Radix Dipsaci Research Articles

Asperosaponin VI promotes progesterone receptor expression in decidual cells via the notch signaling pathway

Recurrent spontaneous abortion (RSA) is a common clinical condition, but its reasons remain unknown in 37–79% of the affected women. The steroid hormone progesterone (P4) is an integral mediator of early pregnancy events, exerting its effects via the progesterone receptor (PR). Dipsaci Radix (DR) has long been used for treating gynecological diseases in Chinese medicine, while its molecular mechanisms and active ingredients are still unclear. We report here the progesterone-like effects of the alcohol extraction and Asperosaponin VI from DR in primary decidual cells and HeLa cell line. We first determined the safe concentration of Asperosaponin VI in the cells with MTT assay and then found by using dual luciferase reporter and Western blotting that Asperosaponin VI significantly increased PR expression. Moreover, we explored the mechanisms of action of the DR extracts and Asperosaponin VI, and the results showed that they could activate Notch signaling, suggesting that they may function by promoting decidualization.

Antiosteoporotic activity of Du–Zhong–Wan water extract in ovariectomized rats

Context Eucommiae Cortex and Radix Dipsaci, occurring in a ratio of 1:1 in Du-Zhong-Wan (DZW), a Chinese herbal medicine, is available as a water extract followed by ethanol precipitation for the treatment of osteoporosis, fractures and menopausal syndrome.Objective This study investigates the protective effects of DZW in ovariectomy (OVX)-induced bone loss in a rat osteopenia model.Materials and methods Sixty Sprague–Dawley rats were randomly divided into the sham-operated group (SHAM) and five OVX subgroups: OVX with vehicle (OVX), 17β-estradiol (E2) and with three graded doses of DZW. Daily oral administration of the different samples started on the fifth week and lasted for 12 weeks, respectively. The body weight, uterus wet weight, serum biochemical parameters, bone mineral density (BMD), bone biomechanical properties, bone microarchitecture and immunohistochemistry were examined.Results Compared with the SHAM group, the DZW treatment significantly reversed the osteoporotic changes in OVX rats. The DZW-H group showed that serum tartrate-resistant acid phosphatase 5b (TRACP-5b) levels reduced by 152.25% (p < 0.01) and osteocalein (OCN) levels dose dependently increased by 118.43% (p < 0.01) as compared with the OVX group. Compared with the OVX group, the DZW at different three dosages of DZW evidently increased the right femur BMD by 112.43, 114.56 and 116.45%, and dramatically promoted bone quality and bone strength (p < 0.05). Further, immunohistochemical evaluation also showed that DZW administration increased ER expression in uteri (p < 0.01).Conclusions DZW exhibits an anti-osteoporotic effect, probably mediated via phyto-estrogenic effects. It might be a potential herbal alternative for the management of postmenopausal osteoporosis.

In vivo screening for anti-osteoporotic fraction from extract of herbal formula Xianlinggubao in ovariectomized mice.

Background and ObjectivesTraditional Chinese Medicine (TCM) Fufang or formula Xianlinggubao (XLGB) is a prescribed TCM drug in China registered for prevention and treatment of osteoporosis. Fufang in TCM is comprised of a group of herbal compounds contributing in group to the treatment efficacy. The present study aims to identify the bioactive fraction(s) in XLGB extract that account(s) dominantly for its osteogenic effects.MethodsThe extract of XLGB formula was separated into three fractions using chromatography, i.e., XLGB-A, XLGB-B and XLGB-C. They were administrated to 4-month old ovariectomized (OVX) mice for 6 weeks to determine which bioactive fraction(s) were more effective for preventing OVX-induced bone loss evaluated by microCT, biomechanical testing and biochemical markers. The main peaks of the key fraction were identified using reference compounds isolated from the fraction. In addition, the effects of the composite compounds in XLGB-B on osteoblasts’ proliferation and mineralization were evaluated in UMR 106 cells.ResultsXLGB-B with a yield of 13.0% from herbal Fufang XLGB was identified as the most potential one among the three fractions for prevention of OVX-induced bone loss confirmed with bone mass, bone microarchitecture, bone strength and bone turnover markers. Nine compounds in HPLC fingerprint were identified in the XLGB-B fraction, including phenylpropanoids from Herba Epimedii, terpenes from Radix Dipsaci and coumarins from Fructus Psoraleae. In addition, the identified compounds effectively promoted proliferation and/or mineralization of osteoblast-like UMR 106 cells in vitro.ConclusionXLGB-B with defined phytochemical structures was screened as the key fraction that demonstrated preventive effects on OVX-induced bone loss in mice. The present study laid down a foundation towards a new generation of herbal Fufang characterized with “less herbal materials for achieving equal treatment efficacy” in development strategy of TCM for prevention of OVX-induced osteoporosis.

