Exploring the Mechanism of Topical Application of Clematis Florida in the Treatment of Rheumatoid Arthritis through Network Pharmacology and Experimental Validation.

Clematis Florida (CF) is a folk medicinal herb in the southeast of China, which is traditionally used for treating osteoarticular diseases. However, the mechanism of its action remains unclear. The present study used network pharmacology and experimental validation to explore the mechanism of CF in the treatment of rheumatoid arthritis (RA). Liquid chromatography-mass spectrometry (LC-MS/MS) identified 50 main compounds of CF; then, their targets were obtained from TCMSP, ETCM, ITCM, and SwissTargetPrediction databases. RA disease-related targets were obtained from DisGeNET, OMIM, and GeneCards databases, and 99 overlapped targets were obtained using a Venn diagram. The protein-protein interaction network (PPI), the compound-target network (CT), and the compound-potential target genes-signaling pathways network (CPS) were constructed and analyzed. The results showed that the core compounds were screened as oleanolic acid, oleic acid, ferulic acid, caffeic acid, and syringic acid. The core therapeutic targets were predicted via network pharmacology analysis as PTGS2 (COX-2), MAPK1, NF-κB1, TNF, and RELA, which belong to the MAPK signaling pathway and NF-κB signaling pathway. The animal experiments indicated that topical application of CF showed significant anti-inflammatory activity in a mouse model of xylene-induced ear edema and had strong analgesic effect on acetic acid-induced writhing. Furthermore, in the rat model of adjuvant arthritis (AA), topical administration of CF was able to alleviate toe swelling and ameliorate joint damage. The elevated serum content levels of IL-6, COX-2, TNF-α, IL-1ß, and RF caused by adjuvant arthritis were reduced by CF treatment. Western blotting tests showed that CF may regulate the ERK and NF-κB pathway. The results provide a new perspective for the topical application of CF for treatment of RA.

The complete chloroplast genome of Clematis florida Thunb. (Ranunculaceae), an ornamental and medicinal plant from Henan province, China.

Clematis florida Thunb. is a herbaceous and perennial plant native to East Asia. The plant is resistant to cold but sensitive to heat. It is an ornamental and medicinal plant that has great commercial potential. Here, we assembled and characterized the complete chloroplast (cp) genome of C. florida. The cp genome of C. florida was characterized by Illumina pair-end sequencing and is 159,606 bp in total length. The genome includes a large single-copy (LSC) region of 79,467 bp, a small single-copy (SSC) region of 18,057 bp, and a pair of inverted repeats (IR) regions of 31,041 bp. The genome contains 135 genes including 91 protein-coding, 36 tRNA, and eight rRNA genes. Phylogenetic analysis based on 18 Clematis species indicates that C. florida is closely related to C. fusca in the Ranunculaceae. The phylogenetic relationships and taxonomic status of C. florida revealed by cp genome were consistent with the previous molecular studies, and can serve as a reference for future studies on molecular biology, evolution, and taxonomy in the genus Clematis.

A new cyclopeptide alkaloid from Clematis Florida.

One new 14-membered ring cyclopeptide alkaloid, plenane A (1), along with six known compounds (2-7), was isolated from the roots of Clematis florida. Their structures were elucidated by means of NMR spectroscopic analysis and mass spectrometry. In addition, their anti- inflammatory effects on lipopolysaccharide-induced RAW264.7 macrophages were evaluated in vitro. The compounds 1, 2 and 6 exhibited potent indirect inhibitory effects, with IC50 values of 40.92, 22.88 and 6.32 µM, respectively.

The in vitro and in vivo anti-inflammatory activities of triterpene saponins from Clematis florida.

Clematis florida is widely used in She Ethnopharmacy in China owing to its significant anti-inflammatory activities. This study aimed to investigate the anti-inflammatory effect of the active fraction of C. florida (CFAF) in an arthritis animal model and its possible mechanism. Pre-inflammatory cytokine levels were examined by ELISA. CFAF can significantly improve the symptoms of arthritis such as paw swelling, arthritic index, and histological condition in AA rat. CFAF can also reduce levels of IL-1ß, TNF-α and IL-6. Further studies showed that triterpene saponins from CFAF induced anti-inflammatory activity inhibited inflammatory mediators by blocking JAK/STAT signalling pathways in the LPS-treated macrophages.

Cytotoxic Activities of Total Saponins from Plena Clematis on Human Tumor Cell Lines In Vitro.

