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As a common malignant tumor, the morbidity and mortality of lung cancer have been rising in recent years. The concept of "premetastatic niche" may lead to a revolutionary change in antitumor metastasis therapeutic strategies. Traditional Chinese medicine with multitargets and lower poisonous agents may be a potentially effective means to intervene in the "premetastatic niche (PMN)" to prevent and treat tumor metastasis. Astragalus polysaccharide (APS) is a substance with strong immune activity in Astragalus membranaceus that has excellent biological activities such as immunomodulation, anti-inflammatory, and antitumor. In this study, we constructed a tumor lung metastasis animal model to explore the intervention mechanism of APS on the premetastatic niche. We found that APS inhibited the formation of the lung premetastatic niche and inhibited the recruitment of myeloid-derived suppressor cells (MDSCs) in the lung. Mechanistically, we showed that the proteins and gene expression of S1PR1, STAT3, and p-STAT3 in the S1PR1/STAT3 signaling pathway were suppressed by APS. In line with the above findings, our results confirmed that APS may inhibit the accumulation of MDSCs in the premetastatic niche through the intervention of the S1PR1-STAT3 signaling pathway to achieve the antitumor effect. [ABSTRACT FROM AUTHOR]

Aim: Periodontitis (PD) is the sixth most prevalent disease around the world and is involved in the development and progression of multiple systemic diseases. Previous studies have reported that PD may aggravate liver injuries. The objective of this study was to investigate whether and how PD affects liver fibrosis. Materials and Methods: Ligature‐induced PD (LIP) was induced in male C57/B6J mice, and sub‐gingival plaques (PL) from patients with PD were applied to mouse teeth. Liver fibrosis was induced by carbon tetrachloride (CCl4) injection. The mice were randomly divided into six groups: Oil, Oil+LIP, Oil+LIP+PL, CCl4, CCl4+LIP, and CCl4+LIP+PL. Alveolar bone resorption was evaluated by methylene blue staining. Hepatic function was analysed by serum alanine aminotransferase and hepatic hydroxyproline. Picrosirius red and α‐smooth muscle actin (SMA) staining were used to evaluate the fibrotic area. RNA sequencing and quantitative RT‐PCR were used to measure gene expression. Western blotting was used to measure protein levels. Flow cytometry was used to analyse the accumulation of immune cells. Mouse microbiota were analysed using 16S rRNA gene sequencing. Results: Mice in the CCl4+LIP+PL group displayed higher serum alanine aminotransferase and hepatic hydroxyproline as well as more Picrosirius red‐positive and α‐SMA‐positive areas in liver samples than those of the CCl4 group, suggesting that PD (LIP+PL) aggravated CCl4‐induced hepatic dysfunction and liver fibrosis. Consistently, the expression of fibro‐genic genes and the protein levels of transforming growth factor β were much higher in the CCl4+LIP+PL group than in the CCl4 group. Flow cytometry revealed that PD increased the accumulation of immune cells, including Kupffer cells, B cells, and Th17 cells, in the liver of mice with CCl4 treatment. PD also increased the expression of inflammatory genes and activated pro‐inflammatory nuclear factor‐kappa B pathway in the livers of CCl4‐injected mice. Moreover, PD altered both oral and liver microbiota in CCl4‐injected mice. Conclusions: PD aggravates CCl4‐induced hepatic dysfunction and fibrosis in mice, likely through the increase of inflammation and alteration of microbiota in the liver. [ABSTRACT FROM AUTHOR]

Objective. This study tends to assess the dose-dependent effects of spironolactone on TGF-β1 expression, atrial fibrosis, and the vulnerability to atrial fibrillation in spontaneously hypertensive rats (SHRs) and tries to clarify the association of atrial fibrosis with the vulnerability to atrial fibrillation. Methods. Forty 20-week-old male SHRs were randomly divided into 4 groups (10 rats per group): 3 spironolactone groups were lower-dose group (10 mg·kg−1·d−1, dissolved in 2 ml saline solution, group SL), medium-dose group (40 mg·kg−1·d−1, dissolved in 2 ml saline solution, group SM), higher-dose group (80 mg·kg−1·d−1, dissolved in 2 ml saline solution, group SH) and one hypertension group (2 ml saline solution for stomach gavage, group H). Ten matched homologous WKY rats were set as the control group (group C). After 7 weeks of gavage, a multiple electroconductive physiological recorder was used to detect atrial electrical parameters, including P-wave duration, PR interval, and atrial effective refractory period (AERP), the inducibility, and duration of atrial fibrillation. HE staining was used to determine myocardial cell size. Masson staining was used to detect the deposition of the interstitial collagen fibers in atrial muscle. The expression of TGF-β1 was detected by immunohistochemistry and western blot. Results. Compared with group C, the myocardial cell size, atrial fibrosis, TGF-β1 expression, P-wave duration, PR interval, AERP, inducibility, and duration of atrial fibrillation in group H were conspicuously increased (p < 0.05); compared with group H, there was no significant difference in the myocardial cell size, atrial fibrosis, TGF-β1 expression, and electrophysiological indexes in group SH upon spironolactone intervention (p > 0.05); compared with group H, the myocardial cell size, atrial fibrosis, the expression of TGF-β1, P-wave duration, PR interval, the inducibility, and duration of atrial fibrillation in the group SL and group SM were all decreased (p < 0.05); compared with group SM, the effect in the group SL was more prominent (p < 0.01). Conclusion. Hypertension can lead to cardiomyocyte hypertrophy, deposition of interstitial fibrosis in myocardial tissue, and an increase in the vulnerability to atrial fibrillation. Spironolactone showed a certain dose-dependent effect in SHRs. Lower-dose spironolactone was superior to higher-dose spironolactone in the aspect of reducing hypertensive atrial fibrosis and TGF-β1 expression, as well as preventing the occurrence of atrial fibrillation. [ABSTRACT FROM AUTHOR]

