Mutation of cysteine 46 in IKK-beta increases inflammatory responses.

Activation of IκB kinase ß (IKK-ß) and nuclear factor (NF)-κB signaling contributes to cancer pathogenesis and inflammatory disease; therefore, the IKK-ß-NF-κB signaling pathway is a potential therapeutic target. Current drug design strategies focus on blocking NF-κB signaling by binding to specific cysteine residues on IKK-ß. However, mutations in IKK-ß have been found in patients who may eventually develop drug resistance. For these patients, a new generation of IKK-ß inhibitors are required to provide novel treatment options. We demonstrate in vitro that cysteine-46 (Cys-46) is an essential residue for IKK-ß kinase activity. We then validate the role of Cys-46 in the pathogenesis of inflammation using delayed-type hypersensitivity (DTH) and an IKK-ß C46A transgenic mouse model. We show that a novel IKK-ß inhibitor, dihydromyricetin (DMY), has anti-inflammatory effects on WT DTH mice but not IKK-ß C46A transgenic mice. These findings reveal the role of Cys-46 in the promotion of inflammatory responses, and suggest that Cys-46 is a novel drug-binding site for the inhibition of IKK-ß.

The pharmacokinetic study of sinomenine, paeoniflorin and paeonol in rats after oral administration of a herbal product Qingfu Guanjiesu capsule by HPLC.

An accurate and reliable high-performance liquid chromatography-diode array detector (HPLC-DAD) method was developed and validated for determination of sinomenine (SI), paeoniflorin (PF) and paeonol (PA), which was further applied to assess the pharmacokinetics of SI, PF and PA in an anti-arthritic herbal product, Qingfu Guanjieshu (QFGJS) capsule, in rats. Successful separation was achieved with a C18 column and a mobile phase composed of acetonitrile and aqueous phase (containing 0.1% formic acid, adjusted with triethylamine to pH 3.5 ± 0.2). The method was validated with excellent precision, accuracy, recovery and stability in calibration ranges from 0.06 to 11.62 µg/mL for SI, from 0.09 to 35.70 µg/mL for PF, and from 0.15 to 4.53 µg/mL for PA (with r(2) > 0.999 for all three compounds). Our results showed that absorption of PF after administration of QFGJS was similar to that after oral administration of PF alone; the absorption of SI was decreased while the absorption of PA was increased after giving QFGJS orally compared with pure compounds. We may conclude that pharmacokinetic studies of complex herbal products are not only necessary but also feasible by using representative bioactive chemicals as indicators of establishing quality control standards and of determining pharmacokinetic behavior of herbal medicines.

The Predicted Proteomic Network Associated with the Antiarthritic Action of Qingfu Guanjieshu in Collagen-II-Induced Arthritis in Rats.

Qingfu Guanjieshu (QFGJS) is an herbal preparation for treating rheumatoid arthritis (RA). Previous studies revealed that QFGJS significantly inhibited experimental arthritis and acute inflammation, accompanied by reduction of proinflammatory cytokines and elevation of anti-inflammatory cytokines. This study aims to identify the targeted proteins and predict the proteomic network associated with the drug action of QFGJS by using 2D gel and MALDI-TOF-MS/MS techniques. Thirty female Wistar rats were evenly grouped as normal and vehicle- and QFGJS-treated CIA rats. The antiarthritic effect of QFGJS was examined with a 19-day treatment course, and the knee synovial tissues of animals from each group were obtained for 2D gel and MALDI-TOF-MS/MS analysis. Results showed that QFGJS significantly ameliorated collagen II-induced arthritis when administrated at 2.8 g/kg body weight for 19 days. 2D gel image analysis revealed 89 differentially expressed proteins in the synovial tissues among the normal and vehicle- and QFGJS-treated CIA rats from over 1000 proteins of which 63 proteins were identified by MALDI-TOF-MS/MS analysis, and 32 proteins were included for classification of functions using Gene Ontology (GO) method. Finally, 14 proteins were analyzed using bioinformatics, and a predicted proteomic network related to the anti-arthritic effect of QFGJS was established, and Pgk1 plays a central role.

