Selection of reference miRNAs for RT-qPCR assays in endometriosis menstrual blood-derived mesenchymal stem cells.

Choosing appropriate reference genes or internal controls to normalize RT-qPCR data is mandatory for the interexperimental reproducibility of gene expression data obtained by RT-qPCR in most studies, including those on endometriosis. Particularly for miRNAs, the choice for reference genes is challenging because of their physicochemical and biological characteristics. Moreover, the retrograde menstruation theory, mesenchymal stem cells in menstrual blood (MenSCs), and changes in post-transcriptional regulatory processes through miRNAs have gained prominence in the scientific community as important players in endometriosis. Therefore, we originally explored the stability of 10 miRNAs expressions as internal control candidates in conditions involving the two-dimensional culture of MenSCs from healthy women and patients with endometriosis. Here, we applied multiple algorithms (geNorm, NormFinder, Bestkeeper, and delta Ct) to screen reference genes and assessed the comprehensive stability classification of miRNAs using RefFinder. Pairwise variation calculated using geNorm identified three miRNAs as a sufficient number of reference genes for accurate normalization. MiR-191-5p, miR-24-3p, and miR-103a-3p were the best combination for suitable gene expression normalization. This study will benefit similar research, but is also attractive for regenerative medicine and clinics that use MenSCs, miRNA expression, and RT-qPCR.

Identification of a rare copy number polymorphic gain at 3q12.2 with candidate genes for familial endometriosis.

Endometriosis is a complex disease that affects 10-15% of women of reproductive age. Familial studies show that relatives of affected patients have a higher risk of developing the disease, implicating a genetic role for this disorder. Little is known about the impact of germline genomic copy number variant (CNV) polymorphisms on the heredity of the disease. In this study, we describe a rare CNV identified in two sisters with familial endometriosis, which contain genes that may increase the susceptibility and progression of this disease. We investigated the presence of CNVs from the endometrium and blood of the sisters with endometriosis and normal endometrium of five women as controls without the disease using array-CGH through the Agilent 2x400K platform. We excluded common CNVs that were present in the database of genomic variation. We identified, in both sisters, a rare CNV gain affecting 113kb at band 3q12.2 involving two candidate genes: ADGRG7 and TFG. The CNV gain was validated by qPCR. ADGRG7 is located at 3q12.2 and encodes a G protein-coupled receptor influencing the NF-kappaß pathway. TFG participates in chromosomal translocations associated with hematologic tumor and soft tissue sarcomas, and is also involved in the NF-kappa B pathway. The CNV gain in this family provides a new candidate genetic marker for future familial endometriosis studies. Additional longitudinal studies of affected families must confirm any associations between this rare CNV gain and genes involved in the NF-kappaß pathway in predisposition to endometriosis.

Identification of a rare copy number polymorphic gain at 3q12.2 with candidate genes for familial endometriosis

Abstract Endometriosis is a complex disease that affects 10-15% of women of reproductive age. Familial studies show that relatives of affected patients have a higher risk of developing the disease, implicating a genetic role for this disorder. Little is known about the impact of germline genomic copy number variant (CNV) polymorphisms on the heredity of the disease. In this study, we describe a rare CNV identified in two sisters with familial endometriosis, which contain genes that may increase the susceptibility and progression of this disease. We investigated the presence of CNVs from the endometrium and blood of the sisters with endometriosis and normal endometrium of five women as controls without the disease using array-CGH through the Agilent 2x400K platform. We excluded common CNVs that were present in the database of genomic variation. We identified, in both sisters, a rare CNV gain affecting 113kb at band 3q12.2 involving two candidate genes: ADGRG7 and TFG. The CNV gain was validated by qPCR. ADGRG7 is located at 3q12.2 and encodes a G protein-coupled receptor influencing the NF-kappaβ pathway. TFG participates in chromosomal translocations associated with hematologic tumor and soft tissue sarcomas, and is also involved in the NF-kappa B pathway. The CNV gain in this family provides a new candidate genetic marker for future familial endometriosis studies. Additional longitudinal studies of affected families must confirm any associations between this rare CNV gain and genes involved in the NF-kappaβ pathway in predisposition to endometriosis.

