Evaluating the inhibitory potential of Withania somnifera on platelet aggregation and inflammation enzymes: An in vitro and in silico study

Context Withania somnifera (L.) Dunal is traditionally used for treating various ailments, but lacks scientific evaluation.

Objective This study evaluates Withania somnifera (WS) for its effect on platelet activity and inflammatory enzymes.

Materials and methods Aqueous and ethanolic (1:1) leaf extracts were subjected to in vitro indirect haemolytic activity using Naja naja venom, human platelet aggregation was quantified for lipid peroxidation using arachidonic acid (AA) as agonist and 5-lipoxygenase (5-LOX) levels were determined using standard spectrometric assays. Further, molecular docking was performed by the ligand fit method using molegro software package (Molegro ApS, Aarhus, Denmark).

Results The study found that aqueous and ethanol extracts have very negligible effect (15%) with an IC₅₀ value of 13.8?mg/mL on PLA₂ from Naja naja venom. Further, extracts of WS also had very little effect (18%) with an IC₅₀ value of 16.6?mg/mL on malondialdehyde (MDA) formation. However, a 65% inhibition of 5-LOX with an IC₅₀ value of 0.92?mg/mL was observed in 1:1 ethanol extracts. The same was evident from SAR model with the active ingredient withaferin A binding predominantly on Phe 77, Tyr 98, Arg 99, Asp 164, Leu 168, Ser 382, Arg 395, Tyr 396 and Tyr 614 with an atomic contact energy value of??128.96 compared to standard phenidone (?103.61). Thus, the current study validates the application of WS for inflammatory diseases.

Conclusion This study reveals the inhibitory potential of W. somnifera on inflammatory enzymes and platelet aggregation. Thus, WS can serve as a newer, safer and affordable medicine for inflammatory diseases.     

~(125)Ⅰ眼镜蛇毒及抗眼镜蛇毒血清在小白鼠体内分布的比较观察

用氯胺-T法125I标记眼镜蛇毒示踪液及抗眼镜蛇毒血清。小白鼠分二组，用125I-眼镜蛇毒、125I-抗眼镜蛇毒血清分别给小白鼠尾静脉注射，于注射后的不同时间测定血、颈部肌肉、肝、肺、肾、心等脏器放射性分布。结果发现：125I-眼镜蛇毒在肌肉及肝、肺等器官中于注射后2小时过高峰，在肾脏中浓度高，在脑未见放射性分布，抗眼镜蛇毒血清与眼镜蛇毒分布相似但在肌肉及肺脏有更高的分布。证明抗眼镜蛇毒血清能跟踪分布眼镜蛇毒。     

中华眼镜蛇毒F组分对沙鼠脑缺血再灌注损伤的保护作用

目的：研究中华眼镜它毒F组分脑缺血再灌注损伤的保护作用,方法：结扎沙鼠双侧颈总动脉1h,造成前脑缺血模型,用比色法测定脑匀浆过氧化脂质的最终产物丙二醛（MDA）的含量及超氧化物歧化酶（SOD）的活性,结果：F组分0．9,0．3,0．1mg/kg可显著抑制脑内脂质过氧化,降低MDA的含量,并提高超化物歧化酶的活,性且呈一定的量效关系,同缺血再灌组比较P＜0．05,同阳性对照药尼莫通比较,P＞0．05,结论：中华眼镜蛇毒F组分对脑缺血再灌注损伤有保护作用,可能与抑制自由基的生成和促进自由基的清除有关。     

中华眼镜蛇毒组分C抗白血病作用的实验研究

目的：研究眼镜蛇毒组分Ｃ对白血病细胞的直接作用及机制。方法：应用ＭＴＴ、ＤＮＡ电泳、流式细胞仪ＲＴ－ＰＣＲ等方法,观察眼镜蛇毒组分Ｃ对ＨＬ６０等９株白血病细胞系的毒性作用、量效关系及白血病细胞经过眼镜蛇毒组分Ｃ处理后的生物化学、Ｂｃｌ－２／Ｂａｘ表达水平变化。结果：眼镜蛇毒组分对９株人白血病细胞系均有明显的抑制作用,ＩＣ５０为０．００４６～４．４０μｇ／ｍｌ,且呈较好的量效关系（ｒ为０．６６～０．     

Naja melanoleuca cobra venom contains two forms of complement-depleting factor (CVF)

Two forms of complement-depleting cobra venom factor (CVFm1 and CVFm2), possessing molecular masses of 142.6 kDa (CVFm1) and 143.1 kDa (CVFm2), according to MALDI mass-spectrometry, were isolated from the Naja melanoleuca cobra venom. As shown by polyacrylamide gel electrophoresis in the presence of SDS, both forms similarly to factor from the Naja kaouthia cobra venom (CVFk) consist of three polypeptide chains with molecular masses of about 70, 50, and 30 kDa, the two large subunits being glycosylated. As determined by MALDI mass-spectrometry, 30 kDa subunits of CVFm1 and CVFm2 have considerably different finger-prints of tryptic digests that suggests differences in their amino acid sequences. A study of activity in vivo has shown no significant differences in C3 consumption by CVFm1, CVFm2 and CVFk in mouse blood. However, as shown by an immunoassay method, they differ in their ability to activate the complement system via C3 conversion, the ratio of these activities for CVFm1:CVFm2:CVFk being 2.5:1.6:1. Kinetic studies using a hemolytic test showed that complement depletion by CVFm1 is faster than that by CVFm2. Thus, for the first time the presence in a single venom of two forms of CVF differing by both amino acid sequence and biological activity has been shown.     

