Cyril Huissoud

Precursor Diversity and Complexity of Lineage Relationships in the Outer Subventricular Zone of the Primate

Long-term ex vivo live imaging combined with unbiased sampling of cycling precursors shows that macaque outer subventricular zone (OSVZ) includes four distinct basal radial glial (bRG) cell morphotypes, bearing apical and/or basal processes in addition to nonpolar intermediate progenitors (IPs). Each of the five precursor types exhibits extensive self-renewal and proliferative capacities as well as the ability to directly generate neurons, albeit with different frequencies. Cell-cycle parameters exhibited an unusual stage-specific regulation with short cell-cycle duration and increased rates of proliferative divisions during supragranular layer production at late corticogenesis. State transition analysis of an extensive clonal database reveals bidirectional transitions between OSVZ precursor types as well as stage-specific differences in their progeny and topology of the lineage relationships. These results explore rodent-primate differences and show that primate cortical neurons are generated through complex lineages by a mosaic of precursors, thereby providing an innovative framework for understanding specific features of primate corticogenesis.

High-dose tranexamic acid reduces blood loss in postpartum haemorrhage

IntroductionOur purpose in conducting this study was to determine whether administration of high-dose tranexamic acid (TA) at the time of diagnosis of postpartum haemorrhage (PPH) could reduce blood loss.MethodsThis was a randomised, controlled, multicentred, open-label trial. Women with PPH >800 mL following vaginal delivery were randomly assigned to receive TA (loading dose 4 g over 1 hour, then infusion of 1 g/hour over 6 hours) or not. In both groups, packed red blood cells (PRBCs) and colloids could be used according to French guidelines. The use of additional procoagulant treatments was permitted only in cases involving intractable bleeding. The primary objective was to assess the efficacy of TA in the reduction of blood loss in women with PPH, and the secondary objectives were the effect of TA on PPH duration, anaemia, transfusion and the need for invasive procedures.ResultsA total of 144 women fully completed the protocol (72 in each group). Blood loss between enrolment and 6 hours later was significantly lower in the TA group than in the control group (median, 173 mL; first to third quartiles, 59 to 377) than in controls (221 mL; first to third quartiles 105 to 564) (P = 0.041). In the TA group, bleeding duration was shorter and progression to severe PPH and PRBC transfusion was less frequent than in controls (P < 0.03). Invasive procedures were performed in four women in the TA group and in seven controls (P = NS). PPH stopped after only uterotonics and PRBC transfusion in 93% of women in the TA group versus 79% of controls (P = 0.016). Mild, transient adverse manifestations occurred more often in the TA group than in the control group (P = 0.03).ConclusionsThis study is the first to demonstrate that high-dose TA can reduce blood loss and maternal morbidity in women with PPH. Although the study was not adequately powered to address safety issues, the observed side effects were mild and transient. A larger international study is needed to investigate whether TA can decrease the need for invasive procedures and reduce maternal morbidity in women with PPH.Trial registrationControlled Trials ISRCTN09968140.

Association between fibrinogen level and severity of postpartum haemorrhage: secondary analysis of a prospective trial

BACKGROUND The aim of the study was to determine whether the fibrinogen level at diagnosis of postpartum haemorrhage (PPH) is associated with the severity of bleeding. METHODS This is a secondary analysis of a population-based study in 106 French maternity units identifying cases of PPH prospectively. PPH was defined by a blood loss exceeding 500 ml during the 24 h after delivery or a peripartum haemoglobin decrease of more than 20 g litre(-1). This analysis includes 738 women with PPH after vaginal delivery. Fibrinogen levels were compared in patients whose PPH worsened and became severe and those whose PPH remained non-severe. Severe PPH was defined as haemorrhage by occurrence of one of the following events: peripartum haemoglobin decrease ≥ 40 g litre(-1), transfusion of concentrated red cells, arterial embolization or emergency surgery, admission to intensive care, or death. RESULTS The mean fibrinogen concentration at diagnosis was 4.2 g litre(-1) [standard deviation (sd)=1.2 g litre(-1)] among the patients without worsening and 3.4 g litre(-1) (sd=0.9 g litre(-1)) (P<0.001) in the group whose PPH became severe. The fibrinogen level was associated with PPH severity independently of other factors [adjusted odds ratio=1.90 (1.16-3.09) for fibrinogen between 2 and 3 g litre(-1) and 11.99 (2.56-56.06) for fibrinogen <2 g litre(-1)]. CONCLUSIONS The fibrinogen level at PPH diagnosis is a marker of the risk of aggravation and should serve as an alert to clinicians.

