X-Message-Number: 30131
Date: Thu, 13 Dec 2007 21:23:18 -0800 (PST)
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Subject: glycerol induced extrinsic toxicity

[Since glycerol does not denature DNA or proteins, other mechanisms must be
operative in its toxicity. Alvarez (8407576) tested the effects of an
antioxidant on glycerol induced cryodamage. Vitamin E was found to be
ineffective, and it was concluded that osmotic injury was the dominant
mode of  glycerol induced cryodamage. However free radicals do
mediate glycerol induced renal failure, and vitamin E (10720895) is now
known to be ineffective in treating this. Astragalus (9387712), carnitine
(10720895), catechin (12967597), grape seed proanthocyanidins (10843237),
quercetin ( 15452363), resveratrol (16538975), and salvia miltorhiza
(9387712) are all effective in reducing glycerol induced renal
damage. Testing these antioxidants on glycerol induced cryodamage would be
a better test to determine if extrinsic toxicity is a significant factor
in cryodamage. Note that glycerol would have to be added extremely slowly
to eliminate confounding osmotic damage. Indeed it would be ironic if it
did turn out that glycerol toxicity was mediated solely by osmotic damage,
as this would mean that a potentially non-toxic vitrification agent has
been readily available all long, but everybody missed this fact, by always
adding glycerol too rapidly.]

J Androl. 1993 May-Jun;14(3):199-209.
Evidence that membrane stress contributes more than lipid peroxidation to
sublethal cryodamage in cryopreserved human sperm: glycerol and other
polyols as sole cryoprotectant.
  Alvarez JG, Storey BT. Department of Obstetrics and Gynecology, University
of Pennsylvania Medical Center, Philadelphia 19104-6080.
  One effect of cryopreservation on human sperm is sublethal cryodamage,
in which cell viability post-thaw is lost more rapidly at later times than
in fresh cells. We hypothesized two modes of sublethal cryodamage: one is
peroxidation-related involving plasma membrane damage due to lipid
peroxidation; the other is membrane stress-related involving membrane
embrittlement during phase transitions occurring during freeze-thaw. If the
peroxidation-related mode contributed substantially to sublethal cryodamage,
the hypothesis predicts that lipid peroxidation inhibitors should reduce
this damage. To test this prediction, we examined the effect of the lipid
peroxidation inhibitors, hypotaurine, bovine serum albumin (BSA), and
alpha-tocopherol (Vit. E) on the time to loss of motility (TLM), taken as a
measure of cell viability over time, for sperm samples cryopreserved in
glycerol plus egg yolk medium. These agents had no effect on TLM of these
samples, indicating that this mode contributes little to sublethal
cryodamage. If the membrane stress-related mode contributed, the hypothesis
predicts rapid recovery of motility in the presence of egg yolk plus
glycerol, but slow recovery in the presence of glycerol alone. It also
predicts that an appropriate polyol may be both necessary and sufficient for
cryopreservation. In the presence of egg yolk plus glycerol, motility
recovery was complete within 5 minutes, but the percent motile cells then
decreased linearly with time. With glycerol alone in the range 3-12%, at 5
minutes post-thaw the percent motile cells was 5-10%, but by 40 minutes
post-thaw had risen to 60-80%, approaching that in the fresh sample, and was
maintained up to 4 hours. In the absence of glycerol, the percentage of
motile cells post-thaw was nil and remained nil up to 4 hours. The polyols,
erythritol, ribitol, and sorbitol had similar effects to that of glycerol,
but the recovery of motility was not as complete. These results indicate
that the membrane stress-related mode contributes substantially to sublethal
cryodamage. They also indicate that glycerol and other polyols can function
alone as cryoprotectants, but that recovery of motility is slow in these
systems.
PMID: 8407576

