Effects and mechanisms of total glucosides of paeony on synoviocytes activities in rat collagen-induced arthritis

The aim of the study was to investigate the effects of TGP, an active compound extracted from the roots of Paeonia lactiflora Pall, on the activities of synoviocytes in rats with collagen-induced arthritis (CIA) and its possible mechanisms. CIA was induced in male Sprague-Dawley (SD) rats immunized with chicken type II collagen (CII) in Freund's complete adjuvant (FCA). Synoviocytes proliferation was determined by 3-(4, 5-2dimethylthiazal-2yl) 2, 5-diphenyltetrazoliumbromide (MTT) assay. Tumor necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1), prostaglandin E(2) (PGE(2)) and cyclic adenosine monophosphate (cAMP) levels in synoviocytes were measured by radioimmunoassay (RIA). E-prostanoid (EP)(2) and EP(4) receptors were analyzed by Western blot analysis. The results showed that TGP significantly inhibited the proliferation of synoviocytes, decreased the production of IL-1, TNF-alpha and PGE(2) and elevated the levels of cAMP. Further study showed that TGP could up-regulate the expression of EP(2) and EP(4). These results indicated that TGP might exert its anti-inflammatory effects through inhibiting the production of pro-inflammatory mediators in synoviocytes of CIA rats, which might be associated with its ability to regulate cAMP-dependent EP(2)/EP(4)-mediated pathway.     

Paeoniflorin inhibits function of synoviocytes pretreated by rIL-1α and regulates EP4 receptor expression

Ethnopharmacological relevance

To investigate the effect of the Paeoniflorin (Pae), a main active component of total glucosides of paeony (TGP) extracted from the root of Paeonia lactiflora, on regulation of synoviocytes cultured from rats collagen-induced arthritis (CIA) in vitro.

Materials and methods

CIA was induced in male Sprague-Dawley rats immunized with chicken type II collagen (CCII) in Freund's complete adjuvant. The levels of interleukin-1 (IL-1), tumor necrosis factor α (TNF-α), prostaglandin E₂ (PGE₂) and cyclic adenosine monophosphate (cAMP) were measured by radioimmunoassay. The proliferation responses was determined by the 3-(4,5-2dimethylthiazal-2yl) 2,5-diphenyltetrazoliumbromide (MTT) assay. Expression of E-prostanoid (EP₄) receptor was detected by Western blotting technique.

Results

Treatment of Pae (2.5, 12.5, 62.5 μg/ml) significantly decreased the production of IL-1 and TNF-α. Recombinant interleukin-1 (rIL-1α) (10 ng/ml) apparently stimulated synoviocyte, thymocyte and splenocyte proliferation, and Pae (12.5, 62.5 μg/ml) inhibited abnormal proliferation responses stimulated by rIL-1α. Moreover, rIL-1α time- and concentration-dependently increased production of PGE₂. The production of PGE₂ produced by synoviocytes from CIA rats significantly inhibited by administration of Pae (12.5, 62.5 μg/ml). rIL-1α (10 ng/ml) decreased cAMP of synoviocytes cells treated for 24 h. Similarly rIL-1α (0.1, 1, 10 ng/ml) induced a concentration-dependent decrease in the production of cAMP at 24 h. Pae (12.5, 62.5 μg/ml) increased the production of cAMP in synoviocytes. The immunoblot, Pae (12.5, 62.5 μg/ml) apparently increased the expression of EP₄ receptor in synoviocytes stimulated by rIL-1α (10 ng/ml).

Conclusions

The present study indicates that Pae might exert its anti-inflammatory effects through suppressing synoviocytes function and regulating immune cells responses in CIA rats, which might be associated with its ability to up-regulate the E-prostanoid (EP₄) receptor protein expression and modulate intracellular cAMP level.     

The expression change of β-arrestins in fibroblast-like synoviocytes from rats with collagen-induced arthritis and the effect of total glucosides of paeony

Aim of the study

To investigate the expression of β-arrestins in fibroblast-like synoviocytes (FLS) from collagen-induced arthritis (CIA) rats and the effect of total glucosides of paeony (TGP).

