CN102550541A

CN102550541A - Organ vitrification freezing and preserving fluid

Info

Publication number: CN102550541A
Application number: CN2010106013581A
Authority: CN
Inventors: 潘峰; 刘迪
Original assignee: Individual
Current assignee: Individual
Priority date: 2010-12-23
Filing date: 2010-12-23
Publication date: 2012-07-11
Anticipated expiration: 2030-12-23
Also published as: CN102550541B

Abstract

The invention relates to the field of medical organ preservation and more particularly discloses a vitrification freezing and preserving fluid for preserving organs for a long term. The organ vitrification freezing and preserving fluid is characterized by comprising dimethyl sulfoxide, acetamide, glycerol, ethylene glycol, propylene glycol, cane sugar, polyethylene glycol and the like, wherein the concentrations of the dimethyl sulfoxide, the acetamide, the glycerol, the ethylene glycol, the propylene glycol and the cane sugar are 0.1-6 mol/L; the concentration of the polyethylene glycol is 1-20 mmol/L; and the pH value is in a range from 5.0 to 8.0. By implementing the vitrification preserving fluid for preserving organs, provided by the invention, the preserving fluid is in a glassy state rather than forming ice crystal in the cooling and freezing process to protect the tissue morphology and structure completeness, completely suppresses energy metabolism of tissue cells under a freezing condition and recovers biological functions of organs after anabiosis. The organ vitrification freezing and preserving fluid, disclosed by the invention, can be used for freezing and preserving kidney and also can be further widely applied to freezing and preserving of other organs for a long term for scientific research and clinic application.

Description

The organ glass freezing is preserved liquid

Technical field

Kidney is the complicated organ of function in the human body, can only {in vitro} conservation under 4 ℃ of perfusion conditions maximum 50 hours.Will be for a long time even forever preserve kidney, must further reduce storage temperature, suppress histiocytic metabolic rate, even cellular metabolism is stopped fully.Kidney if can be preserved at suitable long period under refrigeration, transplants after when needed that the confession kidney employing of freezing preservation is the suitable means " recovery " again, will open up a brand-new prospect for clinical transplantation work.

Freezing and storing method is considered to realize the desirable approach of organ long preservation; Key of problem is that the organ volume is bigger; Ice crystal forms inside and outside in cooling and rewarming process, being difficult to overcome cell; Can not avoid simultaneously the damage of machinery, chemistry, osmotic pressure and biology aspect, so the frozen problem of larger volume organ is still unresolved so far.Vitrifying (Vitrification) is meant that liquid changes the solidification process of amorphous state (glassy state) into.Relation between the glassy solids molecule and the obviously difference of liquid nothing do not form ice crystal, thereby are a kind of desirable freezing preservation approach in the cooling refrigerating process.

Background technology

In the recent period, freezing preservation cell, sperm, embryo even ovary tissue are successively achieved success, but the freezing preservation of complex organ such as kidney but still is a difficult problem that does not break through as yet.The world today along with the continuous increase of kidney transplant technology rapid development and nephrotic's quantity, presses for a large amount of kidneys and stocks and supply.Clinical transplantation is with supplying kidney all preserving more than 0 ℃ at present, though the method comparative maturity, easy, the holding time is very limited.Though Chinese scholars sets about prolonging the kidney holding time from improving sides such as preserving liquid level and conserving appliance; Or in the limited holding time, improve tissue matching and work; Supply the reasonable utilization rate of kidney and the long-term surviving rate after the transplanting with raising, but still can not satisfy clinical demand.In addition, under the situation that donor organ lacks at home, owing to join type, patient's condition is not inconsistent and reasons such as region also can cause certain organ wasting of resources.Can in a country and even All Around The World scope, share human leucocyte antigen (HLA) (HLA) and join the kidney that type coincide; Make each receptor can both in time obtain optimized confession kidney? For Utopian like this hope; Have only through improving kidney preservation technology; Prolong confession kidney holding time to several months even several years, and set up " organ storehouse " (Organ Bank) and could finally realize.

