Simvastatin protects against the development of endometriosis in a nude mouse model

Abstract

Context: Endometriosis is a common condition associated with infertility and pelvic pain in women. Recent in vitro studies have shown that statins decrease proliferation of endometrial stroma (ES) and inhibit angiogenesis.

Objective: The aim was to evaluate effects of simvastatin on development of endometriosis in a nude mouse model.

Methods: Proliferative phase human endometrial biopsies were obtained from healthy donors and established as organ cultures or used to isolate ES cells. To establish endometriosis in the nude mouse, endometrial tissues were maintained in 1 nm estradiol (E) for 24 h and subsequently injected into ovariectomized nude mice. Mice (n = 37) were treated with E (8 mg, SILASTIC capsule implants; made in author laboratory) alone or with E plus simvastatin (5 or 25 mg/kg x d) for 10 d beginning 1 d after tissue injection (from three donors). Mice were killed and examined for disease. Effects of simvastatin on matrix metalloproteinase-3 (MMP-3) were evaluated in cultures of ES cells.

Primary outcome: The number and size of endometriotic implants were measured.

Results: Simvastatin induced a dose-dependent decrease of the number and size of endometrial implants in mice. At the highest dose of simvastatin, the number of endometrial implants decreased by 87%, and the volume by 98%. Simvastatin also induced a concentration-dependent decrease in MMP-3 in the absence and presence of inflammatory challenge (using IL-1alpha).

Conclusions: Simvastatin exerted a potent inhibitory effect on the development of endometriosis in the nude mouse. Mechanisms of action of simvastatin may include inhibition of MMP-3. The present findings may lead to the development of novel treatments of endometriosis involving statins.

Figure 1

Gross (A–E) and microscopic (F–J, hematoxylin and eosin staining) photomicrographs of experimental endometriosis established by proliferative phase human endometrium in nude mice. All mice were implanted with a slow-release estradiol capsule before introduction of human tissues. Mice were killed 4 h to 10 d after human tissue injection. Results are representative of three separate experiments using three different human biopsies. Original magnification: gross, ×15; microscopic, ×40.

Figure 2

Gross morphology of experimental endometriosis established by proliferative phase human endometrium in nude mice. All mice were implanted with a slow-release estradiol capsule before introduction of human tissues. Mice were treated by gavage with vehicle (A), 5 mg/kg simvastatin (B), or 25 mg/kg simvastatin (C). D–F, Hematoxylin and eosin stains of tissues from each treatment group (D, vehicle; E, 5 mg/kg simvastatin; and F, 25 mg/kg simvastatin). Results are representative of three separate experiments using three different human biopsies. Original magnification: ×100. Arrows point to endometrial implants.

Figure 3

Effects of simvastatin on the number and volume of lesions per mouse; each bar represents mean ± sem. Means with no superscripts in common are significantly different (P < 0.05). The figure summarizes three separate experiments on 37 mice: 13 in the control group, 12 treated with simvastatin at 5 mg/kg, and 12 treated with simvastatin at 25 mg/kg.

Figure 4

Western analysis of expression and regulation of MMP-3 protein in human endometrial stromal cells isolated from proliferative phase endometrial biopsies (n = 5). Cells were cultured for 48 h in the presence of estradiol (E, 1 nm), followed by an additional 24 h with E alone or E plus simvastatin (Sim) at 1 or 10 μm. Some cultures were also exposed to IL-1α during the last 6 h of culture. A representative experiment is shown in top panel. Densitometric analysis of all studies is shown in bottom panel. *, Means significantly different from control; †, means significantly different from IL-1α.