Materials and methods

TGP and glucosides of tripterygium wilfordii (GTW) were intragastriclly administrated to collagen-induced arthritis (CIA) rats after immunization. The secondary inflammatory reaction was evaluated by hind paw swelling, polyarthritis index and histopathological changes. Antibodies to type II collagen (CII) were determined by enzyme-linked immunosorbent assay (ELISA). Synoviocyte proliferations were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl (MTT) assay. The expression of β-arrestins in synoviocytes from CIA rats was measured by western blot.

Results

The administration of TGP (25, 50, 100 mg/kg) depressed hind paw swelling and decreased the arthritis scores of CIA rats. TGP improved the pathologic manifestations of CIA. Serum anti-CII antibodies level increased significantly in CIA rats, while TGP had no effect on it. Fibroblast-like synoviocytes (FLS) proliferation was inhibited by TGP (50, 100 mg/kg). On d14, d28 after immunization, β-arrestins expression greatly up-regulated in synoviocytes from CIA rats and then returned to baseline levels on d42 after immunization. TGP (50, 100 mg/kg) significantly reduced the expression of β-arrestins.

Conclusion

An inflammatory process in vivo induces an up-regulation of β-arrestins in synoviocytes from CIA rats while TGP can inhibit this change, which might be one of the important mechanisms for TGP to produce a marked therapeutic effect on RA.     

Therapeutic effects and mechanisms of total flavonoids of Turpinia Arguta Seen on adjuvant arthritis in rats

AIM: To investigate the therapeutic effects and mechanisms of total flavonoids of Turpinia Arguta Seen (TFS) on adjuvant arthritis in rats. METHODS: The model of adjuvant arthritis was induced by injection of Freund's Complete Adjuvant (FCA). Secondary paw swelling of AA rats was measured with volume meter and polyarthritis index were scored. The splenocyte proliferation, (interleukin-1) IL-1 and interleukin-2 (IL-2) production were assayed by cell proliferation assay. Prostaglandin E(2) (PGE(2)) production was determined by radioimmunoassay. RESULTS: TFS (80, 160, 320 mg/kg, i.g.) could significantly inhibit secondary inflammatory reaction (secondary swelling, multiple arthritis, pathologic change of ankle arthritis) in AA rats. The results in vivo showed that the low response of splenocytes to concanavalin A (Con A) and lipopolysaccharide (LPS) and the decreased IL-2 synthesis were restored in AA rats treated with TFS (160, 320 mg/kg, i.g.), while the elevated IL-1 and PGE(2) released from peritoneal macrophages (PMphi) were also reduced. CONCLUSION: TFS has significant therapeutic effect on AA rats, which might be relate to its immunoregulatory actions.     

通痹灵对大鼠佐剂性关节炎滑膜细胞功能的影响

目的：研究通痹灵（ＴＢＬ）对佐剂性关节炎（ＡＡ）大鼠滑膜成纤维增殖及滑膜细胞分泌白细胞介素１（ＩＬ－１），肿瘤坏死因子α（ＴＮＦ－α）和前列腺素Ｅ２（ＰＧＥ２）的影响。     

雷公藤、白芍总苷、苦杏仁水煎剂对佐剂型关节炎大鼠治疗效果及机制的比较

目的比较雷公藤、白芍总苷、苦杏仁水煎剂对佐剂性关节炎(AA)大鼠模型抗炎作用效果及机制差异,为类风湿关节炎的防治提供理论依据。方法建立AA大鼠模型,比较雷公藤、白芍总苷、苦杏仁水煎剂对AA大鼠足踝关节组织病理及血清中促炎因子肿瘤坏死因子α(TNF-α)、免疫细胞间黏附分子(sICAM-1)水平的影响。结果雷公藤、白芍总苷、苦杏仁水煎剂均能有效抑制关节肿胀,减轻炎症的发展,并降低AA大鼠血清中TNF-α、sICAM-1的水平。与雷公藤相比,白芍总苷、苦杏仁水煎剂更能降低血清中TNF-α的水平。结论雷公藤、白芍总苷、苦杏仁水煎剂均可起到抗类风湿关节炎作用,白芍总苷、苦杏仁水煎剂可有效降低血清中TNF-α的水平。     

