Animal
selection
Male SD/Wistar rats should be selected for the
study.
The age of animals should be between 7-8 weeks,
weighing 200 - 250 gm to start the animals.
Animals procurement & randomization
Male SD/Wistar rats, aged 7-8 weeks were procured
from CPCSEA registered vendor and were subjected to quarantined for one week.
After completion of quarantine, animals were randomly allocated into different
groups and acclimatized for 4 days. The animals received water and
standard rodent chow ad libitum for the entire course of the study.
Vancomycin (VCM) preparation
Vancomycin was dissolved in water for injection
(WFI) provided by the same company to prepare a solution of concentration of 50mg/mL. In normal animals, water for injection was injected intraperitoneally.
Drug and vancomycin
administration
·
Prepared vancomycin solution
was administered twice daily at 12-hourly intervals by the intraperitoneal
route.
·
The best dose of vancomycin is
200 mg/kg, b.i.d. of body weight for 10 days for the best induction of acute
kidney injury. If you go for 300 mg/kg b.i.d bodyweight for 10 days, this may
cause extremely severe kidney damage which means serum urea and creatinine
level may increase up to 12 to 15 times as compared to normal animals whereas
at the 200 mg/kg serum urea and creatinine level increases up to 4 to 6 times.
·
In the morning, Vancomycin
should be injected one hour after the drug administration.
·
The dose volume should be
maintained at 5 mL/kg, throughout the experimental period.
Acute Kidney Injury: Symptoms, Causes, Prevention, Risk factors, Diagnosis and Treatment
Care during vancomycin
preparation and injection
·
Vancomycin should be properly
dissolved in water for injection to make the concentration.
·
To properly mix the solution,
use vortex mixing.
·
Prepare the vancomycin
solution just before injection because it starts to degrade after some time
and vancomycin efficacy may reduce.
·
Before dosing, animals should
be properly acclimatized so that Vancomycin can be easily injected without any
complications.
·
The gap between test item
dosing and vancomycin injection should be a minimum of 1 hour.
·
The bodyweight of the animals
should be taken twice daily just before vancomycin induction because vancomycin
may cause a decrease in animal body weight.
Urine & blood collection
and necropsy
·
For urine collection, on day
11 at 09:00 AM animals are placed in metabolic cages after 12 hours of the last
dose of vancomycin.
·
When you place the animals in
metabolic cages, it is a totally new environment for the rat so they feel
scared and stressed and many doses of vancomycin cause kidney damage resulting
in very low urine production and less urine volume may be insufficient for a
urinalysis and biochemical analysis. So, to increase the rat's urine
production, normal RO water (20 mL/kg) must be administered at 0, 4 and 8 hours
after placing the animals in metabolic cages.
·
In the urine collection tube,
a small amount of preservatives (Sodium azide & Thymol crystal) should be
added to prevent bacterial growth because the bacterial growth in urine may alter
the urine parameters.
·
On day 12, in the morning 24 hours
urine will be collected and urinalysis and urine biochemistry was performed. For
Urinalysis, an Erba Urine analyser was used (ERBA LAURA) and a DEKA PHAN LAURA kit
was used for urinalysis. For biochemistry analysis, the ERBA EM200 biochemistry
analyser was used for urine biochemistry analysis. Urea and creatinine were
measured in urine which will additionally be
measured in the serum as well.
Acute Kidney Injury: Symptoms, Causes, Prevention, Risk factors, Diagnosis and Treatment
·
After urine collection,
animals were anaesthetized by using an isoflurane nose cone and blood was withdrawn
by the retro-orbital route. After blood collection, animals were euthanized with
150mg/kg thiopentone and bilateral kidneys will be excised and weighed. The left kidney was processed for histopathological parameters and the right kidney was
subjected to estimation of various oxidative stress and gene expression
parameters.
·
After blood collection, haematology
should be performed because kidney injury causes severe alteration in CBC parameters
mainly such as platelet count, RBC, WBC, haemoglobin etc.
Study Schedule
|
Vancomycin induced AKI |
|||
|
Days |
Date |
Weekdays |
Activities |
|
1 |
|
|
VCM induction in
all except normal control |
|
2 |
|
|
VCM induction in
all except normal control |
|
3 |
|
|
VCM induction in
all except normal control |
|
4 |
|
|
VCM induction in
all except normal control |
|
5 |
|
|
VCM induction in
all except normal control |
|
6 |
|
|
VCM induction in
all except normal control |
|
7 |
|
|
VCM induction in
all except normal control |
|
8 |
|
|
VCM induction in
all except normal control |
|
9 |
|
|
VCM induction in
all except normal control |
|
10 |
|
|
VCM induction in
all except normal control |
|
11 |
|
|
Bodyweight measurement |
Bodyweight and dose-volume recording format
|
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|
Groups |
Animal no. |
Body wt. (g) |
Dose-volume (mL) |
Body wt. (g) |
Dose volume (mL) |
Body wt. (g) |
Dose volume (mL) |
Body wt. (g) |
Dose volume (mL) |
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Test item weighing record format
|
|
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|||
|
Groups/ compound
name |
Conc. (mg/mL) |
Amount of drug
(mg) |
Vol of vehicle
(mL) |
Amount of drug
(mg) |
Vol of vehicle
(mL) |
|
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END OF DOCUMENTS
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