Stimulation of platelet aggregation by affinity captured rhodocytin from the Malayan pit viper Calloselasma rhodostoma.

The receptor protein CLEC-2 on platelet membranes is the target of the endogenous ligand podoplanin found on cancer cells and of rhodocytin, a snake venom component of the Malayan pit viper Calloselasma rhodostoma. Ligand binding results in platelet activation, increased blood coagulation and thrombosis. In an effort to isolate rhodocytin, we have purified CLEC-2 as bait from E. coli. Affinity captured rhodocytin interacted with mammalian CLEC-2 and stimulated platelet aggregation in a dose dependent manner.

Upper Extremity Crotalid Envenomation: A Review of Incidence and Recent Trends in Management of Snakebites.

PURPOSE: The goal of this study was to evaluate the recent trends in the management of upper extremity Crotalid envenomation in the state of Georgia, United States. METHODS: A retrospective review of the Georgia Poison Center database looking at the reported snakebites to the upper extremity between 2015 and 2020 was performed. Patient demographics, timing and location of injury, severity of envenomation, treatment, including use of antivenin and surgical intervention, and reported complications related to the use of antivenin was extracted. RESULTS: A retrospective review of snakebites between 2015 and 2020 showed 2408 snakebite cases with a mean patient age of 37.4 years. Males incurred 62.8% of all bites. The highest incidence was in summer 52.5%, and between the hours of 5 PM to midnight 57.2%. Overall, 1010 (41.9%) of all bites were categorized as venomous snakebites (55.6% copperhead, 20% rattlesnake, 2.4% cottonmouth, and 22% miscellaneous [including 3 Elapid envenomations] or unidentified. The total number of venomous bites to the upper extremity was 575 (56.9%) and 567 patients received antivenin. Envenomation severity was mild in 29%, moderate in 45%, severe in 10%, and undetermined in 16% of cases. Crotalidae polyvalent immune Fab (Ovine) was the main antivenin used, with overall mean initial therapy dose of 6.2 vials and 59% of patients receiving maintenance therapy. Three patients (0.5%) had a severe anaphylactic reaction to antivenin requiring cessation of therapy. Seven patients had acute compartment syndrome of the upper extremity requiring fasciotomy (3 copperhead, 2 rattlesnake, and 2 unidentified). There was no reported mortality during this period. CONCLUSIONS: Hand surgeons should be familiar with the management of upper extremity Crotalid envenomation. Antivenin remains the main treatment for symptomatic patients. Crotalid snakebites rarely require operative intervention. TYPE OF STUDY/LEVEL OF EVIDENCE: Prognostic IV.

Using Haemocoagulase Agkistrodon in Patients Undergoing Transurethral Plasmakinetic Resection of the Prostate: A Pilot, Real-World, and Propensity Score-Matched Study.

Objectives: To compare the clinical outcomes of using different hemostatic agents after transurethral plasmakinetic resection of the prostate (TUPKP) in benign prostatic hyperplasia (BPH) patients. Methods: The patients were divided into 5 groups according to the hemostatic agents used after TUPKP, including the haemocoagulase agkistrodon for injection (HCA), hemocoagulase for injection (HC), hemocoagulase bothrops atrox for injection (HCB), ethylenediamine diaceturate injection (EDD), and tranexamic acid (TXA). Propensity score matching was performed based on age, body mass index, prostate volume, hypertension status, fasting blood glucose, smoking, and drinking history. The hospitalization time, bladder irrigation time, indwelling catheterization time, the patency of urine flow, and blood transfusion records were used as outcome indicators to compare the clinical effects of these five agents. Results: We finally matched 65 pairs receiving HCA or HC, 71 pairs receiving HCA or HCB, 38 pairs receiving HCA or TXA, and 29 pairs receiving HCA or EDD. Compared with HC, HCA given during the perioperative period significantly reduced the median hospitalization time [7.00 days (5.00, 8.00) vs. 9.00 days (8.00, 10.00); p < 0.001] and median catheterization time (109.00 hours [88.00, 129.00] vs. 164.00 hours [114.00, 189.00], p < 0.001). Compared with EDD, the median hospitalization time (7.00 days [6.00, 8.00] vs. 10.00 days [8.00, 11.00]; p < 0.001) and median catheterization time (113.00 hours [95.00, 143.00] vs. 160.00 hours [139.00, 168.00]; p < 0.001) were also significant shorter in HCA group. Compared with HCB, median bladder irrigation time (45.00 hours [27.00, 71.00] vs. 49.00 hours [45.00, 72.00]; p = 0.04) was shorter in the HCA group. However, there were no statistical differences in outcomes between HCA and TXA. Conclusions: HCA probably has an advantage over HC, HCB, and EDD in reducing the hospitalization time, catheterization time, and bladder irrigation time among BPH patients undergoing TUPKP.