With the aging of the population and the extension of life expectancy, osteoporosis is becoming a global epidemic. Although there are several drugs used to treat osteoporosis in clinical practice, such as parathyroid hormone or bisphosphonates, they all have some serious side effects. Therefore, a safer drug is called for osteoporosis, especially for the prevention in the early stage of the disease, not only the treatment in the later stage. Panax notoginseng saponin (PNS), a traditional Chinese herb, has been used as anti-ischemic drug due to its function on improving vascular circulation. In order to verify whether Panax notoginseng saponins (PNS) could be used to prevent osteoporosis, ovariectomy (OVX) was induced in female C57BL/C6J mice, followed by orally administration with 40 mg/kg/d, 80 mg/kg/d, and 160 mg/kg/d of three different dosages of PNS for 9 weeks. Serum biochemical analysis, micro-CT, histological evaluation, and immunostaining of markers of osteogenesis and angiogenesis were performed in the sham, osteoporotic (OVX), and treatment (OVX+PNS) groups. Micro-CT and histological evaluation showed that compared to sham group, the bone mass of OVX group reduced significantly, while it was significantly restored in the moderate-dose PNS (40 mg/kg and 80 mg/kg) treatment groups. The expression of CD31 and osteocalcin (OCN) in the bone tissue of treatment group also increased, suggesting that PNS activated osteogenesis and angiogenesis, which subsequently increased the bone mass. These results confirmed the potential function of PNS on the prevention of osteoporosis. However, in the high dose of PNS (160 mg/kg) group, the antiosteoportic effect had been eliminated, which also suggested the importance of proper dose of PNS for the prevention and treatment of osteoporosis in postmenopausal women. [ABSTRACT FROM AUTHOR]

Background. Accumulating evidence from prospective epidemiological studies has showed that depression disorder (DD) is a risk factor for cancer. The aim of this study is to explore the association of DD and the overall occurrence risk of hepatocellular carcinoma (HCC) and the mechanism. Methods. In this study, 60 mice were randomly divided into four groups: Control group, DD group, HCC group, HCC-DD group. Mice received a chronic dose of reserpine to establish depression model, followed by Diethylnitrosamine and Carbon tetrachloride administration to establish HCC models. Behavioral depression was assessed by sucrose preference test (SPT) and the expression of Serotonin 1A (5-HT1A) receptor in the hippocampal. The expression of Oatp2a1 and Oatp2b1 in the digestive system tissues was detected by PCR and western blotting. Results. Reserpine-administrated mice had a reducing sucrose preference at Day 14 compared with blank mice (P<0.05). The expression of 5-HT1A receptor in the hippocampal was decreased in DD mice compared with blank mice. The survival analysis indicated that the HCC mice with DD have poorer survival rate compared with the HCC mice. Compared with HCC mice, the expression of Oatp2a1 and Oatp2b1 was lower in liver and stomach tissue and higher in hepatic carcinoma and colon tissue of HCC-DD mice (P<0.05), and the expression of Oatp2a1 was higher in the spleen tissue of HCC-DD mice while the expression of Oatp2b1 was lower (P<0.05). However, no difference was found in the expression of Oatp2a1 and Oatp2b1 in the small intestine tissue between HCC group and HCC-DD group. Conclusions. DD was the adverse factors for the overall occurrence risk of HCC. Mechanistically, be the downregulation of Oatp2a1 and Oatp2b1 in liver tissue induced by DD might be involved. [ABSTRACT FROM AUTHOR]

Tumor-associated macrophages (TAMs) with M2 phenotype play an essential role in tumor microenvironment (TME) during the progression and development of numerous cancers and associated with poor prognosis. Thus, regulation of TAMs polarization emerged as a new strategy for tumor immune therapy. According to Traditional Chinese Medicine (TCM) theory, herbs with Qi-tonifying character are involved in improving the defense capacity of immune system. In this study, we screened extracts and ingredients from five Qi-tonifying herbs exhibiting an inhibitory effect on M2 polarization of murine macrophages RAW264.7 induced by IL-4 and IL-13. Among these candidates, total flavonoids from Glycyrrhiza Radix et Rhizoma (TFRG) and ethanol extract of Ginseng Radix et Rhizoma significantly inhibited the expression of Arginase-1 (Arg-1) (above 90% at 100μg/mL), one of the phenotype markers of M2 macrophages. The inhibition of total saponins of Ginseng Radix et Rhizoma, ethanol extract of Cordyceps, ethanol extract of Acanthopanacis senticosi Radix et Rhizoma Seu caulis, and ethanol extract of Astragali Radix reached above 50% at 100μg/mL. The inhibition of ingredients including glabridin, isoliquiritin apioside, lysionotin, cordycepin, astragaloside IV, and calycosin reached above 50% at 50μM. Then, we investigated the molecular mechanisms of TFRG. TFRG abolished the migration of murine breast cancer 4T1 stimulated by the conditioned medium from M2 macrophages (M2-CM). In addition to Arg-1, TFRG also antagonized the IL-4/13-mediated mRNA upregulation of the M2 markers including found in inflammatory zone 1 (FIZZ1), chitinase-3-like protein 3 (YM1), and mannose receptor (CD206) and upregulated the expression of inducible nitric oxide synthase (iNOS), one of the M1 markers. The further exploration showed that TFRG decreased the phosphorylation of STAT6 and increased the expression of miR-155. Our study provides a series of potential immune regulating natural products from five Qi-tonifying herbs on M2 phenotype. For instance, TFRG suppressed M2 polarization of macrophages partly by inactivating STAT6 pathway and enhanced the level of miR-155 to regulate the expressions of M1 and M2 markers. [ABSTRACT FROM AUTHOR]

