Fetal Limb Research Articles

Wnt/β-Catenin Promotes the Osteoblastic Potential of BMP9 Through Down-Regulating Cyp26b1 in Mesenchymal Stem Cells.

All-trans retinoic acid (ATRA) promotes the osteogenic differentiation induced by bone morphogenetic protein 9 (BMP9), but the intrinsic relationship between BMP9 and ATRA keeps unknown. Herein, we investigated the effect of Cyp26b1, a critical enzyme of ATRA degradation, on the BMP9-induced osteogenic differentiation in mesenchymal stem cells (MSCs), and unveiled possible mechanism through which BMP9 regulates the expression of Cyp26b1. ATRA content was detected with ELISA and HPLC-MS/MS. PCR, Western blot, and histochemical staining were used to assay the osteogenic markers. Fetal limbs culture, cranial defect repair model, and micro-computed tomographic were used to evaluate the quality of bone formation. IP and ChIP assay were used to explore possible mechanism. We found that the protein level of Cyp26b1 was increased with age, whereas the ATRA content decreased. The osteogenic markers induced by BMP9 were increased by inhibiting or silencing Cyp26b1 but reduced by exogenous Cyp26b1. The BMP9-induced bone formation was enhanced by inhibiting Cyp26b1. The cranial defect repair was promoted by BMP9, which was strengthened by silencing Cyp26b1 and reduced by exogenous Cyp26b1. Mechanically, Cyp26b1 was reduced by BMP9, which was enhanced by activating Wnt/β-catenin, and reduced by inhibiting this pathway. β-catenin interacts with Smad1/5/9, and both were recruited at the promoter of Cyp26b1. Our findings suggested the BMP9-induced osteoblastic differentiation was mediated by activating retinoic acid signalling, viadown-regulating Cyp26b1. Meanwhile, Cyp26b1 may be a novel potential therapeutic target for the treatment of bone-related diseases or accelerating bone-tissue engineering.

Fetal Hands: A Comprehensive Review of Prenatal Assessment and Diagnosis Over the Past 40 Years.

Fetal skeletal dysplasias involving limbs and hands are rare congenital malformations. Prenatal two-dimensional ultrasound diagnosis of fetal limb defects has a sensitivity of about 30%; however, an increased detection rate may be obtained using three-dimensional (3-D) ultrasound in the rendering mode. 3-D ultrasound may be used as a complementary method providing additional information. Currently, magnetic resonance imaging (MRI), with the emergence of ultrafast imaging techniques and new sequences, allows for better diagnosis of several fetal skeletal dysplasias such as limb reduction defects and neuromuscular disorders. 3-D volumetric images from ultrasound or MRI scan data allow 3-D ultrasound reconstructions of virtual/physical models, and virtual reality can help researchers to improve our understanding of both normal and abnormal fetal limb/hand anatomy. In this article, we review the embryological development of fetal hands and their main anomalies including prenatal diagnostic methods, genetic counseling, the role of orthopedic and plastic surgery reconstruction, and new perspectives in fetal surgery.

Routine Screening for Fetal Limb Abnormalities in the First Trimester

Objective: To evaluate the routine screening for fetal limb abnormalities in the first trimester. Study Design: Prospective study Place and Duration of Study: Department of Radiology and Department of Obstetrics & Gynaecology, Khirpur Medical College, Khairpur Mir’s from 1st October 2020 to 30th September 2022. Methodology: The study enrolled all pregnant women in visiting the Gynaecology OPD for routine check-up. All the cases were referred to ultrasound diagnostic for screening of fetal limb abnormalities. The total sample size of the study was assigned as 1000 fetuses from 980 women as 10 women had twin pregnancies. Limb reduction shortcomings included transverse-limb reduction deficit as well as longitudinal-limb reduction deficit as seen through absent radius and also club hand, absent fibula phocomelia. The diagnosis of the limb shortening was based on the length of long bone if <5th percentile. Lethal skeletal dysplasia suspicion was based on narrow thorax and long bone abnormalities. Results: The mean age of the pregnant women was 27.6±5.3 year. Around 26% of the women were nulliparous. The fetal limb anomaly was presented in 0.9% of the total cases. Within these, 0.7% was those women with single fetus while 0.2% had twin pregnancy. There were 60% of the women who were identified with fetal anomaly on their first screening visit within 1st trimester transverse limb anomaly was observed in 2 case within first trimester whereas 2 cases of club foot were identified in first trimester and one in third trimester. Conclusion: The fetal limb anomaly can be feasibly screened within first trimester. It has a prevalence of 0.9% with transverse limb deficiencies and club foot to be presented in highest frequency. Keywords: Fetal limb abnormalities, First trimester, Prevalence

