Systematical Identification of the Protective Effect of Danhong Injection and BuChang NaoXinTong Capsules on Transcription Factors in Cerebral Ischemia Mice Brain

Abstract

Cerebral ischemia has led to a high rate of both disability and mortality with massive healthcare costs. Although transcriptional regulation is typically mediated by different combinations of TFs, a combined regulatory unit to synergistically activate transcription has remained unclear in cerebral ischemia, especially in different drug treatments. In this study, TFs alterations after 6 h cerebral ischemic injury and repair were performed by a concatenated tandem array of consensus transcription factor response elements (catTFREs), and vital TFs were obtained by TFs-target imbalanced network. Drug intervention used Danhong injection (DHI) and BNC (BuChang NaoXinTong Capsules), which has been widely prescribed in Chinese herb medicine for the treatment of cerebrovascular and cardiovascular diseases. There were 198 TFs identified after 6 h MCAO operation, and six TFs (Sox2, Smad3, FoxO1, Creb1, Egr,1 and Smad4) were considered as critical TFs in response to cerebral ischemia. Moreover, Smad3 was identified as a hub TF among six vital TFs, and the transcription activity of Smad3 was further verified. These 6 TFs were all reversed by DHI or BNC, indicating different medications may regulate different transcription factors through TF synergy. Moreover, validation results indicated that Smad3 was a putative target TF for DHI and BNC-mediated protection against cerebral ischemia. The observations of the present study provide a fresh understanding of biomolecules and possible new avenues for therapeutic interventions, in addition to the new intervention pattern for different treatments for ischemia stroke.

Figure 1

Preventive effect of Ginaton, DHI, and BNC on MCAO mice: neurological score (a), infarct rate (b), and TTC staining of the brain (c). ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001, the model group versus the sham group; ^#p < 0.05, ^##p < 0.01, ^###p < 0.001, the BNC group or DHI group or Ginaton group versus the MCAO group.

Figure 2

The transcriptional activity of TFs in response to MCAO-induced cerebral ischemia. The number of unique peptides identified of peptides in each group (a). Venn of TFs in the sham and model groups (b). Upregulation and downregulation of differently expressed TFs in the model group compared with the sham group (c). Overlapped items of biological process in both the sham and model groups (d).

Figure 3

The protective effect of BNC and DHI on transcriptional activity. Upregulation and downregulation of BNC and DHI (a). Venn of TFs in the sham, model, BNC, and DHI groups (b). Hierarchical clustering of the quantitative information of TFs in the four groups (c). The biological process of different expressions TFs in the four groups (d).

Figure 4

Potential TF synergy of DHI and BNC. TFs-targets imbalanced network related to ischemia (a). GO analysis of six vital TFs (b). PPI network of six vital TFs (c). Verification of Smad3 by EMSA and the value of downstream effector Smad4 mRNA by qPCR (d).