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Effects of andrographolide on the expression of procoagulant and fibrinolytic inhibition factors in rat type Ⅱ alveolar epithelial cells stimulated by lipopolysaccharide / 中华危重病急救医学
Chinese Critical Care Medicine ; (12): 155-160, 2021.
Article in Chinese | WPRIM | ID: wpr-883849
ABSTRACT

Objective:

To determine the effect of andrographolide (AD) on the expression of procoagulant and fibrinolytic inhibitory factors in rat type Ⅱ alveolar epithelial cells (AECⅡ) stimulated by lipopolysaccharide (LPS).

Methods:

The AECⅡ cells RLE-6TN in the logarithmic growth phase were divided into 5 groups the normal control (NC) group, the LPS group, and the 6.25, 12.5, and 25 mg/L AD groups (AD 6.25 group, AD 12.5 group, AD 25 group). The NC group was cultured with RPMI 1640 conventional medium. In the LPS group, 5 mg/L LPS was added to the RPMI 1640 conventional medium for stimulation. Cells in the AD groups were treated with 6.25, 12.5, and 25 mg/L AD in advance for 1 hour and then given LPS to stimulate the culture. The cells and cell culture supernatant were collected 24 hours after LPS stimulation. The protein and mRNA expressions of tissue factor (TF), tissue factor pathway inhibitor (TFPI), and plasminogen activator inhibition-1 (PAI-1) in cells were detected by Western blotting and real-time fluorescent quantitative polymerase chain reaction (RT-qPCR). The levels of procollagenpeptide (PⅢP), thrombin-antithrombin complex (TAT), antithrombin Ⅲ (AT-Ⅲ) and activated protein C (APC) in the cell supernatant were detected by enzyme linked immunosorbent assay (ELISA).

Results:

Compared with the NC group, the protein and mRNA expressions of TF and PAI-1 in the LPS group were significantly increased, and the protein and mRNA expressions of TFPI were significantly reduced. At the same time, the levels of PⅢP and TAT in the cell supernatant were significantly increased, the levels of AT-Ⅲ, APC were significantly reduced. Compared with the LPS group, the protein and mRNA expressions of TF and PAI-1 in AD 6.25 group, AD 12.5 group, AD 25 group were significantly reduced [TF/GAPDH 0.86±0.08, 0.45±0.04, 0.44±0.04 vs. 1.32±0.10, TF mRNA (2 -&Delta;&Delta;Ct) 2.59±0.25, 2.27±0.05, 1.95±0.04 vs. 4.60±0.26, PAI-1/GAPDH 2.11±0.07, 1.45±0.04, 0.86±0.09 vs. 2.56±0.09, PAI-1 mRNA (2 -&Delta;&Delta;Ct) 3.50±0.22, 2.23±0.29, 1.84±0.09 vs. 6.60±0.27, all P < 0.05], while the protein and mRNA expressions of TFPI were significantly increased [TFPI/GAPDH 0.78±0.05, 0.81±0.03, 0.84±0.07 vs. 0.36±0.02, TFPI mRNA (2 -&Delta;&Delta;Ct) 0.46±0.09, 0.69±0.07, 0.91±0.08 vs. 0.44±0.06, all P < 0.05]. Also the levels of PⅢP and TAT in the cell supernatant were significantly reduced, and the levels of AT-Ⅲ and APC were significantly increased [PⅢP (μg/L) 13.59±0.23, 12.66±0.23, 10.59±0.30 vs. 15.82±0.29, TAT (ng/L) 211.57±6.41, 205.69±4.04, 200.56±9.85 vs. 288.67±9.84, AT-Ⅲ (μg/L) 102.95±3.86, 123.92±2.63, 128.67±1.67 vs. 92.93±3.36, APC (μg/L) 1 188.95±14.99, 1 366.12±39.93, 1 451.15±29.69 vs. 1 145.55±21.07, all P < 0.05]. With the increase of the dose of AD, the above-mentioned promotion and inhibition effects became more obvious. In the AD 25 group, TF, PAI-1 protein and mRNA expressions decreased, TFPI mRNA expression increased, PⅢP level in the supernatant decreased and AT-Ⅲ, APC levels increased compared with AD 6.25 group, the difference was statistically significant, and the decrease of PAI-1 protein expression and PⅢP level in the supernatant were also statistically significant compared with AD 12.5 group.

Conclusions:

Andrographolide in the dose range of 6.25-25 mg/L can dose-dependently inhibit the expression and secretion of procoagulant and fibrinolytic inhibitor-related factors in AECⅡ cells RLE-6TN stimulated by LPS, and promote the secretion of anticoagulant factors. 25 mg/L has the most obvious effect.
Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Critical Care Medicine Year: 2021 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Critical Care Medicine Year: 2021 Type: Article