Characterization of the Chemical Constituents in Radix gentianae by Ultra-High Performance Liquid Chromatography Coupled with Quadrupole Time-of-Flight Mass Spectrometry.

Radix gentianae (RG) is a traditional Chinese medicine used for the treatment of acute and chronic hepatitis in clinic. However, the chemical profile of RG is still unconfirmed, which hindered the progress of pharmacological study and clinical application. In this study, ultra-high performance liquid chromatography together with quadrupole time-of-flight mass spectrometry techniques were employed to separate and characterize the chemical constituents in RG. Under the optimized conditions, a total of 60 compounds were rapidly identified or tentatively characterized. Results indicated that iridoid glucosides, flavonoids, organic acids, amino acids, saccharides and nucleosides were major constituents in RG. It is concluded the established method can help to clarify the substance basis and provide useful information for ascertaining the bioactive constituents and action mechanism of RG.

Characterization of chemical constituents in Huangqi Guizhi Wuwu decoction using ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

Huangqi Guizhi Wuwu decoction (HGWWD) is a classic traditional Chinese medicine prescription for the treatment of ischemic stroke, etc. However, the material basis of its efficacy remains unclear, seriously affecting drug development and clinical applications. In the present study, an ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry method was developed to separate and identify the chemical components of HGWWD. A total of 81 compounds were identified and tentatively characterized. Eight compounds were accurately identified by comparing the retention time and mass spectrometry data with those of reference substances, the remaining compounds were characterized by comparing the mass spectrometry data and reference information. Based on the results of compound attribution, 35 compounds were from Astragali Radix, six compounds were from Cinnamomi Ramulus, 23 compounds were from Paeoniae Radix Alba, eight compounds were from Zingiberis Rhizoma Recens and nine compounds were from Jujubae Fructus. The results showed that monoterpenoids, flavonoids, organic acids, triterpenes, amino acids, gingerols, alkaloids, and glycosides were the main chemical components of HGWWD. This analytical method is suitable for characterizing the chemical constituents of HGWWD, and the results provide important information for elucidating its pharmacodynamic material basis and mechanism of action.

PD-1 blockade and radiotherapy combination for advanced Epstein-Barr virus-associated intrahepatic cholangiocarcinoma: a case report and literature review.

Advanced intrahepatic cholangiocarcinoma (ICC) is a rare malignant tumor of biliary epithelial cells, known for its extremely unfavorable prognosis. In the absence of intervention, patients typically survive for less than 5 months. Current guidelines from the Chinese Society of Clinical Oncology (CSCO), National Comprehensive Cancer Network (NCCN), and European Society for Medical Oncology (ESMO) recommend chemotherapy-based systemic therapy as the standard treatment for advanced ICC. However, the first-line regimen, consisting of gemcitabine in combination with cisplatin, generally results in a median survival of approximately one year, which is considered suboptimal. Significant progress has been made in radiotherapy techniques, molecular diagnostics, and tumor immune microenvironments. The integration of immune and radiation therapies has revolutionized treatment strategies for cholangiocarcinoma. Moreover, combined therapeutic regimens have shown promising results in improving survival rates among patients with advanced ICC. In this study, we present a case report of a 70-year-old male patient diagnosed with stage IV ICC, featuring metastases to the retroperitoneal, left adrenal, and left supraclavicular lymph nodes. The patient exhibited a high tumor mutational load, significant microsatellite instability, and hyper-expression of PD-L1 (90%), along with positive Epstein-Barr virus-encoded RNA (EBER). Pembrolizumab, a programmed cell death 1 (PD-1) inhibitor, was administered in conjunction with radiotherapy. As a result, considerable shrinkage and inactivation of the primary foci were observed, accompanied by the disappearance of metastases. Ultimately, the patient achieved complete remission and maintained progression-free survival for 41 months following the initial treatment. To the best of our knowledge, this represents the longest case of complete remission using a combination of immunotherapy and radiotherapy as a first-line regimen for the high tumor mutational load, microsatellite instability, and PD-L1 expression (90%) subtype of Epstein-Barr virus-associated ICC (EBVaICC). These findings suggest that the combination of PD-1 inhibitors with radiotherapy may serve as a promising therapeutic strategy for treating this particular cancer subtype.

