Biology subjects

David, A. L.

Publications and source records attributed to David, A. L..

6 recordsLinked to original sources

bioRxiv2026

Derivation of primary fetal epithelial organoids from cryopreserved human amniotic fluid cells.

Primary fetal human epithelial organoids are canonically derived from tissue samples obtained after termination of pregnancy. Recently, we have demonstrated that these organoids can be consistently generated from amniotic fluid cells isolated prenatally during diagnostic and interventional procedures. Amniotic Fluid-derived Organoids (AFOs) are promising fetal epithelial lung, kidney and small intestine tissue models that bypass some of the ethical and legal constraints associated with obtaining primary fetal tissue. Despite this, the widespread adoption of this technology is limited by the lack of standardised and accessible pipelines for the biobanking, culturing and distribution of organoid-forming amniotic fluid cells. To enable broader AFO use for research and clinical purposes, this work investigates two cryopreservation strategies to optimise AF cell recovery for organoid derivation. In Strategy 1, viable AF cells were sorted before freezing, while in Strategy 2 unsorted AF was frozen, and viable AF cells were sorted for viability after thawing. We present here a comprehensive evaluation of 5 commercially available GMP-compliant freezing media (FM1-5) alongside a standard lab-grade control (FM CT). AFOs were assessed for formation efficiency, morphological characteristics, proliferation capacity, epithelial identity, and tissue type. Our results demonstrate that organoid-forming AF cells can be successfully cryopreserved both pre- and post-sorting. Strategy 1 yielded higher organoid formation efficiency, with AFOs derived from cryopreserved and fresh cells exhibiting comparable expansion potential. Finally, FM4 provided minimal to no decline in survival rate, making it the most effective GMP-grade freezing medium tested. In conclusion, we present two viable cryopreservation strategies adaptable to different laboratory settings and identify optimal GMP-compliant freezing media, to support safe distribution and centralised processing, thereby facilitating scalability and collaborative work on the AFO technology.

cell biology↗

bioRxiv2024

Low maternal plasma apelin in the second trimester is associated with adverse pregnancy outcome in severe, early-onset fetal growth restriction

ContextFetal growth restriction increases adverse pregnancy outcomes such as preterm birth and intrauterine fetal death. Apelin is a secreted peptide expressed in placental syncytiotrophoblast and downregulated in fetal growth restriction. ObjectivesWe tested the hypothesis that adverse pregnancy outcome is associated with low maternal plasma apelin at diagnosis of early-onset fetal growth restriction. MethodsPlasma samples and fetomaternal blood flow Doppler velocimetry measurements were obtained from pregnant women (n=59) at diagnosis of early-onset fetal growth restriction in the second trimester. Plasma apelin was determined by ELISA and pregnancy outcome was recorded. Placental gene expression was analysed after birth by qRT-PCR, compared to term placentas from women with late-onset fetal growth restriction or with appropriate-for-gestational age infants. ResultsAt diagnosis of early-onset fetal growth restriction, plasma apelin concentration was significantly lower in women who delivered extremely preterm (<28 weeks gestation) or had an intrauterine fetal death, compared to women who had a livebirth[≥]28 weeks (P<0.05). Plasma apelin correlated directly with uterine artery volume flow rate and inversely with pulsatility index. Placental gene expression of apelin, but not the apelin receptor or elabela, was lower in women with early-onset fetal growth restriction delivering preterm than in appropriate-for-gestational-age, term control women. ConclusionLow maternal circulating apelin during the second trimester is associated with impaired uteroplacental perfusion and subsequent adverse pregnancy outcome in severe, early-onset fetal growth restriction. Placental apelin deficiency may contribute mechanistically to the pathogenesis of early-onset fetal growth restriction.

physiology↗

bioRxiv2024

Pleotropic effects of a recessive COL1A2 mutation occurring in a mouse model of severe osteogenesis imperfecta

Approximately 85-90% of individuals with Osteogenesis Imperfecta (OI) have dominant pathogenic variants in the COL1A1 or COL1A2 genes. This leads to decreased or abnormal Collagen type I production. Subsequently, bone formation is strongly reduced, causing bone fragility and liability to fractures throughout life. OI is clinically classified in 5 types with the severity ranging from mild to lethal depending on the gene and the type and location of the OI-causative variant and the subsequent effect on (pro) collagen type I synthesis. However, the specific effects on the phenotype and function of osteoblasts are not fully understood. To investigate this, the OI murine model was used, with the oim/oim (OIM) mice closest resembling severely deforming OI type 3 in humans. We showed that in OIM, COL1 mutation results in a multifactorial inhibition of the osteogenic differentiation and maturation as well as inhibition of osteoclastogenesis. The phenotype of differentiated OIM osteoblasts also differs from that of wild type mature osteoblasts, with upregulated oxidative cell stress and autophagy pathways, possibly in response to the intracellular accumulation of type I collagen mRNA. The extracellular accumulation of defective type I collagen fibres contributes to activation of the TGF-{beta} signalling pathway and activates the inflammatory pathway. These effects combine to destabilise the balance of bone turnover, increasing bone fragility. Together, these findings identify the complex mechanisms underlying OI bone fragility in the OIM model of severe OI and can potentially enable identification of clinically relevant endpoints to assess the efficacy of innovative pro-osteogenic treatment for patients with OI.