Altered Differential Expression of Genes and microRNAs Related to Adhesion and Apoptosis Pathways in Patients with Different Phenotypes of Endometriosis.

We aim to investigate the expression of genes (MAPK1 and CAPN2) and microRNAs (miR-30a-5p, miR-7-5p, miR-143-3p, and miR-93-5p) involved in adhesion and apoptosis pathways in superficial peritoneal endometriosis (SE), deep infiltrating endometriosis (DE), and ovarian endometrioma (OE), and to evaluate whether these lesions share the same pathophysiological mechanisms. We used samples of SE (n = 10), DE (n = 10), and OE (n = 10), and endometrial biopsies of these respective patients affected with endometriosis under treatment at a tertiary University Hospital. Endometrial biopsies collected in the tubal ligation procedure from women without endometriosis comprised the control group (n = 10). Quantitative real-time polymerase chain reaction was performed. The expression of MAPK1 (p < 0.0001), miR-93-5p (p = 0.0168), and miR-7-5p (p = 0.0006) was significantly lower in the SE group than in the DE and OE groups. The expression of miR-30a (p = 0.0018) and miR-93 (p = 0.0052) was significantly upregulated in the eutopic endometrium of women with endometriosis compared to the controls. MiR-143 (p = 0.0225) expression also showed a statistical difference between the eutopic endometrium of women with endometriosis and the control group. In summary, SE showed lower pro-survival gene expression and miRNAs involved in this pathway, indicating that this phenotype has a different pathophysiological mechanism compared to DE and OE.

Downregulation of DROSHA: Could It Affect miRNA Biogenesis in Endometriotic Menstrual Blood Mesenchymal Stem Cells?

Menstrual blood mesenchymal stem cells (MenSCs) have gained prominence in the endometriosis scientific community, given their multifunctional roles in regenerative medicine as a noninvasive source for future clinical applications. In addition, changes in post-transcriptional regulation via miRNAs have been explored in endometriotic MenSCs with a role in modulating proliferation, angiogenesis, differentiation, stemness, self-renewal, and the mesenchymal-epithelial transition process. In this sense, homeostasis of the miRNA biosynthesis pathway is essential for several cellular processes and is related to the self-renewal and differentiation of progenitor cells. However, no studies have investigated the miRNA biogenesis pathway in endometriotic MenSCs. In this study, we profiled the expression of eight central genes for the miRNA biosynthesis pathway under experimental conditions involving a two-dimensional culture of MenSCs obtained from healthy women (n = 10) and women with endometriosis (n = 10) using RT-qPCR and reported a two-fold decrease in DROSHA expression in the disease. In addition, miR-128-3p, miR-27a-3p, miR-27b-3p, miR-181a-5p, miR-181b-5p, miR-452-3p, miR-216a-5p, miR-216b-5p, and miR-93-5p, which have been associated with endometriosis, were identified through in silico analyses as negative regulators of DROSHA. Because DROSHA is essential for miRNA maturation, our findings may justify the identification of different profiles of miRNAs with DROSHA-dependent biogenesis in endometriosis.

Uterine administration of C-X-C motif chemokine ligand 12 increases the pregnancy rates in mice with induced endometriosis.

OBJECTIVE: To study the effect of intrauterine injection of C-X-C motif chemokine ligand 12 (CXCL12), also known as a stem cell chemoattractant (stromal cell-derived factor 1), on fertility and endometrial receptivity in mice with endometriosis. DESIGN: Laboratory study. SETTING: Academic Medical Center. ANIMAL(S): Fifty-six mice underwent chemotherapy and bone marrow transplantation. Thirty-six of these mice underwent either surgery to induce endometriosis (n = 20) or sham surgery (n = 16). INTERVENTION(S): Injection of CXCL12 as a potential therapeutic agent to improve fertility in endometriosis. MAIN OUTCOME MEASURE(S): Pregnancy rate, bone marrow-derived cell (BMDC) recruitment and endometrial receptivity markers. RESULT(S): The mice with or without endometriosis received a single uterine injection of either CXCL12 or placebo. Uterine injection of CXCL12 increased the pregnancy rates in a mouse model of endometriosis. Mice were euthanized after delivery, and implantation markers homeobox A11, alpha-v beta-3 integrin, and progesterone receptor were analyzed by immunohistochemistry, whereas green fluorescent protein positive BMDC recruitment was quantified by immunohistochemistry and immunofluorescence. The sham surgery groups without endometriosis had the highest cumulative pregnancy rate (100%) regardless of CXCL12 treatment. The endometriosis group treated with placebo had the lowest pregnancy rate. An increased pregnancy rate was noted in the endometriosis group after treatment with CXCL12. There was also an increase in BMDC recruitment and endometrial expression of progesterone receptor and alpha-v beta-3 integrin in the endometriosis group that received CXCL12 compared with that in the endometriosis group that received placebo. CONCLUSION(S): Uterine injection of CXCL12 increased the pregnancy rates in a mouse model of endometriosis. These results suggest that CXCL12 has a potential role as a therapeutic agent in women with infertility related to endometriosis and potentially other endometrial receptivity defects.