Interleukin 6 promotes BMP9-induced osteoblastic differentiation through Stat3/mTORC1 in mouse embryonic fibroblasts.

Interleukin 6 (IL-6) plays a dual role in regulating bone metabolism, although the concrete mechanism is unclear. Bone morphogenetic protein 9 (BMP9) is one of the most potent osteogenic inducers, and a promising alternative for bone tissue engineering. The relationship between IL-6 and BMP9 in osteogenic differentiation remains to be elucidated, and the osteoblastic potential of BMP9 needs to be enhanced to overcome certain shortcomings of BMP9. In this study, we used real-time PCR, western blot, immunofluorescent stain, fetal limb culture and cranial defects repair model to explore the IL-6 role in BMP9-induced osteogenic differentiation in mouse embryonic fibroblasts (MEFs). We found that the rat serum level of IL-6 was increased in the dexamethasone-induced osteoporosis model, and IL-6 expression was detectable in several progenitor cells and MEFs. BMP9 upregulated IL-6 in MEFs, and the BMP9-induced osteoblastic markers were elevated by IL-6, but reduced by IL-6 knockdown. BMP9 and/or IL-6 both activated mTOR, and the IL-6 effect on BMP9-induced osteoblastic markers and bone formation were reduced greatly by mTOR inhibition. Raptor was up-regulated by IL-6 and/or BMP9 specifically, and the osteoblastic markers induced by IL-6 and/or BMP9 were reduced by Raptor knockdown. Meanwhile, Stat-3 was activated by IL-6 and/or BMP9, and the increase of Raptor or osteoblastic markers by IL-6 and/or BMP9 were reduced by Stat-3 inhibition. The Raptor promoter activity was regulated by p-Stat-3. Our finding suggested that IL-6 can promote the BMP9 osteoblastic potential, which may be mediated through activating Stat-3/mTORC1 pathway.

Ultrasound and fetal magnetic resonance imaging: Clinical performance in the prenatal diagnosis of orofacial clefts and mandibular abnormalities

Cleft lip, with or without cleft palate, is the most common congenital craniofacial anomaly and the second most common birth defect worldwide. Micrognathia is a rare facial malformation characterized by small, underdeveloped mandible and frequently associated with retrognathia. Second- and third-trimester prenatal ultrasound is the standard modality for screening and identification of fetal orofacial abnormalities, with a detection rate in the low-risk population ranging from 0% to 73% for all types of cleft. The prenatal ultrasonography detection can also be performed during the first trimester of pregnancy. Given the potential limitations of obstetric ultrasound for examining the fetal face, such as suboptimal fetal position, shadowing from the surrounding bones, reduce amniotic fluid around the face, interposition of fetal limbs, umbilical cord and placenta, and maternal habitus/abdominal scars, the use of adjunct imaging modalities can enhance prenatal diagnosis of craniofacial anomalies in at-risk pregnancies. Fetal magnetic resonance imaging (MRI) is a potentially useful second-line investigation for the prenatal diagnosis of orofacial malformations with a pooled sensitivity of 97%. In this review, we discuss the role of ultrasound and fetal MRI in the prenatal assessment of abnormalities of the upper lip, palate, and mandible.

The Lower Limb Movements of the Fetus in Uterus: A Narrative Review.

