Banxia Xiexin decoction promotes gastric lymphatic pumping by regulating lymphatic smooth muscle cell contraction and energy metabolism in a stress-induced gastric ulceration rat model.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional Chinese medicine (TCM) formula Banxia Xiexin decoction (BXD) has definite therapeutic effect in treating stress-induced gastric ulceration (SIGU) and many other gastrointestinal diseases, but its effect on gastric lymphatic pumping (GLP) remains unclear. AIM OF THE STUDY: Elucidating the role of GLP in SIGU and BXD treatment, and exploring the molecular mechanisms of GLP regulation. MATERIALS AND METHODS: In vivo GLP imaging were performed on SIGU rat model, and the lymphatic dynamic parameters were evaluated. Gastric antrum tissues and serum were collected for macroscopic, histopathological and ulcerative parameters analysis. Gastric lymphatic vessel (GLV) tissues were collected for RNA-Seq assays. Differentially expressed genes (DEGs) were screened from RNA-Seq result and submitted for transcriptomic analysis. Key DEGs and their derivative proteins were measured by qRT-PCR and WB. RESULTS: GLP was significantly suppressed in SIGU rats. BXD could recover GLP, ameliorate stomach lymphostasis, and alleviate the ulcerative damage. Transcriptome analysis of GLV showed the top up-DEGs were concentrated in smooth muscle contraction signaling pathway, while the top the down-DEGs were concentrated in energy metabolism pathways especially fatty acid degradation pathway, which indicated BXD can promote lymphatic smooth muscle contraction, regulate energy metabolism, and reduce fatty acid degradation. The most possible target of these mechanisms was the lymphatic smooth muscle cells (LSMCs) which drove the GLP. This speculation was further validated by the qRT-PCR and WB assessments for the level of key genes and proteins. CONCLUSIONS: By activating the smooth muscle contraction signaling pathway, restoring energy supply, modulating energy metabolism program and reducing fatty acid degradation, BXD effectively recovered GLP, mitigated the accumulation of inflammatory cytokines and metabolic wastes in the stomach, which importantly contributes to its efficacy in treating SIGU.

Exploring the effect and mechanism of Haizao Yuhu decoction containing three variants of glycyrrhiza on goiter using an integrated strategy of network pharmacology and RNA sequencing.

ETHNOPHARMACOLOGICAL RELEVANCE: Haizao Yuhu decoction (HYD) is a classic Chinese herbal formula described in the surgical monographs of the Ming Dynasty "Waikezhengzong." It has been widely used to treat goiter for approximately 500 years and found to be particularly effective. HYD contains glycyrrhiza and sargassum. This pair of herbs belongs to "18 incompatible medicaments" of traditional Chinese medicine theory. Although these two herbs are opposite, our preliminary study proved that they have superior effect when added into HYD at 2 times the dose of Chinese Pharmacopoeia. However, the species of glycyrrhiza in HYD that are the most effective have not been recorded in ancient Chinese medical texts. According to the Chinese Pharmacopoeia, glycyrrhiza is divided into the following three species: Glycyrrhiza uralensis Fish., G. glabra L., and G. inflata Bat. The effect of HYD containing different species of glycyrrhiza and their mechanisms remain to be further explored. AIM OF THE STUDY: To investigate the effect of HYD containing three species of glycyrrhiza on goiter, and to elucidate the molecular mechanism using network pharmacology combined with RNA sequencing (RNA-seq). MATERIALS AND METHODS: A rat model of goiter was established by 14 days of intragastric gavage of propylthiouracil (PTU), and the rats were treated for 4 weeks with HYD containing three different species of glycyrrhiza. The body weight and rectal temperature of rats were tested weekly. At the end of the experiment, the serum and thyroid tissues of rats were collected. The effect of the three HYDs was assessed based on general observations (including body weight, rectal temperature, and living status of rats), absolute/relative thyroid weight, thyroid function (including triiodothyronine, thyroxine, free triiodothyronine, free thyroxine, and thyroid-stimulating hormone levels), and thyroid tissue pathology. Next, we explored their pharmacological mechanisms using network pharmacology combined with RNA-seq and validated key targets using real-time quantitative reverse-transcription polymerase chain reaction (RT-qPCR), western blotting (WB), and immunofluorescence (IF) assays. RESULTS: The three HYDs reduced the absolute/relative weights of thyroid tissues and improved the pathological structure, thyroid function, and general findings of rats with goiter. Overall, the effect of HYD-G. uralensis Fish. (HYD-U) was better. Results from network pharmacology and RNA-seq jointly suggested that both the pathogenesis of goiter and the mechanism of action of HYD for goiter were related to the phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) pathway. We validated the key targets in the pathway, namely, vascular endothelial growth factor (VEGF) A, VEGF receptor 2, phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) and its encoded protein PI3K (p85), AKT serine/threonine kinase 1 (AKT1), phospho-AKT and cyclin D1 using RT-qPCR, WB, and IF assays. The PI3K-Akt pathway was hyperactivated in rats with PTU-induced goiter, whereas the three HYDs could inhibit the pathway. CONCLUSION: This study confirmed the definite effect of the three HYDs in the treatment of goiter, and HYD-U was found to be more effective. The three HYDs inhibited angiogenesis and cell proliferation in goiter tissue by inhibiting the PI3K-Akt signaling pathway.