白头翁皂苷对佐剂性关节炎大鼠FZD8表达的影响

研究中药白头翁有效成分白头翁皂苷(pulchinenoside,PULC)对佐剂性关节炎(adjuvant arthritis,AA)大鼠Frizzled(FZD)表达的影响。采用足跖注射完全弗氏佐剂制备AA大鼠,原代培养AA大鼠滑膜成纤维样滑膜细胞(fibroblast-like synoviocytes,FLS),Real-time qPCR检测PULC灌胃治疗对AA大鼠FLS FZD8表达的影响,MTT,ELISA检测FZD8敲除对FLS增殖和IL-1,IL-6,IL-8表达的影响,Real-time qPCR检测miR-375在FZD8异常表达中的作用。检测发现经PULC灌胃治疗后,AA大鼠FLS中FZD8表达明显降低,FZD8敲除后FLS增殖明显减弱,IL-1,IL-6,IL-8表达明显减少,PULC灌胃治疗后miR-375表达明显升高,上调表达的miR-375能够抑制FZD8的表达。PULC可能通过上调miR-375表达抑制FZD8的表达。     

鹿瓜多肽口服液对大鼠佐剂性关节炎的治疗作用及其机制

 目的研究鹿瓜多肽口服液(lugua polypeptide solution,LGPS)对大鼠佐剂性关节炎(adjuvant arthritis,AA)的治疗作用,并探讨其可能的作用机制。方法复制大鼠AA模型后第8天,LGPS按3.5,7和14mg·kg^-1给大鼠灌胃治疗,每天1次,连续16d。于给药第4,7,10,13,16及18天测定大鼠左、右足跖容积,计算肿胀率。治疗后第19天测定大鼠腹腔巨噬细胞吞噬功能,关节渗出液中PGE2含量,血液流变学,血清SOD活性及MDA、NO、IL-1β、TNF-α和IL-6含量。结果LGPS能明显降低AA大鼠右后关节的原发性肿胀和左后关节的继发性肿胀(P<0.05～P<0.001),可使腹腔巨噬细胞吞噬功能,全血低(5/s)、中(80/s)、高切(160/s)黏度,血沉,血清MDA、NO、IL-1β、TNF-α及IL-6含量明显降低,血清SOD活性明显提高(P<0.05或P<0.01),对关节渗出液中PGE2含量则无明显影响(P>0.05)。结论LGPS对大鼠AA有明显治疗作用,可能与其抑制巨噬细胞活化,降低血清中IL-1、TNF-α和IL-6含量,提高机体对氧自由基的清除力及改善血液流变性有关。     

白芍总苷对佐剂关节炎大鼠足爪组织病理的影响及MMP-9蛋白表达的抑制作用

目的:探讨白芍总苷(total glucosides of paeonia:TGP)对佐剂关节炎大鼠足爪组织中基质金属蛋白酶-9(MMP-9)蛋白表达的抑制作用和对大鼠足爪组织病理改变的影响及血清中血管内皮生长因子(VEGF)和白介素-1β(IL-1β)含量的影响。方法:建立Ⅱ型胶原诱导的佐剂关节炎大鼠模型,用免疫组化染色法检测足爪组织中MMP-9蛋白的表达,病理切片观察不同剂量的TGP对佐剂关节炎大鼠的治疗效果,用ELISA法检测佐剂关节炎大鼠血清中VEGF和IL-1β的含量。结果:白芍总苷中(50 mg/kg)、大(100 mg/kg)剂量组的胞浆内MMP-9的阳性表达比模型对照组明显降低,TGP中、大剂量治疗组与地塞米松组能明显改变佐剂关节炎大鼠皮下组织细胞排列、炎性细胞浸润及血管增生现象,TGP 25 mg/kg组对改善佐剂关节炎大鼠足部皮肤的病理状况无明显作用。中、大剂量的TGP组和地塞米松组血清VEGF和IL-1β的含量明显降低。结论:TGP对佐剂关节炎大鼠炎症的抑制作用,可能与其下调MMP-9蛋白的表达,抑制炎性细胞浸润及血管增生,降低炎性细胞因子VEGF和IL-1β的产生有关。     

