Unveiling the Therapeutic Potential of Herba Epimedii: Enhancing Bone Healing Through Cytoskeletal Regulation of RhoA/Rock1 Pathway.

Herba Epimedii is widely used to promote bone healing, and their active ingredients are total flavonoids of Epimedium (TFE). Ras homolog gene family member A / Rho-associated protein kinase (RhoA/Rock), an important pathway regulating the cytoskeleton, has been proven to affect bone formation. However, whether TFE promotes bone healing via this pathway remains unclear. In this study, the therapeutic effects of TFE were estimated using micro-computed tomography and hematoxylin and eosin staining of pathological sections. F-actin in osteoblasts was stained to investigate the protective effects of TFE on the cytoskeleton. Its regulatory effects on the RhoA/Rock1 pathway were explored using RT-qPCR and Western blot analysis. Besides, flow cytometry, alkaline phosphatase and nodule calcification staining were performed to evaluate the effects on osteogenesis. The bone healing in rats was improved, the cytoskeletal damage in osteoblasts was reduced, the RhoA/Rock1 pathway was downregulated, and osteogenesis was enhanced after TFE treatment. Thus, TFE can promote bone formation at least partially by regulating the expression of key genes and proteins in the cytoskeleton. The findings of this study provided evidence for clinical applications and would contribute to a better understanding of Epimedium's mechanisms in treating bone defects.

Influences of miR-378a-3p on the Pathogenesis of Allergic Rhinitis via GzmB-Mediated Inflammatory Reaction.

Methods: Totally, 24 BALB/c mice were assigned to the AR group, control group, GzmB group, and blank group (each n = 6). The blank group was normally fed without treatment, and the other three groups were treated by ovalbumin (OVA) to induce AR models, in which the GzmB group was intranasally injected with lentiviral vector suppressing GzmB expression during the second immunization, while the control group was given the GzmB-blank vector. The times of AR pathological behaviours such as sneezing and scratching the nose of mice were observed and counted. The nasal lavage fluid of each mouse was acquired, and then, the mouse was executed by cervical dislocation, followed by collection of blood and nasal mucosa tissues. Then, ELISA was adopted for quantifying immunoglobulin E (IgE), interleukin (IL)-4, IL-6, and histamine (HA), and nasal mucosa tissues were treated by HE and TUNEL staining to observing their histopathological manifestations. PCR and western blot (WB) were adopted for quantifying GzmB and miR-378a-3p. Additionally, with NP69 cells, dual luciferase reporter (DLR) assay was carried out for determining the targeting association of GzmB with miR-378a-3p. Another 24 mice were assigned to the AR group, GzmB group, miR-378a-3p group, and GzmB+ miR-378a-3p group (each n = 6). The AR and GzmB groups were treated as above. The miR-378a-3p group was intervened by lentiviral vector suppressing miR-378a-3p, while the GzmB+ miR-378a-3p group was given GzmB and lentiviral vector suppressing miR-378a-3p meantime. A rescue assay was conducted through repeating the above tests. Results: The times of sneezing and rubbing the nose and the levels of IgE, IL-4, IL-6, and HA were similar between the control and AR groups (all P > 0.05), and these items of the two groups were all higher than those of the blank and GzmB groups (all P < 0.05). However, no notable difference was observed in IL-4 and IL-6 levels between the GzmB and blank groups (both P > 0.05), while higher levels of other detection results were found in the former group than in the latter (all P < 0.05). The staining results revealed obvious congestion, oedema, and necrosis structures in the nasal mucosa epithelium of the control and AR groups and also revealed a large number of infiltrating eosinophils and notable increase of apoptotic nasal mucosa epithelial cells. The GzmB group showed notably improved nasal mucosa tissues, and its infiltration and apoptosis of eosinophils were more notable than those of the blank group, but notably weaker than those of the AR and control groups. Additionally, the PCR and WB results revealed similar miR-378a-3p and GzmB levels in nasal mucosa between the control and AR groups (both P > 0.05), and a notable decrease of miR-378a-3p and a notable increase of GzmB in both groups (both P < 0.05). The DLR assay revealed notably suppressed fluorescence activity of GzmB-WT in NP69 cells after transfection of miR-378a-3p mimics (P < 0.05) and notably down regulated GzmB protein after increase of miR-378a-3p (P<0.05). Finally, the rescue assay revealed that downregulating miR-378a-3p aggravated the pathological changes of AR (P < 0.05) and also completely reversed the impacts of inhibiting GzmB on the pathological behaviours of AR mice. Conclusions: MiR-378a-3p can accelerate the pathological development of AR through targeted inhibition on the release of pro-inflammatory factors such as IgE and HA activated by GzmB, so it is a promising molecular target of AR therapy and offers a novel research direction for the complete cure of AR.