The fetus movements play an important role in fetal well-being. With the continuous advancement of real-time scanning machines, it is feasible to observe the fetus movement in detail. The characteristics of fetal lower limb movements in prenatal examination have not been systematically investigated. This review proposes the patterns of fetal lower limb movements, the maternal influence on fetal lower limb movements, and the application of fetal lower limb movements for the diagnosis of prenatal diseases. A systematic search of literature on the lower limb movements of the fetus in uterus was performed in the databases, namely, Web of Science and Scopus. Thirty-four publications were selected. This review demonstrates that isolated fetal lower limb movements are rare and always accompanied with the movements of other body segments. Detection of the presence of fetal leg movements seems to be of no diagnostic value for fetuses with prenatal diseases. The isolated lower limb movement was statistically significant different between fetuses of low- and high-risk pregnant women. The coordinated movements of the fetal lower limbs and other parts should be considered when analyzing fetal movements in the future study.

Cross-Sectional Observational Study of Typical in utero Fetal Movements Using Machine Learning

Early variations of fetal movements are the hallmark of a healthy developing central nervous system. However, there are no automatic methods to quantify the complex 3D motion of the developing fetus in utero. The aim of this prospective study was to use machine learning (ML) on in utero MRI to perform quantitative kinematic analysis of fetal limb movement, assessing the impact of maternal, placental, and fetal factors. In this cross-sectional, observational study, we used 76 sets of fetal (24–40 gestational weeks [GW]) blood oxygenation level-dependent (BOLD) MRI scans of 52 women (18–45 years old) during typical pregnancies. Pregnant women were scanned for 5–10 min while breathing room air (21% O₂) and for 5–10 min while breathing 100% FiO₂ in supine and/or lateral position. BOLD acquisition time was 20 min in total with effective temporal resolution approximately 3 s. To quantify upper and lower limb kinematics, we used a 3D convolutional neural network previously trained to track fetal key points (wrists, elbows, shoulders, ankles, knees, hips) on similar BOLD time series. Tracking was visually assessed, errors were manually corrected, and the absolute movement time (AMT) for each joint was calculated. To identify variables that had a significant association with AMT, we constructed a mixed-model ANOVA with interaction terms. Fetuses showed significantly longer duration of limb movements during maternal hyperoxia. We also found a significant centrifugal increase of AMT across limbs and significantly longer AMT of upper extremities <31 GW and longer AMT of lower extremities >35 GW. In conclusion, using ML we successfully quantified complex 3D fetal limb motion in utero and across gestation, showing maternal factors (hyperoxia) and fetal factors (gestational age, joint) that impact movement. Quantification of fetal motion on MRI is a potential new biomarker of fetal health and neuromuscular development.

M13, an anthraquinone compound isolated from Morinda officinalis promotes the osteogenic differentiation of MSCs by targeting Wnt/β-catenin signaling

BackgroundMorinda officinalis (MO) is a herb used in Traditional Chinese Medicine (TCM) for the treatment of osteoporosis. M13, a MO-based anthraquinone compound is known to suppress osteoclast activity. However, whether M13 promotes MSCs osteogenic differentiation and its potential mechanism remains unknown. PurposeTo examine the influence of M13 on MSCs proliferation and osteogenic differentiation and elucidate the underlying mechanism. Methods/Study designsThe effect of M13 exposure on MSCs proliferation was assessed via CCK8 assay, clone formation assay, immunofluorescence, RT-qPCR, and Western blot. The M13-mediated osteogenesis in vitro and ex vivo were evaluated via ALP and Alizarin red S staining, osteogenesis-associated gene (Runx2, Col1a1 and Opn) expression, and fetal limb explants culture. Molecular docking was employed for target signal pathway screening. The potential signaling mechanisms of M13-promoted MSCs osteogenic differentiation were analyzed by introducing XAV939 (Wnt/β-catenin signaling inhibitor). ResultsM13 induced certain obvious positive effects on MSCs proliferation and osteogenic differentiation. Treatment with M13 enhanced MSCs viability and clone numbers. Meanwhile, M13 promoted osteogenic gene expression, enhanced ALP intensity and Alizarin red S staining in MSCs. In terms of mechanism, M13 strongly interacted with the docking site of the WNT signaling complex, thereby activating the Wnt/β-catenin pathway. Furthermore, the M13-mediated osteogenic effect was partially inhibited by XAV939 both in vitro and ex vivo, which confirmed that the Wnt/β-catenin axis is a critical regulator of M13-induced osteogenic differentiation of MSCs. ConclusionOur study elucidated for the first time that M13 significantly promoted osteogenic differentiation of MSCs via stimulation of the Wnt/β-catenin pathway in vitro and ex vivo.Our findings offered new additional evidence to support the MO or M13-based therapy of osteoporosis.

