Estrogen antagonizes ASIC1a-induced chondrocyte mitochondrial stress in rheumatoid arthritis.

BACKGROUND: Destruction of articular cartilage and bone is the main cause of joint dysfunction in rheumatoid arthritis (RA). Acid-sensing ion channel 1a (ASIC1a) is a key molecule that mediates the destruction of RA articular cartilage. Estrogen has been proven to have a protective effect against articular cartilage damage, however, the underlying mechanisms remain unclear. METHODS: We treated rat articular chondrocytes with an acidic environment, analyzed the expression levels of mitochondrial stress protein HSP10, ClpP, LONP1 by q-PCR and immunofluorescence staining. Transmission electron microscopy was used to analyze the mitochondrial morphological changes. Laser confocal microscopy was used to analyze the Ca2+, mitochondrial membrane potential (Δψm) and reactive oxygen species (ROS) level. Moreover, ASIC1a specific inhibitor Psalmotoxin 1 (Pctx-1) and Ethylene Glycol Tetraacetic Acid (EGTA) were used to observe whether acid stimulation damage mitochondrial function through Ca2+ influx mediated by ASIC1a and whether pretreatment with estrogen could counteract these phenomena. Furthermore, the ovariectomized (OVX) adjuvant arthritis (AA) rat model was treated with estrogen to explore the effect of estrogen on disease progression. RESULTS: Our results indicated that HSP10, ClpP, LONP1 protein and mRNA expression and mitochondrial ROS level were elevated in acid-stimulated chondrocytes. Moreover, acid stimulation decreased mitochondrial membrane potential and damaged mitochondrial structure of chondrocytes. Furthermore, ASIC1a specific inhibitor PcTx-1 and EGTA inhibited acid-induced mitochondrial abnormalities. In addition, estrogen could protect acid-stimulated induced mitochondrial stress by regulating the activity of ASIC1a in rat chondrocytes and protects cartilage damage in OVX AA rat. CONCLUSIONS: Extracellular acidification induces mitochondrial stress by activating ASIC1a, leading to the damage of rat articular chondrocytes. Estrogen antagonizes acidosis-induced joint damage by inhibiting ASIC1a activity. Our study provides new insights into the protective effect and mechanism of action of estrogen in RA.

Potential therapeutic effect of Shufeng Jiedu capsule and its major herbs on coronavirus disease 2019 (COVID-19): A review.

The outbreak and rapid spread of coronavirus disease 2019 (COVID-19) poses a huge threat to human health and social stability. Shufeng Jiedu capsule (SFJDC), a patented herbal drug composed of eight medicinal plants, is used to treat different viral respiratory tract infectious diseases. Based on its antiviral, anti-inflammatory, and immunoregulatory activities in acute lung injury, SFJDC can be effectively used as a treatment for COVID-19 patients according to the diagnosis and treatment plan issued in China and existing clinical data. SFJDC has been recommended in 15 therapeutic regimens for COVID-19 in China. This review summarizes current data on the ingredients, chemical composition, pharmacological properties, clinical efficacy, and potential therapeutic effect of SFJDC on COVID-19, to provide a theoretical basis for its anti-viral mechanism and the clinical treatment of COVID-19.

Acid-sensing ion channel 1a mediates acid-induced pyroptosis through calpain-2/calcineurin pathway in rat articular chondrocytes.