Coagulation assessment by rotation thrombelastometry in normal pregnancy

We analysed changes in coagulation during normal pregnancy with a novel point-of-care device based on thrombelastometry (ROTEM). We compared the results obtained with those of standard coagulation tests in 104 patients: 20 non-pregnant women (controls) and 84 women in the first (T1, n = 17), second (T2, n = 9) and third (T3, n = 58) trimesters of pregnancy. We measured the clotting time (CT), the maximum clot firmness (MCF), the early clot amplitude at 5 and 15 minutes (CA(5), CA(15)) and the clot lysis index (CLI(30)) with four tests containing specific reagents. (a) The INTEM test involving ellagic acid activated the intrinsic pathway and (b) the EXTEM test using tissue factor triggered the extrinsic pathway; (c) The FIBTEM test based on a platelet inhibitor (cytochalasin D) evaluated the contribution of fibrinogen to clot formation and (d) the APTEM test was similar to the EXTEM but was based on inhibition in vitro of fibrinolysis by aprotinin. CT and CLI(30) were not significantly modified during pregnancy whereas MCF, CA(5) and CA(15) (INTEM, EXTEM, FIBTEM) increased significantly between the second and third trimesters (e.g. median [interquartile range]: MCF-FIBTEM, 13 [11-16] mm vs. 19 [17-23] mm, respectively, in controls and T3, p < 0.001). EXTEM values were not significantly different from those measured with APTEM. There were significant correlations between the results obtained with ROTEM and those from standard coagulation tests. ROTEM analysis showed a marked increase in coagulability during normal pregnancy. ROTEM values may serve as the basis for future studies in pregnant women.

Accuracy of real-time polymerase chain reaction for Toxoplasma gondii in amniotic fluid.

OBJECTIVE: To provide clinicians with information about the accuracy of real-time polymerase chain reaction (PCR) analysis of amniotic fluid for the prenatal diagnosis of congenital Toxoplasma infection. METHODS: This was a prospective cohort study of women with Toxoplasma infection identified by prenatal screening in three centers routinely carrying out real-time PCR for the detection of Toxoplasma gondii in amniotic fluid. The data available were gestational age at maternal infection, types and dates of maternal treatment, results of amniocentesis and neonatal work-up and definitive infectious status of the child. We estimated sensitivity, specificity and positive and negative predictive values both overall and per trimester of pregnancy at the time of maternal infection. RESULTS: Polymerase chain reaction analysis was carried out on amniotic fluid for 261 of the 377 patients included (69%). It was accurate with the exception of four negative results in children who were infected. Overall sensitivity and negative predictive value were 92.2% (95% confidence interval [CI] 81–98%) and 98.1% (95% CI 95–99.5%), respectively. There was no significant association with the trimester of pregnancy during which maternal infection occurred. Specificity and positive predictive values of 100% were obtained for all trimesters. CONCLUSION: Real-time PCR analysis significantly improves the detection of T. gondii on amniotic fluid. It provides an accurate tool to predict fetal infection and to decide on appropriate treatment and surveillance. However, postnatal follow-up remains necessary in the first year of life to fully exclude infection in children for whom PCR results were negative. LEVEL OF EVIDENCE: III

Postpartum hemorrhage: guidelines for clinical practice from the French ă College of Gynaecologists and Obstetricians (CNGOF) in collaboration ă with the French Society of Anesthesiology and Intensive Care (SFAR)