Assessment of lower limb trophism in fetuses with open spina bifida using fractional thigh volume of three-dimensional ultrasound.

To assess the trophism of the lower limbs of fetuses with open spina bifida using fractional thigh volume (TVOL) of three-dimensional (3D) ultrasound. A prospective cross-sectional study was carried out with normal fetuses and with open spina bifida (myelomeningocele and rachischisis) at 26 weeks' gestation. The TVOL (delimitation of five cross-sectional areas of the middle portion of the limb) was evaluated, as well as the subjective assessment of hypotrophy and lower limb movement. Thirty-one fetuses with open spina bifida, 21 with myelomeningocele and 10 with rachischisis, and 51 normal fetuses were included. There were no significant differences in the TVOL between normal and spina bifida fetuses (p=0.623), as well as between normal fetuses, with myelomeningocele and with rachischisis (p=0.148). There was no significant difference in the TVOL of fetuses with spina bifida with or without lower limb hypotrophy (p=0.148). Fetuses with spina bifida and with lower limb movement had higher TVOL values than fetuses without lower limb movement (p=0.002). There were no significant differences in the TVOL measurement of normal and spina bifida fetuses (rachischisis and myelomeningocele). Fetuses with spina bifida without spontaneous movement of the lower limbs had lower TVOL values.

Genetic risk of osteoarthritis operates during human skeletogenesis.

Osteoarthritis (OA) is a polygenic disease of older people resulting in the breakdown of cartilage within articular joints. Although it is a leading cause of disability, there are no disease-modifying therapies. Evidence is emerging to support the origins of OA in skeletogenesis. Whereas methylation quantitative trait loci (mQTLs) co-localizing with OA genome-wide association study signals have been identified in aged human cartilage and used to identify effector genes and variants, such analyses have never been conducted during human development. Here, for the first time, we have investigated the developmental origins of OA genetic risk at seven well-characterized OA risk loci, comprising 39 OA-mQTL CpGs, in human fetal limb (FL) and cartilage (FC) tissues using a range of molecular genetic techniques. We identified significant OA-mQTLs at 14 and 29 CpGs in FL and FC tissues, respectively, and compared our results with aged cartilage samples (AC). Differential methylation was observed at 26 sites between FC and AC, with the majority becoming actively hypermethylated in old age. Notably, 6/9 OA effector genes showed allelic expression imbalances during fetal development. Finally, we conducted ATAC-sequencing in cartilage from the developing and aged hip and knee to identify accessible chromatin regions and found enrichment for transcription factor binding motifs including SOX9 and FOS/JUN. For the first time, we have demonstrated the activity of OA-mQTLs and expression imbalance of OA effector genes during human skeletogenesis. We show striking differences in the spatiotemporal function of these loci, contributing to our understanding of OA aetiology, with implications for the timing and strategy of pharmacological interventions.

Congenital Limb Defects: A Retrospective Cohort Study and Overview of the Literature.

Congenital limb defects are common malformations that are often associated with other organ defects and genetic disorders. Since prenatal detection is challenging and classification is often complex, the aim of this study was to describe a large cohort of fetuses with congenital limb defects and to identify characteristics that are essential for prenatal evaluation, counselling, and management. In this retrospective cohort study, all cases of confirmed fetal limb defects from two centers for prenatal ultrasound between 2001 and 2021 were evaluated. Cases with skeletal dysplasia were excluded from this study. Demographic data, association with genetic disorders, and correlation with maternal parameters were analyzed statistically. 170 fetuses were included in this study. 60% were diagnosed with a reduction anomaly and 40% with a duplication anomaly. The majority of fetuses were male, and in 73.5% of all cases, additional malformations were present. Among the genetic causes, trisomy 13 and 18 were the most common in this cohort. Congenital limb malformations are important markers for complex fetal disorders that warrant referral to specialists in prenatal ultrasound. To improve prenatal detection, care should be taken to visualize all fetal extremities already in early pregnancy.

