Fetal Trachea Research Articles

Continuous Telemetric In Utero Tracheal Pressure Measurements in Fetal Lambs.

Normal in utero lung development and growth rely upon the expansion of airspaces and the controlled efflux of lung liquid into the amniotic space. Infants with congenital diaphragmatic hernia (CDH) also have lung hypoplasia due to occupation of the chest cavity by the stomach and bowel and, in the most severe cases, the liver. Balloon tracheal occlusion reduces the severity of lung hypoplasia in fetuses with CDH but increases the risk of premature birth. Understanding the optimal occlusion pressure and duration required to improve lung hypoplasia with tracheal occlusion is essential to improving in utero corrective treatments for CDH. The study reports a new method for continuous measurement of the intratracheal and amniotic pressures in an unoccluded and occluded fetal lamb surgical model of CDH. Time-pregnant Merino ewes underwent two recovery hysterotomies: the first at ~80 days of gestation to create the CDH, and the second at ~101 days of gestation to occlude the fetal trachea and implant an intratracheal and amniotic pressure measurement device. Lambs were delivered at ~142 days, and the pressure measurement device was removed and cleaned. The data were downloaded and filtered using a 6 h window. Transrespiratory pressure was calculated.

A Tracheal Aspirate-derived Airway Basal Cell Model Reveals a Proinflammatory Epithelial Defect in Congenital Diaphragmatic Hernia.

Rationale: Congenital diaphragmatic hernia (CDH) is characterized by incomplete closure of the diaphragm and lung hypoplasia. The pathophysiology of lung defects in CDH is poorly understood. Objectives: To establish a translational model of human airway epithelium in CDH for pathogenic investigation and therapeutic testing. Methods: We developed a robust methodology of epithelial progenitor derivation from tracheal aspirates of newborns. Basal stem cells (BSCs) from patients with CDH and preterm and term non-CDH control subjects were derived and analyzed by bulk RNA sequencing, assay for transposase accessible chromatin with sequencing, and air-liquid interface differentiation. Lung sections from fetal human CDH samples and the nitrofen rat model of CDH were subjected to histological assessment of epithelial defects. Therapeutics to restore epithelial differentiation were evaluated in human epithelial cell culture and the nitrofen rat model of CDH. Measurements and Main Results: Transcriptomic and epigenetic profiling of CDH and control BSCs reveals a proinflammatory signature that is manifested by hyperactive nuclear factor kappa B and independent of severity and hernia size. In addition, CDH BSCs exhibit defective epithelial differentiation invitro that recapitulates epithelial phenotypes found in fetal human CDH lung samples and fetal tracheas of the nitrofen rat model of CDH. Furthermore, blockade of nuclear factor kappa B hyperactivity normalizes epithelial differentiation phenotypes of human CDH BSCs invitro and in nitrofen rat tracheas invivo. Conclusions: Our findings have identified an underlying proinflammatory signature and BSC differentiation defects as a potential therapeutic target for airway epithelial defects in CDH.

Evaluation of the trachea in fetuses with double aortic arch using prenatal ultrasound: a retrospective cohort study