Summary of the invention

Organ glass freezing according to the invention is preserved liquid; Form by compositions such as dimethyl sulfoxide (DMSO), acetamide, glycerine, ethylene glycol, propane diols, sucrose, polyethylene glycol; Dimethyl sulfoxide (DMSO), acetamide, glycerine, ethylene glycol, propane diols, sucrose concentration are 0.1-6mol/L; Polyethylene glycol concentration is 1-20mmol/L, and the pH value scope is 5.0-8.0.Glass freezing is preserved liquid solution in the cooling refrigerating process and is glassy state, does not form ice crystal, avoids or reduce the freezing infringement of cell, can protect cell structure and shape integrity, suppresses histiocytic energy metabolism simultaneously fully.Metabolic activity in cells is restored after suitable means recovery, can bring into play biological function again.Organ glass freezing according to the invention is preserved liquid, can be used for the freezing preservation (200--30 ℃) of kidney, and the long-term frozen that also can be applicable to other organs (liver, heart etc.) is preserved.

Embodiment

To preserve the perfusion kidney through organ glass freezing according to the invention, be cooled to-80 ℃, change profound hypothermia refrigerator or liquid nitrogen long preservation over to.Be warming up to 4 ℃ during the organ recovery, wash-out is preserved laggard line correlation morphology of liquid and function assessment inspection.

Preserve liquid at-80 ℃ of new zealand white rabbit kidneys of preserving for 2 weeks through organ glass freezing according to the invention, general form is not seen obvious freezing infringement, skin medullary substance clear in structure; The interior capillary button loop of glomerulus is complete under the light microscope; The slight swelling of renal cells, ultra microstructures such as glomerular filtration membrane, sertoli cell and mesangial cell are clear under the electron microscope, and the nuclei dyeing chromaticness is dense; Kytoplasm inner cell organ number is normal; Epithelial cell chamber face brush border microvillus marshalling, vascular endothelial cell and elastic membrane are complete, and demonstration kidney tissue morphology after freezing preservation is not seen obvious destruction.

Preserve liquid at-80 ℃ of new zealand white rabbit kidneys of preserving for 2 weeks through organ glass freezing according to the invention; Carry out phenolsulfonphthalein excretion test (PSP test), indicarminum measuring renal tubular function behind the ex vivo perfusion; PSP15 minute and 120 minutes excretion amounts are all near normal value (25%; 55%), sees blue urine at once after the indicarminum perfusion and discharge, show that kidney has the excretion function through freezing preservation metanephric tubule.

Preserve liquid at-80 ℃ of beasle dog kidneys of preserving for 2 weeks through organ glass freezing according to the invention; The autotransplantation of kidney postoperative carried out CDFI, emission single photon computed tomography (ECT) and the inspections such as (IVP) of vein secretion radiography in 1 month; Blood flow passes through in the color ultra visible transplanting kidney, and IVP and ECT show that renal secretion/filtering function continues to exist.

Claims

1. an organ glass freezing is preserved liquid, it is characterized in that, comprises compositions such as dimethyl sulfoxide (DMSO), acetamide, glycerine, ethylene glycol, propane diols, sucrose, polyethylene glycol.

2. preservation liquid according to claim 1 is characterized in that dimethyl sulfoxide (DMSO), acetamide, glycerine, ethylene glycol, propane diols, sucrose concentration are 0.1-6mol/L, and polyethylene glycol concentration is 1-20mmol/L, and the pH value scope is 5.0-8.0.

3. requiring 1 described preservation liquid according to economic rights, it is characterized in that, storage temperature is-200--30 degree centigrade.

4. according to the arbitrary described preservation liquid of claim 1 to 3, it is characterized in that described organ is kidney, liver, heart, lungs, skin or limbs etc.

5. an organ store method comprises organ is put into preservation liquid, it is characterized in that, comprises compositions such as dimethyl sulfoxide (DMSO), acetamide, glycerine, ethylene glycol, propane diols, sucrose, polyethylene glycol in the said preservation liquid.

6. store method according to claim 5 is characterized in that, organ is carried out glass freezing preserve.