Laboratório de Imunoparasitologia Dr. Mário Endsfeldz Camargo, Instituto de Ciências Biomédicas, Universidade Federal de Uberlândia, 38400-902 Uberlândia, MG, Brazil.
Laboratório de Imunologia, Instituto de Ciências Biológicas e Naturais, Universidade Federal do Triângulo Mineiro, 38025-180 Uberaba, MG, Brazil.
Mediators Inflamm. 2019 Jul 29;2019:5195134. doi: 10.1155/2019/5195134. eCollection 2019.
It has been described that the metalloprotease BmooMP-alpha-I purified from snake venom is able to hydrolyze the TNF molecule. However, this observation has been based mainly on investigation, in addition to molecular modeling and docking approaches. Considering that there is no in vivo study to demonstrate the biological effects of this enzyme, the major aim to the present work was to investigate whether the BmooMP-alpha-I has any anti-inflammatory efficacy by setting up a murine experimental design of colitis induced by dextran sulfate sodium (DSS). For this purpose, C57BL/6 mice were divided into six groups, as follows: (i) animals without intestinal inflammation, (ii) animals without intestinal inflammation treated with BmooMP-alpha-I (50 g/animal/day), and (iii) animals with intestinal inflammation induced by 3% of DSS, (iv) mice with intestinal inflammation induced by DSS and treated with BmooMP-alpha-I enzyme at the 50, 25, or 12.5 g/animal/day dosages by intraperitoneal route. Clinical signs of colitis were observed daily for calculating the morbidity scores, cytokine measurements, and histological features. We observed that the animals treated with different doses of the enzyme presented a remarkable improvement of colitis signs, as confirmed by a significant increase of the intestine length in comparison to the DSS group. Also, no difference was observed between the groups treated with the enzyme or vehicle, as the colon length of these animals was slightly lower than that of the group of healthy animals, without induction of intestinal inflammation. The cytokine quantification in supernatants of intestinal tissue homogenates showed a significant reduction of 38% in IFN-gamma levels, when the animals were treated with 50 g of the BmooMP-alpha-I compared to the animals receiving DSS only. A significant reduction of 39% in TNF levels was also observed in all doses of treatment with BmooMP-alpha-I, in addition to a significant reduction of 35% in the amount of IL-12p40. Histological examinations revealed that the BmooMP-alpha-I 50 g treated group preserved colon architecture and goblet cells and reduced the ulcer area, when compared with DSS mice, which showed typical inflammatory changes in tissue architecture, such as ulceration, crypt dilation, loss of tissue architecture, and goblet cell depletion, accompanied by a significant cell infiltration. In conclusion, our results suggest that the improvement of clinical scores and histological findings related to BmooMP-alpha-I treatment in this experimental model could be attributed to the metalloprotease ability to modulate cytokine production locally at the inflamed intestine. These findings highlight the potential anti-inflammatory role and effectiveness of this enzyme as a therapeutic alternative in this type of immunopathological condition.
据描述，从蛇毒液中纯化的金属蛋白酶 BmooMP-alpha-I 能够水解 TNF 分子。然而，这一观察结果主要基于调查，此外还有分子建模和对接方法。考虑到没有体内研究来证明这种酶的生物学效应，本工作的主要目的是通过建立葡聚糖硫酸钠（DSS）诱导的结肠炎的小鼠实验设计来研究 BmooMP-alpha-I 是否具有抗炎疗效。为此，将 C57BL/6 小鼠分为六组，如下所示：（i）无肠道炎症的动物，（ii）无肠道炎症且用 BmooMP-alpha-I（50 g/动物/天）处理的动物，和（iii）用 3%的 DSS 诱导肠道炎症的动物，（iv）用 DSS 诱导肠道炎症且用 50、25 或 12.5 g/动物/天的剂量通过腹腔途径用 BmooMP-alpha-I 酶处理的小鼠。每天观察结肠炎的临床症状以计算发病率评分、细胞因子测量和组织学特征。我们观察到，用不同剂量的酶处理的动物表现出结肠炎迹象的显著改善，这通过与 DSS 组相比肠长度的显著增加得到证实。此外，用酶或载体处理的动物之间没有观察到差异，因为这些动物的结肠长度略低于没有诱导肠道炎症的健康动物的结肠长度。肠组织匀浆上清液中细胞因子的定量分析显示，与仅接受 DSS 的动物相比，用 50 g 的 BmooMP-alpha-I 处理的动物 IFN-γ水平降低了 38%。在用 BmooMP-alpha-I 处理的所有剂量中，TNF 水平也降低了 39%，此外，IL-12p40 的量降低了 35%。组织学检查显示，与 DSS 小鼠相比，50 g 的 BmooMP-alpha-I 处理组保留了结肠结构和杯状细胞，并减少了溃疡面积，DSS 小鼠的组织结构显示出典型的炎症变化，如溃疡、隐窝扩张、组织结构丧失和杯状细胞耗竭，伴有显著的细胞浸润。总之，我们的结果表明，在这种实验模型中，BmooMP-alpha-I 治疗与临床评分和组织学发现的改善可能归因于金属蛋白酶局部调节炎症肠道中细胞因子产生的能力。这些发现强调了这种酶作为这种免疫病理状况的治疗替代方法的潜在抗炎作用和有效性。