Double aortic arch is the most common form of complete vascular ring. The trachea and/or esophagus could be compressed by the complete vascular ring, which may lead to early respiratory and/or esophageal symptoms in children with double aortic arch. Accurate prenatal assessment of tracheal compression could provide relevant information for perinatal clinical management of double aortic arch and emergency treatment of infants with double aortic arch. The fetal trachea is filled with amniotic fluid and can be clearly visualized with prenatal ultrasound. Previous studies reported the use of prenatal ultrasound to measure the tracheal internal diameters in normal fetuses and showed a linear correlation between the fetal tracheal internal diameters and gestational age. However, to the best of our knowledge, few studies have quantitatively evaluated tracheal compression in fetuses with double aortic arch using ultrasound. This study aimed to evaluate the tracheal compression caused by the vascular ring in fetuses with double aortic arch using prenatal ultrasound and to analyze the relationship between tracheal compression and postnatal clinical symptoms. The data of fetuses with double aortic arch diagnosed with prenatal ultrasound at 2 institutions from January 2011 to April 2021 were retrospectively analyzed. Singleton pregnancies with normal fetuses as the control group were prospectively recruited. The tracheal compression-evaluated by comparing the tracheal internal diameter z scores against the gestational age-was assessed in fetuses with double aortic arch and in normal fetuses. The live-born infants with double aortic arch were divided into symptomatic and asymptomatic groups for the comparison of z scores. The receiver operating characteristic curve for the tracheal internal diameter z score cutoffs and prediction of symptomatic infants with double aortic arch was plotted. Intraobserver and interobserver agreements were investigated. A total of 26 fetuses with double aortic arch were diagnosed, and 14 fetuses (53.8%) with double aortic arch were delivered alive. Among the 14 live-born infants, 7 (50.0%) were symptomatic, whereas 7 (50.0%) were asymptomatic. The tracheal internal diameter z scores were significantly lower in the double aortic arch group than in the normal groups (-0.62±1.36 vs 0.00±0.78; P<.001). The tracheal internal diameter z scores were significantly lower in the symptomatic group than in the asymptomatic group (-1.42±0.92 vs -0.49±0.96; P=.018). The area under the curve was 0.878 (95% confidence interval, 0.689-1.000). Using a tracheal internal diameter z scores cutoff of -1.21, the sensitivity was 71%, and the specificity was close to 100%. The intraclass correlation coefficients of interobserver and intraobserver agreements were 0.987 (95% confidence interval, 0.980-0.992) and 0.975 (95% confidence interval, 0.955-0.987), respectively. The clinical symptoms in infants with double aortic arch were associated with prenatal tracheal compression, which can be prenatally evaluated using ultrasound. If fetuses are diagnosed with double aortic arch, prenatal surveillance of the tracheal internal diameters and comparison with z score reference ranges could provide pertinent information that would aid perinatal clinical management.

TITF1 Screening in Human Congenital Diaphragmatic Hernia (CDH).

TITF1 (Thyroid Transcription Factor-1) is a homeodomain-containing transcription factor. Previous studies showed that Titf1 null mice are characterized by failure of tracheo-oesophageal separation and impaired lung morphogenesis resulting in Pulmonary Hypoplasia (PH). In this study, we aim to evaluate the role of TITF1 in the pathogenesis of congenital diaphragmatic hernia (CDH) in humans. We investigated TITF1 expression in human trachea and lungs and performed direct mutation analysis in a CDH population. We studied 13 human fetuses at 14 to 24 weeks of gestation. Five μm sections were fixed in paraformaldehyde and incubated with anti-TITF1 primary antibody. Positive staining was visualized by biotinylated secondary antibody. We also performed TITF1 screening on genomic DNA extracted from peripheral blood of 16 patients affected by CDH and different degrees of PH, searching for mutations, insertions, and/or deletions, by sequencing the exonic regions of the gene. Histochemical studies showed positive brown staining of fetal follicular thyroid epithelium, normal fetal trachea, and normal fetal lung bronchial epithelium. Fetal esophageal wall was immunohistochemically negative. Molecular genetic analysis showed complete identity between the sequences obtained and the Wild Type (WT) form of the gene in all cases. No mutation, insertion and/or deletion was detected. Although TITF1 is expressed in the human fetal lung and has been considered to have a role in the pathogenesis of PH in CDH, the results of our study do not support the hypothesis that TITF1 mutations play a key role in the etiopathogenesis of CDH.

Randomized Trial of Fetal Surgery for Severe Left Diaphragmatic Hernia

(Engl J Med. 2021;385:107–118) Congenital diaphragmatic hernia (CDH) can lead to serious lifelong complications, but may be treated if diagnosed before birth. In fetoscopic endoluminal tracheal occlusion (FETO), a balloon is inserted into the fetal trachea in utero to stimulate fetal lung growth and is removed prior to birth. One study of 210 fetuses undergoing FETO for severe lung hypoplasia due to isolated CDH found a nearly doubled survival rate among fetuses who received FETO compared with controls (49% vs. 24%). This current study, Tracheal Occlusion to Accelerate Lung Growth (TOTAL), aimed to test whether fetuses with surgical treatment of severe left CDH had superior survival rates compared with fetuses who received expectant prenatal care.

