Effects of Icariin on Modulating Gut Microbiota and Regulating Metabolite Alterations to Prevent Bone Loss in Ovariectomized Rat Model.

Postmenopausal osteoporosis (PMOP) is an estrogen deficiency-induced bone loss, which has been shown an association with an altered gut microbiota (GM). Gut microbiota-bone axis has been recognized as a crucial mediator for bone homeostasis. Icariin (ICA) is an effective agent to delay bone loss by regulating the bone homeostasis. Thus, we hypothesize that ICA can prevent bone loss by modulating GM and regulating metabolite alterations. The effects of ICA on bone metabolism improvement in ovariectomized (OVX) rats and their relationships with the GM and fecal metabolites were investigated. Micro-computed tomography (micro-CT) and hematoxylin-eosin (HE) staining showed a typical bone boss in OVX group, while ICA or estradiol (E2) administration exhibited positive effects on bone micro-architecture improvement. The GM such as Actinobacteria, Gammaproteobacteria, Erysipelotrichi, Erysipelotrichales, Enterobacteriales, Actinomycetales, Ruminococcus and Oscillospira significantly correlated to serum bone Gla-protein (BGP), receptor activator of nuclear factor-κB (RANK), receptor activator of nuclear factor-κB ligand (RANKL), osteoprotegerin (OPG) and tartrate resistant acid phosphatase (TRACP). Further t-test revealed a substantial variation of the GM and fecal metabolites in different treatments. Among them, Lachnoclostridium, Butyricimonas, Rikenella, Paraprevolla, Adlercreutzia, Enterorhabdus, Anaerovorax, Allobaculum, Elusimicrobium, Lactococcus, Globicatella and Lactobacillus were probably the key microbial communities driving the change of bile acid, amino acid and fatty acid, thereby leading to an improvement of PMOP. The significant up-regulation of L-Saccharopine, 1-Aminocyclohexadieneacid and linoleic acid after ICA administration suggested important contributions of amino acid and fatty acid metabolisms in the prevention and treatment of PMOP. Taken together, our study has provided new perspectives to better understand the effects of ICA on PMOP improvement by regulating GM and the associated fecal metabolites. Our findings contribute to develop ICA as a potential therapy for PMOP.

Verification of pain-related neuromodulation mechanisms of icariin in knee osteoarthritis.

Knee osteoarthritis (KOA) is a common disease with no specific treatment. Icariin (ICA) is considered an agent for KOA. This study aimed to confirm the pain-related neuromodulation mechanisms of ICA on KOA. Three experiments were designed: (1) verifying the therapeutic effects of ICA in vivo and in vitro, (2) exploring the potential pain-related neuromodulation pathways involved in ICA treatment by functional magnetic resonance imaging (fMRI) and virus retrograde tracing (VRT) and (3) confirming the pain-related targets by tandem mass tag (TMT)-based quantitative proteomics and bioinformatic analyses. Experiment 1 verified the efficacy of ICA in OA animal and cell models. Experiment 2 found a series of brain regions associated with KOA reversed by ICA treatment, indicating that a pain-related hypothalamic-mediated neuromodulation pathway and an endocannabinoid (EC)-related pathway contribute to ICA mechanisms. Experiment 3 explored and confirmed four pain-related genes involved in KOA and ICA treatment. We confirmed the key role of pain-related neuromodulation mechanisms in ICA treatment associated with its analgesic effect. Our findings contribute to considering ICA as a novel therapy for KOA.

Identification of the Key Role of NF-κB Signaling Pathway in the Treatment of Osteoarthritis With Bushen Zhuangjin Decoction, a Verification Based on Network Pharmacology Approach.

