Biology subjects

Andrews, P. W.

Publications and source records attributed to Andrews, P. W..

8 recordsLinked to original sources

bioRxiv2025

Functional and molecular rescue of aganglionic colon by human enteric nervous system progenitor transplantation in Hirschsprung disease

Hirschsprung disease (HSCR) is a devastating congenital disorder characterised by absence of the enteric nervous system (ENS) in the distal gut. Cell therapy, using human pluripotent stem cell-derived ENS progenitors, offers an attractive alternative to treat this life-limiting disorder. Here we provide an in-depth characterisation of the HSCR phenotype in the B6;129-Ednrbtm1Ywa/J mouse model including molecular analyses highlighting the wide-ranging molecular effects of aganglionosis in HSCR colon. We show that transplantation of hPSC-derived ENS progenitors leads to significant increases in contractile function with formation of extensive donor-derived neuronal networks in colonic explants. Moreover, post-transplantation, we show that integration of ENS progenitors rescues many of the biological pathways impacted in the HSCR aganglionic microenvironment. These novel results provide strong evidence that human ENS progenitor transplantation can modulate critical signalling pathways affected in HSCR and exert positive impacts on contractile function, in aganglionic tissue, supporting further translational development of this regenerative medicine approach.

cell biology↗

bioRxiv2022

Neuroblastoma-associated chromosomal aberrations drive cell identity loss in human neural crest via disruption of developmental regulators

Early childhood tumours arise from transformed embryonic cells, which often carry large copy number alterations (CNA). However, it remains unclear how CNAs contribute to embryonic tumourigenesis due to a lack of suitable models. Here we employ female human embryonic stem cell (hESC) differentiation and single-cell transcriptome and epigenome analysis to assess the effects of chromosome 17q/1q gains, which are prevalent in the embryonal tumour neuroblastoma (NB). We show that CNAs impair the specification of trunk neural crest (NC) cells and their sympathoadrenal derivatives, the putative cells-of-origin of NB. This effect is exacerbated upon overexpression of MYCN, whose amplification co-occurs with CNAs in NB. Moreover, CNAs potentiate the pro-tumourigenic effects of MYCN and mutant NC cells resemble NB cells in tumours. These changes correlate with a stepwise aberration of developmental transcription factor networks. Together, our results sketch a mechanistic framework for the CNA-driven initiation of embryonal tumours.

cancer biology↗

bioRxiv2020

Ectopic FGF23 production induces mineral changes, osteogenic transdifferentiation, and cancer associated microcalcifications

Testicular microcalcifications consist of hydroxyapatite and have been associated with an increased risk of testicular germ cell tumors (TGCTs) but are also found in benign cases such as loss-of-function variants in the phosphate-transporter SLC34A2. Here, we show that fibroblast growth factor 23 (FGF23), a regulator of phosphate homeostasis, is expressed in testicular germ cell neoplasia in situ (GCNIS), embryonal carcinoma (EC), and human embryonic stem cells. FGF23 is not glycosylated in TGCTs and therefore cleaved into a C-terminal fragment which competitively antagonizes full-length FGF23. Here, Fgf23 knockout mice presented with marked calcifications in the epididymis, spermatogenic arrest, and focally germ cells expressing the osteoblast marker Osteocalcin (gene name: BGLAP, protein name: Osteocalcin). Moreover, the frequent testicular microcalcifications in mice with no functional androgen receptor and lack of circulating gonadotropins are associated with lower Slc34a2 and higher Bglap/Slc34a1 (protein name: NPT2a) expression compared with wild-type mice. In accordance, human testicular specimens with microcalcifications also have lower SLC34A2 and a subpopulation of germ cells express phosphate-transporter NPT2a, Osteocalcin, and RUNX2 highlighting aberrant local phosphate handling and expression of bone-specific proteins. Mineral disturbance in vitro using calcium or phosphate treatment induced deposition of calcium-phosphate in a spermatogonial cell line and this effect was fully rescued by the mineralization-inhibitor pyrophosphate. In conclusion, testicular microcalcifications arise secondary to local alterations in mineral homeostasis, which in combination with impaired Sertoli cell function and reduced levels of mineralization-inhibitors due to high alkaline phosphatase activity in GCNIS and TGCTs, facilitate osteogenic-like differentiation of testicular cells and deposition of hydroxyapatite.

molecular biology↗

bioRxiv2020

Defining the signalling determinants of a posterior ventral spinal cord identity in human neuromesodermal progenitor derivatives