The pyroptosis is a causative agent of rheumatoid arthritis, a systemic autoimmune disease merged with degenerative articular cartilage. Nevertheless, the precise mechanism of extracellular acidosis on chondrocyte pyroptosis is largely unclear. Acid-sensing ion channels (ASICs) belong to an extracellular H+ -activated cation channel family. Accumulating evidence has highlighted activation of ASICs induced by extracellular acidosis upregulate calpain and calcineurin expression in arthritis. In the present study, to investigate the expression and the role of acid-sensing ion channel 1a (ASIC1a), calpain, calcineurin, and NLRP3 inflammasome proteins in regulating acid-induced articular chondrocyte pyroptosis, primary rat articular chondrocytes were subjected to different pH, different time, and different treatments with or without ASIC1a, calpain-2, and calcineurin, respectively. Initially, the research results showed that extracellular acidosis-induced the protein expression of ASIC1a in a pH- and time-dependent manner, and the messenger RNA and protein expressions of calpain, calcineurin, NLRP3, apoptosis-associated speck-like protein, and caspase-1 were significantly increased in a time-dependent manner. Furthermore, the inhibition of ASIC1a, calpain-2, or calcineurin, respectively, could decrease the cell death accompanied with the decreased interleukin-1ß level, and the decreased expression of ASIC1a, calpain-2, calcineurin, and NLRP3 inflammasome proteins. Taken together, these results indicated the activation of ASIC1a induced by extracellular acidosis could trigger pyroptosis of rat articular chondrocytes, the mechanism of which might partly be involved with the activation of calpain-2/calcineurin pathway.

ASIC1a induces synovial inflammation via the Ca2+/NFATc3/ RANTES pathway.

Rationale: Synovial inflammation is one of the main pathological features of rheumatoid arthritis (RA) and is a key factor leading to the progression of RA. Understanding the regulatory mechanism of synovial inflammation is crucial for the treatment of RA. Acid-sensing ion channel 1a (ASIC1a) is an H+-gated cation channel that promotes the progression of RA, but the role of ASIC1a in synovial inflammation is unclear. This study aimed to investigate whether ASIC1a is involved in the synovial inflammation and explore the underlying mechanisms in vitro and in vivo. Methods: The expression of ASIC1a and nuclear factor of activated T cells (NFATs) were analyzed by Western blotting, immunofluorescence, and immunohistochemistry both in vitro and in vivo. The Ca2+ influx mediated by ASIC1a was detected by calcium imaging and flow cytometry. The role of ASIC1a in inflammation was studied in rats with adjuvant-induced arthritis (AA). Inflammatory cytokine profile was analyzed by protein chip in RA synovial fibroblasts (RASF) and verified by a magnetic multi-cytokine assay and ELISA. The NFATc3-regulated RANTES (Regulated upon activation, normal T cell expressed and secreted) gene transcription was investigated by ChIP-qPCR and dual-luciferase reporter assay. Results: The expression of ASIC1a was significantly increased in human RA synovial tissues and primary human RASF as well as in ankle synovium of AA rats. Activated ASIC1a mediated Ca2+ influx to increase [Ca2+]i in RASF. The activation/overexpression of ASIC1a in RASF up-regulated the expression of inflammatory cytokines RANTES, sTNF RI, MIP-1a, IL-8, sTNF RII, and ICAM-1 among which RANTES was increased most remarkably. In vivo, ASIC1a promoted inflammation, synovial hyperplasia, articular cartilage, and bone destruction, leading to the progression of AA. Furthermore, activation of ASIC1a upregulated the nuclear translocation of NFATc3, which bound to RANTES promoter and directly regulated gene transcription to enhance RANTES expression. Conclusion: ASIC1a induces synovial inflammation, which leads to the progression of RA. Our study reveals a novel RA inflammation regulatory mechanism and indicates that ASIC1a might be a potential therapeutic target for RA.

Effects of autophagy on apoptosis of articular chondrocytes in adjuvant arthritis rats.

