Evaluation of two endometriosis models by transplantation of human endometrial tissue fragments and human endometrial mesenchymal cells

Abstract

Background: The animal models of endometriosis could be a valuable alternative tool for clarifying the etiology of endometriosis.

Objective: In this study two endometriosis models at the morphological and molecular levels was evaluated and compared.

Materials and methods: The human endometrial tissues were cut into small fragments then they were randomly considered for transplantation into γ irradiated mice as model A; or they were isolated and cultured up to fourth passages. 2×10⁶ cultured stromal cells were transplanted into γ irradiated mice subcutaneously as model B. twenty days later the ectopic tissues in both models were studied morphologically by Periodic acid-Schiff and hematoxylin and eosin staining. The expression of osteopontin (OPN) and matrix metalloproteinase 2 (MMP2) genes were also assessed using real time RT-PCR. 17-β estradiol levels of mice sera were compared before and after transplantation.

Results: The endometrial like glands and stromal cells were formed in the implanted subcutaneous tissue of both endometriosis models. The gland sections per cubic millimeter, the expression of OPN and MMP2 genes and the level of 17-β estradiol were higher in model B than model A (p=0.03).

Conclusion: Our observation demonstrated that endometrial mesenchymal stromal cells showed more efficiency to establish endometriosis model than human endometrial tissue fragments.

Keywords: Endometriosis; Matrix metalloproteinase 2; Osteopontin; Stromal cells.

Figure 1

Phase contrast imaging of cultured human endometrial cells at first passage (A) and fourth passage (B).

Figure 2

Flow cytometric analysis of CD90+ cultured endometrial stromal cells at the end of fourth passage. The histogram is the representative of three independent experiments. The red line shows the isotype control.

Figure 3

The gross morphology of endometriosis lesions in model A (A) and model B (B). As the arrow shows the lesions have cystic morphology

Figure 4

Light microscopy of normal human endometrial tissue were observed (A &B), the endometrial glands (black arrow) with stromal tissue (S) were seen

Figure 5

The morphology of recovered endometriosis lesion obtained from γ-irradiated mice stained by hematoxylin and eosin in model A. The black arrows indicate the endometrial like glands within the subcutaneous tissue of mice with low (A) and high magnification (B-D). The yellow arrows show the stromal cells between glands like sections

Figure 6

The representative photomicrographs of recovered endometriosis lesion stained by hematoxylin and eosin in model B. A lot of gland sections are seen within the subcutaneous tissue (black arrow) and these gland like sections are prominent in model B with low magnification (A & B) and higher magnification (C & D). Yellow arrow head shows the stromal cells between gland sections with high magnification (C). The black arrow head shows the eosinophilic secretion within these gland like structure (D).

Figure 7

Photomicrograph of endometriosis lesions in model B stained with PAS method. It demonstrated the endometrial like glands which lined with cuboidal epithelial cells (black arrows) and PAS positive secretion within them (black arrow heads). The stromal cells between gland sections show the PAS positive reaction (yellow arrow

Figure 8

The expression ratio of OPN and MMP2 to GAPDH gene in normal endometrium as control1 and CD90+ mesenchymal stromal cells as control2 and in endometrial lesion in model A and model B. a: Significant differences with control1; b: Significant differences with control2; c: Statistically difference with model A (p=0.03

Figure 9

The gel electrophoresis of real time RT-PCR product. The ladder was 100bp. OPN: osteopontin; MMP2: metalloproteinase 2; GAPDH as internal control