Radix Dipsaci total saponins stimulate MC3T3-E1cell differentiation via the bone morphogenetic protein-2/MAPK/Smad-dependent Runx2 pathway.

Radix Dipsaci total saponins (RTS) are primary active components of Radix Dipsaci, which is administered orally for the treatment of osteoporosis according to Chinese Medicine. RTS have also been shown to reduce the risk of bone fractures in rats. However, the detailed molecular mechanisms underlying their action remain elusive. In the present study, the ability of RTS to increase alkaline phosphatase activity, osteocalcin levels and the degree of mineralization was investigated in MC3T3‑E1 mouse osteoblast precursor cells. In addition, the associated molecular mechanism was detected. The results revealed that RTS exerted an effect on osteoblastic maturation and differentiation. Induction of differentiation by RTS was associated with an increase in the expression levels of bone morphogenetic protein‑2 (BMP‑2), phosphorylated (P)‑Smad1/5/8, P‑ERK1/2, P‑p38 and Runt‑related transcription factor 2 (Runx2). Blocking BMP‑2 expression with noggin significantly reduced the levels of osteoblastic differentiation and subsequently attenuated the expression levels of P‑Smad1/5/8, P‑ERK1/2, P‑p38 and Runx2. This indicated that RTS induced osteoblastic differentiation through BMP‑2/mitogen‑activated protein kinase/Smad1/5/8‑dependent Runx2 signaling pathways and that it may be a promising agent for enhancing bone formation.

Early Outcome of Ilizarov Ring Fixator in Open Displaced Intra-Articular Calcaneal Fractures

The study was to observe the safety and anti-inflammatory effects of a topical herbal agent. The herbal patch contained extracts of the 3 herbs Flos carthami (c´ eŠ± ), Radix dipsaci (coŒae–·), Rhizoma rhei (a¤§e»ƒ) and Borneolum syntheticum (a†°c‰‡). The Pain score Visual Analogue Scale (VAS) and Foot Function index (FFI) questionnaires were conducted to measure pain and disability. Blood was taken to measure the inflammation cytokines. All the assessments were processed on before and after the treatment intervention. After treatment, the pain score and foot function index were remarkably improved. Inflammation Cytokine IL-8 was decreased. The novel herbal patch used in this study has laboratory evidences of anti-inflammation and pro-angiogenesis. With its mildly penetrative property, it could be a suitable additional tool for patients’ self-care.

Does Topical Agent Help Fracture Healing? A Pilot Study Using a Herbal Patch

To observe the safety and effectiveness of a topical herbal agent used to promote fracture healing, a herbal patch containing extracts of three herbs, viz., Flos Carthami, Radix Dipsaci, Rhizoma Rhei and Borneolum Syntheticum (an enhancer) was applied on ten subjects with un-displaced fifth metatarsal fractures. Pain scores, foot and ankle function questionnaires and regional swelling were carefully assessed and recorded. Peripheral blood was taken to measure the inflammation cytokines. Assessment checks were performed biweekly to enforce effective patch application and compliance. The results showed that pain improved after two weeks and fracture sites swelling had 20% reduction in thickness when measured with an ultrasonic tool. Foot and ankle functional scores markedly improved after six weeks. Radiological examinations revealed early perfect fracture unions. The topical herbal patch was effective in promoting fracture healing. It was well tolerated by the fracture patients. Larger randomized controlled trials would be indicated.