Haizao Yuhu decoctions including three species of glycyrrhiza protected against propylthiouracil-induced goiter with hypothyroidism in rats via the AMPK/mTOR pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Glycyrrhiza and sargassum are among the 18 incompatible medicaments according to traditional Chinese medicine (TCM) theory. Although it contains glycyrrhiza and sargassum, Haizao Yuhu decoction (HYD) is a classic prescription widely used as TCM to treat goiter. According to the Chinese Pharmacopoeia, glycyrrhiza is divided into three varieties: Glycyrrhiza uralensis Fish., Glycyrrhiza glabra L., and Glycyrrhiza inflata Bat. Whether the three varieties of glycyrrhiza have different efficacy or toxicity when applied in the HYD is unknown. AIM OF THE STUDY: To explore whether the HYDs comprising three varieties of glycyrrhiza have different efficacy or toxicity when used to treat goiter in rats and the underlying mechanisms of these HYDs. MATERIALS AND METHODS: For two weeks, the goiter model was replicated by intragastric propylthiouracil (PTU) administration. Samples were divided into the control group, model group, euthyrox group, HYD with glycyrrhiza uralensis (HYD-U) group, HYD with glycyrrhiza glabra (HYD-G) group, and HYD with glycyrrhiza inflata (HYD-I) group. After four weeks of treatment, body weight, rectal temperature, thyroid/liver/kidney coefficient, thyroid/liver/kidney function, thyroid/liver/kidney histomorphology, and thyroid ultrastructure were evaluated. Then, real-time quantitative reverse-transcription polymerase chain reaction (RTqPCR), Western blot, and immunofluorescence analyses were performed to detect genes and proteins affecting autophagy and apoptosis in thyroid cells in the AMP-activated Protein Kinases (AMPK)/Mammalian target of rapamycin (mTOR) pathway. RESULTS: All three HYDs increased thyroid hormones (THs) levels, relieved thyroid pathological tissue and ultrastructure, and activated vital proteins and genes in the AMPK/mTOR pathway. Comparisons among the efficacy of the three HYDs indicated that HYD-U restored the THs most effectively; however, no difference in the anti-goiter effect was observed. Moreover, the three HYDs resulted in no toxicity and promoted the recovery of impaired liver and kidney function caused by PTU. Comparisons among the recovery effects of the three HYDs on the liver and kidney were the same. CONCLUSION: Our experiments demonstrated that the three HYDs had outstanding anti-goiter effects and protected liver and kidney function. Their anti-goiter effects were attributed to AMPK/mTOR pathway-induced autophagy and apoptosis. HYD-U resulted in the best THs recovery. It was further indicated that in our present study, glycyrrhiza and sargassum were compatible in the three HYDs, thereby suggesting their safety of compounding in HYD and providing a basis for the research of the 18 incompatible medicaments.

Comparative Efficacy of Haizao Yuhu Decoction Composed of Different Varieties of Glycyrrhiza in Goiter Rats.

In traditional Chinese medicine, Glycyrrhiza and Sargassum are one pair of the "18 incompatible medicaments," which in theory cannot be used together. However, since ancient times, many reports have described using compounds containing both Glycyrrhiza and Sargassum to treat diseases. Haizao Yuhu Decoction (HYD), which contains both ingredients, is mainly used to treat goiter. Chinese Pharmacopoeia officially recorded three varieties of Glycyrrhiza: Glycyrrhiza uralensis, Glycyrrhiza inflata, and Glycyrrhiza glabra. These three varieties have certain differences in chemical composition and pharmacological effects. The purpose of the present study was to investigate whether the HYD containing different varieties of Glycyrrhiza and Sargassum had different therapeutic effects in rats with goiter and to elucidate the underlying mechanism of any difference. In this study, propylthiouracil (PTU) was used to replicate the goiter model, then HYDs containing different varieties of Glycyrrhiza were used for treatment for four weeks, and then the relevant indicators were tested. The results demonstrated that HYD had antigoiter effects, alleviated the pathological changes in the thyroid tissue, and restored the abnormal serum levels of hormones related to thyroid function induced by PTU. HYD containing Glycyrrhiza uralensis had the best therapeutic effect in rats with PTU-induced goiter. The antigoiter effect of HYD may function through the hypothalamic-pituitary-thyroid (HPT) axis, inhibit the expression of the Tg and NIS genes, and regulate the synthesis of thyroid hormones, thereby reducing the excessive stimulation of TSH in thyroid cells. In addition, HYD also prevented goiter by promoting thyroid cell apoptosis and inhibiting the ERK/RSK1 pathway of cell proliferation. In conclusion, three types of HYD had different therapeutic effects in rats with goiter, which might be caused by the compatibility of different varieties of Glycyrrhiza and Sargassum.