The anteroposterior axial identity of motor neurons (MNs) determines their functionality and vulnerability to neurodegeneration. Thus it is a critical parameter in the design of strategies aiming to produce MNs from human pluripotent stem cells (hPSCs) for regenerative medicine and disease modelling applications. However, the in vitro generation of posterior spinal cord MNs has been challenging. Although the induction of cells resembling neuromesodermal progenitors (NMPs), the bona fide precursors of the mammalian spinal cord, offers a promising solution, the progressive specification of posterior MNs from these cells is not well-defined. Here we determine the signals guiding the transition of human NMP-like cells toward posterior ventral spinal cord neurectoderm. We show that combined WNT-FGF activities drive a posterior dorsal early neural state while suppression of TGFβ-BMP signalling pathways, combined with SHH stimulation, promotes a ventral identity. Based on these results, we define an optimised protocol for the generation of posterior MNs that can efficiently integrate within the neural tube of chick embryos. We expect that our findings will facilitate the functional comparison of hPSC-derived spinal cord cells of distinct axial identities.Competing Interest StatementThe authors have declared no competing interest.View Full Text

developmental biology↗

bioRxiv2019

Nucleosides rescue replication-mediated genome instability of human pluripotent stem cells

Human pluripotent stem cells (PSC) often acquire genetic changes on prolonged culture, which pose concerns for their use in research and regenerative medicine (Amps et al., 2011, Seth et al., 2011). The acquisition of these changes during culture necessarily first requires mutation and then selection of those mutations that provide a growth advantage. Whilst selection accounts for the recurrent nature of the variants commonly reported (Draper et al., 2004, Olariu et al., 2010), the mechanisms of mutation in PSC remain largely elusive. Here we show that, in contrast to somatic cells, human PSC have an increased susceptibility to DNA damage and mitotic errors, both of which are caused by heightened replication stress in PSC and this can be alleviated by culture with exogenous nucleosides. These results reflect the requirement for rapid replication of human PSC enabled by a truncated G1 (Becker et al., 2006, Becker et al., 2010) that impairs the preparation of these cells for the ensuing DNA replication. A similar relationship has been shown in relation to chromosomal instability in cancer cells (Burrell et al., 2013, Wilhelm et al., 2019) but PSC differ by replication stress triggering apoptosis (Desmarais et al., 2012, Desmarais et al., 2016). Nevertheless, evasion of this response still leads to the appearance of genetic variants that are of concern for regenerative medicine. The inclusion of nucleosides into culture media greatly improves the efficiency of human PSC culture and minimises the acquisition of genomic damage.

cell biology↗

bioRxiv2019

Retinoic acid accelerates the specification of enteric neural progenitors from in vitro-derived neural crest

The enteric nervous system (ENS) is derived primarily from the vagal neural crest, a migratory multipotent cell population emerging from the dorsal neural tube between somites 1-7. Defects in the development and function of the ENS give rise to a range of disorders, termed enteric neuropathies and include conditions such as Hirschsprungs disease. Little is known about the signalling that specifies early ENS progenitors. This has, thus far, limited progress in the generation of enteric neurons from human Pluripotent Stem Cells (hPSCs) that could provide a useful tool for disease modelling and regenerative medicine. We describe the efficient and accelerated generation of ENS progenitors from hPSCs, revealing that retinoic acid is critical for the acquisition of both vagal axial identity and early ENS progenitor specification. These ENS progenitors generate enteric neurons in vitro and following in vivo transplantation, achieving long-term colonisation of the ENS in adult mice. Thus, hPSC-derived ENS progenitors may provide the basis for cell therapy for defects in the ENS.

developmental biology↗

bioRxiv2019

Generation and Trapping of a Mesoderm Biased State of Human Pluripotency

We postulate that exit from pluripotency involves intermediates that retain pluripotency while simultaneously exhibiting lineage-bias. Using a MIXL1 reporter, we explored mesoderm lineage-bias within the human pluripotent stem cell compartment. We identified a substate, which at the single cell level coexpresses pluripotent and mesodermal gene expression programs. Functionally these cells could initiate stem cell cultures and exhibited mesodermal bias in differentiation assays. By promoting mesodermal identity through manipulation of WNT signalling while preventing exit from pluripotency using lysophosphatidic acid, we could trap and maintain cells in a lineage-biased stem cell state through multiple passages. These cells correspond to a normal state on the differentiation trajectory, the plasticity of which is evidenced by their reacquisition of an unbiased state upon removal of differentiation cues. The use of cross-antagonistic signalling to trap pluripotent stem cell intermediates with different lineage-bias may have general applicability in the efficient production of cells for regenerative medicine.

cell biology↗

bioRxiv2018

Human axial progenitors generate trunk neural crest cells

The neural crest (NC) is a multipotent embryonic cell population generating distinct cell types in an axial position-dependent manner. The production of NC cells from human pluripotent stem cells (hPSCs) is a valuable approach to study human NC biology. However, the origin of human trunk NC remains undefined and therefore current in vitro differentiation strategies induce only a modest yield of trunk NC cells. Here we show that hPSC-derived axial progenitors, the posteriorly-located drivers of embryonic axis elongation, give rise to trunk NC cells and their derivatives. Moreover, we define the molecular signatures associated with the emergence of human NC cells of distinct axial identities in vitro. Collectively, our findings indicate that there are two routes toward a human post-cranial NC state: the birth of cardiac and vagal NC is facilitated by retinoic acid-induced posteriorisation of an anterior precursor whereas trunk NC arises within a pool of posterior axial progenitors.

developmental biology↗