Prevalence of chronic pelvic pain and primary dysmenorrhea in women of reproductive age in Ecuador.

BACKGROUND: Chronic pelvic pain (CPP) and primary dysmenorrhoea are debilitating conditions that can impair the quality of life of affected women. These conditions are frequently neglected, delaying proper diagnosis and healthcare provision. This study aimed to estimate the prevalence of CPP and primary dysmenorrhoea in Ecuador and identify potential variables associated with their occurrence. METHODS: We conducted a cross-sectional survey in an urban neighbourhood of Quito, the capital of Ecuador. A total of 2397 participants of 14-49 years of age were included. The data were collected through questionnaires administered by trained interviewers.The crude and adjusted prevalence ratios were calculated using a log-binomial regression model. The correlation between pain intensity catastrophising of symptoms were statistically analysed. RESULTS: The prevalence of CPP and primary dysmenorrhoea was 9.8% and 8.9%, respectively. Irritative urinary symptoms, primary dysmenorrhoea, and underlying mental disorders were associated with CPP, while smoking, irritable bowel syndrome, sleep disturbance, dyspareunia, and mental disorders were associated with primary dysmenorrhoea. CONCLUSIONS: The prevalence of CPP and primary dysmenorrhoea in Ecuador was similar to that in other Latin American countries. Primary dysmenorrhoea is a risk factor of CPP, and less than a quarter of women are undergoing treatment for the condition. Our findings reinforce the importance of healthcare interventions in anticipating the diagnosis of these conditions in women of reproductive age.

Overexpression of miR-200b-3p in Menstrual Blood-Derived Mesenchymal Stem Cells from Endometriosis Women.

The key relationship between Sampson's theory and the presence of mesenchymal stem cells in the menstrual flow (MenSCs), as well as the changes in post-transcriptional regulatory processes as actors in the etiopathogenesis of endometriosis, are poorly understood. No study to date has investigated the imbalance of miRNAs in MenSCs related to the disease. Thus, through literature and in silico analyses, we selected four predicted miRNAs as regulators of EGR1, SNAI1, NR4A1, NR4A2, ID1, LAMC3, and FOSB involved in pathways of apoptosis, angiogenesis, response to steroid hormones, migration, differentiation, and cell proliferation. These genes are frequently overexpressed in the endometriosis condition in our group studies. They were the trigger for the miRNAs search. Therefore, a case-control study was conducted with MenSCs of women with and without endometriosis (ten samples per group). Crossing information obtained from the STRING, PubMed, miRPathDB, miRWalk, and DIANA TOOLS databases, we chose to explore the expression of miR-21-5p, miR-100-5p, miR-143-3p, and miR-200b-3p by RT-qPCR. We found an upregulation of the miR-200b-3p in endometriosis MenSCs (P = 0.0207), with a 7.93-fold change (ratio of geometric means) compared to control. Overexpression of miR-200b has been associated with increased cell proliferation, stemness, and accentuated mesenchymal-epithelial transition process in eutopic endometrium of endometriosis. We believe that dysregulated miR-200b-3p may establish primary changes in the MenSCs, thus favoring tissue implantation at the ectopic site.

Eutopic endometrium from women with endometriosis and chlamydial endometritis share immunological cell types and DNA repair imbalance: A transcriptome meta-analytical perspective.

