Barbosa P S F, Martins A M C, Havt A, Toyama Daniela O, Evangelista J S A M, Ferreira D P P, Joazeiro P P, Beriam Luis O S, Toyama Marcos H, Fonteles M C, Monteiro H S A
Institute of Biomedicine and Clinical Research Unit, Department of Physiology and Pharmacology, Faculty of Medicine, Federal University of Ceará, Fortaleza, Ceará, Brazil.
Toxicon. 2005 Sep 15;46(4):376-86. doi: 10.1016/j.toxicon.2005.04.024.
Bothrops jararacussu myotoxin I (BthTx-I; Lys 49) and II (BthTX-II; Asp 49) were purified by ion-exchange chromatography and reverse phase HPLC. In this work we used the isolated perfused rat kidney method to evaluate the renal effects of B. jararacussu myotoxins I (Lys49 PLA2) and II (Asp49 PLA2) and their possible blockage by indomethacin. BthTX-I (5 microg/ml) and BthTX-II (5 microg/ml) increased perfusion pressure (PP; ct120=110.28+/-3.70 mmHg; BthTX I=171.28+/-6.30mmHg; BthTX II=175.50+/-7.20mmHg), renal vascular resistance (RVR; ct120=5.49+/-0.54 mmHg/ml.g(-1)min(-1); BthTX I=8.62+/-0.37mmHg/ml g(-1)min(-1); BthTX II=8.9+/-0.36mmHg/ml g(-1)min(-1)), urinary flow (UF; ct(120)=0.14+/-0.01ml g(-1)min(-1); BthTX I=0.32+/-0.05ml g(-1)min(-1); BthTX II=0.37+/-0.01ml g(-1)min(-1)) and glomerular filtration rate (GFR; ct120=0.72+/-0.10 ml g(-1)min(-1); BthTX I=0.85+/-0.13ml g(-1)min(-1); BthTX II=1.22+/-0.28ml g(-1)min(-1)). In contrast decreased the percent of sodium tubular transport (%TNa(+); ct(120)=79,76+/-0.56; BthTX I=62.23+/-4.12*; BthTX II=70.96+/-2.93*) and percent of potassium tubular transport (%TK(+);ct120=66.80+/-3.69; BthTX I=55.76+/-5.57*; BthTX II=50.86+/-6.16*). Indomethacin antagonized the vascular, glomerular and tubular effects promoted by BthTX I and it's partially blocked the effects of BthTX II. In this work also evaluated the antibacterial effects of BthTx-I and BthTx-II against Xanthomonas axonopodis. pv. passiflorae (Gram-negative bacteria) and we observed that both PLA2 showed antibacterial activity. Also we observed that proteins Also we observed that proteins chemically modified with 4-bromophenacyl bromide (rho-BPB) decrease significantly the antibacterial effect of both PLA2. In conclusion, BthTx I and BthTX II caused renal alteration and presented activity antimicrobial. The indomethacin was able to antagonize totally the renal effects induced by BthTx I and partially the effects promoted by BthTx II, suggesting involvement of inflammatory mediators in the renal effects caused by myotoxins. In the other hand, other effects could be independently of the enzymatic activity of the BthTX II and the C-terminal domain could be involved in both effects promoted for PLA2.
矛头蝮蛇毒肌毒素I（BthTx-I；赖氨酸49）和II（BthTX-II；天冬氨酸49）通过离子交换色谱法和反相高效液相色谱法进行纯化。在本研究中，我们采用离体灌注大鼠肾脏方法，评估矛头蝮蛇毒肌毒素I（赖氨酸49磷脂酶A2）和II（天冬氨酸49磷脂酶A2）对肾脏的影响以及吲哚美辛对其可能的阻断作用。BthTX-I（5微克/毫升）和BthTX-II（5微克/毫升）可增加灌注压（PP；对照120=110.28±3.70毫米汞柱；BthTX I=171.28±6.30毫米汞柱；BthTX II=175.50±7.20毫米汞柱）、肾血管阻力（RVR；对照120=5.49±0.54毫米汞柱/毫升·克⁻¹·分钟⁻¹；BthTX I=8.62±0.37毫米汞柱/毫升·克⁻¹·分钟⁻¹；BthTX II=8.9±0.36毫米汞柱/毫升·克⁻¹·分钟⁻¹）、尿流（UF；对照(120)=0.14±0.01毫升·克⁻¹·分钟⁻¹；BthTX I=0.32±0.05毫升·克⁻¹·分钟⁻¹；BthTX II=0.37±0.01毫升·克⁻¹·分钟⁻¹）和肾小球滤过率（GFR；对照120=0.72±0.10毫升·克⁻¹·分钟⁻¹；BthTX I=0.85±0.13毫升·克⁻¹·分钟⁻¹；BthTX II=1.22±0.28毫升·克⁻¹·分钟⁻¹）。相反，肾小管钠转运百分比（%TNa⁺；对照(120)=79.76±0.56；BthTX I=62.23±4.12*；BthTX II=70.96±2.93*）和肾小管钾转运百分比（%TK⁺；对照120=66.80±3.69；BthTX I=55.76±5.57*；BthTX II=50.86±6.16*）降低。吲哚美辛可拮抗BthTX I促进的血管、肾小球和肾小管效应，并部分阻断BthTX II的效应。在本研究中，还评估了BthTx-I和BthTx-II对西番莲黄单胞菌（革兰氏阴性菌）的抗菌作用，我们观察到两种磷脂酶A2均具有抗菌活性。此外，我们还观察到用4-溴苯甲酰溴（ρ-BPB）化学修饰的蛋白质可显著降低两种磷脂酶A2的抗菌效果。总之，BthTx I和BthTX II可引起肾脏改变并具有抗菌活性。吲哚美辛能够完全拮抗BthTx I诱导的肾脏效应，并部分拮抗BthTx II促进的效应，提示炎症介质参与了肌毒素引起的肾脏效应。另一方面，其他效应可能与BthTX II的酶活性无关，且C末端结构域可能参与了磷脂酶A2促进的两种效应。