Does the Fragrance of Essential Oils Alleviate the Fatigue Induced by Exercise? A Biochemical Indicator Test in Rats.

OBJECTIVE: To study the effect of the essential oils of Citrus sinensis L., Mentha piperita L., Syzygium aromaticum L., and Rosmarinus officinalis L. on physical exhaustion in rats. METHODS: Forty-eight male Wistar rats were randomly divided into a control group, a fatigue group, an essential oil mixture (EOM) group, and a peppermint essential oil (PEO) group. Loaded swimming to exhaustion was used as the rat fatigue model. Two groups were nebulized with EOM and PEO after swimming, and the others were nebulized with distilled water. After continuous inhalation for 3 days, the swimming time, blood glucose, blood lactic acid (BLA), blood urea nitrogen (BUN), superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and malondialdehyde (MDA) in blood were determined. RESULTS: While an increased time to exhaustion and SOD activity were apparent in both the EOM and PEO groups, the BLA and MDA were lower in both groups, in comparison with the fatigue group, and the changes in the EOM group were more dramatic. Additionally, the EOM group also showed marked changes of the rise of blood glucose and the decrease of BUN and GSH-PX. CONCLUSION: The results suggested that the inhalation of an essential oil mixture could powerfully relieve exercise-induced fatigue.

Geniposide, the component of the Chinese herbal formula Tongluojiunao, protects amyloid-ß peptide (1-42-mediated death of hippocampal neurons via the non-classical estrogen signaling pathway.

Tongluojiunao (TLJN) is an herbal medicine consisting of two main components, geniposide and ginsenoside Rg1. TLJN has been shown to protect primary cultured hippocampal neurons. However, its mechanism of action remains unclear. In the present study, primary cultured hippocampal neurons treated with Aß1-42 (10 µmol/L) significantly increased the release of lactate dehydrogenase, which was markedly reduced by TLJN (2 µL/mL), specifically by the component geniposide (26 µmol/L), but not ginsenoside Rg1 (2.5 µmol/L). The estrogen receptor inhibitor, ICI182780 (1 µmol/L), did not block TLJN- or geniposide-mediated decrease of lactate dehydrogenase under Aß1-42-exposed conditions. However, the phosphatidyl inositol 3-kinase or mitogen-activated protein kinase pathway inhibitor, LY294002 (50 µmol/L) or U0126 (10 µmol/L), respectively blocked the decrease of lactate dehydrogenase mediated by TLJN or geniposide. Therefore, these results suggest that the non-classical estrogen pathway (i.e., phosphatidyl inositol 3-kinase or mitogen-activated protein kinase) is involved in the neuroprotective effect of TLJN, specifically its component, geniposide, against Aß1-42-mediated cell death in primary cultured hippocampal neurons.

The improvement of spatial memory deficits in APP/V717I transgenic mice by chronic anti-stroke herb treatment.

In China, herbal medicine has an extensive history for the treatment of cerebrovascular diseases. Clinical studies have shown that stroke patients are more likely to experience significant memory decline in comparison to their healthy counterparts. Cognition is improved in stroke patients treated with herbal medicine active components, Geniposide (GP) and Geniposide Rg1 (GRg1) (together, called TLJN). However, the effect of TLJN in Alzheimer disease remains unknown. Therefore, we investigated the behavioral effect of TLJN in male and female APP/V717I transgenic (Tg) mice. We conducted two different treatment strategies: (1) pretreatment strategy: medically treated at the age of 3 months which lasted for 3 months; (2) early treatment strategy: medically treated at the age of 6 months which lasted for 4 months. In open field test, locomotor activity and anxiety-like behavior were not affected after TLJN administration in Tg mice. In Morris Water Maze test, spatial learning processes in both genders were improved by TLJN treatments. Furthermore, retrieval processes were significantly improved in the pretreatment strategy for only male mice, which also showed a trend for improved retrieval processes with early treatment. In the inhibitory avoidance test, TLJN enhanced learning processes. In addition, gender differences were found in Tg mice exposed to TLJN treatments. In Tg male mice, significant efficacy was seen at high and middle doses, and in Tg female mice, a low dose was more effective.