Michio Kitajima

Maternal serum triglyceride at 24–32 weeks’ gestation and newborn weight in nondiabetic women with positive diabetic screens

Objective To determine whether elevated midpregnancy maternal serum lipid levels predict newborn weight at term and the risk of large for gestational age (LGA) infants in women with positive diabetic screen but normal glucose tolerance test. Methods Japanese gravidas who had positive diabetic screens and normal 75-g oral glucose tolerance tests (GTT) at 24–32 weeks were enrolled. Subjects with complications, including diabetes, hypertension, or fetal anomalies were excluded, as were women with multifetal gestations. Fasting serum triglyceride, free fatty acids, and total cholesterol levels were measured at the time of GTT. We tested the association between maternal variables and birth weight by univariable analysis. We used multivariable analysis to test whether the association between fasting lipids and birth weight was independent of prepregnant maternal body mass index (BMI), maternal weight gain during pregnancy, and plasma glucose levels at GTT. We also used multiple logistic regression analysis to determine whether maternal hyperlipidemia, defined as more than the 75th percentile of each lipid, is a risk factor for having an LGA infant. Results We enrolled 146 subjects. Among measured maternal lipids, only triglyceride levels correlated with birth weight in univariable analysis (r = 0.22, P = .009). Birth weight also was correlated with prepregnant maternal BMI (r = 0.18, P = .04) and fasting plasma glucose levels (r = 0.17, P = .04). The association between maternal fasting triglyceride level and birth weight remained significant after adjusting for prepregnant BMI, maternal weight gain, fasting plasma glucose levels, fetal gender, and gestational age at birth (P = .01). Logistic regression analysis showed that fasting maternal hypertriglyceridemia (over 259 mg/dL) was the significant predictor of LGA infants, independent of prepregnant BMI, maternal weight gain, and maternal plasma glucose levels (odds ratio 11.6; 95% confidence interval 1.1, 122; P = .04). Conclusion In women with positive diabetic screens but normal GTTs, fasting triglyceride levels at 24–32 weeks correlated positively with newborn weight at term, independent of maternal plasma glucose levels and obesity. Maternal fasting serum triglyceride levels in midpregnancy might be an independent predictor of fetal macrosomia in those women.

Changes in tissue inflammation, angiogenesis and apoptosis in endometriosis, adenomyosis and uterine myoma after GnRH agonist therapy

BACKGROUND Information is limited regarding the multifunctional role of GnRH agonist (GnRHa) therapy in reproductive diseases. We investigated the pattern of changes in inflammatory reaction, micro-vessel density and apoptosis in the tissues collected from women with endometriosis, adenomyosis and uterine myoma who were treated with or without GnRHa therapy. METHODS Biopsy specimens were collected from lesions, myometria and corresponding endometria of 45 women with ovarian endometrioma, 35 women with adenomyosis and 56 women with uterine myoma. A fraction of these women were treated with GnRHa therapy for a variable period of 3-6 months before surgery. We performed immunohistochemical analysis of CD68, a macrophage (Mvarphi) marker and von Willebrand factor (VWF), a vessel marker, using respective antibodies. Changes in apoptosis were examined using TdT-mediated dUTP-biotin nick end-labeling assay and by the immunoexpression of activated caspase-3 in tissues after GnRHa therapy. RESULTS The infiltration of CD68-positive Mvarphi and VWF-positive micro-vessel density were significantly decreased in the endometria of women with endometriosis, adenomyosis and uterine myoma in the GnRHa-treated group when compared with that in the non-treated group. Marked decreases in inflammatory and angiogenic responses were observed in lesions and myometria of these diseases. When compared with the non-treated group, a significant increase in apoptotic index (apoptotic cells per 10 mm(2) area) and quantitative-histogram scores of activated caspase-3 after GnRHa therapy were observed in the eutopic endometria, lesions and myometria of these diseases. CONCLUSIONS GnRHa was able to markedly reduce the inflammatory reaction and angiogenesis and to significantly induce apoptosis in tissues derived from women with endometriosis, adenomyosis and uterine myoma. These multiple biological effects at the tissue level may be involved in the regression of these reproductive diseases.

Immunopathogenesis of Pelvic Endometriosis: Role of Hepatocyte Growth Factor, Macrophages and Ovarian Steroids

Endometriosis, a chronic disease characterized by endometrial tissue located outside the uterine cavity is associated with chronic pelvic pain and infertility. However, an in‐depth understanding of the pathophysiology of endometriosis is still elusive. It is generally believed that besides ovarian steroid hormones, the growth of endometriosis can be regulated by innate immune system in pelvic microenvironment by their interaction with endometrial cells and immune cells. We conducted a series of studies in perspectives of pelvic inflammation that is triggered primarily by bacterial endotoxin (lipopolysacccharide) and is mediated by toll‐like receptor 4 and showed their involvement in the development of pelvic endometriosis. As a cellular component of innate immune system, macrophages were found to play a central role in inducing pelvic inflammatory reaction. We further report here that peritoneal macrophages retain receptors encoding for estrogen and progesterone and ovarian steroids also participate in producing an inflammatory response in pelvic cavity and are involved in the growth of endometriosis either alone or in combination with hepatocyte growth factor (HGF). As a pleiotropic growth factor, HGF retains multifunctional role in endometriosis. We describe here the individual and step‐wise role of HGF, macrophages and ovarian steroid hormones and their orchestrated involvement in the immunopathogenesis of pelvic endometriosis.

Changes in serum anti-Müllerian hormone levels may predict damage to residual normal ovarian tissue after laparoscopic surgery for women with ovarian endometrioma.

We measured serum anti-Müllerian hormone levels before and after surgery in women undergoing unilateral and monolocular cystectomy for benign ovarian diseases. Comparing to control benign cysts, we found a significant decline in serum anti-Müllerian hormone levels with consequent depletion of follicles in tissue specimens after surgery for women with ovarian endometrioma.

Immunoexpression of hepatocyte growth factor and c-Met receptor in the eutopic endometrium predicts the activity of ectopic endometrium

OBJECTIVE To investigate the mitogenic and angiogenic activity of the eutopic and ectopic endometrium throughout the menstrual cycle and to examine whether the activity of the eutopic endometrium is useful to predict greater activity of the ectopic endometrium. DESIGN Controlled clinicopathologic study using intact tissue. SETTING Nagasaki University School of Medicine, Nagasaki, Japan. PATIENT(S) Fifteen infertile women with pelvic endometriosis and 10 women without endometriosis undergoing laparoscopy. INTERVENTION(S) Biopsies from the ectopic endometrium and the corresponding eutopic endometrium were collected. Immunohistochemical staining was performed using respective antibodies, and a computer analyzed modified quantitative-histogram (Q-H) score was used to quantify immunostaining. MAIN OUTCOME MEASURE(S) The immunoreactions of hepatocyte growth factor (HGF), its receptor, c-Met, vascular endothelial growth factor (VEGF), proliferating cell nuclear antigen (PCNA), and von Willebrand factor (VWF) in eutopic and ectopic endometrium were examined, and their relation with different revised American Society for Reproductive Medicine (r-ASRM) stages and the morphology of endometriosis was evaluated. RESULT(S) The immunoexpressions of HGF and c-Met were significantly higher in the eutopic endometrium of patients with endometriosis than in that of controls. The Q-H scores of HGF, c-Met, VEGF, PCNA, and microvessel density (MVD) were markedly higher in red peritoneal lesions when compared with other lesions. The Q-H scores did not reveal r-ASRM stage-dependent variation in any of these markers. We observed a significant correlation between the immunoexpressions of HGF, c-Met, and PCNA or microvessel counts. When we combined the Q-H scores of the glandular epithelium and stroma, we found that increased activity of the eutopic endometrium as measured by the immunoreaction of HGF, c-Met, VEGF, PCNA, and MVD was similar to highly active red lesions and was significantly higher than that of controls and other lesions. CONCLUSION(S) Immunoexpression of HGF and c-Met in the eutopic endometrium of patients with pelvic endometrioisis is possibly useful to predict greater activity of the ectopic endometrium.

Escherichia coli contamination of menstrual blood and effect of bacterial endotoxin on endometriosis

To test the hypothesis that bacterial contamination of menstrual blood could be a local biologic event in the development of endometriosis, menstrual blood was cultured and bacterial endotoxin was measured in menstrual blood and peritoneal fluid. Our results suggest that compared with control women, higher colony formation of Escherichia coli in menstrual blood and endotoxin levels in menstrual fluid and peritoneal fluid in women with endometriosis may promote Toll-like receptor 4-mediated growth of endometriosis.

Toll-Like Receptors in Innate Immunity: Role of Bacterial Endotoxin and Toll-Like Receptor 4 in Endometrium and Endometriosis

Macrophages, dendritic cells, and Toll-like receptors (TLRs) are integral components of the innate immune system. This rapidly reactive system responds immediately to infectious or other non-self agents, thereby inducing an inflammatory response to protect the host until the activation of the slower adaptive immune system. The fundamentals of the innate immune system, functional characteristics of TLRs, and signaling pathways of TLR4 are discussed for the easy understanding by readers. Studies showed that the growth and progression of endometriosis continue even in ovariectomized animals. This indicates that besides ovarian steroid hormones, the growth of endometriosis can be regulated by the innate immune system in the pelvic environment. As a component of the innate immune system, increased infiltration of macrophages has been described in the intact tissue and peritoneal fluid of women with endometriosis. In this review article, we discuss the role of bacterial endotoxin and TLR4 in endometrium and endometriosis and outline the involvement of endotoxin in causing adverse reproductive outcome.

Higher activity by opaque endometriotic lesions than nonopaque lesions

Background.  Higher activity by early endometriosis than advanced endometriosis has been reported. However, the pattern of activity in individual colored endometriotic lesions in pelvic cavity is unknown. We investigated the variation in activity of the different colored morphologic lesions as proposed by the current revised American Society of Reproductive Medicine (ASRM) classification in women with endometriosis.

Modified Reduction Surgery for Adenomyosis

We describe the preliminary clinical results of a modified method of reduction surgery for easy approach and effective removal of lesions in women with adenomyosis. Old classical reduction surgery was performed in 5 women with imaging diagnosis of adenomyosis who were selected retrospectively among 104 patients undergoing conservative surgery. A transverse H incision method in the reduction surgery was applied to 6 of 83 patients wishing to preserve fertility. Benefit in operative procedure, complications, patients’ compliance, and pregnancy outcome were analyzed and compared between 5 women with the classical method and 6 women with the H incision method for adenomyosis. No apparent difference was observed in the operation time, blood loss and volume of excised specimens between these two groups. The major complication of perforation during surgery occurred in 2 patients (40%) by the classical method and in only 1 patient (17%) by the H incision technique. The subjective relief of pain was relatively more evident in the modified than in the classical group. There was no case of pregnancy in the classical group; however, 1 patient conceived spontaneously 4 months after operation by this H incision procedure. Our H incision technique may be considered a useful method for an easy surgical approach and satisfactory removal of adenomyotic lesions and may be better than the old classical method of reduction surgery.

Cell proliferation effect of GnRH agonist on pathological lesions of women with endometriosis, adenomyosis and uterine myoma

BACKGROUND We recently demonstrated the effect of gonadotrophin-releasing hormone agonist (GnRHa) on tissue inflammation, angiogenesis and apoptosis in endometriosis, adenomyosis and uterine myoma. Here, we investigated expression of GnRH receptors (GnRHRs) and effect of GnRHa on the proliferation of cells derived from endometria and pathological lesions of women with these reproductive diseases. METHODS Biopsy specimens were collected from lesions and corresponding endometria of 35 women with pelvic endometriosis, 45 women with ovarian endometrioma, 35 women with adenomyosis and 56 women with uterine myoma during laparoscopy or laparotomy. The gene and protein expressions of GnRHR in eutopic/ectopic cells and tissues were examined by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry. The immunoreactivity of GnRHR in tissue was analysed by quantitative-histogram (Q-H) scores. The exogenous effect of GnRHa on cell proliferation was examined by 5-bromo-2-deoxyuridine incorporation assay. The Ki-67-immunoreactive cell proliferation index was analysed in biopsy specimens derived from GnRHa-treated and -non-treated women. RESULTS Types I and II GnRHRs mRNA and proteins were expressed in eutopic endometria and pathological lesions derived from women with endometriosis, adenomyosis and uterine myoma. GnRHR expression was the highest in the menstrual phase when compared with other phases of the menstrual cycle. Higher Q-H scores of GnRHR immunoreaction were found in blood-filled opaque red lesions than in other peritoneal lesions. Exogenous treatment with GnRHa significantly suppressed the proliferation of cells derived from respective endometria and pathological lesions when compared with GnRHa-non-treated cells. CONCLUSIONS Local tissue expression of GnRHR was detected in endometriosis, adenomyosis and uterine myoma. In addition to a hypo-estrogenic effect, a direct anti-proliferative effect of GnRHa may be involved in the regression of these reproductive diseases with consequent remission of clinical symptoms.