1.0 TEST SYSTEM DETAILS:
Species : Mus musculus
(Mice)
Strain : Swiss Albino or C57/bl6
Age : 6-10 weeks
Body weight : 18-22 g
Sex : Male
No. of animals : 8 Animals in each group
Total animals : 56 + 0 Extra (Total = 56)
|
Group No. |
Group Description |
Wound Generation procedure |
Treatment administered |
Quantity and Route |
|
G1 |
Excision wound without
Infection |
500 mm2
full-thickness skin will be exercised from the
dorsal interscapular without Infection |
No Treatment |
10 mg/mm2,
Topical application on the
wound |
|
G2 |
Excision wound with
Infection |
500
mm2 full-thickness skin will be exercised from the
dorsal interscapular region of rats and inoculated with the 0.1 ml S. aureus (approximately 109
CFU). |
No Treatment |
|
|
G3 |
Excision wound with
Infection vehicle control |
Treated with carrier gel |
||
|
G4 |
Reference Control |
Mupirocin ointment - once
daily |
||
|
G5 |
Treated with low
concentration of TFA2 |
TFA2:
3%, twice daily |
||
|
G6 |
Treated with intermediate
dose of TFA2 |
TFA2:
9%, twice
daily |
||
|
G7 |
Treated with high dose TFA2 |
TFA2:
27%, twice
daily |
2.0 METHOD:
Healthy animals will be selected for the study, randomized based on body weight, and will be assigned to 7 groups consisting of 8 animals each.
· Excision wounds without infection and with infection (animals assigned to group G1 and G2 respectively) will receive no treatment.
· Animals of group G3 will serve as Vehicle Control (Excision wounds without infection and with infection), receiving carrier gel used for formulating TFA2.
· Animals of group G4 will be treated with reference formulation-Mupirocin ointment-10 mg/mm2, once daily, topically. Treatment will be initiated immediately after wound generation.
· Animals of group G5-G7 will be treated with TFA2, at different incremental concentrations twice daily, topically. Treatment will be initiated immediately after wound generation.
· Animals allocated to groups G1-G7 will be anesthetized by the combination of (87.5 mg Ketamine + 12.5 mg Xylazine) by intraperitoneal route.
· The rats will be depilated on the back and a predetermined area of 500 mm2 full-thickness skins will be excised in the dorsal interscapular region and the created wound will be left undressed in the open environment.
· To confirm the infection, 0.1 ml of S. aureus (approximately 109 CFU) will be inoculated into the wound. After 24 hours, a bacteria sample will be collected using a sterile saline-moistened swab. The swab will be rolled around the infected wound area to ensure the establishment of S. aureus infection. The wound swab sample will then be inoculated onto nutrient agar for bacteriological examination. Cultures will be incubated at 37 degrees Celsius for up to 24 hours and checked for microbial growth.
· The formulation gels and standard topical preparation will be applied daily until complete healing is observed. In this model, wound contraction and epithelialization period will be monitored. Wound contraction will be measured as percent contraction every 3 days after wound formation using a vernier caliper. From the healed wound, a specimen sample of tissue of normal skin (Collected skin specimen on day 1st from the normal animals after their wound creation) and wounded (on the day of termination of study) site will be collected from each rat and will be fixed in 10% neutral buffered formalin for histopathological whereas some part of wounded skin will be stored at -80°C for the ensuing biochemical and molecular evaluations.
2.0
PARAMETERS TO BE EVALUATED:
·
Body
weight: Twice a week.
·
Skin oxidative
stress parameters: SOD, GSH: GSSG Ratio, MPO.
·
Gene expression
analysis in healed skin or normal skin (collected during wound generation) by Real-Time
PCR: MMP-2, MMP-9, TGF-β1, Col1a1, NF-kB.
·
Histological
analysis of wounded skin (H & E and MT).
·
Wound
contraction TIME
·
Epithelialization
period
·
Hydroxyproline
estimation
·
Estimation
of microbial count every 4th day
·
Protein
analysis by zymography (matrix metalloproteinases (MMPs)) of wound tissue
·
Histopathological
studies
3.0
REFERENCE(S):
1. Hemant Kumar et al.. Pharmacological Investigation of the Wound Healing Activity
of Cestrum nocturnum (L.) Ointment in Wistar Albino Rats. Hindawi Publishing
Corporation Journal of Pharmaceutics Volume 2016, Article ID 9249040, 8 pages
http://dx.doi.org/10.1155/2016/9249040.
2. Pawar RS et al. Wound healing activity of Sida cordifolia
Linn. in rats. Indian J Pharmacol 2013;45:474-8
3. Saleh, M. A., Shabaan, A. A., May, M. & Ali, Y. M. Topical application of indigo-plant leaves extract enhances healing of skin lesion in an excision wound model in rats. J. Appl. Biomed. 20, 124–129 (2022).
4. Rajoo A, Ramanathan S, Mansor SM, Sasidharan S. Formulation and evaluation of wound healing activity of Elaeis guineensis Jacq leaves in a Staphylococcus aureus infected Sprague Dawley rat model. Journal of Ethnopharmacology. 2021 Feb 10;266:113414.
END OF THE DOCUMENT
You may like to read these links:
1. List of OECD Guidelines or Toxicological Studies
2. List of Guidelines Used for Medical Devices Testing
3. What are Bradford Hill's criteria for any chemical identification?
4. Animal Facility Design - Small Laboratory Animals (Rat, Mice, Rabbit, Guinea Pig)
5. List of Chemicals Needed for In-vivo Laboratory
6. OECD Test Guidelines 425 detailedOutlines| AOT425StatPgm Software Installation LD50 Calculation
7. List of Guidelines for Toxicology Animal Studies
8. List of All SOPs and Documents for the Animal House Facility
9. List of All SOPs and Documents for In-vivo Laboratory
Hi. I’m Writer of Researchsop.com. ’ ’ Please share these SOPs to all concern pharma people for their development. I like to fullfill the need of curious people. These things inspire me to make things looks better.
0 comments:
Post a Comment