Rapid identification of crude and sweated Dipsaci Radix based on near-infrared spectroscopy combined with principal component analysis-Mahalanobis distance

In order to discriminate the crude and sweated Dipsaci Radix correctly and rapidly, the crude and sweated Dipsaci Radix were scanned by the NIR spectrometer, and an identifying model was developed by near infrared spectroscopy combined with principal component-Mahalanobis distance pattern recognition method. The pretreated spectra data of 129 crude samples and 86 sweated ones were analyzed through principal component analysis (PCA). The identifying model was developed by choosing the spectrum for 9 881.46-4 119.20 cm(-1) and "SNV + spectrum + S-G" to the original spectral preprocessing with 14 principal components, and then was verified by prediction set, identifying with 100% accuracy. The rapid identification model of the crude and sweated Dipsaci Radix by NIR is feasible and efficient, and could be used as an assistant means for identifying the crude and sweated Dipsaci Radix.

Treatment of Radix Dipsaci extract prevents long bone loss induced by modeled microgravity in hindlimb unloading rats

Context: Radix Dipsaci is a kidney tonifying herbal medicine with a long history of safe use for treatment of bone fractures and joint diseases in China. Previous studies have shown that Radix Dipsaci extract (RDE) could prevent bone loss in ovariectomized rats.Objective: This study investigates the effect of RDE against bone loss induced by simulated microgravity.Materials and methods: A hindlimb unloading rat model was established to determine the effect of RDE on bone mineral density and bone microarchitecture. Twenty-four male Sprague–Dawley rats were divided into four groups (n = 6 per group): control (CON), hindlimb unloading with vehicle (HLU), hindlimb unloading treated with alendronate (HLU-ALN, 2.0 mg/kg/d), and hindlimb unloading treated with RDE (HLU-RDE, 500 mg/kg/d). RDE or ALN was administrated orally for 4 weeks.Results: Treatment with RDE had a positive effect on mechanical strength, BMD, BMC, bone turnover markers, and the changes in urinary calcium and phosphorus excretion. MicroCT analysis showed that RDE significantly prevented the reduction of the bone volume fraction, connectivity density, trabecular number, thickness, tissue mineral density, and tissue mineral content as well as improved the trabecular separation and structure model index.Discussion and conclusion: RDE was demonstrated to prevent the loss of bone mass induced by HLU treatment, which suggests the potential application of RDE in the treatment of microgravity-induced bone loss.

Chromatographic Fingerprint Combined with Content of Asperosaponin VI and Antioxidant Activity for Quality Evaluation of Wine-fried Dipsaci Radix

Dipsaci Radix, the dry root of Dipsacusasper Wall. ex Henry, is a commonly used traditional Chinese medicine (TCM). A novel comprehensive method was proposed for quality evaluation of wine-fried Dipsaci Radix by an integrated data including three aspects of information: high performance liquid chromatography (HPLC) fingerprint, asperosaponin VI content and antioxidant activity (AA). Various indicators including fingerprint similarity, asperosaponin VI content and AA were respectively employed for quality assessment of processed Dipsaci Radix samples. Results showed that considerable differences existed in quality of processed samples with different processing conditions according to three indicators. Among the factors affecting quality of wine-fried Dipsaci Radix, heating temperature was the most influential factor based on analysis of variance (ANOVA), and should be cautiously controlled. The three evaluation indicators respectively used for optimization of processing technology suggested different optimal conditions of wine-frying. Therefore, a combined indicator based on three evaluation indicators was used for determination of optimal processing condition. Multivariate statistical methods such as Hierarchical Clustering Analysis (HCA) and Principal Components Analysis (PCA) were both employed to classify the processed samples for quality evaluation. To more comprehensively evaluate the quality of wine-fried Dipsaci Radix, HPLC fingerprint combined with content of asperosaponin VI and AA may be a reasonable and practical approach.