This study aimed to identify whether the NF-κB signaling pathway plays a key role in the treatment of osteoarthritis (OA) with Bushen Zhuangjin Decoction (BZD) based on a typical network pharmacology approach (NPA). Four sequential experiments were performed: 1) conventional high-performance liquid chromatography (HPLC), 2) preliminary observation of the therapeutic effects of BZD, 3) NPA using three OA-related gene expression profiles, and 4) verification of the key pathway identified by NPA. Only one HPLC-verified compound (paeoniflorin) was identified from the candidate compounds discovered by NPA. The genes verified in the preliminary observation were also identified by NPA. NPA identified a key role for the NF-κB signaling pathway in the treatment of OA with BZD, which was confirmed by conventional western blot analysis. This study identified and verified NF-κB signaling pathway as the most important inflammatory signaling pathway involved in the mechanisms of BZD for treating OA by comparing the NPA results with conventional methods. Our findings also indicate that NPA is a powerful tool for exploring the molecular targets of complex herbal formulations, such as BZD.

Confirmation of inhibitingTLR4/MyD88/NF-κB Signalling Pathway by Duhuo Jisheng Decoction on Osteoarthritis: A Network Pharmacology Approach-Integrated Experimental Study.

This study was conducted to identify whether the TLR4/MyD88/NF-κB signalling pathway plays a vital role in osteoarthritis (OA) treatment with Duhuo Jisheng Decoction (DHJSD) on the basis of a network pharmacology approach (NPA)-integrated experiment. Two experiments were conducted as follow: NPA for DHJSD using six OA-related gene series and the key pathway was screened out using NPA. NPA identified a vital role for the TLR4/MyD88/NF-κB signalling pathway in OA treatment with DHJSD, the conventional western blot analysis and qPCR confirmed it. Furthermore, changes of miR-146a-5p and miR-34a-5p in the cellular models were recovered by DHJSD administration, which synergistically contributed to OA therapy. The toll-like receptor signalling pathway and the NF-κB signalling pathway were meaningfully enriched by the miRNA-regulated gene pathways. This study identified and confirmed the TLR4/MyD88/NF-κB signalling pathway is an essential inflammatory signalling pathway in the DHJSD underlying OA treatment. The results provide a basis for further evaluation of the regulatory mechanism of the drug's efficacy in treating OA.

A simple enzyme-catalyzed reaction induced "switch" type fluorescence biosensor based on carbon nitride nanosheets for the assay of alkaline phosphatase activity.

An enzyme-catalyzed fluorescence "switch" type sensor was constructed for the determination of alkaline phosphatase (ALP) activity by combining the fluorescence quenching effect of Ag+ on ultrathin g-C3N4 nanosheets (CNNSs) with the simple redox reaction of AA and Ag+. Briefly, Ag+ exhibits a significant quenching effect on the fluorescence of CNNSs. Thus the fluorescence signal of the CNNS-Ag+ system is extremely weak even in the presence of l-ascorbic acid-2-phosphate (AAP) ("off" state). When ALP coexists in the system, the enzyme can specifically catalyze the hydrolysis of AAP to form ascorbic acid (AA), which reduces Ag+ to Ag0. In this case, the fluorescence signal of the system is recovered ("on" state). Based on this principle, a signal-enhanced CNNS fluorescence sensor was developed to determine the activity of alkaline phosphatase. The experimental results show that the detection range of alkaline phosphatase is 0.5-20 U L-1, and the detection limit is 0.05 U L-1 (S/N = 3). Meanwhile, this method was used to assay ALP in serum samples.

Data on Tougu Xiaotong capsules may inhibit p38 MAPK pathway-mediated inflammation in vitro.

The Tougu Xiaotong capsule (TXC) is a traditional herbal compound used to treat osteoarthritis (OA) in China. We performed fingerprint analysis with HPLC for the quality control of TXC. Its composition was identified by the comparison of the spectrogram and chromatographic peak of retention time with a reference substance. TXC was found to contain paeoniflorin, isofraxidin, ferulic acid, and rosmarinic acid. The chondrocytes were identified by immunohistochemical staining using collagen II. Chondrocytes that were positive for collagen II were stained brown in the cytoplasm. The toll-like receptor 4 (TLR4) was expressed on the chondrocyte membrane, which was observed using immunofluorescence microscopy. The nuclei were stained blue by 4',6-diamidino-2-phenylindole (DAPI) and TLR4 was stained green. These were observed using laser scanning confocal microscopy. The successful establishment of LPS-exposed chondrocytes was confirmed using enzyme-linked immunosorbent assay (ELISA). Lipopolysaccharide (LPS) administration significantly reduced the levels of interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α), and a maximum effect was observed at 8 h. We believe that these methods will be useful in future investigations of OA. This data article is related to the research article "Tougu Xiaotong capsules may inhibit p38 MAPK pathway-mediated inflammation: In vivo and in vitro verification" [1].