The aim of this study was to identify the key similarities between the eutopic endometrium of women with endometriosis and chlamydia-induced endometritis taking into account tissue microenvironment heterogeneity, transcript gene profile, and enriched pathways. A meta-analysis of whole transcriptome microarrays was performed using publicly available data, including samples containing both glandular and stromal endometrial components. Control samples were obtained from women without any reported pathological condition. Only samples obtained during the proliferative menstrual phase were included. Cellular tissue heterogeneity was predicted using a method that integrates gene set enrichment and deconvolution approaches. The batch effect was estimated by principal variant component analysis and removed using an empirical Bayes method. Differentially expressed genes were identified using an adjusted p-value < 0.05 and fold change = 1.5. The protein-protein interaction network was built using the STRING database and interaction score over 400. The Molecular Signatures Database was used to analyse the functional enrichment analysis. Both conditions showed similarities in cell types in the microenvironment, particularly CD4+ and CD8+ Tem cells, NKT cells, Th2 cells, basophils, and eosinophils. With regards to the regulation of cellular senescence and DNA integrity/damage checkpoint, which are commonly enriched pathways, 21 genes were down-regulated and directly related to DNA repair. Compared to the endometriosis samples, some chlamydial endometritis samples presented a lack of enriched immune pathways. Our results suggest that both conditions show similar distributions of microenvironment cell types, the downregulation of genes involved in DNA repair and cell cycle control, and pathways involved in immune response evasion.

Laparoscopic bowel resection of deep infiltrating endometriosis. Comparative outcomes of a public teaching hospital and a referral private hospital.

PURPOSE: To compare the operative outcomes of laparoscopic surgical treatment for bowel endometriosis in a public teaching hospital versus in a private referral hospital. METHODS: The indications for surgery, type and time of operation, length of hospital stay, need for a temporary stoma, rate of conversion to open surgery, and postoperative complications were evaluated. RESULTS: One hundred eighty-one patients were included (150 patients, 82.9%, in a private hospital). In the private hospital, there were more patients with infertility [56% vs. 29%; P=0.01] as an indication for surgery) and segmental resection was more common in the private hospital (48% vs. 29%, p=0.05). The average operative time (211.9±83.4 minutes vs. 128 ± 55 minutes, p<0.001) as well as the length of hospital stay (3.97±1.7 days vs. 1.56±0.85 days, p<0.001) was higher in the public hospital; the rate of conversion to open surgery was significantly lower in the private hospital (2% vs. 32.3%, p<0.001). Operations performed at the public hospital were associated with higher rates of postoperative complications (Clavien-Dindo II and II) (38.7% x 11.3%, p=0.021; OR 3.2, CI 95% 1.2-8.0). CONCLUSION: Laparoscopic surgery in private centers was associated with reductions in major complications, surgical times, lengths of stay and rates of conversion to open surgery compared to that in public teaching hospitals.

Gynecological Surgery and COVID-19: What is the Impact and How Should I Manage it?

It is estimated that around 28 million surgeries will be postponed or canceled worldwide as a result of this pandemic, causing a delay in the diagnosis and treatment of more than 2 million cancer cases. In Brazil, both the National Health Agency (ANS) and National Health Surveillance Agency (ANVISA) advised the postponement of elective and non-essential surgeries, causing a considerable impact on the number of surgical procedures that decreased by 33.4% in this period. However, some women need treatment for various gynecological diseases that cannot be postponed. The purpose of this article is to present recommendations on surgical treatment during the COVID-19 pandemic.

Estima-se que cerca de 28 milhões de cirurgias sejam postergadas ou canceladas no mundo em decorrência desta pandemia, causando atraso no diagnóstico e tratamento de mais de 2 milhões de casos oncológicos. No Brasil, tanto a ANS (Agencia Nacional de Saúde) como a ANVISA (Agencia Nacional de Vigilância Sanitária) orientaram o adiamento das cirurgias eletivas e não essenciais, tendo um impacto considerável no número de procedimentos cirúrgicos com diminuição de 33,4% neste período no Brasil. No entanto, algumas mulheres necessitam de tratamento para várias doenças ginecológicas, algumas das quais não podem ser adiadas. O objetivo deste artigo é apresentar recomendações sobre o tratamento cirúrgico durante a pandemia de COVID-19.