Rheumatoid arthritis (RA) is a chronic, systemic autoimmune disease that eventually leads to joint deformities and loss of joint function. Previous studies have demonstrated a close relationship between autophagy and the development of RA. Although autophagy and apoptosis are two different forms of programmed death, the relationship between them in relation to RA remains unclear. In this study, we explored the effect of autophagy on apoptosis of articular chondrocytes in vivo and in vitro. Adjuvant arthritis (AA) and acid-induced primary articular chondrocyte apoptosis were used as in vivo and in vitro models, respectively. Articular chondrocyte autophagy and apoptosis were both observed dynamically in AA rat articular cartilage at different stages (15 days, 25 days and 35 days). Moreover, chondrocyte apoptosis and articular cartilage injury in AA rats were increased by the autophagy inhibitor 3-methyladenine (3-MA) and decreased by the autophagy activator rapamycin. In addition, pre-treatment with 3-MA increased acid-induced chondrocyte apoptosis, while pre-treatment with rapamycin reduced acid-induced chondrocyte apoptosis in vitro. These results suggest that autophagy might be a potential target for the treatment of RA.

The role of Ca2+ in acid-sensing ion channel 1a-mediated chondrocyte pyroptosis in rat adjuvant arthritis.

Rheumatoid arthritis is an autoimmune disease with a poor prognosis. Pyroptosis is a type of proinflammatory programmed cell death that is characterised by the activation of caspase-1 and secretion of the proinflammatory cytokines interleukin (IL)-1ß/18. Previous reports have shown that pyroptosis is closely related to the development of some autoimmune diseases, such as rheumatoid arthritis. The decrease in the pH of joint fluid is a main pathogenic feature of RA and leads to excessive apoptosis in chondrocytes. Acid-sensitive ion channels (ASICs) are extracellular H+-activated cation channels that mainly influence Na+ and Ca2+ permeability. In this study, we investigated the role of Ca2+ in acid-sensing ion channel 1a-mediated chondrocyte pyroptosis in an adjuvant arthritis rat model. The expression of apoptosis-associated speck-like protein, NLRP3, caspase-1, ASIC 1a, IL-1ß and IL-18 was upregulated in the joints of rats compared with that in normal rats, but the expression of Col2a in cartilage was decreased. However, these changes were reversed by amiloride, which is an inhibitor of ASIC1a. Extracellular acidosis significantly increased the expression of ASIC1a, IL-1ß, IL-18, ASC, NLRP3 and caspase-1 and promoted the release of lactate dehydrogenase. Interestingly, Psalmotoxin-1 (Pctx-1) and BAPTA-AM inhibited these effects. These results indicate that ASIC1a mediates pyroptosis in chondrocytes from AA rats. The underlying mechanism may be associated with the ability of ASIC1a to promote [Ca2+]i and upregulate the expression of the NLRP3 inflammasome.

[Morphological, microscopic, multiple-component assay and fingerprinting based systematic research on quality evaluation of Moutan Cortex(Paeonia suffruticosa)].

The purpose of this study was to combine morphological, microscopic, UHPLC multiple-component assay and fingerprinting studies in order to evaluate the quality of Moutan Cortex (MC) systematically. The root system of Paeonia suffruticosa was measured to compare the morphological variation and the chemical composition of different grades of MC was discussed according to previous studies. The difference between the main microscopic features of MC powder and the xylem powder is dramatic, the MC powder contains great amount of starch granules and clusters of calcium oxalate, while the xylem powder displays considerable vessels. Interestingly, the growth rings of P. suffruticosa was first reported in the xylem of the root transection, this can help to determine the growth years of the plant. Moreover, through the assay of 16 component, MC produced in Tongling and Bozhou in Anhui province were compared, content of PGG in MC produced in Bozhou was significantly higher than MC produced in Tongling (P<0.01). MC with different growth years, MC with xylem and unprocessed MC and MC decoction pieces were compared respectively by combining the results of 16 compounds assay and fingerprinting. It is proposed that the quality evaluation standard include the assay of paeoniflorin. Above all, the holistic quality difference can be evaluated more comprehensively by combining multiple analytical methods.

Interleukin-1ß and tumor necrosis factor-α augment acidosis-induced rat articular chondrocyte apoptosis via nuclear factor-kappaB-dependent upregulation of ASIC1a channel.

