PTEN/PI3k/AKT Regulates Macrophage Polarization in Emphysematous mice

Abstract

Macrophages play an important role in the pathogenesis of COPD. Macrophage polarization towards the M2 phenotype has been observed in the lung tissues of COPD patients and cigarette smokers. The molecular basis of this process remains unclear, and it has not been completely illuminated in animal models of emphysema. In our study, we combined cigarette smoke (CS) exposure with intraperitoneal injection of cigarette smoke extract (CSE) to build an emphysema model. We found by immunohistochemical staining and flow cytometry that the expression level of CD206 and the ratio of M2 to M1 macrophages was increased in emphysematous mice. We also demonstrated that decreased protein level for phosphatase and tensin homology deleted on chromosome ten (PTEN) and increased total protein levels for phosphorylation -protein kinase B (p-AKT) in the lung tissue of emphysematous mice and in CSE-treated RAW264.7 cells. In both bone marrow-derived macrophages (BMDMs) from emphysematous mice and CSE-treated RAW264.7 cells, we observed by RT-PCR that the mRNA levels of M2 macrophage-related markers and cytokines were increased. Furthermore, M1 macrophage-related markers and cytokines were decreased. Meanwhile we treated BMDMs from emphysematous mice and CSE-treated RAW264.7 cells with the phosphoinositide 3-kinase (PI3K)/Akt inhibitor (LY294002), we observed a reduction in RNA levels of M2 macrophage-related markers and cytokines. In conclusion, we confirmed that macrophage M2 polarization was induced in emphysematous mice generated by CS exposure combined with intraperitoneal injection of CSE. We also showed that M2 polarization was mediated through PTEN/PI3k/AKT pathway activation.

Changes of Foxo3a in PBMCs and its associations with stress hyperglycemia in acute obstructive suppurative cholangitis patients

“…The adherent cells continued to cultivate for 12 h. Then, PBMCs were pretreated with LY294002 (30 μM) (S1105, Selleck, USA) and SC79 (4 μg/ml) (S7863, Selleck, USA) for 1 h, followed by treatment with LPS (100 ng/ml) (L5293, SIGMA, USA) for 2 h. For S1PR2 inhibitor testing, PBMCs were pretreated with JET-013 (1 μM) (S7182, Selleck, the USA) and LPS (100 ng/ml) for 2 h. The total protein and RNA of PBMCs were collected for western blotting (WB) and real-time polymerase chain reaction (RT-PCR). Safe doses of LY294002, SC79 and JET-013 were verified in previous studies [ 39 , 41 – 42 ].…”

Characteristics and potential role of M2 macrophages in COPD

“…The COPD model-building method of CS exposure combined with CES intraperitoneal injection was first reported by our group, and the changes were similar with those of human smokers and COPD patients pathologically and physiopathologically. 13 The long-term smoking-induced mouse COPD model is the most classical method to build COPD model, and the method of CES intraperitoneal injection alone was also reported previously. 19 This new method is more efficient compared to the classical ones.…”

“…The breeding environment and details of the modeling method had been previously described. 13 In short, the whole modeling period was 28 days. Briefly, the model group mice were put to CS exposure for two cycles per day except on days 1, 12 and 23.…”

“…The preparation method of CSE was also described previously. 13 Briefly, each non-filter Furong cigarette (tar: 13 mg, nicotine: 1.0 mg, carbon monoxide: 14 mg/cigarette; China Tobacco Hunan Industrial Co. Ltd., Changsha, China) was burned, and all the smoke was dissolved in 4 mL PBS as the 100% CSE. The solution was utilized for CSE intraperitoneal injection and cell culture before being filtrated via a 0.22-μm pore filter (Thermo Fisher Scientific, Waltham, MA, USA) to remove particles and bacteria.…”

“…Lung function was measured by a small animal spirometer (PLY3211 system; Buxco Electronics, Sharon, CT, USA), following the protocol described previously by our group. 13 In brief, each mouse was weighed first and anesthetized by intraperitoneal injection of 10% chloral hydrate (3 mL/kg) before being tracheostomized. The trachea was cannulated, and the cannula was connected to a computer-controlled small animal spirometer.…”