[Agkistrodon halys venom antitumor component-I inhibits vasculogenic mimicry in triple-negative breast cancer cells in vitro by down-regulating MMP2].

OBJECTIVE: To investigate the inhibitory effect of agkistrodon halys venom antitumor component-I (AHVAC-I) on vasculogenic mimicry (VM) formation in triple-negative breast cancer MDA-MB-231 cells and explore its possible mechanism. METHODS: CCK8 assay was used to determine the optimal concentration of AHVAC-I for cell treatment based on its halfinhibitory concentration (IC50). MDA-MB-231 cells were treated with different concentrations of AHVAC-I or 5-Fu, and the changes in vasomimetic capacity of the cells were examined using Matrigel assay. The expression levels of matrix metalloproteinase-2 (MMP2) and MMP9 in the treated cells were detected using quantitative PCR and Western blotting. RESULTS: Compared with the control treatment with culture medium, treatment with 5, 10 and 20 µg/mL AHVAC-I significantly reduced vasomimetic ability of MDA-MB-231 cells in a dose-dependent manner (P < 0.01). MMP2 supplementation obviously restored the vasomimetic ability of the cells inhibited by AHVAC-I. CONCLUSION: AHVAC-I inhibits VM formation in triplenegative breast cancer cells in vitro by down-regulating MMP2 production.

Evaluation of Signaling Pathways Profiling in Human Dermal Endothelial Cells Treated by Snake Venom Cysteine-Rich Secretory Proteins (svCRiSPs) from North American Snakes Using Reverse Phase Protein Array (RPPA).

Cysteine-Rich Secretory Proteins (CRiSPs) are typically found in many snake venoms; however, the role that these toxins play in the pathophysiology of snakebites is still unclear. Herein, we compared the effects of snake venom CRiSPs (svCRiSPs) from the most medically important species of North American snakes on endothelial cell permeability and vascular permeability. We used reverse phase protein array (RPPA) to identify key signaling molecules on human dermal lymphatic (HDLECs) and blood (HDBECs) endothelial cells treated with svCRiSPs. The results showed that Css-CRiSP isolated from Crotalus scutulatus scutulatus and App-CRiSP from Agkistrodon piscivorus piscivorus are the most potent causes of increase vascular and endothelial permeability in comparison with other svCRiSPs used in this study. We examined the protein expression levels and their activated phosphorylation states in HDLECs and HDBECs induced by App-CRiSP and Css-CRiSP using RPPA. Interestingly, both App-CRiSP and Css-CRiSP induced caveolin-1 expression in HDBECs. We also found that stimulating HDBECs with Css-CRiSP and App-CRiSP significantly induced the phosphorylation of mTOR and Src, respectively. In HDLECs, Css-CRiSP significantly downregulated the expression of N-Cadherin and phospholipase C-gamma, while App-CRiSP significantly enhanced Akt and JNK phosphorylation. These results suggest that the increased endothelial permeability in HDLECs and HDBECs by Css-CRiSP and App-CRiSP may occur through different pathways.

Development of a species-specific PCR assay for authentication of Agkistrodon acutus based on mitochondrial cytochrome b gene

BACKGROUND: Agkistrodon acutus, a traditional Chinese medicine, clinically used in the treatment of rheumatism, tumor, and cardiovascular and cerebrovascular diseases. Due to the unique medicinal value and the difficulty of artificial breeding of Agkistrodon acutus, the supply of Agkistrodon acutus on the market exceeds the demand, and a large number of its adulterants are found on the market. In this study, the cytb gene sequences of Agkistrodon acutus and 9 snakes were compared and analyzed, specific primers were designed, and specific PCR methods were established to detect Agkistrodon acutus medicinal samples on the market. RESULTS: This method was successfully applied to distinguish the snake from other adulterated species, and tested 18 Agkistrodon acutus samples randomly purchased from six cities. Twelve samples were counterfeit and six were genuine. The standard reference material of Agkistrodon acutus was cloned by molecular cloning and sequencing, and the gene sequence difference with other species was significant. It shows that the region could be used as the fingerprint region of the target species. CONCLUSIONS: The proposed method can be used as a species-specific marker and can be highly distinguished from other adulterated snake species, which is helpful to effectively avoid the problem of false sale of Agkistrodon acutus.

