Demetra Socolov

The value of serological markers in the diagnosis and prognosis of endometriosis: a prospective case-control study

OBJECTIVE We analyzed selected well-known and less well-known serum markers that have been proposed for diagnosis and severity assessment of endometriosis, in a case-control study. STUDY DESIGN This prospective study was carried out in a Clinical Department of Gynecology in Iasi, Romania. Study participants included endometriosis patients, and controls in whom laparoscopy had excluded endometriosis. Each case and control was investigated for serum levels of CA125, TNF, IL-1, IL-6 and IL-8. The data were correlated with clinical symptoms and revised American Fertility Society (rAFS) score and stage, and interpreted by Mann-Whitney U-test and ANOVA regression analysis. RESULTS Over the course of 1 year, 24 cases of endometriosis and 24 controls of matched age were selected. The rAFS stages were: stage I, 12.5%; stage II, 16.7%; stage III, 58.3%; and stage IV, 12.5%. CA125 levels were over the cut-off of 35 IU/l in 54% of patients (versus 8% of controls), averaging 67.5 (CI95: ±17.5). The sensitivity and specificity were 54% and 91%, respectively, with a p value of <0.001 (statistically significant). For IL-6, 71% of cases and 87% of controls were above the cut-off of 2 pg/ml, with an average of 11.83 ± 7. The sensitivity and specificity were 71% and 12%, respectively, but the difference was not statistically significant, p = 0.071. Other tested serum markers had no discrimination value. A correlation with severity of endometriosis was seen for CA125 (p = 0.03) but not for IL-6, by ANOVA. CONCLUSION CA125 correlated with endometriosis screening and severity, indicating its superiority as a marker for further, larger studies.

Comparison of the pain experienced by infertile women undergoing hysterosalpingo contrast sonography or radiographic hysterosalpingography

To evaluate the pain and cause of pain experienced by women undergoing hysterosalpingography (HSG) and contrast hysterosalpingo sonography (HyCoSy) with air in a saline solution for the assessment of uterine and tubal patency.

Quantitative analysis of the relationship between microRNA‑124a, -34b and -203 gene methylation and cervical oncogenesis

Cervical cancer is a leading cause of mortality in women. Molecular and epidemiological data have unequivocally confirmed that high-risk human papillomaviruses (HPVs) are a major etiological agent of this malignancy, as host epigenetic alterations are induced in response to viral infection. The present study evaluated the methylation status of CpG islands surrounding miR-124a, miR-34b and miR-203 in 29 cervical cancer precursor lesions, 31 cervical tumors and 30 normal control samples, with the aim of identifying potential markers of cervical cancer. Direct quantitative methylation-specific PCR (qMSP) was used to evaluate the degree of methylation in the samples. HPV DNA was detected and genotyped using the Linear Array HPV Genotyping Test. Data were statistically analyzed using the Kruskal-Wallis test. Differences in miRNA hypermethylation between the tumor and control samples were highly significant for all the genes tested (p<0.0001). Significant results were also obtained regarding the hypermethylation of miR-124a and miR-203 in the precursor lesions compared to the control samples. Among the 29 patients with precursor lesions, 68.97% (20/29) presented high risk (hr)-HPV genotypes and 31.03% (9/29) were diagnosed with low risk (lr)-HPV. Significant results (p=0.0266) were obtained for miR-124a (hr-HPV group, mean 41.32; lr-HPV group, mean 6.74), revealing a strong association between the methylation process and the hr-HPV genotype. Borderline results (p=0.058) were obtained for miR-203 (hr-HPV group, mean 44.05; lr-HPV group, mean 3.33). These results confirm the involvement of epigenetic alterations in cervical oncogenesis. The lr-HPV precursor lesions had a methylation percent pattern similar to that of the normal samples, while the results for the hr-HPV precursor lesions and tumors indicate a possible involvement of the hr-HPV genotype in the miRNA methylation process.

Methylenetetrahydrofolate reductase (MTHFR) polymorphisms and promoter methylation in cervical oncogenic lesions and cancer.