The acute-phase proinflammatory cytokines interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) demonstrate high-level expression and pleiotropic biological effects, and contribute to the progression and persistence of rheumatoid arthritis (RA). Acid hydrarthrosis is also an important pathological characteristic of RA, and the acid-sensing ion channel 1a (ASIC1a) plays a critical role in acidosis-induced chondrocyte cytotoxicity. However, the roles of IL-1ß and TNF-α in acid-induced apoptosis of chondrocytes remain unclear. Rat adjuvant arthritis and primary articular chondrocytes were used as in vivo and in vitro model systems, respectively. ASIC1a expression in articular cartilage was increased and highly colocalized with nuclear factor (NF)-κB expression in vivo. IL-1ß and TNF-α could upregulate ASIC1a expression. These cytokines activated mitogen-activated protein kinase and NF-κB pathways in chondrocytes, while the respective inhibitors of these signaling pathways could partially reverse the ASIC1a upregulation induced by IL-1ß and TNF-α. Dual luciferase and gel-shift assays and chromatin immunoprecipitation-polymerase chain reaction demonstrated that IL-1ß and TNF-α enhanced ASIC1a promoter activity in chondrocytes by increasing NF-κB DNA-binding activities, which was in turn prevented by the NF-κB inhibitor ammonium pyrrolidinedithiocarbamate. IL-1ß and TNF-α also decreased cell viability but enhanced LDH release, intracellular Ca2+ concentration elevation, loss of mitochondrial membrane potential, cleaved PARP and cleaved caspase-3/9 expression, and apoptosis in acid-stimulated chondrocytes, which effects could be abrogated by the specific ASIC1a inhibitor psalmotoxin-1 (PcTX-1), ASIC1a-short hairpin RNA or calcium chelating agent BAPTA-AM. These results indicate that IL-1ß and TNF-α can augment acidosis-induced cytotoxicity through NF-κB-dependent up-regulation of ASIC1a channel expression in primary articular chondrocytes.

Origins, Phytochemistry, Pharmacology, Analytical Methods and Safety of Cortex Moutan (Paeonia suffruticosa Andrew): A Systematic Review.

Cortex Moutan (CM), a well-known traditional Chinese medicine, is commonly used for treating various diseases in China and other eastern Asian countries. Recorded in Pharmacopeias of several countries, CM is now drawing increasing attention and under extensive studies in various fields. Phytochemical studies indicate that CM contains many valuable secondary metabolites, such as monoterpene glycosides and phenols. Ample evidence from pharmacological researches suggest that CM has a wide spectrum of activities, such as anti-inflammatory, anti-oxidant, anti-tumor, anti-diabetic, cardiovascular protective, neuroprotective, hepatoprotective effects. Moreover, various analytical methods were established for the quality evaluation and safety control of CM. This review synopsizes updated information concerning the origins, phytochemistry, pharmacology, analytical method and safety of CM, aiming to provide favorable references for modern CM research and application. In conclusion, continuing pharmacological investigations concerning CM should be conducted to unravel its pharmacological mechanisms. Further researches are necessary to obtain comprehensive and applicable analytical approach for quality evaluation and establish harmonized criteria of CM.

[Study on Appraising Storage Periods of Moutan Cortex and Its Quality by Means of Colorimeter].

Objective: To examine the correlation between storage periods and L*, a* and b* color of Moutan Cortex. Methods: A optical density meter was used for the measurement of reflected light from sieved powder and section samples using the CIE 1976 L~*,a~*,b~* color system. The content of paeonol were determined by HPLC. The correlation between storage periods,paeonol content and color indices of Moutan Cortex was analyzed. Results: The measured color was significantly correlated with storage periods. The color of Moutan Cortex shift toward the red with the increase of storage periods. The storage periods were correlated with paeonol content. The measured color was equably correlated with paeonol content. Conclusion: The correlation between the color of Moutan Cortex and storage periods was found in this study. Measurment of the color of Moutan Cortex can be used to appraise its storage periods and quality.

Total flavonoids of Bidens bipinnata L. ameliorate experimental adjuvant-induced arthritis through induction of synovial apoptosis.

