Low-dose Warfarin Functions as an Immunomodulator to Prevent Cyclophosphamide-induced NOD Diabetes

Warfarin has been used as an anticoagulant for a long time. Recently, the pleiotropic effect of warfarin has been investigated. As low-dose warfarin has been reported to have anti-inflammatory effect through suppression of IL-6 secretion and inhibit the immune-associated signal between Tyro3 and its ligand, Gas6, the effect of low-dose warfarin on autoimmune diabetes in NOD mice was examined. To investigate the anti-inflammatory effect of warfarin, IL-6 secretion by splenocytes was examined in the presence of various concentrations of warfarin. Low concentration of warfarin inhibited IL-6 secretion. mRNA expression of Rse, one of the Tyro3 receptor family members, and Gas6 were analyzed in NOD mice. It was detected in islets, splenocytes and bone-marrow derived dendritic cells. 0.25 mg/l or 0.50 mg/l of warfarin was orally administered to NOD mice as a cyclophosphamide-induced diabetes model. Oral administration of warfarin at much lower doses than those clinically used as an anticoagulant significantly reduced the degree of insulitis and diabetes incidence in this model. We previously demonstrated that anti-FasL Ab-treatment led to complete prevention of autoimmune diabetes in NOD mice. As Fas/FasL signaling is reported to be essential for cyclophosphamide-induced diabetes model, we extracted RNA from lymphocytes of the inguinal lymph nodes of anti-FasL Ab-treated NOD mice and performed real-time PCR to determine expression of Rse gene. Interestingly, the expression of Rse gene related to the blockade of Fas/FasL signaling was reduced to less than half the level of untreated mice. In conclusion, low-dose warfarin is a potential immunomodulator which can prevent autoimmune diabetes. Type 1 diabetes is a chronic autoimmune disease caused by autoreactive T cells promoting the specific destruction of insulin-producing β cells of the pancreatic islets (1,6). Nonobese diabetic (NOD) mouse is an animal model of human autoimmune diabetes (19). In the NOD mouse, diabetes develops as the result of a chronic inflammation that starts with leukocytic infiltration of islets from 3-5 weeks of age and gradually exacerbates until hyperglycemia develops after 16 weeks of age in a high percentage of female mice. Warfarin has been widely used for a long time as an oral anticoagulant agent. In addition, Kater et al. reported the pleiotropic effect of low-dose warfarin related with inflammation, demonstrating that low-dose warfarin inhibited inflammatory signal transduction through suppression of TNF-α induced IL-6 secretion from murine macrophages (12)

Role of FBXW7 in the quiescence of gefitinib-resistant lung cancer stem cells in EGFR-mutant non-small cell lung cancer

Several recent studies suggest that cancer stem cells (CSCs) are involved in intrinsic resistance to cancer treatment. Maintenance of quiescence is crucial for establishing resistance of CSCs to cancer therapeutics. F-box/WD repeat-containing protein 7 (FBXW7) is a ubiquitin ligase that regulates quiescence by targeting the c-MYC protein for ubiquitination. We previously reported that gefitinib-resistant persisters (GRPs) in EGFR-mutant non-small cell lung cancer (NSCLC) cells highly expressed octamer-binding transcription factor 4 (Oct-4) as well as the lung CSC marker CD133, and they exhibited distinctive features of the CSC phenotype. However, the role of FBXW7 in lung CSCs and their resistance to epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors in NSCLC is not fully understood. In this study, we developed GRPs from the two NSCLC cell lines PC9 and HCC827, which express an EGFR exon 19 deletion mutation, by treatment with a high concentration of gefitinib. The GRPs from both PC9 and HCC827 cells expressed high levels of CD133 and FBXW7, but low levels of c-MYC. Cell cycle analysis demonstrated that the majority of GRPs existed in the G0/G1 phase. Knockdown of the FBXW7 gene significantly reduced the cell number of CD133-positive GRPs and reversed the cell population in the G0/G1-phase. We also found that FBXW7 expression in CD133-positive cells was increased and c-MYC expression was decreased in gefitinib-resistant tumors of PC9 cells in mice and in 9 out of 14 tumor specimens from EGFR-mutant NSCLC patients with acquired resistance to gefitinib. These findings suggest that FBXW7 plays a pivotal role in the maintenance of quiescence in gefitinib-resistant lung CSCs in EGFR mutation-positive NSCLC

Combined insulin B:9-23 self-peptide and polyinosinic-polycytidylic acid accelerate insulitis but inhibit development of diabetes by increasing the proportion of CD4+Foxp3+ regulatory T cells in the islets in non-obese diabetic mice.

Insulin peptide B:9-23 is a major autoantigen in type 1 diabetes. Combined treatment with B:9-23 peptide and polyinosinic-polycytidylic acid (poly I:C), but neither alone, induce insulitis in normal BALB/c mice. In contrast, the combined treatment accelerated insulitis, but prevented diabetes in NOD mice. Our immunofluorescence study with anti-CD4/anti-Foxp3 revealed that the proportion of Foxp3 positive CD4(+)CD25(+) regulatory T cells (Tregs) was elevated in the islets of NOD mice treated with B:9-23 peptide and poly I:C, as compared to non-treated mice. Depletion of Tregs by anti-CD25 antibody hastened spontaneous development of diabetes in non-treated NOD mice, and abolished the protective effect of the combined treatment and conversely accelerated the onset of diabetes in the treated mice. These results indicate that poly I:C combined with B:9-23 peptide promotes infiltration of both pathogenic T cells and predominantly Tregs into the islets, thereby inhibiting progression from insulitis to overt diabetes in NOD mice

Optimal lipid-lowering therapy in patients who were functionally deferred percutaneous coronary intervention

Abstract Deferral of percutaneous coronary intervention (PCI) for functionally insignificant stenosis, defined as fractional flow reserve (FFR) > 0.80, is associated with favorable long-term prognoses. The lower-the-better strategy for low-density lipoprotein cholesterol (LDL-C) management is an established non-angioplasty therapy to improve the clinical outcomes of patients undergoing PCI. We examined the optimal LDL-C management in cases of intermediate coronary stenosis with deferred PCI on the basis of FFR values. This observational study included 273 consecutive patients with a single target vessel and deferred PCI with an FFR > 0.80. Patients with an FFR of 0.81–0.85 (n = 93) and those with FFR > 0.85 (n = 180) were classified into the lower ( 0.85 (log-rank, p = 0.003). In patients with an FFR of 0.81–0.85, the lower LDL-C group showed a significantly lower MACCE rate than the higher LDL-C group (log-rank, p = 0.006). However, the event rate did not differ significantly between the two groups in patients with FFR > 0.85 (log-rank, p = 0.84). Uncontrolled LDL-C levels were associated with higher MACCE rates in cases with deferred PCI due to an FFR of 0.81–0.85. This high-risk population for adverse cardiovascular events should receive strict LDL-C-lowering therapy