Alkylphenols from the roots of Ardisia brevicaulis induce G1 arrest and apoptosis through endoplasmic reticulum stress pathway in human non-small-cell lung cancer cells.

From the roots of Ardisia brevicaulis DIELS, two new alkylphenol derivatives, named ardisiphenol E (2) and F (3), have been isolated together with a known alkylphenol, ardisiphenol D (1). The structures of 1-3 were elucidated by chemical and spectroscopic techniques. Compounds 1 and 2 exhibited strong cytotoxicities on two human non-small-cell lung cancer cell lines (H1299 and A549). We found that compounds 1 and 2 upregulated mRNA and protein expressions of endoplasmic reticulum (ER) stress markers including C/EBP homologous protein (CHOP), binding immunoglobulin protein (Bip) and inositol-requiring enzyme 1 (IRE1) indicating 1 and 2 are novel natural ER stress inducers. Treatments with 1 and 5 µM of 1 or 2 triggered G1 arrest in H1299 and A549 cells with concomitant downregulation of ubiquitin fusion degradation protein 1 (Ufd1) and S-phase kinase-associated protein 2 (Skp2) proteins and the accumulation of p27, the key axes of ER stress-mediated G1 arrest. Compounds 1 and 2 also induced apoptosis at high concentrations (10, 20 µM) which was shown to be coupled with the upregulation of CHOP and Bim, the activation of caspase-9, caspase-3 and poly(ADP-ribose) polymerase (PARP) cleavage. These results indicate that compounds 1 and 2 induce ER stress that subsequently causes G1 arrest and apoptosis in human non-small-cell lung cancer cells and they may have potential anticancer effects.

Liposomes prolong the therapeutic effect of anti-asthmatic medication via pulmonary delivery.

PURPOSE: The main objective of this study was to develop a novel aerosolized liposome formulation for pulmonary delivery of anti-asthmatic medication and to explore the relationship between the bioavailability and anti-asthmatic efficacy of such a formulation. Asthma treatment usually requires frequent administration of medication for sustained bronchodilating response. Liposomes are known for their capability for sustained drug release and thus would be a suitable delivery system for anti-asthmatic medication for prolonged therapeutic effect. Salbutamol sulfate (SBS) was chosen as the model drug in this study because of its high water solubility and fast absorption after administration. METHODS: SBS was efficiently encapsulated into liposomes by the vesicular phospholipid gel technique. SBS permeability across the pulmonary membrane of an Asian toad was determined by in vitro study. Intratracheal administration of liposomes labeled with the fluorescent dye 1,1'-dioctadecyltetramethyl indotricarbocyanine iodide (DiR) in a rat model was assessed by a small animal imaging system and pharmacokinetic analysis. Pharmacodynamic analysis was performed in guinea pigs using the Konzett-Rössler method. RESULTS: SBS was efficiently encapsulated into liposomes with encapsulation efficiency as high as 70%. The particle size of the SBS liposome suspension was approximately 57 ± 21 nm. In the in vitro study of permeability across the pulmonary membrane of Asian toads, SBS from liposomes demonstrated a slower transport rate compared to free SBS solution. Pulmonary delivery of liposomes in a rat model showed that the liposomes were effectively distributed in the respiratory tract and lungs, and that the release of SBS from liposomes was sustained for at least 48 hours. Pharmacodynamic analysis in a guinea pig model showed that the anti-asthmatic effect of SBS liposomes persisted for up to 18 hours, whereas that of free SBS solution was less than 8 hours. CONCLUSION: The overall results demonstrated that liposomes could increase the concentration and retention time of SBS in the lungs and therefore prolong its therapeutic effect.

Paeoniflorin reduced acute toxicity of aconitine in rats is associated with the pharmacokinetic alteration of aconitine.

