Research Progress on the Antioxidant Activity of Natural Diarylheptanoids: Mechanisms and Structure-activity Relationships.

Antioxidant research has recently become a popular topic. Medicinal plants are important sources of novel active compounds. Diarylheptanoids, a typical family of secondary plant metabolites, are of great interest owing to their extensive spectrum of biological activities. They possess a unique 1,7-diphenylmethane structural skeleton. Thus, this review summarizes the natural linear or macrocyclic diarylheptanoids with antioxidant activity in the last two decades. In addition, the relationships between the structural characteristics of natural diarylheptanoids and their antioxidant capacity were also discussed. All the available data highlight the potential of natural diarylheptanoids as novel antioxidants.

[Effect and mechanism of Danggui Buxue Decoction-containing serum in mitigating H9c2 cell injury caused by exposure to intermittent low oxygen].

This study aims to explore the effect and mechanism of Danggui Buxue Decoction(DBD)-containing serum in alleviating the H9c2 cell injury caused by the exposure to intermittent low oxygen. H9c2 cells were assigned into five groups: control(CON) group, intermittent low oxygen(IH) group, intermittent low oxygen plus DBD-containing serum(IH+DBD) group, intermittent low oxygen plus the autophagy enhancer rapamycin(IH+RAPA) group, and intermittent low oxygen plus DBD-containing serum and the autophagy inhibitor 3-methyladenine(IH+DBD+3-MA) group. Monodansylcadaverine(MDC) staining was employed to detect the changes of autophagosomes. Cell counting kit-8(CCK-8) assay was employed to determine the activity of myocardial cells, and lactate dehydrogenase(LDH) and creatine kinase(CK) kits were used to measure the LDH and CK levels in the cell culture, which would reflect the degree of cell damage. TdT-mediated dUTP nick-end labeling(TUNEL) staining was used to detect the apoptosis of myocardial cells, and JC-1 fluorescence probe to detect the changes in mitochondrial membrane potential. Western blot was employed to determine the expression levels of the autophagy-related proteins microtubule-associated proteins light chain 3Ⅱ(LC3Ⅱ), microtubule-associated proteins light chain 3Ⅰ(LC3Ⅰ), P62, Parkin and apoptosis related proteins pro caspase-3, caspase-3, B-cell lymphoma-2(Bcl-2), Bcl-2-associated X(Bax). The results showed that compared with the CON group, the IH group showed decreased fluorescence intensity of MDC staining, decreased LC3Ⅱ/LC3Ⅰ ratio, down-regulated Parkin expression, and up-regulated expression of P62. In addition, the IH group showed decreased cell survival rate, increased content of LDH and CK in the culture medium, increased number of TUNEL positive cells, and decreased pro caspase-3/caspase-3 and Bcl-2/Bax ratios and mitochondrial membrane potential. Compared with the IH group, the IH+DBD and IH+RAPA groups showed increased fluorescence intensity of MDC staining, increased LC3Ⅱ/LC3Ⅰ ratio, up-regulated Parkin expression, and down-regulated P62 expression. In addition, the two groups showed increased cell survival rate, reduced content of LDH and CK in the culture medium, decreased number of TUNEL positive cells, and increased pro caspase-3/caspase-3 and Bcl-2/Bax ratios and mitochondrial membrane potential. The IH+DBD+3-MA and IH groups showed no significant differences in the above indicators. Compared with the IH+DBD group, the IH+DBD+3-MA group showed decreased fluorescence intensity of MDC staining, decreased LC3Ⅱ/LC3Ⅰ ratio, down-regulated Parkin expression, and up-regulated P62 expression. In addition, the group had decreased cell survival rate, increased content of LDH and CK in the culture medium, increased number of TUNEL positive cells, decreased pro caspase-3/caspase-3 and Bcl-2/Bax ratios, and declined mitochon-drial membrane potential. To sum up, DBD could promote the mitophagy, inhibit the apoptosis, and alleviated the injury of H9c2 cells exposed to low oxygen.

[Danggui Buxue Decoction improves heart function of chronic intermittent hypoxia mice by promoting mitochondrial autophagy and inhibiting cardiomyocyte apoptosis].