Postpartum haemorrhage (PPH) is defined as blood loss ≥500mL after delivery and severe PPH as blood loss ≥1000mL, regardless of the route of delivery (professional consensus). The preventive administration of uterotonic agents just after delivery is effective in reducing the incidence of PPH and its systematic use is recommended, regardless of the route of delivery (Grade A). Oxytocin is the first-line prophylactic drug, regardless of the route of delivery (Grade A); a slowly dose of 5 or 10 IU can be administered (Grade A) either IV or IM (professional consensus). After vaginal delivery, routine cord drainage (Grade B), controlled cord traction (Grade A), uterine massage (Grade A), and routine bladder voiding (professional consensus) are not systematically recommended for PPH prevention. After caesarean delivery, placental delivery by controlled cord traction is recommended (grade B). The routine use of a collector bag to assess postpartum blood loss at vaginal delivery is not systematically recommended (Grade B), since the incidence of severe PPH is not affected by this intervention. In cases of overt PPH after vaginal delivery, placement of a blood collection bag is recommended (professional consensus). The initial treatment of PPH consists in a manual uterine examination, together with antibiotic prophylaxis, careful visual assessment of the lower genital tract, a uterine massage, and the administration of 5-10 IU oxytocin injected slowly IV or IM, followed by a maintenance infusion not to exceed a cumulative dose of 40IU (professional consensus). If oxytocin fails to control the bleeding, the administration of sulprostone is recommended within 30minutes of the PPH diagnosis (Grade C). Intrauterine balloon tamponade can be performed if sulprostone fails and before recourse to either surgery or interventional radiology (professional consensus). Fluid resuscitation is recommended for PPH persistent after first line uterotonics, or if clinical signs of severity (Grade B). The objective of RBC transfusion is to maintain a haemoglobin concentration (Hb) >8g/dL. During active haemorrhaging, it is desirable to maintain a fibrinogen level ≥2g/L (professional consensus). RBC, fibrinogen and fresh frozen plasma (FFP) may be administered without awaiting laboratory results (professional consensus). Tranexamic acid may be used at a dose of 1 g, renewable once if ineffective the first time in the treatment of PPH when bleeding persists after sulprostone administration (professional consensus), even though its clinical value has not yet been demonstrated in obstetric settings. It is recommended to prevent and treat hypothermia in women with PPH by warming infusion solutions and blood products and by active skin warming (Grade C). Oxygen administration is recommended in women with severe PPH (professional consensus). If PPH is not controlled by pharmacological treatments and possibly intra-uterine balloon, invasive treatments by arterial embolization or surgery are recommended (Grade C). No technique for conservative surgery is favoured over any other (professional consensus). Hospital-to-hospital transfer of a woman with a PPH for embolization is possible once hemoperitoneum is ruled out and if the patients hemodynamic condition so allows (professional consensus).

Novel primate miRNAs coevolved with ancient target genes in germinal zone-specific expression patterns

Major nonprimate-primate differences in cortico-genesis include the dimensions, precursor lineages, and developmental timing of the germinal zones (GZs). microRNAs (miRNAs) of laser-dissected GZ compartments and cortical plate (CP) from embryonic E80 macaque visual cortex were deep sequenced. The CP and the GZ including ventricular zone (VZ) and outer and inner subcompartments of the outer subventricular zone (OSVZ) in area 17 displayed unique miRNA profiles. miRNAs present in primate, but absent in rodent, contributed disproportionately to the differential expression between GZ subregions. Prominent among the validated targets of these miRNAs were cell-cycle and neurogenesis regulators. Coevolution between the emergent miRNAs and their targets suggested that novel miRNAs became integrated into ancient gene circuitry to exert additional control over proliferation. We conclude that multiple cell-cycle regulatory events contribute to the emergence of primate-specific cortical features, including the OSVZ, generated enlarged supragranular layers, largely responsible for the increased primate cortex computational abilities.