BACKGROUND: Bidens bipinnata are widely distributed in China, which have been widely used as a traditional Chinese medicine. The aim of this study was to examine the effect of total flavonoids of Bidens pilosa L. (TFB) on adjuvant arthritis (AA) and its possible mechanisms. METHODS: The macroscopic scoring of paw edema, secondary paw swelling, and polyarthritis index were measured. Histological examination of the joints and the serum concentrations of IL-6, IL-1beta, and TNF-alpha were examined. Apoptosis in synovial tissue was detected. The expression of Caspase 3 cleavage, serves as a marker undergoing apoptosis, was confirmed by Western blot. RESULTS: TFB attenuated the severity of arthritis on paw edema, hind paw volume, and polyarthritis index of AA rats, improved the histological status in AA rats as well. TFB can inhibit the production of IL-6, IL-1beta, and TNF-alpha from serum. Clear DNA ladder formation was observed in DNA extraction of synovium from TFB treated AA rats. The number of apoptosis was increased with TFB treatment in TUNEL assay. TFB treatment on AA rats significantly increased the expression of Caspase 3 in synovium. CONCLUSIONS: Our data suggest that TFB has a significant anti-arthritic effect in AA through the induction of apoptosis in synovial.

Acute, but not chronic, stress increased the plasma concentration and hypothalamic mRNA expression of NUCB2/nesfatin-1 in rats.

Nesfatin-1, a newly discovered satiety peptide, has recently been reported to be involved in the stress response. Stress-induced expression of nesfatin-1 has been reported and few studies focus on its expression in the hypothalamus, which is the center of the stress response. To test our hypothesis that peripheral and hypothalamic nesfatin-1 overexpression should play an important role in the stress response and the associated hyperactivity of hypothalamic-pituitary-adrenal (HPA) axis, acute stress (AS) was induced using water avoidance stress (WAS), and chronic unpredictable mild stress (CUMS) was also induced using 3 consecutive weeks of 7 different stressors. The behavior of CUMS rats was evaluated by an open field test (OFT), sucrose preference test (SPT), and forced swimming test (FST). The activity of the HPA axis was detected by measurement of the plasma corticosterone concentration and hypothalamic mRNA expression of corticotropin-releasing-hormone (CRH). The plasma concentration and hypothalamic mRNA expression of nesfatin-1 were measured with an enzyme-linked immunosorbent assay (ELISA) and real-time fluorescent quantitative PCR, respectively. The results showed that both AS and CUMS increased the plasma corticosterone concentration and hypothalamic CRH mRNA expression. Depression-like behavior was induced in CUMS rats, as indicated by a decreased movement distance, frequency of rearing and grooming in the OFT, and sucrose preference index and increased immobility in the FST. Moreover, the AS rats showed increased plasma concentration and hypothalamic mRNA expression of nesfatin-1, which were positively correlated with the plasma corticosterone concentration and hypothalamic CRH expression, respectively. These results indicated that acute stress, but not chronic stress, increased the plasma concentration and hypothalamic mRNA expression of NUCB2/nesfatin-1 in rats.

[Chemical constituents from Bidens bipinnata].

