Genetic variation at mouse and human ribosomal DNA influences associated epigenetic states

Background: Ribosomal DNA (rDNA) displays substantial inter-individual genetic variation in human and mouse. A systematic analysis of how this variation impacts epigenetic states and expression of the rDNA has thus far not been performed. Results: Using a combination of long- and short-read sequencing, we establish that 45S rDNA units in the C57BL/6J mouse strain exist as distinct genetic haplotypes that influence the epigenetic state and transcriptional output of any given unit. DNA methylation dynamics at these haplotypes are dichotomous and life-stage specific: at one haplotype, the DNA methylation state is sensitive to the in utero environment, but refractory to post-weaning influences, whereas other haplotypes entropically gain DNA methylation during aging only. On the other hand, individual rDNA units in human show limited evidence of genetic haplotypes, and hence little discernible correlation between genetic and epigenetic states. However, in both species, adjacent units show similar epigenetic profiles, and the overall epigenetic state at rDNA is strongly positively correlated with the total rDNA copy number. Analysis of different mouse inbred strains reveals that in some strains, such as 129S1/SvImJ, the rDNA copy number is only approximately 150 copies per diploid genome and DNA methylation levels are < 5%. Conclusions: Our work demonstrates that rDNA-associated genetic variation has a considerable influence on rDNA epigenetic state and consequently rRNA expression outcomes. In the future, it will be important to consider the impact of inter-individual rDNA (epi)genetic variation on mammalian phenotypes and diseases

The Movember Prostate Cancer Landscape Analysis: an assessment of unmet research needs

Prostate cancer is a heterogeneous cancer with widely varying levels of morbidity and mortality. Approaches to prostate cancer screening, diagnosis, surveillance, treatment and management differ around the world. To identify the highest priority research needs across the prostate cancer biomedical research domain, Movember conducted a landscape analysis with the aim of maximizing the effect of future research investment through global collaborative efforts and partnerships. A global Landscape Analysis Committee (LAC) was established to act as an independent group of experts across urology, medical oncology, radiation oncology, radiology, pathology, translational research, health economics and patient advocacy. Men with prostate cancer and thought leaders from a variety of disciplines provided a range of key insights through a range of interviews. Insights were prioritized against predetermined criteria to understand the areas of greatest unmet need. From these efforts, 17 research needs in prostate cancer were agreed on and prioritized, and 3 received the maximum prioritization score by the LAC: first, to establish more sensitive and speci

Mechanical ventilation of the very immature lung: mechanisms of injury and repair

Very preterm infants often require mechanical ventilation (MV) due to respiratory insufficiency. However, respiratory support that includes MV can injure the immature lung and contribute to the development of bronchopulmonary dysplasia (BPD). Structural changes in the lungs that are associated with MV and BPD can persist, resulting in deficits in lung function in children and young adults. Multiple factors are known to contribute to the development of BPD including MV, the use of supplemental oxygen, infection and impaired nutrition, so it has been difficult to determine the effects of MV alone. Using our unique model of MV-induced lung injury in the fetal sheep we are able to investigate the effects of MV alone on the immature lung. To determine if MV-induced injury in the very immature lung persists or resolves in the absence of further ventilation, we exposed very immature fetal sheep to a brief period (2h) of MV and examined the lungs 24h and 15d later. These studies were conducted in sheep in both the saccular and early-alveolar stages of lung development. Knowledge about the susceptibility to MV-induced injury at different stages of lung development will provide important information regarding the factors contributing to BPD in preterm infants. At both stages of lung development brief, injurious MV caused severe lung injury at 24h. Lungs displayed reduced secondary septal crest density, atelectasis, disorganised extracellular matrix deposition and hemorrhage. Bronchioles had thickened, injured epithelium and often contained luminal debris. MV caused differences in lung injury manifestation between saccular and early-alveolar stage lungs. Fifteen days after MV was performed saccular and early-alveolar stage lungs spontaneously repaired and displayed no signs of injury or remodelling. In order to determine if injury and/ or repair mechanisms are active in our model of MV-induced injury 24h after brief MV mRNA and protein expression of cytokines, early response genes and a subset of putative repair genes were evaluated. Cytokine and early response gene mRNA levels were not elevated and the protein deposition of early response genes was also not increased. The mRNA expression levels of two putative repair genes, metallothionein and urokinase plasminogen activator receptor, were significantly increased. Protein deposition of metallothionien was also increased in ventilated lungs. This study has shown for the first time that 24h after MV of the very immature lung, while lungs remain severely injured and remodelled that (1) lung repair processes are likely to have commenced, (2) manifestation of acute phase lung injury has ceased, and (3) the saccular and early-alveolar stage lungs are likely to undergo repair by similar mechanisms. These findings demonstrate that the immature lung does have the capacity to repair in the absence of continued MV. Furthermore I have identified possible mechanisms of repair in the developing lung, which are worthy of further investigation

