Systematic identification of the interventional mechanism of Qingfei Xiaoyan Wan (QFXYW) in treatment of the cytokine storm in acute lung injury using transcriptomics-based system pharmacological analyses

Figures & data

Table 1. Primers for RT-qPCR.

Gene	Sequence (5’-3’)	PCR product (bp)
β-actin (NM_007393.3)	Forward: TGTTACCAACTGGGACGACA	165
	Reverse: GGGGTGTTGAAGGTCTCAAA
IL-6 (NM_031168.1)	Forward: TCCAGTTGCCTTCTTGGGAC	140
	Reverse: GTGTAATTAAGCCTCCGACTTG
TNF-α (NM_013693.2)	Forward: TAGCCAGGAGGGAGAACAGA	127
	Reverse: TTTTCTGGAGGGAGATGTGG
TLR2 (NM_011905.3)	Forward: AAGAGGAAGCCCAAGAAAGC	80
	Reverse: CAATGGGAATCCTGCTCACT
CCL2	Forward: CCCAATGAGTAGGCTGGAGA	125
(NM_011333.3)	Reverse: TCTGGACCCATTCCTTCTTG
CXCL1	Forward: TGTTGTGCGAAAAGAAGTGC	91
(NM_008176.3)	Reverse: TACAAACACAGCCTCCCACA
CXCL2	Forward: AAGTTTGCCTTGACCCTGAA	180
(NM_009140.2)	Reverse: AGGCACATCAGGTACGATCC
CXCL10	Forward: TCCTTGTCCTCCCTAGCTCA	124
(NM_021274.2)	Reverse: ATAACCCCTTGGGAAGATGG
IL-1β	Forward: GACCTTCCAGGATGAGGACA	183
(NM_008361.3)	Reverse: AGCTCATATGGGTCCGACAG
MMP-9	Forward: TGAATCAGCTGGCTTTTGTG	191
(NM_013599.4)	Reverse: ACCTTCCAGTAGGGGCAACT

Figure 1. Effects of QFXYW on leukocytes (A) and neutrophils (B). (Compared with the control group, ^#p < 0.01, ^##p < 0.05; compared with the ALI group, *p < 0.05; n = 6). ALI: acute lung injury; QFXYW: Qingfei Xiaoyan Wan.

Figure 2. Histopathological changes of lung tissues of mice in each group (A) Control group; (B) ALI group; (C) DEX group; (D–F) QFXYW (1.3, 2.6, and 3.9 g/kg). ALI: acute lung injury; DEX: dexamethasone; QFXYW: Qingfei Xiaoyan Wan.

Figure 3. Transcriptomics analysis of QFXYW treated ALI. (A) Volcano map of the control group; (B) Volcano map of the ALI group; (C) Heatmap of the control, DEX, QFXYW, and ALI groups. (D) Venn diagram of differentially expressed genes. ALI: acute lung injury; DEX: dexamethasone; QFXYW: Qingfei Xiaoyan Wan.

Figure 4. The process used for the screening of hub genes.

Table 2. Topological parameter of hub genes.

No.	Genes	Protein names	Target class	Degree
1	CXCL10	C-X-C motif chemokine ligand 10	Signalling	30
2	CXCL2	C-X-C motif chemokine ligand 2	Signalling	30
3	CXCL1	C-X-C motif chemokine ligand 1	Signalling	29
4	IL-6	Interleukin 6	None	24
5	CCL5	C-C motif chemokine ligand 5	Signalling	23
6	CCL4	C-C motif chemokine ligand 4	Signalling	23

Figure 5. Functional enrichment analysis of DEGs between the control vs. ALI and QFXYW vs. ALI groups. (A) Biological process (BP) of DEGs in the control vs. ALI group and QFXYW vs. ALI group; (B) Cellular component (CC) of DEGs in the control vs. ALI group and QFXYW vs. ALI group; (C) Molecular function (MF) of DEGs in the control vs. ALI group and QFXYW vs. ALI group; (D) KEGG of DEGs in the control vs. ALI group and QFXYW vs. ALI group. ALI, acute lung injury; DEG: differentially expressed genes; KEGG: Kyoto Encyclopaedia of Genes and Genomes; QFXYW: Qingfei Xiaoyan Wan.

Figure 6. Effects of the QFXYW on the mRNA expression in LPS induced ALI (A) IL-6; (B) TNF-α; (C) CCL2; (D) CXCL1; (E) MMP-9; (F) IL-10; (G) TLR2; (H) IL-1β; and (I) CXCL10 (Compared with the control group, ^#p < 0.01, ^##p < 0.05; compared with the ALI group, *p < 0.05; n = 3) LPS, lipopolysaccharide; ALI: acute lung injury; QFXYW: Qingfei Xiaoyan Wan.

Figure 7. Effect of QFXYW on the protein expression of IL-6 and TNF-α. (A) IL-6 in the serum; (B) TNF-α in the serum; (C) IL-6 in the lung tissue; (D) TNF-α in the lung tissue; (E) IL-6 in the BALF; (F) TNF-α in the BALF. BALF: bronchoalveolar lavage fluid; QFXYW: Qingfei Xiaoyan Wan.

Figure 8. Effect of QFXYW on the production of multiple cytokines in the serum of the mice with ALI. (A) Cytokines and chemokines screened using cytokine protein assay; (B) Optical density detected by Image. ALI: acute lung injury; QFXYW: Qingfei Xiaoyan Wan.

Figure 9. Effect of QFXYW on the production of multiple cytokines in the lung tissue of mice with ALI. (A) Cytokines and chemokines screened using cytokines protein assay; (B) Optical density detected by Image. ALI: acute lung injury; QFXYW: Qingfei Xiaoyan Wan.