Acetoacetate and β-hydroxybutyrate reduce mouse embryo viability via differential metabolic and epigenetic mechanisms

Research questionDoes the ketone acetoacetate (AcAc) alone, or combined with β-hydroxybutyrate (βOHB), impact mouse embryo development, metabolism, histone acetylation and viability? DesignPronucleate mouse oocytes were cultured in vitro in G1/G2 media supplemented with ketones (AcAc or AcAc + βOHB) at concentrations representing those in maternal serum during pregnancy (0.04 mmol/l AcAc, 0.1 mmol/l βOHB), standard diet consumption (0.1 mmol/l AcAc, 0.25 mmol/l βOHB), ketogenic diet consumption (0.8 mmol/l AcAc, 2 mmol/l βOHB) and diabetic ketoacidosis (2 mmol/l AcAc, 4 mmol/l βOHB). Day 5 blastocysts were assessed for cell allocation, glucose metabolism and histone acetylation. Day 4 blastocysts exposed to 0.8 mmol/l AcAc + 2 mmol/l βOHB were transferred to standard-fed recipient females, and E14.5 fetal and placental development assessed. ResultsExposure to 2 mmol/l AcAc or 0.8 mmol/l AcAc + 2 mmol/l βOHB did not impair blastocyst development, but significantly increased glucose consumption (P = 0.001 each), lowered glycolytic flux (P = 0.01, P < 0.001) and elevated trophectoderm (TE) histone 3 lysine 27 acetylation (H3K27ac; P < 0.001 each) compared with unexposed controls. Preimplantation AcAc + βOHB exposure reduced post-implantation fetal development by 25% (P = 0.037), and delayed female-specific fetal limb development (P = 0.019) and estimated fetal age (P = 0.019) compared with controls. ConclusionPreimplantation exposure to ketones affects underlying metabolism and histone acetylation in blastocysts that are associated with persistent, female-specific perturbations in fetal development. A periconceptional diet that elevates ketone concentrations may impair human embryonic viability.

Resveratrol Synergistically Promotes BMP9-Induced Osteogenic Differentiation of Mesenchymal Stem Cells.

Background Mesenchymal stem cells (MSCs) differentiate into osteocytes, adipocytes, and chondrocytes. Resveratrol and bone morphogenetic protein 9 (BMP9) are known osteogenic induction factors of MSCs, but the effect of both resveratrol and BMP9 on osteogenesis is unknown. Herein, we explored whether resveratrol cooperates with BMP9 to improve osteogenic induction. Methods The osteogenic induction of resveratrol and BMP9 on C3H10T1/2 cells was evaluated by detecting the staining and activity of the early osteogenic marker alkaline phosphatase (ALP). In addition, the late osteogenic effect was measured by the mRNA and protein levels of osteogenic markers, such as osteopontin (OPN) and osteocalcin (OCN). To assess the bone formation function of resveratrol plus BMP9 in vivo, we transplanted BMP9-infected C3H10T1/2 cells into nude mice followed by intragastric injection of resveratrol. Western blot (WB) analysis was utilized to elucidate the mechanism of resveratrol plus BMP9. Results Resveratrol not only enhanced osteogenic induction alone but also improved BMP9-induced ALP at 3, 5, and 7 d postinduction. Both the early osteogenic markers (ALP, Runx2, and SP7) and the late osteogenic markers (OPN and OCN) were significantly increased when resveratrol was combined with BMP9. The fetal limb explant culture further verified these results. The in vivo bone formation experiment, which involved transplanting BMP9-overexpressing C3H10T1/2 cells into nude mice, also confirmed that resveratrol synergistically enhanced the BMP9-induced bone formation function. Resveratrol phosphorylated adenosine monophosphate- (AMP-) activated protein kinase (AMPK) and stimulated autophagy, but these effects were abolished by inhibiting AMPK and Beclin1 using an inhibitor or siRNA. Conclusions Resveratrol combined with BMP9 significantly improves the osteogenic induction of C3H10T1/2 cells by activating AMPK and autophagy.

