ inhibited this induction, a possible mechanism of the potentiation of the anti-cancer effect of 5-FU.ConclusionOur findings suggest that the combination therapy with CQ should be a novel therapeutic modality to improve efficacy of 5-FU-based chemotherapy, possibly by inhibiting autophagy-dependent resistance to chemotherapy.

IL-17A is a T cell-derived proinflammatory cytokine that contributes to the pathogenesis of rheumatoid arthritis. Recently, six related molecules have been identified to form the IL-17 family, as follows: IL-17A, IL-17B, IL-17C, IL-17D, IL-17E, and IL-17F. Whereas IL-17A and IL-17F up-regulate IL-6 in synovial fibroblasts, IL-17B and IL-17C are reported to stimulate the release of TNF-α and IL-1β from the monocytic cell line, THP-1 cell. However, their detailed function remains to be elucidated. We report in this study the effects of IL-17 family on the collagen-induced arthritis (CIA) progression by T cell gene transfer and bone marrow chimeric mice. The mRNA expressions of IL-17 family (IL-17A, IL-17B, IL-17C, and IL-17F) and their receptor (IL-17R and IL-17Rh1) genes in the arthritic paws of CIA mice were elevated compared with controls. Although IL-17A and IL-17F were expressed in CD4+ T cells, IL-17B and IL-17C were expressed in the cartilage and in various cell populations in the CIA arthritic paws, respectively. In vitro, IL-17A, IL-17B, IL-17C, and IL-17F induced TNF-α production in mouse peritoneal exudate cells. In vivo, adoptive transfer of IL-17B- and IL-17C-transduced CD4+ T cells evidently exacerbated arthritis. Bone marrow chimeric mice of IL-17B and IL-17C exhibited elevated serum TNF-α concentration and the high arthritis score upon CIA induction. Moreover, neutralization of IL-17B significantly suppressed the progression of arthritis and bone destruction in CIA mice. Therefore, not only IL-17A, but also IL-17B and IL-17C play an important role in the pathogenesis of inflammatory arthritis.

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Human neutrophil antigens (HNAs) play an important role in a variety of clinical conditions including immune‐mediated neutropenia, non‐hemolytic transfusion reactions, and transfusion‐related acute lung injury. The aim of this study was to investigate the frequency distribution of HNAs‐1 to ‐5 among the Japanese population. We analyzed samples from 570 healthy Japanese by molecular and serologic techniques to estimate the gene frequencies of HNAs‐1 to ‐5. DNA samples were obtained and typed for the HNA‐1 (n = 523), ‐3 (n = 570), ‐4 (n = 570), and ‐5 (n = 508), by molecular techniques. The HNA‐1 genotype was determined by using a commercial polymerase chain reaction‐reverse sequence‐specific oligonucleotide probes (PCR‐rSSOP) kit. The HNA‐3 to ‐5 genotypes were determined by the PCR‐sequence specific primer (PCR‐SSP), previously described, with a small modification. The HNA‐2a phenotype was determined in 301 donors by granulocyte immunofluorescence test. In Japanese, the gene frequencies of HNA‐1a, ‐1b, and ‐1c were 0.623, 0.377, and 0.000, respectively. The frequency of HNA‐2a phenotype was 0.987, and the gene frequencies of HNA‐3a and ‐3b were 0.654 and 0.346, respectively. HNA‐4a and ‐4b were found at 1.000 and 0.000, respectively, and HNA‐5a and ‐5b at 0.840 and 0.160, respectively. We describe, for the first time, the frequencies of all HNAs (HNA‐1 to ‐5) among the Japanese population. This study will be helpful for the prediction of the risk of alloimmunization to HNA, especially to determine the risk of HNA alloantibody production by transfusion of HNA incompatible blood and feto‐maternal incompatibility.

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The first major event when a medical device comes in contact with blood is the adsorption of plasma proteins. Protein adsorption on the material surface leads to the activation of the blood coagulation cascade and the inflammatory process, which impair the lifetime of the material. Various efforts have been made to minimize protein adsorption and platelet adhesion. Recently, diamond-like carbon (DLC) has received much attention because of their antithrombogenicity. We recently reported that coating silicon substrates with fluorine-doped diamond-like carbon (F-DLC) drastically suppresses platelet adhesion and activation. Here, we evaluated the protein adsorption on the material surfaces and clarified the relationship between protein adsorption and platelet behaviors, using polycarbonate and DLC- or F-DLC-coated polycarbonate. The adsorption of albumin and fibrinogen were assessed using a colorimetric protein assay, and platelet adhesion and activation were examined using a differential interference contrast microscope. A higher ratio of albumin to fibrinogen adsorption was observed on F-DLC than on DLC and polycarbonate films, indicating that the F-DLC film should prevent thrombus formation. Platelet adhesion and activation on the F-DLC films were more strongly suppressed as the amount of fluorine doping was increased. These results show that the F-DLC coating may be useful for blood-contacting devices.