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Objectives There are three major subtypes of breast cancer, ER+, HER2+ and triple-negative breast cancer (TNBC), namely ER−, PR−, HER2−. TNBC is the most aggressive breast cancer with poor prognosis and no target drug up to now. Actinomycin D (ActD) is a bioactive metabolite of marine bacteria that has been reported to have antitumor activity. The aim of study is to investigate whether ActD has a synergetic effect on TNBC with Doxorubicin (Dox), the major chemotherapeutic drug for TNBC, and explore the underlying mechanism. Methods TNBC cell lines HCC1937, MDA-MB-436 and nude mice were used in the study. Drug synergy determination, LDH assay, MMP assay, Hoechst 33342 staining, Flow cytometry, Flexible docking and CESTA assay were carried out. The expression of proteins associated with apoptosis was checked by Western blot and siRNA experiments were performed to investigate the role of P53 and PUMA induced by drugs. Results There was much higher apoptosis rate of cells in the ActD + Dox group than that in ActD group or Dox group. Expression of MDM2 and BCL-2 was reduced while expression of P53, PUMA and BAX were increased in the groups treated with ActD + Dox or Dox compared to the control group. Furthermore, P53 siRNA or PUMA siRNA tremendously abrogated the cell apoptosis in the groups treated by ActD, Dox and ActD + Dox. Flexible docking and CESTA showed that ActD can bind MDM2. Conclusions ActD had a synergetic effect on TNBC with Dox via P53-dependent apoptosis and it may be a new choice for treatment of TNBC. [ABSTRACT FROM AUTHOR]

Objective: To analyze the expression of type Ⅱ inflammatory-related cytokines in nasal secretions of patients with eosinophilic chronic rhinosinusitis with nasal polyps（ECRSwNP）, and to preliminarily explore the role of type Ⅱ inflammatory cytokines in nasal secretions in predicting ECRSwNP. Methods: A prospective analysis was made of 91 patients with CRSwNP who underwent endoscopic sinus surgery in Peking University Third Hospital from November 2020 to June 2021. All the selected patients had their SNOT-22 score, Lund-Mackay score and blood eosinophilia collected before surgery. Percentage and absolute value; the nasal secretions of patients were collected before operation, and enzyme-linked immunosorbent assay was used to detect the typeⅡinflammatory cytokines（IL-4, IL-5, IL-13, IL-25, IL-33, Eotaxin-3, periostin）, intraoperative nasal polyp tissue was collected for eosinophil count. According to the proportion of eosinophils in the tissue≥10%, they were divided into ECRSwNP group and nECRSwNP group. The clinical baseline data and type Ⅱ inflammatory cytokines were compared between the two groups, and the related factors of ECRSwNP were evaluated by univariate logistic regression analysis. The receiver operating characteristic（ROC） curve was used to evaluate the predictive potential of each clinical index. Results: The SNOT-22 score, Lund-Mackay score, blood eosinophil percentage and absolute value in the ECRSwNP group were higher than those in the nECRSwNP group（ P <0.05）. In the nECRSwNP group（ P <0.01）. Logistic regression analysis found that IL-5, Eotaxin-3 and blood eosinophil percentage were risk factors for ECRSwNP（ P <0.05）. ROC analysis found that IL-5, Eotaxin-3 and blood eosinophil percentage had predictive diagnostic value（ P <0.01）, among which blood eosinophil percentage had the greatest predictive value（AUC=0.756）. The prediction model composed of Eotaxin-3, SNOT-22 score, sinus CT Lund-Mackay score, blood eosinophil percentage and blood eosinophil absolute value had better prediction effect on ECRSwNP（AUC=0.873）. Conclusion: Type Ⅱ inflammatory cytokines IL-5 and Eotaxin-3 in nasal secretions may be involved as biomarkers for early diagnosis of ECRSwNP., Competing Interests: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose., (Copyright© by the Editorial Department of Journal of Clinical Otorhinolaryngology Head and Neck Surgery.)

Adenosine triphosphate (ATP) is stored as lysosomal vesicles in marginal cells of the stria vascular in neonatal rats, but the mechanisms of ATP release are unclear. Primary cultures of marginal cells from 1-day-old Sprague-Dawley rats were established. P2Y 2 receptor and inositol 1,4,5-trisphosphate (IP3) receptor were immunolabelled in marginal cells of the stria vascular. We found that 30 μM ATP and 30 μM uridine triphosphate (UTP) evoked comparable significant increases in the intracellular Ca 2+ concentration ([Ca 2+ ] i ) in the absence of extracellular Ca 2+ , whereas the response was suppressed by 100 μM suramin, 10 μM 1-(6-(17β-3-methoxyester-1,3,5(10)-trien-17-yl)amino)-hexyl)-1H-pyrrole-2,5-dione(U-73122), 100 μM 2-aminoethoxydiphenyl borate (2-APB) and 5 μM thapsigargin (TG), thus indicating that ATP coupled with the P2Y 2 R-PLC-IP3 pathway to evoke Ca 2+ release from the endoplasmic reticulum (ER). Incubation with 200 μM Gly-Phe-β-naphthylamide (GPN) selectively disrupted lysosomes and caused significant increases in [Ca 2+ ] I ; this effect was partly inhibited by P2Y 2 R-PLC-IP3 pathway antagonists. After pre-treatment with 5 μM TG, [Ca 2+ ] i was significantly lower than that after treatment with P2Y 2 R-PLC-IP3 pathway antagonists under the same conditions, thus indicating that lysosomal Ca 2+ triggers Ca 2+ release from ER Ca 2+ stores. Baseline [Ca 2+ ] i declined after treatment with the Ca 2+ chelator 50 μM bis-(aminophenolxy) ethane-N,N,N',N'-tetra-acetic acid acetoxyme-thyl ester (BAPTA-AM) and 4 IU/ml apyrase. 30 μM ATP decrease of the number of quinacrine-positive vesicles via lysosome exocytosis, whereas the number of lysosomes did not change. However, lysosome exocytosis was significantly suppressed by pre-treatment with 5 μM vacuolin-1. Release of ATP and β-hexosaminidase both increased after treatment with 200 μM GPN and 5 μM TG, but decreased after incubation with 50 μM BAPTA-AM, 4 IU/ml apyrase and 5 μM vacuolin-1. We suggest that ATP triggers Ca 2+ release from the ER, thereby contributing to secretion of lysosomal ATP via lysosomal exocytosis. Lysosomal stored Ca 2+ triggers Ca 2+ release from the ER directly though the IP3 receptors, and lysosomal ATP evokes Ca 2+ signals indirectly via the P2Y 2 R-PLC-IP3 pathway., (Copyright © 2018 Elsevier Ltd. All rights reserved.)