Erxian herbal pair enhances bone formation in infected bone nonunion models and attenuates lipopolysaccharide-induced osteoblastinhibition by regulating miRNA-34a-5p.

Bacterium-induced inflammatory responses cause bone nonunion. Although antibiotics suppress infection, bone loss after antibacterial treatment remains a critical challenge. Erxian herbal pair (EHP) has been proven effective in promoting bone formation. Our study aimed to investigate the effect of EHP on bone repair after anti-infection treatment, explore its effect on a lipopolysaccharide (LPS)-induced osteoblast. We evaluated effects of EHP on bone repair with Micro-CT, and morphology detecting. Chemical constituents of EHP and EHP-containing serum (EHP-CS) were identified by UHPLC-Q/TOF-MS. In addition, osteoblast induced by LPS was established and administrated with EHP-CS. Cell proliferationwas assessed by MTT. Target prediction identified SMAD2 as a potential target of miRNA-34a-5p. MiRNA mimic, inhibitor and siRNA were transiently transfected into osteoblasts. The mRNA levels and protein expressions of miRNA-34a-5p, BMP2, Runx2, SMAD2 were assessed. The results showed that the main biocactivity ingredients in EHP-CS were Baohuoside Ι and Orcinol Glucoside. EHP could promote bone remolding after anti-infection therapy and restore the activity of LPS-induced osteoblasts. Moreover, miRNA-34a-5p was dramatically downregulated and SMAD2 was upregulated after LPS stimulation, while EHP resisted the inhibition of LPS by promoting miRNA-34a-5p, ALP, and BMP2 expressions. Whereas downregulation of miRNA-34a-5p reversed these effects. Silencing endogenous SMAD2 expression markedly promoted BMP2 and ALP activity and enhanced osteogenesis. Taken together, EHP restored LPS-induced bone loss by regulating miRNA-34a-5p levels and repressing its target gene SMAD2. EHP might be a potential adjuvant herbal remedy for the treatment of bone nonunion, and miRNA-34a-5p is a novel target for controlling bone and metabolic diseases.

Timosaponin AIII attenuates inflammatory injury in AGEs-induced osteoblast and alloxan-induced diabetic osteoporosis zebrafish by modulating the RAGE/MAPK signaling pathways.

BACKGROUND: Advanced glycation end products (AGEs) deposition causes inflammatory injury in osteoblasts and contributes to diabetic osteoporosis. The receptor for advanced glycation end product/mitogen-activated protein kinase pathway (RAGE/MAPK) signaling pathway is closely linked to the pathogenesis of diabetic osteoporosis. Timosaponin AIII, a steroidal saponin isolated from Anemarrhena asphodeloides Bunge (Asparagaceae), shows anti-inflammatory and anti-osteoporosis effects. PURPOSE: The present study was aimed to investigate the therapeutic effects of timosaponin AIII on diabetic osteoporosis and whether its effect is dependent on protecting osteoblasts against AGEs-induced injury via RAGE/MAPK signaling suppression. METHODS: An alloxan-induced diabetic osteoporosis zebrafish model was applied to investigate the effects of timosaponin AIII in vivo, and alendronate was used as a positive control. Moreover, related mechanisms were explored in primary rat osteoblasts. Molecular docking was applied to investigate the interactions between timosaponin AIII and RAGE. RESULTS: Timosaponin AIII treatment reversed alloxan-induced reduction in the mineralized area of the larvae head skeleton, accompanied by a decreased level of triglyceride and total cholesterol in the zebrafish. Additionally, AGEs significantly influenced RAGE expression, alkaline phosphatase activity, interleukin 1ß expression, interleukin 6 expression, and tumor necrosis factor-α expression, and increased cell apoptosis. Timosaponin AIII significantly downregulated AGEs-induced interleukin 1ß, interleukin 6, and tumor necrosis factor-α levels, and upregulated alkaline phosphatase and osteocalcin levels. Timosaponin AIII also significantly reduced the expression of RAGE and had additive effects on downstream P38, extracellular signal-regulated kinase and c-Jun N-terminal kinase in AGEs-induced osteoblast. Molecular docking predicted that hydrogen and hydrophobic interactions occurred between timosaponin AIII and RAGE. CONCLUSION: These data clarified that timosaponin AIII attenuates diabetic osteoporosis via a novel mechanism involved suppressing the RAGE/MAPK signaling pathway. Our finding highlights the potential value of timosaponin AIII as an anti-diabetic osteoporosis agent.

