Evaluation of Ki-67 antigen expression in the vaginal epithelium of castrated female rats treated with raloxifene and tamoxifen: a comparative study.

OBJECTIVE: The aim of this study was to evaluate the effects of raloxifene and tamoxifen on Ki-67 antigen expression in the vaginal epithelium of castrated rats. MATERIAL AND METHODS: Thirty-nine virgin, adult, castrated female Wistar-Hannover rats were randomly divided into three groups: Group I (control, n = 13), Group II (raloxifene, n = 13) and Group III (tamoxifen, n = 13). After confirmation of their hypoestrogenic state, the rats were given 0.5 ml of propylene glycol (vehicle), 750 µg of raloxifene or 250 µg of tamoxifen, respectively, by gavage, for 30 days. On the 31st day, the rats were euthanized and their vaginas removed and fixed in 10% buffered formalin for of Ki-67 immunohistochemical evaluation. Data were analyzed using Levene's test and Tukey's method (p < 0.05). RESULTS: Mean Ki-67 expression in groups I, II and III was 27 ± 2.6, 32.3 ± 1.9 and 43.7 ± 3.5, respectively. In Group III (tamoxifen), there was a greater proportion of stained cells compared to Groups I and II (p < 0.0003), with no statistically significant difference between Groups I and II (p = 0.3626). CONCLUSIONS: The present results show that tamoxifen significantly increased cell proliferation in the vaginal epithelium of the castrated rats and no difference between the raloxifene and control groups.

Effects of raloxifene on the mammary epithelium of female rats in persistent estrus.

OBJECTIVE: The aim of this study was to evaluate the morphological and morphometric alterations produced by raloxifene in the mammary epithelium of female rats in persistent estrus. MATERIALS AND METHODS: Twenty-one female Wistar-Hannover rats in persistent estrus induced by 1.25 mg of testosterone propionate were randomly divided into two groups: Group I (n = 10), receiving only water and used as control; Group II (experimental, n = 11), treated with 3 mg of raloxifene daily for 21 days. The first abdominoinguinal pair of mammary glands was extirpated and processed for morphological and morphometric evaluation. The data were statistically analysed using Student's t-test (p < 0.05). RESULTS: Morphology revealed signs of epithelial atrophy, and morphometry showed a significant reduction in the mean number of ducts and alveoli in the experimental group (12.82 +/- 0.42 and 2.91 +/- 0.53, respectively) when compared with the control group (28.70 +/- 1.15 and 7.20 +/- 0.57, respectively). This difference was statistically significant, both for the ducts and for the alveoli (p < 0.001). CONCLUSION: The present results indicate that, at the dose and during the time of treatment used, raloxifene induced atrophy of the mammary epithelium of rats in persistent estrus.

Vascular endothelial growth factor expression in the urethral epithelium of castrated adult female rats treated with tamoxifen.

OBJECTIVE: The aim of this study was to evaluate the effects of tamoxifen on vascular endothelial growth factor (VEGF) expression in the urethral epithelium of castrated rats. MATERIALS AND METHODS: Thirty-six adult, castrated, female Wistar-Hannover rats were randomly divided into two groups: group I (n = 16, control), receiving only propylene glycol, and group II (n = 20, tamoxifen), treated with 250 microg/day of tamoxifen for 30 consecutive days by gavage. On the 31st day, the animals were sacrificed and the urethras were immediately removed, separated into the proximal and distal segments and processed for VEGF immunohistochemistry. The data were analysed using Student's t-test (p < 0.05). RESULTS: The mean percentage of VEGF expression in the epithelium of the proximal urethra of the animals in groups I and II was 64.47+/-3.70 and 74.69+/-3.03, respectively (p < 0.03), whereas the mean percentage of VEGF expression in the distal urethral epithelium of the animals in groups I and II was 53.49+/-4.64 and 68.57+/-3.67, respectively (p < 0.01). CONCLUSIONS: Our results indicate that, at the dose and during the time of treatment used, tamoxifen increased VEGF expression in the urethral epithelium of castrated rats.

Evaluation of insulin-like growth factor-I in postmenopausal women with breast cancer treated with raloxifene.