High performance liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight–mass spectrometry as a powerful analytical strategy for systematic analysis and improved characterization of the major bioactive constituents from Radix Dipsaci

Radix Dipsaci (RD), the dried root of Dipsacus asper, is commonly used as a traditional Chinese medicine for the treatment of bone diseases and functions in strengthening bone and healing bone fractures. Nevertheless, the high polarity, non chromophores and low abundance of multiple compounds in this plant bring difficulty for their isolation and structural determination by traditional chromatographic and spectroscopic techniques, which hindered the use of RD in clinical practice and retarded the process of RD modernization. In this work, a sensitive and rapid high-performance liquid chromatography coupled with electrospray time-of-flight–mass spectrometry (HPLC-ESI-QTOF–MS/MS) was employed to rapidly separate and identify the multiple minor constituents in RD. Separation was performed an Agilent poroshell 120 EC-C18 column (2.1mm×100mm, i.d., 2.7μm) with 0.1% formic acid aqueous solution and acetonitrile as the mobile phase under gradient conditions. As a result, 36 major constituents including dipsacus saponins, iridoid glycosides and caffeoyl quinic acid derivatives were identified or tentatively characterized from the RD, 11 of which had not been previously reported to the best of our knowledge. In conclusion, the HPLC-ESI-QTOF–MS/MS is feasible and credible technique to separate and identify the constituents in complex matrices of traditional Chinese medicines.

Metabolites profile of Xian-Ling-Gu-Bao capsule, a traditional Chinese medicine prescription, in rats by ultra performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry analysis

Xian-Ling-Gu-Bao capsule (XLGB), a well-known traditional Chinese medicine prescription (TCMP), is widely used for the treatment of osteoporosis. However, due to lack of metabolism research, the effective material of XLGB is still unknown. It entails a huge obstacle for the clinical-safe medication administration and quality control of XLGB. To explore the metabolic fate of multiple components of XLGB, herein, we proposed a “representative structure based homologous xenobiotics identification” (RSBHXI) strategy based on ultra performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UPLC/Q-TOF-MS) and mass defect filter (MDF) technique. A total of 147 XLGB-related xenobiotics were identified or tentatively characterized in rat biofluids after oral administration of XLGB, including 134 (57 prototype components and 77 metabolites) in plasma, 93 (37 prototype components and 56 metabolites) in urine and 118 (46 prototype components and 72 metabolites) in bile. Our results indicated that prenylated flavonol glycosides from Herba epimedii, prenylated flavonoids from Fructus psoraleae, saponins from Radix dipsaci and Rhizoma anemarrahenae, as well as tanshinones from Radix Salviae Miltiorrhizae were major absorbed chemical components of XLGB. Hydrolysis, glucuronidation and sulfation were major metabolic reactions of XLGB. As more xenobiotics were detected in bile than those in urine, it demonstrated that multiple components of XLGB underwent comprehensive hepatobiliary excretion. The present study expands our knowledge about the metabolism of XLGB which will be conducive to revealing its in vivo pharmacological material basis. In addition, the application of RSBHXI strategy provides a new approach for metabolite identification of TCMPs and other complex mixture.

Quantitative determination and pattern recognition analyses of bioactive marker compounds from Dipsaci Radix by HPLC

In this study, quantitative and pattern recognition analyses were developed using HPLC/UV for the quality evaluation of Dipsaci Radix. For quantitative analysis, five major bioactive compounds were assessed. The separation conditions employed for HPLC/UV were optimized using ODS C18 column (250 × 4.6 mm, 5 μm) with a gradient of acetonitrile and water as the mobile phase at a flow rate of 1.0 mL/min and a detection wavelength of 212 nm. These methods were fully validated with respect to linearity, accuracy, precision, recovery, and robustness. The HPLC/UV method was applied successfully to the quantification of five major compounds in the extract of Dipsaci Radix. The HPLC analytical method for pattern recognition analysis was validated by repeated analysis of 17 Dipsaci Radix and four Phlomidis Radix samples. The results indicate that the established HPLC/UV method is suitable for quantitative analysis.