Nephron. 2000 Mar;84(3):243-7.
Effect of vitamin E and pentoxifylline on glycerol-induced acute renal
failure.
  Akpolat T, Akpolat I, Ozt rk H, Sarikaya S, Cosar AM, Bedir A, Kandemir B.
Department of Nephrology, Ondokuz Mayis University School of Medicine,
Samsun, Turkey.
  The pathogenesis of acute renal failure may involve, among other causes,
ischemia, vascular congestion, arachidonic acid pathways, and reactive
oxygen metabolites. The aim of this study is to evaluate the effects of
pentoxifylline and vitamin E on the prevention of experimental acute renal
failure induced by glycerol. Eighty-five Sprague-Dawley rats weighing
170-230 g were included in the study. The rats were randomly divided into
four groups: group 1 was given 1 ml saline; group 2, glycerol; group 3,
glycerol plus vitamin E, and group 4, glycerol plus pentoxifylline. Extent
of histological renal tubular necrosis and regeneration in each animal were
graded. Blood urea nitrogen, serum creatinine, and creatine kinase
concentrations were measured. Mean blood urea nitrogen and serum creatinine
concentrations and tubular injury scores were significantly lower in group 1
than in groups 2-4 (p < 0.001), but there were no significant differences
among groups 2-4. We conclude that postinsult administration of vitamin E
and pentoxifylline does not have a beneficial effect on prevention and
severity of acute renal failure and that controlled, multicenter studies
involving a large number of patients are needed to clarify this subject.
Copyright 2000 S. Karger AG, Basel
PMID: 10720895

Clin Exp Pharmacol Physiol. 2006 Jan-Feb;33(1-2):119-24.
Protective effects of L-carnitine on myoglobinuric acute renal failure in
rats.
  Aydogdu N, Atmaca G, Yalcin O, Taskiran R, Tastekin E, Kaymak K.
Department of Physiology, Faculty of Medicine, University of Trakya, Edirne,
Turkey.
  1. Muscle injury (rhabdomyolysis) is one of the causes of acute renal
failure (ARF). Iron, free radicals and nitric oxide (NO) play a critical
role in the pathogenesis of glycerol-induced myoglobinuric ARF. L-Carnitine
is an anti-oxidant and prevents the accumulation of end-products of lipid
peroxidation. Therefore, the aim of the present study was to investigate the
effects of L-carnitine on myoglobinuric ARF induced by intramuscular (i.m.)
hypertonic glycerol injection. 2. Sprague-Dawley rats were divided into
three groups. Rats in group 1 (n = 8) were given saline, whereas those in
groups 2 (n = 10) and 3 (n = 10) were injected with glycerol (10 mL/kg,
i.m.). Concomitant with and 24 h after glycerol injection, L-carnitine (200
mg/kg, i.p.) was administered to group 3 rats. Forty-eight hours after
glycerol injection, blood samples and kidney tissues were taken from
anaesthetised rats. 3. Plasma creatine kinase (CK) activity, urea,
creatinine and NO levels, as well as kidney tissue superoxide dismutase
(SOD), catalase (CAT) and glutathione peroxidase (GPx) enzyme activity and
malondialdehyde (MDA) and glutathione (GSH) levels, were determined. In the
kidney tissue, histopathological changes and iron accumulation in the
tubular epithelium were also investigated. 4. Glycerol treatment caused
severe ARF: a marked renal oxidative stress, significantly increased CK
activity, urea and creatinine levels and decreased plasma NO levels.
Histopathological findings in group 2 rats confirmed that there was renal
impairment by cast formation and tubular necrosis and a marked increase in
iron accumulation in the tubular epithelium. All these factors were
significantly improved by L-carnitine supplementation. 5. These results may
indicate that L-carnitine treatment protects against functional, biochemical
and morphological damage and iron accumulation in glycerol-induced
myoglobinuric ARF in rats. In this model, the protective effect of
L-carnitine treatment may provide a new insight into the treatment of
rhabdomyolysis-related ARF.
PMID: 16445710

