ABSTRACT
Ovarian cancer (OC) is the most common gynecological malignancy. Chemotherapy failure is a major challenge in OC treatment. Targeting autophagy is a promising strategy to enhance the cytotoxicity of chemotherapeutic agents. In this study, we found that costunolide (CTD) inhibits autophagic flux and exhibits high therapeutic efficacy for OC treatment in an in vitro model. Mechanistically, CTD inactivates AMPK/mTOR signaling to inhibit autophagy initiation at the early stage and blocks mTORC1-dependent autophagosome-lysosome fusion at the late stage during autophagy by disrupting SNARE complex (STX17-SNAP29-VAMP8) formation, resulting in lethal autophagy arrest in OC cells. Furthermore, CTD sensitizes OC cells to cisplatin (CDDP) by blocking CDDP-induced autophagy both in vitro and in vivo. Together, our data provide novel mechanistic insights into CTD-induced autophagy arrest and suggest a new autophagy inhibitor for effective treatment of OC.
Subject(s)
Cisplatin , Ovarian Neoplasms , Humans , Female , Cisplatin/pharmacology , Cisplatin/metabolism , AMP-Activated Protein Kinases/metabolism , Lysosomes/metabolism , Signal Transduction , Autophagy , TOR Serine-Threonine Kinases/metabolism , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/metabolism , Qb-SNARE Proteins/metabolism , Qb-SNARE Proteins/pharmacology , Qc-SNARE Proteins/metabolism , Qc-SNARE Proteins/pharmacology , R-SNARE Proteins/metabolismABSTRACT
Respiratory burst function of neutrophils is thought to play a pivotal role in the development of pathologies such as indirect (extra-pulmonary) acute lung injury (iALI), as well as sepsis. The current study was conducted to determine the effect of an HIV transactivator of transcription (TAT)-fusion protein containing a soluble N-ethylmaleimide-sensitive factor attachment protein receptor domain from synaptosome-associated protein-23 (SNAP-23) on the shock/sepsis- and sepsis-enhanced neutrophil burst capacity using the clinical relevant two-hit iALI mouse model and the classical cecal ligation and puncture (CLP) septic model. TAT-SNAP-23 significantly decreased the blood neutrophil respiratory burst in vitro, and also in vivo in CLP and hemorrhaged mice. We found that the neutrophil influx to the lung tissue, as measured by myeloperoxidase levels and neutrophil-specific esterase(+) cells, was also decreased in the TAT-SNAP-23-treated group. Consistent with this, treatment of TAT-SNAP-23 significantly reduced the disruption of lung tissue architecture and protein concentration of bronchoalveolar lavage fluid in iALI mice compared with vehicle-treated iALI mice. In addition, although TAT-SNAP-23 did not alter the extent of local cytokine/chemokine expression, the in vitro migration capacity of neutrophils was blunted from septic and hemorrhagic mice. These data support our hypothesis that TAT-SNAP-23 reduces neutrophil dysfunction in iALI and sepsis by inhibiting neutrophil respiratory burst.
Subject(s)
Acute Lung Injury/etiology , Gene Products, tat/pharmacology , Neutrophils/drug effects , Qb-SNARE Proteins/pharmacology , Qc-SNARE Proteins/pharmacology , Sepsis/complications , Shock, Septic/complications , Acute Lung Injury/physiopathology , Animals , Bronchoalveolar Lavage Fluid , Chemotaxis, Leukocyte/drug effects , Hemorrhage/immunology , Mice , Mice, Inbred C57BL , N-Ethylmaleimide-Sensitive Proteins/pharmacology , Peroxidase/metabolism , Respiratory Burst/drug effects , Sepsis/physiopathology , Shock, Septic/physiopathologyABSTRACT
Plasma cells (PC) are B-lymphocytes terminally differentiated in a postmitotic state, with the unique purpose of manufacturing and exporting Igs. Despite the importance of this process in the survival of vertebrates, no studies have been made to understand the molecular events that regulate Ig exocytosis by PC. The present study explores the possible presence of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) system in human PC, and examines its functional role in Ig secretion. Syntaxin-2, Syntaxin-3, Syntaxin-4, vesicle-associated membrane protein (VAMP)-2, VAMP-3, and synaptosome-associated protein (SNAP)-23 could be readily detected in normal human PC obtained from intestinal lamina propria and blood, as well as in human PC lines. Because SNAP-23 plays a central role in SNAREs complex formation, it was chosen to examine possible functional implications of the SNARE system in PC Ig secretion. When recombinant SNAP-23 fusion protein was introduced into the cells, a complete abolishment of Ig production was observed in the culture supernatants of PC lines, as well as in those of normal PC. These results provide insights, for the first time, into the molecular machinery of constitutive vesicular trafficking in human PC Ig secretion and present evidence indicating that at least SNAP-23 is essential for Ab production.