ETHNOPHARMACOLOGICAL RELEVANCE: To investigate the influence of paeoniflorin (major bioactive component of Paeonia lactiflora Pall.) on the pharmacokinetic behavior of aconitine (major toxic and bioactive component of Aconitum carmichaeli Debx.) and potential detoxifying effect of paeoniflorin on the acute toxicity of aconitine, which may provide in depth understanding to the toxicity reduction effect of Paeonia lactiflora to Aconitum carmichaeli. MATERIALS AND METHODS: Ultra high performance liquid chromatography coupled with triple quadrupole mass spectrometer (UHPLC-MS/MS) was employed to determine the plasma content of aconitine. Aconitine was administrated by oral to SD rats at the dosage of 200 µg/kg with or without paeoniflorin given by intraperitoneal injection at the dosage of 20 mg/kg. Plasma samples were collected for determination and analysis of pharmacokinetic parameters of aconitine. The LD(50) of aconitine and acute animal death induced by aconitine were examined when aconitine was given alone or jointly with paeoniflorin in ICR mice. RESULTS: A sensitive, accurate, precise, reliable and repeatable UHPLC-MS/MS method was successfully established for determination of the plasma content of aconitine in 12.5 µL plasma sample. The lower limit of quantification of aconitine was 0.01 ng/mL. Compared with the SD rats that were orally administrated with aconitine alone, the rats received aconitine and co-administrated with paeoniflorin by peritoneal injection showed a remarkably lower C(max) (5.69 ng/mL vs 9.66 ng/mL, P<0.05) and delayed T(max) (70 min vs 46 min, P<0.05), as well as a trend of reduction in AUC(0-t) (1082.75 ng-min/mL vs 1650.27 ng-min/mL, P=0.395). The LD(50) values of aconitine coadministered with 120 or 240 mg/kg of paeoniflorin were obviously increased to 2.30 and 2.15 mg/kg against 1.80 mg/kg of aconitine by oral administration alone. Mice treated with paeoniflorin (240 mg/kg) and aconitine (1.8 mg/kg) together revealed a significant decreased death rate than that received aconitine treatment alone (15% vs 50%, P<0.05). CONCLUSIONS: The acute oral toxicity of aconitine in rats was significantly reduced by paeoniflorin; this might result from the alterations of pharmacokinetic behavior of aconitine in the animals by coadministration of paeoniflorin.

Ginseng protects rodent hearts from acute myocardial ischemia-reperfusion injury through GR/ER-activated RISK pathway in an endothelial NOS-dependent mechanism.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginseng (Panax ginseng C.A. Meyer) is widely used in Asian communities for treating cardiovascular diseases. However, the mechanism by which it protects the myocardium in ischemia-reperfusion (I/R) injury remains unclear. In this study, we aim to investigate whether a standardized ginseng extract (RSE) protects rodent hearts against I/R injury and if glucocorticoid and/or estrogen receptor-mediated activation of Akt and Erk1/2 (the reperfusion injury salvage kinase pathway, RISK) and subsequent nitric oxide (NO) synthesis signaling are involved in this effect. MATERIALS AND METHODS: Rats or gene-deleted mice were subjected to 30 min ischemia by occluding the left anterior descending coronary artery and 90 min reperfusion. Infarct size, serum level of creatine kinase (CK), lactate dehydrogenase (LDH), and NO, expression and phosphorylation of glucocorticoid receptor (GR), estrogen receptor (ER), phosphatidylinositol-3 kinase (PI3K), Akt, NO synthase (NOS), extracellular signal-regulated kinase (Erk) 1/2, p38, and c-Jun NH2 terminal kinases (JNK) were examined in rat or mice treated with or without RSE in the absence or presence of pharmacological inhibitors. RESULTS: RSE significantly reduced infarct size in a dose-dependent manner and reduced the incidence of arrhythmia, increased serum NO production, reduced serum activities of creatine kinase and lactate dehydrogenase. The infarct size reduction effect of RSE was abolished by RU468 (an inhibitor of GR), tamoxifen (an inhibitor of ER), LY294002 (an inhibitor of PI3K), Akt inhibitor IV (an inhibitor of Akt protein kinase), U0126 (an inhibitor of Erk1/2) and NG-nitro-l-arginine methyl ester hydrochloride (an inhibitor of NOS), but not actinomycin D (an inhibitor of transcription process). RSE also significantly increased the activation of GR/ER, PI3K-Akt-eNOS cascades and Erk1/2 signaling in rat heart. However, RSE did not markedly reduce infarct size in endothelium NOS(-/-) mice. This differs from its effect in inducible NOS(-/-) and wild type mice, suggesting that endothelium NOS is required for the beneficial effect of RSE on the heart. CONCLUSION: Our findings showed for the first time that RSE protects hearts subjected to acute I/R injury and the infarct size reduction effect of RSE is associated with GR and/or ER-mediated Akt and Erk1/2 activation in an endothelium NOS-dependent manner.