Gynecological surgery and Covid-19: what is the impact and how should I manage it? / Cirurgia ginecológica e Covid-19: qual impacto e como devo conduzir?

Abstract It is estimated that around 28 million surgeries will be postponed or canceled worldwide as a result of this pandemic, causing a delay in the diagnosis and treatment of more than 2 million cancer cases. In Brazil, both the National Health Agency (ANS) and National Health Surveillance Agency (ANVISA) advised the postponement of elective and non-essential surgeries, causing a considerable impact on the number of surgical procedures that decreased by 33.4% in this period. However, some women need treatment for various gynecological diseases that cannot be postponed. The purpose of this article is to present recommendations on surgical treatment during the COVID-19 pandemic.

Resumo Estima-se que cerca de 28 milhões de cirurgias sejam postergadas ou canceladas nomundo em decorrência desta pandemia, causando atraso no diagnóstico e tratamento de mais de 2 milhões de casos oncológicos. No Brasil, tanto a ANS (Agencia Nacional de Saúde) comoa ANVISA (Agencia Nacional de Vigilância Sanitária) orientaram o adiamento das cirurgias eletivas e não essenciais, tendo um impacto considerável no número de procedimentos cirúrgicos comdiminuição de 33,4% neste período no Brasil.No entanto, algumasmulheres necessitam de tratamento para várias doenças ginecológicas, algumas das quais não podem ser adiadas. O objetivo deste artigo é apresentar recomendações sobre o tratamento cirúrgico durante a pandemia de COVID-19.

Impact of Bevacizumab on Experimentally Induced Endometriotic Lesions: Angiogenesis, Invasion, Apoptosis, and Cell Proliferation.

Endometriosis is responsible for pain symptoms with great impact on the patient's quality of life. Several medication lines have been studied aiming at its definitive treatment. Among them, angiogenesis inhibitor factors may be effective given that angiogenesis has fundamental role in the establishment and growth of endometriotic lesions. In this study, we investigated the influence of bevacizumab, anti-factor drug of endothelial growth (anti-VEGF), used at two different dosages, in experimental endometriosis induced in rats. After the induction of endometriosis lesions in rats, they were divided in 3 groups: control group, no treatment, and two other groups were treated with different dosages of the same medication for 4 weeks. At the end of the treatment, endometriotic lesions were removed and evaluated regarding area of lesions, presence of endometrial tissue in microscopy, positivity for anti-VEGF antibody in immunohistochemistry, and gene expression of Pcna, Mmp9, Tp63, and Vegfa. Bevacizumab acted by reducing the area of lesions in the groups that received medication (p = 0.002) and reducing gene expression to Tp63 in lesions (p = 0.04). There was no significant result in other evaluations. We observed that there was significant reduction of the area of lesions among groups, suggesting that bevacizumab has a positive effect on disease control. The gene expression of Tp63 was significantly lower in the group that received high dose of the drug when compared with the other two groups; therefore, we concluded that bevacizumab acts by reducing cell proliferation and differentiation in lesions, constituting a real option for treating endometriosis.

Transcriptome meta-analysis reveals differences of immune profile between eutopic endometrium from stage I-II and III-IV endometriosis independently of hormonal milieu.