Network pharmacology associated anti-influenza mechanism research of Qingjie-Tuire Granule via STAT1/3 signaling pathway.

Qingjie-Tuire (QT) granule was approved for clinical use and its combination was reported to treat influenza infection. To explore its active component and mechanism, the components of QT granule were retrieved from UPLC-UC-Q-TOF/MS analysis. The genes corresponding to the targets were retrieved using GeneCards and TTD database. The herb-compound-target network was constructed by Cytoscape. The target protein-protein interaction network was built using STRING database. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of QT granule to IAV were performed for further study. The regulation to different signaling transduction events and cytokine/chemokine expression of QT granule was evaluated using Western blotting and real-time qPCR. Totally, 47 compounds were identified and effect of QT granule on cell STAT1/3 signaling pathways was confirmed by A549 cell model. The efficiency of QT granule on host cell contributes to its clinical application and mechanism research.

Infant exposure to trace elements in breast milk, infant formulas and complementary foods from southern China.

Excessive intake of essential trace elements or exposure to potentially toxic elements above certain thresholds may cause adverse health effects in humans. To date, there is scarce evidence concerning Chinese infant exposure to trace elements and the associated risks. In this study, we collected 61 breast milk, 54 infant formula and 90 complementary food samples from southern China to investigate the levels of cobalt (Co), chromium (Cr), copper (Cu), selenium (Se), zinc (Zn), arsenic (As), cadmium (Cd), nickel (Ni) and lead (Pb). The concentrations of these elements in the breast milk samples ranged from under the limit of detection (

High humidity aggravates the severity of arthritis in collagen-induced arthritis mice by upregulating xylitol and L-pyroglutamic acid.

BACKGROUND: Humidity was an unfavorable factor for patients with rheumatoid arthritis (RA). RA disease activity was severe in high humidity conditions. However, there is no evidence to demonstrate the effects of humidity on arthritis in the animal experiments and explore its relevant mechanism. METHODS: Using the DBA/1 mice, this study addressed the effects of a high humidity (80 ± 5%) on arthritis in collagen-induced arthritis (CIA) mice. Then, this study used the gas chromatography-mass spectrometer (GC-MS) to explore alterations in serum metabolome caused by the high humidity. Furthermore, xylitol and L-pyroglutamic acid, which were both significantly upregulated by the high humidity, were selected to further study their effects on arthritis in the CIA mice. RESULTS: The high humidity (80 ± 5%) could aggravate arthritis variables including increasing arthritis score and swelling, serum autoantibodies (anti-COII and anti-CCP), and proinflammatory cytokines (IL-6, IL-17A, and G-CSF). In addition, the high humidity could cause significant alterations in serum metabolome in the CIA mice. Xylitol and L-pyroglutamic acid were the representative serum metabolites that were significantly upregulated by the high humidity. Further experiments demonstrated that the supplementation of 0.4 mg/mL xylitol in drinking water after inducing the CIA model and 2.0 mg/mL in drinking water before inducing the CIA model could both aggravate arthritis in the CIA mice. CONCLUSIONS: These data demonstrated that high humidity was not beneficial for arthritis development and its mechanism might be associated with xylitol and L-pyroglutamic acid.

Tissue Distribution Study of Poloxamer188 in Rats by Ultra-High-Performance Liquid Chromatography Quadrupole Time of Flight/Mass Spectrometry with MSALL-Based Approach.

Poloxamer188 (PL188), as one of the most commonly used pharmaceutical excipients, has unique physicochemical properties and good biocompatibility, and so is playing an increasingly extensive role in the field of medicine. Currently, there are few studies on the tissue distribution of PL188 in vivo. In this study, the LC-MS method based on MSALL technique of quadrupole time of flight mass spectrometry for absolute quantitative analysis of poloxamer 188 in biological substrates was established for the first time. The tissue distribution of poloxamer188 in SD rats were studied using the established quantitative analysis method. To explore the distribution of PL188 in organs and tissues, PL188 was administered via rat tail vein at a dose of 5 mg/kg. Eight kinds of tissues including heart, liver, spleen, lung, kidney, stomach, muscle and brain of rats were collected at 0.25 h, 1 h and 4 h after administration. Tissue distributions showed the highest level was observed in kidney, then in stomach, which indicated PL188 mainly bioaccumulated in the kidney. This study can provide references for the further study of PL188.

