“Cryopreservation Of Human Body Organs

Introduction Cryopreservation is a process where cell or organ preserved by cooling to sub zero temperatures. Cryoprotectant solutions are used in cryopreservation. e.g. Phosphate buffered sucrose solution.

Defination Cryonics: It is the technique used to store human bodies at extremely low temperatures. Cryogenics: It is defined as, "The branches of physics and engineering that involve the study of very low temperatures, how to produce them, and how materials behave at those temperatures."

Principles of organ preservation Remove the organ Organ is cooled rapidly and warm ischemia time is minimized Transferred in cryoprotectant solution The organ is maintained in hypothermic state and stored until it is ready to be transplant

Techniques of organ preservation Hypothermic preservation. Freezing and thawing. Vitrification.

Hypothermic preservation Two techniques of hypothermic preservation: simple cold storage: continuous hypothermic perfusion:

Freezing and thawing When ice is used in this tech. it can damge to structual integrity of the organ. Thermal strees occurin at low temp. Osmotic movement can also causes mechanical stresses.

Vitrification This is the process of taking an aqueous solution and making it into an amorphous solid. Vitrification can also be achieved by adding solutes that develop a structure in water that must be broken down for crystalline growth.

Preservation solutions Euro-Collins solutions Ross-Marshall citrate solutions Phosphate-buffered sucrose solution University of Wisconsin solution Celsior solution Kyoto ET solution

Euro-Collins solutions Containing high concentrations of potassium, magnesium, phosphate, sulphate, and glucose. The solution is adequate for use in preserving the heart, liver, and lung.

Ross-Marshall citrate solutions Ross-Marshall citrate solutions were developed as alternatives to the Collins solutions. Their electrolytic compositions are similar except that citrate replaces phosphate, and mannitol replaces glucose. The citrate acts as a buffer and chelates with magnesium to form an impermeable molecule that helps in stabilizing the extracellular environment.

Phosphate-buffered sucrose solution This solution contains sucrose 140 mmol/L and sodium hydrogen and dihydrogen phosphate as buffers. In experimental studies, it preserve human kidneys for 3 days.

University of Wisconsin solution developed for liver, kidney, and pancreas preservation. The solution has an osmolality of 320 mmol/kg and pH 7.4 at room temperature and is composed of the following:

Composition of UW solution Potassium ……………………..135 mmol/L Sodium………………………….. 35 mmol/L Magnesium……………………. 5 mmol/L Lactobionate ………………...100 mmol/L Phosphate ……………………..25 mmol/L Sulphate ………………………..5 mmol/L Raffinose ……………………….30 mmol/L Adenosine ……………………..5 mmol/L Allopurinol ……………………1 mmol/L Glutathione …………………..3 mmol/L Insulin …………………………..100 U/L Dexamethasone …………....8 mg/L Hydroxyethyl starch (HES)50 g/L Bactrim …………………………0.5 ml/L

Celsior solution Celsior is a recently developed extracellular-type, low-viscosity. Composition: Sodium…………………………………….. 100 mmol/L Potassium…………………………………. 15 mmol/L Magnesium………………………………. 13 mmol/L Calcium …………………………………….0.25 mmol/L Lactobionate ……………………………..80 mmol/L Glutathione ……………………………….3 mmol/L Glutamate…………………………………. 20 mmol/L Mannitol ……………………………………60 mmol/L Histidine…………………………………… 30 mmol/L

Kyoto ET solution Researchers at Kyoto University developed a new solution that contains a high sodium concentration, a low potassium concentration, trehalose, and gluconate. Composition: Sodium ………………..100 mmol/L Potassium……………. 44 mmol/L Phosphate……………. 25 mmol/L Trehalose ……………..41 mmol/L HES ………………………30 gm/L Gluconate …………….100 mmol/L

Cryopreservation of liver: . Pathological area of P3's liver, showing numerous cavities reminiscent of ice crystal spaces. Scale bar = 40 microns. H&E (hematoxylin and eosin stain), Karnovsky's. "Island" of well-preserved cellular structure in the liver. H&E, Karnovsky's primary fixative. Scale bar = 40 microns.

Cryopreservation of kidney(Renal medula):

Cryopreservation of heart: Cross-section of myofibrils, displaying normal shape and density. Cardiac blood vessel with fundamentally preserved structure.

Needs and benefits of organ Cryopreservation: In organ transplantation to store the isolated organ until it s transplantation without damage.

Major problems in Cryopreser- vation : Vascular damage may be caused due ice formation. Fracture of frozen tissue may be due thermal stress during warming. Formation of cytoplasm glass due to ultra -rapid cooling.

References: Day J.G and McLellan M.R.(eds) (1995) Cryopreservation and freeze-drying protocols. Meth. Mol. Biol. 38: 254pp. Meryman, H.T., Ice Crystal Formation in Frozen Tissues. Lecture and Review Series, Naval Medical Research Institute, No. 53-3, 25-48,1953. Lillehei, R.C., et al., In vitro preservation of whole organs by hypothermia and hyperbaric oxygenation, Cryobiology, 1, 181-193, 1964. Toledo-Pereyra, L.H., and MacKenzie, G.H., Freezing of human kidneys, initial in vitro observations, The American Surgeon, 48, 232-236, 1982.

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