Anti-inflammatory and analgesic effect of plumbagin through inhibition of nuclear factor-κB activation.

Plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone) (PL) is a naturally occurring yellow pigment found in the plants of the Plumbaginaceae, Droseraceae, Ancistrocladaceae, and Dioncophyllaceae families. It has been reported that PL exhibits anticarcinogenic, anti-inflammatory, and analgesic activities. However, the mechanism underlying its anti-inflammatory action remains unknown. In the current study, we investigated and characterized the anti-inflammatory and analgesic effects of PL orally administrated in a range of dosages from 5 to 20 mg/kg. We also examined the role of nuclear factor κB (NF-κB) and proinflammatory cytokines and mediators in this effect. The results showed that PL significantly and dose-dependently suppressed the paw edema of rats induced by carrageenan and various proinflammatory mediators, including histamine, serotonin, bradykinin, and prostaglandin E(2). PL reduced the number of writhing episodes of mice induced by the intraperitoneal injection of acetic acid, but it did not reduce the writhing episode numbers induced by MgSO(4) in mice or prolong the tail-flick reaction time of rats to noxious thermal pain. Mechanistic studies showed that PL effectively decreased the production of the proinflammatory cytokines interleukin 1ß, interleukin 6, and tumor necrosis factor α. It also inhibited the expression of the proinflammatory mediators inducible nitric-oxide synthase and cyclooxygenase 2, whereas it did not inhibit the expression of cyclooxygenase 1. Further studies demonstrated that PL suppressed inhibitor of κBα phosphorylation and degradation, thus inhibiting the phosphorylation of the p65 subunit of NF-κB. This study suggests that PL has a potential to be developed into an anti-inflammatory agent for treating inflammatory diseases.

Development of liposomal salbutamol sulfate dry powder inhaler formulation.

The purpose of our study was to develop a formulation of liposomal salbutamol sulfate (SBS) dry powder inhaler (DPI) for the treatment of asthma. Liposomes of high encapsulation efficiency (more than 80%) were prepared by a vesicular phospholipid gel (VPG) technique. SBS VPG liposomes were subjected to lyophilization using different kinds of cryoprotectants in various mass ratios. Coarse lactose (63-106 microm) in different mass ratios was used as a carrier. Magnesium stearate (0.5%) was added as a lubricator. The dry liposomal powders were then crushed by ball milling and sieved through a 400-mesh sieve to control the mean particle size at about 10 microm. The effects of different kinds of cryoprotectants and the amount of lactose carrier on the fine particle fraction (FPF) of SBS were investigated. The results showed that the developed formulation of liposomal dry powder inhaler was obtained using lactose as a cryoprotectant with a mass ratio of lyophilized powder to carrier lactose at 1 : 5; 0.5% magnesium stearate was used as a lubricator. The value of FPF for SBS was 41.51+/-2.22% for this formulation. Sustained release of SBS from the VPG liposomes was found in the in vitro release study. The study results offer the promising possibility of localized pulmonary liposomal SBS delivery in the anhydrous state.

