Biomedical subjects
Publications and source records attributed to E C Jesudason.
Wilms' tumour commonly presents with an abdominal mass and gross haematuria. Here, we present the novel application of paediatric renal arterial embolisation to control life-threatening haematuria in Wilms' tumour.
Congenital diaphragmatic hernia (CDH) is a common birth defect which continues to challenge paediatric surgeons and intensivists. Affecting approximately 1:2500 births, a baby with CDH is born every 24-36 hours in the UK.
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Lung hypoplasia is central to the poor prognosis of babies with congenital diaphragmatic hernia (CDH). Prolapse of abdominal organs through a diaphragmatic defect has traditionally been thought to impair lung growth by compression. The precise developmental biology of CDH remains unresolved. Refractory to fetal correction, lung hypoplasia in CDH may instead originate during embryogenesis and before visceral herniation. Resolving these conflicting hypotheses may lead to reappraisal of current clinical strategies. Genetic studies in murine models and the fruitfly, Drosophila melanogaster are elucidating the control of normal respiratory organogenesis. Branchless and breathless are Drosophila mutants lacking fibroblast growth factor (FGF) and its cognate receptor (FGFR), respectively. Sugarless and sulphateless mutants lack enzymes essential for heparan sulphate (HS) biosynthesis. Phenotypically, all these mutants share abrogated airway branching. Mammalian organ culture and transgenic models confirm the essential interaction of FGFs and HS during airway ramification. Embryonic airway development (branching morphogenesis) occurs in a defined spatiotemporal sequence. Unlike the surgically-created lamb model, the nitrofen rat model permits investigation of embryonic lung growth in CDH. Microdissecting embryonic lung primordia from the nitrofen CDH model and normal controls, we demonstrated that disruption of stereotyped airway branching correlates with and precedes subsequent CDH formation. To examine disturbed branching morphogenesis longitudinally, we characterised a system that preserves lung hypoplasia in organ culture. We tested FGFs and heparin (an HS analogue) as potential therapies on normal and hypoplastic lungs. Observing striking differences in morphological response to FGFs between normal and hypoplastic lung primordia, we postulated abnormalities of FGF/HS signalling in the embryonic CDH lung. Evaluating this hypothesis further, we examined effects of an HS-independent growth factor (epidermal growth factor, EGF) on hypoplastic lung development. Visible differences in morphological response indicate an intrinsic abnormality of hypoplastic lung primordia that may involve shared targets of FGFs and EGE. These studies indicate that lung hypoplasia precedes diaphragmatic hernia and may involve disturbances of mitogenic signalling pathways fundamental to embryonic lung development. What does this imply for human CDH? Fetal surgery may be 'too little, too late' to correct an established lung embryopathy. In utero growth factor therapy may permit antenatal lung rescue. Prevention of the birth defect by preconceptual prophylaxis may represent the ultimate solution.
It has been hypothesised that the liver induces lung hypoplasia in congenital diaphragmatic hernia (CDH) by non-compressive intrathoracic growth rather than traditional mass herniation. Utilising a co-culture system, we tested the capacity of liver cells to inhibit lung growth by contact rather than compression. Heart, liver, and lungs were microdissected from normal rat embryos (n > 20 from at least three litters) on day 13.5 of gestation. Monolayer cultures of enzymatically dispersed livers and hearts were established at the same cell density. Lung primordia were cultured in direct contact with hepatic cells or partitioned from them by a permeable polytetrafluoroethylene membrane. This permits the contributions of diffusable factors and cell contact to be distinguished. Lungs were similarly cultured in direct contact with or partitioned from cardiac cells. Lungs cultured in isolation served as further controls. Daily inspection permitted assessment of in-vitro lung growth. Growth of lungs in direct contact with hepatic cells was equivalent to that of lungs partitioned from liver cells. Lungs in direct contact with cardiac cells and lungs partitioned from cardiac cells were also not inhibited compared to lungs cultured in isolation. Early lung development is thus not inhibited by humoral or contact-mediated interactions with embryonic liver cells. Lung hypoplasia in CDH is therefore unlikely to originate from contact inhibition with the developing liver. An intrinsic pulmonary defect may better explain hypoplastic lung development in CDH.