白芍总苷对大鼠胶原性关节炎及其免疫功能的影响

 目的研究白芍总苷(TGP)对大鼠胶原性关节炎(CIA)及其免疫功能的影响。方法采用鸡Ⅱ型胶原诱导大鼠CIA模型,关节评分法检测关节炎发生情况,ELSIA法检测大鼠血清中抗CⅡ抗体的水平,MTT法检测T细胞和B细胞增殖反应,IL-1、IL-2活性的测定采用小鼠淋巴细胞增殖法。结果大鼠于致炎后第14天开始出现多发性关节炎症状,体重减轻,血清中抗CⅡ抗体水平升高,TGPig给药,连续14d,能够明显抑制CIA大鼠的AI,促进大鼠体重的恢复,降低血清中抗CⅡ抗体水平。进一步研究表明,TGP体内、外给药均能抑制CIA大鼠增高的T、B淋巴细胞增殖反应,还能显著降低刀豆蛋白A(ConA)诱导的CIA大鼠脾淋巴细胞产生IL-2的能力以及脂多糖(LPS)诱导的腹腔巨噬细胞产生IL-1的能力。结论TGP对CIA大鼠具有治疗作用,其机制可能与其调节CIA大鼠异常的免疫功能,平衡细胞因子的产生有关。     

益肾蠲痹丸对类风湿性关节炎的药效学研究

目的：使用弗氏完全佐剂制作大鼠关节炎模型，评价益’肾蠲痹丸对类风湿性关节炎的药效。方法：使用弗氏完全佐剂造模后，测量益肾蠲痹丸对对侧足爪肿胀和脏器指数的影响：ELISA法检测其对血清肿瘤坏死因子Ⅱ（TNF-Ⅱ），白介素1β（IL—1β），白介素-6（IL-6）的影响；分光光度法测量总一氧化氮合酶（tNOS）和诱导型一氧化氮合酶（iNOS）活性：并用Westernblot法检测益肾蠲痹丸对凋亡蛋白Bcl-2表达的影响。结果：益肾蠲痹丸可有效抑制大鼠佐剂性关节炎所引起的足跖肿胀，且对胸腺脾脏指数无明显影响。益肾蠲痹丸可有效抑制血清中TNF-Ⅱ，IL-1β，IL-6含量：抑制血清中iNOS活性；下调Bcl-2在滑膜组织中的表达。结论：益肾蠲痹丸的抗类风湿关节炎的作用与其调节主要炎症因子，下调炎症部位Bcl-2蛋白表达有关。     

马钱子总碱囊泡凝胶对AA大鼠的治疗作用

目的:观察马钱子总碱囊泡凝胶对佐剂性关节炎(AA)大鼠的治疗作用及其机制。方法:SD大鼠随机分为9组:正常组、AA模型组、马钱子总碱囊泡凝胶高、中、低剂量(25,12.5,6.25 mg·kg-1)组、马钱子总碱药物凝胶高、中、低剂量(25,12.5,6.25 mg·kg-1)组和扶他林对照组(双氯芬酸二乙胺乳胶剂,50 mg·kg-1)。除正常组外,其余各组均以弗氏完全佐剂(FCA)每鼠左后足跖皮内注射0.1 mL致炎,各给药组在大鼠足趾部分均匀涂抹药物并用油纸包裹。足容积法测量继发侧足肿胀度,进行疼痛评分和多发性关节炎评分,HE染色观察关节病理损伤,酶联免疫吸附测定法(ELISA)测定白细胞介素1(IL-1)、肿瘤坏死因子α(TNF-α)、前列腺素(PGE2)、白细胞介素6(IL-6)和血管内皮细胞生长因子(VEGF)的表达。结果:与模型组相比,囊泡凝胶和药物凝胶可明显减轻AA大鼠的继发侧足爪肿胀度(17～21 d)。马钱子总碱囊泡凝胶和药物凝胶可有效抑制大鼠多发性关节炎指数,减轻关节病理学损伤,马钱子总碱囊泡凝胶还可降低滑膜组织匀浆上清液IL-1,TNF-α,PGE2,IL-6水平和提高VEGF水平。结论:马钱子总碱囊泡凝胶可拮抗大鼠AA发展,其作用机制与抑制炎症介质的生成密切相关。     

