Autophagy is a major degradative process responsible for the disposal of cytoplasmic proteins and dysfunctional organelles via the lysosomal pathway. During the autophagic process, cells form double-membraned vesicles called autophagosomes that sequester disposable materials in the cytoplasm and finally fuse with lysosomes. In the present study, we investigated the inhibition of autophagy by a synthesized compound, MHY1485, in a culture system by using Ac2F rat hepatocytes. Autophagic flux was measured to evaluate the autophagic activity. Autophagosomes were visualized in Ac2F cells transfected with AdGFP-LC3 by live-cell confocal microscopy. In addition, activity of mTOR, a major regulatory protein of autophagy, was assessed by western blot and docking simulation using AutoDock 4.2. In the result, treatment with MHY1485 suppressed the basal autophagic flux, and this inhibitory effect was clearly confirmed in cells under starvation, a strong physiological inducer of autophagy. The levels of p62 and beclin-1 did not show significant change after treatment with MHY1485. Decreased co-localization of autophagosomes and lysosomes in confocal microscopic images revealed the inhibitory effect of MHY1485 on lysosomal fusion during starvation-induced autophagy. These effects of MHY1485 led to the accumulation of LC3II and enlargement of the autophagosomes in a dose- and time- dependent manner. Furthermore, MHY1485 induced mTOR activation and correspondingly showed a higher docking score than PP242, a well-known ATP-competitive mTOR inhibitor, in docking simulation. In conclusion, MHY1485 has an inhibitory effect on the autophagic process by inhibition of fusion between autophagosomes and lysosomes leading to the accumulation of LC3II protein and enlarged autophagosomes. MHY1485 also induces mTOR activity, providing a possibility for another regulatory mechanism of autophagy by the MHY compound. The significance of this study is the finding of a novel inhibitor of autophagy with an mTOR activating effect.

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Fronto-ethmoidal mucocoeles have the capacity to destroy bone. Sinus lining tissue has been obtained at surgery from patients with mucocoeles, from those with chronic sinusitis undergoing endoscopic sinus surgery and from patients undergoing craniofacial resection. Tissues have been frozen, sectioned, and subjected to immunohistochemical examination with monospecific antibodies for the presence of the potent osteolytic cytokines interleukins-1 and -6 and tumour necrosis factor alpha. In addition, the chemotactic intercrine--interleukin-8 was investigated. The presence of the cytokine-inducible vascular endothelial adhesion receptors--Inter-Cellular Adhesion Molecule (ICAM)-1 and E-Selectin (also known as Endothelial Leukocyte Adhesion Molecule--ELAM) was also determined. Normal sinus tissue showed no immunoreactivity with the antibodies to these various moieties. Surprisingly, only a small proportion of tissues from patients with chronic sinusitis showed the presence of cytokines or vascular adhesion receptors. In contrast, all specimens of fronto-ethmoidal mucocoeles showed positive staining for IL-1 alpha and beta and for ICAM-1 and E-selectin. IL-1 immunostaining was restricted to the epithelial cell population not being found in infiltrating leukocytes. In 40% of mucocoeles infiltrating macrophage-like cells showed the presence of tumour necrosis factor alpha. The presence of the potent osteolytic cytokine--IL-1 in all specimens of fronto-ethmoidal mucocoeles coupled to the finding of the IL-1-inducible adhesion molecules ICAM-1 and E-Selectin argues strongly that IL-1 is released from the epithelial cells and that this cytokine may be the factor causing the erosion of bone overlying the expanding mucocoele. The nature of the signals inducing cytokine synthesis remain, however, unidentified.

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Approximately 20% of HER2 positive breast cancer develops disease recurrence after adjuvant trastuzumab treatment. This study aimed to develop a molecular prognostic model that can reliably stratify patients by risk of developing disease recurrence. Using miRNA microarrays, nine miRNAs that differentially expressed between the recurrent and non-recurrent patients were identified. Then, we validated the expression of these miRNAs using qRT-PCR in training set (n = 101), and generated a 2-miRNA (miR-4734 and miR-150-5p) based prognostic signature. The prognostic accuracy of this classifier was further confirmed in an internal testing set (n = 57), and an external independent testing set (n = 53). Besides, by comparing the ROC curves, we found the incorporation of this miRNA based classifier into TNM stage could improve the prognostic performance of TNM system. The results indicated the 2-miRNA based signature was a reliable prognostic biomarker for patients with HER2 positive breast cancer.