Matrine induces cell anergy in human Jurkat T cells through modulation of mitogen-activated protein kinases and nuclear factor of activated T-cells signaling with concomitant up-regulation of anergy-associated genes expression.

Induction of immunotolerance has become a new strategy for treating autoimmune conditions in recent decades. However, so far there is no ideal therapeutics available for clinical use. Medicinal herbs are a promising potential source of immunotolerance inducers. In the current study, we sought first to optimize conditions for a validated cellular model of human Jurkat cells; and then used this model to screen bioactive compounds derived from medicinal plants for inducing T cell anergy in comparison with the effect of well-known T cell anergy inducer, ionomycin. The results showed that passage of the cells, and concentration and stimulation time of ionomycin on the cells could influence the ability of T cell anergy induction. Matrine, a small molecule derived from the root of Sophora flavescens AIT., was demonstrated to be effective in inducing T cell anergy in human Jurkat cells. The cells exposed to matrine showed markedly decreased mRNA expression of interleukin-2, an indicator of T cell anergy, when the cells were stimulated by antigens, anti-OKT3 plus anti-CD28. Mechanistic study showed that ionomycin and matrine could up-regulate the anergy-associated gene expressions of CD98 and Jumonji and activate nuclear factor of activated T-cells (NFAT) nuclear translocation in absence of cooperation of AP-1 in Jurkat cells. Pre-incubation with matrine or ionomycin could also shorten extracellular signal-regulated kinase (ERK) and suppress c-Jun NH(2)-terminal kinase (JNK) expression on the anergic Jurkat cells when the cells were stimulated with anti-OKT-3 plus anti-CD28 antibodies. Thus, matrine is a strong candidate for further investigation as a T cell immunotolerance inducer.

Pseudolaric acid B suppresses T lymphocyte activation through inhibition of NF-kappaB signaling pathway and p38 phosphorylation.

Pseudolaric acid B (PAB) is a major bioactive component of the medicinal plant Pseudolarix kaempferi. Traditional medicine practitioners in Asia have been using the roots of this plant to treat inflammatory and microbial skin diseases for centuries. In the current study, in vitro immunosuppressive effect of PAB and the underlying mechanisms have been investigated. The results showed that PAB dose-dependently suppressed human T lymphocyte proliferation, IL-2 production and CD25 expression induced by co-stimulation of PMA plus ionomycin or of anti-OKT-3 plus anti-CD28. Mechanistic studies showed that PAB significantly inhibited nuclear translocation of NF-kappaB p65 and phosphorylation and degradation of IkappaB-alpha evoked by co-stimulation of PMA plus ionomycin. PAB could also suppress the phosphorylation of p38 in the MAPKs pathway. Based on these evidences, we conclude that PAB suppressed T lymphocyte activation through inhibition of NF-kappaB and p38 signaling pathways; this would make PAB a strong candidate for further study as an anti-inflammatory agent.

Sinomenine ameliorates arthritis via MMPs, TIMPs, and cytokines in rats.

Rheumatoid arthritis (RA) is a chronic, inflammatory autoimmune disease that results in progressive joint destruction and substantial morbidity. The stem of the Chinese medicinal plant, Sinomenium acutum Rehder & Wilson (Family Menispermaceae), has been used to treat various rheumatic and arthritic diseases, of which the major bioactive component is sinomenine. We investigated the nature and molecular mechanisms of the anti-arthritic effect of sinomenine on collagen-induced arthritis in female Wistar rats. The results showed that sinomenine markedly suppressed the incidence and disease progression of established CIA, showing as dramatic reduction of paw swelling, ESR, and arthritic scores. Sinomenine suppressed the production of proinflammatory cytokines IL-1beta and IL-6 in serum, inhibited the protein expressions and activities of MMP-2 and MMP-9, and elevated the protein expressions and activities of TIMP-1 and TIMP-3 in rat paw tissues.

An optimized high-performance liquid chromatography (HPLC) method for benzoylmesaconine determination in Radix Aconiti Lateralis Preparata (Fuzi, aconite roots) and its products.