An Implantable Electronic Device for Monitoring Fetal Lung Pressure in a Lamb Model of Congenital Diaphragmatic Hernia

Tracheal occlusion (TO) is a fetal treatment option for congenital diaphragmatic hernia. It aims to reduce lung hypoplasia and hypertension by occluding the trachea with a subsequent increase in pulmonary pressure. However, how TO affects lung pressure is unknown, and no devices are available for its measurement. We developed an implantable device for real-time wireless monitoring and long-term logging of lung pressure in ovine fetuses. The device comprised a microcontroller unit, two pressure sensors, an inertial measurement unit (IMU), a radio frequency unit, and an antenna. <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">In vitro</i> and <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">in vivo</i> tests were performed to evaluate device performances. A diaphragmatic hernia was created in ovine fetuses via hysterotomy. Nine devices were implanted at 100 days of gestation and remained until delivery (~142 d). One pressure sensor measured pressure in the amniotic sac and the other in the fetal trachea. <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">In vitro</i> pressure accuracy was median (interquartile range) 0.31% (0.1%). <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">In vitro</i> device lifetime was ~60 days. The <italic xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">in vivo</i> device recording time was 26.0 (3.9) days and it was well tolerated by the fetus and the ewe. Data telemetry operated throughout the device lifetime. At delivery, amniotic fluid, device encapsulation, and lamb skin did not present evidence of damages. Pressure data in four lambs were analyzed. Fast pressure changes due to the spontaneous fetal activity and long-term mean pressure changes resulting from TO were detected. The IMU allowed for the assessment of changes in fetal position. This device allows long-term monitoring of mean lung pressure, spontaneous breathing activity, and movements in physiological and pathological ovine fetal models.

Optimization of cultivation parameters for bovine respiratory syncytial virus strain Vologda/2019

There are currently many controversial issues in the study of bovine respiratory syncytial infection. In this regard, it is relevant to study the biological properties of the virus, optimize the methods of its cultivation and select the most technologically advanced methods of designing diagnostic and prevention tools for this disease. The aim of this work was to select sensitive cell systems and to optimize the cultivation parameters in selected cell cultures. The Vologda/2019 strain of the bovine respiratory syncytial infection virus isolated from biological material obtained from a calf with respiratory symptoms was used in the experiment. The strain was adapted to the continuous cell culture derived from bovine turbinate tissue (BT) and deposited in the State collection of microorganism strains at FGBI “ARRIAH”. It was established that the continuous cell lines of fetal bovine trachea (FBT) and calf kidney (RBT) are the most sensitive cell systems for the reproduction of the bovine respiratory syncytial virus strain Vologda/2019, the maximum accumulation of the virus was observed in these cell cultures. The cytopathic activity of the virus in the FBT cell culture ranged from 4.78 ± 0.18 to 5.50 ± 0.16 lg TCID 50 /cm 3 , and in the RBT cell culture – from 4.00 ± 0.23 to 4.75 ± 0.20 lg TCID 50 /cm 3 at days 4–5 of cultivation. It was determined that in case of multiplicity of inoculation of FBT and RBT cell cultures with the virus at 0.1 lg TCD 50 /cell and the use of 2% glutamine in the maintenance nutrient medium, as well as 2% horse or cattle blood serum, it is possible to obtain virus material with high cytopathic activity.

