Bioflavonoids attenuate renal proximal tubular cell injury during cold preservation in Euro-Collins and University of Wisconsin solutions.
BACKGROUND
Cold ischemia and reperfusion during kidney transplantation are associated with release of free oxygen radicals and damage of renal tubular cells. Bioflavonoids may diminish cold storage-induced injury due to antioxidant and iron chelating activities. This study was designed to delineate the renoprotective mechanisms of bioflavonoids and to define the structural features conferring cytoprotection from cold injury.


METHODS
LLC-PK1 cells were preincubated for three hours with bioflavonoids and cold stored in University of Wisconsin (UW)- or Euro-Collins (EC)-solution for 20 hours. After rewarming, cell viability was assessed by the lactate dehydrogenase (LDH) release, MTT-test, and amino acid transport activity. Lipid peroxidation was assessed from the generation of thiobarbituric acid-reactive substances.


RESULTS
Twenty-hours of cold storage of LLC-PK1 cells resulted in a substantial loss of cell integrity that was more pronounced in the EC (LDH release, 93.6 +/- 1.6%) than the UW solution (67.2 +/- 6.9%; P < 0.0001). Pretreatment with quercetin significantly enhanced cell survival (LDH release, 5.4 +/- 2.7% for UW and 8.4 +/- 4.2% for EC) in a concentration dependent manner. Structure-activity studies revealed similar renoprotection for kaempferol, luteolin and fisetin, unlike myricetin, morin, apigenin, naringenin, catechin, silibinin and rutin. Lipid peroxidation was reduced (UW alone, 2.7 +/- 1.2 vs. UW+quercetin 0.5 +/- 0.2 nmol/mg protein, P < 0.01), and l-threonine uptake completely sustained by pretreatment with quercetin, kaempferol, luteolin, and fisetin. However, renoprotection by fisetin was rapidly lost during rewarming. Protective properties of bioflavonoids were governed by the number and arrangement of hydroxyl substitutes, electron-delocalization, sterical planarity, and lipophilicity of the basic diphenylpyran skeleton.


CONCLUSIONS
Cold storage-induced renal tubular cell injury is ameliorated by bioflavonoids. Renoprotective effects of bioflavonoids are defined by structure, suggesting that flavonoids are incorporated into membrane lipid bilayers and interfere with membrane lipid peroxidation.
Ahlenstiel
Thurid
T
Department of Medicine, Division of Nephrology and Hypertension, University Hospital of Saarland, Homburg/Saar, Germany.
Burkhardt
Gunther
G
Köhler
Hans
H
Kuhlmann
Martin K
MK
D000241:Adenosine / Q000009:adverse effects*; D000493:Allopurinol / Q000009:adverse effects*; D026905:Amino Acid Transport Systems / Q000378:metabolism; D000818:Animals; D015925:Cryopreservation; D005419:Flavonoids / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology*; D005978:Glutathione / Q000009:adverse effects*; D006358:Hot Temperature; D006982:Hypertonic Solutions / Q000009:adverse effects*; D007328:Insulin / Q000009:adverse effects*; D007687:Kidney Tubules, Proximal / Q000378:metabolism / Q000473:pathology*; D007700:Kinetics; D018374:LLC-PK1 Cells; D015227:Lipid Peroxidation; D047311:Luteolin; D019188:Organ Preservation Solutions / Q000009:adverse effects*; D009994:Osmolar Concentration; D011794:Quercetin / Q000493:pharmacokinetics; D011887:Raffinose / Q000009:adverse effects*; D013329:Structure-Activity Relationship; D013552:Swine; D013997:Time Factors
D026905:Amino Acid Transport Systems; C037096:Euro-Collins' solution; D005419:Flavonoids; D006982:Hypertonic Solutions; D007328:Insulin; D019188:Organ Preservation Solutions; C055206:University of Wisconsin-lactobionate solution; D000493:Allopurinol; D011794:Quercetin; D005978:Glutathione; D000241:Adenosine; D047311:Luteolin; D011887:Raffinose