BACKGROUND: Benzoylmesaconine (BMA) is the main Aconitum alkaloid in Radix Aconiti Lateralis Preparata (Fuzi, aconite roots) with potent pharmacological activities, such as analgesia and anti-inflammation. The present study developed a simple and reliable method using BMA as a marker compound for the quality control of processed aconite roots and their products. METHODS: After extraction, a high-performance liquid chromatography (HPLC) determination of BMA was conducted on a RP-C18 column by gradient elution with acetonitrile and aqueous phase, containing 0.1% phosphoric acid adjusted with triethylamine to pH 3.0. RESULTS: A distinct peak profile was obtained and separation of BMA was achieved. Method validation showed that the relative standard deviations (RSDs) of the precision of BMA in all intra-day and inter-day assays were less than 1.36%, and that the average recovery rate was 96.95%. Quantitative analysis of BMA showed that the content of BMA varied significantly in processed aconite roots and their products. CONCLUSION: This HPLC method using BMA as a marker compound is applicable to the quality control of processed aconite roots and their products.

In vivo anti-inflammatory and analgesic activities of a purified saponin fraction derived from the root of Ilex pubescens.

The root of Ilex pubescens (Mao-Dong-Qing in Chinese, MDQ) has been commonly used for treatment of cardiovascular and inflammatory diseases in the Chinese medical system. The current studies aimed to investigate the anti-inflammatory and analgesic effects as well as the underlying mechanisms of a purified saponin fraction (PSF) derived from MDQ. PSF was found to significantly suppress the paw edema of rats induced by histamine given intraperitoneally at dosages ranging from 12.5-100 mg/kg. Meanwhile, PSF given orally at dosages of 200 and 100 mg/kg significantly inhibited acetic acid-induced abdominal writhing response of mice and prolonged the time required for mouse tail flick after exposure to a source of radiant heat. The mechanistic studies showed that cyclooxygenase-2 (COX-2) protein expression in carrageenan-injected paw tissues of rats was markedly attenuated by intraperitoneal injection of PSF at dosages of 12.5 to 100 mg/kg. PSF could also markedly inhibit production of proinflammatory cytokines, especially IL-1 beta, IL-6 and TNF-alpha, but enhance production of anti-inflammatory cytokines of IL-4 and IL-10 in the carrageenan-injected paw tissues in rats. These effects resulted in an overall attenuation of the ratio of proinflammatory/anti-inflammatory cytokines and, ultimately suppression of the paw edema. In conclusion, the current study has demonstrated the in vivo anti-inflammatory and analgesic effects of PSF, and suggested that the molecular mechanisms might be associated with inhibition of the elevated expression of COX-2 protein and the overproduction of the proinflammatory cytokines, as well as augmentation of the anti-inflammatory cytokines IL-4 and IL-10 in the carrageenan-injected paw tissues of rats.

Study on the pharmacokinetics and metabolism of paeonol in rats treated with pure paeonol and an herbal preparation containing paeonol by using HPLC-DAD-MS method.

Paeonol, a principal bioactive component of the Chinese herb Moutan Cortex with anti-inflammatory and analgesic effects, was comparatively studied to determine its pharmacokinetic behavior and metabolic profile in rat following oral administration of the pure paeonol alone and an herbal preparation "Qingfu Guanjieshu" (QFGJS) containing paeonol. An HPLC-DAD method was developed and validated for determining the concentration of paeonol in rat plasma. The in vivo time curves and AUC of paeonol at three doses were increased in a dose-dependent manner, while the pharmacokinetic parameters of paeonol in QFGJS at a comparable dosage were significantly elevated in comparison with those of pure paeonol. By using LC-Q/TOF-MS technique, four metabolites of paeonol were identified in plasma at 5min after dosing, with T(max) around 20min after treatment with the pure paeonol or QFGJS. Interestingly, relative concentrations of metabolites P2, P3 and P5 were markedly increased in plasma of rats treated with QFGJS compared with those of pure paeonol. These results indicate that other components in QFGJS could effectively influence the pharmacokinetic behavior and metabolic profile of paeonol in rats. The current studies emphasize the significance of the research toward an understanding of pharmacokinetic interactions of the co-existing components in the herbal preparations.