The study established a chronic intermittent hypoxia(CIH) model in mice to investigate the effects of Danggui Buxue Decoction(DBD) on mitochondrial autophagy and cardiomyocyte apoptosis and explore its protective effect and mechanism on cardiac function of CIH mice. Forty C57 BL/6 N male mice were randomly divided into the control(CON) group, CIH group, CIH+DBD group, and DBD group, with 10 mice in each group. CIH was induced by filling the hypoxic chamber with N_2(90 s) to reduce the O_2 concentration to 5% and then filling the hypoxic chamber with O_2(90 s) to restore O_2 concentration to 21%, 3 min per cycle, and the CIH treatment continued for 35 d, 8 h per day. Mice in the CIH+DBD and DBD groups were treated with intragastric administration of DBD every day, while those in the CON and CIH groups with the same volume of normal saline. The cardiac function of mice was measured by echocardiography. The pathological changes in myocardium were observed after HE staining, followed by the observation of cardiomyocyte apoptosis by Tunel staining. The expression of apoptosis-related proteins pro-caspase-3, caspase-3, Bcl-2, and Bax and autophagy-related proteins LC3Ⅱ, LC3Ⅰ, P62, parkin, and cytochrome C(Cytc) was detected by Western blot. The mitochondrial membrane potential was observed using JC-1 fluorescent probe. Compared with the CON group, the CIH group exhibited remar-kably lowered left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), elevated left ventricular end-systolic volume(LVESV) and end-diastolic volume(LVEDV), disordered myocardial fiber arrangement, increased number of TUNEL-positive cells, decreased pro-caspase-3/caspase-3, Bcl-2/Bax, and LC3Ⅱ/LC3Ⅰ ratios, parkin, mitochondrial Cytc expression, and mitochondrial membrane potential, and up-regulated P62 and Cytc expression. Compared with the CIH group, DBD increased LVEF, LVFS, pro-caspase-3/caspase-3, Bcl-2/Bax, and LC3Ⅱ/LC3Ⅰ ratios, and parkin expression, as well as mitochond-rial Cytc expression, and mitochondrial membrane potential, decreased LVESV, LVEDV, and the number of Tunel-positive cells, and improved the myocardial fiber arrangement. DBD has a protective effect on the heart function of CIH mice. It improves the heart function possibly by promoting mitochondrial autophagy to ameliorate mitochondrial function and inhibiting the cardiomyocyte apoptosis.

Deciphering the Q-markers of nourishing kidney-yin of Cortex Phellodendri amurense from ZhibaiDihuang pill based on Chinmedomics strategy.

BACKGROUND: Cortex Phellodendri amurensis (CPA) has high medicinal value in the treatment of kidney-yin deficiency diseases. However, due to the lack of research on the therapeutic material basis of CPA, the current quality control standard for CPA is defective, and the effect of the nourishing kidney-yin of CPA was limited. PURPOSE: Based on the principle of correspondence between the syndrome and prescriptions, we studied the CPA in ZhibaiDihuang pill (ZBDH) to identify quality markers (Q-markers) of CPA in ZBDH for treating kidney-yin deficiency and seek the potential Q-markers of CPA under nourishing kidney-yin effect combined with the analysis of single CPA. METHODS: Taking Chinmedomics as the core strategy, metabonomics analysis and effective component identification were performed by UPLC-MS. RESULTS: A total of 121 chemical components of ZBDH were identified, among which the contents of berberine, palmatine, jatrorrhizine and magnoflorine changed the most obviously with the addition of CPA. Forty-five components were identified in the blood in the markedly effective state, including berberine, palmatine, jatrorrhizine and magnoflorine. The therapeutic material basis of ZBDH in the treatment of kidney-yin deficiency was found, and 6 components were found to derive from CPA, including magnoflorine and jatrorrhizine. In addition, seventeen components were identified in the blood in the single CPA treatment, including berberine, palmatine, jatrorrhizine and magnoflorine. CONCLUSIONS: Magnoflorine and jatrorrhizine were the Q-markers of CPA for treating kidney-yin deficiency in the formula of ZBDH and they were also potential Q-markers of the nourishing kidney-yin of CPA.

Effect and Safety of Penetrating Moxibustion in Treatment of Migraine without Aura: A Randomized Controlled Trial.