Qingda granule attenuates angiotensin II-induced cardiac hypertrophy and apoptosis and modulates the PI3K/AKT pathway.

Qingda granule (QDG), simplified from Qingxuan Jiangya Decoction, is a well-known traditional Chinese medicine formula that has been used for decades to treat hypertension. However, the cardioprotective effects of QDG on Ang II-induced hypertension remain unknown. This study aimed to investigate the effects of QDG on hypertension-induced cardiac hypertrophy and apoptosis, as well as explore its underlying mechanisms. Mice were infused with Ang II (500 ng/kg/min) or saline solution as control, then administered oral QDG (1.145 g/kg/day) or saline for two weeks. QDG treatment attenuated the elevation in blood pressure caused by Ang II, as well as the decreased left ventricle ejection fractions and fractional shortening. Moreover, QDG treatment significantly alleviated the Ang II-induced elevation of the ratio of heart weight to tibia length, as well as cardiac injury, hypertrophy, and apoptosis. In cultured H9C2 cells stimulated with Ang II, QDG partially reversed the increase in cell surface area and number of apoptotic cells, up-regulation of hypertrophy markers ANP and BNP, and activation of caspases-9 and -3. QDG also partially reversed Ang II-induced accumulation of reactive oxygen species (ROS), depolarization of the mitochondrial membrane, release of cytochrome C, up-regulation of Bax, and decrease in levels of p-PI3K, p-AKT, and Bcl-2. These results suggest that QDG can significantly attenuate Ang II-induced hypertension, cardiac hypertrophy and apoptosis, and it may exert these effects in part by suppressing ROS production and activating the PI3K/AKT signaling pathway.

Efficacy and safety of retention enema with traditional Chinese medicine for ulcerative colitis: A meta-analysis of randomized controlled trials.

OBJECTIVE: To assess the efficacy and safety of retention enema with traditional Chinese medicine (TCM) for ulcerative colitis (UC) through a meta-analysis of published studies. METHODS: Literatures were retrieved from five electronic databases. Quality evaluation and meta-analysis were respectively conducted using the Cochrane collaboration and RevMan5.3. Overall quality of evidence was evaluated using GRADE system. Effect sizes were pooled using random effect models. RESULTS: Seventeen RCTs were included. Compared with routine pharmacotherapies (RPs), TCM enema exhibited a statistically significant difference in clinical efficacy and reduction of the recurrence rate. The results of qualitative description for other endpoints, such as improvements in anabrosis, ulcer, diarrhea, and hematochezia, suggested that TCM enema had better efficacy than RPs. Furthermore, the incidence of side effects in TCM was lower than that in RPs. CONCLUSION: This meta-analysis confirmed the efficacy and safety of TCM enema for improving UC symptoms. However, further well-designed researches are needed.

Multi-model ensemble simulated non-point source pollution based on Bayesian model averaging method and model uncertainty analysis.

Watershed models are cost-effective and powerful tools for evaluating and controlling non-point source pollution (NPSP), while the reliability of watershed models in a management context depends largely on inherent uncertainties in model predictions. The objective of this study is to present the use of multi-model ensemble applied to streamflow, total nitrogen (TN), and total phosphorus (TP) simulation and quantify the uncertainty resulting from model structure. In this study, three watershed models, which have different structures in simulating NPSP, were selected to conduct watershed monthly streamflow, TN load, and TP load ensemble simulation and 90% credible intervals based on Bayesian model averaging (BMA) method. The result using the observed data of the Yixunhe watershed revealed that the coefficient of determination and Nash-Sutcliffe coefficient of the BMA model simulate streamflow, TN load, and TP load were better than that of the single model. The higher the efficiency of a single model is, the greater the weight during the BMA ensemble simulation is. The 90% credible interval of BMA has a high coverage of measured values in this study. This indicates that the BMA method can not only provide simulation with better precision through ensemble simulation but also provide quantitative evaluation of the model structure through interval, which could offer rich information of the NPSP simulation and management.

