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M S Ametani

Publications and source records attributed to M S Ametani.

10 recordsLinked to original sources
PubMed1999

Biphasic mechanism for hypothermic induced loss of protein synthesis in hepatocytes.

BACKGROUND: A complication in liver transplantation is increased clotting times due to inhibition of protein synthesis resulting from prolonged hypothermic preservation. Protein synthesis is also blocked in cold preserved hepatocytes. In this study, the mechanism of inhibition of protein synthesis in cold preserved hepatocytes was investigated. METHODS: Hepatocytes prepared from rat liver were cold preserved in University of Wisconsin solution for 4, 24, and 48 hr. Protein synthesis was measured as incorporation of radiolabeled leucine into acid precipitable proteins. Hepatocytes were treated with antioxidants (dithiothreitol, trolox or deferoxamine, nitric oxide synthase inhibitor (N(G)-monomethyl-L-arginine monoacetate), steroids (dexamethasone or methylprednisolone), methods to keep adenosine triphosphate high (aerobic storage), and cytoskeletal disrupting agents (cytochalasin D or colchicine). RESULTS: There was a 26% decrease in protein synthesis after only 4 hr of cold storage and a further 25% decrease at 24 hr. Antioxidants, elevated adenosine triphosphate, and N(G)-monomethyl-L-arginine monoacetate did not affect the rate of loss of protein synthesis. Protein synthesis was not due to inhibition of amino acid transport or lack of amino acids in the storage medium. Steroid pretreatment of hepatocytes had no effect on the loss of protein synthesis occurring in the first 4 hr of storage but did suppress the loss occurring during the next 44 hr of storage. Cytoskeletal disrupting agents, added to freshly isolated cells, inhibited protein synthesis. CONCLUSION: The mechanism of loss of protein synthesis in cold preserved liver cells is not mediated by: (1) oxygen free radical generation or improved by antioxidant therapy, (2) nitric oxide generation in hepatocytes, (3) an adenosine triphosphate-sensitive destruction of cell viability, and (4) decreased permeability of amino acids or loss of amino acids from the cells. Loss of protein synthesis due to hypothermic storage appears biphasic. The first phase, occurring within 4 hr of storage, may be the result of the effects of hypothermia on the cell cytoskeletal system and may be untreatable. The second phase, which occurs during the next 24 to 48 hr is sensitive to steroid pretreatment. This phase may be amenable to improved preservation methodology. Improved preservation of the liver may require the use of steroids to conserve protein synthetic capabilities.

Adenosine↗
PubMed1997

Preservation of bladder tissue in different solutions for varying times.

OBJECTIVES: To evaluate three popular storage media and the effect of 24-hour cold storage on bladder tissue. METHODS: Guinea pig bladders were stored in three solutions: UW solution (a media used for transplant organs), Reznikoff solution [cell culture medium], and Krebs' solution with and without aeration. RESULTS: Cell potassium and sodium concentrations and total tissue water (a measurement of cell swelling) are important parameters for evaluating tissue damage. Reznikoff solution and Krebs' solution without gases maintained tissues for 24 hours with the least tissue damage; these solutions require no special equipment or attention. Twenty-four hour uniterrupted aeration of Krebs' solution caused the greatest degree of cell swelling with possible redistribution of receptors and required adjustment and regulation of the preservation apparatus. UW solution induced dehydration of cells, required the longest recovery period after cold storage, and is far more expensive than the other solutions. CONCLUSIONS: Reznikoff solution caused consistent relative changes in smooth muscle receptors and was superior to aerated Krebs' and UW solutions for 24-hour bladder tissue storage. It is unnecessary to aerate Krebs' solution during 24-hour cold storage.

Adenosine↗
PubMed1997

The effect of calcium in the UW solution on preservation of the rat liver.

In this study we investigated the effect of calcium addition to the UW solution on the quality of the preserved rat liver as judged by normothermic isolated perfusion. Rat livers were cold stored in UW solution containing varying concentrations of calcium chloride (0, 0.5, 1.5, 5.0 mM) for periods of 0, 24 and 48 hours. At the end of the preservation period the livers were reperfused for 90 minutes at 37 degrees C with Krebs Henseleit Buffer. The quality of preservation was assessed by quantification of enzyme release, bile production and protein synthesis. The addition of 1.5 mM calcium to the UW solution suppressed the incidence of damage in the 24 hour cold stored liver similar to control livers (0 hours preserved). LDH release were significantly reduced from 22.1 +/- 7.3 (units/hr/g) in regular UW to 9.4 +/- 0.8 (units/hr/g) in UW plus 1.5 mM calcium. AST release also was suppressed by the addition of calcium to the UW. Bile production was enhanced by the addition of calcium; from 21.3 +/- 0.6 (mg/hr/g) in regular UW to 46.3 +/- 5.9 (mg/hr/g) in UW plus 1.5 mM calcium. Protein synthesis was reduced to 38% of control after 24 hr cold storage and was unchanged by the addition of calcium to the preservation solution. Although the addition of calcium to the UW solution improved the preservation of the 24 hour cold stored liver it did not offer the same degree of protection to the 48 hour preserved liver. Therefore, calcium addition may be one agent for improving preservation for short term cold storage of the liver but longer term storage will require other modifications as well.