Preparation and application of polyvinyl alcohol-decorated cell membrane chromatography for screening anti-osteoporosis components from Liuwei Dihuang decoction-containing serum.

Cell membrane chromatography is a powerful tool for screening active components from complex matrices. New cell membrane carriers need to be developed to increase the coverage of cell membranes on the surface of stationary phases, thereby improving cell membrane chromatographic retention. In this work, we prepared polyvinyl alcohol-poly dimethyl diallyl ammonium chloride modified silica gel as a cell membrane carrier. Osteoblast cell was used as cell membrane source, which was widely used to evaluate the osteogenic activity of drugs. The new cell membrane chromatographic stationary phase was used to screen anti-osteoporosis components in Liuwei Dihuang decoction-containing serum. The chemical structures of the new modified materials were characterized by scanning electronic microscopy, Fourier transform infrared spectroscopy, and elemental analysis characterization. Compared with the common cell membrane column, the cell membrane coverage of this modified material was increased by 30%, and thus provided a stronger retention effect in positive drugs. Nineteen metabolites in rat serum samples were retained on the cell membrane chromatography and identified by ultra-high-performance liquid chromatography/time-of-flight mass spectrometry. Among those, four components (morroniside, catalpol, loganin, and acteoside) were selected for in vitro pharmacodynamics validation. They significantly increased the osteoblast proliferation. The new modified material was successfully applied to screen anti-osteoporosis components from Liuwei Dihuang Decoction-containing serum.

Molten salt synthesis and luminescence of Dy3+ -doped Y3 Al5 O12 phosphors.

Dy3+ -doped Y3 Al5 O12 phosphors were prepared at a relatively low temperature using molten salt synthesis. The phase of the prepared Dy3+ -doped Y3 Al5 O12 phosphors was confirmed using X-ray powder diffraction. Results indicated that Dy3+ doping did not change the Y3 Al5 O12 phase. Following excitation at 352 nm, emission spectra of the Dy3+ -doped Y3 Al5 O12 phosphors consisted of blue, yellow, and red emission bands. The influence of Dy3+ concentration and excitation wavelength on emission was investigated. The ratio of yellow light to blue light varied with change in Dy3+ doping concentration, due to changes in the structure around Dy3+ . Emission intensities also changed when the excitation wavelength was changed. This variation is luminescence generated a system for tunable white light for Dy3+ -doped Y3 Al5 O12 phosphors.

Preparation and characterization of a novel colorimetric indicator film based on gelatin/polyvinyl alcohol incorporating mulberry anthocyanin extracts for monitoring fish freshness.

This work aimed to develop a novel colorimetric indicator film for monitoring of food freshness based on gelatin/polyvinyl alcohol matrix incorporated with anthocyanin extracts from mulberry. The color of anthocyanin extracts solutions obviously changed from bright red to dark green in the pH range of 2.0-11.0. FTIR spectra and isothermal titration calorimetry showed that the anthocyanin extracts were successfully combined with gelatin/polyvinyl alcohol matrix by hydrogen binding and electrostatic interaction, which enhanced the stability of anthocyanin. The scanning electric microscopy showed that the compatibility between polyvinyl alcohol and gelatin were improved owing to the addition of anthocyanin extracts. With the anthocyanin extracts addition from 0 to 45 mg/100 mL mixed solution, the tensile strength decreased from 30.80 to 21.01 MPa and the elongation at break increased from 589.22% to 905.86%. The color response of film in buffer solution of different pH were in accordance with anthocyanin extracts solutions, and its color changes were clearly visible with naked eye. Finally, the film was evaluated by a test on monitoring fish spoilage, which presented visible color changes due to volatile nitrogenous compounds formed over time. These results showed that this developed film could be used as an effective method for the monitoring of food freshness.

