The advantages of adding hyperbaria, low flow perfusion, and protein colloid to hypothermia at 0° C for kidney preservation are described. The possibilities of longer preservation at higher temperatures are discussed

Dienst, S. G.

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Henry Ford Hospital Medical JournalVolume 18 | Number 2 Article 106-1970In Vitro Kidney Preservation TechniquesS. G. DienstFollow this and additional works at: https://scholarlycommons.henryford.com/hfhmedjournalPart of the Life Sciences Commons, Medical Specialties Commons, and the Public HealthCommonsThis Article is brought to you for free and open access by Henry Ford Health System Scholarly Commons. It has been accepted for inclusion in HenryFord Hospital Medical Journal by an authorized editor of Henry Ford Health System Scholarly Commons.Recommended CitationDienst, S. G. (1970) "In Vitro Kidney Preservation Techniques," Henry Ford Hospital Medical Journal : Vol. 18 : No. 2 , 149-152.Available at: https://scholarlycommons.henryford.com/hfhmedjournal/vol18/iss2/10Henry Ford Hosp. Med. Journal VoL 18, No. 2, 1970 NOTES AND COMMENTS In Vitro Kidney Preservation Techniques S. G. Dienst, M.D.* The advantages of adding hyperbaria, low flow perfusion, and protein colloid to hypothermia at 0° C for kidney preservation are described. The possibilities of longer preservation at higher temperatures are discussed. The preferred method of handling the donor kidney during transplanta-tion continues to be rapid cooling by flushing the kidney's vascular bed with cold solutions, then holding its temper-ature at 0° C by surface cooling. With certain adjuncts, this technique offers good preservation of renal function for eight hours. A longer period of reliable preservation would allow for transpor-tation and wider use of potential human donor kidneys. Reports of successful 24-hour preservation, using continuous plasma perfusion and oxygenation at temperatures of 8° to 10°C,' have en-couraged studies of these techniques and various modifications of them. Hypothermic preservation in its sim-plest form consist of flushing the kid-ney with cold isomotic electrolyte solu-tion such as Ringer's lactate. Heparin is added to prevent intravascular clot-ting. After the blood has been flushed from the vascular bed, the kidney is immersed in a partiaUy frozen saline solution or "saline slush." It is stored * General Surgery Division I I This work was sponsored in part by The McGraw Foundation. in this solution for periods as long as six to eight hours until anastomosed to the vessels of the recipient. In our ex-perience the kidney thus handled does not retain all of its potential function. Certain additives and adjuncts have improved the immediate response and final function of the kidney. Table I Agents added to initial kidney flushing solution for preservation at 0 C. ADDITIVES RATIONALE Potassium Ptiosphate (or Chloride) Magnesium Phenoxybenzamine Glucose and Citrate Hydrocortisone Cnlorpromazine (Thorazine! formal oncotic pressure Reduce extra-intracellular ion gradients To conserve mitochondrial ATP and DPN* Reduce arterial spasm Oxidative substrates Stabilize lysozome and cellular membranes • Adenosine Triphosphate and Diphosphopyridine Nucleotide Albumin or plasma is used to main-tain a normal oncotic pressure gradient between the extra and intracellular compartments. This minimizes cellular swelling during hypothermia. This is better accomplished with albumin* * In practice, concentrated "Albumisol" or other human albumin is added to the intra-cellular electrolyte solution to give a six gram percent protein solution. 149 Dienst than with dextran. Kidneys perfused with albumin solution at 0° C have an immediate uniform pink color when re-vascularized. On the other hand, kid-neys perfused with dextran in an elec-trolyte solution have a persistent bluish cast and do not warm rapidly, indicat-ing poor perfusion. Also, angiograms of the kidneys perfused with dextran indicate obstruction of small arteries, particularly in the medullary portion.^ Composition of the Perfusate Potassium phosphate or chloride is added to reduce the intra-extraceUular gradient of sodium and potassium. This is done to conserve the energy normally expended to maintain these gradients. Also, it has been shown that intracellu-lar potassium is rapidly lost from cells into cold perfusing solutions.^ Magne-siimi is added to reduce intracellular losses, to inhibit ATPase, and to mini-mize the loss of ATP and DPN from mitochondria. Free magnesium is also necessary for the activity of many res-piratory enzymes. Phenoxybenzamine is a long acting alpha-adrenergic block-ing agent used to prevent arterial spasm. Recent proponents for the use of pheno-xybenzamine now advocate giving it systemically to a cadaver donor. The use of phenoxybenzamine, magesium, and potassium is taken from the recent work of Collins, Bravo-Shugar-man and Teraski.