OBJECTIVE: To observe the clinical effect of penetrating moxibustion on migraine without aura (MO) patients. METHODS: Totally 60 MO patients from the Acupuncture Clinic of the Third Affiliated Hospital of Henan University of Chinese Medicine were collected from November 2015 to February 2017. All patients were assigned to a treatment group and a control group using a random number table, 30 cases in each group. The treatment group was treated with penetrating moxibustion, and the control group was treated with mild moxibustion, thrice a week for 4 consecutive weeks. The total effective rate, Visual Analogue Scale (VAS) scores, headache intensity, and Migraine Specific Quality of Life Questionnaire (MSQ) scores of patients after treatment were compared between the two groups. The moxibustion sensation and reaction after moxibustion were observed, and the adverse reactions were evaluated. All patients were followed up at 4 and 16 weeks after treatment. RESULTS: The total effective rate of the treatment group was significantly higher than that of the control group (93.33% vs. 80.00%, P<0.05). The improvement of VAS scores, headache intensity, and the role restrictive and role preventive scores in MSQ in the treatment group was better than those in the control group (P<0.05). The person-time of moxibustion sensations of itching, numbness and cold as well as flushing and sweating after moxibustion in the treatment group was all significantly higher than that in the control group (P<0.01). There was no significant difference in safety evaluation between the two groups (P>0.05). CONCLUSIONS: Penetrating moxibustion can significantly relieve pain and improve quality of life of MO patients. After penetrating moxibustion, flushing and sweating of patients were obvious, and the curative effect was superior to the mild moxibustion.

[Effect of xiaotan huayu liqiao traditional Chinese medicine compound on myocardial fibrosis in rats with chronic intermittent hypoxia and its mechanism].

Objective: To explore the role of transforming growth factor-ß (TGF-ß) signaling pathway in xiaotan huayu liqiao traditional Chinese medicine compound (XC)'s anti-myocardial fibrosis in chronic intermittent hypoxia (CIH) rats. Methods: Forty SD rats were randomly divided into normoxia group, oxygen + traditional Chinese medicine compound group ( TCMC), Chronic intermittent hypoxia model group (CIH), TCMC + CIH, 10 in each group. CIH cabin was built by filling with nitrogen and oxygen. Firstly, the volume fraction of oxygen in the cabin reduced from 21% to 9% in 90 s by filling the cabin with nitrogen. And then it gradually rose to 21% by reoxygenating in 90s, as a cycle. CIH and TCMC+CIH group rats were placed in the CIH device, while normoxia and TCMC group rats were placed in the normal oxygen chamber. In addition, rats in TCMC +CIH group and TCMC group were treated with XC crude drug (24 g/kg) daily by gavage, while rats in CIH group and normoxia group were given equal volume normal saline. Using sirius red staining, the collagen in myocardial interstitium was visualized. The protein expressions of collagen I, collagen III and fibronectin were detected by Western blot, p-Smad3, p-Smad2 and TGF-ß protein in the TGF-ß/Smads signaling pathway were also analyzed by Western blot. The mRNA expressions of matrix metalloproteinase-2 (MMP-2) and tissue inhibitors of metalloproteinase -2(TIMP-2) were measured by real-time quantitative polymerase chain reaction (PCR). Results: Compared with the rats exposed to normoxia, the CIH rats showed obvious collagen deposition, protein expressions of collagen I, collagen III and fibronectin were significantly increased in the myocardial tissue (P＜0.01). The protein expression levels of TGF-ß, p-smad2 and p-smad3 in the myocardial tissue of the CIH rats were also significantly increased (P＜0.01). The up-regulation of TIMP-2 mRNA in the myocardial tissues resulted in the decrease of MMP-2 mRNA(P＜0.01). XC reduced myocardial fibrosis of CIH rats and inhibited the expressions of collagen I and collagen III and fibronectin protein (P＜0.05,P＜0.01,P＜0.05, respectively). The further mechanism study showed that XC inhibited the expression of TGF-ß (P＜0.01), which down-regulated the expressions of p-smad2, p-smad3 and TIMP-2 (P＜0.05). Conclusion: XC could reduce the expression of TGF-ß and smad2/3 phosphorylation, down-regulate the expression of TIMP-2, which would inhibit the formation of myocardial fibrosis in CIH rats, and improve the myocardial function of CIH rats.

[The effects of Xiaotan Huayu Liqiao formula on mesenteric artery function in rats exposed to chronic intermittent hypoxia].