Eutopic endometrium appears to be crucial for endometriosis development. Despite of the evident importance, data regarding the cellular microenvironment remain unclear. Our objective was to explore the tissue microenvironment heterogeneity, transcripts, and pathways that are enriched in all phases of the menstrual cycle by analysing publicly deposited data derived from whole transcriptome microarrays of eutopic endometria of women with and without endometriosis. A meta-analysis of the transcriptome microarrays was performed using raw data available from a public database. Eligibility criteria included eutopic endometrium samples from women with endometriosis and healthy controls without any pathological condition reported the presence of an adequately reported normal menstrual phase, and samples containing both glandular and stromal components. Raw data were processed using a robust multiarray average method to provide background correction, normalisation, and summarisation. The batch effect was estimated by principal variant component analysis and removed using an empirical Bayes method. Cellular tissue heterogeneity was inferred using the xCell package. Differentially expressed genes were identified based on a 5% adjusted p value and a 2.0-fold change. Pathways were identified by functional enrichment based on the Molecular Signatures Database, a p value of < 5%, and an FDR q value of ≤ 25%. Genes that were more frequently found in pathways were identified using leading edge analysis. In a manner independent of cycle phase, the subpopulations of activated dendritic cells, CD4 T effector memory phenotype cells, eosinophils, macrophages M1, and natural killer T cells (NKT) were all higher in stage I-II endometriosis compared to those in healthy controls. The subpopulations of M2 macrophages and natural killer T cells were elevated in eutopic endometriums from women with stage III-IV endometriosis, and smooth muscle cells were always more prevalent in healthy eutopic endometriums. Among the differently expressed genes, FOS, FOSB, JUNB, and EGR1 were the most frequently mapped within the interaction networks, and this was independent of stage and cycle phase. The enriched pathways were directly related to immune surveillance, stem cell self-renewal, and epithelial mesenchymal transition. PI3K AKT mTOR, TGF signalling, and interferon alpha/gamma responses were enriched exclusively in stage III-IV endometriosis. The cellular microenvironments and immune cell profiles were different between eutopic endometriums from women with stage I-II and stage III-IV endometriosis, and these differences were independent of the hormonal milieu. Specifically, a pro-inflammatory profile was predominant in stage I-II endometriosis, and M1-M2 polarization into eutopic endometrium may be crucial for the progression of the disease. The higher prevalence of NKT cells in eutopic endometriums from women with endometriosis that was independent of cycle phase or staging suggested a sustained stress and/or damage to these eutopic endometriums. Based on this, the results of this meta-analysis are important for identifying challenges and opportunities for future research.

Analysis of body composition and pain intensity in women with chronic pelvic pain secondary to endometriosis / Análise da composição corporal e intensidade de dor em mulheres com dor pélvica crônica secundária a endometriose

Abstract Objective To determine the average body composition (percentage of body fat), the anthropometric markers, and the intensity of clinical pain in women with a clinical diagnosis of chronic pelvic pain (CPP) secondary to endometriosis. Methods A case-control study performed with 91 women, 46 of whom with CPP secondary to endometriosis and 45 of whom with CPP secondary to other causes. They underwent an evaluation of the anthropometric parameters by means of the body mass index (BMI), the perimeters (waist, abdomen, hip), and the percentage of body fat (%BF), which were assessed on a body composition monitor by bioimpedance; the intensity of the clinical pain was evaluated using the visual analog scale (VAS), and the symptoms of anxiety and depression, using the hospital's anxiety and depression scale (HAD). Results The groups did not differ in terms of mean age, BMI, %BF or regarding the available waist-to-hip ratio (WHR). The mean intensity of the clinical pain by the VAS was of 7.2 ± 2.06 in the group with CPP secondary to endometriosis, and of 5.93 ± 2.64 in the group with CPP secondary to other causes (p = 0.03), revealing significant differences between the groups. Conclusion We concluded that, despite the difference in the pain score assessed between the two groups, there was no difference regarding body composition and anthropometry.

Resumo Objetivo Determinar a composição corporal média (porcentagem de gordura corporal), os marcadores antropométricos, e a intensidade de dor clínica em mulheres com diagnóstico clínico de dor pélvica crônica (DPC) secundária a endometriose. Métodos Um estudo de caso-controle realizado com 91 mulheres, 46 das quais com DPC secundária a endometriose, e 45 das quais com DPC secundária a outras causas. As pacientes foram submetidas à avaliação dos parâmetros antropométricos por meio do índice de massa corporal (IMC), dos perímetros (cintura, abdômen, quadril), e do percentual de gordura corporal (%GC), que foram avaliados emmonitor de composição corporal por bioimpedância; a intensidade clínica da dor foi avaliada usando-se a escala visual analógica (EVA), e os sintomas de ansiedade e depressão, usando a escala de ansiedade e depressão do hospital (EADH). Resultados Os grupos não diferiram quanto à idade média, ao IMC, ao %GC, nem quanto à relação da cintura-quadril (RCQ) disponível. Amédia da intensidade clínica da dor pela EVA foi de 7,2 ± 2,06 no grupo com DPC secundária a endometriose, e de 5,93 ± 2,64 no grupo com DPC secundária a outras causas (p = 0,03), revelando diferenças significativas entre os grupos. Emrelação à EADH, ambos os grupos estavam acima da média de corte. Conclusão Concluímos que, apesar da diferença no escore de dor avaliado entre os dois grupos, não houve diferença com relação à composição corporal e à antropometria.

