Nitric oxide augments oridonin-induced efferocytosis by human histocytic lymphoma U937 cells via autophagy and the NF-κB-COX-2-IL-1β pathway.
We previously demonstrated that oridonin-induced autophagy enhanced efferocytosis (phagocytosis of apoptotic cells) by macrophage-like U937 cells through activation of the inflammatory pathways. In this study, exposure of U937 cells to 2.5 μM oridonin caused up-regulation of inducible nitric oxide synthase (iNOS) expression and continuous endogenous generation of nitric oxide (NO), which was reversed by pre-treatment with the inhibitors of nitric oxide synthase 1400 W (dihydrochloride) or L-NAME (hydrochloride). NO donor sodium nitroprusside (SNP) and efferocytosis irritant lipopolysaccharide (LPS) could also exert NO generation and iNOS expression. Moreover, oridonin-induced stimulation of efferocytosis was significantly suppressed by 1400 W or L-NAME. In addition, 1400 W or L-NAME impaired oridonin-induced autophagy. Inhibition of autophagy with 3-methyladenine (3MA) or Beclin-1 siRNA attenuated the uptake of apoptotic cells with a slight increase in the production of NO. The pro-inflammatory cytokine interleukin-1β (IL-1β) has been reported to be involved in oridonin-induced efferocytosis in U937 cells and interact with NO to contribute to inflammatory responses. 1400 W or L-NAME blocked the secretion of IL-1β and the activation of NF-κB and COX-2. Provision of SNP or LPS in place of oridonin resulted in the similar enhancement of efferocytosis, autophagy, the release of IL-1β and the expression of signal protein. NO augmented the oridonin-induced efferocytosis by mediating autophagy and activating the NF-κB-COX-2-IL-1β pathway. Inhibition of NF-κB or COX-2 in turn decreased the production of NO and the expression of iNOS. There exists a positive feedback loop between NO generation and NF-κB-COX-2-IL-1β pathway.
China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University , 103 Wenhua Road, Shenyang , People's Republic of China.
Linghe
L
Zang
Hao
H
He
Yuanchao
Y
Ye
Weiwei
W
Liu
Simiao
S
Fan
Shin-Ichi
S
Tashiro
Satoshi
S
Onodera
Takashi
T
Ikejima
D001343:Autophagy / Q000187:drug effects* / Q000502:physiology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D045786:Diterpenes, Kaurane / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D053583:Interleukin-1beta / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D016403:Lymphoma, Large B-Cell, Diffuse / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection; D020298:U937 Cells
D045786:Diterpenes, Kaurane; D053583:Interleukin-1beta; D016328:NF-kappa B; C011959:oridonin; D009569:Nitric Oxide; D051546:Cyclooxygenase 2