Loss of the Hematopoietic Stem Cell Factor GATA2 in the Osteogenic Lineage Impairs Trabecularization and Mechanical Strength of Bone

The transcription factor GATA2 is required for expansion and differentiation of hematopoietic stem cells (HSCs). In mesenchymal stem cells (MSCs), GATA2 blocks adipogenesis, but its biological relevance and underlying genomic events are unknown. We report a dual function of GATA2 in bone homeostasis. GATA2 in MSCs binds near genes involved in skeletal system development and colocalizes with motifs for FOX and HOX transcription factors, known regulators of skeletal development. Ectopic GATA2 blocks osteoblastogenesis by interfering with SMAD1/5/8 activation. MSC-specific deletion of GATA2 in mice increases the numbers and differentiation capacity of bone-derived precursors, resulting in elevated bone formation. Surprisingly, MSC-specific GATA2 deficiency impairs the trabecularization and mechanical strength of bone, involving reduced MSC expression of the osteoclast inhibitor osteoprotegerin and increased osteoclast numbers. Thus, GATA2 affects bone turnover via MSC-autonomous and indirect effects. By regulating bone trabecularization, GATA2 expression in the osteogenic lineage may contribute to the anatomical and cellular microenvironment of the HSC niche required for hematopoiesis.Peer reviewe

Tubular von Hippel-Lindau knockout protects against rhabdomyolysis-induced AKI

In akuten Nierenversagen (AKI) unterschiedlicher Genese finden sich Belege für eine regionale Hypoxie der Niere. In allen bisher untersuchten AKI-Modellen fand sich jedoch nur eine geringe und zeitlich begrenzte Hypoxieanpassung durch Hypoxie-induzierbare Transkriptionsfaktoren (HIF). Nephronsegmente mit dem höchsten Maße an morphologischen Zellschäden besitzen die geringste Fähigkeit zur HIF-Aktivierung. Präkonditionelle HIF-Aktivierung schützt vor AKI, dass mittels Klemmen der Nierenarterie oder des Nierenstiels erzeugt wird. Die Mechanismen dieser HIF-vermittelten Protektion sind weitgehend unbekannt. Ebenso bleibt unklar, ob solche Mechanismen auch in anderen AKI- Formen wirksam sein können. Die vorliegende Arbeit zeigt, dass eine selektive Hochregulation von HIF in Nierentubuli durch einen Pax8-rtTA-abhängigen und induzierbaren Knockout des von Hippel-Lindau Proteins (VHL) vor Rhabdomyolyse- induziertem AKI schützt. In diesem AKI-Modell ist die HIF-Aktivierung umgekehrt proportional dem tubulären Zellschaden, der in proximalen Tubuli zu 5% Nekrosen am Tag 1 und 40% Nekrosen am Tag 2 führt. Eine genomweite Microarray-Analyse zeigt, dass in diesem AKI-Modell etwa 20% des aktiven Genoms signifikant (p<0,01) reguliert ist. Rhabdomyolyse führt in VHL-KO- Mäusen in Vergleich zu Tieren ohne VHL-KO zu einer starken und anhaltenden HIF-Aktivierung in proximalen Tubuli. In den ersten 48h nach Auslösung der Rhabdomyolyse zeigt sich in VHL-KO-Tieren eine Nephroprotektion anhand niedrigerer Werte für Plasmakreatinin/Plasmaharnstoff, Caspase-3-Protein und Tubulusnekrose. Am Tag 1 nach Auslösung der Rhabdomyolyse ist der tubuläre Schaden größtenteils sublethal und daher potentiell reversibel. Zu diesem Zeitpunkt finden wir in VHL-KO-Tieren Hinweise für aktivierte Glykolyse und Glukoseaufnahme, Autophagie, Gefäßerweiterung und Protonenelimination, wie anhand von qPCR, Pathway-Enrichment-Analyse und Immunhistochemie ersichtlich. Zusammenfassend unterstützen unsere Daten die Ansicht, dass die HIF-bedingte AKI-Protektion auf einer Umstellung des Metabolismus hin zu einer anaeroben Energiegewinnung gründet.Regional renal hypoxia is a common feature of acute kidney injury (AKI) of different causes. However, in all AKI models tested so far, adaptation to hypoxia through hypoxia-inducible transcription factors (HIF) is very limited. Moreover, renal cell types most prone to injury seem to have the least capacity of up-regulating HIF under the respective AKI condition. Pre- conditional HIF activation has been demonstrated to protect from AKI induced by ischemia-reperfusion injury. But, the mechanisms responsible for renal protection are largely unknowns. Equally, it remains unclear if HIF based rescue mechanisms can be effective in other AKI forms, as well. The present work shows that selective HIF activation in renal tubules through Pax8-rtTA based inducible knockout of von Hippel-Lindau protein (VHL) protects from rhabdomyolysis-induced AKI. In this model, HIF activation inversely correlates with tubular injury, leading to 5% necrosis at d1 and 40% necrosis at d2 compared with controls. Genome wide microarray analysis reveals that roughly 20% of the active genome is significantly (p<0.01) regulated in this AKI form. Compared with control animals, VHL-KO mice subjected to rhabdomyolysis substantially activate HIF in portions of the nephron at risk for acute injury. In the first 48 h after induction of rhabdomyolysis, in VHL-KO mice kidney protection is reflected by lower plasma creatinine/urea, caspase-3 protein, and tubular necrosis. At d1 after rhabdomyolysis tubular injury is mostly sublethal, and hence, potentially reversible. We provide evidence that at this time point VHL-KO leads to activated glycolysis, enhanced cellular glucose uptake and utilization, autophagy, vasodilation, and proton removal, as demonstrated by qPCR, pathway enrichment analysis and immunohistochemistry. In conclusion, our data support the view that HIF based protection from AKI relies on a shift of metabolism towards anaerobic energy supply