Adenosine↗
PubMed1996

Effect of hypothermia on intracellular Ca2+ in rabbit renal tubules suspended in UW-gluconate preservation solution.

Altered cellular calcium (Ca) homeostasis may be important in mediating hypothermic injury in preserved kidneys. In this study the effect of hypothermic (5 degrees C) storage on ionized intracellular Ca concentration ([Ca]i) in rabbit tubules was examined using Indo-1. Tubules were stored up to 250 min in UW-gluconate solution containing either 0.0, 0.5, 1.5, or 5.0 mM Ca (yielding about 3.6, 62, 371, and 1,010 microM ionized solution Ca (Ca2+) at 5 degrees C, respectively). [Ca]i increased to about 1,600 nM within 1 min after suspension in UW solution followed by a decrease in [Ca]i during the subsequent 60 min in all groups, suggesting mitochondrial Ca sequestration. Thereafter, [Ca]i either 1) increased in tubules incubated with 1.5 and 5.0 mM Ca to levels greater than 2,500 nM; 2) decreased to about 800 nM in tubules incubated with 0.5 mM Ca and then remained stable; or 3) continued to decrease in tubules incubated with 0.0 mM added Ca to reach an apparent steady-state concentration of about 175 nM after 180 min of incubation. The early spike in [Ca]i was unaffected by adding EGTA (solution Ca2+ = 50 nM). Ryanodine eliminated the [Ca]i spike, indicating that cooling in UW-gluconate solution caused release of endoplasmic reticulum Ca. This study shows that [Ca]i initially increases after exposure to UW-gluconate solution and appears to be transiently buffered through intracellular, probably mitochondrial, sequestration. Saturation of cellular buffer mechanisms resulted in a sustained dependence of [Ca]i on extracellular Ca2+. These results support the hypothesis that the effect of Ca on kidney viability is related to solution-induced alterations in [Ca]i.

Adenosine↗
PubMed1993

Cytoprotective effect of glycine in cold stored canine renal tubules.

Reperfusion injury has been suggested to cause delayed graft function in renal transplantation. Methods to reduce reperfusion injury could lead to improved clinical renal transplantation. Glycine has been shown to suppress reperfusion injury in rabbit renal tubules and rat hepatocytes. In this study we have determined the effects of glycine on viability of isolated canine renal tubules. Renal tubules were cold stored at 4 degrees C under hypoxic conditions for up to 96 h in the UW solution and rewarmed to 37 degrees C for up to 2 h under oxygenated conditions to simulate reperfusion of an organ after cold static storage. Short-term storage (24 to 48 h) did not cause membrane injury (leakage of lactate dehydrogenase (LDH)) on rewarming. However, after 72 and 96 h cold storage reperfusion injury was evident and LDH leakage increased from about 25% to 59 +/- 3% and 71 +/- 2% at 72 and 96 h cold storage, respectively. The presence of 3 mM glycine in the reperfusion medium suppressed injury to cold-stored renal tubules. After cold storage for 72 and 96 h LDH leakage was reduced to control concentrations (31 +/- 3% and 29 +/- 1%, respectively). After cold storage for 72 h there was a reduction in ATP concentration in rewarmed renal tubules (3 nmol/mg protein at 48 h to 1.25 nmol/mg protein at 72 h). Also, there was a loss of mitochondrial functions including decreased stimulation of oxygen consumption by uncoupling of oxidative phosphorylation. Although glycine suppressed LDH leakage in renal tubules cold stored for 72 h it had no effect on the regeneration of ATP or mitochondrial functions, which remained depressed.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1990

Important components of the UW solution.