CULTURAL PROPERTIES OF BOVINE RESPIRATORY SYNCYTIAL VIRUS STRAIN AB1908

Cattle respiratory diseases are some of the most spread pathologies that can cause economic damage, resulting from fi nancial losses and costs of treatment and diagnostics. One of the major factors contributing to respiratory pathology development is bovine respiratory syncytial infection. The analysis of serological testing, performed by the FGBI “ARRIAH” Reference Laboratory for Cattle Diseases in 2017–2018, showed that respiratory syncytial virus seroprevalence in animals of dairy farms is 60%. Herewith, it was noted that the most susceptible animals to this infection are calves under one year of age. The eff ectiveness of bovine respiratory syncytial infection control measures depends on timely diagnosis; that is why reliable and accurate diagnostic tools are needed, including optimal techniques of virus isolation from pathological material. For successful virus isolation from clinical samples, it is necessary to adhere strictly to optimal parameters of this agent cultivation. This paper presents data on study of bovine respiratory syncytial virus strain AB 1908 cultural properties. The tests performed showed that a continuous bovine turbinate (BT) cell line, continuous bovine fetal trachea (FBT) cell line and continuous bovine calf kidney (RBT) cell line are sensitive for cultivation of this agent and can be used to prepare viral suspension, needed for further research. Virus titre in BT cell culture was 4.33 ± 0.16 – 4.66 ± 0.12 lg TCID50/ cm3, in RBT cell culture – 4.33 ± 0.33 – 4.7 ± 0.36 lg TCID50/cm3 and in FBT cell culture – 4.13 ± 0.20 – 4.78 ± 0.17 lg TCID50/cm3. The following virus cultivation optimal parameters were also determined during this study: the age of the culture for virus inoculation should be 1–2 days and multiplicity of inoculation should be 0.1 TCID50/cell.

Feasibility of quantitative measurement of fetal trachea and bilateral bronchial angles by ultrasound to diagnose heterotaxy syndrome in middle and late pregnancy

Objective To analyze the feasibility of prenatal ultrasound quantitative measurement of the angles between trachea and bilateral bronchi as a new diagnostic tool for fetuses with heterotaxy syndrome. Methods The angles between trachea and bilateral bronchi were measured at 18-34+6 gestational weeks for 200 fetuses with normal ultrasound findings (normal group) and 41 fetuses with heterotaxy syndrome [35 right atrial isomerisms (right group) and six left atrial isomerisms (left group)] diagnosed by ultrasound and confirmed after abortion or induction (case group) in Chongqing Health Center for Women and Children from October 2015 to December 2018. Scatter plots of left/right bronchus angle (α/β) ratios were drawn. Differences in bilateral bronchus angles between the three groups were statistically analyzed using one-way analysis of variance and paired t-test. Results In the normal group, scatter plots showed the α/β ratios were relatively constant at 0.8-1.0 with larger β than α (t=-33.14, P 0.05] and presented as bilateral right bronchial angle; in the left group, the β value decreased and was close to the α value [(147.38±3.16)° vs (148.82±5.56)°, t=-1.18, P>0.05] and presented as bilateral left bronchial angle. There were significant differences in the α and β values among the normal group, the right and left groups (all P<0.05), and the bronchial angles of the right group were both greater than those of the left group (all P<0.05). Conclusions Prenatal ultrasound measurement of the angles between trachea and bilateral bronchi can be used as an indirect indicator for the diagnosis of heterotaxy syndrome. Key words: Heterotaxy syndrome; Ultrasonography, prenatal; Trachea; Bronchi

Z scores of the fetal trachea and bronchial dimension.

To develop Z scores for the trachea and main bronchi in normal fetuses. This was a prospective cross-sectional study in 823 normal singleton fetuses. The tracheal diameter immediately proximal to the bifurcation and the left and right main bronchial diameters were measured from their inner to inner edge in the coronal view. Z scores were created for the trachea and main bronchial diameters using gestational age (GA), femur length (FL), and biparietal diameter (BPD) as independent variables. Between gestational weeks 20 and 40, the inner diameters of the trachea, left principal bronchi (LPB), and right principal bronchi (RPB) increased from 1.8 to 4.7mm, 0.8 to 2.2mm, and 0.9 to 2.3mm, respectively. A simple linear regression equation was fitted to model the mean of each diameter. There was however significant heteroscedasticity of the standard deviation (SD) with increasing GA, FL, or BPD. Eventually, the following formula was used to calculate Z scores for the diameters: [measured value-equation for mean]/equation forSD. We have developed Z scores for the fetal trachea and main bronchi by applying standard statistical methods. These Z scores may be useful to evaluate the early development of the respiratory system.