Pulmonary hypoplasia (PH) is a leading contributor to the lethality of congenital diaphragmatic hernia (CDH). Studies now suggest that PH arises prior to visceral herniation. Growth factors (GF) are pivotal to this embryonic lung growth. With striking in-vitro effects on lung morphogenesis, GF are under investigation as therapies for PH. Heparin modulates the kinetics of heparan-sulphate binding ligands that drive lung development. We hypothesised that heparin may rescue PH by favourable alteration of endogenous pulmonary GF activity. Normal and hypoplastic lung primordia were microdissected on day 13.5 of gestation and cultured for up to 78 h in plain media with and without heparin. In-vitro morphological development was studied by serial measurements of terminal bud count, lung area, and lung perimeter. Nitrofen-exposed lungs cultured with heparin showed no significant improvements in terminal bud count, lung area, and lung perimeter at 30, 54, and 78 h compared to untreated hypoplastic lungs maintained in vitro. In normal lungs heparin demonstrated no sustained significant morphological effects compared to untreated control lungs. In this study, heparin did not stimulate branching morphogenesis of normal or hypoplastic lungs in our organ culture system. Known at higher concentrations to inhibit smooth-muscle proliferation, heparin may ameliorate pulmonary vascular hypermuscularisation with the prospect of benefiting CDH infants on extracorporeal membrane oxygenation. Future studies will address the impact of exogenous GF on hypoplastic lung development in organ culture.
BACKGROUND/PURPOSE: Lung hypoplasia, a leading contributor to the lethality of congenital diaphragmatic hernia (CDH), has been attributed to compression of the fetal lung by herniated abdominal viscera. Contested findings in experimental CDH suggest that lung malformation may precede diaphragmatic hernia. To address this unresolved question, we studied the pattern and progress of embryonic lung development in vivo and in vitro before diaphragmatic herniation in the nitrofen CDH model. METHODS: Sprague-Dawley rats were fed nitrofen on day 9.5 of pregnancy to induce pulmonary hypoplasia and CDH in newborns (term, day 22). Control rats received olive oil. Embryonic lungs were microdissected on day 13.5 gestation, 24 hours after lung primordia bud from the foregut (normal diaphragmatic closure, day 16.5). In vivo airway branching was measured by counting terminal lung buds at this stage. Lungs were cultured for up to 78 hours and longitudinal in vitro development studied by serial measurements of terminal bud count, area, and perimeter. RESULTS: At 13.5 days of gestation in vivo, nearly 99% of normal lungs (n = 130) had > or = 6 terminal lung buds. In contrast, 36% of the nitrofen-exposed lungs (n = 170) fell short of this developmental milestone with less than 6 terminal buds (P < .001). In vitro, the nitrofen lungs had reduced area compared with controls after 6, 30, and 54 hours (P = .001, P < .001, and P = .001, respectively). Bud count and epithelial perimeter were reduced in the nitrofen lungs after 6 and 30 hours in vitro (P < .001 and P = .01 v normal terminal bud count; P < .001 and P= .002 v normal perimeter). CONCLUSIONS: Before diaphragmatic herniation, nitrofen-exposed embryos in vivo have a 36% frequency of reduced airway branching that strikingly parallels the 30% to 40% term incidence of CDH in this model. The authors propose that this early lung anomaly is restricted to a subset of nitrofen-exposed embryos in which it is integral to an emerging CDH phenotype. In vitro data confirm that nitrofen-induced pulmonary hypoplasia precedes visceral herniation and persists to allow experimental manipulation in culture. The developmental biology underlying lung hypoplasia in CDH may now be addressed using this organ culture system.