Mechanical ventilation of the very immature lung: mechanisms of injury and repair

Very preterm infants often require mechanical ventilation (MV) due to respiratory insufficiency. However, respiratory support that includes MV can injure the immature lung and contribute to the development of bronchopulmonary dysplasia (BPD). Structural changes in the lungs that are associated with MV and BPD can persist, resulting in deficits in lung function in children and young adults. Multiple factors are known to contribute to the development of BPD including MV, the use of supplemental oxygen, infection and impaired nutrition, so it has been difficult to determine the effects of MV alone. Using our unique model of MV-induced lung injury in the fetal sheep we are able to investigate the effects of MV alone on the immature lung. To determine if MV-induced injury in the very immature lung persists or resolves in the absence of further ventilation, we exposed very immature fetal sheep to a brief period (2h) of MV and examined the lungs 24h and 15d later. These studies were conducted in sheep in both the saccular and early-alveolar stages of lung development. Knowledge about the susceptibility to MV-induced injury at different stages of lung development will provide important information regarding the factors contributing to BPD in preterm infants. At both stages of lung development brief, injurious MV caused severe lung injury at 24h. Lungs displayed reduced secondary septal crest density, atelectasis, disorganised extracellular matrix deposition and hemorrhage. Bronchioles had thickened, injured epithelium and often contained luminal debris. MV caused differences in lung injury manifestation between saccular and early-alveolar stage lungs. Fifteen days after MV was performed saccular and early-alveolar stage lungs spontaneously repaired and displayed no signs of injury or remodelling. In order to determine if injury and/ or repair mechanisms are active in our model of MV-induced injury 24h after brief MV mRNA and protein expression of cytokines, early response genes and a subset of putative repair genes were evaluated. Cytokine and early response gene mRNA levels were not elevated and the protein deposition of early response genes was also not increased. The mRNA expression levels of two putative repair genes, metallothionein and urokinase plasminogen activator receptor, were significantly increased. Protein deposition of metallothionien was also increased in ventilated lungs. This study has shown for the first time that 24h after MV of the very immature lung, while lungs remain severely injured and remodelled that (1) lung repair processes are likely to have commenced, (2) manifestation of acute phase lung injury has ceased, and (3) the saccular and early-alveolar stage lungs are likely to undergo repair by similar mechanisms. These findings demonstrate that the immature lung does have the capacity to repair in the absence of continued MV. Furthermore I have identified possible mechanisms of repair in the developing lung, which are worthy of further investigation

Mechanical ventilation injury and repair in extremely and very preterm lungs

BACKGROUND: Extremely preterm infants often receive mechanical ventilation (MV), which can contribute to bronchopulmonary dysplasia (BPD). However, the effects of MV alone on the extremely preterm lung and the lung’s capacity for repair are poorly understood. AIM: To characterise lung injury induced by MV alone, and mechanisms of injury and repair, in extremely preterm lungs and to compare them with very preterm lungs. METHODS: Extremely preterm lambs (0.75 of term) were transiently exposed by hysterotomy and underwent 2 h of injurious MV. Lungs were collected 24 h and at 15 d after MV. Immunohistochemistry and morphometry were used to characterise injury and repair processes. qRT-PCR was performed on extremely and very preterm (0.85 of term) lungs 24 h after MV to assess molecular injury and repair responses. RESULTS: 24 h after MV at 0.75 of term, lung parenchyma and bronchioles were severely injured; tissue space and myofibroblast density were increased, collagen and elastin fibres were deformed and secondary crest density was reduced. Bronchioles contained debris and their epithelium was injured and thickened. 24 h after MV at 0.75 and 0.85 of term, mRNA expression of potential mediators of lung repair were significantly increased. By 15 days after MV, most lung injury had resolved without treatment. CONCLUSIONS: Extremely immature lungs, particularly bronchioles, are severely injured by 2 h of MV. In the absence of continued ventilation these injured lungs are capable of repair. At 24 h after MV, genes associated with injurious MV are unaltered, while potential repair genes are activated in both extremely and very preterm lungs

Mechanical ventilation injury and repair in extremely and very preterm lungs.

