Harvesting Peritoneal Fat Depots: A Technique to Study Ovarian Cancer Colonization of Peritoneal Adipose Tissues in Mouse Models

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Preparing Animals for Experimental Studies

1. Allow animals to acclimate to new housing and environment, to recover from potential physiologic effects of transport and handling.
   Note: The following technique is applicable to all commercially available strains of mice. The number of animals to be used for an experiment is dependent on the study design and should be done with consultation from a statistician.

2. Identification and Isolation of Peritoneal Fat Depots

1. Euthanize animals via CO₂ overdose and a secondary physical method to confirm death. 
2. As harvesting peritoneal fat depots constitutes internal organ removal (secondary method), make a midline incision close to the inguinal papillae through the peritoneal wall to expose the internal organs (Figure 1A).
3. For the Omental Fat (OM), expose the omental/pancreatic complex by extending the spleen from the peritoneal cavity with forceps (Figure 1C). Release omentum from the pancreas and spleen by closely trimming all tissue connections. Ensure that any remaining pancreatic tissue is excised.
4. For the Gonadal Fat (GF), use forceps to lift the gonadal fat surrounding the ovaries and excise by cutting tissue connections (Figure 1A upper right).
5. For the Uterine Fat (UF), use forceps to lift the uterine fat surrounding the uterine horns and excise by cutting tissue connections (Figure 1A lower right).
6. For the Mesenteric Fat (MF), cut the junction between the small intestine and the pylorus. Use forceps to firmly grip the free end of the small intestine, and slowly "peel" it away from the mesenteric fat. Release the mesenteric fat from the mesenteric root using dissecting scissors (Figure 1A lower left).
7. For the Splenoportal Fat (SF), lift the distal end of the spleen using forceps to expose the thin fatty band of tissue connecting the hilum of the spleen to the pancreas. Excise the splenoportal fat by first releasing it from the pancreas and then carefully dissecting it from the spleen (Figure 1B).

Figure 1. The relative locations and structures of the main peritoneal adipose depots. (A) Gross anatomic dissection showing the relative location of four of the five primary sources of peritoneal fat. The omentum (OM; outlined), gonadal fat (GF), uterine fat (UF), mesentery (MY), ovary (ov), uterine horns (uh), and small intestines (si) are shown. (B) Splenoportal fat (SP; outlined) exposed by lifting the spleen with forceps. (C) The mouse omentum shown dissected free from the pancreas to improve visualization. (D) Relative sizes and structures of excised peritoneal fat; the mesentery is shown attached to the mesenteric root. (E) Histologic evaluation of peritoneal fat for the presence of milky spots – (A) adipocytes, (MS) milky spot.