马钱子配伍苏木对佐剂关节炎大鼠滑膜组织IL-1β、IL-62及IL-10的影响

目的：通过观察马钱子配伍苏木（简称马苏）对佐剂关节炎（AA）大鼠滑膜中IL-1β、IL-6、IL-10含量的影响,探讨马苏配伍后的抗炎机制并寻求最佳配比。方法：按随机数字表法将91只Wistar大鼠分为空白组,模型组,马钱子单煎剂组,马苏1：6、1：12、1：18、1：24煎剂组。除空白组外于大鼠右后足跖垫部皮内注射完全弗氏佐剂（CFA）0．1mL,复制AA大鼠模型,测定AA大鼠滑膜中IL-1β、IL-6、IL-10含量的变化。结果：马苏1：18煎剂组能显著降低AA大鼠滑膜中IL-1β、IL-6的含量（P〈0．05）,且能升高AA大鼠滑膜中IL-10的含量（P〈0．05）。推测马苏是通过降低AA大鼠滑膜中IL-1β含量、减少炎性因子IL-6的分泌,升高保护性因子IL-10含量发挥其抗炎作用。结论：马苏对AA具明显的抗炎作用,其最佳配比为马苏1：18煎剂。     

赤雹根总皂苷对类风湿性关节炎大鼠血清细胞因子的影响

目的研究赤雹根总皂苷对类风湿性关节炎大鼠血清IL-1β、IL-6、TNF-α的影响。方法以弗氏完全佐剂诱导大鼠佐剂性关节炎,将造模成功50只大鼠随机分为模型组,赤雹根总皂苷160,80,40 mg/(kg·d)组和雷公藤12 mg/(kg·d)组,另取10只正常大鼠作正常对照。连续灌胃给药21 d。观测各组大鼠继发关节肿胀度和关节炎指数;给药后第22天,处死大鼠,用酶联免疫吸附法(ELISA)检测血清IL-1β、IL-6、TNF-α含量。结果赤雹根总皂苷能明显降低佐剂性关节炎大鼠继发侧的关节肿胀度和关节炎指数(P<0.01);与正常对照组相比,模型组大鼠血清IL-1β、IL-6、TNF-α含量明显升高(P<0.01);赤雹根总皂苷灌胃治疗后,各组大鼠血清IL-1β、IL-6、TNF-α含量较模型组明显降低(P<0.01)。结论赤雹根总皂苷对大鼠类风湿性关节炎有治疗作用,且该作用与抑制IL-1β、IL-6、TNF-α的合成有关。     

Effects of total glucosides from paeony (Paeonia lactiflora Pall) roots on experimental atherosclerosis in rats

Ethnopharmacological relevance

Total glucosides of paeony (TGP), compounds extracted from the roots of Paeonia lactiflora Pall, have been used as an anti-inflammatory drug for the treatment of rheumatoid arthritis (RA) in China. Inflammation plays a critical role in the development of atherosclerotic vascular disease. Risk of cardiovascular diseases is significantly higher in patients with RA than in normal population. It has a great significance to study the effects of TGP on atherosclerosis.

Aim of the study

To investigate the effects of TGP on atherosclerosis induced by excessive administration of vitamin D and cholesterol in rats and study the mechanisms involved.

Materials and methods

Atherosclerosis was induced by excessive administration of vitamin D and cholesterol in rats. TGP was intragastrically administered for 15 weeks. The serum concentrations of total cholesterol (TC), triglyceride (TG), low density lipoprotein-cholesterol (LDL-C) and high density lipoprotein-cholesterol (HDL-C) were measured by automatic biochemistry analyzer. Apolipoprotein A1 (ApoA1) and apolipoprotein B (ApoB) were determined by immunoturbidimetry method, tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and C-reactive protein (CRP) were measured by enzyme-linked immunosorbent assay (ELISA) method. The morphological changes of aorta were observed with optical microscopy.

Results

Compared to controls, TGP significantly lowered the serum level of TC, TG, LDL-C, ApoB, TNF-alpha, IL-6 and CRP, increased the ratios of HDL-C/LDL-C and ApoA1/ApoB, decreased the intima-media thickness (IMT) of abdominal aortal wall and improved the morphological change of the aorta.

Conclusions

TGP may attenuate the development of atherosclerotic disease. The beneficial effects are associated with its lowering blood lipids and inhibiting the expression of inflammatory cytokines.     