Tougu Xiaotong capsules may inhibit p38 MAPK pathway-mediated inflammation: In vivo and in vitro verification.

ETHNOPHARMACOLOGICAL RELEVANCE: Tougu Xiaotong capsules (TXC) are an herbal compound commonly used to treat osteoarthritis (OA) in China. AIM OF THE STUDY: We attempted to verify TXC's therapeutic effects and mechanisms related to the p38 mitogen-activated protein kinase (MAPK) pathway in vivo and in vitro. MATERIALS AND METHODS: TXC's therapeutic effects were assessed by observing cartilage degeneration and inflammatory factors in a modified Hulth's model (in vivo) and a lipopolysaccharides (LPS)-exposed cellular model (in vitro). The expression of biomarkers related to p38 MAPK pathway-mediated inflammation was also investigated. RESULTS: TXC treatment reversed cartilage degeneration related biomarkers (ADAMTS 4, ADAMTS 5, Col I, Col V, MMP 3, MMP 9, and MMP 13) and inflammation factors (IL-1ß, TNF-α, and IL-6) in both the animal and cellular OA models. Expression of p-p38 MAPK was downregulated following TXC administration, and changes to microRNAs in the cellular models were recovered. These results indicated that the p38 MAPK pathway-related mechanism may involve therapeutic effects of TXC. CONCLUSIONS: This study verified TXC's efficacy to treat OA in vivo and in vitro and suggests that p38 MAPK pathway-related mechanisms may be involved in TXC's therapeutic effects.

Signal-on fluorescence assay for pyrophosphate ions based on DNA-stabilized silver nanoclusters.

Pyrophosphate anion (P2 O7 4- , PPi) is considered as a potential biomarker for arthritic diseases because high levels of PPi may result in calcium pyrophosphate dehydrate crystal deposition diseases. In this study, a simple fluorescence method for PPi was demonstrated by organic integration of the efficient fluorescence quenching ability of copper ions to DNA-scaffolded silver nanoclusters and the strong affinity of PPi towards copper ions. This simple fluorescence sensor showed a low detection limit (0.28 µM based on signal/noise = 3) towards the detection of PPi. Practical application of this method was also validated by detection of PPi in the synovial fluid.

An in vitro validation of the therapeutic effects of Tougu Xiaotong capsule on tunicamycin-treated chondrocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Tougu Xiaotong capsule (TXC) is a Chinese herbal compound that belongs to a range of Chinese herbs functioning as 'kidney invigorators and liver softeners' commonly used to treat osteoarthritis (OA) in China. AIMS OF THE STUDY: The aims of the present study are to confirm the therapeutic effects of TXC in an OA cell model and to determine the mechanisms involved in such effects. MATERIALS AND METHODS: A tunicamycin (Tm)-exposed OA cell model was employed, and the effects of TXC were confirmed by observing cell viability and apoptosis. The reduced cell viability and increased apoptosis caused by Tm were improved by TXC, confirming the cellular protection of TXC. We then investigated the expression of biomarkers related to the endoplasmic reticulum (ER) stress pathway, including microRNA-211 (miR-211), a regulator in the ER stress pathway. RESULTS: Downregulation of X-box binding protein 1 (Xbp-1) and miR-211 expression following Tm administration was reversed by TXC. Moreover, the upregulation by Tm of the expression levels of binding immunoglobulin protein, Xbp-1, activating transcription factor 4, C/EBP-homologous protein, Caspase-9 and Caspase-3 was downregulated by TXC. These results indicated that the ER stress pathway-related mechanism may play a potential role in the therapeutic effects of TXC. CONCLUSIONS: The present study provides evidence of the therapeutic effects of TXC at the cell level and describes a cellular model for establishing the mechanisms of the effects of TXC used in the treatment of OA.

Glucosamine promotes chondrocyte proliferation via the Wnt/ß­catenin signaling pathway.