OBJECTIVE: To investigate the effects of Xiaotan Huayu Liqiao formula (the Chinese Medicine) on mesenteric artery function in rats exposed to chronic intermittent hypoxia (CIH), and to explore the related mechanism. METHODS: Forty-eight male SD rats were randomly divided into four groups as Normoxia, CIH, Formula+CIH and formula group. Rats were exposed to normoxia in the Normoxia and Formula group, or intermittent hypoxia in CIH or Formula+CIH group. Xiaotan Huayu Liqiao formula was given at 24g/kg by intragastric administration before intermittent hypoxia exposure. The pathological changes of mesenteric artery were determined by HE staining, and the relaxation of mesenteric artery (induced by acetylcholine(ACh) and L-arginine(L-Arg)) was recorded by microvascular ring technique. Serums of all rats were collected (0 d and 21 d) and the content of NO was detected by ELISA. The levels of endothelial nitric oxide synthase (eNOS) and p-eNOS were measured by Western blot. RESULTS: Compared with Normoxia group, the mesenteric arterial endothelial injury and media thickening were observed and the relaxation of mesenteric artery was significantly reduced in rats exposed to CIH. The level of NO in serum and the ratio of p-eNOS/eNOS were also decreased in the CIH group. Xiaotan Huayu Liqiao formula administration improved the pathologic changes and dilatation function of mesenteric artery, increased the levels of NO and p-eNOS. Compared with Normoxia group,all the results were not observed significant difference in Formula group. CONCLUSION: Xiaotan Huayu Liqiao formula increased the bioavailability of NO, and ameliorated the CIH induced mesenteric artery function injury.

Herbs-partitioned moxibustion alleviates aberrant intestinal epithelial cell apoptosis by upregulating A20 expression in a mouse model of Crohn's disease.

BACKGROUND: A20 inhibits intestinal epithelial cell apoptosis in Crohn's disease, and herbs-partitioned moxibustion (HPM) has been demonstrated to be an effective treatment for Crohn's disease. However, the mechanism by which HPM reduces intestinal epithelial cell apoptosis in Crohn's disease has not been thoroughly elucidated to date. AIM: To elucidate whether HPM exerts its effects by upregulating A20 to affect intestinal epithelial cell apoptosis in a Crohn's disease mouse model. METHODS: In this study, mice with A20 deletion in intestinal epithelial cells (A20IEC-KO) were utilized to establish a Crohn's disease mouse model with 2,4,6-trinitrobenzene sulfonic acid (TNBS) administration, as well as wild-type mice. Mice were randomly divided into normal control (NC), model control (MC), mesalazine (MESA), and HPM groups. The morphology of the colonic mucosa was observed by hematoxylin-eosin staining, and serum endotoxin and apoptosis of epithelial cells were evaluated by enzyme-linked immunosorbent assay and terminal dUTP nick-end labeling assay accordingly. The protein expression levels of A20 and tumor necrosis factor receptor 1 (TNFR1)-related signaling molecules were evaluated by Western blot, and co-expression of A20 and TNFR1-associated death domain (TRADD) and co-expression of A20 and receptor-interacting protein 1 (RIP1) were observed by double immunofluorescence staining. RESULTS: The intestinal epithelial barrier was noted to have an improvement in the HPM group of wild-type (WT) mice compared with that in A20IEC-KO mice. Compared with A20 IEC-KO HPM mice, serum endotoxin levels and apoptosis percentages were decreased (P < 0.01), A20 expression levels were increased (P < 0.01), and expression of TNFR1, TRADDD, and RIP1 was decreased in the HPM group of WT mice (P TNFR1 < 0.05, P TRADD < 0.01, P RIP1 < 0.01). Both of the co-expression of A20/TRADD and A20/RIP1 showed a predominantly yellow fluorescence in the HPM group of WT mice, while a predominantly red fluorescence was noted in the HPM group of A20IEC-KO mice. CONCLUSION: Our findings suggest that HPM in treating Crohn's disease functions possibly via upregulation of the A20 expression level, resulting in downregulation of TNFR1, TRADD, and RIP1 to alleviate increased cell apoptosis in the intestinal epithelial barrier in Crohn's disease.

[Herbal cake-partitioned moxibustion improves intestinal epithelial barrier dysfunction by suppressing TNF-α-mediated apoptosis pathway of intestinal epithelium in rats with Crohn's disease].

