Mechanism of Buzhong Yiqitang in Repairing Brain Developmental Abnormalities in Offspring of Pregnant Rats with Subclinical Hypothyroidism
10.13422/j.cnki.syfjx.20251137
- VernacularTitle:补中益气汤改善妊娠期亚临床甲减后代脑发育异常的作用机制
- Author:
Yan MA
1
;
Xiaojiao LYU
2
;
Yangling HUANG
3
;
Xiande MA
3
;
Tianshu GAO
4
;
Peiwei CONG
5
;
Wei CHEN
6
Author Information
1. The Second Clinical Medical College of Liaoning University of Traditional Chinese Medicine （TCM）， Shenyang 110847， China
2. School of Integrative Medicine， Liaoning University of TCM， Shenyang 110847， China
3. Teaching Experiment Center of Liaoning University of TCM， Shenyang 110847， China
4. The Affiliated Hospital of Liaoning University of TCM， Shenyang 110032， China
5. Key Laboratory of Ministry of Education for TCM Viscera-State Theory and Applications， Liaoning University of TCM， Shenyang 110847， China
6. The Second Affiliated Hospital of Liaoning University of TCM， Shenyang 110031， China
- Publication Type:Journal Article
- Keywords:
subclinical hypothyroidism during pregnancy;
offspring brain development;
Buzhong Yiqitang;
mitochondrial oxidative stress;
methyltransferases
- From:
Chinese Journal of Experimental Traditional Medical Formulae
2025;31(16):24-34
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo evaluate the pharmacological effect of Buzhong Yiqitang on brain development in offspring of rats with subclinical hypothyroidism （SCH） during pregnancy and explore its potential mechanism. MethodsForty-eight SPF female SD rats were divided into sham operation group （n=8） and model group （n=40）. The rat model of subclinical hypothyroidism （SCH） was constructed by total thyroidectomy combined with postoperative subcutaneous injection of levothyroxine （L-T4）. The modeled rats were randomly allocated into model， low-， medium-， and high-dose （5.58， 11.16， 22.32 g∙kg-1， respectively） Buzhong Yiqitang， and euthyrox （4.5×10-6 g∙kg-1） groups， with 8 rats in each group. These rats were co-housed with normal male rats for mating. Drug administration started 2 weeks before pregnancy and continued until delivery. Hematoxylin-eosin staining and Golgi-cox staining were used to observe pathological changes in the hippocampal tissue of offspring rats. Western blot was employed to detect the effects of Buzhong Yiqitang on the protein levels of cytochrome C oxidase subunitⅠ （COX）Ⅰ and COXⅣ in the hippocampal tissue of offspring rats. A colorimetric method was used to measure the mitochondrial adenosine triphosphate （ATP） content in the hippocampal tissue of offspring rats. For in vitro experiments， a hydrogen peroxide （H2O2）-induced oxidative damage model was established with rat pheochromocytoma cells （PC12）. Interventions included the DNA methyltransferase inhibitor （SGI-1027）， Buzhong Yiqitang-medicated serum， and euthyrox-medicated serum. The cell counting kit-8 （CCK-8） assay was used to examine the effect of Buzhong Yiqitang on cell proliferation. Immunofluorescence staining was performed to evaluate the effect on tubulin beta 3 class Ⅲ （TUBB3） in PC12 cells. Western blot was employed to assess the effects on the protein levels of DNA methyltransferases （TETs and DNMTs） in PC12 cells. The fluorescent probe 2′，7′-dichlorodihydrofluorescein diacetate （DCFH-DA）， luciferase assay， and JC-1 staining were employed to assess the effects of Buzhong Yiqitang on the levels of reactive oxygen species （ROS） and ATP and the mitochondrial membrane potential in PC12 cells. ResultsCompared with the sham group， the model group showed a reduction in the number of hippocampal neurons， incomplete pyramidal cell bodies， loose arrangement， shortened average dendrite length， decreased dendritic complexity and dendritic spine density， and reduced expression levels of COXⅠ and COXⅣ and content of ATP in the brain tissue （P<0.05， P<0.01）. Compared with the model group， after administration of Buzhong Yiqitang and euthyrox， hippocampal neurons exhibited regular arrangement， complete morphology， extended dendrite， increased dendritic complexity and dendritic spine density， and restored expression levels of COXⅠ and COXⅣ and content of ATP （P<0.05， P<0.01）， with the medium-dose Buzhong Yiqitang group showing the best therapeutic effect. In the PC12 cell model of oxidative damage， Buzhong Yiqitang increased the cell viability （P<0.01）， enhanced neuronal differentiation， down-regulated the expression levels of DNMTs （P<0.05）， up-regulated the expression levels of TETs （P<0.05）， decreased the ROS content （P<0.01）， and restored the ATP content and mitochondrial membrane potential （P<0.01）. ConclusionBuzhong Yiqitang protects brain development in offspring of pregnant rats with SCH. It mainly acts on the oxidative stress and mitochondrial dysfunction resulted from abnormal mtDNA methylation， with DNMTs and TETs as the key proteins for its effects.