The present study investigated the mechanism underlying the effects of glucosamine (GlcN) on the proliferation of chondrocytes isolated from the knee cartilage of Sprague­Dawley rats. Chondrocytes were treated with various concentrations of GlcN or without GlcN. The effects of GlcN on chondrocyte proliferation were determined using reverse transcription­polymerase chain reaction, western blot analysis and immunohistochemistry. The results indicated that GlcN significantly improved chondrocyte viability, accelerated G1/S transition during progression of the cell cycle and promoted the expression of cell cycle regulatory proteins, including cyclin D1, cyclin­dependent kinase (CDK)4 and CDK6, thus indicating that GlcN may promote chondrocyte proliferation. Furthermore, GlcN upregulated the expression levels of Wnt­4, Frizzled­2 and ß­catenin, and downregulated the expression of glycogen synthase kinase­3. GlcN also promoted ß­catenin translocation; ß­catenin is able to activate numerous downstream target genes, including cyclin D1. To determine the role of the Wnt/ß­catenin signaling pathway in chondrocyte proliferation, the Wnt/ß­catenin signaling pathway was inhibited using Dickkopf­1 (DKK­1), after which chondrocytes were treated with GlcN. The results demonstrated that the expression levels of ß­catenin and cyclin D1 were decreased in chondrocytes treated with DKK­1 and GlcN. These results suggested that GlcN may promote chondrocyte proliferation via the Wnt/ß­catenin signaling pathway.

Psoralen promotes the expression of cyclin D1 in chondrocytes via the Wnt/ß-catenin signaling pathway.

Psoralen (PSO), the active ingredient of Fructus Psoraleae (FP) the dried ripe fruit of Psoralea corylifolia L., has been commonly used in traditional Chinese medicine (TCM) for the treatment of osteoarthritis (OA). We found that PSO activates cartilaginous cellular functions of rat chondrocytes in vitro. However, the effect of PSO on chondrocyte proliferation and the precise mechanisms involved remain to be elucidated. We investigated the effects of PSO on chondrocytes isolated from Sprague­Dawley (SD) rats and evaluated involvement of the Wnt/ß-catenin signaling pathway. The viability of chondrocytes treated with PSO was increased in a dose- and time-dependent manner, as assessed by MTT assay. We found that the gene expression and protein levels of Wnt-4, Frizzled-2, ß-catenin and cyclin D1 in the PSO-treated chondrocytes were significantly upregulated, while the gene expression and protein level of glycogen synthase kinase-3ß (GSK-3ß) were downregulated, compared with the untreated chondrocytes. By immunofluorescence, we also found that PSO induced ß-catenin nuclear translocation. Importantly, the expression of ß-catenin and cyclin D1 was partly inhibited by Dickkopf-1 (DKK-1), an inhibitor of the Wnt/ß-catenin signaling pathway. Additionally, Col-II expression in chondrocytes was increased after treatment with PSO. Taken together, these results indicate that PSO promotes chondrocyte proliferation by activating the Wnt/ß-catenin signaling pathway, and it may play an important role in the treatment of OA.

Highly sensitive antibody-aptamer sensor for vascular endothelial growth factor based on hybridization chain reaction and pH meter/indicator.

Vascular endothelial growth factor (VEGF) is a crucial signaling protein for the tumor growth and metastasis, which is also acted as the biomarkers for various diseases. In this research, we fabricate an aptamer-antibody sensor for point-of-care test of VEGF. Firstly, target VEGF is captured by antibody immobilized on the microplate, and then binds with aptamer to form the sandwich structure. Next, with the assist of glucose oxidase (GOx)-functionalized ssDNAs, hybridization chain reaction occurs using the aptamer as the primer. Thus, GOx are greatly gathered on the microplate, which catalyzes the oxidization of glucose, leading to the pH change. As a result, the detect limit at a signal-to-noise was estimated to be 0.5pg/mL of target by pH meter, and 1.6pg/mL of VEGF was able to be distinguished by naked eyes. Meanwhile, this method has been used assay VEGF in the serum with the satisfactory results.

Cibotium barometz polysaccharides stimulate chondrocyte proliferation in vitro by promoting G1/S cell cycle transition.