Simiao Decoction Alleviates Gouty Arthritis by Modulating Proinflammatory Cytokines and the Gut Ecosystem.

Simiao decoction, a classical traditional Chinese medicine (TCM) formula, has been widely used for thousands of years due to its safety and efficiency in treating gouty arthritis. Utilizing serum proinflammatory cytokines and gut ecosystems, this study elucidated the mechanisms of alleviating gouty arthritis by Simiao decoction. Simiao decoction (4.0, 8.0, and 16.0 g/kg) was orally administered to gouty arthritis mice and febuxostat was given as a positive control. The spleen, kidney, and liver indexes indicated that Simiao decoction was safe for the treatment of gouty arthritis in C57BL/6 mice. Besides, our study demonstrated that Simiao decoction was effective for reducing the level of serum uric acid and decreasing MPO, XOD, and ADA activity, as well as alleviating gouty-related symptoms, such as foot swelling and pain. Moreover, Simiao decoction could also reduce some specific serum proinflammatory cytokines including IL-1ß, IL-9, IFN-γ, MIP-1α and MIP-1ß. We then surveyed the effects of Simiao decoction on the gut ecosystems in a systematic manner by combining network pharmacology, ELISA, western blot, and illumina sequencing. In the murine of model of gouty arthritis, Simiao decoction could suppress NLRP3 inflammasomes expression, reduce gut apoptosis through modulating TNF-α, Caspase 8, and AIFM1 protein expressions, affect lipid metabolism by regulating APOB, LPL, PPARα protein expressions and restore gut microbiota via reducing potential pathogens. Overall, these findings suggested that Simiao decoction was an effective therapeutic drug for gouty arthritis and the gut ecosystem might act as a potential anti-inflammatory target of Simiao decoction.

Qingda granules attenuate hypertensive cardiac remodeling and inflammation in spontaneously hypertensive rats.

Qingda granules (QDG) are derived from QingXuanJiangYa Decoction (QXJYD) a traditional Chinese medication that has been used to treat hypertension for more than 60 years. QXJYD has been shown to be effective in rat models of hypertension. However, the effects of QDG on hypertension remain largely unknown. In the current study, baicalin was identified as one of the main components of QDG using Ultra Performance Liquid Chromatography (UPLC) analysis. We investigated the effects of QDG on blood pressure, cardiac remodeling, and cardiac inflammation. QDG (0.8 g/kg/day) treatment attenuated the elevated blood pressure in spontaneously hypertensive rats (SHRs). Moreover, QDG treatment reduced the degree of myocardial fiber disarray, degeneration and necrosis of myocardial cells, expression of ANP and BNP, as well as collagen content of SHRs. Moreover, we further assessed the effect of QDG treatment on cardiac inflammation and found that QDG treatment reduced CD68 protein expression, decreased levels of IL-6 and TNF-α in both serum and cardiac tissues, as well as suppressed activation of NF-κB pathway in cardiac tissues of SHRs. Differential expressed metabolites (DEMs) analysis identified 41 increased and 51 decreased metabolites in the cardiac tissues of SHRs after QDG treatment. In summary, QDG treatment of SHRs attenuated the elevated blood pressure and ameliorated cardiac remodeling and inflammation, in part, through suppression of NF-κB pathway and DEMs, which provide a basis for other therapeutic uses of this TCM.

A steroidal saponin isolated from Allium chinense simultaneously induces apoptosis and autophagy by modulating the PI3K/Akt/mTOR signaling pathway in human gastric adenocarcinoma.

