Ladislav Krofta

Absolute and Relative Quantification of Placenta-Specific MicroRNAs in Maternal Circulation with Placental Insufficiency–Related Complications

Placental insufficiency-related complications are one of the leading causes of maternal and perinatal morbidity and mortality. This study investigated the quantification of placenta-specific microRNAs (miRNAs) in the maternal circulation during gestation in a cohort of women with normally progressing pregnancies, the differentiation between placental insufficiency-related complications and normally progressing pregnancies, and the differentiation between placental insufficiency and normally progressing pregnancies during the early stages of gestation. Both absolute and relative quantification of placenta-specific miRNAs (ie, miR-516-5p, miR-517*, miR-518b, miR-520a*, miR-520h, miR-525, and miR-526a) was determined in 50 women with normally progressing pregnancies, 32 with complicated pregnancies [21 with preeclampsia with or without intrauterine growth retardation (IUGR) and 11 with IUGR], and 7 women with pregnancies at various gestational stages who later developed preeclampsia and/or IUGR using real-time PCR and a comparative C(T) method relative to normalization factor (ie, geometric mean of ubiquitous miR-16 and let-7d). Both quantification approaches revealed significant increases in extracellular placenta-specific miRNA levels over time in women with normally progressing pregnancies; however, they were not able to differentiate between normally progressing and complicated pregnancies at the time of preeclampsia and/or IUGR onset. Nevertheless, significant elevation of extracellular miRNA levels was observed during early gestation (ie, within the 12th to 16th weeks) in pregnancies with later onset of preeclampsia and/or IUGR. Early gestation extracellular miRNA screening can differentiate between women with normally progressing pregnancies and those who may later develop placental insufficiency-related complications.

A prospective randomized comparison of vaginal hysterectomy, laparoscopically assisted vaginal hysterectomy, and total laparoscopic hysterectomy in women with benign uterine disease

OBJECTIVES To compare the clinical results of three minimally invasive hysterectomy techniques: vaginal hysterectomy (VH), laparoscopically assisted vaginal hysterectomy (LAVH), and total laparoscopic hysterectomy (TLH). STUDY DESIGN A prospective, randomized study was performed at a tertiary care center between March 2004 and October 2005. A total of 125 women indicated to undergo hysterectomy for benign uterine disease were randomly assigned to three different groups (40 VH, 44 LAVH, and 41 TLH). Outcome measures, including operating time, blood loss, rate of complications, inflammatory response, febrile morbidity, consumption of analgesics, and length of hospital stay, were assessed and compared between groups. RESULTS Vaginal hysterectomy had the shortest operating time (66 min) and smallest drop in hemoglobin. However, there were technical problems with salpingo-oophorectomy from the vaginal approach (3/20 cases) and this group had a significantly higher rate of febrile complications (20%) compared to LAVH (2.3%) and TLH (7.3%). The increase in inflammatory markers was higher in vaginal hysterectomy patients. Laparoscopically assisted vaginal hysterectomy had an acceptable operating time (85 min), a low complication rate, lack of severe post-operative complications, and the lowest consumption of analgesics. However, it had the highest blood loss. Total laparoscopic hysterectomy had the longest operating time (111 min) and severe complications occurred only in this group. Conversions to another hysterectomy method occurred in all three groups, most of these conversions were to LAVH. CONCLUSIONS Based on our results, in women with non-malignant disease of the uterus, LAVH and VH seem to be the preferred hysterectomy techniques for general gynecological surgeons. Vaginal hysterectomy had the shortest operating time and least drop in hemoglobin, making it a suitable method for women for whom the shortest duration of surgery and anesthesia is optimal. LAVH is a versatile procedure, combining the advantages of both the vaginal and laparoscopic approach, and is preferable in cases when oophorectomy is required. Total laparoscopic hysterectomy did not appear to offer any significant benefits over the other two methods and should be strictly indicated in women where neither VH nor LAVH are feasible and should only be performed by very experienced laparoscopists.

