RESUMEN
Biobanking of turkey ovarian tissue has the potential to play a crucial part in preserving female genetics. To date, ovarian tissue has only been vitrified using a standard protocol, with immediate analyses after warming, therefore, long-term cryoinjury is unknown. Long-term cryoinjury was investigated here by in-ovo culturing, fresh (non-vitrified), a purposefully suboptimal poor vitrification (PV), and the standard vitrified (StV) protocol. Assessments were performed via cellular morphological changes and mRNA gene expression differences, immediately (day 0) or after 2, 4, or 6 days of in-ovo culturing. On day 0, the mRNA levels of heat-shock protein A2 (HSPA2) were lowest in the fresh tissue, and increased 5-fold in the StV treatment, and 18-fold in the PV treatment. Whereas, by day 6, growth determining factor 9 (GDF9) mRNA levels within the fresh tissue were over 3-fold and 21-fold higher than StV and PV treatments, respectively. After 6 days of in-ovo culture the follicle density was highest in the fresh ovarian tissue (4701 ± 950 #/mm3), followed by the StV (1601 ± 300 #/mm3), with PV having the lowest density (172 ± 145 #/mm3). This shows that although the density of follicles was higher in StV versus PV, a considerable number (â¼65 %) were lost compared to the fresh treatment. Additionally, the HSPA2 expression could be an early screening tool, whereas GDF9 expression could be a late screening tool, used to assess turkey ovarian tissue vitrification protocols. We conclude that the StV protocol should be further optimized to try and improve follicle numbers post-warming.
Asunto(s)
Bancos de Muestras Biológicas , Criopreservación , Femenino , Humanos , Criopreservación/métodos , Ovario , Vitrificación , Expresión Génica , ARN Mensajero/genéticaRESUMEN
BACKGROUND: Cyanide is a rapid acting, lethal, metabolic poison and remains a significant threat. Current FDA-approved antidotes are not amenable or efficient enough for a mass casualty incident. OBJECTIVE: The objective of this study is to evaluate short and long-term efficacy of intramuscular aqueous dimethyl trisulfide (DMTS) on survival and clinical outcomes in a swine model of cyanide exposure. METHODS: Anesthetized swine were instrumented and acclimated until breathing spontaneously. Potassium cyanide infusion was initiated and continued until 5 min after the onset of apnea. Subsequently, animals were treated with intramuscular DMTS (n = 11) or saline control (n = 10). Laboratory values and DMTS blood concentrations were assessed at various time points and physiological parameters were monitored continuously until the end of the experiment unless death occurred. A subset of animals treated with DMTS (n = 5) were survived for 7 days to evaluate muscle integrity by repeat biopsy and neurobehavioral outcomes. RESULTS: Physiological parameters and time to apnea were similar in both groups at baseline and at time of treatment. Survival in the DMTS-treated group was 90% and 30% in saline controls (p = 0.0034). DMTS-treated animals returned to breathing at 12.0 ± 10.4 min (mean ± SD) compared to 22.9 ± 7.0 min (mean ± SD) in the 3 surviving controls. Blood collected prior to euthanasia showed improved blood lactate concentrations in the DMTS treatment group; 5.47 ± 2.65 mmol/L vs. 9.39 ± 4.51 mmol/L (mean ± SD) in controls (p = 0.0310). Low concentrations of DMTS were detected in the blood, gradually increasing over time with no elimination phase observed. There was no mortality, histological evidence of muscle trauma, or observed adverse neurobehavioral outcomes, in DMTS-treated animals survived to 7 days. CONCLUSION: Intramuscular administration of aqueous DMTS improves survival following cyanide poisoning with no observed long-term effects on muscle integrity at the injection site or adverse neurobehavioral outcomes.
Asunto(s)
Antídotos , Sulfuros , Animales , Antídotos/farmacología , Antídotos/uso terapéutico , Cianuros , Humanos , Cianuro de Potasio , PorcinosRESUMEN
Smoke inhalation from a structure fire is a common route of cyanide poisoning in the U.S. Cyanide inhibits cellular respiration, often leading to death. Its rapid distribution throughout the body can result in injuries to multiple organs, and cyanide victims were reported to experience myocardial infarction and other cardiac complications. However, molecular mechanisms of such complications are yet to be elucidated. While FDA-approved CN antidotes such as sodium thiosulfate and hydroxocobalamin are clinically used, they have foreseeable limitations during mass casualty situations because they require intravenous administration. To facilitate the development of better antidotes and therapeutic treatments, a global view of molecular changes induced by cyanide exposure is necessary. As an exploratory pursuit, we performed oligonucleotide microarrays to establish cardiac transcriptomes of an animal model of nose-only inhalation exposure to hydrogen cyanide (HCN), which is relevant to smoke inhalation. We also profiled cardiac transcriptomes after subcutaneous injection of potassium cyanide (KCN). Although the KCN injection model has often been used to evaluate medical countermeasures, this study demonstrated that cardiac transcriptomes are largely different from that of the HCN inhalation model at multiple time points within 24 h after exposure. Pathway analysis identified that HCN-induced transcriptomes were enriched with genes encoding mediators of pathways critical in modulation of cardiac complications and that a large number of such genes were significantly decreased in expression. We utilized the upstream regulatory analysis to propose drugs that can be potentially employed to treat cyanide-induced cardiac complications.
Asunto(s)
Lesiones Cardíacas/complicaciones , Cianuro de Hidrógeno/envenenamiento , Cianuro de Potasio/envenenamiento , Animales , Cianuro de Hidrógeno/administración & dosificación , Exposición por Inhalación , Inyecciones Subcutáneas , Masculino , Ratones , Cianuro de Potasio/administración & dosificaciónRESUMEN
CONTEXT: Cyanide (CN) is a metabolic poison, halting ATP synthesis by inhibiting complex IV of the electron transport chain. If exposed at high enough concentrations, humans and most animals can die within minutes. Because time is a crucial factor in survival of CN poisoning, a rapidly bioavailable, nontoxic, easy to administer CN medical countermeasure could improve morbidity/mortality in a mass CN exposure scenario. The most likely route of exposure to CN is via inhalation. OBJECTIVE: This study examined the efficacy of a new formulation for dimethyl trisulfide (DMTS), a countermeasure which has shown promise as a treatment for CN poisoning, using both inhalation and injection models of CN exposure. METHODS: We developed a model of acute CN inhalation intoxication, using the highly toxic agent system from CH Technologies for nose-only exposure. Both continuous and discontinuous HCN exposure paradigms were implemented. For comparison, we also utilized a potassium cyanide (KCN) injection model. In all experiments, DMTS was administered as a cyanide countermeasure via intramuscular injection in unanesthetized mice. RESULTS: We found DMTS administration to be highly protective against both subcutaneous KCN and HCN inhalation toxicity. In the KCN injection model, DMTS afforded protection against 3.73 times the LD50 dose of KCN. In our HCN inhalation exposure model, mice challenged with LC50 HCN doses for the duration of either 10- or 40-minute exposure paradigms demonstrated improved survival in the presence of DMTS treatment (87.5% and 90.0% survival, respectively). Animals in the DMTS treatment groups of both lethal exposure models similarly exhibited improvement in observed toxic signs. CONCLUSION: We show that a newly developed formulation of DMTS is efficacious within two lethal CN exposure mouse models (inhalation and injection) and is highly effective by intramuscular injection. Within these HCN studies, we demonstrate efficacy of DMTS in both continuous and discontinuous inhalation exposure models.