Traditional Chinese Medicine Jianpijiedu decoction (JPJD) could improve the general status of liver cancer patients in clinics, especially the symptoms of decreased food intake and diarrhea. In this study, our results showed that the survival rate of the liver cancer with food restriction and diarrhea (FRD-LC) rats was lower than the liver cancer (LC) rats, and the tumor volume of the FRD-LC rats was higher than the LC rats. It was also shown that the high dose of JPJD significantly improved the survival rate, weight, and organ weight when compared with FRD-LC-induced rats. Moreover, JPJD administration upregulated the mRNA and protein levels of ABCC2 and downregulated the mRNA and protein levels of OATP1B2 in liver tissues. However, opposite results were observed in the cancer tissues. In conclusion, the study indicated that the Chinese Medicine JPJD could contribute to the rats with liver cancer which were pretreated with food restriction and diarrhea by regulating the expression of ABCC2 and OATP1B2 in liver tissues and cancer tissues. [ABSTRACT FROM AUTHOR]

The present study determined the effects of electroacupuncture on interstitial cells of Cajal and investigated whether changes in the stem cell factor pathway were involved. Animals were assigned to normal, diabetic, diabetic plus sham stimulation, diabetic plus low-frequency stimulation, and diabetic plus high-frequency stimulation groups. Electroacupuncture was performed daily for 8 weeks. In vitro contractility of colonic muscle strips were studied. Expression of c-kit (the marker of interstitial cells of Cajal) and stem cell factor were measured. The results showed that (1) contraction of colonic muscle strips was significantly elevated in low- and high-frequency stimulation groups and (2) in contrast to the diabetic group, the expressions of c-kit and stem cell factor were markedly increased in the low- and high-frequency stimulation groups. These results indicate that both low- and high-frequency stimulation can promote the contractility of colonic muscle strips partially through increasing the number of interstitial cells of Cajal, and these effects could be mediated by an elevated endogenous stem cell factor. [ABSTRACT FROM PUBLISHER]

Fasting and especially intermittent fasting have been shown to be an effective intervention in many diseases, such as obesity and diabetes. The fasting-mimicking diet (FMD) has recently been found to ameliorate metabolic disorders. To investigate the effect of a new type of low-protein low-carbohydrate FMD on diabetes, we tested an FMD in db/db mice, a genetic model of type 2 diabetes. The diet was administered every other week for a total of 8 weeks. The intermittent FMD normalized blood glucose levels in db/db mice, with significant improvements in insulin sensitivity and β cell function. The FMD also reduced hepatic steatosis in the mice. Deterioration of pancreatic islets and the loss of β cells in the diabetic mice were prevented by the FMD. The expression of β cell progenitor marker Ngn3 was increased by the FMD. In addition, the FMD led to the reconstruction of gut microbiota. Intermittent application of the FMD increased the genera of Parabacteroides and Blautia while reducing Prevotellaceae, Alistipes and Ruminococcaceae. The changes in these bacteria were also correlated with the fasting blood glucose levels of the mice. Furthermore, intermittent FMD was able to reduce fasting blood glucose level and increase β cells in STZ-induced type 1 diabetic mouse model. In conclusion, our study provides evidence that the intermittent application of an FMD is able to effectively intervene in the progression of diabetes in mice. [ABSTRACT FROM AUTHOR]

Zanthoxylum armatum DC. (ZADC) is a traditional medicinal plant with various pharmacological activities and is widely used in China, Japan, India, and other regions. Previous studies have revealed that the methanol extract of ZADC can cause neurotoxicity symptoms in rats, such as drooling, decreased appetite, decreased movement, and increased respiratory rate. However, the basis of these toxic substances and the mechanism of neurotoxicity remain unclear. To evaluate the effects of ZADC on nerve cells and their damage mechanisms and discuss the possible toxic substance basis. The ethyl acetate extract of ZADC is obtained by extracting the methanol extract of ZADC with ethyl acetate. The Q-Orbitrap LC-MS/MS method was employed to analyze the chemical composition of the EA extract of ZADC. SH-SY5Y cells were incubated with different concentrations of the ethyl acetate extract of ZADC. The cytotoxicity of the extract was evaluated using CCK-8, LDH, and ROS assays, and the oxidative stress status of cells was assessed using MDA, GSH, and SOD. Cell apoptosis was detected using flow cytometry. Damage to mitochondrial function was evaluated by labeling mitochondria, ATP, and MMP with fluorescence. Cyto-C, Caspase-3, Caspase-9, Apaf-1, Bax, and reduced Bcl2 expression were measured to evaluate the activation of the mitochondrial apoptosis pathway. Finally, NAC intervention was used to detect changes in the relevant indicators. The activation of mitochondrial apoptosis pathway was evaluated by measuring Cyto-C, Caspase-3, Caspase-9, Apaf-1, and Bax and Bcl2 expression. Finally, NAC intervention was utilized to detect changes in the relevant indicators. After treating SY-SY5Y cells with EA extract from ZADC, cell viability decreased significantly, and the intracellular ROS level increased in a dose-dependent manner. Meanwhile, ZADC can cause cellular oxidative stress and increase MDA and SOD concentrations while decreasing GSH concentrations. It can also shorten the mitochondrial cristae and decrease the number of mitochondria. In contrast, it can reduce ATP synthesis in the mitochondria and mitochondrial membrane potential (MMP). Furthermore, it increased the apoptosis rate and the expression of Cyto-C, Caspase-3, Caspase-9, Apaf-1, and Bax and reduced Bcl2 expression. NAC intervention alleviated the reduction in SH-SY5Y cell survival and the accumulation of reactive oxygen species induced by the EA extract in ZADC. It also inhibits signaling pathways dominated by proteins, such as Cyto-C, reducing cell apoptosis and cytotoxicity. A total of 46 compounds were identified in the extracts. The results suggest that EA extract of ZADC can induce the mitochondrial apoptotic pathway by accumulating ROS in cells, leading to apoptosis. Antioxidants had a good inhibitory and protective effect against cell damage caused by the EA extract of ZADC. The neurotoxic components of ZADC may be organic acids and compounds containing amino groups. [Display omitted] • Zanthoxylum armatum DC. induces cell apoptosis and produces neurotoxicity. • Zanthoxylum armatum DC. causes intracellular ROS accumulation and oxidative stress. • ROS mediated mitochondrial apoptosis pathway is the main mechanism of ZADC toxicity. • NAC intervention can alleviate the neurotoxicity caused by Zanthoxylum armatum DC. [ABSTRACT FROM AUTHOR]