genetics↗

bioRxiv2023

Single cell-guided prenatal derivation of primary epithelial organoids from the human amniotic and tracheal fluids.

Despite advances in prenatal diagnosis, it is still difficult to predict severity and outcomes of many congenital malformations. New patient-specific prenatal disease modelling may optimise personalised prediction. We and others have described the presence of mesenchymal stem cells in amniotic fluid (AFSC) that can generate induced pluripotent stem cells (iPSCs). The lengthy reprogramming processes, however, limits the ability to define individual phenotypes or plan prenatal treatment. Therefore, it would be advantageous if fetal stem cells could be obtained during pregnancy and expanded without reprogramming. Using single cell analysis, we characterised the cellular identities in amniotic fluid (AF) and identified viable epithelial stem/progenitor cells of fetal intestinal, renal and pulmonary origin. With relevance for prenatal disease modelling, these cells could be cultured to form clonal epithelial organoids manifesting small intestine, kidney and lung identity. To confirm this, we derived lung organoids from AF and tracheal fluid (TF) cells of Congenital Diaphragmatic Hernia (CDH) fetuses and found that they show differences to non-CDH controls and can recapitulate some pathological features of the disease. Amniotic Fluid Organoids (AFO) allow investigation of fetal epithelial tissues at clinically relevant developmental stages and may enable the development of therapeutic tools tailored to the fetus, as well as to predicting the effects of such therapies.

cell biology↗

bioRxiv2020

Impaired Fetal Lung Development can be Rescued by Administration of Extracellular Vesicles Derived from Amniotic Fluid Stem Cells

Incomplete lung development, also known as pulmonary hypoplasia, is a recognized cause of neonatal death and poor outcome for survivors. To date, there is no effective treatment that promotes fetal lung growth and maturation. Herein, we describe a novel stem cell-based approach that enhances fetal lung development via the administration of extracellular vesicles (EVs) derived from amniotic fluid stem cells (AFSCs). In experimental models of pulmonary hypoplasia, administration of AFSC-EVs promoted lung branching morphogenesis and alveolarization, and stimulated pulmonary epithelial cell and fibroblast differentiation. This regenerative ability was confirmed in two models of injured human lung cells, where human AFSC-EVs obtained following good manufacturing practices restored pulmonary epithelial homeostasis. AFSC-EV beneficial effects were exerted via the release of RNA cargo, primarily miRNAs, that regulate the expression of genes involved in fetal lung development. Our findings suggest that AFSC-EVs hold regenerative ability for underdeveloped fetal lungs, demonstrating potential for therapeutic application. One Sentence SummaryFetal lung regeneration via administration of extracellular vesicles derived from amniotic fluid stem cells

developmental biology↗

bioRxiv2019

Stem cell delivery to kidney via minimally invasive ultrasound-guided renal artery injection in mice

Cell-based therapies are promising treatments for various kidney diseases. However, the major hurdle in initiating therapeutic responses is the inefficiency of injection routes to deliver cells to the kidney parenchyma. Systemic injection, such as intravenous injection only delivers a small proportion of cells to the kidney. Whereas direct delivery, such as renal artery injection require surgical procedures. A minimally invasive renal artery injection was therefore developed to enhance cell delivery to kidney. In this study, luciferase expressing human adipocyte derived stem cells (ADSC) were labelled with gold nanorods (GNR) and injected into the renal artery using ultrasound guidance. The ADSCs were tracked using bioluminescence and photoacoustic imaging serially over 7 days. Imaging confirmed that the majority of signal was within the kidney, indicative of successful injection and that signal remained stable for 3 days. Histology showed co-localization of GNRs with ADSC staining throughout the kidney with no indication of injury caused by injection. These findings demonstrate that ultrasound-guided renal artery injection is feasible in mice and can successfully deliver a large proportion of cells which are retained within the kidney for 3 days. Therefore, the techniques developed here will be useful for optimising cell therapy in kidney diseases.

cell biology↗