Urinary proteomics reveals biological processes related to acute kidney injury in Bothrops atrox envenomings

Abstract

Acute kidney injury (AKI) is a critical systemic complication caused by Bothrops envenoming, a neglected health problem in the Brazilian Amazon. Understanding the underlying mechanisms leading to AKI is crucial for effectively mitigating the burden of this complication. This study aimed to characterize the urinary protein profile of Bothrops atrox snakebite victims who developed AKI. We analyzed three groups of samples collected on admission: healthy subjects (controls, n = 10), snakebite victims who developed AKI (AKI, n = 10), and those who did not evolve to AKI (No-AKI, n = 10). Using liquid-chromatography tandem mass spectrometry, we identified and quantified (label-free) 1190 proteins. A panel of 65 proteins was identified exclusively in the urine of snakebite victims, with 32 exclusives to the AKI condition. Proteins more abundant or exclusive in AKI's urine were associated with acute phase response, endopeptidase inhibition, complement cascade, and inflammation. Notable proteins include serotransferrin, SERPINA-1, alpha-1B-glycoprotein, and NHL repeat-containing protein 3. Furthermore, evaluating previously reported biomarkers candidates for AKI and renal injury, we found retinol-binding protein, beta-2-microglobulin, cystatin-C, and hepcidin to be significant in cases of AKI induced by Bothrops envenoming. This work sheds light on physiological disturbances caused by Bothrops envenoming, highlighting potential biological processes contributing to AKI. Such insights may aid in better understanding and managing this life-threatening complication.

Fig 1. Venn Diagram showing the common and exclusive proteins identified in the urine of B. atrox snakebite victims.

Fig 2. Biological pathways of proteins exclusively found in the urine of Bothrops snakebite victims, shared by both clinical outcomes.

Fig 3. Distribution of the 10 most abundant proteins exclusive to Bothrops snakebite victims’ urine between No-AKI and AKI patients.

Protein listed: Apolipoprotein C-II (APOC2), Myoglobin (MB), Carboxypeptidase (B2CPB2), Dermcidin (DCD), Component C8 gamma chain (C8G), SH3 domain-binding glutamic acid-rich-like protein (SH3BGRL), Insulin-like growth factor binding protein 6 (IGFBP6), Osteoglycin (OGN), Serglycin (SRGN), Complement C2 (C2), and Coagulation factor IX (F9).

Fig 4. Protein-protein interactions within the group of proteins exclusively found in the urine of Bothrops snakebite patients with acute kidney injury.

Colors represent the associated biological processes: Red–Acute phase response (GO:0006953); Green–Mixed processes, including complement and coagulation cascades, and lipoprotein particles (CL:19706); Blue–Endopeptidase inhibitor activity (GO:00048630).

Fig 5. Differential abundance of proteins in the urine of Bothrops snakebite victims with distinct outcomes regarding acute kidney injury.

Each protein was represented in the graph as a dot, plotted based on the (-"Log")2(" p-value) on the x-axis and Log2 (Fold change) on the y-axis. Red dots indicate proteins that did not meet the q-value cutoff or the fold change cutoff. Green dots represent proteins that met the fold change but not the q-value. Blue dots signify proteins that satisfied both fold change and q-value (0.05). The proteins plotted include alpha-1B-glycoprotein (A1BG), NHL repeat-containing protein 3 (NHLRC3), serotransferrin (TF), alpha-trypsin 1 (SERPINA1), molybdopterin synthase sulfur carrier subunit (MOCS2), fructose-bisphosphate aldolase B (ALDOB), neurosecretory protein VGF (VGF), putative phospholipase B-like 2 (PLBD2), lysosome-associated membrane glycoprotein 2 (LAMP2), tumor necrosis factor receptor superfamily member 19L (RELT), coagulation factor V (F5) and filaggrin (FLG).

Fig 6. Distribution of literature-appointed biomarkers for kidney injury in Bothrops snakebite urine samples.

The proteins listed are retinol-binding protein (RBP4), beta-2-microglobulin (B2M), alpha-1-acid glycoprotein 1 (ORM1), cystatin-C (CST3), alpha-2-HS-glycoprotein (AHSG), hepcidin (HAMP), clusterin (CLU) and fatty acid-binding protein (L-FABP), neutrophil gelatinase-associated lipocalin (NGAL), insulin-like growth factor-binding protein 7 (IGFBP7), protein S100-A9 (S100A9), vascular cell adhesion protein 1 (VCAM1) and protein S100-A8 (S100A8).