About Study Insights: Göttingen Minipigs are increasingly selected for all aspects of biomedical research and are fully recognized as a reliable and established animal model by all regulatory authorities worldwide. This section aims at providing an insight into the wide use of Göttingen Minipigs within biological research.
Background
Ellegaard Göttingen Minipigs was chosen as key partner for model development by Tissue-Link for establishment of a pre-clinical pig model in excessive bleeding to support the development of our innovative bio surgical devices.
This section aims at providing an insight into the wide use of Göttingen Minipigs within biological research. If you know of an interesting study, you are welcome to reach out.
Unmet medical need
Excessive hemorrhage is the leading cause of preventable civilian- and combat related deaths. In addition, excessive hemorrhage increases mortality, extend intensive care unit treatments and increases societal costs for patients surviving severe bleeding. Early control is the most effective strategy for treating hemorrhage and therefore a key target for new development of a safe and effective therapy.
Solution
Tissue-Link is developing hemostatic products encompassing a construct of the human protein, tissue factor, as first-in-class. Tissue factor initiates and amplifies clot development whereby bleeding is stopped effectively including bleeding from large arteries which is high pressure bleedings. TL-101 is our first non-resorbable dressing, which is able to control excessive life-threatening bleedings. Tissue-Link’s methodology circumvents the risk previously known by use of tissue factor as a treatment option through an innovative linking of tissue-factor to a dressing.
Research challenge
To validate the hemostatic efficacy of TL-101 Tissue-Link, a robust and relevant bleeding model in a relevant species is needed. The coagulation among species is different and compared to humans most animals are hypercoagulable and therefore models, in smaller animals such as rats, can be difficult to evaluate both in relation to safety as well as efficacy.
A femoral artery lesion model in regular swine has been used by e.g. the US army for efficacy evaluation (Kheirabadi et al. 2011). In this model the femoral artery is damaged by a 6 mm arteriotomy followed by free bleeding 30 seconds, followed by typical 3 minutes compression with the examined device. With the most used hemostat in the US army QuikClot (not indicated for arterial bleedings), the survival rate and hemostatic success is around 30% in this model.
We wanted to develop a model with smaller animals keeping the significance of species selection intact and therefore initiated a collaboration with Ellegaard Göttingen Minipigs by use of Göttingen Minipigs. Due to anatomic differences, the use of the femoral artery was difficult in minipigs as it is located rather deep in the femoral region and would require complicated surgery. Therefore, Kirsten Rosemary Jacobsen and a team from Ellegaard Göttingen Minipigs suggested to explore the use of the carotid artery instead as the bleeding site as an alternative.
Method
The experiment was approved by the Danish Animal Experiments Inspectorate (license number: 2020-15-0201-00569) and all animal procedures were performed in accordance with the Guide for the Care and Use of Laboratory Animals in an AAALAC international approved barrier facility. A study outline as seen in image 2 was developed and the first study took place in 2020.
Coagulation status of the animals were evaluated by thromboelastographic parameters (TEG). The minipigs received a minute dose of 25iu/kg heparin to humanize their coagulation status. Anesthesia was induced by Zoletil 50 (tiletamin 125mg, zolazepam 125mg) + 6.25 ml xylazine (20 mg/ml) + 1.25 ml Ketamine (100 mg/ml) + 2.5 ml butorphanol (10 mg/m), dose 1 ml/10 kg IM. Once anesthetized, animals were intubated and prepared for surgery lege artis. Anesthesia was maintained by isoflurane inhalation, 0-2%, to effect and the following parameters are evaluated and maintained within normal range: FiO2, MV, RR, EtCO2, HAR, SpO2, IBP and body temperature. IV access was obtained by a midline catheter either in the ear vein or v. femoralis. A pressure catheter was placed in the femoral artery, and ringers Acetate was provided at various rates to maintain a mean arterial pressure (MAP) above 65mmHg.
For the carotid artery lesion skin, subcutaneous tissue were dissected and the left carotid artery was identified. The vessel was cleaned from connecting tissue and non-invasive holding sutures were placed underneath the artery to allow to be lifted more dorsal for ease of precise puncture. A few drops of lidocaine (20mg/ml lidocainhydrochlorid monohydrat) were added a few minutes prior to puncture to allow for maximum vasodilation. A vascular puncture was made with a 14G syringe in a 45-degree angle and twisted with a visual significant lesion as a result. After the lesion was confirmed visually, the artery was lowered to its normal position and upon releasing the artery 30 sec of free bleeding.
Thereafter, the test device was compressed at the lesion for 5 minutes. After compression the device was removed and the cavity checked for hemostasis. We studied TL-101, placebo, and the active comparator QuikClot. Placebo and TL-101 were blinded but QuikClot was visually and material wise different from the other devices.
Experience with model development and value for Tissue-Link
Our experience with the model development together with Ellegaard Göttingen Minipigs has been very positive. We have achieved convincing preclinical results in a new model, a relevant species and an arterial bleeding model. We can now build our further device development on this convincing model. Ellegaard Göttingen Minipigs has been very supportive, knowledgeable, and flexible in their support. We have also enjoyed the opportunity for a true collaboration, where we were allowed to be participants – It has truly been teamwork.