A multicenter, randomized, prospective, controlled study comparing sacrospinous fixation and transvaginal mesh in the treatment of posthysterectomy vaginal vault prolapse

OBJECTIVE The objective of the study was to compare recurrence and complication rates for sacrospinous fixation (SSF) and prolene mesh techniques for the primary treatment of posthysterectomy vaginal vault prolapse. STUDY DESIGN Patients undergoing surgery for vault prolapse were included in a multicenter, randomized, controlled study comparing SSF or total mesh (Prolift; Gynecare/Ethicon, Somerville, NJ). The examination included pelvic organ prolapse quantification, urodynamics, ultrasound, and quality-of-life (QoL) questionnaires before and 3 and 12 months after surgery. RESULTS Of 168 randomized patients, 83 underwent SSF and 85 mesh repair. Prolapse recurrence after 12 months occurred in 39.4% of the SSF group and in 16.9% of the mesh group (P = .003). The mesh exposure rate was 20.8%. No difference in QoL improvement as well as of de novo stress urinary incontinence and overactive bladder onset was found. CONCLUSION Mesh exposure occurrence was balanced against a lower prolapse recurrence rate in the patients undergoing mesh surgery compared with those undergoing SSF.

Circulating C19MC MicroRNAs in Preeclampsia, Gestational Hypertension, and Fetal Growth Restriction

The objective of the study was to identify the profile of circulating C19MC microRNAs (miR-516-5p, miR-517∗, miR-518b, miR-520a∗, miR-520h, miR-525, and miR-526a) in patients with established preeclampsia (n = 63), fetal growth restriction (n = 27), and gestational hypertension (n = 23). We examined the correlation between plasmatic concentrations and expression levels of microRNAs and the severity of the disease with respect to clinical signs, requirements for the delivery, and Doppler ultrasound parameters. Using absolute and relative quantification approaches, increased extracellular C19MC microRNA levels (miR-516-5p, P = 0.037, P = 0.009; miR-517∗, P = 0.033, P = 0.043; miR-520a∗, P = 0.001, P = 0.009; miR-525, P = 0.026, P = 0.01; miR-526a, P = 0.03, P = 0.035) were detected in patients with preeclampsia. The association analysis pointed to no relationship between C19MC microRNA plasmatic concentrations and expression profile and identified risk factors for a poorer perinatal outcome. However, the dependence between the levels of plasmatic C19MC microRNAs and the pulsatility index in the middle cerebral artery and the values of cerebroplacental ratio was demonstrated. The study brought the interesting finding that the upregulation of miR-516-5p, miR-517∗, miR-520a∗, miR-525, and miR-526a is a characteristic phenomenon of established preeclampsia.

Bilateral avulsion of the puborectal muscle: magnetic resonance imaging‐based three‐dimensional reconstruction and comparison with a model of a healthy nulliparous woman

Obstetric trauma to the puborectal muscle seems to be an important cause of pelvic floor dysfunction in women. Due to the complicated three‐dimensional (3D) arrangement of the pelvic structures, two‐dimensional images are not sufficient to demonstrate its relationships in a complex fashion. Thus, we aimed to create a 3D computer model to visualize the normal female pelvic floor anatomy and to compare this with the anatomy after bilateral avulsion of the puborectal muscle following delivery.

Cardiovascular and Cerebrovascular Disease Associated microRNAs Are Dysregulated in Placental Tissues Affected with Gestational Hypertension, Preeclampsia and Intrauterine Growth Restriction.

Aims To demonstrate that pregnancy-related complications are associated with alterations in cardiovascular and cerebrovascular microRNA expression. Gene expression of 32 microRNAs (miR-1-3p, miR-16-5p, miR-17-5p, miR-20a-5p, miR-20b-5p, miR-21-5p, miR-23a-3p, miR-24-3p, miR-26a-5p, miR-29a-3p, miR-33a-5p, miR-92a-3p, miR-100-5p, miR-103a-3p, miR-122-5p, miR-125b-5p, miR-126-3p, miR-130b-3p, miR-133a-3p, miR-143-3p, miR-145-5p, miR-146a-5p, miR-155-5p, miR-181a-5p, miR-195-5p, miR-199a-5p, miR-208a-3p, miR-210-3p, miR-221-3p, miR-342-3p, miR-499a-5p, and miR-574-3p) was assessed in placental tissues, compared between groups (35 gestational hypertension, 80 preeclampsia, 35 intrauterine growth restriction and 20 normal pregnancies) and correlated with the severity of the disease with respect to clinical signs, delivery date, and Doppler ultrasound parameters. Initially, selection and validation of endogenous controls for microRNA expression studies in placental tissues affected by pregnancy-related complications have been carried out. Results The expression profile of microRNAs was different between pregnancy-related complications and controls. The up-regulation of miR-499a-5p was a common phenomenon shared between gestational hypertension, preeclampsia, and intrauterine growth restriction. Preeclamptic pregnancies delivering after 34 weeks of gestation and IUGR with abnormal values of flow rate in the umbilical artery demonstrated up-regulation of miR-1-3b. Preeclampsia and IUGR requiring termination of gestation before 34 weeks of gestation were associated with down-regulation of miR-26a-5p, miR-103a-3p and miR-145-5p. On the other hand, some of microRNAs (miR-16-5p, miR-100-5p, miR-122-5p, miR-125b-5p, miR-126-3p, miR-143-3p, miR-195-5p, miR-199a-5p, miR-221-3p, miR-342-3p, and miR-574-3p) were only down-regulated or showed a trend to down-regulation just in intrauterine growth restriction pregnancies requiring the delivery before 34 weeks of gestation. Conclusion Epigenetic changes induced by pregnancy-related complications in placental tissue may cause later onset of cardiovascular and cerebrovascular diseases in offspring.