Allium chinense, as a side dish on Asian table, is often used in folk medicine for its health benefits. (25R)-5α-spirostan-3ß-yl-3-O-acetyl-O-ß-d-glucopyranosyl-(1 â†’ 2)-O-[ß-d-glucopyranosyl-(1 â†’ 3)]-O-ß-d-glucopyranosyl-(1 â†’ 4)-ß-d-galactopyranoside (A-24) is a bioactive steroidal saponin isolated from Allium chinense. Previously, we have shown that A-24 has cytotoxic effects on cancer cells, but not on normal cells. To further explore the underlying mechanisms, in this study, we investigated the anticancer activity of A-24 in human gastric cancer cell lines in terms of cell proliferation, colony formation, cell cycle, induction of apoptosis/autophagy, and PI3K/Akt/mTOR pathway. A-24 showed dose-dependent cytotoxicity in SGC-7901 and AGS cell lines, it induced intrinsic mitochondrial pathway of apoptosis as well as autophagy, G2/M phase arrest and modulation of cyclinB1, p-cdc2, p-wee1 and p-Histone H3 expression. Furthermore, A-24 downregulated the phosphorylation of Akt at Ser473 and mTOR at Ser2448 in PI3K/Akt/mTOR pathway, and its downstream substrates p-p70S6K and p-4EBP1 in a dose-dependent manner. In addition, the pre-treatment of tumor cells with 3-methyladenine (3-MA) and LY294002 increased A-24-induced apoptosis. Collectively, these findings highlight the significance of downregulation of PI3K/Akt/mTOR pathway in A-24-induced apoptosis and autophagy, and the potential application of A-24 as a novel candidate in the treatment of human gastric adenocarcinoma.

Huoxin pill attenuates myocardial infarction-induced apoptosis and fibrosis via suppression of p53 and TGF-ß1/Smad2/3 pathways.

Huoxin Pill (HXP), a Traditional Chinese Medicine, is used widely to treat patients with coronary heart disease and angina pectoris in China. However, the underlying protective mechanism of HXP on cardiac apoptosis and fibrosis has never been evaluated. Therefore, the aim of this study was to investigate the role of HXP in a myocardial infarction (MI) mouse model. The mice were randomly divided into 3 groups and subjected to surgical ligation of the left anterior descending (LAD) coronary artery or sham surgery (n = 6 for each group) and treated with HXP (50 mg/kg/day) or saline by gavage for 2 weeks. At 2 weeks post MI, we found that HXP significantly enhanced myocardial function and attenuated the increase of heart weight index (HWI) and pathological changes in MI mice. RNA-sequencing and KEGG pathway analyses identified 660 differentially expressed genes and multiple enriched signaling pathways including p53 and TGF-ß. In support of these findings, HXP attenuated cardiac apoptosis and decreased p53 and Bax protein expression, while increasing Bcl-2 protein expression in cardiac tissues of MI mice. Furthermore, HXP treatment inhibited cardiac fibrosis and significantly down-regulated TGF-ß1 protein expression and Smad2/3 phosphorylation in cardiac tissues. In summary, HXP can improve cardiac function in mice after MI by attenuating cardiac apoptosis and fibrosis partly via supression of the p53/Bax/Bcl-2 and TGF-ß1/Smad2/3 pathways.

Efficacy of traditional Chinese medicine for chronic gastritis: A meta-analysis of randomized controlled trials.

BACKGROUND: To systematically evaluate efficacy of traditional Chinese medicine (TCM) in treating chronic gastritis (CG). METHODS: Data sources from PubMed, Embase, Springer Link, China National Knowledge Infrastructure, Chinese Scientific Journals Database, Chinese Biomedicine Database, and Wan-fang database were searched up to July 5, 2018. Review Manager software version 5.3, the Cochrane Collaboration's risk of bias tool, and the Grading of Recommendations Assessment, Development, and Evaluation profiler software were conducted for this meta-analysis. RESULTS: Sixteen studies involving 1673 participants (906 vs 767) were included in this study. Pooled data showed significant statistical differences between TCM groups and current routine pharmacotherapy (RP) groups in overall clinical efficacy (odds ratio [OR] 4.65; 95% confidence interval [CI] 3.29, 6.56; P < .00001), efficacy under endoscopy (OR 2.46; 95% CI 1.12, 5.43; P = .03), stomach distension (mean difference [MD] -0.37; 95% CI -0.56, -0.19; P < .0001), stomachache (standardized MD [SMD] -0.80; 95% CI -1.45, -0.14; P = .02), and belching (SMD -2.00; 95% CI -3.80, -0.20; P = .03). However, acid regurgitation (SMD -0.71; 95% CI -1.69, 0.28; P = .16) and anorexia (SMD -0.75; 95% CI -2.30, 0.80; P = .35) showed no significant statistical differences between 2 groups. In addition, incidence of adverse reactions of TCM groups was lower than that of RP groups. CONCLUSION: Evidence from this meta-analysis suggests that TCM could be more efficacious than current RP in treating CG. But further standardized research of rigorous design should be needed to further validate its efficacy.

