Acute phase reactions in Daboia siamensis venom and fraction-induced acute kidney injury: the role of oxidative stress and inflammatory pathways in in vivo rabbit and ex vivo rabbit kidney models

Abstract Background: This study examines the direct nephrotoxic effects of Daboia siamensis venom (RVV) and venom fractions in in vivo and isolated perfused kidneys (IPK) to understand the role of inflammation pathways and susceptibility to oxidative stress in venom or fraction-induced acute renal failure. Methods: We administered RVV and its venom fractions (PLA2, MP, LAAO, and PDE) to rabbits in vivo and in the IPK model. We measured oxidative stress biomarkers (SOD, CAT, GSH, and MDA) in kidney tissue, as well as inflammatory cytokines (TNF-α, IL-1β, IFN-γ, IL-4, IL-5, and IL-10), MDA and GSH levels in plasma and urine. We also calculated fractional excretion (FE) for pro-/anti-inflammatory cytokines and oxidative stress biomarkers, including the ratios of pro-/anti-inflammatory cytokines in urine after envenomation. Results: In both kidney models, significant increases in MDA, SOD, CAT, and GSH levels were observed in kidney tissues, along with elevated concentrations of MDA and GSH in plasma and urine after injecting RVV and venom fractions. Moreover, RVV injections led to progressive increases in FEMDA and decreases in FEGSH. The concentrations of IL-4, IL-5, IL-10, IFN-γ, and TNF-α in plasma increased in vivo, as well as in the urine of the IPK model, but not for IL-1β in both plasma and urine after RVV administrations. Urinary fractional excretion of TNF-α, IL-1β, IFN-γ, IL-4, IL-5, and IL-10 tended to decrease in vivo but showed elevated levels in the IPK model. A single RVV injection in vivo disrupted the balance of urinary cytokines, significantly reducing either the TNF-α/IL-10 ratio or the IFN-γ/IL-10 ratio. Conclusion: RVV induces renal tubular toxicity by increasing oxidative stress production and elevating inflammatory cytokines in urine. During the acute phase of acute kidney injury, the balance of urine cytokines shifts toward anti-inflammatory dominance within the first two hours post-RVV and venom fractions.

Clinical and laboratory features distinguishing between Deinagkistrodon acutus and Daboia siamensis envenomation

There are 6 species of venomous snakes in Taiwan. Two of them, Deinagkistrodon acutus (D. acutus) and Daboia siamensis (D. siamensis), can cause significant coagulopathy. However, a significant proportion of patients with snakebites cannot identify the correct snake species after envenomation, which hampers the application of antivenom. Hence, the differential diagnosis between the two snakebites by clinical presentations is important. This study aims to compare their clinical and laboratory features for the purpose of differential diagnosis between the two snakebites. Methods: We retrospectively reviewed the medical records of patients who arrived at the emergency department due to D. acutus or D. siamensis envenomation, between 2003 and 2016, in one medical center in eastern Taiwan. Since these snakebites are rare, we also included 3 cases reported from another hospital in central Taiwan. Results: In total, 15 patients bitten by D. acutus and 12 patients by D. siamensis were analyzed. Hemorrhagic bulla formation and the need for surgical intervention only presented for D. acutus envenomation cases (Both 53.3% vs. 0.0%, P= 0.003). As to laboratory features, lower platelet counts (20.0 × 103/µL [interquartile range, 14-66 × 103/µL] vs. 149.0 × 103/µL [102.3-274.3 × 103/µL], P = 0.001), lower D-dimer level (1423.4 µg/L [713.4-4212.3 µg/L] vs. 12,500.0 µg/L [2351.4-200,000 µg/L], P = 0.008), higher proportion of patients with moderate-to-severe thrombocytopenia (platelet count < 100 × 103/µL) (80% vs. 16.7%, odds ratio (OR) = 20.0, 95% CI, 2.77-144.31; P = 0.002), and lower proportion of patients with extremely high D-dimer (> 5000 ng/mL) (16.7% vs. 66.7%, adjusted OR = 0.1 (95% CI, 0.01-0.69; P = 0.036) were found among cases of D. acutus envenomation compared to D. siamensis envenomation. The combination of hemorrhagic bulla, thrombocytopenia, and a lack of extremely high D-dimer had good discriminatory power (area under the curve (AUC) = 0.965; 95% CI, 0.904-1.00) for distinguishing D. acutus from D. siamensis envenomation. Conclusions: The presentation of moderate to severe thrombocytopenia (platelet count < 100 × 103/µL) and hemorrhagic bulla formation may indicate D. acutus envenomation. However, the envenomed patient with extremely high D-dimer levels may indicate a D. siamensis envenomation. These findings may help diagnose and select the right antivenom in patients with unknown snakebites who present significant coagulopathy.(AU)

