| Purpose:Arisaematis rhizoma preparatum (ARP) wasis a product obtained by dried rhizome of Araceae Arisaema processing with ginger and alum, which was used to phlegm dampness, dispelling wind and relieving convulsion, dispelling node and other aspects, especially had a good effects of clinical for rheumatoid arthritis (RA). Currently, substance basis and mechanism of toxicity of RA has been thorough research, but the material basis and mechanism of ARP haven’t study thoroughly, mechanism of original (increasing) effect of processing of RA haven’t break. So, we choose to study the anti rheumatoid arthritis, through by tracking of active site isolation of ARP, investigate ARP of anti RA substance basis and mechanism of action and the active ingredient in vivo absorption and excretion of were preliminary studied. Hope to provide research foundation of original (increasing) effect of processing of RA, and clarify the material basis and mechanism of ARP.Method:1 Study on effect of ARP extracts anti rheumatoid arthritis. Type Ⅱ collagen induced a arthritis (CIA) rat model was built, and scores of paw swelling, arthritis index, rat weight, index of immune organs, serum inflammation factor IL-1β, IL-6, IL-10 and TNF-a level were used to investigate the anti rheumatoid arthritis effect of ARP extract (100,200,400 mg/kg), in vivo.2 Extractive parts isolation and active screening of ARP. A semipermeable membrane (Mw:5000) was used to divided ARP into high molecular material component and small molecules material component. The active of anti-inflammatory and inhibition the proliferation of CD4+T cells as the index were screening the effect of extractive site.3 Chemical compositions isolation and active screening of ARP. ARP macromolecules were divided into polysaccharide and non-polysaccharide site, and the further separation and purification of the polysaccharide by DEAE-52 and Sephadex column to obtain homogeneous polysaccharide. The active of anti-inflammatory and inhibition the proliferation of CD4+T cells as the index were screening the effect of extractive site.4 The preliminary study of anti-inflammatory mechanism of ARP. The protein expression of p38, JNK and ERK 1/2, I kappa B, NF kappa B p65 were detected by Western blotting of ARP polysaccharide (ARPA) on LPS induced RAW264.7 cells.5 The preliminary study of immunoregulation mechanism of ARP. The morphology of Jurkat was staining by hoechest 33258 and PI, cell apoptosis rate and cycle were investigated by flow cytometry, apoptosis signal pathway protein expression of caspase 8, caspase 9, caspase 3, Bcl-2 and Bax by Western blotting.6 Research of ARPA on rheumatoid arthritis. The Complete Freund’s adjuvant (CFA) was used to induced arthritis (AA) rats model, paw swelling, arthritis index, rat weight, index of immune organs, serum inflammation factor IL-1β, TNF-a level and histopathological observation as evaluation index, investigate the effects of ARPA (20, 40,60 mg/kg) anti rheumatoid arthritis effect, in vivo.7 The preliminary pharmacokinetics of ARPA. The stability of ARPA labeled by fluorescein fluorescein (FITC) was detect in biological samples. To establish a method for the quantitative detection of fluorescein labeled ARPA, to determine the pharmacokinetics of ARPA in rats by fluorescence spectrophotometry and calculate the pharmacokinetic parameters, the excretion of drug in rats was studied also.8 The content changes of ARPA after processing. Through optimization the preparation process of ARPA, preparation polysaccharide with a batch of Rhizoma Arisaematis and ARISAEMATIS RHIZOMA PREPARATUM, and compared with the characterization of ARPA. Simultaneous determination the content of ARPA in Rhizoma Arisaematis and ARISAEMATIS RHIZOMA PREPARATUM.Results:1 ARP doses of 400 mg/kg and 200 mg/kg has obvious improvement of CIA rat paw swelling, can reduce the CIA rat spleen index, can inhibit rat peripheral blood promoting inflammatory factor TNF alpha, IL-1 beta and IL-6 levels, while increasing anti inflammatory cytokine IL-10 levels.2 ARP macromolecules group divided the anti-inflammatory activity and inhibit the activity of CD4+ T cell proliferation and in a concentration dependent manner; small molecular substances group almost not on Jurkat cell proliferation rate is affected, and have a certain degree of promoting effect of the development of inflammation.3 Six homogeneous polysaccharides from ARP total polysaccharide, respectively ARPA, ARPB, ARPC-I, ARPC-II, ARPD and ARPE. ARPA and ARPB both have obvious anti-inflammatory activity, and inhibit the proliferation of ARPA and ARPC-II is the strongest.4 The possible mechanisms of anti-inflammatory activity. ARPA may inhibit I kappa B degradation and regulation of NF-kappa B p65 binding, which could synthesis of inflammatory factors. At the same time, it could inhibiting the phosphorylation of p38 and ERK1/2, then impact the activation of downstream nuclear transcription factor. The expression of phosphorylated JNK has not reduced.5 The possible mechanism of immunoregulation activity. ARPA can make Jurkat cells arrest in G2/M phase, resulting in inhibition of cell proliferation and apoptosis induced by Jurkat. ARPA induces apoptosis is through activate of Caspase-3 and caspase-9, and not the caspase8. Further research of the mitochondrial pathway of apoptosis, ARPA can significantly increased Pro apoptotic protein Bax expression and down-regulation of the expression of anti apoptotic protein Bcl-2.6 ARPA at dose of 60mg/kg can significantly decreased paw swelling and total score of AA rats, decreased thymus index, inhibit rat peripheral blood promoting inflammatory factor TNF-a, IL-1β levels, and synovial biopsy results show that rats given 60mg/kg of ARPA treatment, the inflammatory cell infiltration was significantly reduced.7 ARPA combined with tyramine, then FITC was labeling with ARPA-TYR. ARPA-TYR-FITC has final stability in PBS solution, plasma and urine. The plasma, urine and feces of rats were existed in the form of the prototype after oral administration. ARPA-TYR-FITC has a long elimination half-life, the larger the apparent volume of distribution in the body; mainly through feces excreted in the form of the original drug.8 The content of ARPA in Rhizoma Arisaematis and ARISAEMATIS RHIZOMA PREPARATUM. The stable ARPA can be prepared by the optimized process, and the content of ARPA in Rhizoma Arisaematis is 1.4 times higher than that of the processed products.Conclusion:Homogeneous polysaccharide A is one of the material basis of anti rheumatoid arthritis activity of Arisaematis Rhizoma Preparatum, even after oral administration of ARPA, a small portion of the absorbed blood was absorbed into the blood in rats., and mainly in the form of the original drug through feces excreted. The mechanism of action were through regulation of MAPK/NF-nuclear factor kappa B inflammatory signal pathway to improve the external inflammatory symptoms of RA. At the same time, ARPA was induce the apoptosis of CD4+T cells through the mitochondrial pathway. Polysaccharide A exist in Arisaematis Rhizoma and Arisaematis Rhizoma Preparatum, the content of ARPA was decreased about 28% through processing. |
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