Thalidomide Reduces Cell Proliferation in Endometriosis Experimentally Induced in Rats.

OBJECTIVE: To analyze the effect of thalidomide on the progression of endometriotic lesions experimentally induced in rats and to characterize the pattern of cell proliferation by immunohistochemical Proliferating Cell Nuclear Antigen (PCNA) labeling of eutopic and ectopic endometrium. METHODS: Fifteen female Wistar rats underwent laparotomy for endometriosis induction by resection of one uterine horn, isolation of the endometrium and fixation of a tissue segment to the pelvic peritoneum. Four weeks after, the animals were divided into 3 groups: control (I), 10mg/kg/day (II) and 1mg/kg/day (III) intraperitoneal thalidomide for 10 days. The lesion was excised together with the opposite uterine horn for endometrial gland and stroma analysis. Eutopic and ectopic endometrial tissue was submitted to immunohistochemistry for analysis of cell proliferation by PCNA labeling and the cell proliferation index (CPI) was calculated as the number of labeled cells per 1,000 cells. RESULTS: Group I showed a mean CPI of 0.248 ± 0.0513 in the gland and of 0.178 ± 0.046 in the stroma. In contrast, Groups II and III showed a significantly lower CPI, that is, 0.088 ± 0.009 and 0.080 ± 0.021 for the gland (p < 0.001) and 0.0945 ± 0.0066 and 0.075 ± 0.018 for the stroma (p < 0.001), respectively. Also, the mean lesion area of Group I was 69.2 mm2, a significantly higher value compared with Group II (49.4 mm2, p = 0.023) and Group III (48.6 mm2, p = 0.006). No significant difference was observed between Groups II and III. CONCLUSION: Thalidomide proved to be effective in reducing the lesion area and CPI of the experimental endometriosis implants both at the dose of 1 mg/kg/day and at the dose of 10 mg/kg/day.

OBJETIVO: Analisar o efeito da talidomida na progressão de lesões endometrióticas induzidas experimentalmente em ratas e caracterizar o padrão de proliferação celular pela marcação imunohistoquímica de Antígeno Nuclear de Célula Proliferativa (PCNA) no endométrio eutópico e ectópico. MéTODOS: Quinze ratas Wistar foram submetidas a laparotomia para indução de endometriose por ressecção de um corno uterino, isolamento do endométrio e fixação de um segmento do tecido ao peritônio pélvico. Após quatro semanas, os animais foram divididos em 3 grupos: controle (I), 10 mg/kg/dia (II) e 1 mg/kg/dia (III) de talidomida intraperitoneal por um período de 10 dias. As lesões foram resseccionadas juntamente com o corno uterino oposto para análise da glândula endometrial e do estroma. O tecido endometrial eutópico e ectópico foi submetido à imunohistoquímica para análise da proliferação celular por marcação com PCNA e o índice de proliferação celular (CPI) foi calculado como o número de células marcadas por 1.000 células. RESULTADOS: O grupo I apresentou média de CPI de 0,248 ± 0,0513 na glândula e de 0,178 ± 0,046 no estroma. Em contraste, os grupos II e III apresentaram CPI significativamente menor, isto é, 0,088 ± 0,009 e 0,080 ± 0,021 para a glândula (p < 0,001) e 0,0945 ± 0,0066 e 0,075 ± 0,018 para o estroma (p < 0,001), respectivamente. Além disso, a área de lesão média do Grupo I foi de 69,2 mm2, valor significativamente maior em relação ao Grupo II (49,4 mm2, p = 0,023) e Grupo III (48,6 mm2, p = 0,006). Não houve diferença estatisticamente significante entre os Grupos II e III. CONCLUSãO: A talidomida mostrou-se eficaz na redução da área da lesão e CPI dos implantes de endometriose experimental tanto na dose de 1 mg/kg/dia quanto na dose de 10 mg/kg/dia.