Derivation and cloning of a novel rhesus embryonic stem cell line stably expressing tau-green fluorescent protein

Embryonic stem cells (ESC) have the ability of indefinite self‐renewal and multilineage differentiation, and they carry great potential in cell‐based therapies. The rhesus macaque is the most relevant preclinical model for assessing the benefit, safety, and efficacy of ESC‐based transplantations in the treatment of neurodegenerative diseases. In the case of neural cell grafting, tracing both the neurons and their axonal projections in vivo is essential for studying the integration of the grafted cells in the host brain. Tau‐Green fluorescent protein (tau‐GFP) is a powerful viable lineage tracer, allowing visualization of cell bodies, dendrites, and axons in exquisite detail. Here, we report the first rhesus monkey ESC line that ubiquitously and stably expresses tau‐GFP. First, we derived a new line of rhesus monkey ESC (LYON‐ES1) that show marker expression and cell cycle characteristics typical of primate ESCs. LYON‐ES1 cells are pluripotent, giving rise to derivatives of the three germ layers in vitro and in vivo through teratoma formation. They retain all their undifferentiated characteristics and a normal karyotype after prolonged culture. Using lentiviral infection, we then generated a monkey ESC line stably expressing tau‐GFP that retains all the characteristics of the parental wild‐type line and is clonogenic. We show that neural precursors derived from the tau‐GFP ESC line are multipotent and that their fate can be precisely mapped in vivo after grafting in the adult rat brain.

Effects of a continuous low-dose clonidine epidural regimen on pain, satisfaction and adverse events during labour: a randomized, double-blind, placebo-controlled trial.

Background and objective Epidural clonidine has been proposed as an adjunct for anaesthetic mixtures during labour. Administered as a bolus, clonidine may have side effects such as sedation and hypotension; its continuous infusion could be attractive in this respect. We, therefore, conducted a randomized, double-blind trial using patient-controlled epidural analgesia with a background infusion using a low dose of clonidine during labour. Methods A total of 128 healthy parturients in active labour received a patient-controlled epidural analgesia solution of 0.0625% levobupivacaine and sufentanil 0.25 μg ml−1 with or without clonidine 2 μg ml−1. Ninety-five parturients were analysed. The pain score over time was evaluated as well as drug volume utilization; supplementation bolus and side effects were recorded. The primary endpoint was maternal satisfaction [ClinicalTrials.gov Identifier (NCT00437996)]. Results Three patients in the control group failed to achieve satisfactory epidural analgesia owing to a technical issue. Although the primary endpoint was not statistically significant, analgesia was more pronounced and obtained earlier in the clonidine group. The area under the curve for the visual analogue pain score was significantly lower in the clonidine group. In this group, hourly doses of levobupivacaine and sufentanil were reduced (13.9 ± 4.3 vs. 16.3 ml ± 4.0; P = 0.005) as well as rescue supplementation and pruritus incidence (18 vs. 46%; P = 0.004). Maternal blood pressure was significantly lower, over time, in the clonidine group but remained within the normal range. Sedation was similar in both groups (4.3 vs. 2.0%; not significant). Conclusion The addition of clonidine to epidural levobupivacaine and sufentanil for patient-controlled epidural analgesia in labour improved analgesia, reduced the supplementation rate and reduced pruritus without improvement in maternal satisfaction. Blood pressure was significantly lower in the clonidine group over time but without clinical consequence.

La mise en œuvre des codes « couleur » réduit le délai décision-naissance des césariennes urgentes

OBJECTIVES To assess the efficiency of a new tool designed to shorten the decision-to-delivery interval (DDI) for emergency C-sections (CS). MATERIALS AND METHODS DDI comparisons during three 6-month periods in a third level maternity. In stage A we evaluated the spontaneous DDI, in stage B the DDI was measured after the introduction of a color-code communication tool related to the degree of urgency for CS (amber code indicated urgent CS with an ideal DDI of 30 min and red code for very urgent CS with an ideal DDI of 15 min). In stage C we assessed the impact of the color-codes related protocols implementation. RESULTS Two hundred and fifty-three C-sections were included (211 urgent CS and 42 very urgent CS). Mean DDI decreased significantly from 42 min to 24 min between period A and period C for amber codes (corresponding to 43.2% and 82.1% of the objectives respectively) and from 24.9 min to 10.7 min for red codes (20% et 83.3% of the objectives). CONCLUSION This study suggests that color-codes and their related application protocols significantly shorten the DDI during the evaluation periods.