EVALUATION OF EFFICACY OF TEST FORMULATION(S) IN RAT MODEL OF Α-NAPHTHYL-ISOTHIOCYANATE (ANIT)-INDUCED CHOLESTASIS
1.0
TEST SYSTEM DETAILS:
Species : Rattus
norvegicus
Strain : Wistar/Sprague Dawley
Age : 7-8 weeks
Sex : Male/Female
No. of animals : 8 /Group
Total animals : 48
Study duration : 21 days
2.0
ALLOCATION OF GROUPS:
|
Group No. |
Group
Description |
Disease- Induction agent administered |
Treatment
administered |
Dose
Volume and Route |
|
G1 |
Normal Control |
Olive oil; p.o x 1 day |
0.5% MC, b.i.d. |
5 mL/kg, p.o. |
|
G2 |
Disease Control |
100 mg/kg of ANIT in Olive
oil; p.o. × 1 day |
0.5% MC, b.i.d. |
|
|
G3 |
Reference Control |
Ursodeoxycholic acid 100 mg/kg, q.d. in 0.5% MC |
||
|
G4 |
Treated with low dose of TF |
TF -X1
mg/kg, b.i.d. in 0.5% MC |
||
|
G5 |
Treated with intermediate dose of TF |
TF -X2
mg/kg, b.i.d. in 0.5% MC |
||
|
G6 |
Treated with high dose of TF |
TF -X3
mg/kg, b.i.d. in 0.5% MC |
Abbreviations: MC – Methyl Cellulose, p.o.- per os, b.i.d. –
bis in die, q.d.: quaque die. X1, X2 and X3 are defined as the incremental
doses of the Test formulations.
3.0
METHOD:
·
Healthy
animals will be selected, randomized based on body weight, and allocated into 6,
different groups consisting of 8 animals each.
·
Cholestasis
will be induced by oral administration of ANIT 100 mg/kg (dissolved in olive
oil) in animals allocated to G2-G6. G1 animals will serve as the normal control
and will receive olive oil orally.
·
Animals
of the Group G1 0.5% MC, p.o., q.d.
·
Disease
control (G2) animals will receive 0.5% MC, p.o., q.d.
·
Animals
of group G3 will be treated with reference drug Ursodeoxycholic acid 100 mg/kg,
p.o., q.d.
·
Animals
of group G4-G6 will be treated with Test Formulation at different dose levels ranging
from 10-1000 mg/kg, q.d., five days prior to and 10 days
post-ANIT administration.
·
On the last
day i.e. on day 11 post-ANIT administration, the animals will be euthanized after an overdose
of thiopentone. After suitable anesthesia but before the animal dies, blood
will be collected from the retro-orbital plexus, and after that liver tissue will
be collected. One lobe of the liver will be fixed in 10% neutral buffered
formalin for histopathology and the remaining part will be stored at -80°C
for biochemical and molecular evaluations.
4.0
END POINT PARAMETER(S):
·
Serum
hepatic biomarkers: AST, ALT, γGT, ALP, DB, TB, cholesterol, and TBA
·
Expression of TNF-α, TGF-β1, IL- 1β, IL-10, IL-18, TIMP-1, BCL2 and BCL2
associated X (BAX).
·
Protein
expression of NF-κB, Nrf2, HO-1, NLRP3, Caspase-1, and TLR4.
·
Fecal
Microbiota: Bifidobacterium, Lactobacillus, Enterococcus faecalis, E. Coli.
·
Histological Analysis of liver tissue (H&E and MT Staining).
5.0
REFERENCE(S):
1. Yu, H. et al. Efficacy of Zhuyu
Pill Intervention in a Cholestasis Rat Model: Mutual Effects on Fecal
Metabolism and Microbial Diversity. Front. Pharmacol. 12, 1–15
(2021).
2. Takitani,
K. et al. Altered expression of retinol metabolism-related genes in an
anit-induced cholestasis rat model. Int. J. Mol. Sci. 19, 1–14
(2018).
3. Chen, L. et
al. Paeoniflorin Protects against ANIT-Induced Cholestatic Liver Injury in
Rats via the Activation of SIRT1-FXR Signaling Pathway. Evidence-based
Complement. Altern. Med. 2021, (2021).
4. Yun, H. et
al. Pingkui Enema Alleviates TNBS-Induced Ulcerative Colitis by Regulation
of Inflammatory Factors , Gut Bifidobacterium , and Intestinal Mucosal Barrier
in Rats. 2020, (2020).
5. Ou, Q. Q. et al. Yinzhihuang attenuates ANIT-induced
intrahepatic cholestasis in rats through upregulation of Mrp2 and Bsep
expressions. Pediatr. Res. 79, 589–595 (2016).
END OF DOCUMENT
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