A novel translational model for fetoscopic intratracheal delivery of nanoparticles in piglets

The aim of this study was to assess the feasibility of fetal tracheal injection in the late-gestational pig to target the airways. Following laparotomy and hysterotomy, fetoscopy was performed in pregnant sows to access the fetal trachea. Two volumes of fluospheres were injected (1 and 3 mL). Fluosphere distribution to the different lung lobes was investigated by microscopy. Possible fetal airway injury, caused by the surgical procedure or intratracheal injection, was investigated. Lung morphology and fetal lung volumes were calculated by micro computed tomography (μCT). Intratracheal administration was successfully performed in 20/21 fetuses. Analysis by confocal microscopy demonstrated that 3 mL, and not 1 mL, most efficiently targeted all lung lobes. On high-resolution μCT, total airway volume was estimated at 2.9 mL; strengthening that 3 mL is appropriate to target all lung lobes. No procedural damage was evidenced in the lungs or trachea. Intratracheal injection of nanoparticles is feasible in the pregnant pig and does not cause procedural lung damage. Using an injection volume of 3 mL, all lung lobes were efficiently targeted. This nanoparticle delivery model to fetal airways opens perspectives for therapeutic interventions. © 2016 John Wiley & Sons, Ltd.

Patterned, tubular scaffolds mimic longitudinal and radial mechanics of the neonatal trachea

Tracheal damage, abnormality or absence can result from the growth of tumors or from Congenital High Airway Obstruction Syndrome. No optimal or routine treatment has been established for tracheal repair, despite numerous attempts with natural and artificial prostheses. The fetal trachea is comprised of cartilaginous rings connected by an elastomeric tissue. In an effort to design an engineered trachea replacement, we have synthesized 2-hydroxyethyl methacrylate hydrogels with moduli of 67±3.1kPa (soft) and 13.0±1.8MPa (hard). Given the criteria for longitudinal extensibility and lateral rigidity applied during respiration, we evaluated a series of patterned hydrogels with different sizes of hard and soft segments to mimic fetal tracheas. A 1:2 ratio of soft:hard segments resulted in a construct capable of 11.0±1% extension within the elastic range. Tubular constructs with this ratio required similar load/length for cyclic compression as ovine trachea samples. Achieving biomimetic mechanical properties in a trachea replacement may be essential for achieving normal respiration in recipient patients. Statement of SignificanceFetal abnormalities or tumors can result in tracheal absence or damage. Despite numerous attempts with natural and artificial replacements, there is still no routine treatment for tracheal repair. The literature recognizes the importance of tracheal lateral rigidity and longitudinal extensibility for normal respiration. Achieving closely matched mechanical properties may provide proper function and help decrease implant fibrosis and subsequent occlusion. In this study, we evaluated the mechanics of a series of patterned, tubular hydrogels with different ratios of hard and soft segments to mimic alternating cartilage and ligament sections in fetal tracheas. We compared our results to that of sheep trachea. This is the first report to assess both radial rigidity and longitudinal extensibility in an engineered trachea construct.