Integrated pathological cell fishing and network pharmacology approach to investigate main active components of Er-Xian decotion for treating osteoporosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Oxidative damage to osteoblasts was a key factor in the development of osteoporosis. Er-Xian Decotion (EXD) is widely used in China for the treatment of osteoporosis, which has a variety of antioxidant active ingredients. EXD may be an important source of protection against oxidative damage in osteoblasts, but the anti-osteoporotic active components of EXD is currently unclear. AIM OF THE STUDY: This work established an effective and reliable drug screening method to find main active ingredients in EXD (M-EXD) that can protect osteoblasts against oxidative stress and achieve anti-osteoporosis effects. MATERIALS AND METHODS: H2O2-induced osteoblast cell fishing with UHPLC-QTOF/MS was firstly used to discover the potential active components from EXD. Afterword, the EXD compound-osteoporosis target network was constructed using network pharmacology, thus potentially anti-osteoporosis ingredients were founded, and their combination were defined as the M-EXD. Finally, pharmacology effects of M-EXD was evaluated by ovariectomized rats, prednisolone induced-zebrafish and H2O2-induced osteoblasts. RESULTS: 40 candidate active ingredients in EXD were initially screened out via pathological cell fishing. According to network pharmacology result, M-EXD consisted of 13 ingredients since they had a close relationship with 65 osteoporosis-related targets. Pharmacological evaluation showed that M-EXD significantly ameliorated oxidative stress in H2O2-induced osteoblast model, evidently reversed the activity of ALP, ROS, GSH-px, NO and MDA compared with the model group. M-EXD showed better anti-oxidative activities than individual ingredients, presenting obvious synergetic effects. In osteoporosis rat and zebrafish models, M-EXD also demonstrated good anti-osteoporotic properties by mitigating the osteoporosis bone loss and increasing serum bone morphogenetic protein 2, and reversing osteocalcin expression in bone tissue. It significantly ameliorated oxidative stress in the in-vivo models. Moreover, M-EXD and EXD showed similar anti-osteoporotic and anti-oxidative properties, while the rest components of EXD had no satisfactory anti-osteoporotic efficacy. CONCLUSIONS: Our work successfully identified the main active components in EXD, which could represent the efficacy of EXD on treating osteoporosis, and meanwhile, it also provided an effective strategy to investigate active ingredients from natural medicines, which might be helpful for drug development and application.

Treatment with Vancomycin Loaded Calcium Sulphate and Autogenous Bone in an Improved Rabbit Model of Bone Infection.

Bone infection results from bacterial invasion, which is extremely difficult to treat in clinical, orthopedic, and traumatic surgery. The bone infection may result in sustained inflammation, osteomyelitis, and eventual bone non-union. Establishment of a feasible, reproducible animal model is important to bone infection research and antibiotic treatment. As an in vivo model, the rabbit model is widely used in bone infection research. However, previous studies on rabbit bone infection models showed that the infection status was inconsistent, as the amount of bacteria was variable. This study presents an improved surgical method for inducing bone infection on a rabbit, by blocking the bacteria in the bone marrow. Then, multi-level evaluations can be carried out to verify the modelling method. In general, debriding necrotic tissue and implantation of vancomycin-loaded calcium sulphate (VCS) are predominant in antibiotic treatment. Although calcium sulphate in VCS benefits osteocyte crawling and new bone growth, massive bone defects occur after debriding. Autogenous bone (AB) is an appealing strategy to overcome bone defects for the treatment of massive bone defects after debriding necrotic bone. In this study, we used the tail bone as an autogenous bone implanted in the bone defect. Bone repair was measured using micro-computed-tomography (micro-CT) and histological analysis after animal sacrifice. As a result, in the VCS group, bone non-union was consistently obtained. In contrast, the bone defect areas in the VCS-AB group were decreased significantly. The present modeling method described a reproducible, feasible, stable method to prepare a bone infection model. The VCS-AB treatment resulted in lower bone non-union rates after antibiotic treatment. The improved bone infection model and the combination treatment of VCS and autogenous bone could be helpful in studying the underlying mechanisms in bone infection and bone regeneration pertinent to traumatology orthopedic applications.

Effects of alkaloids from Sophora flavescens on osteoblasts infected with Staphylococcus aureus and osteoclasts.