To investigate the chemical constituents of the whole plants of Bidens bipinnata, the separation and purification of constituents were performed by chromatography on macroporous resin, silica gel, MCI and Sephadex LH-20. Their structures were elucidated by spectroscopic data as quercetin (1), quercetin-3-0-alpha-L-rhamnoside (2), keampferol-3-O-beta-D-glucopyranoside (3), keampferol-3-O-alpha-L-rhamnoside (4), 3', 5-dyhydroxy-3, 6, 4'-trimethoxyl -7-O-beta-D-glucopyranoside flavonoid (5), 7, 8, 3', 4'-tetraflavanone(6), (2S)- and (2R)-isookanin-7-O-beta-D- glucopyranoside (7a/7b), (2S)- and (2R)-3'-methoxy-isookanin-8-O-beta-D-glucopyranoside (8a/8b), 6, 7, 3', 4'-tetrahydroxyaurone(9), maritimetin (10), esculetin (11), 3-O-caffeoyl-2-methyl-d-erythrono-1, 4-lactone (12), (7S, 8R) balanophonin-4-O-beta-D-glucopyranoside (13), eugenyl-O-beta-apiofuranosyl-( 1"-6') -O-beta-glucopyranoside (14), and (+)-syringaresinol-4'-O-beta-D-glucopyranoside (15). Compounds 8, 13, 14, and 15 were isolated from this genus for the first time. Compounds 1 and 6 were potent inhibitors against HSC-T6 cells in vitro and compounds 1, 2, 6, and 7 were capable of decreasing the inflammatory cytokine production of macrophage cells in vitro.

[Analysis of fingerprint and bioactive components of Bidens biternata by HPLC].

OBJECTIVE: To establish HPLC fingerprint of Bidens biternata from different habitats and determine the contents of hyperoside, isoquercetin, astragalin and bipinnatapolyacetylpside. METHODS: Analysis was carried on Hypersil ODS C18 column (4.6 mm x 250 mm, 5.0 microm) with acetonitrile and 3% acetic acid as the mobile phase in a gradient elution. The contents of 4 components were determined simultaneously. RESULTS: The fingerprint of 10 populations were established and the data were analyzed by the similarity evaluation software. There were almost no differences between the similarities of 10 population, but the contents of 4 main compoerls were different among them. CONCLUSION: This method is stable and reliable which could be applied in quality assessment.

Synthesis, characterization, drug-loading capacity and safety of novel pH-independent amphiphilic amino acid copolymer micelles.

A novel block copolymer containing two polymeric components, poly(L-aspartic acid)-b-poly (L-phenylalanine) (PAA-PPA), was synthesized and its potential for the preparation of copolymer micelles with a poorly water-soluble drug was investigated in this study. The chemical structure and physical properties of PAA-PPA were characterized by FTIR, 1H NMR and TG. The degree of polymerization of PAA-PPA was calculated by analyzing the relative area of N-CH signal and C-CH3 of 1H NMR spectra. The critical micelle concentration (CMC) of the PAA-PPA achieved a minimum of 11.1 mg/L. Studies on the drug-free PAA-PPA solutions showed PAA-PPA aggregation into micellar type in the sub-150 nm size range. Furthermore, the size of the PAA-PPA micelles was found to be pH-independent between the pH range of 4.0 and 8.0, which could be favorable to avoid the limitation of the size change at the specified pH value seeking drug stability. 4-amino-2-trifluoromethyl-phenyl retinate (ATPR) was studied as a poorly water-soluble model drug. The drug-loading and entrapment efficiency of the ATPR-loaded PAA-PPA micelles were 30.9 wt% and 87.9 %, respectively. The high drug-loading and entrapment efficiency were due to the synergistic effect of the micellar encapsulation and the binding interaction between drug and PAA-PPA. The ATPR-loaded PAA-PPA micelles showed a narrow size distribution, low zeta potential, high drug-loading capacity and good stable. The PAA-PPA was safer than Tween-80 and Cremophor EL (CrmEL) as an injectable pharmaceutical adjuvant for ATPR as indicated by the hemolysis and cytotoxicity studies. The novel amphiphilic amino acid copolymer can be considered as a prospective injectable delivery system for ATPR in terms of the pH-independent, greater drug-loading capacity and safety.

Inhibition of platelet aggregation by curdione from Curcuma wenyujin essential Oil.