Observation and measurement of trachea and bronchus in normal fetuses by prenatal ultrasonography

Objective To explore the methodology of observing trachea and bronchus in the normal fetuses by prenatal ultrasound and to summarize its ultrasonographic features, then to measure relevant parameters and to establish their normal reference value range. Methods Nine hundred and seventy normal fetuses from 20 to 39+ 6 weeks of gestation were observed by ultrasound at the coronal section of trachea and bronchus.The subjects were classified into five groups according to gestational age to calculate demonstration rate: group A, 20-23 weeks; group B, 24-27 weeks; group C, 28-31 weeks; group D, 32-35 weeks, group E, 36-39 weeks. The fetuses whose trachea and bronchus were clearly shown were grouped by each gestational week to measure the diameters of trachea and left/right bronchus and the angle between them. Results There were 565 fetuses whose trachea and bronchus could be demonstrated clearly (58.2%). The numbers and demonstration rates of each group were following: group A 184(53.5%), group B 88(79.3%), group C 109(68.1%), group D 97(48.3%) and group E 87(55.4%). The results of both group B and C were significantly higher than others (P 0.05). The angle between trachea and left bronchus was smaller than that of trachea and right bronchus(P<0.05). Conclusions It is feasible to observe fetal trachea and bronchus at the coronal view of thorax. The appropriate gestational age for ultrasonic observation is 24-31+ 6 weeks. The diameter of trachea and bronchus and the angle between them can be potential valuable for assessing the growth of trachea and bronchus prenatally. The fact that the angle between trachea and left bronchus was smaller than that of trachea and right bronchus may provide a new clue for judging visceral position. Key words: Ultrasonography, prenatal; Fetus; Trachea; Bronchi

Prenatal Observation of Fetal Trachea and Bilateral Bronchi using Ultrasonography

ABSTRACT The fetal airway tract can be observed using the planes of view in cardiac screening. In three-vessel-tracheal view, the crosssection of the trachea is identified adjacent to the right-posterior side of the middle transverse aorta. In three-vessel view, the cross-section of the right bronchus is observed adjacent to the right side of the ascending aorta. In aortic and ductal arch views, the short axis of the left bronchus is confirmed inward of the arch. In oblique three-vessel view, the right bronchus is seen on top of the ascending aorta coursing toward the azygos vein. To exclude the vessels, absence of blood flow signals in the trachea and bronchi should be reconfirmed by using pulsed and color Doppler imagings. In the whole ‘Y-shaped’ image, the right main bronchus depicts the more linear extension from the trachea than the left bronchus and the first lobar bronchus that branches near the carina to the right upper lobe. Segmental bronchi also can be depicted in both bilateral bronchi. Several new modalities, including three-dimensional (3D) ultrasound, simultaneous visualization of the fetal trachea and esophagus, and fluid flow waveforms in the fetal airway tract, makes feasible detailed prenatal assessment, prediction of neonatal respiratory condition, and most especially identification of anatomical deformity and pathologic entities. How to cite this article Aoki S. Prenatal Observation of Fetal Trachea and Bilateral Bronchi using Ultrasonography. Donald School J Ultrasound Obstet Gynecol 2014;8(4):353-361.

Difficulties in the Hemodynamic Assessment of the Newborn with Prenatally Treated Congenital Diaphragmatic Hernia

Congenital diaphragmatic hernia (CDH) is a malformation with a diverse clinical picture. Its severity can be assessed prenatally using the lung area to head circumference ratio (LHR). LHR lower than 1 as associated with high mortality is an indication for prenatal treatment involving occlusion of fetal trachea. Before such treatment other congenital malformations must be excluded. Assessment of circulatory system in these patients is difficult. Heart is compressed by viscera in the thorax that decreases blood return to the left atrium, and structures of the left heart may be smaller than in healthy fetuses. The echocardiographic picture may be similar to some congenital defects of the left heart. Differentiation between structural and functional cardiac abnormalities is difficult but essential for diagnostic and therapeutic decisions in the most severe type of CDH. The authors present problems with interpretation of hemodynamic disturbances in a neonate with prenatally diagnosed and treated severe CDH.