Industrial-Scale Preparation of Akebia Saponin D by a Two-Step Macroporous Resin Column Separation

A simple and efficient procedure for the industrial preparation of akebia saponin D, one of the bioactive compounds commonly found in the well-known Chinese Medicinal herb DipsaciRadix, was developed. First, HPD-722 was selected from among 10 kinds of macroporous absorption resins. Following this step, the purity of akebia saponin D was increased about 10 times from 6.27% to 59.41%. In order to achieve a higher purity, ADS-7 was chosen from among five kinds of macroporous absorption resins, and the purity of akebia saponin D was increased from 59.41% to 95.05%. The result indicated HPD-722 and ADS-7 were the most suitable resins to purify akebia saponin D from DipsaciRadix. Under these conditions, large-scale preparation of akebia saponin D was carried out successfully. The preparation method is simple, efficient, and has been demonstrated to be effective for large scale preparations of akebia saponin D from DipsaciRadix.

Therapeutic effects of radix dipsaci, pyrola herb, and cynomorium songaricum on bone metabolism of ovariectomized rats

BackgroundThe objective of this study was to evaluate the effects of herbal medicines, such as Radix Dipsaci (RDD), Pyrola Herb (PHD), and Cynomorium songaricum decoction (CSD), on osteoporotic rats induced by ovariectomy (OVX).MethodsOVX or sham operations were performed on 69 virgin Wistar rats that were divided into six groups: sham (sham, n = 12), OVX control group (OVX, n = 12), and OVX rats with treatments (diethylstilbestrol, E2, n = 12; RDD, n = 11, PHD, n = 11, and CSD, n = 11). Non-surgical rats served as normal control (NC, n = 12). The treatments began four weeks after surgery and lasted for 12 weeks. Bone mass and bone turnover were analyzed by histomorphometry. Levels of protein expression and mRNA of OPG and RANKL in osteoblasts (OB) and bone marrow stromal cells (bMSC) were evaluated by immunohistochemistry and in situ hybridization.ResultsCompared to NC and sham rats, trabecular bone formation was significantly reduced in OVX rats, but restored in E2-treated rats. Treatment with either RDD or PHD enhanced trabecular bone formation remarkably. No significant change of bone formation was observed in CSD-treated rats. OPG expression of protein and mRNA was reduced significantly in OB and bMSC of OVX control rats. RANKL expression of protein and mRNA was increased significantly in OB and bMSC of OVX control rats. These effects were substantially reversed (increased in OPG and decreased in RANKL) by treatment with E2, RDD, or PHD in OB and bMSC of OVX rats. No significant changes in either OPG or RANKL expression were observed in OB and bMSC of OVX rats treated with CSD.ConclusionsOur study showed that RDD and PHD increased bone formation by stimulating overexpression of OPG and downregulation of RANKL in OB and bMSC. This suggests that RDD and PHD may be used as alternative therapeutic agents for postmenopausal osteoporosis.

The beneficial effect of Radix Dipsaci total saponins on bone metabolism in vitro and in vivo and the possible mechanisms of action

The purpose of this study is to investigate the anti-osteoporotic effects of Radix Dipsaci total saponins (RTS). We showed that RTS was able to improve bone properties by either an increase of osteoblastic activity or a decrease in osteoclastic activity. Radix Dipsaci has long been used as an anti-osteoporotic drug. The present study investigates the anti-osteoporotic effects of RTS. Three-month-old female rats were randomly assigned into a sham-operated group (sham) and five ovariectomy (OVX) subgroups, namely, OVX with vehicle (OVX), OVX with 17β-ethinylestradiol (E(2)), and OVX with graded doses of RTS (50, 100, or 200 mg/kg/d). RTS and E(2) were administered orally, daily from 1 week after OVX treatment for 4 months. Bone mass, turnover, and strength were evaluated by dual-energy X-ray absorptiometry, biochemical markers, and the three-point bending test. The trabecular bone microarchitecture was assessed by microCT. In vitro experiments were performed to determine the potential molecular mechanisms of the anti-osteoporotic effect of RTS. RTS prevented the loss of bone mass induced by OVX. The preventive effect on bone loss was primarily indicated by decreasing levels of bone turnover markers and confirmed by the changes in urinary calcium and phosphorus excretion. The treatment also enhanced the biomechanical strength of bone and prevented the deterioration of trabecular bone microarchitecture. RTS induced MC3T3-E1 and primary osteoblastic cell maturation and differentiation and increased bone formation by increasing BMP-2 synthesis. In addition, RTS inhibited osteoclastogenesis through an increase in osteoprotegrin and a decrease in NF-kB ligand expression in vitro. RTS treatment can effectively suppress the loss of bone mass induced by OVX and in vitro evidence suggests this could be through actions on both osteoblasts and osteoclasts.