Qing Gan Zi Shen Decoction (QGZS) is a traditional Chinese formula. It has been extensively used for decades in the treatment of hypertension combined with metabolic diseases, but its cardioprotective effects and underlying mechanisms are poorly understood. To explore the cardioprotective effects and potential mechanisms of QGZS in an animal model of obese hypertension. In this study, spontaneously hypertensive rats (SHRs) were utilized as an animal model to examine the effects of a high-fat diet and two concentrations of QGZS. Echocardiography, hematoxylin eosin (H&E) staining, and wheat germ agglutinin (WGA) staining were employed to assess the cardiac structure and function of the SHRs throughout a 16-week therapy period. Furthermore, Western blotting (WB) and immunofluorescence (IF) were employed to identify the levels of Nrf2 expression in the mitochondria, cytoplasm, and nucleus of the myocardium. Additionally, transmission electron microscopy and enzyme-linked immunosorbent assay (ELISA) were utilized to measure mitochondrial morphology and pro-inflammatory cytokine levels, respectively. Furthermore, Western blotting (WB), immunohistochemistry (IHC), and immunofluorescence (IF) techniques were employed to quantify the levels of marker proteins associated with myocardial fibrosis, cardiac inflammation, oxidative stress, and mitochondrial dysfunction. QGZS inhibited weight gain and depressed systolic and mean arterial pressures in high-fat-fed SHRs. Echocardiographic results demonstrated that QGZS prevented the increase in left ventricular mass, restricted the growth of left ventricular diameter, and improved ejection fraction (EF), fractional shortening (FS), and the ratio of early diastolic peak velocity of transmitral flow (E) to late diastolic peak velocity (A) in high-fat-fed SHRs. This suggested that QGZS prevented ventricular remodeling and protected cardiac systolic and diastolic functions. H&E and WGA staining showed that QGZS improved cardiomyocyte disorders and restricted cardiomyocyte hypertrophy. The underlying mechanisms, QGZS attenuated the oxidative stress state, including reducing the generation of reactive oxygen species (ROS) in the myocardium, revitalizing the antioxidant enzyme system, and protecting mitochondrial function. Moreover, QGZS alleviated the pro-inflammatory state in high-fat-fed SHRs. What's more, QGZS significantly increased the expression level of Nrf2 in nuclei and mitochondria in rat heart tissues, exerting a proximate Nrf2 agonist effect. QGZS exerted cardioprotective effects, in part due to its increasing expression of Nrf2 protein in the heart, which promoted Nrf2 nuclear expression. [Display omitted] • In high-fat-induced obese SHRs, Qing Gan Zi Shen reversed ventricular remodeling and protected cardiac function. • In high-fat-induced obese SHRs, Qing Gan Zi Shen alleviated ventricular remodeling and myocardial fibrosis. • QGZS actived Nrf2 to play a role in reducing cardiac inflammation, inhibiting cardiac oxidative stress, and protecting mitochondrial function. [ABSTRACT FROM AUTHOR]

Zanthoxylum armatum DC. (Z. armatum) is an herbal medicine with various active ingredients and pharmacological effects. However, modern studies found that Z. armatum is hepatotoxic. The liver is the target organ for toxic effects and an important site for lipid metabolism. The effects of Z. armatum on lipid level and metabolism in the liver are still unclear. This study aimed to analyze hepatic lipid levels, lipid metabolites and metabolic pathways of action of Z. armatum based on lipidomics, to investigate the potential hepatotoxic mechanism of Z. armatum. Different doses (62, 96, and 150 mg/kg) of the methanolic extract of Z. armatum (MZADC) were administered to ICR mice by gavage. The hepatotoxicity of MZADC was assessed by the liver index, serum biochemical measurements, and histopathological examination. Lipid levels measured by the serum lipid index were evaluated in the mice. Lipidomics was used to screen for differential lipid metabolism markers and lipid metabolism pathways in the liver. Western blot analysis was performed to investigate the effects of MZADC on the liver. Liver index values and serum alanine transaminase and aspartate transaminase levels were increased in the MZADC group. Histopathology examination revealed hepatocyte necrosis, watery degeneration of the hepatocytes, and hepatic cord rupture in the livers of mice. Serum levels of low-density lipoprotein cholesterol, cholesterol, and triglycerides were elevated, and high-density lipoprotein cholesterol levels were decreased. Lipidomics screening for markers of differential lipid metabolism in the liver, and altered profiles of differential metabolites indicated that glycerophospholipid metabolism, linoleic acid metabolism, alpha-linolenic acid metabolism, glycosylphosphatidylinositol-anchored biosynthesis, sphingolipid metabolism and arachidonic acid metabolic pathways were significantly associated with MZADC-induced liver injury. Western blots confirmed that the protein expression of LC3, Beclin-1, ATG5, ATG12 and ATG16L1 was decreased, and p62 was increased in the MZADC group. The proportion of p-PI3K/PI3K and p-AKT/AKT was increased. The liver injury induced by MZADC involved many different lipid metabolites and lipid metabolic pathways, which may be related to autophagy. This study provides a new perspective on the hepatotoxicity study of Z. armatum and provides a reference for the safe application of Z. armatum in the medicine and food fields. [Display omitted] • Based on lipidomics, this study revealed hepatotoxicity induced by Zanthoxylum armatum DC. • The metabolites of liver lipid difference induced by Zanthoxylum armatum DC. may serve as potential biomarkers. • Glycerophospholipid metabolism pathways were significantly associated with Zanthoxylum armatum DC. induced liver injury. [ABSTRACT FROM AUTHOR]