Cibotium barometz polysaccharides (CBPS) are one of the most important bioactive components extracted from the Cibotium barometz plant, which belongs to the Dicksoniaceae family. It has been widely used for the treatment of orthopedic diseases in traditional Chinese medicine. However, the molecular mechanisms behind the therapeutic effects of CBPS remain to be clarified. In the present study, the concentration of CBPS was detected by phenol-vitriol colorimetry. Furthermore, the effects stimulated by CBPS on the viability and G1/S cell cycle transition in primary chondrocytes from Sprague-Dawley rats were investigated. A cell viability assay demonstrated that chondrocyte proliferation may be enhanced by CBPS in a dose­ and time­dependent manner. The mechanism underlying the promotion of chondrocyte cell cycle was suggested to involve the stimulation of G1 to S phase transition. To further confirm the results, reverse transcription­quantitative polymerase chain reaction and western blot analyses were used to detect the expression of mRNA and protein levels of cyclin D1, cyclin­dependent kinase 4 and retinoblastoma protein. The results suggested that CBPS may stimulate chondrocyte proliferation via promoting G1/S cell cycle transition. Since osteoarthritis is characterized by deficient proliferation in chondrocytes, the present study indicates that CBPS may potentially serve as a novel method for the treatment of osteoarthritis.

Understanding the diverse functions of Huatan Tongluo Fang on rheumatoid arthritis from a pharmacological perspective.

Huatan Tongluo Fang (HTTLF) is a traditional herbal formula that can resolve phlegm and dredge collaterals. HTTLF has also been used to treat rheumatoid arthritis (RA); however, the mechanism underlying the therapeutic effects of HTTLF on RA has not been clearly elucidated at the molecular level. In the present study, an integrated model of system pharmacology containing chemical space analysis, potential active compound prediction and compound-target-disease network was constructed to investigate the molecular mechanisms of HTTLF. The compounds from HTTLF dispersed well in the chemical space. Most of the compounds from HTTLF had similar chemical spaces to drug/drug-like compounds associated with RA, according to the MDL Drug Data Report. A total of 127 potentially active compounds and 17 targets of RA were identified. Among them, 50 compounds interacted with ≥2 targets, while 77 compounds interacted with only one target. In addition, 17 targets were associated with 82 diseases that belonged to 26 categories. These results indicate that HTTLF has diverse chemical spaces and polypharmacology with regards to the treatment of RA. In addition, HTTLF demonstrated therapeutic potential against diverse diseases other than RA, including osteoarthritis, atherosclerosis and brain cancer. This study provides a novel platform for understanding how HTTLF treats RA; this is beneficial for explaining the diverse functions of HTTLF with regards to RA, and may help develop novel compounds with desirable therapeutic targets to treat RA.

Bushen Zhuangjin decoction inhibits TM-induced chondrocyte apoptosis mediated by endoplasmic reticulum stress.