The UW solution for preservation of the liver, kidney, and pancreas contains a number of components, and the importance of each of these has not been fully resolved. In the studies reported here the importance of glutathione and adenosine is demonstrated in isolated cell models (rabbit renal tubules and rat liver hepatocytes) of hypothermic preservation and reperfusion and in dog renal transplantation. Glutathione in the UW solution is necessary for the preservation of the capability of the cell to regenerate ATP and maintain membrane integrity. Adenosine in the UW solution provides the preserved cell with substrates for the regeneration of ATP during the reperfusion period following cold storage. The omission of GHS from the UW solution results in poorer renal function in the 48 hr dog kidney preservation-transplant model. The role of other components of the UW solution is discussed including lactobionic acid; other impermeants; and the colloid, hydroxyethyl starch. It is concluded that the development of improved preservation solutions will require a more detailed understanding of the mechanism of injury due to cold storage and, once obtained, solutions more complex than the UW solution may be required for improved long-term storage of organs.

Adenosine↗
PubMed1985

Effects of short-term hypothermic perfusion and cold storage on function of the isolated-perfused dog kidney.

The isolated-perfused dog kidney was used as a model to measure the effects of short-term hypothermic preservation on renal function and metabolism. Kidneys were cold-stored in Collins' solution, hypotonic citrate, or phosphate-buffered sucrose for 4 and 24 hr, or were continuously perfused for 4 and 24 hr with a synthetic perfusate. Following preservation kidneys were perfused with an albumin-containing perfusate at 37 degrees C for 60 min for determination of renal function. The results indicate that many of the effects of short-term preservation on renal function in dog kidneys are similar to results reported for rat and rabbit kidneys. Cold storage for 4 hr resulted in a large decrease in GFR (57%), but only a small decrease in Na reabsorption (from 97 to 87%). Cold storage for 24 hr caused a further decline in renal function (GFR = 95% decrease, Na reabsorption = 49-64%). Results were similar for all cold storage solutions tested. Perfusion for 4 hr was less damaging to renal function than cold storage. The GFR decreased only 14% and urine formation and Na reabsorption were practically normal. After 24 hr of hypothermic perfusion, the GFR was reduced by 79%, urine flow was normal, and Na reabsorption was 78%. There were no obvious biochemical correlates (adenine nucleotides, tissue edema, or electrolyte concentration) with the loss of renal function during short-term preservation. The results suggest that the isolated-perfused dog kidney can be used to test the effects of preservation on renal function, and yields results similar to those obtained using small animal models.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Stimulation of ATP synthesis in hypothermically perfused dog kidneys by adenosine and PO4.

During continuous hypothermic perfusion of dog kidneys there occurs a gradual decrease in ATP from about 1.4 to 0.6 mumol/g wet wt after 5 days of preservation. The loss of ATP can be prevented by including both adenosine (10 mM) and PO4 (25 mM) in the perfusate. Under these conditions kidney cortex ATP levels were more than double control values--3.5 mumol/g wet wt. Both adenosine and PO4 were necessary since omission of one substance resulted in no net synthesis of ATP. Furthermore, these high levels of ATP were obtained only if adequate concentrations of adenosine were maintained during perfusion. Following 3 days of perfusion the adenosine level in the perfusate decreased to about 1 mM and under this condition ATP levels were low. Adenosine levels were maintained in the perfusate by two methods: (1) addition of fresh perfusate or (2) pretreatment of the kidney with the adenosine deaminase inhibitor--deoxycoformycin. The increased levels of ATP appear directly related to the availability of nucleotide precursors and the presence of inhibitors of the enzymes involved in the catabolism of nucleotides and nucleosides (PO4 and deoxycoformycin). Mitochondrial activity was similar in kidneys with high or low ATP levels following 5 days of preservation.

Adenine Nucleotides↗
PubMed1984

Effects of hypothermic perfusion of kidneys on tissue and mitochondrial phospholipids.

The changes in the level of phospholipids in kidney tissue and isolated mitochondria from dog kidneys perfused hypothermically (6-8 degrees C) for 1, 3, and 5 days were compared. Following 1 day of perfusion there was no change in total tissue phosphatidylserine (PS), a 25% decrease in the level of phosphatidylethanolamine (PE), and a 16% decrease in phosphatidylcholine (PC). No further decrease was observed with longer perfusion times. In fact, an increase in the level of PE occurred between the third and fifth days. Mitochondria isolated from perfused kidneys also showed a slight decrease in PE and PC following 1 day, no further change at 3 days, and an increase at Day 5. The loss of tissue phospholipids does not appear related to the viability of perfused kidneys. The major loss occurs within 1 day of perfusion and kidneys perfused up to 3 days are fully viable. Five-day perfused kidneys are nonviable, but show no greater loss of phospholipids than the viable 1- or 3-day perfused kidneys.

Animals↗