INTRODUCTION: Curdione, one of the major sesquiterpene compounds from Rhizoma Curcumae, has been shown to exhibit multiple bioactive properties. In this study, we investigated the anti-platelet aggregation and antithrombotic activities of curdione with different methods both in vitro and in vivo. The purpose of the study was to explore an inhibitor of platelet aggregation, which promised to be a preventive or therapeutic agent for various vascular diseases. MATERIALS AND METHODS: Curdione was isolated from the essential oil of Curcuma wenyujin using the silica gel column chromatography method. The effects of curdione on human platelet aggregation induced by thrombin (0.3 U/ml), platelet-activating factor (PAF, 0.375 µg/ml), adenosine diphosphate (ADP, 10 µM) and arachidonic acid (AA, 0.1mg/ml) were tested in vitro, and the potential mechanisms underlying such activities were investigated. We also tested the antithrombotic effect of curdione in a tail thrombosis model. RESULTS AND CONCLUSIONS: Curdione preferentially inhibited PAF- and thrombin- induced platelet aggregation in a concentration-dependent manner (IC(50): 60-80 µM), whereas much higher concentrations of curdione were required to inhibit platelet aggregation induced by ADP and AA. Curdione also inhibited P-selectin expression in PAF-activated platelets. Moreover, curdione caused an increase in cAMP levels and attenuated intracellular Ca(2+) mobilization in PAF-activated platelets. In vivo, we also found that curdione showed significant antithrombotic activity. Therefore, we conclude that the inhibitory mechanism of curdione on platelet aggregation may increase cAMP levels and subsequently inhibit intracellular Ca(2+) mobilization. Furthermore, the effect observed in the tail thrombosis model might be explained completely by increased vasodilation. These results indicate that curdione may be one of the main bioactive constituents in Rhizoma Curcumae that removes blood stasis.

Effects of leonurine hydrochloride on medically induced incomplete abortion in early pregnancy rats.

OBJECTIVES: To determine the effect of leonurine hydrochloride (LH) on abnormal bleeding induced by medical abortion. STUDY DESIGN: Rats had incomplete abortions induced in early pregnancy using mifepristone in combination with misoprostol. After abortion, rats were treated with LH for 7 days, and the duration and volume of uterine bleeding were observed. Approximately 30min after the last treatment, the animals were killed and the uterine shape was observed. The sinistro-uteri were suspended in organ baths to record the contraction curves, including the frequency and tension for 10min; the dextro-uteri were fixed with formaldehyde for pathologic evaluation. In addition, blood samples were collected from the femoral artery for the measurement of estradiol (E2) and progesterone (P) levels by radioimmunoassay. RESULTS: In in vivo experiments, compared with the model group, LH treatment markedly reduced the volume of bleeding and intrauterine residual, and significantly shortened the duration of bleeding. From the contraction curve, LH notably reinforced the frequency and tension of uterine contractions. LH remarkably elevated the serum estradiol level in rats, but had no obvious effect on progesterone level. CONCLUSIONS: LH has an inhibitory effect on bleeding caused by incomplete abortion; the mechanism may be related to up-regulation of the E2 level, leading to an increase in uterine contractions and evacuation of intrauterine residuum.

Inhibition of acid-sensing ion channels in articular chondrocytes by amiloride attenuates articular cartilage destruction in rats with adjuvant arthritis.

OBJECTIVE: The aim of this study was to examine whether drugs such as amiloride that block acid sensing ion channels (ASICs) could attenuate articular cartilage destruction in adjuvant-induced arthritis (AA). METHODS: Articular chondrocytes were isolated from the normal rats, and intracellular calcium ([Ca(2+)]i) was analyzed with laser scanning confocal microscopy. The cell injury was analyzed with lactate dehydrogenase release assay and electron microscopy. Amiloride or phosphate buffered saline was administered daily to AA rats for 1 week from the time of arthritis onset. Morphology of the articular cartilage was examined by hematoxylin and eosin staining, and Mankin score was calculated. The expression level of type II collagen (COII) and aggrecan mRNA and proteins in the articular cartilage was evaluated by real-time PCR and Western blotting, respectively. RESULTS: The rapid decrease in extracellular pH (6.0) induced a conspicuous increase in [Ca(2+)]i in the articular chondrocytes. Amiloride reduced this increase in [Ca(2+)]i, and inhibited acid-induced articular chondrocyte injury. Amiloride significantly decreased Mankin scores in the articular cartilage in AA rats. COII and aggrecan mRNA and protein expression in the articular cartilage was significantly increased by amiloride. CONCLUSION: These findings represent some experimental evidence of a potential role for ASICs in the pathogenesis of articular cartilage destruction in rheumatoid arthritis.