Progress of ultrasonic application in fetal trachea and bronchus

临床上胎儿气管与支气管病变较少见,但发病后大多病情凶险,胎儿存活率很低[1-2].目前产前常规超声检查规范尚未涉及气管与支气管,对正常胎儿这一区域的超声形态了解较少,偶尔发现病变多是依赖间接超声征象,故病变检出率低,检出时间较晚,存在一定的漏诊率.现对胎儿气管与支气管的解剖特征、先天性病变及超声研究进展进行综述。

Pulmonary alveolar and vascular morphometry after gel plug occlusion of the trachea in a fetal rabbit model of CDH

PurposeTracheal occlusion (TO) induces lung growth in congenital diaphragmatic hernia (CDH) but is also associated with drawbacks. We devised a temporary gel plug that induced lung growth when placed in the fetal trachea. This study evaluates the effects of temporary versus permanent TO on histologic radial alveolar count (RAC) and vascular morphometrics. MethodsExperimental CDH was created surgically in 64 New Zealand White rabbit fetuses on gestational day (GD) 24. On GD 27, these fetuses were randomized to intratracheal instillation of a fibrin gel plug (GP), tracheal suture ligation (SL), intratracheal instillation of normal saline (NS), or sham amniotomy (SH). Non-manipulated fetuses served as controls (NM). Histologic lung sections were assessed blindly for RAC and relative arterial adventitial thickness (%AT) as a variable for vascular remodelling. Results were statistically compared. ResultsRAC was significantly lower in the ipsilateral lung of SH fetuses than in the contralateral lung (p = 0.011). Mean RAC was higher after SL (p < 0.001) and GP (p = 0.03) compared to SH. Furthermore, %AT was higher in GP (50 ± 28, p < 0.001) and SL (45 ±2 6, p = 0.003) fetuses than in controls (36 ± 19). ConclusionTemporary and permanent TO leads to increased RAC; this effect was more pronounced with permanent TO. Both interventions were associated with an increased %AT. These findings may explain the adverse clinical effects of TO, despite causing accelerated lung growth.

Human Amnion Epithelial Cells Reduce Fetal Brain Injury in Response to Intrauterine Inflammation

Intrauterine infection, such as occurs in chorioamnionitis, is a principal cause of preterm birth and is a strong risk factor for neurological morbidity and cerebral palsy. This study aims to examine whether human amnion epithelial cells (hAECs) can be used as a potential therapeutic agent to reduce brain injury induced by intra-amniotic administration of lipopolysaccharide (LPS) in preterm fetal sheep. Pregnant ewes underwent surgery at approximately 110 days of gestation (term is approx. 147 days) for implantation of catheters into the amniotic cavity, fetal trachea, carotid artery and jugular vein. LPS was administered at 117 days; hAECs were labeled with carboxyfluorescein succinimidyl ester and administered at 0, 6 and 12 h, relative to LPS administration, into the fetal jugular vein, trachea or both. Control fetuses received an equivalent volume of saline. Brains were collected 7 days later for histological assessment of brain injury. Microglia (Iba-1-positive cells) were present in the brain of all fetuses and were significantly increased in the cortex, subcortical and periventricular white matter in fetuses that received LPS, indicative of inflammation. Inflammation was reduced in fetuses that received hAECs. In LPS fetuses, the number of TUNEL-positive cells was significantly elevated in the cortex, periventricular white matter, subcortical white matter and hippocampus compared with controls, and reduced in fetuses that received hAECs in the cortex and periventricular white matter. Within the fetal brains studied there was a significant positive correlation between the number of Iba-1-immunoreactive cells and the number of TUNEL-positive cells (R² = 0.19, p < 0.001). The administration of hAECs protects the developing brain when administered concurrently with the initiation of intrauterine inflammation.

A Novel Surgical Approach for Intratracheal Administration of Bioactive Agents in a Fetal Mouse Model

