EVALUATION OF TEST FORMULATION ON THE PLATELET COUNTS IN CYCLOPHOSPHAMIDE-INDUCED THROMBOCYTOPENIC MODEL IN RAT
1.0 TEST SYSTEM DETAILS:
Species : Rattus norvegicus (Rat)
Age : 06-08 weeks
Sex : Male
No. of animals : 8 /Group
Total animals : 48
2.0 ALLOCATION OF GROUPS:
|
Group No. |
Group
Description |
Disease
Induction agent administered |
Treatment administered |
Dose
Volume and Route |
|
G1 |
Normal Control |
Normal Saline |
0.5% MC, p.o., q.d. |
5 ml/kg, p.o. |
|
G2 |
Disease Control |
Intraperitoneal Injection with
50 mg/day for 3 consecutive days. |
0.5% MC, p.o., q.d. |
|
|
G3 |
Treated orally with a low
dose of TF1 |
TF1 mg/kg, in 0.5% MC., q.d. |
||
|
G4 |
Treated orally with
intermediate dose 1 of TF1 |
TF1 mg/kg, in 0.5% MC., q.d. |
||
|
G5 |
Treated orally with
intermediate dose 2 of TF1 |
TF1 mg/kg, in 0.5% MC., q.d. |
||
|
G6 |
Treated orally with high
dose of TF1 |
TF1 mg/kg, in 0.5% MC., q.d. |
Abbreviations: MC-Methyl Cellulose, p.o.-per os. q.d.: quaque die, in die. X1, X2, X3, and X4 are the incremental doses of test formulations. The dose range will be from 10 mg/kg to 1000 mg/kg, q.d.
3.0 METHOD:
· After the completion of quarantine, healthy animals will be selected for the study. Subsequently, they will be randomized based on body weight and allocated into 10, different groups consisting of 8 animals each.
· Post-randomization, animals will be acclimatized for 5 days in an experimental room earmarked for the experiment.
· The animals assigned to the normal control group (G1) will be treated with normal saline i.p injection.
· Thrombocytopenic condition will be induced in group G2 and drug dose groups G3-G6.
· Animals of Groups G1 and G2, designated as Normal-control and disease-control respectively, will be administered 0.5% MC, p.o., q.d.
· Animals of group G2-G6 will be treated with cyclophosphamide (50 mg/kg) i.p for three consecutive days.
·Animals of group G3-G6 will be treated with TF1 orally at different dose levels ranging from 10-1000 mg/kg, q.d.
· On day 15, i.e. after 14 days of treatment of both formulations, animals will be sacrificed under an overdose of thiopentone anesthesia. After suitable anesthesia but before the animal dies, blood will be collected and serum will be separated for estimation of blood parameters. Immediately after the animal dies, the spleen will be harvested and weighed. One portion will be fixed in 10% neutral buffered formalin for histopathological analysis.
4.0 PARAMETERS TO BE EVALUATED:
· Body weight: Twice a week
· Weight of the spleen.
· Platelet count
· Platelet distribution width
· Mean platelet volume
· Platelet large cell ratio
· Bleeding and clotting time
· Histological analysis of all the spleen (Hematoxylin & Eosin-stained).
5.0 REFERENCES:
5.1 Hong NIE, Kong-yan, Zhang, Feng, Yang Wu YS, Zhou JY and Ye WC. Establishment of a mouse thrombocytopenia model induced by cyclophosphamide. Zoological research. 2009, Dec. 30(6):645-652.
5.2 Babu E, Basu D. Platelet large cell ratio in the differential diagnosis of abnormal platelet counts. Indian J Pathol Microbio. 2004, 47(2): 202-205.
5.3 Zhang Q, Peng HJ, Yu XB, Xu PL. Establishment of a mouse model of Cyclophosphamide-induced thrombocytopenia and determination of platelet function of this model. J Fist Mil Med Univ, 23(12): 1277-1282.
5.4 Sailor G, Hirani K, Parmar G, Maheshwari R, Singh R, Seth AK. Platelet augmentation potential of polyherbal formulation in cyclophosphamide-induced thrombocytopenia in Wistar rats. Folia Medica. 2021;63(1):67-73.
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