ABSTRACT
Snake envenomation (Bothrops genus) is common in tropical countries and acute kidney injury is one of the complications observed in Bothrops snakebite with relevant morbidity and mortality. Here, we showed that Bothropoides pauloensis venom (BpV) decreased cell viability (IC50 of 7.5 µg/mL). Flow cytometry with annexin V and propidium iodide showed that cell death occurred predominantly by apoptosis and late apoptosis, through caspases 3 and 7 activation, mitochondrial membrane potential collapse and ROS overproduction. BpV reduced perfusion pressure, renal vascular resistance, urinary flow, glomerular filtration rate, percentage of sodium, chloride or potassium tubular transportation. These findings demonstrated that BpV cytotoxicity on renal epithelial cells might be responsible for the nephrotoxicity observed in isolated kidney.
Subject(s)
Bothrops , Crotalid Venoms/toxicity , Kidney Tubules/drug effects , Kidney/drug effects , Animals , Apoptosis/drug effects , Caspase 3/metabolism , Caspase 7/metabolism , Cell Survival/drug effects , Dogs , Epithelial Cells/drug effects , Flow Cytometry , Glomerular Filtration Rate/drug effects , In Vitro Techniques , Madin Darby Canine Kidney Cells , Male , Membrane Potential, Mitochondrial/drug effects , Rats, Wistar , Reactive Oxygen Species/metabolism , Toxicity Tests , Vascular Resistance/drug effectsABSTRACT
Mammalian natriuretic peptides (NPs) have been extensively investigated for use as therapeutic agents in the treatment of cardiovascular diseases. Here, we describe the isolation, sequencing and tridimensional homology modeling of the first C-type natriuretic peptide isolated from scorpion venom. In addition, its effects on the renal function of rats and on the mRNA expression of natriuretic peptide receptors in the kidneys are delineated. Fractionation of Tityus serrulatus venom using chromatographic techniques yielded a peptide with a molecular mass of 2190.64 Da, which exhibited the pattern of disulfide bridges that is characteristic of a C-type NP (TsNP, T. serrulatus Natriuretic Peptide). In the isolated perfused rat kidney assay, treatment with two concentrations of TsNP (0.03 and 0.1 µg/mL) increased the perfusion pressure, glomerular filtration rate and urinary flow. After 60 min of treatment at both concentrations, the percentages of sodium, potassium and chloride transport were decreased, and the urinary cGMP concentration was elevated. Natriuretic peptide receptor-A (NPR-A) mRNA expression was down regulated in the kidneys treated with both concentrations of TsNP, whereas NPR-B, NPR-C and CG-C mRNAs were up regulated at the 0.1 µg/mL concentration. In conclusion, this work describes the isolation and modeling of the first natriuretic peptide isolated from scorpion venom. In addition, examinations of the renal actions of TsNP indicate that its effects may be related to the activation of NPR-B, NPR-C and GC-C.
Subject(s)
Kidney/drug effects , Natriuretic Peptide, C-Type/isolation & purification , Scorpion Venoms/isolation & purification , Scorpions/chemistry , Amino Acid Sequence , Animals , Brazil , Cyclic GMP/genetics , Cyclic GMP/metabolism , Down-Regulation , Glomerular Filtration Rate , Kidney/metabolism , Male , Molecular Sequence Data , Natriuretic Peptide, C-Type/chemistry , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Rats, Wistar , Receptors, Atrial Natriuretic Factor/genetics , Receptors, Atrial Natriuretic Factor/metabolism , Scorpion Venoms/chemistry , Sequence Alignment , Up-RegulationABSTRACT
In a variety of animal models, uroguanylin causes diuresis, natriuresis and kaliuresis and is found in larger concentrations in the urine compared to controls after oral salt intake or in conditions of excess salt and fluid retention. It has been proposed that uroguanylin functions as an intestinal natriuretic hormone following intake of meals high in salt content. In the present work, we examined if 10 days of salt ingestion resulted in an enhanced response to uroguanylin in the isolated perfused rat kidney. Rats were given normal water, 1% NaCl (HS1%), or 2% NaCl (HS2%) for 10 days, at which time the right kidneys were surgically removed and perfused with a modified Krebs-Henseleit solution for 30 min. After a 30-min control period, the kidneys were perfused with a modified Krebs-Henseleit solution containing 0.06 microM uroguanylin for an additional 90 min. Compared to vehicle-matched time controls, 0.06 microM uroguanylin perfusion of kidneys from rats maintained on HS2% resulted in a significantly increased urine flow (UF; from 0.17+/-0.01 to 0.23+/-0.01, after 60 min, n=6, P<0.05), fractional Na(+) excretion (%E(Na+); from 16.6+/-0.7 to 30+/-2, after 60 min, n=6, P<0.05), fractional K(+) excretion (%E(K+); from 20.5+/-0.58 to 37.4+/-2.1, after 60 min, n=6, P<0.05), and fractional Cl(-) excretion increased from 18.16+/-0.52 to 35.2+/-2.0 at 60 min, n=6, P<0.05. With the exception of a significant increase in the %E(K)(+), no other effect was observed in the kidneys from the rats maintained on HS1%, and no significant effects were seen in those that were maintained on normal water. The effect of a higher dose (0.6 microM) of uroguanylin on urinary flow, sodium or potassium excretion was also significantly increased by 2% NaCl (HS2%) treatment (P<0.05). We also observed an expressive upregulation of the GC-C and a slight downregulation of the GC-A receptor in high-salt treated rats. These data demonstrate that prolonged salt ingestion primes the kidney to enhanced renal responses to uroguanylin.