Chronic osteomyelitis is primarily caused by infection with Staphylococcus aureus (S. aureus). Antibiotics are commonly administered; however, it is a challenge to promote bone healing. The aim of this study was to investigate the in vitro effects of alkaloids from the herbal remedy Sophora flavescens (ASF) on rat calvarial osteoblasts (ROBs) infected with S. aureus and healthy osteoclasts. Cell proliferation and alkaline phosphatase, interleukin-6, and tumour necrosis factor-α activity was measured in infected ROBs; tartrate-resistant acid phosphatase was evaluated in osteoclasts via enzyme-linked immunosorbent assay. The mRNA and protein expression levels of bone morphogenetic protein 2, runt-related transcription factor 2, osteoprotegerin, and receptor activator of nuclear factor kappa-B ligand were assessed in infected ROBs through reverse transcription-polymerase chain reaction and western blotting analysis, respectively. Results indicated that ASF increased the viability of uninfected ROBs and infected ROBs treated with vancomycin via regulation of bone morphogenetic protein 2, runt-related transcription factor, osteoprotegerin, and receptor activator of nuclear factor kappa-B ligand mRNA and protein expression levels. In addition, the secretion of the inflammatory factor tumour necrosis factor-α was decreased and alkaline phosphatase activity was increased, inhibiting the viability of osteoclasts and tartrate-resistant acid phosphatase activity. Therefore, the herbal remedy ASF has potential as a new treatment for chronic osteomyelitis.

Icariin Enhances Bone Repair in Rabbits with Bone Infection during Post-infection Treatment and Prevents Inhibition of Osteoblasts by Vancomycin.

Vancomycin is an effective antibiotic for treatment of bone infection caused by Staphylococcus aureus, however, a high local concentration of vancomycin might induce a delay in bone union. Icariin has been reported to suppress osteoclastogenes and promote osteogenesis. Our study aimed to investigate the effect of icariin on bone repair after anti-infection treatment in vivo and to explore the resisting effect of icariin on rat calvarial osteoblasts (ROBs) inhibited with high doses of vancomycin. Rabbits with bone infection of S. aureus were treated with implanted vancomycin-calcium sulfate (VCS) and icariin at 10.86 mg/kg/day for consecutive 8 weeks. Micro-CT, morphology, blood biochemistry were evaluated. In addition, ROBs were treated with vancomycin and icariin at different doses. Cell proliferation and differentiation capabilities, BMP2, Runx2, OPG, RANKL mRNA levels and protein expression were assessed. The results indicated that high dose of vancomycin significantly decreased bone mass and inhibited osteocalcin secretion; icariin increased these indicators compared with the single vancomycin treatment. Over 0.1 mg/mL of vancomycin inhibited the proliferation and differentiation of ROBs, while icariin resisted the inhibition of vancomycin by regulating cell cycle and promoting the Alkaline phosphatase (ALP) activity. Moreover, icariin promote bone formation by up-regulating BMP2/Runx2 and OPG/RANKL pathways. Icariin exhibited osteoplastic properties on osteoblasts that had been inhibited with high doses of vancomycin. Therefore, icariin is helpful for post-infection treatment of bone infection.

A Network Pharmacology Approach to Determine the Active Components and Potential Targets of Curculigo Orchioides in the Treatment of Osteoporosis.

BACKGROUND Osteoporosis is a complex bone disorder with a genetic predisposition, and is a cause of health problems worldwide. In China, Curculigo orchioides (CO) has been widely used as a herbal medicine in the prevention and treatment of osteoporosis. However, research on the mechanism of action of CO is still lacking. The aim of this study was to identify the absorbable components, potential targets, and associated treatment pathways of CO using a network pharmacology approach. MATERIAL AND METHODS We explored the chemical components of CO and used the five main principles of drug absorption to identify absorbable components. Targets for the therapeutic actions of CO were obtained from the PharmMapper server database. Pathway enrichment analysis was performed using the Comparative Toxicogenomics Database (CTD). Cytoscape was used to visualize the multiple components-multiple target-multiple pathways-multiple disease network for CO. RESULTS We identified 77 chemical components of CO, of which 32 components could be absorbed in the blood. These potential active components of CO regulated 83 targets and affected 58 pathways. Data analysis showed that the genes for estrogen receptor alpha (ESR1) and beta (ESR2), and the gene for 11 beta-hydroxysteroid dehydrogenase type 1, or cortisone reductase (HSD11B1) were the main targets of CO. Endocrine regulatory factors and factors regulating calcium reabsorption, steroid hormone biosynthesis, and metabolic pathways were related to these main targets and to ten corresponding compounds. CONCLUSIONS The network pharmacology approach used in our study has attempted to explain the mechanisms for the effects of CO in the prevention and treatment of osteoporosis, and provides an alternative approach to the investigation of the effects of this complex compound.

