Nonobese diabetic/severe combined immunodeficient murine xenograft model for human uterine leiomyoma  Guiwen Wang, M.D., Hiroshi Ishikawa, M.D., Ph.D., Kunizui Sone, M.D., Tatsuya Kobayashi, M.S., J. Julie Kim, Ph.D., Takeshi Kurita, Ph.D., Makio Shozu, M.D., Ph.D.  Fertility and Sterility  Volume 101, Issue 5, Pages 1485-1492.e3 (May 2014) DOI: 10.1016/j.fertnstert.2014.01.054 Copyright © 2014 American Society for Reproductive Medicine Terms and Conditions

Figure 1 Sex steroid–dependent growth of the subrenal UL xenograft in NOD/SCID mice. Estrogen and P were SC injected into each mouse during the grafting procedure and sequentially injected every week, every 2 weeks, and every 3 weeks. The representative gross appearance and estimated volume of the UL xenografts are shown. The estimated volume of the xenografts was calculated as follows: (4/3π) × a × b × c, where π is the ratio of the circumference to the diameter, and a, b, and c represent each radius of an ellipsoidal body. (A) Every week administration of E and P; (B) every 2 weeks administration of E and P; (C) every 3 weeks administration of E and P; (D) no administration of E or P; and (E) a kidney immediately after xenograft implantaion. Data are presented as means ± SD. Fertility and Sterility 2014 101, 1485-1492.e3DOI: (10.1016/j.fertnstert.2014.01.054) Copyright © 2014 American Society for Reproductive Medicine Terms and Conditions

Figure 2 Gross appearance, volume, histology, and expression of ERα and PR of the UL xenografts in NOD/SCID mice. (A) Subrenal and SC xenografts were histologically evaluated by hematoxylin and eosin staining. The ERα and PR expression levels were evaluated by immunohistochemical analysis using specific antibodies. (B) Estimated volumes of the subrenal and SC UL genografts. Each xenograft contained 5.0 × 105 cells. Xenografts with double the number of cells consisted of 1.0 × 106 cells. Data are presented as means ± SD. *P<.05. Fertility and Sterility 2014 101, 1485-1492.e3DOI: (10.1016/j.fertnstert.2014.01.054) Copyright © 2014 American Society for Reproductive Medicine Terms and Conditions

Figure 3 A representative gene expression profile of the UL xenograft. Messenger RNA expression levels of 12 representative genes were measured by quantitative real-time PCR. The value of each transcript was divided by that of the ACTB transcript. Data are presented as the fold change divided by the value of myometrial tissue and as means ± SEM. Data were analyzed using the Mann–Whitney U test, and a P value <.05 was considered statistically significant. L = leiomyoma; LX = leiomyoma xenograft; M = myometrium. Fertility and Sterility 2014 101, 1485-1492.e3DOI: (10.1016/j.fertnstert.2014.01.054) Copyright © 2014 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 1 Subrenal xenografts in NOD/SCID and SCID mice. The UL xenografts increased in volume beneath the kidney capsule in response to exogenous sex steroid administration in both NOD/SCID mice (A, C, E) and SCID mice (B, D, F). The xenografts shown in a horizontal direction originated from the same leiomyoma tissue: A and B, C and D, and E and F, respectively. Fertility and Sterility 2014 101, 1485-1492.e3DOI: (10.1016/j.fertnstert.2014.01.054) Copyright © 2014 American Society for Reproductive Medicine Terms and Conditions