Journal
BMC PULMONARY MEDICINE
Volume 16, Issue -, Pages -Publisher
BIOMED CENTRAL LTD
DOI: 10.1186/s12890-016-0247-8
Keywords
MK2; HuR; ICAM-1; IL-8; HPMEC
Categories
Funding
- National Natural Science Foundation of China [81270138, 81300052]
- Natural Science Foundation of Jiangsu Province [BK20130402]
- Medicine and Technology Foundation of Chinese PLA [CWS12J008]
- China Postdoctoral Science Foundation [2015 M570420]
- Jiangsu provincial health and Family Planning Commission Science Foundation [H 201558]
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Background: Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are characterized by acute lung inflammation. Intercellular adhesion molecule-1 (ICAM-1) and interleukin-8 (IL-8) play an important role in the development of these diseases. Mitogen-activated protein kinase (MAPK) p38/activated protein kinase 2 (MK2) regulates the expression of ICAM-1 and IL-8 in human lung microvascular endothelial cells (HPMECs) stimulated by tumor necrosis factor-a (TNF-alpha); however, the underlying molecular mechanism remains unclear. Here, we show that human antigen R (HuR), an RNA binding protein which binds preferentially to AU-rich elements (AREs) and stabilizes mRNAs, regulates TNF-alpha-induced ICAM-1 expression in the MK2/HuR signaling pathway. Method: MK2 and HuR were silenced respectively in HPMECs and then HPMECs were stimulatied with TNF-alpha. Nucleo-cytoplasmic shuttling of HuR was detected by subcellular fractionation and confocal microscopy in MK2 knockdown HPMECs. In HuR silencing cells, protein and mRNA levels of ICAM-1 and IL-8 were measured by western blot analysis, ELISA and real-time PCR; mRNA stabilization were measured by real-time PCR after actinomycin D (ActD) blocking transcription. Furthermore, we performed neutrophil adhesion assay to assess the adhering capacity after HuR silencing. Results: MK2 were subjected to a knockdown by interfering RNA, the mRNA and protein levels of HuR in human pulmonary microvascular endothelial cells (HPMECs) were not affected. However, after the stimulation of TNF-alpha, silencing MK2 inhibited HuR accumulation to cytoplasm from nucleus in HPMECs. Consequently, knockdown of HuR by RNA interference in HPMECs, there was reduction in the stability of ICAM-1 mRNA and ICAM-1 protein level. This event was accompanied by a decrease in the adhesion of neutrophils towards HPMECs. Nevertheless, HuR silencing had no effect on the mRNA and protein levels of IL-8. Conclusion: These results indicate that MK2 post-transcriptionally regulates TNF-alpha-induced ICAM-1 expression by altering the cytoplasmic localization of HuR in HPMECs.
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