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and acquired diseases. Apart from congenital or inherited abnormalities with the requirement for long-term expression of the delivered gene, several non-inherited perinatal conditions, where short-term gene expression or pharmacological intervention is sufficient to achieve therapeutic effects, are considered as potential future indications for this kind of approach. Candidate diseases for the application of short-term prenatal therapy could be the transient neonatal deficiency of surfactant protein B causing neonatal respiratory distress syndrome1,2 or hyperoxic injuries of the neonatal lung3. Candidate diseases for permanent therapeutic correction are Cystic Fibrosis (CF)4, genetic variants of surfactant deficiencies5 and α1-antitrypsin deficiency6. Generally, an important advantage of prenatal gene therapy is the ability to start therapeutic intervention early in development, at or even prior to clinical manifestations in the patient, thus preventing irreparable damage to the individual. In addition, fetal organs have an increased cell proliferation rate as compared to adult organs, which could allow a more efficient gene or stem cell transfer into the fetus. Furthermore, in utero gene delivery is performed when the individual's immune system is not completely mature. Therefore, transplantation of heterologous cells or supplementation of a non-functional or absent protein with a correct version should not cause immune sensitization to the cell, vector or transgene product, which has recently been proven to be the case with both cellular and genetic therapies7. In the present study, we investigated the potential to directly target the fetal trachea in a mouse model. This procedure is in use in larger animal models such as rabbits and sheep8, and even in a clinical setting9, but has to date not been performed before in a mouse model. When studying the potential of fetal gene therapy for genetic diseases such as CF, the mouse model is very useful as a first proof-of-concept because of the wide availability of different transgenic mouse strains, the well documented embryogenesis and fetal development, less stringent ethical regulations, short gestation and the large litter size. Different access routes have been described to target the fetal rodent lung, including intra-amniotic injection10-12, (ultrasound-guided) intrapulmonary injection13,14 and intravenous administration into the yolk sac vessels15,16 or umbilical vein17. Our novel surgical procedure enables researchers to inject the agent of choice directly into the fetal mouse trachea which allows for a more efficient delivery to the airways than existing techniques18.

A Novel Surgical Approach for Intratracheal Administration of Bioactive Agents in a Fetal Mouse Model

Prenatal pulmonary delivery of cells, genes or pharmacologic agents could provide the basis for new therapeutic strategies for a variety of genetic and acquired diseases. Apart from congenital or inherited abnormalities with the requirement for long-term expression of the delivered gene, several non-inherited perinatal conditions, where short-term gene expression or pharmacological intervention is sufficient to achieve therapeutic effects, are considered as potential future indications for this kind of approach. Candidate diseases for the application of short-term prenatal therapy could be the transient neonatal deficiency of surfactant protein B causing neonatal respiratory distress syndrome(1,2) or hyperoxic injuries of the neonatal lung(3). Candidate diseases for permanent therapeutic correction are Cystic Fibrosis (CF)(4), genetic variants of surfactant deficiencies(5) and α1-antitrypsin deficiency(6). Generally, an important advantage of prenatal gene therapy is the ability to start therapeutic intervention early in development, at or even prior to clinical manifestations in the patient, thus preventing irreparable damage to the individual. In addition, fetal organs have an increased cell proliferation rate as compared to adult organs, which could allow a more efficient gene or stem cell transfer into the fetus. Furthermore, in utero gene delivery is performed when the individual's immune system is not completely mature. Therefore, transplantation of heterologous cells or supplementation of a non-functional or absent protein with a correct version should not cause immune sensitization to the cell, vector or transgene product, which has recently been proven to be the case with both cellular and genetic therapies(7). In the present study, we investigated the potential to directly target the fetal trachea in a mouse model. This procedure is in use in larger animal models such as rabbits and sheep(8), and even in a clinical setting(9), but has to date not been performed before in a mouse model. When studying the potential of fetal gene therapy for genetic diseases such as CF, the mouse model is very useful as a first proof-of-concept because of the wide availability of different transgenic mouse strains, the well documented embryogenesis and fetal development, less stringent ethical regulations, short gestation and the large litter size. Different access routes have been described to target the fetal rodent lung, including intra-amniotic injection(10-12), (ultrasound-guided) intrapulmonary injection(13,14) and intravenous administration into the yolk sac vessels(15,16) or umbilical vein(17). Our novel surgical procedure enables researchers to inject the agent of choice directly into the fetal mouse trachea which allows for a more efficient delivery to the airways than existing techniques(18).