Inhibitory effects of the root extract of Dipsacus asperoides C.Y. Cheng et al T.M.Ai on collagen-induced arthritis in mice

Ethnopharmacological relevanceDipsaci radix, the dried root of Dipsacus asperoides C.Y. Cheng et al T.M.Ai is used as a medicinal plant in oriental clinics for the treatment of bone diseases and functions by strengthening bone and healing bone fractures. Aim of the studyThis study investigated the therapeutic efficacy of Dipsaci radix in treating rheumatoid arthritis using a type II collagen (CII)-induced arthritis (CIA) mouse model. Materials and methodsArthritis was induced in male DBA/1 mice by immunization with CII. Dipsaci radix water (DR-W) extract at 50mg/kg and 100mg/kg was orally administered from days to after the induction of arthritis. Arthritic score, serum levels of anti-CII IgG2a, the inflammatory mediator prostaglandin E2 (PGE2), and inflammatory cytokines (TNF-α, IL-1β and IL-6), and histological changes in the ankle joint were analyzed in CIA mice. ResultsArthritic induction increased the arthritic score, as well as serum levels of anti-CII IgG2a antibody, PGE2, TNF-α, IL-1β and IL-6 in mice. However, administration of DR-W extract in CIA mice significantly reduced arthritic scores and serum levels of anti-CII IgG2a antibody, PGE2, TNF-α, IL-1β and IL-6 compared with those in vehicle-treated CIA mice. Furthermore, histopathological improvement in joint architecture was also observed in DR-W extract-treated CIA mice. ConclusionsDR-W extract has anti-inflammatory and anti-arthritic effects in arthritic mice. This suggests that Dipsaci radix might be used as a therapeutic agent for the treatment of human arthritis.

Determination of saponins in Dipsaci Radix by HILIC-HPLC

To develop an HILIC-HPLC method for simultaneous analysis of three main active triterpenoid saponins including dipsacus asponin VI, dipsacus asponin X and dipsacus asponin XII. To evaluate the medical material from various habitats in China, different commercial grades or parts of plant. The HPLC was conducted on Venusil HILIC column (4.6 mm x 250 mm, 5 microm) with acetonitrile and water as the mobile phase at 25 degrees C, which was in gradient elution. The flow rate was 1 mL x min(-1); and the detection wavelength was 203 nm. The contents of dipsacus asponin VI, X and XII were 0.77%-14.31%, 0.39%-3.19% and 0.41%-1.49% respectively in different provinces of China, that were related to locations. The contents of saponins from Yunnan and Sichuan provinces were higer than those of Hubei and Guizhou. Thick roots, which were high-class products, contained less saponins than thin ones. In addition, the contents of stem, residual parts of stem and fibrous roots were fewer than main roots. This method is simple, sensitive and accurate. It could be used to determine the contents of dipsacus asponin VI, X, XII and evaluate the quality of dipsacus asperoides.

