Korean J Physiol Pharmacol.  2018 Jul;22(4):369-377. 10.4196/kjpp.2018.22.4.369.

Isoliquiritigenin attenuates spinal tuberculosis through inhibiting immune response in a New Zealand white rabbit model

• Wang W
• Yang B
• Cui Y
• Zhan Y

• KMID: 2414261
• DOI: http://doi.org/10.4196/kjpp.2018.22.4.369

Abstract

Spinal tuberculosis (ST) is the tuberculosis caused by Mycobacterium tuberculosis (Mtb) infections in spinal curds. Isoliquiritigenin 4,2"²,4"²-trihydroxychalcone, ISL) is an anti-inflammatory flavonoid derived from licorice (Glycyrrhiza uralensis), a Chinese traditional medicine. In this study, we evaluated the potential of ISL in treating ST in New Zealand white rabbit models. In the model, rabbits (n=40) were infected with Mtb strain H37Rv or not in their 6th lumbar vertebral bodies. Since the day of infection, rabbits were treated with 20 mg/kg and 100 mg/kg of ISL respectively. After 10 weeks of treatments, the adjacent vertebral bone tissues of rabbits were analyzed through Hematoxylin-Eosin staining. The relative expression of Monocyte chemoattractant protein-1 (MCP-1/CCL2), transcription factor κB (NF-κB) p65 in lymphocytes were verified through reverse transcription quantitative real-time PCR (RT-qPCR), western blotting and enzyme-linked immunosorbent assays (ELISA). The serum level of interleukin (IL)-2, IL-4, IL-10 and interferon γ (IFN-γ) were evaluated through ELISA. The effects of ISL on the phosphorylation of IκBα, IKKα/β and p65 in NF-κB signaling pathways were assessed through western blotting. In the results, ISL has been shown to effectively attenuate the granulation inside adjacent vertebral tissues. The relative level of MCP-1, p65 and IL-4 and IL-10 were retrieved. NF-κB signaling was inhibited, in which the phosphorylation of p65, IκBα and IKKα/β were suppressed whereas the level of IκBα were elevated. In conclusion, ISL might be an effective drug that inhibited the formation of granulomas through downregulating MCP-1, NF-κB, IL-4 and IL-10 in treating ST.

Keyword

Granuloma; Inflammatory cytokines; MCP-1; NF-κB; Spinal tuberculosis

MeSH Terms

Blotting, Western
Bone and Bones
Chemokine CCL2
Enzyme-Linked Immunosorbent Assay
Glycyrrhiza
Granuloma
Interferons
Interleukin-10
Interleukin-4
Interleukins
Lymphocytes
Medicine, Chinese Traditional
Mycobacterium tuberculosis
New Zealand*
Phosphorylation
Rabbits
Real-Time Polymerase Chain Reaction
Reverse Transcription
Transcription Factors
Tuberculosis
Tuberculosis, Spinal*
Chemokine CCL2
Interferons
Interleukin-10
Interleukin-4
Interleukins
Transcription Factors

Figure

• 

  Fig. 1 Hematoxylin-eosin staining of adjacent vertebral bone tissues of rabbits.(A) Sham operated control group. (B) Spinal tuberculosis experimental group. (C) Spinal tuberculosis experimental with 20 mg/kg Isoliquiritigenin treatment. (D) Spinal tuberculosis experimental with 100 mg/kg Isoliquiritigenin treatment. Arrow: the granulomatous inflammation with Langhans giant cells surrounding the caseous necrosis.

• 

  Fig. 2 The effect of ISL on the expression of monocyte chemotactic protein 1 (MCP-1) and nuclear factor kappa B (NF-κB) in lymphocytes.(A) Relative mRNA expression of monocyte chemotactic protein 1 (MCP-1) and nuclear factor kappa B (NF-κB), (B) protein expressions of MCP-1 and NF-κB detected by ELISA, (C) Western blotting of the protein expressions of MCP-1 and NF-κB, (D) Protein expressions of MCP-1 and NF-κB. *p<0.05 compared to the sham control groups, #p<0.05 compared to spinal tuberculosis experimental group (n=10). Control, sham operated control group; ST, Spinal tuberculosis experimental group; 20 mg/kg, Spinal tuberculosis experimental with 20 mg/kg ISL treatment; 100 mg/kg, Spinal tuberculosis experimental with 100 mg/kg ISL treatment.

• 

  Fig. 3 The effect of ISL on serum levels of IFN-γ, IL-2, IL-4 and IL-10 detected by ELISA.*p<0.05 compared to the sham control groups, #p<0.05 compared to spinal tuberculosis experimental group (n=10). Control, sham operated control group; ST, Spinal tuberculosis experimental group; 20 mg/kg, Spinal tuberculosis experimental with 20 mg/kg ISL treatment; 100 mg/kg, Spinal tuberculosis experimental with 100 mg/kg ISL treatment.

• 

  Fig. 4 The effect of ILG on degradation and phosphorylation of IκBα.(A) Western blotting of the protein expressions of IκBα and P-IκBα in lymphocytes, (B) IκBα/β-actin relative expression level, (C) P-IκBα/β-actin relative expression level. *p<0.05 compared to the sham control groups, #p<0.05 compared to spinal tuberculosis experimental group (n=10). Control: sham operated control group, ST: Spinal tuberculosis experimental group, 20 mg/kg: Spinal tuberculosis experimental with 20 mg/kg ISL treatment, 100 mg/kg: Spinal tuberculosis experimental with 100 mg/kg ISL treatment.

• 

  Fig. 5 The effect of ISL on phosphorylation of the nuclear translocation of NF-κB subunit p65 and IKKα/β in lymphocytes.(A) phosphorylation of the protein expressions of p65 and IKKα/β detected by western blotting, (B) P-p65/β-actin relative expression level, (C) IKKα/β/β-actin relative expression level. *p<0.05 compared to the sham control groups, #p<0.05 compared to spinal tuberculosis experimental group (n=10). Control, Sham operated control group; ST, Spinal tuberculosis experimental group; 20 mg/kg, Spinal tuberculosis experimental with 20 mg/kg ISL treatment; 100 mg/kg, Spinal tuberculosis experimental with 100 mg/kg ISL treatment.

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