BACKGROUND: Extremely preterm infants often receive mechanical ventilation (MV), which can contribute to bronchopulmonary dysplasia (BPD). However, the effects of MV alone on the extremely preterm lung and the lung's capacity for repair are poorly understood. AIM: To characterise lung injury induced by MV alone, and mechanisms of injury and repair, in extremely preterm lungs and to compare them with very preterm lungs. METHODS: Extremely preterm lambs (0.75 of term) were transiently exposed by hysterotomy and underwent 2 h of injurious MV. Lungs were collected 24 h and at 15 d after MV. Immunohistochemistry and morphometry were used to characterise injury and repair processes. qRT-PCR was performed on extremely and very preterm (0.85 of term) lungs 24 h after MV to assess molecular injury and repair responses. RESULTS: 24 h after MV at 0.75 of term, lung parenchyma and bronchioles were severely injured; tissue space and myofibroblast density were increased, collagen and elastin fibres were deformed and secondary crest density was reduced. Bronchioles contained debris and their epithelium was injured and thickened. 24 h after MV at 0.75 and 0.85 of term, mRNA expression of potential mediators of lung repair were significantly increased. By 15 days after MV, most lung injury had resolved without treatment. CONCLUSIONS: Extremely immature lungs, particularly bronchioles, are severely injured by 2 h of MV. In the absence of continued ventilation these injured lungs are capable of repair. At 24 h after MV, genes associated with injurious MV are unaltered, while potential repair genes are activated in both extremely and very preterm lungs

Morphological and injury analysis of bronchioles in saccular stage MV and control lungs after 24 h and 15 d.

<p>Light micrographs show cellular intraluminal debris in MV110+24 h lungs (A, B), intact epithelium of C110+24 h bronchiole (C) and bronchiole with denuded epithelium in MV110+24 h lung (D). The basement membrane perimeter of bronchioles was not different between all saccular stage MV and control lungs (E). Epithelial thickness of bronchioles was greater in MV110+24 h fetuses than in controls; however, it was lower in MV110+15 days fetuses relative to 15 days controls (F). The proportion of bronchioles that contained debris within the lumen was increased in MV110+24 h and MV110+15 d fetuses compared with age-matched controls (G). Values that do not share a common letter are significantly different from each other (P&lt;0.05, scale bar = 10 µm in A and B and 20 µm in C and D).</p

Injury analysis of saccular stage control and ventilated bronchioles after 24 h and 15 d.

<p>Mild injury: 45° bronchiole epithelium detached or absent; moderate: 45°–180° bronchiole epithelium detached or absent; severe: 180° bronchiole epithelium detached or absent. Injured data represent total no. of mild, moderate, and severely injured bronchioles. MV110+24 h lungs had a higher proportion of injured bronchioles relative to all other groups, of which most were classified as severely injured (p&lt;0.05).</p

Relative expression of potential repair gene mRNA in saccular and early alveolar stage lungs 24 h after MV.

<p>Metallothionein (A) and Urokinase Plasminogen Activator Receptor (B) mRNA expression was significantly increased in MV saccular and early alveolar stage lungs after 24 h when compared to controls (p&lt;0.05). Relative expression of Delta-Like Homolog Drosophila (C) and Heat Shock 10 kDa Protein (D) mRNA was not different between control and MV lungs at 24 h in the saccular or early-alveolar stage lung.</p

Lung morphometry, collagen and elastin density, percent tissue space and secondary septal crest density in saccular stage MV and control lungs after 24 h and 15 d.

<p>Light micrographs stained with hemotoxylin and eosin depicting lung morphology in C110+24 h (A), MV110+24 h (B), C110+15 d (C) and MV110+15 d (D) lung tissue. At 24 h after MV lung tissue showed signs of heterogeneous injury with regional hypercellularity and atelectasis (arrow, B). Tissue space fraction was increased in MV110+24 h lungs compared to controls (M, p&lt;0.05). Collagen fibres (black staining) are shown in C110 d+24 h (E), MV110+24 h (F), C110+15 d (G) and MV110+15 d (H) and elastin deposits (brown staining) in C110+24 h (I), MV110+24 h (J), C110+15 d (K) and MV110+15 d (L). Collagen fibres (brown staining) were not straight in MV110+24 h lungs (arrow, F), compared to controls at both ages and MV110+15 lungs (E,G,H). Collagen (N) and elastin density (O) was not different between MV lungs and their matched control group. Secondary septal crest density was reduced in MV110+24 h lungs (arrow, J) compared to those in C110+24 h group (I, P). Scale bar = 100 µm for A–D and 20 µm for E–L. Values that do not share a common letter are significantly different.</p