逐痹胶囊对大鼠佐剂性关节炎的影响

目的:观察逐痹胶囊对大鼠佐剂性关节炎(adjuvantarthritis,AA)的影响及作用机制。方法:用福氏完全佐剂(CFA)诱导大鼠AA模型,MTT法检测脾淋巴细胞增殖能力,免疫组织化学方法检测踝关节滑膜白细胞介素1(IL-1),白细胞介素6(IL-6)含量,并且测量原发及继发性关节肿胀程度。结果:生药52,26,13g·kg^-1体重灌胃大鼠,逐痹胶囊显著抑制AA大鼠原发及继发踝关节炎性肿胀;降低脾淋巴细胞增殖能力及踝关节滑膜内IL1,IL-6含量。结论:逐痹胶囊对大鼠AA有一定抑制作用。     

消风Ⅱ号胶囊对佐剂性关节炎大鼠炎性组织中前列腺素E2及一氧化氮含量的影响

目的 通过观察消风Ⅱ号胶囊对佐剂性关节炎(AA)大鼠炎性组织中前列腺素E2(PGE2)及一氧化氮(N0)含量的影响,探讨其治疗类风湿关节灸作用机理.方法 将AA大鼠随机分为模型组及消风Ⅱ号胶囊低(188 mg/kg)、高剂量组(375 mg/kg),同时设正常组.致炎第7日开始灌胃给予药物,灌胃剂量为1.0 mL/100 g体质量,模型组给予等体积的蒸馏水,连续21d.给药结束后次日处死大鼠,测量从大鼠炎性组织中PGE2及NO含量.结果 消风Ⅱ号胶囊低、高剂量组均可抑制大鼠非佐剂注射侧炎性组织中PGE2的生物合成,抑制率分别为41.2％和33.6％,抑制大鼠佐剂注射侧炎性组织中NO含量,抑制率分别为39.9％和29.7％,与模型组相比,差异有统计学意义(P＜0.001,P＜0.01).结论 消风Ⅱ号胶囊抗关节炎作用通过抑制PGE2和NO的合成或释放,使局部炎性组织中PGE2及NO含量下降而发挥作用.     

清络通痹颗粒对佐剂性关节炎大鼠滑膜细胞的影响

目的探讨清络通痹颗粒对佐剂性关节炎大鼠滑膜细胞的影响.方法建立佐剂性关节炎(AIA)大鼠模型,同时设正常对照组.自第7天开始给药,持续21 d.取滑膜,检测滑膜细胞增殖能力和滑膜细胞分泌IL-1的量.结果清络通痹颗粒7.2、14.4 g/kg可明显抑制AIA大鼠的滑膜细胞增殖反应(P＜0.01).结论一定剂量的清络通痹颗粒可抑制AIA大鼠滑膜细胞的过度增生,这与其调节与滑膜增生相关的细胞因子有关.     

Anti-inflammatory and anti-allergic properties of the essential oil and active compounds from Cordia verbenacea

The anti-inflammatory and anti-allergic effects of the essential oil of Cordia verbenacea (Boraginaceae) and some of its active compounds were evaluated. Systemic treatment with the essential oil of Cordia verbenacea (300-600mg/kg, p.o.) reduced carrageenan-induced rat paw oedema, myeloperoxidase activity and the mouse oedema elicited by carrageenan, bradykinin, substance P, histamine and platelet-activating factor. It also prevented carrageenan-evoked exudation and the neutrophil influx to the rat pleura and the neutrophil migration into carrageenan-stimulated mouse air pouches. Moreover, Cordia verbenacea oil inhibited the oedema caused by Apis mellifera venom or ovalbumin in sensitized rats and ovalbumin-evoked allergic pleurisy. The essential oil significantly decreased TNFalpha, without affecting IL-1beta production, in carrageenan-injected rat paws. Neither the PGE(2) formation after intrapleural injection of carrageenan nor the COX-1 or COX-2 activities in vitro were affected by the essential oil. Of high interest, the paw edema induced by carrageenan in mice was markedly inhibited by both sesquiterpenic compounds obtained from the essential oil: alpha-humulene and trans-caryophyllene (50mg/kg, p.o.). Collectively, the present results showed marked anti-inflammatory effects for the essential oil of Cordia verbenacea and some active compounds, probably by interfering with TNFalpha production. Cordia verbenacea essential oil or its constituents might represent new therapeutic options for the treatment of inflammatory diseases.