OBJECTIVE: To investigate the anti-proliferative effects of saponins prepared from Plena Clematis (PC) cultured in Fujian Province, China on 4 human tumor cell lines and its possible anti-tumor mechanism. METHODS: The growth inhibition assays of saponins on human esophageal squamous carcinoma cell line (EC9706), human hepatoma cell line (HepG-2), human oral cancer cell line (KB) and human gastric cancer cell line (BGC-823) were evaluated in vitro by thiazolyl blue (MTT) method. The inhibitory effects on EC9706 treated with different concentrations of saponins (15.62, 31.25, 62.50, 125, 250 and 500 µg/mL) were performed in vitro by MTT method. The morphology and nuclear staining with acridine orange/ethidium bromide of EC9706 cells treated with saponins were illustrated under an inverted phase fluorescence microscope. The apoptotic effects of saponins were further evaluated by annexin-V/propidium iodide dual staining experiment to examine the occurrence of phosphatidylserine externalization onto the cell surface by a flflow cytometer. RESULTS: MTT assay showed that the saponins could inhibit the proliferation of 4 tumor cell lines. Among them, the maximum inhibition rate of 73.1% was detected in EC9706 cells at the saponins concentration of 250 µg/mL for 24 h. Further investigation indicated that the saponins induced EC9706 cells apoposis. The EC9706 cells presented apoptotic characteristics when treated with saponins, including that the morphologies of EC9706 cells were appeared round-shaped with higher refraction, and the cell nuclear stained orange with EB after 250 µg/mL saponins exposure. The flow cytometry analysis results showed that the induction of cell cycle arrest in apoptotic system may participate in the anti-proliferative activity of saponins on EC9706 cells. CONCLUSION: The saponins from PC exhibited significant cytotoxicity against human EC9706, KB, BGC-823, and HepG-2 cells and might be beneficial to development of ethnic pharmaceutical plant for potential anti-tumor drugs.

GC-MS analysis of volatile oil from traditional She medicine Clematis Florida var. plena flower / 中草药

Objective: Clematis florida var. plena is a traditional She medicine. In this paper, we aim to study the main chemical components of essential oil of C. florida var. plena flower. The study will provide reference for large-scale reasonable cultivation and quality standard establishment of C. florida var. plena. Methods: C. florida var. plena flower's essential oil was extracted with steam distillation and it was identified by GC-MS. Area normalization method was used to measure the percentage of each components. Results: In the experiment, 32 element were detected in essential oil of C. florida var. plena flower, among which 20 chemical components were identified. There are 13 chemical components over 1%, and the top five compounds are palmitic acid (26.94%), phytol (10.58%), linoleic acid (6.13%), pentadecane (4.54%) and n-tricosane (3.84%). Conclusion: GC-MS was first used to analyze the main chemical components of C. florida var. plena's flower essential oil. The study provides scientific basis for cultivation, production, development, utilization of C. florida var. plena.

Cytotoxic Activities of Total Saponins from Plena Clematis on Human Tumor Cell Lines In Vitro / 中国结合医学杂志

OBJECTIVE@#To investigate the anti-proliferative effects of saponins prepared from Plena Clematis (PC) cultured in Fujian Province, China on 4 human tumor cell lines and its possible anti-tumor mechanism.@*METHODS@#The growth inhibition assays of saponins on human esophageal squamous carcinoma cell line (EC9706), human hepatoma cell line (HepG-2), human oral cancer cell line (KB) and human gastric cancer cell line (BGC-823) were evaluated in vitro by thiazolyl blue (MTT) method. The inhibitory effects on EC9706 treated with different concentrations of saponins (15.62, 31.25, 62.50, 125, 250 and 500 μg/mL) were performed in vitro by MTT method. The morphology and nuclear staining with acridine orange/ethidium bromide of EC9706 cells treated with saponins were illustrated under an inverted phase fluorescence microscope. The apoptotic effects of saponins were further evaluated by annexin-V/propidium iodide dual staining experiment to examine the occurrence of phosphatidylserine externalization onto the cell surface by a flflow cytometer.@*RESULTS@#MTT assay showed that the saponins could inhibit the proliferation of 4 tumor cell lines. Among them, the maximum inhibition rate of 73.1% was detected in EC9706 cells at the saponins concentration of 250 μg/mL for 24 h. Further investigation indicated that the saponins induced EC9706 cells apoposis. The EC9706 cells presented apoptotic characteristics when treated with saponins, including that the morphologies of EC9706 cells were appeared round-shaped with higher refraction, and the cell nuclear stained orange with EB after 250 μg/mL saponins exposure. The flow cytometry analysis results showed that the induction of cell cycle arrest in apoptotic system may participate in the anti-proliferative activity of saponins on EC9706 cells.@*CONCLUSION@#The saponins from PC exhibited significant cytotoxicity against human EC9706, KB, BGC-823, and HepG-2 cells and might be beneficial to development of ethnic pharmaceutical plant for potential anti-tumor drugs.

