【作者】 胡一梅；

【导师】 艾儒棣；

【作者基本信息】 成都中医药大学 ， 中医外科学， 2013， 博士

【摘要】 目的：观察马齿苋提取物对湿疹大鼠皮肤的治疗效果,探讨单品马齿苋的药效以及促进湿疹病变部位修复的作用机理。方法：1.SD大鼠70只,按体重随机分为7个组,每组10只,分别作为正常组、模型组、Caspase-14基因敲除组、阳性组、马齿苋提取物高、中、低剂量组。除正常组外,用电推剪及电剃须刀去除腹部毛发,面积约2cm×3cm,并涂5%的2,4-二硝基氯苯(DNCB)30μl于皮肤首次致敏；第5天,在右背部2cm×2cm剃毛区涂0.2%DNCB50μl进行二次抗原攻击,左背部对称的剃毛区不用药物作为对照部位。以后每隔3d激发1次,连续3次。于第9天观察大鼠右背部较左背部明显出现红斑,水肿,丘疹,糜烂或渗出,表明造模成功。实验第21天断颈处死,剪取造模皮肤,用分析天平称重测量湿疹大鼠皮肤肿胀度；QRT-PCR检测各实验组大鼠皮肤Caspase-14和Filaggrin基因表达；免疫组化检测TNF-α、IL-22、IL-4、IFN-γ积分光密度。2.另取大鼠50只,随机分为5组：正常对照组,阳性对照组,马齿苋低剂量组、中剂量组、高剂量组。各组按规定的剂量外用给药。实验当日,用0号砂纸擦伤大鼠剃毛处,局部再涂各组相应药物1次,末次涂药后10min,开始于创面处滴0.01%磷酸组胺每只0.05mL,此后每隔3min依0.01%,0.02%,0.03%,0.04%…浓度递增,每一浓度滴加后观察3min内大鼠是否回头舔右后足背,按此时磷酸组胺总量计算各组大鼠的致痒阂。结果：1.与空白对照组相比,高剂量组小鼠皮肤肿胀度明显小于空白对照组(P<0.01),中剂量组、阳性药物组小鼠皮肤肿胀度也小于模型组(P<0.05)。2.与正常组比较,模型组和Caspase-14基因敲除组大鼠皮肤组织细胞中Caspase-14和Filaggrin基因的2-ΔΔCT值极显著减小(P<0.01),与模型组和Caspase-14基因敲除组比较,马齿苋三个剂量组大鼠皮肤组织细胞中Caspase-14和Filaggrin基因的2-ΔΔCT值极显著增大(P<0.01)。3.与正常组比较,模型组大鼠皮肤组织TNF-α免疫组织化学检测积分光密度(IOD)极显著增大(P<0.01)；模型组大鼠皮肤组织IL-22免疫组织化学检测积分光密度(IOD)极显著增大(P<0.01)；模型组大鼠皮肤组织IL-4免疫组织化学检测积分光密度(IOD)极显著增大(P<0.01)；模型组大鼠皮肤组织IFN-γ免疫组织化学检测积分光密度(IOD)显著减小(P<0.05)。4.磷酸组胺对不同剂量组的致痒阈记录显示,马齿苋提取物中剂量组的致痒阈高于阳性对照组(P<0.05),高剂量组明显高于阳性对照组(P<0.05)。结论：马齿苋提取物可以使血管收缩,抑制毛细血管通透性,抑制炎性肿胀,从而减轻湿疹局部的瘙痒、水肿症状。马齿苋提取物还可显著上调湿疹动物皮肤屏障系统中Caspase-14和Filaggrin基因的表达,提示马齿苋通过促进Caspase-14基因的表达而加速ProFilaggrin去磷酸化成为Filaggrin,进而加速Filaggrin水解成各种天然保湿因子,维护皮肤屏障功能。同时,马齿苋提取液通过降低TNF-a和IL-22在湿疹发病过程中的表达,提高IFN-γ的分泌抑制IL-4的表达,增强了机体的细胞免疫功能。更多还原

【Abstract】 Objective To investigate the impact of the Purslane on the eczema skin, explore the effect and mechanism of single Purslane on the repairing lesion site of eczemaMethods1.70SD rats were randomly divided into control group, model group, Caspase-14gene knockout group, positive group and high, medium, low dose of Purslane group. Except The normal group, the hair of rat’s abdomen were shaved by electric clippers and electric shaver, the area is2cm X3cm, then,2,4-dinitrochlorobenzene was used to make rats acur first sensitization. After5days,2,4-dinitrochlorobenzene was used again on the right back of rat, the area is2cm X2cm, the left back as the control site. To use excitation on rat every3days,3times in a row. At the9th day, the right back has erythema, edema, erosion or exudative papules, it showed eczema model was made successful.21days later, all animals were sacrificed, eczema skins were taken out, analytical balance was used to measure swelling of eczema skin, QRT-PCR was used to detect the expression of Caspase-14and Filaggrin gene in each experimental group rat skin. Immunohistochemical detection was ued to measure TNF-α、IL-22、IL-4. IFN-γ2.50SD rats were randomly divided into control group, positive group and high, medium, low dose of Purslane group. The recommended external dosage were given. At the experimental day,0standard Sandpaper was used to scrape the right leg of rat, then, all drug were applied again, after10minutes, the0.01%histamine phosphate was applied on the scraping area, more and more concentrated histamine phosphate were used every3minutes,0.01%,0.02%,0.03%,0.04%...The concentration is the itch threshold when the rat lick the right foot back. Results:1. Compared with the blank control group, swelling of high dose group was significantly less than the control group (P<0.01), The medium dose group, positive drug group is lower than model group (P<0.05).2. Compared with the normal group, Caspase-14and Filaggrin cells’ value in skin tissue of model group and knockout was significantly decreased (P<0.01). Compared with the medel group Caspase-14gene knockout group, Caspase-14and Filaggrin gene in the three dose of Purslane groups was significantly increased (P<0.01).3. Compared with the normal group, IOD of TNF-α, IL-22, IL-4of model group was significantly increased (P<0.01), the IOD of IFN-γ was significantly decreased,(P<0.01); but the IOD of TNF-α, IL-22, IL-4of high and medium group was decreased (P<0.05), IFN-γ was decreased,(P<0.05);4. The histamine phosphate in different dose groups of itching threshold records showed the medium group was higher than positive group(P<0.05), the high group was significantly higher than positive group(P<0.01),Conclusion Purslane extract can constrict the blood vessels, inhibit capillary permeability, inhibit the inflammatory swelling, reduce the itching and swelling symptoms of eczema edema. Purslane extract can raise the expression of Caspase-14and Filaggrin of skin barrier system, it means Purslane extract can improve gene expressionthe of Caspase-14to accelerate dephosphorylation of ProFilaggrin to transformed into filaggrin, accelerating the hydrolysis of Filaggrin into a variety of natural moisturizing factor to enhance skin protective function. At the same time, Purslane extract decrease the expression of TNF-a and IL-22in the eczema, and increase the secretion of IFN-γ to inhibit IL-4, then, the cellular immune function was enhance.更多还原