Changes in water column and sedimentary environments at the onset of the Messinian salinity crisis: Insights from an onshore deep-water section (Govone, NW Italy)

The data set was used to trace the impact of the advent of the late Miocene Messinian salinity crisis (MSC) 5.97 Ma ago in the northern Mediterranean (Piedmont Basin, NW Italy) on the water column and sedimentary environments. We analysed shale/marl couplets deposited from about 6.07 to 5.92 Ma in distal areas of the Piedmont Basin (Govone section; 44°48′08″N; 8°07′34″E). The analyses included X-ray diffraction (XRD), bulk-rock carbon (δ13C) and oxygen (δ18O) stable isotopes, total organic (TOC) and inorganic (TIC) carbon, and lipid biomarkers. XRD analyses aimed to investigate carbonate mineralogy, revealing the occurrence of calcite and dolomite. Dolomite represents the only carbonate phase after the MSC onset, with an overall increase indicated by TIC investigations. Stable isotope ratios suggested the involvement of microbial activity in the precipitation of dolomite (δ13C) and helped the reconstruction of the physical characteristic of the fluids from which the carbonate phase precipitated (δ18O). Lipid biomarker analyses focused on the acyclic terpenoid lycopane, the cyclic terpenoid tetrahymanol, the isoprenoid diphytanyl glycerol diether C20-20 archaeol, isoprenoid glycerol dibiphytanyl glycerol tetraethers (GDGTs), and the carotenoid isorenieratane. The lipids provided new information on the archaeal community thriving in the distal areas of the Piedmont Basin and allowed the reconstruction of physical and chemical properties of the water column (stratification, photic zone euxinia, redox conditions). Ultimately, the data set revealed how physicochemical properties of the water column controlled the stratigraphic architecture of the sediments deposited during the first stage of the MSC

Deeply conserved chordate noncoding sequences preserve genome synteny but do not drive gene duplicate retention

Animal genomes possess highly conserved cis-regulatory sequences that are often found near genes that regulate transcription and development. Researchers have proposed that the strong conservation of these sequences may affect the evolution of the surrounding genome, both by repressing rearrangement, and possibly by promoting duplicate gene retention. Conflicting data, however, have made the validity of these propositions unclear. Here, we use a new computational method to identify phylogenetically conserved noncoding elements (PCNEs) in a manner that is not biased by rearrangement and duplication. This method is powerful enough to identify more than a thousand PCNEs that have been conserved between vertebrates and the basal chordate amphioxus. We test 42 of our PCNEs in transgenic zebrafish assays—including examples from vertebrates and amphioxus—and find that the majority are functional enhancers. We find that PCNEs are enriched around genes with ancient synteny conservation, and that this association is strongest for extragenic PCNEs, suggesting that cis-regulatory interdigitation plays a key role in repressing genome rearrangement. Next, we classify mouse and zebrafish genes according to association with PCNEs, synteny conservation, duplication history, and presence in bidirectional promoter pairs, and use these data to cluster gene functions into a series of distinct evolutionary patterns. These results demonstrate that subfunctionalization of conserved cis-regulation has not been the primary determinate of gene duplicate retention in vertebrates. Instead, the data support the gene balance hypothesis, which proposes that duplicate retention has been driven by selection against dosage imbalances in genes with many protein connections

Pathophysiology of Unilateral Ischemia-Reperfusion Injury: Importance of Renal Counterbalance and Implications for the AKI-CKD Transition

Unilateral ischemia-reperfusion (UIR) injury leads to progressive renal atrophy and tubulointerstitial fibrosis (TIF) and is commonly used to investigate the pathogenesis of the acute kidney injury-chronic kidney disease transition. Although it is well known that contralateral nephrectomy (CNX), even 2 wk post-UIR injury, can improve recovery, the physiological mechanisms and tubular signaling pathways mediating such improved recovery remain poorly defined. Here, we examined the renal hemodynamic and tubular signaling pathways associated with UIR injury and its reversal by CNX. Male Sprague-Dawley rats underwent left UIR or sham UIR and 2 wk later CNX or sham CNX. Blood pressure, left renal blood flow (RBF), and total glomerular filtration rate were assessed in conscious rats for 3 days before and over 2 wk after CNX or sham CNX. In the presence of a contralateral uninjured kidney, left RBF was lower (P \u3c 0.05) from 2 to 4 wk following UIR (3.6 + 0.3 mL/min) versus sham UIR (9.6 + 0.3 mL/min). Without CNX, extensive renal atrophy, TIF, and tubule dedifferentiation, but minimal pimonidazole and hypoxia-inducible factor-1α positivity in tubules, were present at 4 wk post-UIR injury. Conversely, CNX led (P \u3c 0.05) to sustained increases in left RBF (6.2 ∓ 0.6 mL/min) that preceded the increases in glomerular filtration rate. The CNX-induced improvement in renal function was associated with renal hypertrophy, more redifferentiated tubules, less TIF, and robust pimonidazole and hypoxia-inducible factor-1α staining in UIR injured kidneys. Thus, contrary to expectations, indexes of hypoxia are not observed with the extensive TIF at 4 wk post-UIR injury in the absence of CNX but are rather associated with the improved recovery of renal function and structure following CNX