Carbon nanotube-polymer composite for effervescent pipette tip solid phase microextraction of alkaloids and flavonoids from Epimedii herba in biological samples.

An effervescent pipette tip solid phase microextraction based on carbon nanotube-polymer composite microspheres was developed for the simultaneous extraction and determination of four alkaloids (magnoflorine, epiberberine, palmatine, and jatrorrhizine) and four flavonoids (epimedin A/B/C and icariin) from Epimedii herba in biological samples. In this work, the polymeric sorbent was prepared by incorporating carbon nanotube in sulfonated polystyrene-divinylbenzene microspheres. During the extraction, the dispersion of the sorbents at milligram level was achieved by the in situ generation of carbon dioxide. The effervescence enhanced the interaction between the sorbent and the analytes. Langmuir and Freundlich models were used to evaluate adsorption processes. Under the optimum analytical conditions, the method showed good linearity (3-300µgL-1), acceptable precision (RSD <5%), low limits of quantification (1.02-2.98µgL-1) and satisfactory recoveries (90.05-99.85%). The proposed method was successfully applied to the analysis of alkaloids and flavonoids from Epimedii herba in cell culture fluids, rat urine and feces obtained at the different time intervals.

Antioxidant property of water-soluble polysaccharides from Poria cocos Wolf using different extraction methods.

Poria cocos Wolf is a popular traditional medicinal plant that has invigorating activity. Water-soluble polysaccharides (PCPs) are its main active components. In this study, four different methods were used to extract PCPs, which include hot water extraction (PCP-H), ultrasonic-assisted extraction (PCP-U), enzyme-assisted extraction (PCP-E) and microwave-assisted extraction (PCP-M). Their chemical compositions and structure characterizations were compared. In vitro antioxidant activities were studied on the basis of DPPH radical, hydroxyl radical, reducing power and metal chelating ability. The results showed that PCPs were composed of mannose, glucose, galactose, and arabinose, and had typical IR spectra characteristics of polysaccharides. Compared with other PCPs, PCP-M had lower neutral sugar content, higher mannose content and higher uronic acid content. The molecular weight were determined as PCP-E

Pipette tip solid-phase extraction and high-performance liquid chromatography for the determination of flavonoids from Epimedii herba in rat serum and application of the technique to pharmacokinetic studies.

Epimedii herba is a traditional Chinese medicine for the treatment of osteoporosis. Epimedin A, B and C and icariin are the primary effective ingredients of this medicine. In this study, a simple and low-cost method based on pipette tip solid-phase extraction, high-performance liquid chromatography separation, and diode array detection has been developed for the simultaneous analysis of four flavonoids (epimedin A, B and C and icariin) from Epimedii herba in rat serum samples. In this novel extraction configuration, the sorbents were placed between a filter (hollow fiber) and the pipette tip. Pipette tip solid-phase extraction has several advantages compared to conventional extraction methods: faster extraction time (6.0min); lower sample volume (100µL); lower solvent volume (100µL); and less solvent waste. Under the optimum extraction conditions, the method showed good linearity (0.05-10.0µgmL(-1)), acceptable intra- and inter precision (RSD<6%), low limits of quantification (0.027-0.045µgmL(-1)) and satisfactory relative recoveries (98.63-103.18%). This method was successfully applied to investigate the pharmacokinetics of the major flavonoids in Epimedii herba extract after oral administration to rats (10gkg(-1) body weight). The primary pharmacokinetic parameters for rats were determined as follows: Cmax, 0.45-4.11µgmL(-1); Tmax, 0.21-0.26h; t1/2α, 0.06-0.12h; t1/2ß, 2.02-3.48h; AUC0-∞: 0.50-2.58µghmL(-1); CL, 19.53-44.72Lkg(-1)h(-1); and MRT0-∞, 2.25-3.77h. The developed method has the potential to promulgate the pharmacokinetics and provide more information for clinical applications.