Ren Fail. 2006;28(2):161-9.
Protective effect of resveratrol, a polyphenolic phytoalexin on
glycerol-induced acute renal failure in rat kidney.
  Chander V, Chopra K. Pharmacology Division, University Institute of
Pharmaceutical Sciences, Panjab University, Chandigarh, India.
  Rhabdomyolysis-induced myoglobinuric acute renal failure (ARF) accounts
for about 10% to 40% of all cases of ARF. Reactive oxygen intermediates
have been demonstrated to play an etiologic role in myoglobinuric renal
failure.
This study was designed to investigate the effect of resveratrol, a
polyphenolic phytoalexin in glycerol-induced ARF in rats. Seven groups of
rats were employed in this study, group I served as control; group II was
given 50% glycerol (8 mL/kg, intramuscularly); groups III IV, and V were
given glycerol plus resveratrol (2 mg/kg, 5 mg/kg, and 10 mg/kg p.o. route,
respectively) 60 min prior to the glycerol injection; group VI received
L-NAME (10 mg/kg, i.p.) along with glycerol and resveratrol (5 mg/kg), group
VII animals received L-NAME (10 mg/kg) 30 min prior to glycerol
administration. Renal injury was assessed by measuring plasma creatinine,
blood urea nitrogen, creatinine, and urea clearance. The oxidative stress
was measured by renal malondialdehyde levels and reduced glutathione levels,
and by enzymatic activity of catalase, glutathione reductase, and superoxide
dismutase. Tissue and urine nitrite levels were measured as an index of
total nitric oxide levels. Glycerol treatment resulted in a marked
decrease in tissue and urine nitric oxide levels, renal oxidative stress,
and significantly deranged the renal functions along with deterioration of
renal morphology. Pre treatment of animals with resveratrol (5 and 10
mg/kg) 60 min prior to glycerol injection markedly attenuated the fall in
nitric oxide levels, renal dysfunction, morphologic alterations, reduced
elevated thiobarbituric acid reacting substances, and restored the
depleted renal antioxidant enzymes. This protection afforded by
resveratrol was significantly reversed by cotreatment of L-NAME along with
resveratrol, clearly indicating that resveratrol exerts its protective
effect through nitric oxide release along with the antioxidative effect in
glycerol-induced ARF.
PMID: 16538975

Pharmacol Res. 2003 Nov;48(5):503-9.
Catechin, a natural antioxidant protects against rhabdomyolysis-induced
myoglobinuric acute renal failure.
  Chander V, Singh D, Chopra K. Pharmacology Division, University Institute
of Pharmaceutical Sciences (U.I.P.S.), Punjab University, Chandigarh 160014,
India.
  Rhabdomyolysis-induced myoglobinuric acute renal failure accounts for
about 10-40% of all cases of acute renal failure (ARF). Reactive oxygen
intermediates have been demonstrated to play an etiological role in
myoglobinuric renal failure. This study was performed to explore the
protective effect of catechin-a natural antioxidant in an experimental model
of myoglobinuric ARF in rats. Four groups of rats were employed in this
study, group 1 served as control, group 2 was given 50% glycerol (8 ml
kg(-1), i.m.), group 3, glycerol plus catechin (40 mg kg(-1), p.o. for 4
days, twice a day) and group 4 was given only catechin (40 mg kg(-1), p.o.),
respectively. Renal injury was assessed by measuring serum creatinine, blood
urea nitrogen (BUN), creatinine, and urea clearance. The oxidative stress
was measured by renal malondialdehyde (MDA) levels, reduced glutathione
levels and by enzymatic activity of catalase, glutathione reductase (GR) and
superoxide dismutase (SOD). Glycerol administration resulted in a marked
renal oxidative stress, significantly deranged the renal functions as well
as renal morphology. All these factors were significantly improved by
catechin treatment. Catechin, due to its antioxidative activity, reduced the
toxicity of myoglobin in the renal tissues, and thus exerted a
renoprotective effect in this rhabdomyolysis mimicking model.
PMID: 12967597

Ren Fail. 2000 May;22(3):255-66.
Reversal of experimental myoglobinuric acute renal failure with
bioflavonoids from seeds of grape.
  Stefanovic V, Savic V, Vlahovic P, Cvetkovic T, Najman S, Mitic-Zlatkovic
M. Institute of Nephrology and Hemodialysis, University School of Medicine,
Nis, Yugoslavia.
  Rhabdomyolysis may account for about 10% of all cases of acute renal
failure (ARF). This study was performed to explore the protective
influence of proanthocyanidins from seeds of grape in an experimental
model of myoglobinuric ARF. Rats were injected with 50% glycerol (8 mL/kg,
im) followed immediately and daily in the next three days by ip
proanthocyanidins (20 mg/kg) or saline. After 96 h rats were sacrificed and
kidney morphology, kidney cortex peptidase activities, and malondialdehyde
(MDA) content were determined. A moderate renal failure was produced by
glycerol injection with blood urea of 31.8+/-11.0 vs. 7.68+/-0.24 m mol/L,
and serum creatinine of 153. +/-38.2 vs. 39.6+/-9.0 micromol/L, in
glycerol-induced ARF vs. control rats, respectively. Rats that received
proanthocyanidins in addition to glycerol had significantly lower (p < 0.01)
blood urea and serum creatinine levels compared to those receiving glycerol
alone. These functional differences between the glycerol and glycerol plus
proanthocianidins groups were also confirmed histologically. Kidney cortex
dipeptidylpeptidase IV (DPP IV) activity was not significantly changed in
glycerol-induced ARF, however, markedly increased after proanthocyanidins
treatment. Kidney cortex malondialdehyde content was found significantly
increased in glycerol-induced ARF over control level, and was markedly
reduced by proanthocyanidin treatment. Taken together, these results provide
strong evidence for the protective role of proanthocyanidins from seeds of
grape in glycerol-induced ARF. The effect is probably due to the antioxidant
activity of proanthocyanidins and to increased expression of kidney cortex
DPP IV with effective degradation of TNF-alpha. This may provide therapeutic
opportunities of preventing and/or treating myoglobinuric ARF in humans.
PMID: 10843237