Chondrocyte apoptosis triggered by endoplasmic reticulum (ER) stress plays a vital role in the pathogenesis of osteoarthritis (OA). Bushen Zhuangjin decoction (BZD) has been widely used in the treatment of OA. However, the cellular and molecular mechanisms responsible for the inhibitory effects of BZD on chondrocyte apoptosis remain to be elucidated. In the present study, we investigated the effects of BZD on ER stress-induced chondrocyte apoptosis using a chondrocyte in vitro model of OA. Chondrocytes obtained from the articular cartilage of the knee joints of Sprague Dawley (SD) rats were detected by immunohistochemical staining for type Ⅱ collagen. The ER stress-mediated apoptosis of tunicamycin (TM)­stimulated chondrocytes was detected using 4-phenylbutyric acid (4­PBA). We found that 4­PBA inhibited TM-induced chondrocyte apoptosis, which confirmed the successful induction of chondrocyte apoptosis. BZD enhanced the viability of the TM-stimulated chondrocytes in a dose- and time-dependent manner, as shown by MTT assay. The apoptotic rate and the loss of mitochondrial membrane potential (ΔΨm) of the TM-stimulated chondrocytes treated with BZD was markedly decreased compared with those of chondrocytes not treated with BZD, as shown by 4',6-diamidino-2-phenylindole (DAPI) staining, Annexin V-FITC binding assay and JC-1 assay. To further elucidate the mechanisms responsible for the inhibitory effects of BZD on TM­induced chondrocyte apoptosis mediated by ER stress, the mRNA and protein expression levels of binding immunoglobulin protein (Bip), X­box binding protein 1 (Xbp1), activating transcription factor 4 (Atf4), C/EBP­homologous protein (Chop), caspase­9, caspase-3, B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were measured by reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis. In the TM-stimulated chondrocytes treated with BZD, the mRNA and protein expression levels of Bip, Atf4, Chop, caspase-9, caspase-3 and Bax were significantly decreased, whereas the mRNA and protein expression levels of Xbp1 and Bcl-2 were significantly increased compared with the TM­stimulated chondrocytes not treated with BZD. Additionally, all our findings demonstrated that there was no significant difference between the TM­stimulated chondrocytes treated with BZD and those treated with 4­PBA. Taken together, our results indicate that BZD inhibits TM­induced chondrocyte apoptosis mediated by ER stress. Thus, BZD may be a potential therapeutic agent for use in the treatment of OA.

Duhuo Jisheng decoction inhibits endoplasmic reticulum stress in chondrocytes induced by tunicamycin through the downregulation of miR-34a.

Our previous study showed that Duhuo Jisheng decoction (DHJSD) inhibited chondrocyte apoptosis by the mitochondria-dependent signaling pathway. Endoplasmic reticulum (ER) stress is upstream of the mitochondria-dependent signaling pathway and has been shown to promote chondrocyte apoptosis that occurs in osteoarthritis (OA). The present study aimed to evaluate whether DHJSD inhibits the chondrocyte apoptosis by regulating ER stress. DHJSD enhanced the viability of tunicamycin (TM)­exposed chondrocytes, a model of ER stress-induced apoptosis, in a dose­ and time­dependent manner, as shown by MTT assay. The present results showed that DHJSD and sodium 4-phenylbutyrate (PBA), an ER stress inhibitor, reduced TM­induced chondrocyte apoptosis by 4',6-diamidino­2-phenylindole staining. To gain insight into the mechanisms of DHJSD that are responsible for enhancing the viability and inhibiting TM­induced chondrocyte apoptosis, the associated mRNA expressions and protein levels were detected by reverse transcription­polymerase chain reaction (RT­PCR) and western blot analysis, respectively. The results showed that the expression levels of Xbp1, Xbp1s and Bcl­2 were increased, and the expression levels of Bip, Atf4, Chop, Bax, caspase­9 and ­3 were decreased in the TM­exposed chondrocytes treated with DHJSD or PBA compared with that in the TM­exposed chondrocytes. To identify the possible mechanisms, the expression of miR­34a was examined by the TaqMan microRNA assay, and was downregulated in the TM­exposed chondrocytes treated with DHJSD or PBA compared with that in the TM-exposed chondrocytes. DHJSD inhibits ER stress in chondrocytes induced by exposure to TM by downregulating miR­34a, suggesting that DHJSD may be a potential therapeutic agent for OA.

A bio-inspired sensor coupled with a bio-bar code and hybridization chain reaction for Hg(2+) assay.

In this article, a bio-inspired DNA sensor is developed, which is coupled with a bio-bar code and hybridization chain reaction. This bio-inspired sensor has a high sensitivity toward Hg(2+), and has been used to assay Hg(2+) in the extraction of Bauhinia championi with good satisfaction.

Overexpression of Indian hedgehog partially rescues short stature homeobox 2-overexpression-associated congenital dysplasia of the temporomandibular joint in mice.