Nanoelemental selenium alleviated the mercury load and promoted the formation of high-molecular-weight mercury- and selenium-containing proteins in serum samples from methylmercury-poisoned rats.

Selenite (Se4+) has been found to counteract the neurotoxicity of methylmercury (MeHg) in MeHg-poisoned rats. However, Se4+ has narrow range between its toxic and beneficial effects. Nanoelemental selenium (SeNPs) was found to be less toxic than other forms of Se such as Se4+. In this study, the effects of SeNPs on the load of mercury (Hg) in rats were investigated. Hyphenated technique based on size-exclusion chromatography coupled with UV and inductively coupled plasma mass spectrometry (SEC-ICP-MS) detection and synchrotron radiation X-ray fluorescence spectroscopy (SR-XRF) were used to analyze the Hg-Se-containing proteins in the serum from MeHg-poisoned rats. The Hg-Se-containing fractions monitored by UV and ICP-MS were further characterized by MALDI-TOF-MS. Elevated serum Hg and Se levels were found in MeHg-poisoned rats after SeNPs treatment. Three main Hg-containing bands with molecular weights (MWs) of 25, 62 and 140 kDa were detected in the control samples. Treatment with SeNPs increased the Hg content in proteins at 62 and 170 kDa and decreased the Hg content at 25 kDa. The fraction with 25 kDa was assigned to metallothioneins (MTs), and fractions with 40 and 75 kDa were assigned to albumin. This study showed that the low-toxicity SeNPs could reduce the Hg load in the tissues and promote the formation of high molecular weight Hg- and Se-containing proteins in MeHg-poisoned rats.

Selenoprotein P as the major transporter for mercury in serum from methylmercury-poisoned rats.

Selenium (Se) has been found to promote weight gain, decrease hepatic damage, but redistribute mercury (Hg) in brains and livers in methylmercury (MeHg)-poisoned rats. The aims of the present work were to examine the effects of Se on the levels of Hg in serum and the role of serum selenoproteins in binding with Hg in MeHg-poisoned rats. The concentration of Se, Hg and MeHg were studied using ICP-MS and CVAFS. The Hg- and Se-binding selenoproteins were separated and quantified using affinity chromatography with post-column isotope dilution analysis using both enriched 78Se and 199Hg. It was found that Se treatment reduced Hg levels in serum in MeHg-poisoned rats. Among the three separated selenoproteins, the amounts of SelP-bound Hg and Se increased to 73% and 93.6%, from 64.4% and 89.3% of the total Hg and Se, respectively after Se treatment, suggesting that SelP acts as a major transporter for Hg and pool for Se in serum. Over 90% of the total Hg was MeHg in serum, and the molar ratios of MeHg to Se as 1:4 and 1:9 in the formed MeHg-Se-SelP complex in the control and the Se treatment group, respectively. The elevated Se level binding with SelP facilitated the Hg extraction from tissues and organs, as well as its redistribution in brains and livers through blood circulation in the MeHg-poisoned rats. Together, our findings provide direct evidence that serum SelP is the major Hg transporter in MeHg-poisoned rats.

Suppressing Nanoparticle-Mononuclear Phagocyte System Interactions of Two-Dimensional Gold Nanorings for Improved Tumor Accumulation and Photothermal Ablation of Tumors.

The clearance of nanoparticles (NPs) by mononuclear phagocyte system (MPS) from blood leads to high liver and spleen uptake and negatively impacts their tumor delivery efficiency. Here we systematically evaluated the in vitro and in vivo nanobio interactions of a two-dimensional (2D) model, gold (Au) nanorings, which were compared with Au nanospheres and Au nanoplates of similar size. Among different shapes, Au nanorings achieved the lowest MPS uptake and highest tumor accumulation. Among different sizes, 50 nm Au nanorings showed the highest tumor delivery efficiency. In addition, we demonstrated the potential use of Au naonrings in photoacoustic imaging and photothermal therapy. Thus, engineering the shape, surface area, and size of Au nanostructures is important in controlling NP-MPS interactions and improving the tumor uptake efficiency.