BACKGROUND: The objective of this study was to evaluate serum IGF-I levels in postmenopausal women with breast cancer treated primarily with raloxifene. METHODS: Twenty-two postmenopausal patients with operable, stage I or II, estrogen receptor-positive carcinomas participated in this study. Following confirmation of diagnosis, the patients received 60 mg of raloxifene for 28 days prior to definitive surgery. Blood samples were collected for evaluation of serum IGF-I levels prior to initiating medication and following a 28-day treatment course. Student's t-test for paired samples was used in the statistical analysis. Significance was established at p < 0.05. RESULTS: Mean serum IGF-I levels pre- and post-raloxifene treatment were 143.7 +/- 9.7 ng/ml and 94.8 +/- 7.6 ng/ml, respectively. This reduction in serum IGF-I levels following treatment with raloxifene was statistically significant (p < 0.001). CONCLUSION: Raloxifene significantly reduced serum IGF-I levels in postmenopausal women with breast cancer.

Morphological and morphometric analysis of the effects of intralesional tamoxifen on keloids.

The aim of this study was to evaluate the effect of intralesional tamoxifen on keloids, particularly on the concentration of fibroblasts, dermal inflammatory infiltrate, and collagen degeneration. A prospective study was carried out to evaluate keloids in 13 patients of both genders pre- and post-treatment with intralesional tamoxifen. Two samples of keloid lesions were obtained by 4-mm punch biopsies during the study: the first at the time of diagnostic confirmation of keloid and the other eight weeks later at the end of intralesional tamoxifen treatment. The biopsy samples were placed in 10% buffered formalin for HE staining and morphological and morphometric study. The degree of collagen fiber reduction and inflammatory infiltration were analyzed. Student's t-test was used for statistical analysis of the mean number of fibroblasts before and following tamoxifen treatment ( P < 0.05). The degree of collagen fiber reduction and inflammatory infiltration were absent before treatment and present in 100% of cases after treatment, while the mean number of fibroblasts was significantly lower after intralesional tamoxifen treatment ( P < 0.0001). We conclude that intralesional administration of tamoxifen promoted an inflammatory stimulus and collagen fiber reduction as well as a significant reduction in the number of fibroblasts that produce collagen. Impact statement Effective treatment of keloid that is a commonly recurrent dermatosis is very difficult, even after standard treatment. Standard treatment consists of partial resection of the lesion (shaving excision), in addition to local corticosteroid injection. Therefore, there is interest in alternative forms of topical treatment, e.g., selective estrogen receptor modulators, particularly tamoxifen has demonstrated in vitro studies to be a promising drug. Nevertheless, there is scarcity of publications on the effects of intralesional tamoxifen on keloids have been found, leading us to the conception of the present study. In this study, tamoxifen has proven to be an interesting alternative drug for the topical treatment of keloid, allowing us to conclude that the intralesional application of tamoxifen in keloids promotes a variable but ever-present inflammatory stimulus, associated with intense reduction of collagen fiber, in addition to a significant decrease in the number of fibroblasts that produce collagen and are involved in disease maintenance.

Effect of a gel containing pilocarpine on vaginal atrophy in castrated rats.

OBJECTIVES: To evaluate the effect of Carbopol gel formulations containing pilocarpine on the morphology and morphometry of the vaginal epithelium of castrated rats. METHODS: Thirty-one female Wistar-Hannover rats were randomly divided into four groups: the control Groups I (n=7, rats in persistent estrus; positive controls) and II (n=7, castrated rats, negative controls) and the experimental Groups, III (n=8) and IV (n=9). Persistent estrus (Group I) was achieved with a subcutaneous injection of testosterone propionate on the second postnatal day. At 90 days postnatal, rats in Groups II, III and IV were castrated and treated vaginally for 14 days with Carbopol gel (vehicle alone) or Carbopol gel containing 5% and 15% pilocarpine, respectively. Next, all of the animals were euthanized and their vaginas were removed for histological evaluation. A non-parametric test with a weighted linear regression model was used for data analysis (p<0.05). RESULTS: The morphological evaluation showed maturation of the vaginal epithelium with keratinization in Group I, whereas signs of vaginal atrophy were present in the rats of the other groups. Morphometric examinations showed mean thickness values of the vaginal epithelium of 195.10±12.23 µm, 30.90±1.14 µm, 28.16±2.98 µm and 29.84±2.30 µm in Groups I, II, III and IV, respectively, with statistically significant differences between Group I and the other three groups (p<0.0001) and no differences between Groups II, III and IV (p=0.0809). CONCLUSION: Topical gel formulations containing pilocarpine had no effect on atrophy of the vaginal epithelium in the castrated female rats.

Does Leishmaniasis disease alter the parenchyma and protein expression in salivary glands?