* This "intracel-lular" solution has been used success-fully as a flushing solution but not yet as a continuously flowing perfusate. Hydrocortisone and chlorpromazine are sometimes added to reduce the breakdown of lysozome and cell mem-branes. Glucose is added as an anaero-bic supply of DPNH (hydrogen ions) through glycolysis. Citrate provides a Kreb's cycle substrate. For preservation longer than two hours, intermittent low flow perfusion and hyperbaria are carried out with the simple Swenko* apparatus. Until now, this perfusate has been a saline base solution with only four mEq/1 of mag-nesium and no added potassium. A flow of 200 milliliters per hour under three atmospheres of oxygen delivers 8 miUiliters of dissolved oxygen. This is more than is used by the human kidney of average weight at 0°C to 1 °C. Aside from increasing oxygen delivery, hy-perbaria somehow extends preservation through its pressure effect, possibly by minimizing cellular edema. The slow flow of perfusate also supplies constant substrate and lowers the very high PC02's which occur. The uniformly better results with perfusion and hyper-baria suggest that the kidney at 0°C is capable of respiration and sufficient energy production to keep its molecu-lar organization intact. Figure 1 shows a typical response of the BUN in a pa-tient after receiving a kidney stored under hyperbaria. Current Techniques During the 20 to 40 minutes of vas-cular anastomosis, the kidney is grad-ually warming toward the temperature in the operative field. The greatest tem-perature increase is evidenced in the kidney cortex, which normally has the highest metabolic rate. To prevent this warming during the ischemia period, a double wall silastic kidney cooling jacket has been designed. This allows for the circulation of cold solution around the kidney during anastomosis. * Swenko Incorporated, Minneapolis. 150 In Vitro Kidney Preservation Techniques Mg % 0 0 8 0 2 3 4 5 6 DAYS HYPOTHERMIA AND HYPERBARIA 8 HOURS Figure 1 Chart shows the ability to clear urea of the donor kidney stored under hyperbaria. Its ability to preserve function in the transplanted kidney is being evaluated in the laboratory. A human size cooling jacket* has been used during transplant of one cadaver (five hours total stor-age) and one living related kidney do-nation. Both patients had receive Man-nitol. There was prompt diuresis of over a liter during the first hour. Blood urea nitrogen and serum creatinines were normal within 36 hours and both kidneys later had above normal cre-atinine clearances. The use of the "intracellular" solu-tion before storage at 0°C* for a period of 24 hours has not been uniformly successful in the laboratory. After auto-transplantation, these canine kidneys perfused well and produced urine im-mediately. However, even the best re-sults show moderately elevated BUNs and creatinines which began to climb rapidly after five to seven days. The final specimens showed diffuse paren-chymal damage. Perfusion of the kidney at higher temperatures appears to be the method which is now evolving for longer preser-vation. Figure 2 outlines a system in-corporating oxygenation and dialysis of the perfusate. In this Extracorporeal circuit only cryoprecipitated plasma, after the method of Belzer,^  or heavily citrated plasma or serum wUl flow through the kidney for 24 hours or more. In our experience microclotting is an important factor in the vascular obstruction which is prone to occur. Also, perfusion pressures over 50 to 60 millimeters of mercury are associated with increasing edema and resistance to flow. Generally, low pressures and low temperatures (less than 8°C) yield the best perfusion. Effects of Temperature Temperature seems to be the most important variable in these experi-ments. With plasma perfusate, the tem-perature is limited to less than 14°C because plasma alone cannot transport the larger volumes of oxygen required at higher temperatures. This narrow range, 6°C to 14°C, has the disad-vantage of retarding respiration and 02 CO;: "5 '7~ PLASI l lA I' f /J ANE MEMBR /CUPROPH B R A N E / 7 T^J K Mn Na GLUCOSE AMINO AGIOS * Extracorporeal, New Jersey. PO4 Cl HCO3 KIDNEY PERFUSION Figure 2 Diagrammatic presentation of the method for perfusing kidneys at 10°C. The perfusate is serum or more commonly cryoprecipitated plasma which is continuously oxygenated and dialyzed. 151 Dienst phosphorylation and yet does not give the full stabilizing effect of keeping these systems near freezing. This com-promise may prove to be the limiting factor in the method. Current experiments are designed to measure respiration by oxygen con-sumption while varying the tempera-ture. The temperatures wUl be carried to between 20 °C and 28°C by adding red cells or free hemoglobin to the per-fusate for increased oxygen transport. - ConlrolleD level; of Melaholic Intermeiliates - Programmed lemperalure — Oxygen Delivery and Electrolyte KIDNEY PRESERVATION METHODS Figure 3 Effective preservation time which can be achieved with different techniques. Perfusion at physiological flow rates and moderately reduced temperatures appears to be the most promising method for reliable preservation beyond 12 hours. REFERENCES 1. Belzer, F. O., et al: Etiology of rising perfusion pressure in isolated organ perfusion. Ann Surg 168:382-91, Sept 1968. 2. Dienst, S. G., and Krieg, M. A.: Initial studies of isolated kidney perfusion. Henry Ford Hosp Med J 17:165-70, Fall 1969. 3. Keeler, R., et al: The problem of renal preservation. Brit J Urol 38:653-6, 1966. 4. Collins, G. M.; Bravo-Shugarman, M., and Terasaki. P. I . : Kidney preservation for trans-portation. Lancet 2:1219-22, Dec 1969. 152 Henry Ford Hospital Detroit, Michigan 48202 Address Correction requested-Forwarding and return postage guaranteed. iJjo Non-Profit Org. U. S. Postage P A I D Detroi t , Mich. Permit No. 6785 ro Pr in ted in U.S.A. Hi mt 

In Vitro Kidney Preservation Techniques

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In Vitro Kidney Preservation Techniques

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