The aim of this study was to investigate the role of methylenetetrahydrofolate reductase (MTHFR) polymorphisms and MTHFR methylation pattern in cervical lesions development among women from Romania, a country with high prevalence of human papillomavirus (HPV) cervical infections. To achieve this goal, blood samples and cervical cytology specimens (n = 77)/tumour tissue specimens (n = 23) were investigated. As control, blood and negative cytological smears (n = 50) were used. A statistically significant association was found between T allele of C677T polymorphism and cervical lesions, heterozygote women presenting a threefold increased risk (normal/cervical lesions and tumours: wild homozygote 34/41 (0.68/0.41), heterozygote 14/51 (0.28/0.51), mutant homozygote 2/8 (0.04/0.08); OR = 3.081, P = 0.0035). Using χ square test for the control group, the HPV‐negative and HPV‐positive patients with cervix lesions, a significant correlation between viral infection and T allele of C677T polymorphism (P = 0.0287) was found. The MTHFR promoter was methylated in all HGSIL and tumour samples, significant differences being noted between HPV‐positive samples, control group and cases of cervical dysplastic lesions without HPV DNA (P < 0. 0001) and between samples from patients with high‐risk (hr)HPV versus low‐risk (lr)HPV (P = 0.0026). No correlations between polymorphisms and methylation were observed. In Romania, individuals carrying T allele are susceptible for cervical lesions. MTHFR promoter methylation is associated with cervical severity lesions and with hrHPV.

Bacteriological agents which play a role in the development of infertility.

This study was carried out to determine the prevalence of the bacterial agents Chlamydia trachomatis (C. trachomatis), Neisseria gonorrhoeae (N. gonorrhoeae), Mycoplasma hominis (M. hominis) and Ureaplasma urealyticum (U. urealyticum) and the conditions which may play a role in the development of female infertility, in the county of Iaşi in North-Eastern Romania. Cervical and blood samples were collected from 176 infertile women and 45 pregnant women in the third trimester. Classical methods and real time PCR were applied to each cervical sample to detect the presence of these sexually transmitted microorganisms; the ELISA method was applied to blood samples to detect C. trachomatis antibodies (IgA, IgM and IgG). The proportion of C. trachomatis IgG was significantly higher in the infertile group (23.8%) than in the pregnant group (4.4%), p < 0.05. For C. trachomatis antigen (Ag) and N. gonorrhoeae Ag no differences were observed between the two groups. The prevalence of mycoplasma genital infections was higher in the pregnant group (U. urealyticum - 53.3% and M. hominis - 20%) than in the infertile group (U. urealyticum - 39.7% and M. hominis - 7.3%). Higher rate of co-infection with C. trachomatis and mycoplasma were observed among the infertile women (25.7%) than among the pregnant women (7.7%). This combination could be involved in the appearance of pelvic inflammatory disease (PID) and its sequela, including infertility. C. trachomatis IgG determination still remains the gold standard for the diagnosis of PID and should be used as a screening test for the prediction of tubal damage in infertile women. In view of the large number of cases involving the co-existence of genital infections with C. trachomatis, M. hominis and U. urealyticum, it is clearly necessary to perform screening for all three microorganisms among all women of reproductive age but especially those who are infertile.

Methylation pattern of methylene tetrahydrofolate reductase and small nuclear ribonucleoprotein polypeptide N promoters in oligoasthenospermia: a case-control study.