BACKGROUND/PURPOSE: Current treatment for lethal pulmonary hypoplasia in congenital diaphragmatic hernia (CDH) may be hampered by uncertainty over its origin. Herniation of abdominal organs into the chest was thought to produce lung hypoplasia by compression. Experimental CDH models suggest that disturbed lung growth precedes these events. Mammalian development comprises cell differentiation, proliferation and programmed cell death or apoptosis. Could lung hypoplasia in CDH result from alterations in these processes well before visceral herniation takes place? The aim of this study was to compare cell proliferation and apoptosis in normal and hypoplastic embryonic lungs before normal diaphragmatic closure using a CDH model. METHODS: Sprague-Dawley rats were given 100 mg of nitrofen on day 9.5 of pregnancy to create lung hypoplasia and CDH in newborns (term, 22 days). Control rats received olive oil. Cell proliferation in embryonic lung specimens was measured by bromodeoxyuridine (BrdU) incorporation at 13.5 to 15.5 days' gestation, before normal diaphragmatic closure in this species (day 16.5). Apoptosis was measured by the in situ end-nick labelling (TUNEL) method in lung sections obtained from rat embryos of 13.5 to 16.5 days' gestation. RESULTS: High levels of cell proliferation were seen in both normal control and nitrofen-exposed lungs. However, 24 hours before normal diaphragmatic closure, nitrofen-exposed lungs had significant reductions in cell proliferation on day 15.5 of gestation (P = .009 v controls). Apoptosis occurred at low levels throughout the developmental stages examined (< 0.3%) without significant differences encountered between the study groups. CONCLUSIONS: These findings have shown high rates of cell division during normal lung development before diaphragmatic closure. Decreased levels over this critical period in gestation may contribute to early lung anomalies in the nitrofen CDH model. Strategies to promote cell proliferation in the fetal lung may therefore hold future promise in human CDH. Apoptosis appears not to play a major role in hypoplastic lung development. Therapies to inhibit apoptosis would seem unlikely to improve this early lung growth.
BACKGROUND/PURPOSE: Pulmonary hypoplasia, a leading contributor to the lethality of congenital diaphragmatic hernia (CDH), precedes diaphragmatic malformation in the nitrofen model and persists to allow experimental manipulations in organ culture. Fibroblast growth factors (FGFs) are crucial to early lung development. Acidic FGF (FGF-1) binds to all FGF receptors and enhances in vitro branching morphogenesis. Basic FGF (FGF-2) is localized to developing airway epithelium, basement membrane, and extracellular matrix. Heparin (HEP) modulates FGF kinetics and inhibits smooth muscle proliferation in lung primordia. The aim of this study was to examine the morphological effects of fibroblast growth factors and heparin on lung hypoplasia in an organ culture model. METHODS: Sprague-Dawley rats were fed nitrofen on day 9.5 of pregnancy to induce lung hypoplasia and CDH in newborns. Control rats received olive oil. Normal and hypoplastic lung primordia were microdissected on day 13.5 of gestation and cultured up to 78 hours in plain media with or without FGF-1 or FGF-2, with or without HEP. In vitro morphological development was studied by serial measurements of terminal bud count, lung area, and lung perimeter. RESULTS: Over 120 fetal lung specimens were studied (n > or = 4 per group). Significant increases in area, perimeter, and bud count were seen in normal lungs cultured with FGF-1 plus HEP compared with control media (P < .05). In the nitrofen lungs, FGF1 plus HEP yielded reductions in all parameters compared with those in control media (P < .05), whereas FGF-2 produced significant expansion in lung area but marked reductions in bud count and lung perimeter divided by square root of area (P < .05). Heparin did not produce substantial or sustained alteration of morphology in normal or hypoplastic lungs. CONCLUSIONS: These observations may indicate an intrinsic abnormality of FGF processing in the hypoplastic nitrofen lung before diaphragmatic malformation. Heparin did not rescue abnormal lung development. Mechanisms underlying the differential effects of these agents now need to be explored to target fetal lung growth and improve the dismal prognosis of human CDH.
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