Hypervascularity has been considered as one of the major features of many solid tumors. Green tea is one of the commonly drink resources in China, and its active component, Epigallocatechin gallate (EGCG), exhibits antiangiogenic activities in various experimental tumor models. However, EGCG has many shortages, e.g., relatively unstable, low lipid solubility, poor bioavailability, and short duration of action. To overcome the shortages of EGCG for antiangiogenic antitumor usage, our study developed a novel EGCG derivate, Y 6 (5,3′,4′,3″,4″,5″-6-0-ethyl-EGCG). The underlying mechanism was also elucidated. we evaluated the effects of EGCG, Y 6 on HCC and angiogenesis in vivo and in vitro. Moreover, to understand their antitumor mechanisms, key factors within angiogenesis-related signaling pathways (MAPK/ERK1/2, PI3K/AKT, HIF-1 VEGF) were analyzed by using western blot, immunohistochemistry (IHC), quantitative real-time quantitative PCR (RT-PCR). HepG2 xenograft model and the chorioallantoic membrane (CAM) were used to investigate the effects of Y 6 and EGCG on tumors and anti-angiogenesis in vivo. Micro-vessel density (MVD) was analyzed by IHC of CD34 staining. IHC, qRT-PCR and Western blot were used to detect the expression of HIF-1α and VEGF protein in tumor tissues. The protein levels of MAPK/ERK1/2, PI3K/AKT, HIF-1α, and VEGF in tumor tissues were detected by western blot. Our results demonstrated that both EGCG and Y 6 displayed antiangiogenetic and antitumor effects against HCC cells in vitro and in vivo. We found that rather than equal amount of EGCG, Y 6 displayed better abilities in inhibiting the growth of HCC tumor cells, as well as inhibiting the growth of neovascularization in the chick embryos and HepG2 xenograft tumors bearing-mice, based on the data obtained from MTT assay, immunohistochemistry (IHC), chick chorioallantoic membrane (CAM) assays. In the comparison of equivalent dose of EGCG, qRT-PCR data showed that Y 6 induced more significant decrease of the mRNA levels of HIF-1α and VEGF in supernatant-treated SMMC-7721 cells under hypoxic condition, as well as in the in xenograft tumor tissues; whereas Y 6 also significantly reduced the protein levels of MAPK/ERK1/2, PI3K/AKT, HIF-1α, and VEGF to a greater extent than EGCG, determined by western blotting assay. our work suggests that the new EGCG derivate Y 6 could significantly inhibit tumor growth and angiogenesis which is possibly involved with the signaling intervention of MAPK/ERK1/2 and PI3K/AKT/HIF-1α/VEGF pathways, and is supposed to be a potential therapeutic reagent for anti-angiogenesis treatment of solid tumors. Different concentrations of Y6 and EGCG group compared with the control group, the inhibition of chorioallantoic membrane (CAM) angiogenesis was significant. It indicated that it is a potential new vascular inhibitor, which may be a new treatment option for anti-angiogenic therapy in solid tumors. Image 1 [ABSTRACT FROM AUTHOR]

Rhei Radix et Rhizoma (rhubarb), as the preferred representative of cathartic drugs of traditional Chinese medicine (TCM), has a long history of medicinal use and multifarious functions that produce a wide range of dosage. In modern times, rhubarb and its prescriptions are not only used to treat common clinical diseases, but also achieve good results in the treatment of acute, dangerous, severe and difficult diseases. However, rhubarb also has an alias called "General", which means that its efficacy is relatively rapid. The present study was conducted to simultaneously elucidate dosage-effect relationship of rhubarb of different efficacy, "Removing accumulation with purgation" (E1) and "Clearing heat and purging fire" (E2), providing reference for the safe and effective usage of rhubarb. Three-week-old rats were randomly divided into the normal control group (Con.), model group (Mod.) and rhubarb groups with six doses (0.135, 0.27, 0.81, 1.35, 4.05, 8.1 g/kg). We established a constipation model with gastrointestinal accumulated heat induced by dyspepsia, taking defecation characteristics observed by metabolic cages, alvine pushing rate, gastrointestinal hormones in serum, etc. , as indicators of E1, and taking TG, Na+-K+-ATPase, inflammatory factors and proteins, etc. , as indicators of E2. The factor analytic approach was used to systematically evaluate the two effects and analyze the corresponding dosage-effect relationship. The levels of Gas, AchE, TG, Na+-K+-ATPase, TNF-α, IL-1β, (p-)NF-κB p65, (p-)p38, (p-)ERK and p-JNK in model rats increased significantly while the levels of defecation, fecal water content, MTL, SS, ET, NTS, VIP, JNK and TLR4 decreased. Compared with the Mod., in rhubarb groups, the increase of faeces, alvine pushing rate, most gastrointestinal hormones, etc. , reflected the therapeutic efficacy of E1, and the reduction of TG, Na+-K+-ATPase, expression levels of inflammatory indexes, etc. , reflected the impact of E2. After the analysis, the effective threshold dose ranging from 0.67 to 5.37 g/kg (corresponding to 7.44–59.67 g in the clinic) was in the EC 20 -EC 80 range for E1 treatment and from 0.78 to 5.60 g/kg (equivalently clinical 8.67–62.22 g) was for E2 treatment. And the 1.6- and 1.2-fold rhubarb highest dose of Chinese Pharmacopoeia might be the optimal doses for E1 and E2 respectively. In general, however, the concentration of rhubarb liquid for overall efficacy is suggested to be between middle and highest dose of Chinese Pharmacopoeia. A constipation model was used to elucidate two main effects of rhubarb, which was consistent with the characteristics of TCM syndrome. In the wide range of rhubarb dosage, low doses might have little or no effect and although high concentrations of rhubarb liquid enhanced curative efficacy, it would also have certain side effects on the body. Therefore, scientific-based experiments and rational analysis by mathematical models could contribute to the safe and effective application of rhubarb in the clinic. Image 1 [ABSTRACT FROM AUTHOR]