Gestational hypertension, preeclampsia and intrauterine growth restriction induce dysregulation of cardiovascular and cerebrovascular disease associated microRNAs in maternal whole peripheral blood.

AIMS To demonstrate that pregnancy-related complications are associated with alterations in cardiovascular and cerebrovascular microRNA expression. Gene expression of 29 microRNAs (miR-1-3p, miR-16-5p, miR-17-5p, miR-20a-5p, miR-20b-5p, miR-21-5p, miR-23a-3p, miR-24-3p, miR-26a-5p, miR-29a-3p, miR-92a-3p, miR-100-5p, miR-103a-3p, miR-122-5p, miR-125b-5p, miR-126-3p, miR-130b-3p, miR-133a-3p, miR-143-3p, miR-145-5p, miR-146a-5p, miR-181a-5p, miR-195-5p, miR-199a-5p, miR-210-3p, miR-221-3p, miR-342-3p, miR-499a-5p, and miR-574-3p) was assessed in maternal whole peripheral blood, compared between groups (39 gestational hypertension, 68 preeclampsia, 33 intrauterine growth restriction and 20 normal pregnancies) and correlated with the severity of the disease with respect to clinical signs, delivery date, and Doppler ultrasound parameters. Initially, selection and validation of endogenous controls for microRNA expression studies in patients affected by pregnancy-related complications have been carried out. RESULTS The expression profile of microRNAs was different between pregnancy-related complications and controls. The down-regulation of miR-100-5p, miR-125b-5p and miR-199a-5p was a common phenomenon shared between gestational hypertension, preeclampsia, and intrauterine growth restriction. Moreover, IUGR pregnancies induced down-regulation of miR-17-5p, miR-146a-5p, miR-221-3p and miR-574-3p in maternal circulation. Irrespective of the severity of the disease, preeclampsia was associated with the dysregulation of miR-100-5p and miR-125b-5p and IUGR with dysregulation of miR-199a-5p. Preeclampsia requiring termination of gestation before 34 weeks was associated with down-regulation of miR-146a-5p, miR-199a-5p and miR-221-3p. Weak negative correlation between miR-146a-5p and miR-221-3p expression and the pulsatility index in the umbilical artery was found. Additional microRNAs (miR-103a-3p, miR-126-3p, miR-195-5p and miR-499a-5p) showed a trend to down-regulation in appropriate pregnancy-related complications. CONCLUSION Epigenetic changes are induced by pregnancy-related complications in maternal whole peripheral blood.