Hemocoagulase agkistrodon can prevent bleeding and induce hypofibrinogenemia in hepatic disease cases
.

Hemocoagulase is often used for hemostasis in patients with bleeding and hemorrhagic diseases, and to avoid or stanch bleeding after surgery. Herein, three patients with hepatic diseases suffering from hypofibrinogenemia were treated with hemocoagulase agkistrodon (HCA) in Peking University People's Hospital during September 2018. All the 3 patients were chronic hepatitis B patients: Patient 1 presented with hepatic carcinoma and chronic hepatitis B, and right hepatectomy was performed; patient 2 presented with chronic hepatitis B and gastrointestinal bleeding; patient 3 presented with chronic hepatitis B, acute liver failure with hematemesis, and was awaiting liver transplantation. All three patients were percutaneously injected with HCA to prevent late-onset bleeding. After HCA was discontinued, coagulation was restored to > 60 mg/dL on day 6, without injection of fibrinogen. HCA significantly reduced the need for fibrinogen in patients with hepatic diseases, and the level of fibrinogen should be carefully monitored in clinical applications.

Venous hematology, biochemistry, and blood gas analysis of free-ranging Eastern Copperheads (Agkistrodon contortrix) and Eastern Ratsnakes (Pantherophis alleghaniensis).

Hematology, plasma biochemistry, and blood gas analysis were performed on venous samples obtained from free-ranging Eastern Copperheads (Agkistrodon contortrix) and Eastern Ratsnakes (Pantherophis alleghaniensis) in central North Carolina during a mark-recapture study conducted from April to October 2015 at the North Carolina Zoo. Blood samples were collected from 31 (15 male and 16 female) free-ranging copperheads and 34 (20 male and 14 female) free-ranging ratsnakes at the beginning and end of restraint. Restraint was performed for morphometric measurements, sex determination, and identification via placement of intracelomic passive integrated transponder (PIT) tags and marking of ventral scutes with a handheld electrocautery unit. Blood gas analytes were measured at the beginning of restraint and compared to analytes measured at the end to evaluate for changes secondary to handling. Total restraint time prior to the first blood sampling was 1.4 ± 0.4 mins (mean ± SD) and 1.0 ± 0.2 mins (mean ± SD) and restraint time prior to second blood sampling was 12.5 ± 2.4 mins (mean ± SD) and 13.5 ± 3.4 mins (mean ± SD) for copperheads and ratsnakes, respectively. Blood lactate concentrations at the beginning of restraint were similar for both species. Lactate concentrations increased significantly and pH decreased significantly for both species at the end of restraint when compared to the beginning of restraint. Furthermore, lactate concentrations at the end of restraint were significantly elevated in ratsnakes compared to copperheads. This study provides guidelines for interpretation of venous hematology, plasma biochemistry, and blood gas values for free-ranging copperheads and ratsnakes in central North Carolina and demonstrates the physiological response to venous blood gas analytes secondary to capture and restraint.

[Transcriptome analysis of venom gland and identification of functional genes for snake venom protein in Agkistrodon acutus].

Agkistrodon acutus is a traditional Chinese herb medicine which has immunological regulation,anti-tumor,anti-inflammatory and analgesic effects,which is mainly used for the treatment of rheumatoid arthritis,ankylosing spondylitis,sjogren's syndrome and tumors. In order to excavate more important functional genes from A. acutus,the transcriptome of the venom gland was sequenced by the Illumina Hi Seq 4000,and 32 862 unigenes were assembled. Among them,26 589 unigenes were mapped to least one public database. 2 695 unigenes were annotated and assigned to 62 TF families,and 5 920 SSR loci were identified. The majority of mapped unigenes was from Protobothrops mucrosquamatus in the NR database,which revealed their closest homology. Three secretory phospholipase A_2 with different amino acid sequences showed similar spatial structures and all had well-conserved active sites. The 3 D structural models of C-type lectin showed conserved glycosylation binding sites( Asn45). This study will lay the foundation for the further study of the function of snake venom protein,and promoting the development and utilization of genome resources from A. acutus.