Aconitum species, with a long history of traditional application, were applied to treat rheumatism, arthritis, stroke, and pain in Chinese medical practice. However, misuse of Aconitum species may induce central nervous toxic effects, such as numbness, vomiting, and even coma. Aconitine has been proved to be the main toxic component of Aconitum plants. Neurotoxicity is the main toxic effect of aconitine, while the underlying mechanism of aconitine remains unclear. The purpose of the study is to explore the effects and molecular mechanism of ferroptosis caused by aconitine in vivo and in vitro. Six-dpf zebrafish larvae and SH-SY5Y cells were treated with different concentrations of aconitine for 24 h. Inhibitors treatment, e.g. pretreatment with Necrostain-1 (Nec-1) and Z-VZD-FMK for 12 h, or with Ferrostain-1 (Fer-1) for 4 h, were involved in the identification of aconitine-induced ferroptosis. Transient transfection experiment was conducted to explore the effects of SLC7A11 in the process of aconitine-induced ferroptosis. The effects of aconitine on morphological changes, lipid peroxidation, ferrous ion, and ferroptosis were detected by transmission electron microscope, flow cytometry, confocal microscopy, enzyme-linked immunosorbent assay and western blotting. In SH-SY5Y cells, morphological changes including shrunken mitochondria, increased mitochondrial membranes density and ruptured mitochondrial membranes were captured in aconitine-treated group. The cell viability and GSH content dose-dependently declined, levels of lipid reactive oxygen species (ROS), malondialdehyde (MDA), and ferrous ion significantly increased after aconitine exposure for 24 h. Ferroptosis inhibitor Fer-1 pretreatment effectively increased cell viability, GSH content, and decreased levels of MDA and lipid peroxidation, suggesting that aconitine induced ferroptosis. In addition, the protein expression of SLC7A11 and GPX4 were improved after Fer-1 preincubation, which indicated that aconitine triggered ferroptosis via the inhibition of SLC7A11 and the inactivation of GPX4. Ferroptotic characteristics, including GSH depletion and lipid peroxidation accumulation, were alleviated via overexpression of SLC7A11 to increase protein expression of GPX4. In zebrafish experiment, GSH depletion, lipid peroxidation accumulation, iron overload, and the decreased protein expression of SLC7A11 and GPX4 were also induced in zebrafish larvae after aconitine exposure. Taken together, aconitine triggered ferroptotic cell death via inhibiting SLC7A11/GPX4 signal pathway in vivo and in vitro. All results indicated that aconitine triggered ferroptosis of SH-SY5Y cells and zebrafish larvae nerve cells, which involved the inhibition of SLC7A11/GPX4 signal pathway mediated by lipid peroxidation damage and iron overload. [Display omitted] • •Aconitine (AC) induced ferroptosis in SH-SY5Y cells and brain of zebrafish • GSH depletion and lipid peroxide accumulation were critical for aconitine-induced ferroptosis. • SLC7A11/GPX4 signal axis was involved in aconitine-caused ferroptotic cell death. • Overexpression of SLC7A11 alleviated aconitine-triggered ferroptosis via upregulating GPX4. [ABSTRACT FROM AUTHOR]