Leishmaniasis is considered a serious public health problem in several regions in Brazil and worldwide. This research aimed to perform a histopathological and proteomic study of parotid, submandibular and sublingual glands of BALB/c mice infected by Leishmania (L) infantum chagasi using histological, immunohistochemical and epifluorescence techniques. Twelve isogenic BALB/c male mice, around six- to eight-weeks old, were separated into two groups: the animals of the control group were injected with 0.15 ml of NaCl, while those in the experimental group were inoculated with 5 × 10(6) amastigote forms of Leishmania (L) infantum chagasi by the ip route. After 50 days, animals were euthanized and major salivary glands were collected to perform histological, immunohistochemical and epifluorescence techniques using anti-Caspase-2, anti-Ki-67 and anti-ß-catenin antibodies, respectively. The histological and morphometric evaluation showed clusters of mononuclear inflammatory cells and a higher area and perimeter of the parotid gland. However, none of the salivary glands had morphophysiological impairment. There was no immunoreactivity to the anti-caspase-2 antibody and Ki67 expression in acinar and ductal cells in both groups. According to the immunofluorescence staining, the ß-catenin antibodies did not show nuclear expression, suggesting no uncontrolled proliferation. The data obtained in this study showed population and morphological stability of major salivary glands after 50 days post-infection by Leishmania (L) infantum chagasi.

Evaluation of bone metastases from breast cancer by bone scintigraphy and positron emission tomography/computed tomography imaging.

OBJECTIVE: The aim of this study was to compare bone scintigraphy (BS) and positron emission tomography/computed tomography (PET/CT) for the detection of bone metastases from breast cancer. STUDY DESIGN: Twenty patients with breast cancer and bone pain were submitted to both bone scintigraphy and 18-F-fluorodeoxyglucose PET/CT imaging between July 2012 and June 2013. Scintigraphy was performed following an intravenous injection of technetium-99m-methylene diphosphonate (99mTc-MDP) around 10 days before the PET/CT scan, performed using an intravenous injection of 18-F-fluorodeoxyglucose followed by whole-body computed tomography (CT) to characterize metastases by both methods. Student's t-test for paired samples was used in the comparative data analysis, with significance at p<0.05. RESULTS: CT identified 429 metastatic implants in the 20 patients, with scintigraphy showing 244 of these lesions (57%) and PET/CT showing 307 (72%); however, there was no statistically significant difference between the mean number of lesions detected per patient with the two imaging modalities (p=0.367). CONCLUSION: In the present study, no difference was found between PET/CT and bone scintigraphy in the detection of bone metastases from breast cancer.

Effect of tamoxifen on extracellular signal-regulated kinases in the urethra of castrated female rats.

OBJECTIVE: The aim was to evaluate the effects of tamoxifen in activating extracellular signal-regulated kinases (ERKs) 1 and 2 in the urethras of castrated female rats. STUDY DESIGN: Twelve castrated adult female rats were divided into a control group (n=6) in which the animals received vehicle, and the experimental group (n=6) in which the rats received tamoxifen 250 µg/day by gavage for 28 days. Then, the animals were sacrificed and their urethras removed. Proteins were extracted, quantified and processed by Western blot analysis with specific phospho-ERK1 and 2 antibodies. Data were analyzed using Student's t-test (p<0.05). RESULTS: A significant increase occurred in phospho-ERK1 levels in the experimental group compared to the control group (p<0.01), while no difference was found in phospho-ERK2 levels between the groups (p=0.313). CONCLUSION: The present results indicate that, at the doses and during the time of treatment used, tamoxifen significantly increased phospho-ERK1 levels in the urethras of castrated female rats.

The effect of raloxifene on telomerase expression in breast carcinoma samples from postmenopausal women.

OBJECTIVE: This study aims to evaluate the effect of raloxifene on the expression of human telomerase reverse transcriptase (hTERT) in invasive ductal breast carcinoma of postmenopausal women. STUDY DESIGN: This study included 20 postmenopausal women with invasive, stage II, estrogen receptor-positive ductal carcinoma diagnosed by incisional biopsy, who received 60 mg of raloxifene orally for 28 days prior to definitive surgery. On the 29th day of treatment, definitive surgery was performed and a second tumor sample was taken for analysis. The catalytic subunit of telomerase (hTERT) was evaluated semiquantitatively by immunohistochemistry in the tumor samples obtained prior to and following raloxifene use and the results were analyzed using the McNemar test (p<0.05). RESULTS: The samples of 17 patients (85%) were classified as positive for telomerase expression prior to raloxifene treatment, while only 6 (30%) remained positive following raloxifene treatment (p<0.0026). CONCLUSION: In the present study, raloxifene significantly reduced the expression of hTERT in estrogen receptor-positive breast tumors from postmenopausal women.