Combinative method using HPLC quantitative and qualitative analyses for quality consistency assessment of a herbal medicinal preparation.

A selective and efficient quality consistency assessment system was developed for monitoring the manufacturing processes of a Chinese herbal preparation, qingfu guanjieshu (QFGJS) capsule, and for assessing its stability over time. This system is based on quantitative determination of four marker compounds, i.e., sinomenine, paeoniflorin, paeonol, and curcumin, and on qualitative fingerprinting analysis of QFGJS using high-performance liquid chromatography-photodiode array detection (HPLC-DAD) method. The separation was performed on a Phenomenex ODS column by gradient elution with acetonitrile and aqueous phase (containing 0.1% phosphoric acid, adjusted with triethylamine to pH 3.5+/-0.2) at a flow-rate of 1.0 ml/min. In fingerprinting analysis, the chemical characteristics of four herbs present in QFGJS (excluding radix Aconiti Lateralis Preparata) were present in the HPLC chromatographic file. In addition, quantitative determination of hypaconitine was carried out with our published HPLC method as a supplement for quality control of the radix Aconiti Lateralis Preparata in QFGJS. The results showed that the contents of these five marker compounds and HPLC fingerprint profiles of three batches of QFGJS products collected at 3 months after production in the stability testing were relatively consistent. This well-developed method could be used for quality assessment of the complex preparations of herbal medicine.

Suppression of the onset and progression of collagen-induced arthritis in rats by QFGJS, a preparation from an anti-arthritic Chinese herbal formula.

QFGJS is an herbal preparation, and its pronounced effectiveness in treating adjuvant-induced arthritis (AIA) has been previously demonstrated. We herein aimed to confirm its anti-arthritic effect on collagen-induced arthritis (CIA) in rats. CIA was established in female Wistar rats with intradermal injection of type II bovine collagen at the base of the tail of animals. CIA rats were treated daily with oral administration of different doses of QFGJS beginning on the day of the induction of arthritis (day 0, the prophylactic treatment) or on the day after the onset of arthritis (day 13, the therapeutic treatment) until day 30. The results showed that prophylactic treatment with QFGJS significantly suppressed the onset of arthritis, and therapeutic treatment with QFGJS markedly reduced paw swelling and ESR levels even in the established CIA. Radiologic and histopathologic changes in the arthritic joints were also significantly reduced in the QFGJS-treated versus vehicle-treated rats. Moreover, the serum levels of pro-inflammatory cytokines TNF-alpha, IL-1beta, and IL-6 were markedly lowered in the QFGJS-treated rats. Hence, our studies demonstrate the quality, safety, and effectiveness of QFGJS as an anti-arthritic agent, which makes QFGJS a strong candidate for further clinical trials on rheumatoid arthritis (RA) patients.

Suppressive effects of JCICM-6, the extract of an anti-arthritic herbal formula, on the experimental inflammatory and nociceptive models in rodents.

JCICM-6, the extract of an anti-arthritic herbal formula composed of medicinal herbs of Sinomenium acutum, Aconitum carmichaeli DEBX., Curcuma Longa L., Paeonia lactiflora PALL., and Paeonia suffruticosa ANDR., was examined in the effectiveness and mechanism in reducing experimentally-induced inflammation and nociception using nine animal models. JCICM-6 was extracted from herbs and purified with Amberlite XAD-7HP adsorbent resin and analyzed with HPLC-fingerprint for quality consistency. In acute inflammatory models, the paw edema of rats was induced by subcutaneous injection of carrageenan or pro-inflammatory mediators, including histamine, serotonin, bradykinin, and prostaglandin E(2) (PGE(2)) into the right hind paws of animals; while the ear edema of mice was induced by applying arachidonic acid or 12-O-tetradecanoylphorbol 13-acetate (TPA) on the ear surface. In nociceptive models, the tail-flick response induced by radiant heat stimulation was measured and the numbers of abdominal writhing episodes of mice induced by intraperitoneal injection of acetic acid were recorded. JCICM-6 orally administered in a range of dosages from 0.438 g to 1.75 g/kg significantly and dose-dependently suppressed the paw edema of rats induced by carrageenan or various pro-inflammatory mediators and the ear edema of mice induced by arachidonic acid or TPA. JCICM-6 also significantly prolonged the reaction time of rats to radiant heat stimulation and reduced the numbers of writhing episodes of mice. These results indicated that JCICM-6 possesses significant anti-inflammatory and analgesic effects, which implies that it would be a potential candidate for further investigation as a new anti-arthritic botanical drug for humans.