Alterations in DNA methylation patterns in several genes may lead to abnormal male sexual development and infertility. This study investigated the promoter methylation status of MTHFR and SNRPN in infertile men from Romania by quantitative methylation-specific PCR in order to investigate possible correlations with sperm abnormalities. The study groups included patients (n=27) with a median age of 31 years (range 26-41 years) as well as controls (n=11) with a median age of 30 years (range 24-37 years) recruited from couples seeking advice for infertility. DNA was isolated from sperm samples and promoter methylation was assessed using direct. Significant trends were detected for both genes that indicate a tendency towards promoter hypermethylation in spermatozoa with low motility (MTHFR P=0.0032, r=0.23; SNRPN P=0.0003, r=0.32) and poor morphology (MTHFR P=0.0012, r=0.27; SNRPN P=0.0003, r=0.33) but no trend was found in cases of low sperm count (MTHFR r=0.007; SNRPN r=0.06). The data indicate that the methylation patterns of the promoters of MTHFR and SNRPN are associated with changes in sperm motility and morphology, which could lead to male infertility. A large number of studies are now focused on the causes of male infertility. Among these are epigenetic modifications, which are important contributors to reproductive pathology in the male by providing dynamic changes of the phenotype according to the environmental and metabolic factors. The most known epigenetic modification is DNA methylation and alterations in this pattern in several genes could induce male infertility. The present study aims to investigate the promoter methylation status of the genes for methylene tetrahydrofolate reductase (MTHFR) and small nuclear ribonucleoprotein polypeptide N (SNRPN) in infertile males from Romania, in order to establish a correlation with sperm parameters. MTHFR is an enzyme involved in the folate pathway and in de novo nucleotide biosynthesis but also a good example for gene-environment interaction in phenotype development. SNRPN is involved in both somatic cell expression and inheritance of the imprint and the methylation pattern of its gene seems to correlate not only with imprinted disorders but also with infertility. Our study includes patients (n=27, median age 31 years, range 26-41 years) recruited from men seeking advice for couple infertility and control group (n=11, median age 30.5 years, range 24-37 years). The data we obtained indicated significant correlations between hypermethylation of the investigated genes and sperm motility and morphology. No significant correlation between DNA methylation and sperm number was found. Our data suggest that methylation pattern of MTHFR and SNRPN is linked with sperm anomalies of motility and morphology and therefore male infertility.

Levonorgestrel releasing-intrauterine system for the treatment of menorrhagia and/or frequent irregular uterine bleeding associated with uterine leiomyoma

ABSTRACT Objective To evaluate the effectiveness of the levonorgestrel releasing-intrauterine system (LNG-IUS) in the treatment of menorrhagia and/or frequent irregular uterine bleeding in women with uterine myomas. Study design Prospective study whereby 102 women with intramural myomas (in a few cases associated with submucous or subserous myomas), suffering from menorrhagia and/or frequent irregular uterine bleeding, were evaluated by means of the Pictorial Blood Assessment Chart (PBAC; Higham score) and ultrasound, three, six and 12 months after insertion of a LNG-IUS. Results The PBAC score dropped (from a mean value of 231.7 to 17.6 at 12 months). The duration of menstrual bleeding diminished significantly (p < 0.001). Uterine volume decreased from a mean of 145 cm³ to 129 cm³ at 12 months (p = 0.01). Changes in the volume of the myomas were not statistically significant (p = 0.23). Satisfaction rate was good in 91 cases (89%), fair in four cases (4%), and poor in seven cases (7%). During the one-year period of follow-up, 11 cases of expulsion or removal of the LNG-IUS were recorded. Conclusion The LNG-IUS is effective in controlling heavy menorrhagia and/or frequent irregular uterine bleeding related to the presence of myomas, but has no significant effect on the size of the tumours.

Type-specific human papillomavirus detection in cervical smears in Romania.

Anton G, Peltecu G, Socolov D, Cornitescu F, Bleotu C, Sgarbura Z, Teleman S, Iliescu D, Botezatu A, Goia CD, Huica I, Anton A‐C. Type‐specific human papillomavirus detection in cervical smears in Romania. APMIS 2010.