Anticancer properties of phospholipase A2 fromDaboia siamensis venom on human skin melanoma cells

Background Phospholipase A2 (PLA2) is a major component of theDaboia siamensis venom, which is able to hydrolyse the membrane of various cells. For this reason, the activity of PLA2was investigated regarding its pharmaceutical properties. This study was conducted to explore the pharmacological properties of a PLA2from Daboia siamensis (dssPLA2) venom on human skin melanoma cell line (SK-MEL-28). Methods dssPLA2 was isolated by ion exchange and gel filtration columns. Various concentrations of dssPLA2were investigated for cytotoxic activity and inhibition of migration on SK-MEL-28 cells. Cell death analysis, mRNA expression levels of Notch I-III and BRAF V600E genes were also determined. Results dssPLA2 exhibited cytotoxicity on SK-MEL-28 for 24 and 72 h as compared with untreated cells. However, it had no toxic effects on CCD-1064sk cells under the same conditions. dssPLA2 (0.25 and 0.5 μg/mL) induced 17.16 and 30.60 % of apoptosis, while activated 6.53 and 7.05 % of necrotic cells. dssPLA2 at 0.25, 0.5, 1 and 2 μg/mL could inhibit migration on SK-MEL-28 cells for 24 h by 31.06, 41.66, 50 and 68.75 %, respectively. The action of dssPLA2 significantly reduced the levels of Notch I and BRAF V600E genes expression on SK-MEL-28 cells compared with untreated cells at 72 h. Conclusions This study indicates that dssPLA2 had potential effects of apoptosis, necrosis, cytotoxicity and inhibition of migration on SK-MEL-28 cells. dssPLA2 could possibly be a selective agent that targets cancer cells without affecting normal cells.(AU)

Anticancer properties of phospholipase A2 fromDaboia siamensis venom on human skin melanoma cells

Phospholipase A2 (PLA2) is a major component of theDaboia siamensis venom, which is able to hydrolyse the membrane of various cells. For this reason, the activity of PLA2was investigated regarding its pharmaceutical properties. This study was conducted to explore the pharmacological properties of a PLA2from Daboia siamensis (dssPLA2) venom on human skin melanoma cell line (SK-MEL-28). Methods dssPLA2 was isolated by ion exchange and gel filtration columns. Various concentrations of dssPLA2were investigated for cytotoxic activity and inhibition of migration on SK-MEL-28 cells. Cell death analysis, mRNA expression levels of Notch I-III and BRAF V600E genes were also determined. Results dssPLA2 exhibited cytotoxicity on SK-MEL-28 for 24 and 72 h as compared with untreated cells. However, it had no toxic effects on CCD-1064sk cells under the same conditions. dssPLA2 (0.25 and 0.5 g/mL) induced 17.16 and 30.60 % of apoptosis, while activated 6.53 and 7.05 % of necrotic cells. dssPLA2 at 0.25, 0.5, 1 and 2 g/mL could inhibit migration on SK-MEL-28 cells for 24 h by 31.06, 41.66, 50 and 68.75 %, respectively. The action of dssPLA2 significantly reduced the levels of Notch I and BRAF V600E genes expression on SK-MEL-28 cells compared with untreated cells at 72 h. Conclusions This study indicates that dssPLA2 had potential effects of apoptosis, necrosis, cytotoxicity and inhibition of migration on SK-MEL-28 cells. dssPLA2 could possibly be a selective agent that targets cancer cells without affecting normal cells.