In vitro and in vivo antioxidant and antimutagenic activities of polyphenols extracted from hops (Humulus lupulus L.).

BACKGROUND: Hops (Humulus lupulus L.) contain 40-140 mg g(-1) polyphenols. The objective of this study was to determine the phenolic composition of a high-purity (total phenolic content = 887 mg g(-1) ) hop polyphenol extract (HPE) and evaluate its antioxidant activities in vivo and in vitro and its antimutagenic activity. The antioxidant activity of HPE was compared with the activity of green tea polyphenols. RESULTS: The phenolic compositions of HPE were more than 55% proanthocyanidins and more than 28% flavonoid glycosides. In vitro, HPE effectively scavenged α,α-diphenyl-ß-picrylhydrazyl, hydroxyl and superoxide anion radicals, and inhibited DNA oxidative damage. In vivo, oral HPE at a polyphenol dose of 200-800 mg kg(-1) body weight significantly prevented a bromobenzene-induced decrease in liver superoxide dismutase and glutathione peroxidase activity, and decreased levels of liver thiobarbituric acid reactive substances in bromobenzene-treated mice. An oral dose of 20-80 mg kg(-1) body weight HPE significantly reduced the frequency of bone marrow micronuclei induced by cyclophosphamide. The antioxidant activities of hop polyphenols in vitro and in vivo were higher than green tea polyphenols at the same concentration. CONCLUSION: Hop polyphenols had the same or higher antioxidant activity than tea polyphenols. Hop polyphenols might be useful as natural antioxidants and antimutagens.

Qindan capsule changes adventitial collagen synthesis in spontaneously hypertensive rats.

OBJECTIVE: To investigate the effect of Qindan capsule (QC) on collagen synthesis and the mechanism underlying the process in spontaneously hypertensive rats (SHRs). METHODS: Twentyfour SHRs were divided into three groups: the hypertension model group, the QC treatment group, and the losartan treatment group. Eight Wistar Kyoto (WKY) rats were used as the normal control group. The systolic blood pressure (SBP) of the rats was monitored, and the thoracic aorta adventitia of the rats was segregated. The expressions of transforming growth factor 1 (TGF-ß1), Smad3, and collagens I and were measured by histological staining and reverse transcription polymerase chain reaction. RESULTS: The SBP was significantly higher in the model group than in the normal control group (P<0.01). However, a significant SBP-lowering effect was observed in QC or losartan treatment groups (P<0.05 or P<0.01) after 3 weeks of treatment. QC-treated rats showed a decrease of approximately 40 mm Hg, and the losartan-treated rats showed a decrease of approximately 50 mm Hg at the end of treatment compared with the beginning of treatment. The protein and gene levels of TGF-ß1, Smad3, and collagens I and in the model group were significantly increased compared with those in the normal control group (P<0.01). However, the levels were significantly decreased in the QC or losartan treatment group compared with the model group (P<0.05 or P<0.01). However, there was no significant difference between the QC and losartan treatment groups (P<0.05). CONCLUSIONS: QC could exert its antihypertensive effect through down-regulating TGF-ß1-stimulated collagen expressions. The TGF-ß1/Smad3 signaling pathway may be involved in this process.

Hyodeoxycholic acid improves HDL function and inhibits atherosclerotic lesion formation in LDLR-knockout mice.

We examined the effects of a natural secondary bile acid, hyodeoxycholic acid (HDCA), on lipid metabolism and atherosclerosis in LDL receptor-null (LDLRKO) mice. Female LDLRKO mice were maintained on a Western diet for 8 wk and then divided into 2 groups that received chow, or chow + 1.25% HDCA, diets for 15 wk. We observed that mice fed the HDCA diet were leaner and exhibited a 37% (P<0.05) decrease in fasting plasma glucose level. HDCA supplementation significantly decreased atherosclerotic lesion size at the aortic root region, the entire aorta, and the innominate artery by 44% (P<0.0001), 48% (P<0.01), and 94% (P<0.01), respectively, as compared with the chow group. Plasma VLDL/IDL/LDL cholesterol levels were significantly decreased, by 61% (P<0.05), in the HDCA group as compared with the chow diet group. HDCA supplementation decreased intestinal cholesterol absorption by 76% (P<0.0001) as compared with the chow group. Furthermore, HDL isolated from the HDCA group exhibited significantly increased ability to mediate cholesterol efflux ex vivo as compared with HDL of the chow diet group. In addition, HDCA significantly increased the expression of genes involved in cholesterol efflux, such as Abca1, Abcg1, and Apoe, in a macrophage cell line. Thus, HDCA is a candidate for antiatherosclerotic drug therapy.