Protective effects of total flavonoids of Bidens bipinnata L. against carbon tetrachloride-induced liver fibrosis in rats.

Bidens bipinnata L. is well known in China as a traditional Chinese medicine and has been used to treat hepatitis in clinics for many years. In a previous study we found that total flavonoids of Bidens bipinnata L. (TFB) had a protective effect against carbon tetrachloride (CCl4)-induced acute liver injury in mice. Now this study was designed to investigate its therapeutic effect against CCl4-induced liver fibrosis in rats and to determine, in part, its mechanism of action. The liver fibrosis model was established by subcutaneous injection of 50% CCl4 twice a week for 18 weeks. TFB (40, 80 and 160 mg kg(-1)) was administered by gastrogavage daily from the 9th week. The results showed that TFB (80 and 160 mg kg(-1)) treatment for 10 weeks significantly reduced the elevated liver index (liver weight/body weight) and spleen index (spleen weight/body weight), elevated levels of serum transaminases (alanine aminotransferase and aspartate aminotransferase), hyaluronic acid, type III procollagen and hepatic hydroxyproline. In addition, TFB markedly inhibited CCl4-induced lipid peroxidation and enhanced the activity of the antioxidant enzymes superoxide dismutase and glutathione peroxidase. Moreover, TFB (80 and 160 mg kg(-1)) treatment improved the morphologic changes of hepatic fibrosis induced by CCl4 and suppressed nuclear factor (NF)-kappaB, alpha-smooth muscle actin (SMA) protein expression and transforming growth factor (TGF)-beta1 gene expression in the liver of liver fibrosis of rats. In conclusion, TFB was able to ameliorate liver injury and protect rats from CCl4-induced liver fibrosis by suppressing oxidative stress. This process may be related to inhibiting the induction of NF-kappaB on hepatic stellate cell activation and the expression of TGF-beta1.

Protective effects of total flavonoids of Bidens pilosa L. (TFB) on animal liver injury and liver fibrosis.

The hepatoprotective effects of total flavonoids of Bidens pilosa L. (TFB), a traditional Chinese medicine were evaluated in carbon tetrachloride (CCl(4))-induced liver injury in mice and rats. Total flavonoids of Bidens pilosa L. (25, 50 and 100mg/kg) were administered via gavage daily for 10 days to CCl(4)-treated mice as well as TFB (30, 60 and 90mg/kg) administered for 6 weeks to CCl(4)-treated rats. Liver index (liver weight/body weight), serum levels of transaminases (alanine aminotransferase, ALT and aspartate aminotransferase, AST), hepatic malondialdehyde (MDA) content, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were evaluated following the 10 days treatment in mice. In addition histopathologic changes and nuclear factor-kappaB (NF-kappaB) expression of the liver were detected with hematoxylin-eosin (HE) and immunohistochemistry methods, respectively. The results showed that TFB (50 and 100mg/kg) effectively reduced the CCl(4)-induced elevated liver index, serum ALT, AST levels, hepatic MDA content, and restored hepatic SOD, GSH-Px activities in acute liver injury mice. TFB (60 and 90mg/kg) treatment significantly inhibited NF-kappaB activation in liver fibrosis of rats. The histopathological analysis suggested that TFB reduced the degree of liver injury in mice and severity of liver fibrosis in rats. These results suggested that TFB had a protective and therapeutic effect on animal liver injury, which might be associated with its antioxidant properties and inhibition of NF-kappaB activation.