1. Fertility and Sterility
Local application of danazol-loaded hyaluronic acid hydrogel to endometriosis in a rat model 
Kazuhito Nomura, M.D., Koichi Murakami, M.D., Ph.D., Makio Shozu, M.D., Ph.D., Tsuyoshi Nakama, Ph.D., Nobuhiko Yui, Ph.D., Masaki Inoue, M.D., Ph.D. 
Fertility and Sterility 
Volume 85, Pages (April 2006)
DOI: /j.fertnstert
Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

2. FIGURE 1
A [3H]-thymidine incorporation assay of cultured epithelial cells from rat endometrium. Rat endometrial epithelial cells were cultured after enzymatic digestion. Culture medium was changed to Dulbecco’s Modified Eagle’s Medium (DMEM)–F12 (1:1) containing 0.2% bovine serum albumin (BSA) at approximately 50% confluence. After 24 hours of incubation, culture medium was replaced with DMEM–F12 (1:1, phenol red free) supplemented with 5% dextran charcoal–treated fetal bovine serum (FBS) with or without sodium hyaluronate (1 mg/mL). Examined reagents were added to the medium, and incubation was continued for 24 hours. Incorporated [3H]-thymidine was counted 3 hours after addition of 1 μCi [3H]-thymidine. Each bar represents mean ± SE of three different experiments, each performed with cells in triplicate wells. (A) Dose-dependent effects of Danazol (DZ) on [3H]-thymidine incorporation. (B) Effects of E2 on [3H]-thymidine incorporation in DZ-treated epithelial cells. *P<.05 compared with control (Kruskal-Wallis test). **P<.01 compared with control (Kruskal-Wallis test).
Nomura. A DDS for endometriosis. Fertil Steril 2006.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

3. FIGURE 2
Microscopic appearance of endometrioma at 4 weeks after injection (HE stain, ×200; immunostaining for macrophage, ×200). (A) Control group: endometrial epithelial cells are cuboidal and contain normal volumes of cytoplasm. (B) Immunohistochemical staining of macrophages in the control group: many macrophages are found in the endometriotic stromal lesion beneath the epithelial cells. (C) Hyaluronic acid (HA) gel group: epithelial cells resemble those in the control group. (D) High-dose Danazol-loaded hyaluronic acid hydrogel (DZ-HA) gel group: endometrial epithelial cells show atrophic changes (decreased cytoplasm, perinuclear halo, and condensed nuclei). (E) Oophorectomized group: epithelial cells resemble those in DZ-HA gel groups.
Nomura. A DDS for endometriosis. Fertil Steril 2006.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

4. FIGURE 2
Microscopic appearance of endometrioma at 4 weeks after injection (HE stain, ×200; immunostaining for macrophage, ×200). (A) Control group: endometrial epithelial cells are cuboidal and contain normal volumes of cytoplasm. (B) Immunohistochemical staining of macrophages in the control group: many macrophages are found in the endometriotic stromal lesion beneath the epithelial cells. (C) Hyaluronic acid (HA) gel group: epithelial cells resemble those in the control group. (D) High-dose Danazol-loaded hyaluronic acid hydrogel (DZ-HA) gel group: endometrial epithelial cells show atrophic changes (decreased cytoplasm, perinuclear halo, and condensed nuclei). (E) Oophorectomized group: epithelial cells resemble those in DZ-HA gel groups.
Nomura. A DDS for endometriosis. Fertil Steril 2006.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

5. FIGURE 3
Immunohistochemical findings of endometrioma at 4 weeks after treatment (counterstained using hematoxylin, ×200). (A) Control group: scattered apoptotic cells contain ssDNA-positive condensed nuclei and perinuclear halos. (B) High-dose Danazol-loaded hyaluronic acid hydrogel (DZ-HA) gel group: endometrial epithelium with increased number of apoptotic cells, containing ssDNA-positive condensed nuclei and perinuclear halos. (C) Oophorectomized group: epithelial cells display atrophic changes but few apoptotic cells are apparent.
Nomura. A DDS for endometriosis. Fertil Steril 2006.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

6. FIGURE 4
Changes of endometrioma volume after Danazol-loaded hyaluronic acid hydrogel (DZ-HA) gel treatment. A 0.1-mL aliquot of HA gel or DZ-HA gel was injected after evacuation of cyst contents at week 0. Size of the endometrioma was measured weekly using micrometer calipers. Size at gel injection (week 0) was considered 100%. Each point represents mean ± SE of cyst size in each group. Control: untreated; OVX: oophorectomy conducted at week 0. *P<.05 compared with control (Scheffé). **P<.01 compared with control (Scheffé).
Nomura. A DDS for endometriosis. Fertil Steril 2006.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions