Ivo Noci

HLA-G expression in early embryos is a fundamental prerequisite for the obtainment of pregnancy.

Different mechanisms mediated by the expression of the HLA‐class Ib HLA‐G products are suggested to account for the induction of immune tolerance against the paternal antigens of the fetus during pregnancy. Soluble HLA‐G antigens, mainly produced by cytotrophoblast cells at the materno‐fetal interface and circulating in the body fluids, show a capacity analogous to that of membrane‐boundstructures to inhibit NK cells. In the present report we have investigated, using specific ELISA, the presence of sHLA‐G molecules in culture supernatants of early embryos obtained by in vitro fertilization (IVF) before transfer. The data obtained from the analysis of 285 supernatants corresponding to 101 IVF procedures (43 IVF, 58 intracytoplasmic sperm injection) identify two groups of patients on the basis of sHLA‐G antigen presence. No differences in clinical parameters were observed between the groups, but positive embryo implantations occurred only in women showing sHLA‐G molecules in culture supernatants (Fishers exact p value 2.56×10–3). The results obtained indicate that expression of HLA‐G products in embryo cells is a mandatory, but not sufficient, prerequisite for the development of pregnancy.

HERG potassium channels are more frequently expressed in human endometrial cancer as compared to non-cancerous endometrium

HERG K+channels, besides contributing to regulate cardiac and neuronal excitability, are preferentially expressed in tumour cell lines of different histogenesis, where their role in the development and maintenance of the neoplastic phenotype is under study. We show here that both herg gene and HERG protein are expressed with high frequency in primary human endometrial cancers, as compared to normal and hyperplastic endometrium. RT-PCR and immunohistochemistry, using specific anti-HERG antibodies developed in our laboratory, were applied to tissue specimens obtained from 18 endometrial cancers and 11 non-cancerous endometrial tissues. herg RNA and HERG protein are expressed in 67% and 82%, respectively, of cancerous, while in only 18% of non-cancerous tissues. In particular, no expression was found in endometrial hyperplasia. Moreover, electrophysiological experiments confirmed the presence of functioning HERG channels on the plasma membrane of tumour cells. On the whole, these data are the first demonstration of the presence of HERG channels in primary human neoplasias, and could candidate HERG as a potential tool capable of marking cancerous versus hyperplastic endometrial growth.

Identification, localization and functional activity of oxytocin receptors in epididymis

Oxytocin (OT) is a neurohypophysial hormone with unclear physiological functions in the male. Several previous studies indicated that OT might have a role in the ejaculatory process, stimulating sperm release from the epididymal storage. In this study we investigated on the presence and function of OT receptor (OTR) in rabbit and human epididymis. By using RT-PCR, Western and binding studies, we found that OTR gene and protein is expressed in the human epididymis and stimulates in vitro contractility. The immunolocalization of OTR suggests that the receptor is not only present in the smooth muscle cells of the human epididymis but also in the epithelial compartment. Experiments performed in rabbit epididymal epithelial (rEE) cells in culture indicate that OT induces the release of an other potent stimulator of epididymal contractility, endothelin-1 (ET-1), Blocking the ET(A) subtype of the ET-1 receptors, by using a specific antagonist (BQ-123), partially counteracts the contractile effect of OT, suggesting positive interactions between the two peptides in regulating epididymal contractility. Finally, to investigate whether an acute OT administration increases sperm release also in humans, we treated oligozoospermic patients with an intravenous bolus of OT (2.5 IU), just before sperm collection. In a small, single blind study, we found that OT almost doubled sperm retrieval when compared with vehicle administration. Our results indicate that OT might have physiological functions also in the male, controlling epididymal motility and sperm progression through the male genital tract.

Non-classic sHLA class I in human oocyte culture medium.

Soluble human leukocyte antigen (sHLA) class I molecules have been described in all human fluids. These molecules play a significant role in immune function. sHLA has been shown to produce tolerance and to induce apoptosis in cytotoxic alloreactive T cells. They are also present in the supernatant of many cultured cells. Similarly, non-classic HLA class I antigens in soluble form are present in human fluids. Among these, HLA-G is the most important because of its location in fetal tissue that suggests maternal immunological tolerance of the fetal semiallograft. In our present study we show that using two monoclonal antibodies, w6/32 and TP25.99, in the enzyme-linked immunosorbent assay allows the detection of non-classic sHLA class I molecules in the medium from human embryo cultures. The sample were collected from oocytes cultures. Oocyte donors were 11 women attending the in vitro fertilization program. The results showed a significant association (chi2 = 9.66, p = 0.002) between sHLA antigens and the oocyte cleavage rate measured 48 h after fertilization.

Ultrasonographic and clinical correlates of seminal plasma interleukin-8 levels in patients attending an andrology clinic for infertility

This study was aimed at evaluating the association between seminal plasma interleukin-8 (sIL-8) and colour-Doppler ultrasound (CDU) characteristics of the male genital tract in a series of patients fulfilling the criteria of male accessory gland infections (MAGI). Of 250 subjects seeking medical care for couple infertility, 79 (mean age: 36.4 ± 7.5 years) met the criteria of MAGI and scored higher than the rest of the sample on the National Institutes of Health-Chronic Prostatitis Symptom Index score. All patients underwent simultaneous hormone evaluation and seminal analysis (including sIL-8), along with scrotal and transrectal CDU before and after ejaculation. After adjusting for age, sIL-8 in patients with MAGI was significantly related to several abnormal semen and CDU parameters. In particular, leucocytospermia was closely associated with sIL-8. Ejaculate volume, unlike other semen or hormonal parameters, was negatively associated with sIL-8. When scrotal CDU was performed, sIL-8 was positively related to CDU inhomogeneous, hypo-echoic, hyper-echoic epididymis and to epididymal calcifications. In addition, a positive correlation among sIL-8, hyperaemic epididymis and an increased size of epididymal tail was found. When transrectal CDU was performed, an association among sIL-8 and hyper-echoic seminal vesicles, dilated ejaculatory ducts and duct calcifications was also observed. Finally, sIL-8 was positively related to prostate CDU abnormalities such as calcifications, inhomogeneous/hypo-echoic texture, hyperaemia and high arterial blood flow. No association was found with testis parameters. In conclusion, sIL-8 levels in patients with MAGI are associated with several parameters and CDU abnormalities of epididymis, seminal vesicles, ejaculatory ducts and prostate, but not of the testis. Furthermore, sIL-8 positively correlates with CDU signs of ejaculatory duct inflammatory subobstruction.

Elevated body mass index correlates with higher seminal plasma interleukin 8 levels and ultrasonographic abnormalities of the prostate in men attending an andrology clinic for infertility

Background: Obesity is associated with a systemic, low-grade inflammatory state. Although the relationship between obesity and semen parameters or prostate diseases has been previously investigated, the association between body mass index (BMI), prostate inflammatory diseases and color-Doppler ultrasound (CDU) of the male genital tract (MGT) has been poorly studied. Aim: To evaluate the association between BMI and CDU features of the MGT, signs and symptoms of prostate inflammation, semen parameters. Materials/Subjects and methods: We studied 222 men seeking medical care for couple infertility. According to the World Health Organization classification, subjects were divided into 3 groups: normal weight (no.=131, BMI=18.5–24.9 kg/m2), overweight (no.=71, BMI=25.0–29.9 kg/m2), obese (no.=20, BMI≥30.0 kg/m2). All patients underwent simultaneous testosterone evaluation and seminal analysis, including interleukin 8 (sIL-8), along with scrotal and transrectal CDU, before and after ejaculation. Prostatitis symptoms were evaluated by National Institutes of Health-Chronic Prostatitis Symptom Index questionnaire. Results: After adjusting for age and testosterone levels, higher BMI was significantly related to higher prostate volume and several CDU features of the prostate, including macro-calcifications, inhomogeneity, higher arterial peak systolic velocity (the latter adjusted also for blood pressure), but not with abnormalities of testis, epididymis, seminal vesicles. Furthermore, higher BMI and BMI class were significantly related to higher sIL-8, a reliable surrogate marker of prostate inflammatory diseases, even after adjustment for age. Conversely, no associations among BMI, clinical symptoms of prostatitis or semen parameters were observed. Conclusions: Subjects with higher BMI might develop CDU and biochemical signs suggestive of prostate inflammation, although not clinically overt.

Hormonal patterns, steroid receptors and morphological pictures of endometrium in hyperstimulated IVF cycles

OBJECTIVE The purpose of this contribution is to investigate the pathophysiology of the abnormal endometrial development in hyperstimulated IVF cycles. STUDY DESIGN In 12 IVF-patients who did not have embryo transfer because of failure of oocyte fertilization, serum values of 17 beta-estradiol, progesterone, FSH, LH, total and free testosterone, and androstenedione were measured on the pick-up day and were evaluated with respect to the values normally expressed in the day of ovulation; in the endometrial specimens collected 2 days later, at the time of embryo replacement, estrogen and progesterone receptors were immunohistochemically determined and dating by the Noyes method was performed. RESULTS 17 beta-Estradiol values are constantly higher, and progesterone levels are, only in four cases, higher than expected for the day of ovulation in a natural cycle. These hormonal patterns can only partially explain the pattern of steroid receptors: progesterone receptors are expressed sparsely both in glands and stroma, while estrogen receptors are abundant in the glands and absent in the stroma. In 11 of 12 patients an abnormal endometrial development with stromal advancement was observed: this morphological picture of the endometrium could partially be explained only in the four cases presenting high progesterone levels by serum values and endometrial receptor content of estrogen and progesterone. CONCLUSIONS The abnormal endometrial development in hyperstimulated IVF cycles could only in part be explained by estrogen and progesterone, and other factors have to be considered.

Lack of Histocompatibility Leukocyte Antigen-G expression in early embryos is not related to germinal defects or impairment of interleukin-10 production by embryos

The expression of Histocompatibility Leukocyte Antigen (HLA)-G molecules is a mandatory prerequisite for the development of pregnancy but no hypotheses have yet been advanced regarding the lack of HLA-G modulation expression in a percentage of early embryos obtained by in vitro fertilization (IVF). One possible hypothetical model assumes that the absence of regulation of HLA-G or impaired interleukin (IL)-10 secretion could be related to germinal defects. We investigated the presence of soluble HLA-G antigens in supernatants of single embryo cultures from couples admitted to a second fertilization procedure; these couples showed a complete absence of HLA-G modulation in the first cycles embryo supernatants (0/31). The results obtained in the second IVF cycle showed embryo supernatants positive for HLA-G (14/40), suggesting that the previous lack of antigen modulation is independent of germinal defects. Furthermore, since it has been reported that oocytes and early embryos can secrete IL-10, an anti-inflammatory cytokine produced by type 2 helper T cells that induces upregulation of HLA-G expression in monocytes and trophoblasts, we investigated the levels of IL-10 and soluble HLA-G in 40 embryo culture supernatants from 21 IVF cycles. No associations were observed between the presence of IL-10 and the production and concentrations of soluble HLA-G, or between IL-10 levels and pregnancy outcome. These results indicate that the lack of HLA-G production in early embryos is not related to germinal defects or to impairment in embryo IL-10 secretion but could be ascribed to possible uncorrected fertilization processes.

I. Aging of the human endometrium: a basic morphological and immunohistochemical study

OBJECTIVE To evaluate if human endometrium presents morphological variations suggestive of an age-related decline in endometrial receptivity. STUDY DESIGN Peri-implantation endometrium of younger (<30 years of age: n = 13) and older (>40 years of age: n = 17) normally menstruating women was studied. Endometrial specimens were routinely fixed in buffered formalin and embedded in paraffin. Sections (5 mu m) were stained with hematoxylin-eosin, periodic acid-Schiff (PAS) and Trichrome conforming to Masson according to conventional histologic examination. Several consecutive sections were used for the following immunohistochemical study: vascular localization (CD34), cellular proliferation index (PCNA), progesterone and estrogen receptors. RESULTS Using both the traditional morphological evaluation and monoclonal antibodies, no significant differences were found between the endometria of women <30 years of age and those of women >40. CONCLUSIONS Our results suggest that human endometrium does not age, at least while cyclic hormonal stimulation and menstruation are present.

Tamoxifen and Oligospermia

Thirty normogonadotropic oligospermic males were administered Tamoxifen (20 mg/day) for 4 months. A complete evaluation of seminal parameters before and after treatment was performed. An increase of both mean sperm concentration and total sperm count was noted after treatment with Tamoxifen (p less than 0.05), whereas no variation was observed for semen volume, sperm morphologic characteristics, or sperm motility. Hormonal patterns of 5 of these subjects before, during, and after 3 months of treatment with Tamoxifen showed a progressive increase of gonadotropin (especially LH) and 17 beta E2 values; testosterone variations were minimal, and PRL showed a slight decrease.