Effect and Mechanism of Total Saponins from Clematis Florida Thunb. in Adjuvant-induced Arthritis Rats / 中国药师

Objective:To investigate the effect of total saponins from Clematis florida Thunb. on rat model of adjuvant arthritis ( AA) . Methods:Wistar rats were randomly divided into six groups:the normal control group, the model control group, the positive control group ( indomethacin, 10 mg?kg-1 ) , high, medium and low dose groups of total saponins from Clematis florida Thunb. with ten rats in each group. The rat model of AA was established with Freund's complete adjuvant and the effects of drugs on body weight and foot swelling of AA rats were observed. After the animals were sacrificed, the serum levels of superoxide dismutase ( SOD) , mal-onaldehyde ( MDA) , nitric oxide ( NO) and glutathion peroxidase ( GSH-PX) were determined. The serum levels of interleukin-8 ( IL-8), IL-10 and tumor necrosis factor-α(TNF-α) were detected by ELISA. Results:On the 8th day of drug administration, total sapo-nins from Clematis florida Thunb. could effectively alleviate the changes in the body weight and foot swelling of the rats to a certain ex-tent when compared with the model control group (P <0. 05 or P <0. 01). There were no significant differences between the high dose group and the positive control group (P>0. 05), and there were significant differences between the low dose group and the high/medium dose groups (P<0. 05). The levels of SOD, MDA, NO and GSH-PX in serum of AA rats were significantly affected by the total saponins from Clematis florida Thunb. . Except for NO indicator of low dose group, the differences between the other groups and the model control group were statistically significant (P<0. 05 or P<0. 01), and there were significant differences in MDA, NO and GSH-PX levels between high dose group and low dose group (P<0. 05). Compared with those in the model control group, the serum levels of IL-8 and TNF-αin each total saponins from Clematis florida Thunb. group significantly decreased and the content of IL-10 sig-nificantly increased (P<0. 05 or P<0. 01). There was no significant differences in the levels of IL-10 and TNF-α between the high dose group and the positive control group (P>0. 05), and the differences in IL-8 and IL-10 between high dose group and low dose group were statistically significant (P<0. 05). Conclusion: The total saponins from Clematis florida Thunb. have good therapeutic effect on AA, and the underlying mechanism may be related to the reduction of lipid peroxidation and the inhibition of pro-inflammatory cytokines.

Effect and Mechanism of Total Saponins from Clematis Florida Thunb. in Adjuvant-induced Arthritis Rats / 中国药师

Objective:To investigate the effect of total saponins from Clematis florida Thunb. on rat model of adjuvant arthritis ( AA) . Methods:Wistar rats were randomly divided into six groups:the normal control group, the model control group, the positive control group ( indomethacin, 10 mg?kg-1 ) , high, medium and low dose groups of total saponins from Clematis florida Thunb. with ten rats in each group. The rat model of AA was established with Freund's complete adjuvant and the effects of drugs on body weight and foot swelling of AA rats were observed. After the animals were sacrificed, the serum levels of superoxide dismutase ( SOD) , mal-onaldehyde ( MDA) , nitric oxide ( NO) and glutathion peroxidase ( GSH-PX) were determined. The serum levels of interleukin-8 ( IL-8), IL-10 and tumor necrosis factor-α(TNF-α) were detected by ELISA. Results:On the 8th day of drug administration, total sapo-nins from Clematis florida Thunb. could effectively alleviate the changes in the body weight and foot swelling of the rats to a certain ex-tent when compared with the model control group (P <0. 05 or P <0. 01). There were no significant differences between the high dose group and the positive control group (P>0. 05), and there were significant differences between the low dose group and the high/medium dose groups (P<0. 05). The levels of SOD, MDA, NO and GSH-PX in serum of AA rats were significantly affected by the total saponins from Clematis florida Thunb. . Except for NO indicator of low dose group, the differences between the other groups and the model control group were statistically significant (P<0. 05 or P<0. 01), and there were significant differences in MDA, NO and GSH-PX levels between high dose group and low dose group (P<0. 05). Compared with those in the model control group, the serum levels of IL-8 and TNF-αin each total saponins from Clematis florida Thunb. group significantly decreased and the content of IL-10 sig-nificantly increased (P<0. 05 or P<0. 01). There was no significant differences in the levels of IL-10 and TNF-α between the high dose group and the positive control group (P>0. 05), and the differences in IL-8 and IL-10 between high dose group and low dose group were statistically significant (P<0. 05). Conclusion: The total saponins from Clematis florida Thunb. have good therapeutic effect on AA, and the underlying mechanism may be related to the reduction of lipid peroxidation and the inhibition of pro-inflammatory cytokines.

Determination of Active Ingredient Oleanolic Acid in She Medicine Clematis Florida var. Plena by HPLC / 中国药师

Objective:To establish a method for the determination of oleanolic acid in She medicine Clematis florida var. plena. Methods:The HPLC method was carried on a Lanbo-Kromasil C18 chromatography column using acetonitrile-water ( 86∶ 14 ) as the mobile phase. The detection wavelength was 205 nm, the flow rate was 1. 0 ml·min-1 , and the column temperature was 25 ℃. Re-sults:The calibration curve of oleanolic acid was linear within the range of 0. 52-15. 60μg (r=0. 999 9). The average recovery of oleanolic acid was 92. 13%(RSD=3. 0%, n=9). Conclusion:The method is simple, rapid, accurate and reproducible, which can be used as an ideal method for the quality control of She medicine Clematis florida var. plena. .