The effects of qindan-capsule-containing serum on the TGF-ß1/ERK signaling pathway, matrix metalloproteinase synthesis and cell function in adventitial fibroblasts.

CONTEXT: Qindan capsule (QC), a compound used in traditional Chinese medicine, has been used as an anti-hypertensive agent in clinical settings for years. Our previous studies have shown that QC can improve the morphological index of the artery, down-regulate the collagen volume fraction in the media and inhibit the transformation of smooth muscle cells. However, the detailed mechanisms underlying its effects require further investigation, which might provide more scientific evidence for the clinical treatment of hypertensive vascular remodeling (VR). OBJECTIVE: We investigated the effects of QC-containing serum on the TGF-ß1/ERK signaling pathway, cell proliferation, migration, the cell cycle, apoptosis and matrix metalloproteinase synthesis (MMPs) in rat aortic adventitial fibroblasts (AFs). MATERIALS AND METHODS: AFs were cultured through tissue explants in vitro. The levels of extracellular signal-regulated kinase 1/2 (ERK1/2), phospho-ERK1/2 (p-ERK1/2), connective tissue growth factor (CTGF), MMP2 and MMP9 expression were measured by western blotting and RT-PCR. The proliferation and migration of AFs were measured by MTT and transwell migration assays. Cell cycle progression and apoptosis in AFs were analyzed by flow cytometry. RESULTS: The proliferation and migration rates of AFs treated with transforming growth factor ß1 (TGF-ß1) for 24 h were 2.4 ± 0.75 and 2.2 ± 0.06 times higher than those of untreated AFs, and increases in the expression of p-ERK1/2 (3.7 ± 0.15 times), CTGF (3.3 ± 0.24 times), MMP2 (5.7 ± 0.37 times) and MMP9 (5.4 ± 0.46 times) (p < 0.05) were observed. Treatment with QC-containing serum significantly down-regulated cell proliferation (1.9 ± 0.06 times), migration (1.6 ± 0.05 times) and the expression of p-ERK1/2 (1.3 ± 0.75 times), CTGF (1.8 ± 0.64 times), MMP2 (1.6 ± 0.65 times) and MMP9 (1.4 ± 0.46 times) (p < 0.05). We also found that QC-containing serum down-regulated the percentage of cells in the G1 phase by 1.6 ± 0.43 times and increased early-phase apoptosis by 2.3 ± 0.33 times (p < 0.05) in AFs. CONCLUSIONS: QC effectively inhibits the proliferation and migration of AFs and changes cell bioactivity and MMPs, possibly through the TGF-ß/ERK/CTGF signaling pathway. Our findings may provide new insights into the potential function of QC in preventing or treating hypertension.

Silibinin restores paclitaxel sensitivity to paclitaxel-resistant human ovarian carcinoma cells.

BACKGROUND: Drug resistance and tumor metastasis are the main causes of treatment failure and mortality in cancer patients. Silibinin, a naturally occurring flavanone, has been shown to be a potent sensitizer for apoptosis induced by a variety of anticancer drugs. In this study, whether silibinin could overcome chemoresistance and reduce the invasiveness of A2780/taxol cells was investigated. MATERIALS AND METHODS: A2780 and A2780/taxol cells were treated with silibinin alone and in combination with paclitaxel. Cell viability was determined by MTT assay while apoptosis and cell cycle progression were assessed by flow cytometric analysis. Matrigel invasion assays assessed the invasive activity. Protein and mRNA levels influenced by the treatment were studied by Western blots and quantitative real-time PCR. RESULTS: Silibinin enhanced the sensitivity of A2780/taxol cells to paclitaxel, increased paclitaxel-induced apoptosis and G2/M arrest consistent with the down-regulation of survivin and P-glycoproteins. A2780/taxol cells demonstrated a two-fold increase in invasiveness ability compared to A2780 cells, whereas the invasive potential was reduced dramatically by silibinin. CONCLUSION: These results suggest silibinin in combination with paclitaxel may be a beneficial chemotherapeutic strategy, especially in patients with tumors refractory to paclitaxel alone.