Small biparietal diameter in the first trimester and pregnancy outcome

The biparietal diameter (BPD) should be measured as part of the routine first-trimester ultrasound examination1. Several recent reports have suggested that identification of a small BPD at 11–14 weeks’ gestation, adjusted for crown–rump length, may pick up 60–70% of fetuses with spina bifida, with a 5–10% false-positive rate2–4. However, introducing a policy of BPD measurement at an early gestational age, when termination of pregnancy may be considered, may result in unnecessary anxiety in women and health practitioners if a small BPD is noted. The association between small BPD in the first trimester and adverse pregnancy outcome is controversial; the finding has been variously associated with chromosomal anomalies5 and small-for-gestational-age (SGA) newborns. This prompted us to analyze the outcomes of fetuses with small BPD measurements on the first-trimester scan. We performed a retrospective study of women who had undergone routine first-trimester ultrasound examination at Necker Hospital, Paris, between 1 January 2009 and 1 January 2012. Of 6584 patients, 437 (6.6%) were lost to follow-up. Of the remaining 6147 patients with known pregnancy outcomes, 799 (13.0%), 432 (7.0%) and 246 (4.0%) had a fetus with a BPD below the 10th, 5th and 2.5th percentile, respectively. In the subgroup lost to follow-up, 66 fetuses (15.1%) had a BPD below the 10th percentile (P = 0.27). We found a strong association between BPD ≤ 10th percentile and risk of spina bifida (n = 5/9; odds ratio (OR) 8.41 (95% confidence interval (CI) 1.97–37.2), P = 0.0029), using a significance level of P < 0.05. This measurement was not associated with other adverse outcomes, including early miscarriage (n = 1/29; OR, 0.24 (95% CI, 0.01–1.63), P = 0.16), intrauterine fetal death (n = 7/45; OR, 1.57 (95% CI, 0.63–3.75), P = 0.32), fetal and newborn malformations (n = 30/201; OR, 1.18 (95% CI, 0.79–1.75), P = 0.31), abnormal karyotype (n = 8/53; OR, 1.19 (95% CI, 0.52–2.64), P = 0.64), termination of pregnancy (n = 20/124; OR 1.29 (95% CI, 0.77–2.15), P = 0.29), SGA (n = 125/868; OR, 1.15 (95% CI, 0.93–1.42), P = 0.18) or preterm delivery (n = 88/597; OR, 1.18 (95% CI, 0.92–1.50), P = 0.18). There was a similar lack of association between first-trimester BPD and adverse outcomes for measurements below the 5th and 2.5th percentiles (Table S1). Given the low incidence of adverse pregnancy outcomes after 11 weeks, detecting a 50% increase in the composite adverse outcome risk (from 2 to 3%, for example) would require more than 3500 fetuses with BPD measurements under the 10th percentile. However, our study does not support any strong association between a small BPD in the first trimester and adverse pregnancy outcomes, other than for spina bifida. A small BPD at 11–14 weeks’ gestation should prompt a careful examination of the fetal spine. In the absence of a neural tube defect, a small first-trimester BPD does not seem to be associated with other adverse outcomes, and patients can be immediately reassured, even if the value is below the 5th or 2.5th percentile.

Vitamin D Status: A Different Story in the Very Young versus the Very Old Romanian Patients

Background In Romania (latitude 48°15’N to 43°40’N), vitamin D supplementation is common practice mostly in infants 0-1 year old. No published information is available regarding epidemiological data on vitamin D status in the Romanian population for a wide age range and geographical territory. In this context, we aimed to evaluate the seasonal and age variation of vitamin D status in a large Romanian population. Methods 6631 individuals from across Romania had performed 7544 vitamin D assessments (2012-2014) in a chain of private laboratories. Vitamin D (25-hydroxyvitamin D2 and 25-hydroxyvitamin D3) was measured using High Performance Liquid Chromatography. Vitamin D levels were classified as severe deficiency<10ng/mL, deficiency 10-20ng/mL, insufficiency 21-29ng/mL, sufficiency≥30ng/mL and potentially harmful>100ng/ml. Results Male to female ratio was 1:2.9. Age ranged from 0 to 85 years. Mean vitamin D levels increased from April (26.3ng/ml) to September (35.6ng/ml) and decreased from October (33.5ng/ml) to March (24.4 ng/ml). Overall 40% had sufficient vitamin D, while the rest were insufficient 33%, deficient 22%, severely deficient 4% and 1% potentially harmful (of them 81% under 1 year old). Males compared to females showed higher percentages of sufficiency (47% vs. 38%). Children 0- 2 years presented the highest percentage of vitamin D sufficiency (77%). Lowest percentages (21%) of sufficiency were in people 80-84 years. Conclusion In Romania, suboptimal vitamin D levels are common (59%), especially in older age, wintertime and in women. Vitamin D supplementation would be most warranted from January to April in the Romanian population. 25-hydroxyvitamin D levels>100ng/ml were relatively prevalent in children 0-1 year old (17.3%). This was attributed to supplementation errors and the fact that high-risk individuals were more likely to visit for medical check-up. Nonetheless, it stresses the need to increase awareness of the importance of preventing Vitamin D supplementation administration errors in the young.