1.Study Group

alain tenenbaum

N= 29 Wistar rats
Weight mean : 421g

Origin of animals : Biotério Central of Campus of Ribeirão Preto of São Paulo University
Maintenance : Experimental Pathology Lab of Pathology Department on Medicine School of Ribeirão Preto – USP
Animals keeping : polypropylen boxes
Animals food : basic diet of lab and water
Light conditions : natural light/dark cycles under controlled basic conditions
Experimentators : A.Tenenbaum,R.Mené,Kim
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2.Animals Sacrifice and Histologic Samples

microtome

The mice were euthanized in CO2 chambers.

  • Incision of the skin with a blade nº 23
  • Finding and isolation of the pretibial muscle using Metzenbaum scissors
  • section of tendons with a Mayo scissors.
  • Fixation of Portions of the muscles on sections of cork oak
  • Embedding using 50% formaldehyde solution - 10% buffered and 50% of pure paraffin for conventional optical microscopic study.
  • Staining Sections as routine histological process.
  • Microtome used : Leika RM 2155
  • Coloration of muscle slices with

-hematoxilin-eosin
-Masson´s trichrome
-and red picro-sirius.

  • Histologic examination was realized with a conventional microscope (Olympus BH-2).
  • Observation under polarized lampe
  • Photographic documentation was realized with a Leika DMR microscope joined to a digital camera Leika CD 300F and compatible PC.

3.Material

boxes rats   6 boxes with 5 rats/ box 

  • Animals were divided in 6 boxes, so that boxes from number 1 to 5 had 5 rats and box number 6 had 4 rats 
  • All animals were submitted to a walking evaluation by footprint impression on special paper , and previous immersion of back legs on a soap solution. 
  • Box 1 had the control group, that received the application of 0,1 ml saline solution ( NaCl 0,9%) on the right pretibial muscle. 
  • The mice of boxes 2 to 5 received 0,1 ml of Endopeels Carbolic Acid  on the same muscle as control group.

  • Box 6 was constituted of 4 rats, and each one received 0,5 ml of Endopeels carbolic acid injected on the subcutaneous layer. 

  • All the animals were submitted to walking evaluation before and after their respective injections.

4.Methods:Animals are sacrified after 10 days, 1 month, 3 months, 7 months

 

Box Number Number of Rats Product Injected Quantity Where Injected How Injected Days of Sacrifice
1 5 NaCl 0.9% 0.1 ml right pretibial muscle IM 10-30-90-120-205
2 5 Endopeel 0.1 ml right pretibial muscle IM 10-30-90-120-205
3 5 Endopeel 0.1 ml right pretibial muscle IM 10-30-90-120-205
4 5 Endopeel 0.1 ml right pretibial muscle IM 10-30-90-120-205
5 5 Endopeel 0.1 ml right pretibial muscle IM 10-30-90-120-205
6 4 Endopeel 0.5 ml above the right pretibial muscle SC     30-90-120-205

5.Control (NaCl 0.9%)

CONTROL Left Pretibial Right Pretibial after 0.1 ml NaCl 0.9% IM
  left leg right leg
    Nothing to declare for saline solution injection