Age-related Variation in Snake Venom: Evidence from Two Snakes (Naja atra and Deinagkistrodon acutus) in Southeastern China

In this study we explored electrophoretic profiles, enzymatic activities and immunoreactivity of neonate and adult venoms from two snakes （Naja atra and Deinagkistrodon acutus） coexisting in southeastern China. Age-related variation in electrophoretic profiles was found in both species and proteolytic and fibrinogenolytic activity was higher in neonate than adult venoms. Neonate D. acutus venom had higher 5＇ nucleotidase, PLA2, hyaluronidase and gelatinolytie activity, but lower esterolytic activity, than adult venom. Neonate and adult D. acutus venoms showed identical phosphomonoesterase, LAO and fibrinolytic activities. Neonate N. atra venom had higher phosphomonoesterase and LAO activity, but lower 5＇ nucleotidase, PLA2, hyaluronidase and Ache activities than adult venom. Neonate and adult N. atra venoms showed similar gelatinolytic activity. Further, age-dependent immunoreactivity was found in both species, and cross-reactions between homologous venoms and antiserums were closely related to venom composition. We speculate that age-related variation in venom characteristics is possibly driven by evolutionary forces associated with ontogenetic shifts in dietary habits, competition and predation pressure.

Increased Efficacy of Antivenom Combined with Hyperbaric Oxygen on Deinagkistrodon acutus Envenomation in Adult Rats

Background：Snakebites are a neglected threat to global human health with a high morbidity rate.The present study explored the efficacy of antivenom with hyperbaric oxygen （HBO） intervention on snakebites,which could provide the experimental basis for clinical adjuvant therapy.Methods：Male Sprague-Dawley rats （n =96） were randomized into four groups：the poison model was established by injecting Deinagkistrodon acutus （D.acutus） venom （0.8 LD50） via the caudal vein;the antivenom group was injected immediately with specific antivenom via the caudal vein after successful establishment of the envenomation model;and the antivenom ＋ HBO group was exposed to HBO environment for 1 h once at predetermined periods of 0 h,4 h,12 h,and 23 h after antivenin administration.Each HBO time point had six rats;the control group was left untreated.The rats in the experimental group were euthanized at the corresponding time points after HBO therapy,and brain tissue and blood were harvested immediately.Hematoxylin and eosin （H＆E） staining was used to investigate the pathological changes in the rat brain.Immunohistochemistry （IHC）,real-time polymerase chain reaction （PCR）,and Western blotting were used to detect the expression ofNestin mRNA and protein in the subventricular zone （SVZ） of the brain.The levels of coagulation function （prothrombin time,activated partial thromboplastin time [APTT],and fibrinogen） and oxidation/antioxidation index （malondialdehyde [MDA] and superoxide dismutase [SOD]） were analyzed.Data were analyzed using one-way analysis of variance.Results：The brain tissue from rats in the poison model was observed for pathological changes using H＆E staining.Tissues showed edema,decreased cell number,and disordered arrangement in the SVZ in the snake venom group.The antivenom-HBO intervention significantly alleviated these observations and was more prominent in the antivenom ＋ HBO group.The serum levels of SOD and MDA in the snake venom group were increased and the antivenom-HBO intervention further increased the SOD levels but significantly decreased the MDA levels;however,this was enhanced within 1 h after HBO administration （MDA：F=5.540,P=0.008,SOD：F=7.361,P =0.000）.Activated partial thromboplastin time （APTT） was significantly abnormal after venom administration but improved after antivenom and was even more significant in the antivenom ＋ HBO group 5 h after envenomation （F =25.430,P =0.000）.Only a few nestin-positive cells were observed in the envenomation model.The expression levels were significant in the antivenom and antivenom ＋ HBO groups within 1 and 5 h after envenomation and were more significant in the antivenom ＋ HBO group as determined by IHC,real-time PCR,and Western blotting （P 〈 0.05）.D.acutus envenomation has neurotoxic effects in the brain of rats.Conclusions：Antivenin and HBO,respectively,induced a neuroprotective effect after D.acutus envenomation by attenuating brain edema,upregulating nestin expression in SVZ,and improving coagulopathy and oxidative stress.The intervention efficacy of antivenom with HBO was maximum within 5 h after envenomation and was more efficacious than antivenom alone.

鱼腥草地上部分和根部提取物抑制尖吻蝮蛇毒的比较

研究鱼腥草Houttuynia cordata Thunb地上部分提取物和根部提取物对尖吻蝮Deinagkistrodon acutus蛇毒抑制作用。采用体外实验检测两种提取物对尖吻蝮蛇毒中主要酶类(蛋白水解酶、磷脂酶A2(PLA 2)、透明质酸酶等)的抑制作用;采用体内实验检测其对尖吻蝮蛇毒诱导体内毒性(出血、水肿、组织坏死和致死毒性)的抑制作用。鱼腥草地上部分提取物对尖吻蝮蛇毒蛋白水解酶、磷脂酶A2和透明质酸酶活性具有显著的抑制作用,同时能有效抑制促凝活性和纤维蛋白原水解活性;在体内,能够使尖吻蝮蛇毒引起的出血、水肿、组织坏死和致死毒性明显减弱。相比之下,鱼腥草根部提取物也表现出类似的抑制模式,但在相同浓度下其抑制效果均低于地上部分提取物。植物化学成分分析表明,两种提取物中均含有生物碱、蒽醌类物质和蛋白质;地上部分提取物中含有黄酮、多酚、单宁、三萜、甾醇和皂苷类物质,而根部提取物中缺少这些物质。实验结果首次验证了鱼腥草地上部分及根部对尖吻蝮蛇毒的抑制作用,并证实地上部分比根部在抑制毒液的主要酶类和体内毒性方面更为有效,表明鱼腥草地上部分可以作为一种潜在资源用于开发治疗毒蛇咬伤的解毒剂。