Root bark of Sambucus Williamsii Hance promotes rat femoral fracture healing by the BMP-2/Runx2 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Sambucus Williamsii Hance (SWH) is a plant from a family of Caprifoliaceae, which has a long medical history of use as an effective folk treatment for fracture bruises. AIM OF THE STUDY: To evaluate the effects of 50% ethanol extracts of root-bark of Sambucus Williamsii Hance(EE-rbSWH) on fracture healing of rats and explore its mechanism of actions related to the BMP-2 signaling pathway. MATERIALS AND METHODS: EE-rbSWH was orally administered at the doses of 340 and 680mg/kg to adult Sprague-Dawley rats with operation of open femur fracture completely for 2, 4 and 8 weeks. And the rats of sham operation and Model groups were administered Vehicle (distilled water 0.8mL/200g/day). Firstly, the bone X-ray morphology and bone mineral density(BMD) of the fracture site were observed and measured after anesthesia the rats at weeks 2, 4, and 8 after surgery, then the serum levels of alkaline phosphatase(ALP) and osteocalcin (BGP) were measured; Secondly, the tissue morphology of the fracture site was observed after sacrificed the rats; Thirdly, the formation of mineralized nodules in bone marrow stromal cells(BMSC) were evaluated at week 2; Lastly, the genes levels of BMP-2 and Runx2 in the femur were detected at week 2 and 4, and the proteins expression of BMP-2 signaling pathway (BMP-2, BMPRIB, BMPRII and Runx2) in the femur also were detected at week 2. RESULTS: EE-rbSWH remarkably accelerated fracture healing by promoting bone formation at all the time points of fracture healing. Mainly by increasing the BMD level at the fracture site, the levels of serum ALP and BGP, and also the numbers increasing of calcified nodules in BMSC. The mechanism studies, EE-rbSWH can promote fracture healing by enhancing the expressions of BMP-2 and Runx2 mRNA, and also the proteins of BMP-2, BMPRIB, BMPRII and Runx2 at the fracture site of rats. CONCLUSIONS: Our results suggested that 50% ethanol extracts of root-bark of Sambucus Williamsii Hance can accelerate fracture healing by recruitment of osteoblasts at the fracture site and through up-regulation of the BMP-2 signaling pathway.

Sambucus Williamsii Hance Promotes MC3T3-E1 Cells Proliferation and Differentiation via BMP-2/Smad/p38/JNK/Runx2 Signaling Pathway.

The 50% ethanol elution fractions of root-bark of Sambucus Williamsii Hance (rbSWH) evaluated the effect of proliferation and differentiation on preosteoblast MC3T3-E1 cell, and the mechanism of actions. We found that rbSWH(30, 60, and 90 µg/mL) can enhance cell proliferation by MTT assay and promote alkaline phosphatase (ALP) and bone Gla protein (BGP) activities, type I collagen (Col-I) synthesis, and mineralization nodule formation in primary cultured osteoblasts. The results showed that rbSWH can increase mRNA levels of BMP-2 and Runx2 using real-time reverse transcription-quantitative polymerase chain reaction, whereas the BMP-2 antagonist Noggin attenuated the increase of ALP activity induced by rbSWH, indicating that BMP-2 expression was required for the action of rbSWH in osteoblastic. We also found that rbSWH can enhance the expressions of BMP-2, BMPRIB, BMPRII, phosphorylation of Smad, JNK and p38, and Runx2 proteins by western blotting. In addition, pretreatment of cells with p38 inhibitor (SB203580) or JNK inhibitor (SP600125) can antagonize the elevation of BMP-2 expression, ALP activity, and cell viability induced by rbSWH. Taken together, our results provided an evidence that rbSWH can promote MC3T3-E1 cell proliferation and differentiation via BMP-2/Smad/p38/JNK/Runx2 signaling pathway.