OBJECTIVE: To observe the effect of herbal cake-partitioned moxibustion (Moxi) on tumor necrosis factor (TNF)-α/ TNF receptor 1 (TNFR1)-associated death domain (TRADD) / Fas-associated death domain (FADD) pathway-mediated apoptosis of intestinal epithelial cells in Crohn's disease (CD) rats, so as to explore its underlying mechanisms in the treatment of CD. METHODS: Forty-eight SD male rats were randomly divided into normal, model, Moxi and medication groups (n＝12 rats in each). The CD model was established by intra-annual perfusion of 2，4，6-trinitrobenzene sulfonic acid (TNBS) solution (TNBS∶50% alcohol＝2∶1, 3 mL/kg), once every 7 days, 4 times altogether. For rats of the Moxi group, moxibustion was given to "Tianshu" (ST25) and "Qihai" (CV6), two moxa-cones every time, once daily for 10 days. For rats of the medication group, intragastric perfusion of mesalazine solution was given twice daily for 10 days. After the treatment, the colonic epithelium tissue was sampled. The epithelial cells were purified and cultured to establish an in vitro intestinal epithelial barrier, and added with TNF-α (a pro-inflammatory factor, 100 ng/mL) in the culture medium for 24 h for making an increased epithelial permeability model. The permeability of intestinal epithelial cell barrier was evaluated by detecting the fluorescence yellow transmittance of the TNF-α-incubated cell medium. Western blot was used to detect the expression levels of TNFR1, TRADD, receptor-interacting protein 1 (RIP1), FADD and zinc finger protein A20 (A20, a ubiquitination enzyme for inhibiting activation of TRADD and RIP1) of the cultured intestinal epithelium cells. The apoptosis of the TNF-α-incubated intestinal epithelial cells was detected by flow cytometry. RESULTS: After modeling and compared with the normal group, the fluorescence yellow transmittance of intestinal epithelia cells, apoptosis rate, and expression levels of TNFR1, TRADD, and RIP1 proteins were significantly increased (P<0.001, P<0.01), and the expression of A20 was significantly decreased (P<0.01) in the model group. In comparison with the model group, the fluorescence yellow transmittance of intestinal epithelial cells, the apoptosis rate and expression levels of TRADD, RIP1 and FADD were remarkably down-regulated (P<0.001, P<0.01), and the expression of A20 was significantly up-regulated (P<0.01) in both the Moxi and medication groups. CONCLUSION: Herbal cake-partitioned moxibustion may down-regulate the permeability of intestinal epithelial barrier and the apoptosis of intestinal epithelial cells by way of suppressing TNF-α-mediated cellular apoptosis pathway of intestinal epithelium in CD rats.

Effect of Bizhongxiao decoction and its dismantled formulae on IL-1 and TNF levels in collagen-induced arthritis in rat synovial joints.

BACKGROUND: Rheumatoid arthritis (RA), a chronic autoimmune disease, affects sufferers in many different ways. Treatment of this chronic condition is particularly challenging. Traditional Chinese Medicine (TCM) provides alternatives. Bizhongxiao decoction (BZX) is a TCM complex, which has been used clinically for many years to treat RA. The purpose of this study is to compare the effects of BZX decoction and its dismantled formulae on IL-1 and TNF-1 levels in rats with RA, and to elucidate its mechanism of action. METHODS: Ninety healthy normal female SD rats were randomly divided into six groups: normal (control), model, BZX decoction, and the three dismantled formulae (I: heat-clearing and detoxication, II: dissipating dampness, and III: blood circulation promotion). Apart from the normal (control) group, the rats in each group were injected subcutaneously with bovine type II collagen and complete Freund adjuvant to establish a collagen-induced arthritis model, so that inhibition of foot swelling in the rats by BZX decoction and its dismantled formulae could be observed. Immunohistochemistry was used to assess the levels of the inflammatory cytokines IL-1 and TNF in synovial joints at various time points. RESULTS: Twenty-one days after the model was established, the levels of TNF and IL-1 were significantly higher in the model group, BZX decoction group and dismantled formula groups I, II and III than in the normal controls (P < 0.05). The levels of these cytokines were significantly higher in the model group than the BZX decoction or the three dismantled formula groups (P <0.01). At longer times, the TNF and IL-1 levels in model group rose gradually; those in the BZX decoction and dismantled formula groups were gradually reduced. The cytokine levels in the BZX decoction group were lower than in the three dismantled formula groups and continued to decline. CONCLUSIONS: BZX decoction and the three dismantled formulae examined down-regulated the inflammatory factors IL-1 and TNF in collagen-induced arthritis rat models, but BZX exerted the strongest effect.