Scutellaria baicalensis (SB) and Coptis chinensis (CC) are widely used traditional Chinese medicine (TCM) for "heat-clearing and damp-drying" and "purging fire and detoxifying". SB-CC are commonly used as a herbal pair for synergistic treatment of various diseases such as bacteria-related infections, metabolic syndromes, and some inflammatory disorders. This herbal pair is commonly used in many famous TCM formula, like Huang-Lian-Jie-Du, Gegen-Qinlian, Banxia Xiexin decoction. Aryl hydrocarbon receptor (AHR) plays an essential role in the disposition of both xenobiotics and endogenous substances through the induction of cytochrome P450 1A (CYP1A) enzymes. Regulation of the AHR-CYP1A axis is increasingly implicated in drug-drug and drug-herb interactions. Research on SB-CC for regulatory effect on the AHR-CYP1A axis is only limited to few compounds. This study aimed to systematically investigate the regulatory effect of SB-CC and its main constitutes on the AHR-CYP1A axis in vitro and in vivo. The livers of mice treated with SB-CC extract were subjected to RNA-sequencing (RNA-seq). The key target genes related to drug metabolism were screened, and the differential expression genes (DEGs) were validated by qRT-PCR, Western blot, and enzyme activity assay. Luciferase reporter gene, qRT-PCR, and Western blot assays were used to determine whether SB-CC and their main constituents could activate AHR and regulate CYP1A expression in HepG2 cells. The effect of SB-CC on the pharmacokinetics of phenacetin, a CYP1A substrate, were further observed in mice to test the net effect of SB-CC on CYP1A functions. The potential CYP1A inhibitors in SB-CC were screened and their inhibitory mechanisms were also studied using human liver microsomes. AHR and drug metabolism system, especially CYP1A1 and CYP1A2, were strongly affected in the liver of SB-CC-treated mice. These results were further validated by the findings that SB-CC increased CYP1A's mRNA, protein expression and activity in mouse liver. In HepG2 cells, SB, CC, baicalin, baicalein, chrysin, oroxylin A, berberine, coptisine and epiberberine increased CYP1A1 mRNA expression in an AHR-dependent way. Interestingly, SB-CC treatment for 14 days only slightly increased the systemic exposure of paracetamol in mice. In the CYP1A inhibition assay, SB, CC, baicalin, baicalein, wogonoside, wogonin, chrysin, oroxylin A, scutellarein, columbamine, coptisine, palmatine, epiberberine, and berberrubine inhibited CYP1A activity in different degree. These results suggested that SB-CC exerted dual regulatory effect on the AHR-CYP1A axis by increasing CYP1A expression but simultaneously inhibiting CYP1A activity, which may contribute to a tight modulation of AHR signaling for homeostatic control. [Display omitted] [ABSTRACT FROM AUTHOR]

Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease with unknown etiology. Oxytropis falcata Bunge (O. falcata) is a 1–35 cm high perennial clustered herb, also known as edaxia, has viscosity and a special smell, and is mainly distributed in the western areas of China. The root of O. falcata has a diameter of 6 mm, is straight and deep, dark red and its stems are shortened, woody and multibranched. O. falcata has heat-clearing, detoxification, analgesic, anti-inflammatory, antibacterial, hemostatic and antitumor activities. Furthermore, O. falcata has excellent anti-inflammatory and analgesic effects, and it is one of the three major anti-inflammatory drugs in Tibetan medicine, known as "the king of herbs". Total flavonoids of Oxytropis falcata Bunge (FOFB) were previously extracted, and their pharmacological activities are consistent with those of the whole herb. In this study, FOFB was extracted from O. falcata by ethanol extraction, and the mechanism of FOFB on IPF was verified by in vivo and in vitro experiments. In this study, we aimed to observe the effects of FOFB on idiopathic pulmonary fibrosis. In in vivo experiments, an IPF rat model was established by bleomycin induction. The rats were treated with FOFB (100, 200, 400 mg kg−1·d−1) for 4 weeks. Masson staining and the expression of TGF-β, p-Smad2, p-Smad3 and Smad7 in the lung tissue of rats were detected. In in vitro experiments, we perfused normal rats with FOFB (100, 200, 400 mg kg−1·d−1) and obtained the corresponding drug-containing serum. The HFL-1 cell model induced by TGF-β1 was used to detect the corresponding indices through intervention with drug-containing serum. The best intervention time for drug-containing serum was detected by the CCK-8 method. Changes in apoptosis, cytoskeleton and rough endoplasmic reticulum structure were detected. Finally, the expression of TGF-β, p-Smad2, p-Smad3 and Smad7 in cells was examined. In vivo, Masson staining indicated that the degree of pulmonary fibrosis increased significantly, the expression of TGF-β, p-smad2 and p-Smad3 increased significantly, and the expression of Smad7 decreased in the model group. We found that the degree of pulmonary fibrosis gradually decreased and that the inhibition of the TGF-β/Smad signaling pathway became more obvious with increasing FOFB dose. FOFB (400 mg kg−1·d−1) significantly improved the degree of pulmonary fibrosis in rats. In in vitro experiments, the CCK-8 results showed that 120 h was the best intervention time for drug-containing serum. In the model group, there was no obvious apoptosis or changes in microfilaments and microtubules, the number of rough endoplasmic reticulum increased, and the expression of TGF-β, p-Smad2 and p-Smad3 increased significantly, while the expression of Smad7 decreased significantly. We found that with the increase in drug-containing serum concentration, the apoptosis, cytoskeleton and degree of destruction of the rough endoplasmic reticulum in the HFL-1 cell model also increased, and the inhibition of the TGF-β/Smad signaling pathway became more pronounced; the effect of the drug-containing serum administered with FOFB (400 mg kg−1·d−1) was the most significant. The results suggest that FOFB can improve the occurrence and development of IPF. The effect of FOFB on IPF may be mediated by inhibition of the TGF-β1/Smad signaling pathway. [Display omitted] • FOFB has good anti-inflammatory, anti-fibrosis and anti-tumor effects. • Pirfenidone and Nintedanib treat IPF with high side effects. • FOFB might improve the symptoms of idiopathic pulmonary fibrosis. Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease with unknown etiology. O. falcata has heat-clearing, detoxification, analgesic, anti-inflammatory, antibacterial, hemostatic and antitumor activities. Furthermore, O. falcata has excellent anti-inflammatory and analgesic effects, and it is one of the three major anti-inflammatory drugs in Tibetan medicine, known as "the king of herbs". Total flavonoids of Oxytropis falcata Bunge (FOFB) were previously extracted, and their pharmacological activities are consistent with those of the whole herb. In this study, FOFB was extracted from O. falcata by ethanol extraction, and the mechanism of FOFB on IPF was verified by in vivo and in vitro experiments. [ABSTRACT FROM AUTHOR]