Endopelvic Fascia in Women : Shape and Relation to Parietal Pelvic Structures

OBJECTIVE: The endopelvic fascia is a confluent suspensory apparatus of the female pelvic organs. The aim of the study was to construct a three-dimensional model of the endopelvic fascia, defining its shape and its connections to the surrounding parietal structures. METHODS: We created a three-dimensional multiple-source computer model to simultaneously visualize and analyze all the structures within the female pelvic floor. This model integrates data from magnetic resonance imaging of 15 nulliparas under age 30 with no symptoms of pelvic floor dysfunction. The model also includes data from direct observation in the dissection laboratory and in surgical rooms, together with the relevant scientific literature. RESULTS: The endopelvic fascia has the shape of a semifrontally oriented septum, which surrounds the vagina and part of the uterine cervix and divides the pelvic floor into the anterior and posterior compartments. This confluent septum has specific connections to the pubic bone, anterior perineal membrane, perineal body, and superior fascia of the levator ani muscle. Additionally, the uterosacral part of the septum has three subdivisions— the “vascular part,” the “neural part,” and the true uterosacral ligament. Each of these subdivisions has a different physical link to the parietal structures. Three-dimensional illustrations and schemes were created to facilitate the understanding of the anatomy of these complex structures. CONCLUSION: Connecting descriptions of the geometry of the organs visible by magnetic resonance imaging with descriptions of their individual connections to the endopelvic fascia gave us unique information about the three-dimensional representation of the anatomy of the female lesser pelvis. The endopelvic fascia divides the lesser pelvis in a manner that is similar to the way the urorectal septum divides the embryonic cloaca. LEVEL OF EVIDENCE: III

TVT-S for surgical treatment of stress urinary incontinence: prospective trial, 1-year follow-up

Introduction and hypothesisThe aim of this study was to evaluate the effectiveness of and morbidity associated with the tension-free vaginal tape-secur (TVT-S) procedure in women with stress urinary incontinence (SUI).MethodsWe performed a prospective trial, examining 86 women with primary SUI. Eighty-two patients had a 1-year follow-up (dropout rate = 4.6%). The preoperative evaluation included urinalysis, urodynamic studies, and validated questionnaires. The 1-year outcome evaluation also included a 1-h pad testing.ResultsAt the 1-year follow-up, 43 (52.4%) women were objectively cured, and 14 (17.1%) women were objectively improved. Subjectively, 49 (59.7%) patients did not experience urine loss, and 18 (22.2%) women improved in this respect. Postoperative de novo urge incontinence symptoms developed in 24.4% (n = 20) of patients. Vaginal defect healing occurred in 6.1% (n = 5) of patients, and one (1.2%) case of urethral erosion was reported.ConclusionObjective and subjective cure rates following TVT-S are inferior to other tape procedures.

Quantification of extracellular DNA using hypermethylated RASSF1A, SRY, and GLO sequences - evaluation of diagnostic possibilities for predicting placental insufficiency.

This study evaluated quantification of fetal extracellular DNA in maternal plasma for differentiation between cases at risk of onset of placental-insufficiency-related complications and normal pregnancies. Using real-time polymerase chain reaction, fetal (sex-determining region Y [SRY] and hypermethylated RASSF1A sequence) and total (beta-globin [GLO] gene) extracellular DNA was examined in 70 normal pregnancies, 18 at risk of placental-insufficiency-related pregnancy complications, 24 preeclampsia with or without (w or w/o) intrauterine growth retardation (IUGR) (median 34.0 week), and 11 IUGR (median 28.5 week). IUGR was diagnosed when estimated fetal weight was below the 10th percentile for evaluated gestational age. Although increased levels of extracellular DNA were detected in pregnancies with preeclampsia w or w/o IUGR relative to controls (RASSF1A, p < 0.001; SRY, p = 0.009; GLO, p < 0.001), quantities of fetal extracellular DNA in IUGR were not statistically significant (RASSF1A, p = 0.21; SRY, p = 0.2). RASSF1A, SRY, and GLO achieved 93.1%, 93.6%, and 92.1% accuracy for differentiation between normal pregnancy and preeclampsia w or w/o IUGR. Lower sensitivity was observed for pregnancies with onset of IUGR (RASSF1A, 60.0%; SRY, 80.0%; GLO, 72.7%), but did not influence final accuracy (RASSF1A, 91.6%; SRY, 92.5%; GLO, 89.5%). Among 18 patients at risk, 8 pregnancies involving 3 female and 5 male fetuses developed preeclampsia (n = 4), IUGR (n = 3), and chronic placentopathy causing hypoxia (n = 1). Elevation of extracellular DNA was demonstrated in 3/5 (SRY), 1/8 (hypermethylated RASSF1A), and 4/8 (GLO) patients at the earliest 26 weeks and at the latest 2 weeks before the onset of symptoms. These data indicate that fetal and total extracellular DNA concentrations can be significantly elevated in plasma of patients who later developed placental-insufficiency-related pregnancy complications. However, this is strongly individualized, and not a rule for all cases, and probably depends on the actual occurrence of excessive placental trophoblast apoptosis.