Embryological approach to the morphology of the ligamentum mucosum of the human knee joint

PurposeThe ligamentum mucosum is a ligamentous structure within the synovial layer of the knee joint capsule. It usually arises from the infrapatellar fat pad and is inserted into the intercondylar notch of the femur. In recent years, more attention has been paid to this structure because of its clinical significance. Despite the growing attention, the complex morphology of the ligamentum mucosum has led to many misunderstandings among the scientists and clinicians. Therefore, the main objective of this study was to characterize the morphology of the ligamentum mucosum and its attachment points, classify it, and evaluate its prevalence. MethodsClassical anatomical dissection was performed on 70 lower limbs of human fetuses fixed in 10% formalin solution. Morphology was carefully assessed, and morphometric measurements were performed. The types revealed were classified. ResultsThe ligamentum mucosum was present in 100% of all 70 specimens examined. Two types were detected. Type I (65.71%) - "cord-like". This type was the most common and is characterized by a single longitudinal ligament. The proximal attachment of the ligamentum mucosum was located in the middle of the infrapatellar fat, and the distal attachment was at the tip of the intercondylar notch. Type II (34.29%) - "vertical septum". This type is less common, but its morphology is much more complex compared to type I. It is wider and thicker than type I. Proximally, it is attached to the infrapatellar fat pad and distally through the entire intercondylar notch down to the anterior surface of the anterior cruciate ligament. It divides the joint cavity into medial and lateral sides. ConclusionThe ligamentum mucosum exhibits high morphologic variability, and the view that it is a vestigial remnant of the embryologic development of the knee should be reconsidered. We have proposed an anatomical classification of its types in human fetuses, which is a valuable addition to existing knowledge and will hopefully change the approach of clinicians and scientists.

Amniotic band syndrome: A case series

Introduction: Amniotic band sequence (ABS) is a constellation of congenital anomalies that can be seen in infants without any known genetic mutations. Amniotic band syndrome is thought to be responsible for 1/70 still births. There are two leading theories for proposed pathogenesis of ABS, commonly referred to as “extrinsic” and “intrinsic” models. Case Series: Our case series depicts four deliveries at two large military hospitals. Each of our patient case demonstrates the spectrum of ABS and supports the extrinsic theory as the primary mechanism. The first case resulting in lateral encephalocele with a complex facial cleft and subsequent intrauterine fetal demise. The second patient case resulting in fetal unilateral lower extremity limb reduction. The third patient case resulting in premature prelabor rupture of membranes and delivery at 29 weeks with pedal amputation. The fourth and final patient case resulted in a lethal limb-body-wall defect with extravasation of visceral organs and a sacral myelomeningocele. Conclusion: All four of our cases lend support to the extrinsic theory as the primary pathological mechanism. Each of our patients had risk factors for potential hypoxic injury and subsequent amniotic band formation.

Isolated Congenital Giant Fetal Upper Limb and Trunk Hemangioma Diagnosed Prenatally Using 4D Ultrasonography and Regressed by Age of 5 Months: a Case Report

Isolated Congenital Giant Fetal Upper Limb and Trunk Hemangioma Diagnosed Prenatally Using 4D Ultrasonography and Regressed by Age of 5 Months: a Case Report

A study of primary ossification centers in the hind limbs of Awasi sheep fetuses by double stains method and radiography.

Forty Local Awasi sheep fetuses were collected from several areas in Ninevah Governorate, further than local slaughterhouse. Their estimated ages calculated depending on crown-rump length with the aid of Richardson formula. The study aimed to detect the site and the time of appearance of ossification centers in the hind limb bones of Awasi sheep fetuses by using modified double staining method to demonstrate cartilage and bone , or by using radiography. The first ossification centers appeared in the hind limb bones was the primary center of the tibia and femur ,metatarsus with ilium then ischium at 45 , 46 , 48, 50-52 days of fetal age respectively. The ossification centers of pes phalanges appeared in the following sequence ; distal phalanx ,followed by the proximal phalanx and end by middle phalanx , at 58 ,60, 64 days respectively. Ossification centers of tarsal bones began to appear in calcaneus at 64 day , talus at 77 day , central & 4th tarsal bones at 96 day ,, followed by centers of 3rd & 2nd tarsal bones at 100 day while, the center of 1st tarsal bone appeared lastly at 105 day.Femur , tibia and metatarsus studied statistically by follow up the relative increase of their total lengths and length of their ossified part of diaphysis , the results show high significant variance in their growth and ossification of the three bones in first studied period (46-60 days) in comparison with the next sixth successive fortnightly periods