The effects of sinomenine on intestinal absorption of paeoniflorin by the everted rat gut sac model.

Paeoniflorin and sinomenine, derived from the root of Paeonia lactiflora Pall. (family Ranunculaceae) and the stem of Sinomenium acutum Rehder & Wilson (family Menispermaceae), respectively, have been, and are currently, widely used for treatment of rheumatic and arthritic diseases in China and Japan. Our previous studies demonstrated that sinomenine could significantly improve the bioavailability of paeoniflorin in rats, but the underlying mechanisms remain unknown. The present study aims to investigate the intestinal kinetic absorptive characteristics of paeoniflorin as well as the absorptive behavior influenced by co-administration of sinomenine using an in vitro everted rat gut sac model. The results showed a good linear correlation between the paeoniflorin absorption in sac contents and the incubation time from 0 to 90 min. However, the concentration dependence showed that a non-linear correlation exists between the paeoniflorin absorption and its concentrations from 10 to 160 microM, and the absorption was saturated at about 80 microM of the drug. Sinomenine at 16 and 136 microM concentrations could significantly enhance the absorption of paeoniflorin (20 microM) by 1.5- and 2.5-fold, respectively. Moreover, two well-known P-glycoprotein inhibitors, verapamil and quinidine, could significantly elevate the absorption of paeoniflorin by 2.1- and 1.5-fold, respectively. Furthermore, sinomenine in a pattern, which influenced paeoniflorin's absorption, manifested as similar to that of P-glycoprotein inhibitors. In conclusion, sinomenine significantly enhance the intestinal absorption of paeoniflorin, subsequently improve the bioavailability of paeoniflorin. The mechanism underlying the improvement of paeoniflorin's bioavailability was proposed that sinomenine could decrease the efflux transport of paeoniflorin by P-glycoprotein.

Pharmacokinetic interaction of paeoniflorin and sinomenine: pharmacokinetic parameters and tissue distribution characteristics in rats and protein binding ability in vitro.

The root of Paeonia lactiflora and the stem of Sinomenium acutum are two herbs widely used in Chinese herbal medicine for the treatment of inflammatory and arthritic diseases. Studies on the interaction of the active constituents of these herbs, i.e., paeoniflorin and sinomenine, in pharmacokinetic parameters, tissues distribution, and protein binding ability could provide empirical data to support their clinical application. Following oral administration to rats, the pharmacokinetic alterations were compared. The results showed that the pharmacokinetic parameters (Cmax, Tmax, AUC, MRT, C(L), and Vd) of paeoniflorin were markedly enhanced when co-administrated with sinomenine. At 45 min after oral administration, the concentrations of paeoniflorin in the main internal organs were significantly increased when co-administrated with sinomenine. These phenomena were not ascribable to the alteration of the protein binding ability of paeoniflorin by sinomenine because obvious interactions of paeoniflorin and sinomenine in protein binding abilities in vitro to rat and rabbit plasma, human albumin, and alpha-1-acid-glycoprotein were not observed. However, with respect to the in vivo influence of paeoniflorin on sinomenine, the results showed that co-administration of paeoniflorin did not affect the pharmacokinetic parameters and tissue distribution of sinomenine.