The Danggui-Yimucao herb pair (DY) is a classic combination in Chinese herbal formulas, consisting of the root of Angelica sinensis (Oliv.) Diels and the aerial parts of Leonurus japonicus Houtt. DY first appeared in "Zhulinsi fuke mifang" in the Jin Dynasty, and it has a long history as a drug for the treatment of abortion. However, its underlying immunomodulatory mechanisms involved are still unclear. In this study, network pharmacology and pharmacological experiments were used to explore the role and mechanism of DY in the treatment of medical abortion. Network pharmacology was used to establish the relationship between the components of DY and abortion-related targets, and to enrich important pathways and biological process for verification. ELISA was used to assess progesterone levels. Flow cytometry was used to detect the degree of differentiation of Th1/Th2 cells. Immunohistochemical methods and qPCR were used to measure the expression levels of T-bet, GATA-3 and IL-4. Through the prediction analysis of network pharmacology, we found that key pathway for DY treatment of abortion, such as anemia, pelvic infection, immune disorders, and coagulation disorders, was Th1/Th2 cell differentiation pathway. The pharmacological results revealed that DY greatly corrected the imbalance of Th cell subsets in abortion mice, significantly inhibited the differentiation of Th2 cells, and resulted in an increase in the Th1/Th2 ratio. In addition, the concentration of progesterone in the serum of mice after abortion was significantly reduced. We also found that DY upregulated spleen T-bet and downregulated IL-4 gene expression in mice. Besides, immunohistochemical results showed that DYE could up-regulate T-bet but inhibit GATA-3 expression. Our results showed that after RU486-induced abortion, progesterone and Th1/Th2 paradigm were disordered in mice, but DY could make mice recover more quickly, which indicated that DY had great development value in immunoregulation. [Display omitted] [ABSTRACT FROM AUTHOR]

Background: Angiogenesis has a key role in numerous disease processes. One of the most important angiogenic factors is vascular endothelial growth factor (VEGF-A), whereas thrombospondin-1 (TSP-1) is a major antiangiogenic factor. Recent studies have shown that VEGF-A as well as TSP-1 is regulated by transforming growth factor-beta1 (TGF-beta1), but the mechanism remains unclear.Methods: We examined the role of TGF-beta1 and its signaling pathways in mediating expression of these two molecules. Rat proximal tubular cells (NRK52E) were stimulated with TGF-beta1 to induce VEGF-A and TSP-1 synthesis. To clarify roles of receptor-activated Smads (R-Smads), we blocked Smad signaling using overexpression of the inhibitory Smad, Smad7, and by using fibroblasts from wild-type or knockout mice. To confirm the antiantigenic role of Smads, soluble Flt-1 regulation in response to TGF-beta1 was also examined. In addition, the effect of conditioned media from NRK52E and Smad knockout cells was examined on endothelial cell proliferation.Results: Induction of VEGF-A and TSP-1 by TGF-beta1 in NRK52E cells was associated with activation of pathway-restricted R-Smads (Smad2 and 3) and blocking these Smads by overexpression of Smad7 blocked their induction. By using of Smad knockout cells, Smad3 was shown to have a key role in the stimulation of VEGF-A expression whereas Smad2 was critical for TSP-1 expression. Consistent with the hypothesis that Smad2 has an antiangiogenic function, we also demonstrated that Smad2, but not Smad3, mediated the expression of VEGF-A antagonist, soluble VEGF-A receptor sFlt-1, in response to TGF-beta1. Conditioned media from NRK52E, which was stimulated by TGF-beta1 for 24 hours, did not induce endothelial cell proliferation. However, conditioned media from Smad2 knockout induced endothelial cell proliferation, whereas endothelial cell proliferation was inhibited by Smad3 knockout-derived conditioned media.Conclusion: R-Smads have distinct roles in mediating the expression of pro- and antiangiogenic growth factors in response to TGF-beta1. [ABSTRACT FROM AUTHOR]

Sjögren's syndrome (SS) is an autoimmune disease and can cause gastrointestinal disorders such as constipation and intestinal inflammation. As a kind of medicinal material, Paeonia lactiflora Pall has a variety of pharmacological effects, and it is also an indispensable component in many pharmaceutical preparations, which has been widely concerned by the medical and pharmaceutical circles. Total glucosides of paeony (TGP) is a mixture of biologically active compounds extracted from the root of Paeonia lactiflora Pall and has therapeutic effects on a variety of autoimmune diseases. To investigate the therapeutic effect of TGP on constipation and intestinal inflammation in mice modeled by SS, and to provide a basis for clinical research. The SS model was set up by submandibular gland (SMG) immune induction method and then treated with TGP for 24 weeks. The fecal characteristics were observed and the fecal number and moisture content were measured. Colonic pathology was observed by H&E staining. The levels of serum P substance (SP), vasoactive intestinal peptide (VIP), interleukin (IL)-1β, tumor necrosis factor (TNF)-α, nuclear factor (NF)-κB, nitric oxide (NO), and nitric oxide synthase (NOS) were determined by enzyme linked immunosorbent assay (ELISA) and microplate method, respectively. Reverse transcription polymerase chain reaction (RT-PCR) was employed to analyze the mRNA expression of c-kit and stem cell factor (SCF) in colon. Compared with the model group, the dry and rough condition of the feces was improved, and the fecal gloss, number and moisture content significantly increased after the administration of TGP capsules. Meanwhile, TGP treatment improved colonic pathological damage, inhibited the serum concentrations of NO, NOS, IL-1β, TNF-α, NF-κB and SP, increased serum VIP concentration, and up-regulated mRNA expression of SCF and c-kit in colon. TGP could obviously attenuate SS-mediated constipation and intestinal inflammation in mice by acting on some intestinal motility related factors and inflammatory factors. Image 1 [ABSTRACT FROM AUTHOR]