The role of short stature homeobox 2 (shox2) in the development and homeostasis of the temporomandibular joint (TMJ) has been well documented. Shox2 is known to be expressed in the progenitor cells and perichondrium of the developing condyle. A previous study by our group reported that overexpression of shox2 leads to congenital dysplasia of the TMJ via downregulation of the Indian hedgehog (Ihh) signaling pathway, which is essential for embryonic disc primordium formation and mandibular condylar growth. To determine whether overexpression of Ihh may rescue the overexpression of shox2 leading to congenital dysplasia of the TMJ, a mouse model in which Ihh and shox2 were overexpressed (Wnt1-Cre; pMes-stop shox2; pMes-stop Ihh mice) was utilized to assess the consequences of this overexpression on TMJ development during post-natal life. The results showed that the developmental process and expression levels of runt-related transcription factor 2 and sex determining region Y-box 9 in the TMJ of the Wnt1-Cre; pMes-stop shox2; pMes-stop Ihh mice were similar to those in wild­type mice. Overexpression of Ihh rescued shox2 overexpression-associated reduction of extracellular matrix components. However, overexpression of Ihh did not inhibit the shox2 overexpression-associated increase of matrix metalloproteinases (MMPs) MMP9, MMP13 and apoptosis in the TMJ. These combinatory cellular and molecular defects appeared to account for the observed congenital dysplasia of TMJ, suggesting that overexpression of Ihh partially rescued shox2 overexpression­associated congenital dysplasia of the TMJ in mice.

Jiedu Xiaozheng Yin decoction inhibits hepatoma cell proliferation by inducing apoptosis via the mitochondrial-mediated pathway.

Jiedu Xiaozheng Yin decoction (JXY) is a type of Chinese traditional medicine, which has been used to treat various types of cancer. The present study explored the mechanisms underlying the anticancer activity of JXY. The effects of ethyl acetate extraction of JXY (EE-JXY) were evaluated on the HepG2 human hepatoma cell line in vitro and in vivo. Following treatment of the HepG2 cells with EE-JXY for 24 h, cell viability, apoptosis, mitochondrial membrane potential, caspase enzyme activity and the expression levels of apoptotic-associated proteins (Bcl-2 and Bax) were detected by MTT, flow cytometry, ELISA and western blotting respectively. In addition, HepG2 cells were subcutaneously transplanted into BALB/c nude mice, and the tumor bearing mice were treated with either EE-JXY (0.06 g/kg) or normal saline for 21 days. Tumor volume and weight were measured and recorded. The apoptotic index, and the expression levels of Bax and cytochrome c were determined with immunohistochemical staining. Treatment with EE-JXY inhibited the proliferation of HepG2 cells, and reduced cell viability in a dose-and time-dependent manner. Furthermore, EE-JXY induced HepG2 cell apoptosis, as demonstrated by a loss of plasma membrane asymmetry and externalization of phosphatidylserine, collapse of mitochondrial membrane potential, activation of caspase-9 and caspase-3, and an increased ratio of pro-apoptotic Bax to anti-apoptotic Bcl-2. Furthermore, EE-JXY inhibited tumor growth and increased the apoptotic index of tumors in tumor-bearing mice. In conclusion, the results of the present study suggest that JXY inhibits HepG2 cell proliferation through mitochondrion-mediated apoptosis, which may partially explain its anticancer activity.

Deciphering the underlying mechanisms of Diesun Miaofang in traumatic injury from a systems pharmacology perspective.

Diesun Miaofang (DSMF) is a traditional herbal formula, which has been reported to activate blood, remove stasis, promote qi circulation and relieve pain. DSMF holds a great promise for the treatment of traumatic injury in an integrative and holistic manner. However, its underlying mechanisms remain to be elucidated. In the present study, a systems pharmacology model, which integrated cluster ligands, human intestinal absorption and aqueous solution prediction, chemical space mapping, molecular docking and network pharmacology techniques were used. The compounds from DSMF were diverse in the clusters and chemical space. The majority of the compounds exhibited drug-like properties. A total of 59 compounds were identified to interact with 16 potential targets. In the herb-compound-target network, the majority of compounds acted on only one target; however, a small number of compounds acted on a large number of targets, up to a maximum of 12. The comparison of key topological properties in compound-target networks associated with the above efficacy intuitively demonstrated that potential active compounds possessed diverse functions. These results successfully explained the polypharmacological mechanism underlying the efficiency of DSMF for the treatment of traumatic injury as well as provided insight into potential novel therapeutic strategies for traumatic injury from herbal medicine.