ACP-TX-I and ACP-TX-II, Two Novel Phospholipases A2 Isolated from Trans-Pecos Copperhead Agkistrodon contortrix pictigaster Venom: Biochemical and Functional Characterization.

This work reports the purification and biochemical and functional characterization of ACP-TX-I and ACP-TX-II, two phospholipases A2 (PLA2) from Agkistrodon contortrix pictigaster venom. Both PLA2s were highly purified by a single chromatographic step on a C18 reverse phase HPLC column. Various peptide sequences from these two toxins showed similarity to those of other PLA2 toxins from viperid snake venoms. ACP-TX-I belongs to the catalytically inactive K49 PLA2 class, while ACP-TX-II is a D49 PLA2, and is enzymatically active. ACP-TX-I PLA2 is monomeric, which results in markedly diminished myotoxic and inflammatory activities when compared with dimeric K49 PLA2s, confirming the hypothesis that dimeric structure contributes heavily to the profound myotoxicity of the most active viperid K49 PLA2s. ACP-TX-II exhibits the main pharmacological actions reported for this protein family, including in vivo local myotoxicity, edema-forming activity, and in vitro cytotoxicity. ACP-TX-I PLA2 is cytotoxic to A549 lung carcinoma cells, indicating that cytotoxicity to these tumor cells does not require enzymatic activity.

Potential use of 13-mer peptides based on phospholipase and oligoarginine as leishmanicidal agents.

Phospholipase A2 toxins present in snake venoms interact with biological membranes and serve as structural models for the design of small peptides with anticancer, antibacterial and antiparasitic properties. Oligoarginine peptides are capable of increasing cell membrane permeability (cell penetrating peptides), and for this reason are interesting delivery systems for compounds of pharmacological interest. Inspired by these two families of bioactive molecules, we have synthesized two 13-mer peptides as potential antileishmanial leads gaining insights into structural features useful for the future design of more potent peptides. The peptides included p-Acl, reproducing a natural segment of a Lys49 PLA2 from Agkistrodon contortrix laticinctus snake venom, and its p-AclR7 analogue where all seven lysine residues were replaced by arginines. Both peptides were active against promastigote and amastigote forms of Leishmania (L.) amazonensis and L. (L.) infantum, while displaying low cytotoxicity for primary murine macrophages. Spectrofluorimetric studies suggest that permeabilization of the parasite's cell membrane is the probable mechanism of action of these biomolecules. Relevantly, the engineered peptide p-AclR7 was more active in both life stages of Leishmania and induced higher rates of ethidium bromide incorporation than its native template p-Acl. Taken together, the results suggest that short peptides based on phospholipase toxins are potential scaffolds for development of antileishmanial candidates. Moreover, specific amino acid substitutions, such those herein employed, may enhance the antiparasitic action of these cationic peptides, encouraging their future biomedical applications.

The extract from Agkistrodon halys venom protects against lipopolysaccharide (LPS)-induced myocardial injury.

BACKGROUND: Snake venoms contain various bioactive constituents which possess potential therapeutic effects. The aim of this work was to investigate the effect of the extract from Agkistrodon halys venom on lipopolysaccharide (LPS)-induced myocardial injury. METHODS: Thirty male Sprague-Dawley rats were randomly assigned to three groups (10 rats per group): control group, LPS group and LPS + extract group. Rats in control and the LPS groups were intravenously injected with sterile saline solution, and rats in the LPS + extract group with the extract. After 2 h, rats of the control group were intraperitoneally injected sterile saline solution, and rats in the LPS and the LPS + extract groups were treated with LPS (20 mg per kg body weight). Levels of creatine kinase (CK) and lactate dehydrogenase (LDH) in serum were determined. Anti-inflammation of the extract was analyzed via determination of TNF-α and IL-6 in serum, and expression of TNF-α, IL-6, COX-2 and p-ERK protein in hearts. Heme oxygenase-1 (HO-1) and p-NF-κB protein expression in hearts, superoxide dismutase (SOD) activity and malondialdehyde (MDA) level in serum were used to evaluate the anti-oxidative properties of the extract. RESULTS: Extract pretreatment significantly decreased the level of serum CK and LDH, reduced the generation of inflammatory cytokines such as TNF-α and IL-6, and also reduced serum level of MDA in the LPS + extract group compared with the LPS group. In addition, the extract increased SOD activity in serum, HO-1 protein expression in hearts, and decreased TNF-α, IL-6, COX-2, p-NF-κB and p-ERK1/2 protein expression. CONCLUSION: Our results suggested that beneficial effect of this extract might be associated with an improved anti-oxidation and anti-inflammatory effect via downregulation of NF-κB/COX-2 signaling by activating HO-1/CO in hearts.