Effects of the Dichloromethane Fraction of Dipsaci Radix on the Osteoblastic Differentiation of Human Alveolar Bone Marrow-Derived Mesenchymal Stem Cells

Dipsaci Radix is the dried root of Dipsacus asper Wall. It has been used in Korean herbal medicine to treat bone fractures. In this study, we examined the effect of the dichloromethane fraction of Dipsaci Radix (DR(DM)) on the osteoblastic differentiation of human alveolar bone marrow-derived MSCs (ABM-MSCs). The ABM-MSCs were isolated from healthy subjects and cultured in vitro, followed by phenotypic characterization. They showed a fibroblast-like morphology and expressed CD29, CD44, CD73, and CD105, but not CD34. Calcified nodules were generated in response to both dexamethasone (DEX) and DR(DM). There was a significant increase in the alkaline phosphatase (ALP) activity and protein expression of bone sialoprotein (BSP) and osteocalcin (OC) in response to DEX and DR(DM) as compared to control. These results provide evidence for the osteogenic potential of cultured ABM-MSCs in response to DR(DM). Also, an active single compound was additionally isolated from DR(DM). The single compound (hederagenin 3-O-(2-O-acetyl)-α-L-arabinopyranoside) also significantly increased ALP activity and the level of protein expression of BSP and OC. These results highlight the possible clinical applications of DR(DM) and hederagenin 3-O-(2-O-acetyl)-α-L-arabinopyranoside in bone regeneration.

Radix Dipsaci does not improve tendon healing in a rat model of patellar tendon donor site injury

To explore whether Radix Dipsaci (RD) exhibits beneficial effects on tendon healing. An attempt was made to explore the in vitro effects of a hot water extract of RD on gene expression of procollagen Type I (COL1A1), procollagen Type III (COL3A1) and decorin in cultured tendon fibroblasts, and its in vivo effects in a well-established rat model of patellar tendon donor site injury. It was found that gene expression of COL3A1 and decorin in cultured tendon fibroblasts was significantly increased by RD, but that COL1A1 was not affected. In vivo studies showed that RD increased blood vessels in the wound but did not significantly affect the expression of COL1A1, COL3A1 and decorin at day 14 post-injury. The ultimate tensile stress of the healing tendon was not significantly improved by either local injection or oral administration of hot water extract of RD (P > 0.05). The present findings imply that RD per se does not significantly improve tendon healing. Further investigation of RD in a herbal formula may be necessary to test its efficacy in tendon injuries.

In vitro & in vivo assessment of a herbal formula used topically for bone fracture treatment

Aim of the study A novel topical paste used for fracture healing (FH), consisting of the extracts of six herbs, Radix Dipsaci, Ramulus Sambucus Williamsii, Rhizoma Notoginseng, Flos Carthami, Rhizoma Rhei and Fructus Gardeniae, was developed according to the classical theory of traditional Chinese medicine. This study aimed to determine the effectiveness of this formula, and some of its important chemical components in the promotion of fracture healing. The transdermal transport of FH was also examined. Materials and methods The osteogenic, angiogenic and nitric oxide suppressing effects of FH and its important chemical marker components were assessed by using osteoblastosacroma UMR-106 cells, human umbilical vein endothelial cells (HUVEC) and murine macrophage RAW264.7 cells, respectively. The bone healing effects of the FH paste and its transdermal absorption were determined using a rabbit fracture model. The callus sizes, bone specific alkaline phosphatase levels and biomechanical properties of the healed bone were assessed. Results FH significantly increased the cell proliferation in UMR-106 and HUVEC cells and inhibited the nitric oxide production in murine macrophage in dose-dependent manner. Its important chemical components asperosaponin VI, ginsenoside Rg1 and emodin were shown to be acting positively in the respective in vitro studies. FH paste significantly improved the bone healing in the rabbit fracture model, as was indicated by the increases in callus size at weeks 2–5, and the elevations in bone specific alkaline phosphatase activities at weeks 5–6. The analysis using LC/MS/MS also showed the presence of important chemical marker components of the FH formula in the plasma after 8 weeks of topical treatment. Conclusion This study presents the first scientific evidence of the efficacy of a herbal paste in the promotion of fracture healing. There were evidences of transdermal transport of the chemical components, control the inflammation through nitric oxide inhibition, promotion of angiogenesis, and bone healing in the in vitro tests, as well as in the experimental animal.