Effect of a gel containing pilocarpine on vaginal atrophy in castrated rats

OBJECTIVES: To evaluate the effect of Carbopol gel formulations containing pilocarpine on the morphology and morphometry of the vaginal epithelium of castrated rats. METHODS: Thirty-one female Wistar-Hannover rats were randomly divided into four groups: the control Groups I (n=7, rats in persistent estrus; positive controls) and II (n=7, castrated rats, negative controls) and the experimental Groups, III (n=8) and IV (n=9). Persistent estrus (Group I) was achieved with a subcutaneous injection of testosterone propionate on the second postnatal day. At 90 days postnatal, rats in Groups II, III and IV were castrated and treated vaginally for 14 days with Carbopol gel (vehicle alone) or Carbopol gel containing 5% and 15% pilocarpine, respectively. Next, all of the animals were euthanized and their vaginas were removed for histological evaluation. A non-parametric test with a weighted linear regression model was used for data analysis (p<0.05). RESULTS: The morphological evaluation showed maturation of the vaginal epithelium with keratinization in Group I, whereas signs of vaginal atrophy were present in the rats of the other groups. Morphometric examinations showed mean thickness values of the vaginal epithelium of 195.10±12.23 μm, 30.90±1.14 μm, 28.16±2.98 μm and 29.84±2.30 μm in Groups I, II, III and IV, respectively, with statistically significant differences between Group I and the other three groups (p<0.0001) and no differences between Groups II, III and IV (p=0.0809). CONCLUSION: Topical gel formulations containing pilocarpine had no effect on atrophy of the vaginal epithelium in the castrated female rats.

Analysis of Ki-67 and Bcl-2 protein expression in normal colorectal mucosa of women with breast cancer.

The aim of this study was to evaluate Ki-67 and Bcl-2 protein expression in the normal colorectal mucosa adjacent to adenomatous polyps in women with breast cancer. A cross-sectional, controlled study was conducted in 35 women with and without breast cancer who had adenomatous colorectal polyps. The patients were divided into two groups: Group A (a control group of women without breast cancer, n=18) and Group B (a study group of women with breast cancer, n=17). A sample of normal colonic mucosa was collected at a distance of 5 cm from the polypoid lesion to evaluate immunohistochemical expression of the Ki-67 and Bcl-2 proteins. Student's t-test and the chi-square test were used to analyse Ki-67 and Bcl-2 expression, respectively. Statistical significance was established at p<0.05. The mean percentage of Ki-67-stained nuclei in Groups A and B was 25.12+/-2.08 and 41.50+/-1.85, respectively (p<0.001), whereas the percentage of cases with cells expressing Bcl-2 in Groups A and B was 17.6% and 82.4%, respectively (p<0.003). In the present study, greater proliferative activity and greater expression of the antiapoptotic protein Bcl-2 was found in the normal colorectal mucosa of women with breast cancer.

Preoperative pelvic lymphoscintigraphy is of limited usefulness for sentinel lymph node detection in cervical cancer.

OBJECTIVES: To analyze concordance between preoperative lymphoscintigraphy and intraoperative lymphatic mapping (ILM) for sentinel lymph node identification using technetium 99m-labeled-dextran 500 (99m-Tc) and patent blue dye in patients with early cervical cancer undergoing radical hysterectomy and pelvic lymphadenectomy, as well as to evaluate sentinel lymph node (SLN) detection. STUDY DESIGN: Forty-seven patients underwent surgical treatment for cervical cancer. For SLN identification, 99m-Tc and blue patent were injected into the cervix on the eve and day of surgery, respectively. Preoperative pelvic lymphoscintigraphy was performed in all patients after 99m-Tc injection. Concordance between preoperative lymphoscintigraphy and ILM was evaluated. RESULTS: Of the 56 patients who underwent preoperative lymphoscintigraphy, 43 (81.13%) had at least one lymph node identified. Bilateral lymph nodes were identified in 21 (37.5%) patients. Sentinel lymph nodes detected on ILM had been previously found on preoperative lymphoscintigraphy in 66.7%, 67.2% and 0% in the right, left and central locations, respectively. In 14 patients (25%), only one lymph node was identified on preoperative lymphoscintigraphy, but more than one sentinel lymph node was detected on intraoperative mapping. In nine (16.1%) patients, lymphoscintigraphy showed only unilateral lymph nodes, but ILM identified bilateral sentinel lymph nodes. CONCLUSION: The combination of patent blue and radionuclide techniques produced excellent results for SLN detection in cervical cancer. Preoperative lymphoscintigraphy does not offer any advantage over ILM for SLN identification.