Does This Patient Have a Severe Snake Envenomation?: The Rational Clinical Examination Systematic Review.

Importance: Venomous snakebite severity ranges from an asymptomatic dry bite to severe envenomation and death. The clinical evaluation aids in prognosis and is essential to determine the risks and potential benefits of antivenom treatment. Objectives: To identify historical features, clinical examination findings, basic laboratory testing, and clinical grading scales that will risk-stratify patients with pit viper snake envenomation for severe systemic envenomation, severe tissue injury, and/or severe hematologic venom effects. Data Sources: We conducted a structured search of PubMed (1966-October 3, 2017) and Embase database (1980-October 3, 2017) to identify English-language studies that evaluated clinical features predictive of severe envenomation. Study Selection: We included studies that evaluated the test performance of at least 1 clinical finding with an acceptable reference standard of severe envenomation for venomous snakes of the Western Hemisphere. Only studies involving the most common subfamily, Crotalinae (pit vipers), were evaluated. Seventeen studies with data were available for abstraction. Data Extraction and Synthesis: The clinical features assessed and severity outcome measures were extracted from each original study. We assessed severity in 3 categories: systemic toxicity, tissue injury, and hematologic effects. Differences were resolved by author consensus. Results: The pooled prevalence of severe systemic envenomation was 14% (95% CI, 9%-21%). The pooled prevalence of severe tissue injury and severe hematologic venom effects were 14% (95% CI, 12%-16%) and 18% (95% CI, 8%-27%), respectively. Factors increasing the likelihood of severe systemic envenomation included the time from bite to care of 6 or more hours (likelihood ratio [LR], 3.4 [95% CI, 1.1-6.4]), a patient younger than 12 years (LRs, 3.2 [95% CI, 1.5-7.1] and 2.9 [95% CI, 1.3-6.2]), large snake size (LR, 3.1 [95% CI, 1.5-5.7]), and ptosis (LRs, 1.4 [95% CI, 1.0-2.1] and 3.8 [95% CI, 1.8-8.3]). Envenomation by the genus Agkistrodon (copperhead and cottonmouth), as opposed to rattlesnakes, decreased the likelihood of severe systemic envenomation (LR, 0.28 [95% CI, 0.10-0.78]). Initial hypofibrinogenemia (LR, 5.1 [95% CI, 1.7-15.0]) and thrombocytopenia (LR, 3.7 [95% CI, 1.9-7.3]) increased the likelihood of severe hematologic venom effects. Other clinical features from history, physical examination, or normal laboratory values were not discriminative. Conclusions: Clinical features can identify patients at increased risk of severe systemic envenomation and severe hematologic venom effects, but there are few features that are associated with severe tissue injury or can confidently exclude severe envenomation. Physicians should monitor patients closely and be wary of progression from nonsevere to a severe envenomation and have a low threshold to escalate therapy as needed.

Hemostatic Effect of Hemocoagulase Agkistrodon on Surgical Wound in Breast Cancer Surgery.

