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“Cryopreservation is just crude freezing,” is still a common misconception for cryonics. In reality, a multitude of calculated steps have to be made between legal death and possible future revival. One of them is the “cryoprotection procedure”, in which we perfuse cryoprotectant agents to shield the patients’ cells from damage. Why and how exactly are we doing this? Find out in this article.

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Why is Perfusion Needed?

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To understand why perfusion with cryoprotectants is needed, we first have to take a look at what happens to the body during cryopreservation. 

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Cryopreserved patients are most commonly stored in cryogenic storage dewars, submerged in liquid nitrogen at -196°C. Right after legal death, from the initiation of the procedure onwards, the body temperature is constantly lowered. When cooled down below -130°C, patients go through the so-called “glass transition temperature”. Here, vitrification occurs, a glass-like state in which all metabolic activity in the body is paused.

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If we were to cool down to such extreme temperatures without taking precautions, the water inside our body would quickly turn to ice - something we definitely don’t want. Ice crystal formation inside the body irrevocably leads to cellular damage, as they cause cells to dehydrate and shrink. We try to circumvent this effect as much as possible. For that reason, an early step of our procedure is to remove the blood from a patient’s body after legal death, then substitute it with cryoprotective agents.

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How Does Perfusion Work?

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After our SST Team reaches the patient, and legal death has officially been diagnosed, the cryoprotection procedure begins. First on the list is stabilization, where the patient is cooled down to a certain degree so the cryoprotection process can start. The team then needs to surgically access the chest cavity. This is done by classic median sternotomy. Once the heart and its surrounding structures are dissected carefully and quickly, cannulation can take place. To do this, the SST team cannulates the aorta and vena cava (largest vein in the body), and attach our perfusion system. A pump takes over the job of the heart to continuously pump blood, while our team is able to pump the blood out of the body and substitute it with our custom-made cryoprotectants. This process continues until the perfect concentration of cryoprotective agents is reached. Generally, a larger concentration correlates with less freezing of the cells, yet different concentrations are needed at different stages of introduction to the system. Afterwards, we close the circulation again and continue cooling.

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Step-By-Step Perfusion

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Perfusion is not quite as straight-forward as the overview might suggest. As is so often the case, the devil lies in the details. We could easily run into problems if we didn’t pay careful attention. Let’s take another step back and look at the perfusion from top to bottom.

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Cryoprotectant Storage & Roller Pumps

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Before usage, cryoprotectants need to stay cooled constantly. Therefore, we store them in special perfusion bags at cooled temperatures. The first three concentrations are stored at 4°C inside a fridge; the highest concentration at -20°C in a freezer. Once deployed, the bags are hung up and attached to the perfusion system, where the cryoprotective agents are led into a reservoir. From there a roller pump, the same kind used in cardiopulmonary bypass systems, helps take the blood out of the body, and at the same time perfuse the cryoprotectant in.

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Important Filters

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Before the substance reaches the body, it goes through a heat exchanger and arterial filter. These remove any sort of impurities that may have been introduced to the solution by mistake (cryoprotectants are produced under sterile conditions).

It then goes through static and dynamic bubble traps used to prevent micro bubbles from getting through. If bubbles enter the bloodstream, they would act the same as a blood clot (embolus), which can not easily be removed and may prevent cryoprotectants from reaching every nook and cranny like they are supposed to. This must not happen at all costs, hence why bubble filters are used.

After these steps, the cryoprotectant gets perfused directly into the body.

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During the entire perfusion, a pressure gauge is making sure that blood pressure is kept at a normal level. The pressure we work with is very low for human standards, at around 80-90 mmhg up to a maximum of 120 mmhg.

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Closing the Circle

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The process of pumping cryoprotectants through the system is continued until the perfect concentration is reached. Afterwards, the circuit is closed and the body is cooled down further to optimal temperatures. 

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Voilà, the perfusion is done. 

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Excess agents are being properly disposed of, following the completion of the cryoprotection procedure.

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Conclusion

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As you can see, we do a bit more than just “freezing humans” at Tomorrow Bio. The perfusion step is but one of many that help lead to a high quality cryopreservation.

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Tomorrow Bio will continue to look into further optimization of this system by regularly testing out new components. If a new part proves useful, it may be added to the system further down the line. This way we are aiming to make the perfusion process even faster and more efficient in the future.

Are you interested in finding out more about cryopreservation? Feel free to schedule a call with us! We are always happy to talk to you!

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