Korean J Physiol Pharmacol.  2019 Mar;23(2):103-111. 10.4196/kjpp.2019.23.2.103.

Beneficial effects of andrographolide in a rat model of autoimmune myocarditis and its effects on PI3K/Akt pathway

• Zhang Q
• Hu LQ
• Li HQ
• Wu J
• Bian NN
• Yan G

• KMID: 2438084
• DOI: http://doi.org/10.4196/kjpp.2019.23.2.103

Abstract

The study is to investigate effects of andrographolide on experimental autoimmune myocarditis (EAM). Lewis rats were immunized on day 0 with porcine cardiac myosin to establish EAM. The EAM rats were treated with either andrographolide (25, 50, 100 mg/kg/day) or vehicle for 21 days. An antigen-specific splenocytes proliferation assay was performed by using the cells from control rats immunized with cardiac myosin. Survival rates, myocardial pathology and myocardial functional parameters (left ventricle end-diastolic pressure, ± dP/dt and left ventricular internal dimension) of EAM rats received andrographolide were significantly improved. Andrographolide treatment caused an decrease in the infiltration of CD3⁺ and CD14⁺ positive cells in myocardial tissue. Moreover, andrographolide treatment caused a reduction in the plasma levels of tumor necrosis factor-alpha, interleukin-17 (IL-17) and myosin-antibody, and an increase in the level of IL-10 in EAM rats. Oral administration of andrographolide resulted in the decreased expression of p-PI3K, p-Akt without any change of PI3K and Akt. Further results indicate andrographolide significantly inhibited myosin-induced proliferation in splenocytes, and this effect was inhibited by co-treatment of SC79 (Akt activator). Our data indicate andrographolide inhibits development of EAM, and this beneficial effect may be due to powerful anti-inflammatory activity and inhibitory effect on PI3K/Akt pathway.

Keyword

Akt; Andrographolide; Autoimmune myocarditis; PI3K

MeSH Terms

Administration, Oral
Animals
Cardiac Myosins
Interleukin-10
Interleukin-17
Models, Animal*
Myocarditis*
Pathology
Plasma
Rats*
Survival Rate
Tumor Necrosis Factor-alpha
Cardiac Myosins
Interleukin-10
Interleukin-17
Tumor Necrosis Factor-alpha

Figure

• 

  Fig. 1 Effects of andrographolide on mortality of EAM rats.(A) Survival curves of EAM rats. (B) Representative echocardiogram of M-mode on day 21. Control group includes six rats, whereas other groups include ten rats for each. Con: control; EAM: experimental autoimmune myocarditis; Los: losartan.

• 

  Fig. 2 Effects of andrographolide on myocardial histopathology of EAM rats.(A) Con group; (B) EAM group; (C) andrographolide 25 mg/kg; (D) andrographolide 50 mg/kg; (E) andrographolide 100 mg/kg; (F) losartan 10 mg/kg; (G) pathological scores were evaluated by professional staff. Results are presented as the mean ± S.D. N = 6–9. Con: control; EAM: experimental autoimmune myocarditis; Los: losartan. ##p < 0.01, compared with Con; *p < 0.05, **p < 0.01, compared with EAM.

• 

  Fig. 3 Effects of andrographolide on the numbers of CD3+ positive cells in myocardium of EAM rats.(A) Con group; (B) EAM group; (C) andrographolide 25 mg/kg; (D) andrographolide 50 mg/kg; (E) andrographolide 100 mg/kg; (F) the numbers of CD3+ positive cells were determined in ten randomly selected fields by a professional staff, who didn't know grouped details. Results are presented as the mean ± S.D. N = 6–9. Con: control; EAM: experimental autoimmune myocarditis; ##p < 0.01, compared with Con; *p < 0.05, **p < 0.01, compared with EAM.

• 

  Fig. 4 Effects of andrographolide on the numbers of CD14+ positive cells in myocardium of EAM rats.(A) Con group; (B) EAM group; (C) andrographolide 25 mg/kg; (D) andrographolide 50 mg/kg; (E) andrographolide 100 mg/kg; (F) the numbers of CD14+ positive cells were determined in ten randomly selected fields by a professional staff, who didn't know grouped details. Results are presented as the mean ± S.D. N = 6–9. Con: control; EAM: experimental autoimmune myocarditis; ##p < 0.01, compared with Con; *p < 0.05,**p < 0.01, compared with EAM.

• 

  Fig. 5 Effects of andrographolide on the levels of inflammatory cytokines and myosin antibody in EAM rats.(A) TNF-alpha; (B) IL-10; (C) IL-17; (D) myosin antibody. Results are presented as the mean ± S.D. N = 6–9. EAM: experimental autoimmune myocarditis; ##p < 0.01, compared with Control; *p < 0.05, **p < 0.01, compared with EAM.

• 

  Fig. 6 Effects of andrographolide on PI3K/Akt pathway.(A) Representative stripes; (B, C) the stripes were quantitatively analyzed by Image J. Results are showed as the mean ± S.D. N = 6–9. Con: control; EAM: experimental autoimmune myocarditis. ##p < 0.01, compared with Con; **p < 0.01, compared with EMA.

• 

  Fig. 7 Effects of andrographolide on proliferation of myosin-treated splenetic cells.(A) Splenetic cells were collected from EAM rats, and treated with andrographolide in the presence and absence of SC79 for 48 h. Myosin-specific proliferative responses were measured by MTT colorimetric technique. (B1) typical p-Akt stripes; (B2) the stripes were quantitatively analyzed by image j. Results are presented as the mean ± S.D. N = 3. Con: control; EAM: experimental autoimmune myocarditis. ##p < 0.01, compared with splenetic cells without treatment; *p < 0.05, **p < 0.01, compared with splenetic cells only treated with myosin (20 µg/ml); Δp < 0.05, compared with the cells treated with myosin and andrographolide (40 µM).

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