Objective To evaluate the hemostatic effect of hemocoagulase agkistrodon on surgical wound in breast cancer surgery. Methods Totally 60 patients undergoing breast cancer surgery were enrolled in this prospective,randomized,double-blinded,and controlled study. All the patients met the inclusion and exclusion criteria and signed the informed consent. Hemocoagulase agkistrodon (2 U) was injected 20 minutes before surgery and 4 and 24 hours after surgery in the intervention group (n=30),whereas normal saline was used instead in the control group (n=30). The volume of intraoperative bleeding,wound drainage volume 1-3 days after surgery,and total drainage volume were recorded. Meanwhile,the change of blood coagulation function,treatment safety,and clinical outcomes were observed. Results The intra-operative hemorrhage volume of the intervention group [(95.0±48.3)g] was significantly lower than that of the control group [(144.8±105.4)g] (t=-2.07,P=0.044). The volume of total drainage of the intervention group [(166.7±71.2)g] was significantly lower than that of the control group [(251.4±166.3)g] (t=-2.29,P=0.029). The hemoagglutination indicators were similar in the two groups and no complication such as thrombosis occurred. The length of hospital stay of the intervention group [(15.00±3.53)d] was similar to that of the control group [(15.92±2.32)d] (t=-1.057,P=0.297). No research drug-related adverse event was occurred in our study. Conclusion Hemocoagulase agkistrodon has good hemostatic effect for patients undergoing breast cancer surgery without increasing the risk of thrombosis.

Agkistrodon bilineatus. Experimentación pura / Agkistrodon bilineatus. Pure experimentation

El presente trabajo muestra una experimentación pura basada en la preparación homeopática del veneno de la serpiente Agkistrodon bilineatus, algo que ha demostrado su eficacia en diferentes aplicaciones clínicas que datan de la época de Constantine Hering, uno de los pioneros de la Homeopatía en Estados Unidos. Los resultados son alentadores, toda vez que los órganos con mayor número y diversidad de síntomas fueron el sistema nervioso central, los ojos, así como los aparatos respiratorio y digestivo. En conclusión, este derivado del veneno de la serpiente en referencia, preparado homeopáticamente, puede emplearse debido a que se generó una patogenesia concreta en un grupo de experimentadores.

MANAGEMENT OF OSTEOMYELITIS CAUSED BY SALMONELLA ENTERICA SUBSP. HOUTENAE IN A TAYLOR'S CANTIL (AGKISTRODON BILINEATUS TAYLORI) USING AMIKACIN DELIVERED VIA OSMOTIC PUMP.

An adult female Taylor's cantil (Agkistrodon bilineatus taylori) presented with marked spinal and mandibular osteomyelitis that cultured positive for Salmonella enterica subsp. houtenae, serovar IV 43:z4,z32:-. Progression of osteomyelitis was arrested by treatment using amikacin (0.026 mg/kg per hour) delivered via subcutaneous osmotic pump for 10 mo, replacing the pump every 4 wk. No adverse effects on renal function were appreciated throughout the course of therapy. Amikacin therapy was discontinued after improvement with treatment, but 5 mo later, bony lesions worsened, and an additional abscess formed at the previous pump site. The animal's condition declined and euthanasia was elected. Postmortem examination confirmed marked osteomyelitis with Salmonella infection of same serovar as the initial biopsy. This report highlights the pathogenicity of the S. enterica subsp. houtenae serovar and the ability to deliver effective amikacin dosage via osmotic pump to arrest osteomyelitis due to salmonellosis in a venomous snake.

The characterization of trans-pecos copperhead (Agkistrodon contortrix pictigaster) venom and isolation of two new dimeric disintegrins.

The vast amounts of toxins within the venom of snakes, while known to cause medical emergencies, display various biological functions. Trans-pecos copperhead (Agkistrodon contortrix pictigaster) crude venom separated by cation-exchange chromatography showed several fractions with fibrinolytic, hemorrhagic, gelatinase and platelet activities. Venom fractions 1, 2, 4, 5, and 12-17 contained fibrinolytic activity. Venom fractions 1, 2, 5 and 12-14 had hemorrhagic activity. Fractions 1, 2, 12, 13 and 17 contained gelatinase activity. Reverse-Phase C18 High Performance Liquid Chromatography was also used to purify and isolated disintegrins from this venom. Anti-platelet aggregation activity of the C18 fractions collected and performed on whole human blood showed that they inhibited platelet aggregation in presence of several agonists. Results from both SDS-PAGE and N-terminal sequencing determined that pictistatin 1 obtained from the Trans-Pecos copperhead venom was a dimeric disintegrin, and pictistatin 2 was a heterodimeric disintegrin. The molecules with anti-platelet activity could be considered in the development of more effective drugs, for numerous blood-related diseases such as stroke, heart attacks, thrombosis, and other medical conditions. In this study, we are presenting the first report of the purification, isolation, and partial characterization of two new dimeric disintegrins isolated from the venom of trans-pecos copperhead.