Proteomic characterization of venom of the medically important Southeast Asian Naja sumatrana (Equatorial spitting cobra)

The proteome of Naja sumatrana (Equatorial spitting cobra) venom was investigated by shotgun analysis and a combination of ion-exchange chromatography and reverse phase HPLC. Shotgun analysis revealed the presence of 39 proteins in the venom while the chromatographic approach identified 37 venom proteins. The results indicated that, like other Asiatic cobra venoms, N. sumatrana contains large number of three finger toxins and phospholipases A₂, which together constitute 92.1% by weight of venom protein. However, only eight of the toxins can be considered as major venom toxins. These include two phospholipases A₂, three neurotoxins (two long neurotoxins and a short neurotoxin) and three cardiotoxins. The eight major toxins have relative abundance of 1.6–27.2% venom proteins and together account for 89.8% (by weight) of total venom protein. Other venom proteins identified include Zn-metalloproteinase-disintegrin, Thaicobrin, CRISP, natriuretic peptide, complement depleting factors, cobra venom factors, venom nerve growth factor and cobra serum albumin. The proteome of N. sumatrana venom is similar to proteome of other Asiatic cobra venoms but differs from that of African spitting cobra venom. Our results confirm that the main toxic action of N. sumatrana venom is neurotoxic but the large amount of cardiotoxins and phospholipases A₂ are likely to contribute significantly to the overall pathophysiological action of the venom. The differences in toxin distribution between N. sumatrana venom and African spitting cobra venoms suggest possible differences in the pathophysiological actions of N. sumatrana venom and the African spitting cobra venoms, and explain why antivenom raised against Asiatic cobra venom is not effective against African spitting cobra venoms.     

Anti-necrosis potential of polyphenols against snake venoms

Polyphenols from the extracts of Areca catechu L. and Quercus infectoria Oliv. inhibited phospholipase A₂, proteases, hyaluronidase and L-amino acid oxidase of Naja naja kaouthia Lesson (NK) and Calloselasma rhodostoma Kuhl (CR) venoms by in vitro tests. Both extracts inhibited the hemorrhagic activity of CR venom and the dermonecrotic activity of NK venom by in vivo tests. The inhibitory activity of plant polyphenols against local tissue necrosis induced by snake venoms may be caused by inhibition of inflammatory reactions, hemorrhage, and necrosis. The result implies the therapeutic potential of plant polyphenols against necrosis in snakebite victims.     

中华眼镜蛇毒组分H对实验性动脉粥样硬化的影响

本实验在新西兰兔食饵性动脉粥样硬化模型上,观察了中华眼镜蛇毒组分H预防性给药和治疗性给药对血清胆固醇、过氧化脂质及一氧化氮的影响。发现组分H(2mg·D-1)与2%胆固醇同时喂服或喂胆固醇4周后以组分H(4mg·D-1)治疗数周,均能有效抑制高脂所致的一氧化氮降低和过氧化脂质升高,并能保护超氧化物歧化酶活性和降低血清总胆固醇及甘油三酯的水平。     

消斑肽对C57BL/6J小鼠动脉粥样硬化斑块的影响

恨现从中华眼镜蛇毒素中提取一种多肽能抑制血管平滑肌细胞地殖,并在一定范围内杀伤快速增殖的平滑肌细胞。为验证消斑肽是否具有消除动脉粥样硬化斑块和预防再狭窄的药物效应,选用动脉粥样硬化敏感株近交系Ｃ５７ＢＬ／６Ｊ小鼠,经致动脉粥样硬化饮食饲喂１７周。使用消斑肽腹腔注射３．２μｇ／ｇ治疗４周,在主动脉窦部连续切片,油经Ｏ染色并在显微镜下观察,使用Ｒｏｂｅｒｔｓ ＆ Ｔｈｏｍｐｓｏｎ评分判定斑块消退情况。     

中华眼镜蛇毒抗瘤多肽对人卵巢癌HO-8910细胞的增殖抑制和Fas/FasL表达的影响

目的 探讨中华眼镜蛇毒抗瘤多肽对人卵巢癌HO-8910细胞株的抑制作用及Fas/FasL表达的影响.方法 用MTT比色法检测不同浓度中华眼镜蛇毒抗瘤多肽对HO-8910细胞的增殖抑制,RT-PCR和免疫组织化学观察原癌基因Fas/FasL表达的变化.结果 中华眼镜蛇毒抗瘤多肽对HO-8910细胞增殖抑制作用成剂量依赖性,RT-PCR和免疫组化结果显示,随着中华眼镜蛇毒抗瘤多肽作用剂量的增加,Fas/FasL基因的表达无明显变化.结论 中华眼镜蛇毒抗瘤多肽对人卵巢癌HO-8910细胞系的增殖具有明显的抑制作用,并对原癌基因Fas/FasL的表达无抑制作用.