Pharmacology. 2005 Jan;73(1):49-56. Epub 2004 Sep 27.
Reversal of experimental myoglobinuric acute renal failure in rats by
quercetin, a bioflavonoid.
  Chander V, Singh D, Chopra K. Division of Pharmacology, University
Institute of Pharmaceutical Sciences, Panjab University, Chandigarh, India.
  The occurrence of acute renal failure (ARF) following rhabdomyolysis has
been put at between 10 and 40% of cases, and accounts for between 3 and 15%
of all cases of ARF. Reactive oxygen intermediates have been demonstrated to
play an etiological role in myoglobinuric renal failure. This study was
performed to explore the protective effect of quercetin, a bioflavonoid, in
an experimental model of myoglobinuric ARF in rats. Four groups of rats were
employed in this study: group 1 served as control, group 2 was given 50%
glycerol (8 ml/kg, i.m.), group 3 was given glycerol + quercetin (2 mg/kg,
i.p.), and group 4 was given glycerol + DMSO (the solvent for quercetin, 5
ml/kg, i.p.). Renal injury was assessed by measuring serum creatinine, blood
urea nitrogen, creatinine and urea clearance. The oxidative stress was
measured by renal malondialdehyde levels, reduced glutathione levels and by
enzymatic activity of catalase, glutathione reductase, and superoxide
dismutase. Glycerol administration resulted in a marked renal oxidative
stress, significantly deranged the renal functions as well as renal
cytoarchitecture. All these factors were significantly improved by quercetin
treatment. Because of its radical-scavenging and iron-chelating properties,
quercetin protected the kidney against the glycerol-induced oxidative stress
and resultant renal dysfunction. Based on these results, this study confirms
the role of oxidative stress and demonstrates the renoprotective potential
of quercetin in this rhabdomyolysis-mimicking model. 2005 S. Karger AG,
Basel.
PMID: 15452363

Zhonghua Wai Ke Za Zhi. 1996 May;34(5):311-4.
[Experimental study of the protective effects of astragalus and salvia
miltiorrhiza bunge on glycerol induced acute renal failure in rabbits]
[Article in Chinese]
  Hu L, Yu T, Jia Z. Department of Urological Surgery, General Hospital of
Lanzhou, Peoples Liberation Army.
  The astragalus and salvia miltiorrhiza bunge alcohol extracts were used in
preventive treatment of glycerol induced acute renal failure (ARF) in
rabbits. The experimental rabbits were divided: astragalus group (AB),
salvia miltiorrhia bunge group (SM), two extracts mixture group (ABSM), and
normol saline control group (C). The extracts, mixtures and normol saline
were administrated before and after the induced ARF. The counts of Na, K,
creatinine in urine and blood and urinary AAP, NAG were determinated during
24-14 days periodically in all 4 groups, and the renal tissues were taken
from same periods for pathomorphological studies by microscopy and
transmission electromicroscopy. The damage of the glycerol induced ARF was
not only in the convolute tubules but also in the glomerulus. The glomerulor
filtration rate reduced abruptly, and oligouria or auria developed. The
study of renal functions and renal morphogy showed that the AB and ABSM
groups were damaged more lesser and promptely repaired than the SM and C
groups in the early stage. No glomerular sclerosis was noted in the AB, ABSM
groups in later stage, but it occurred sporadically or diffusely in the SM,
C groups. Therefore, astragalus is an ideal protective drug of traditional
chinese medicine for ARF.
PMID: 9387712

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