Computation of Fetal Kicking in Various Fetal Health Examinations: A Systematic Review.

Fetal movement has always been considered an essential indicator to evaluate the health of the unborn fetus. Many factors affect fetal movement. The frequency of fetal kicking is an important measurement of whether fetal development is progressing and healthy. Various instruments and methods of detecting fetal movement have been used and each method has its advantages and disadvantages. Although limited by the fetal environment in utero, the finite element method and musculoskeletal model can be used to calculate fetal lower limb movement. This review aims to summarize the current detection techniques for fetal movement, especially in the lower limbs. These will be outlined by describing the different measurements of fetal movement, and the related biomechanical analyses of fetal lower limb skeletogenesis and the associated muscular development to better evaluate and calculate the movements of the fetus in the womb.

179 A novel misense mutation of col2a1 gene as a cause of short extremeties in two first cousins

Introduction: Etiopathogenesis of fetal limb abnormalities involves genetic disorders, intrauterine factors, and maternal disease or exposure, while in many cases it remains unknown. Aim: We present two related cases (first cousins) of short limbed fetuses caused by a novel mutation of COL2A1 gene (NM_001844.5). Case 1 description: A 29.24-year-old pregnant woman with an uncomplicated pregnancy, presented for b-scan. Limbs were found to be well below the 3rd centile for gestational age. Initial fetal genetic work-up (targeted) was negative and the rest of the maternal follow-up was unremarkable. A healthy baby was delivered and the pediatric follow-up for an SGA child was appropriate. On second pregnancy a normal baby was born. Case 2 description: A 33.01-year-old pregnant woman with an uncomplicated pregnancy, presented for 3rd trimester Doppler scan. Limbs were found to be well below the 1rd centile for gestational age. Given that the two cases were related (first cousins) whole exome sequencing (WES) was performed on Case 2. WES revealed a novel heterozygous missense mutation c.1132G>A (p. Gly378Ser) of COL2A1 gene (NM_001844.5). Subsequently targeted genetic sequencing for the mutation was performed on Case 1 and the same novel mutation was found, making it potentially pathogenic. Targeted sequencing revealed the same mutation in the mother of Case 1 and the Father of Case 2 (siblings). Discussion: Heterozygous mutations of COL2A1 have been associated with broad clinical spectrum (collagenopathy type II), ranging from lethal to the fetus to mild appearing in adult life as early arthritis. More frequently they appear as skeletal dysplasia with ocular, hearing, and craniofacial findings. Conclusion: Ultrasonography has allowed increasingly accurate detection of skeletal abnormalities in utero. Efficient genetic assessment as part of an MDT approach can lead to a quick diagnosis and elucidation of novel genetic causes and appropriate pediatric follow-up of the child.

Foetal Limb in Bladder: A rare complication of vesicouterine fistula

Vesicouterine fistula(VUF) is a rare urogenital fistula and more rarer is the pregnancy in its presence. Even if pregnancy occurs the outcomes appears to be very poor .Most of the cases are related to iatrogenic bladder injury during caesarean section. There is very limited literature on the pregnancy with VUF associated with herniation of foetus or foetal part(s) into the bladder cavity. Here we report a case of pregnancy in a known case of untreated VUF. She presented at 23 weeks of gestation with features of premature rupture of membrane and herniation of foetal left lower limb inside the bladder cavity. Her pregnancy ended up with hysterotomy and removal of non-viable foetus along with fistula repair. So, regardless of the severity of the signs and symptoms associated with VUF clinicians should convince the patients for repair of the fistula especially if future pregnancy is contemplated .