The Synergistic Mechanism of Total Saponins and Flavonoids in Notoginseng-Safflower against Myocardial Infarction Using a Comprehensive Metabolomics Strategy

Abstract

Notoginseng and safflower are commonly used traditional Chinese medicines for benefiting qi and activating blood circulation. A previous study by our group showed that the compatibility of the effective components of total saponins of notoginseng (NS) and total flavonoids of safflower (SF), named NS-SF, had a preventive effect on isoproterenol (ISO)-induced myocardial infarction (MI) in rats. However, the therapeutic effect on MI and the synergistic mechanism of NS-SF are still unclear. Therefore, integrated metabolomics, combined with immunohistochemistry and other pharmacological methods, was used to systematically research the therapeutic effect of NS-SF on MI rats and the synergistic mechanism of NS and SF. Compared to NS and SF, the results demonstrated that NS-SF exhibited a significantly better role in ameliorating myocardial damage, apoptosis, easing oxidative stress and anti-inflammation. NS-SF showed a more significant regulatory effect on metabolites involved in sphingolipid metabolism, glycine, serine, and threonine metabolism, primary bile acid biosynthesis, aminoacyl-tRNA biosynthesis, and tricarboxylic acid cycle, such as sphingosine, lysophosphatidylcholine (18:0), lysophosphatidylethanolamine (22:5/0:0), chenodeoxycholic acid, L-valine, glycine, and succinate, than NS or SF alone, indicating that NS and SF produced a synergistic effect on the treatment of MI. This study will provide a theoretical basis for the clinical development of NS-SF.

Keywords: metabolomics; myocardial infarction; notoginseng; safflower; synergistic.

Figure 1

Evaluation of the heart functions of rats in every group. (A) The M-mode echocardiograms for rats in different groups. (B) The parameters of ECG-M for rats in different groups (mean ± SD, n = 6). ^## p < 0.01, when versus sham group; * p < 0.05, ** p < 0.01, when versus model group. Normal: normal group; Sham: sham-operated group; Model: model group; Positive group: positive control group; NS: total saponins of notoginseng group; SF: total flavonoids of safflower group; NS–SF: the compatibility of NS and SF group.

Figure 2

Effects of NS, SF and NS–SF on myocardial fibrosis and apoptosis induced by MI in rat heart tissues. (A) H&E staining of left ventricular (LV) tissue showed pathological and morphological changes in different groups (magnification, ×400). The necrotic myocardium is labeled with a black arrow, and inflammatory cells are labeled with a yellow arrow. (B) The results of TUNEL staining in different groups. Arrows indicate apoptotic cardiomyocyte nuclei. (C) Apoptosis rate of myocardial cells in each group (mean ± SD, n = 6). (D) The serum levels of MI-associated biochemical markers of rats in different groups (mean ± SD, n = 6). ^## p < 0.01, when versus sham group; * p < 0.05, ** p < 0.01, when versus model group. Normal: normal group; Sham: sham-operated group; Model: model group; Positive group: positive control group; NS: total saponins of notoginseng group; SF: total flavonoids of safflower group; NS–SF: the compatibility of NS and SF group.

Figure 3

Effects of NS, SF and NS–SF on plasma anti-inflammatory and antioxidant activities of MI-induced cardiac hypertrophy in rats (mean ± SD, n = 6), (A) MDA, (B) SOD, (C) GSH-Px, (D) TNF-α, (E) IL-6, and (F) IL-1α. ^## p < 0.01, when versus sham group; p < 0.05, * p < 0.01, when versus model group. Normal: normal group; Sham: sham-operated group; Model: model group; Positive group: positive control group; NS: total saponins of notoginseng group; SF: total flavonoids of safflower group; NS–SF: the compatibility of NS and SF group.

Figure 4

The PCA score plot of rats in different groups. (A) Positive mode by UPLC−QTOF/MS. (B) Negative mode by UPLC−QTOF/MS (C) ¹H NMR. Normal: normal group; Sham: sham−operated group; Model: model group; Positive group: positive control group; NS: total saponins of notoginseng group; SF: total flavonoids of safflower group; NS–SF: the compatibility of NS and SF group.

Figure 5

The OPLS-DA of the plasma samples from sham and model rats. (A) Positive mode by UPLC-QTOF/MS. (B) A combination plot of S-plot and VIP values in positive mode. (C) Negative mode by UPLC-QTOF/MS. (D) A combination plot of S-plot and VIP values in negative mode. (E) ¹H NMR. (F) A combination plot of S-plot (green squares) and VIP (blue squares) values in ¹H NMR. Sham: sham-operated group; Model: model group.

Figure 6

Schematic representation of the effects of NS, SF, and NS–SF on the metabolic network of the vital MI-altered metabolites. Sham: sham-operated group; Model: model group; Positive group: positive control group; NS: total saponins of notoginseng group; SF: total flavonoids of safflower group; NS–SF: the compatibility of NS and SF group.