Zhilong Huoxue Tongyu Capsules Ameliorate Early Brain Inflammatory Injury Induced by Intracerebral Hemorrhage via Inhibition of Canonical NFкβ Signalling Pathway

Maryam Mazhar; Guoqiang Yang; Linshen Mao; Pan Liang; Ruizhi Tan; Li Wang; Houping Xu; Luyin Yang; Wei Ren; Sijin Yang

doi:10.3389/fphar.2022.850060

Zhilong Huoxue Tongyu Capsules Ameliorate Early Brain Inflammatory Injury Induced by Intracerebral Hemorrhage via Inhibition of Canonical NFкβ Signalling Pathway

Front Pharmacol. 2022 Mar 31:13:850060. doi: 10.3389/fphar.2022.850060. eCollection 2022.

Authors

Maryam Mazhar^{1

2}, Guoqiang Yang^{3

4}, Linshen Mao^{1

2}, Pan Liang¹, Ruizhi Tan³, Li Wang³, Houping Xu⁵, Luyin Yang¹, Wei Ren¹, Sijin Yang^{1

2}

Affiliations

¹ National Traditional Chinese Medicine Clinical Research Base and Drug Research Center, The Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, China.
² Institute of Integrated Chinese and Western Medicine, Southwest Medical University, Luzhou, China.
³ Research Center for Integrated Chinese and Western Medicine, The Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, China.
⁴ Research Unit of Molecular Imaging Probes, Department of Radiologic Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, Thailand.
⁵ Preventive Treatment Center, The Affiliated Traditional Chinese Medicine Hospital of Southwest Medical University, Luzhou, China.

Abstract

Background: Intracerebral hemorrhage (ICH) is a debilitating and fatal condition with continuously rising incidence globally, without effective treatment available. Zhilong Huoxue Tongyu (ZLHXTY) capsule is a traditional Chinese medicine that is used for ICH treatment in China. However, the evidence based mechanism is not clear. Purpose: To study the protective effects of ZLHXTY capsules against ICH pathogenesis via targetting nuclear factor kappa β (NFкβ) canonical signalling pathway. Methods: C57BL/6 J mice ICH models using autologous blood injection were used to study the effect of ZLHXTY (1.4 g/kg P.O.) after 24 and 72 hrs of ICH induction. The neurological scoring, corner turn test and balance beam with scoring was performed to assess neurological damage. Hematoxylin/eosin and nissl staining was used for histopathological evaluation. Levels of TNFα, NFкB, iNOS, COX2, IL1, IL6 were measured using real time qPCR and western blotting. Protein levels of IKKβ and IкBα were analyzed through western blotting. Immunofluorescence for co-expression of NeuN/TNFα, NeuN/NFкB, Iba1/TNFα, and Iba1/NFкB was also performed. Results: Treatment with ZLHXTY capsules after ICH ameliorated inflammatory brain injury after 24 and 72 h; revealed by neurological scoring, hematoxylin/eosin and nissl staining. The qPCR and western blot analyses demonstrated significant downregulation of TNFα, NFкB, iNOS, COX2, IL1β and IL6. Further, the IKKβ and IкBα revealed significant downregulation and upregulation respectively in western blot. Immunofluorescence also revealed attenuated expression of TNFα and NFкB in neurons and also low expression of Iba1. Conclusion: ZLHXTY capsules elicit its neuroprotective effect by targetting the NFкβ canonical signalling pathway, thereby ameliorating the ICH induced brain injury.

Keywords: TNFα-NFкB signalling; inflammatory brain injury; inflammatory cytokines; intracerebral hemorrhage; traditional Chinese medicine.