Hemocoagulase agkistrodon-induced anaphylactic shock: A case report and literature review.

Hemocoagulase agkistrodon for injection is the national first-class new drug of China with good hemostatic function and safety for capillary hemorrhage in abdominal incision of surgical patients. Adverse drug reactions (ADRs) to hemocoagulase agkistrodon are rarely reported. In this paper, we describe a case of a 41-year-old woman who developed anaphylactic shock attributed to hemocoagulase agkistrodon before colon cancer surgery. Based on the Naranjo ADR probability score, a "probable" cause and effect relationship existed for this case. Although the cause of anaphylactic reaction (hemocoagulase or excipient) and exact mechanism of hemocoagulase agkistrodon-induced anaphylactic reaction are unknown, attention should be drawn to potential ADRs in clinical use.

Effect of Iron and Carbon Monoxide on Fibrinogenase-like Degradation of Plasmatic Coagulation by Venoms of Six Agkistrodon Species.

Annually, thousands suffer poisonous snakebite, often from defibrinogenating species. It has been demonstrated that iron and carbon monoxide change the ultrastructure of plasma thrombi and improve coagulation kinetics. Thus, this investigation sought to determine whether pre-treatment of plasma with iron and carbon monoxide could attenuate venom-mediated catalysis of fibrinogen obtained from Agkistrodon species with fibrinogenase activity. Human plasma was pre-treated with ferric chloride (0-10 µM) and carbon monoxide-releasing molecule-2 (CORM-2, 0-100 µM) prior to exposure to 0.5-11 µg/ml of six different Agkistrodon species' venom. The amount of venom used for experimentation needed to decrease coagulation function of one or more kinetic parameters by at least 50% of normal values for (e.g. half the normal speed of clot formation). Coagulation kinetics were determined with thrombelastography. All six snake venoms degraded plasmatic coagulation kinetics to a significant extent, especially prolonging the onset to clot formation and diminishing the speed of clot growth. Pre-treatment of plasma with iron and carbon monoxide attenuated these venom-mediated coagulation kinetic changes in a species-specific manner, with some venom effects markedly abrogated while others were only mildly decreased. Further in vitro investigation of other pit viper venoms that possess fibrinogenolytic activity is indicated to identify species amenable to or resistant to iron and carbon monoxide-mediated attenuation of venom-mediated catalysis of fibrinogen. Lastly, future pre-clinical investigation with animal models (e.g. rabbit ear-bleed model) is planned to determine whether iron and carbon monoxide can be used therapeutically after envenomation.

Balancing the Expression and Production of a Heterodimeric Protein: Recombinant Agkisacutacin as a Novel Antithrombotic Drug Candidate.

Agkisacucetin extracted from the venom of Agkistrodon acutus has been demonstrated to be a promising antithrombotic drug candidate in clinical studies due to its function as a novel platelet membrane glycoprotein (GP) Ib inhibitor. Agkisacucetin is a heterodimeric protein composed of α- and ß-subunits with seven disulphide bonds. Both subunits form inactive homodimeric products, which cause difficulties for recombinant production. In this study, Agkisacucetin α- and ß-subunits were inserted sequentially into the chromosome of Pichia pastoris at the mutant histidinol dehydrogenase gene and ribosomal DNA repeat sites, respectively. By optimizing the gene copies and productivity of each subunit by drug screening, we successfully obtained a recombinant strain with balanced expression of the two subunits. Using this strain, a yield greater than 100 mg/L recombinant Agkisacucetin in fed-batch fermentation was reached. The recombinant Agkisacucetin possessed extremely similar binding affinity to recombinant GPIb and human platelets in in vitro assays, and its ristocetin-induced platelet aggregation activity ex vivo was identical to that of the extracted native Agkisacucetin, demonstrating that the yeast-derived Agkisacucetin could be an effective alternative to native Agkisacucetin. Moreover, this study provides an effective strategy for balancing the expression and production of heterodimeric proteins in P. pastoris.