Initiation of cyst formation in autosomal dominant polycystic kidney disease (ADPKD) occurs when kidney tubule cells are rendered null for either PKD1 or PKD2 by somatic 'second hit' mutations. ...Subsequent cyst progression remodels the organ through changes in tubule cell shape, proliferation and secretion. The kidney develops inflammation and fibrosis. We constructed a mouse model in which adult inactivation of either Pkd gene can be followed by reactivation of the gene at a later time. Using this model, we show that re-expression of Pkd genes in cystic kidneys results in rapid reversal of ADPKD. Cyst cell proliferation is reduced, autophagy is activated and cystic tubules with expanded lumina lined by squamoid cells revert to normal lumina lined by cuboidal cells. Increases in inflammation, extracellular matrix deposition and myofibroblast activation are reversed, and the kidneys become smaller. We conclude that phenotypic features of ADPKD are reversible and that the kidney has an unexpected capacity for plasticity controlled at least in part by ADPKD gene function.
The coumarin-hemicyanine hybrid is an internal charge transfer (ICT) dye that features a donor-π-acceptor motif and displays two well-resolved absorption bands at ∼350 nm for the donor coumarin and ...∼550 nm for the fully-conjugated dye. Herein, we demonstrate that preferential excitation of the dye at 350 nm, ascribed to a twisted conformation, gives rise to two well-resolved emission bands at ∼450 nm for the coumarin and ∼650 nm for the planar dye that are highly quenched in water. Unexpectedly, the coumarin blue emission was found to 'turn-on' to aprotic and viscous solvents and respond to binding bovine serum albumin (BSA) in aqueous media. In contrast, the red ICT emission was sensitive to protic solvent polarity and responded to binding the G-quadruplex (GQ) DNA structure produced by the heme binding aptamer (PS2.M). Our studies also demonstrate the influence of the alkyl group attached to the dye quaternary nitrogen atom in determining the microenvironment sensitivity and ability to differentiate biopolymer types.
There are no clinically relevant treatments available that improve function in the growing population of very preterm infants (less than 32 weeks' gestation) with neonatal brain injury. Diffuse white ...matter injury (DWMI) is a common finding in these children and results in chronic neurodevelopmental impairments. As shown recently, failure in oligodendrocyte progenitor cell maturation contributes to DWMI. We demonstrated previously that the epidermal growth factor receptor (EGFR) has an important role in oligodendrocyte development. Here we examine whether enhanced EGFR signalling stimulates the endogenous response of EGFR-expressing progenitor cells during a critical period after brain injury, and promotes cellular and behavioural recovery in the developing brain. Using an established mouse model of very preterm brain injury, we demonstrate that selective overexpression of human EGFR in oligodendrocyte lineage cells or the administration of intranasal heparin-binding EGF immediately after injury decreases oligodendroglia death, enhances generation of new oligodendrocytes from progenitor cells and promotes functional recovery. Furthermore, these interventions diminish ultrastructural abnormalities and alleviate behavioural deficits on white-matter-specific paradigms. Inhibition of EGFR signalling with a molecularly targeted agent used for cancer therapy demonstrates that EGFR activation is an important contributor to oligodendrocyte regeneration and functional recovery after DWMI. Thus, our study provides direct evidence that targeting EGFR in oligodendrocyte progenitor cells at a specific time after injury is clinically feasible and potentially applicable to the treatment of premature children with white matter injury.
Macrophages are a source of both proinflammatory and restorative functions in damaged tissue through complex dynamic phenotypic changes. Here, we sought to determine whether monocyte-derived ...macrophages (MDMs) contribute to recovery after acute sterile brain injury. By profiling the transcriptional dynamics of MDMs in the murine brain after experimental intracerebral hemorrhage (ICH), we found robust phenotypic changes in the infiltrating MDMs over time and demonstrated that MDMs are essential for optimal hematoma clearance and neurological recovery. Next, we identified the mechanism by which the engulfment of erythrocytes with exposed phosphatidylserine directly modulated the phenotype of both murine and human MDMs. In mice, loss of receptor tyrosine kinases AXL and MERTK reduced efferocytosis of eryptotic erythrocytes and hematoma clearance, worsened neurological recovery, exacerbated iron deposition, and decreased alternative activation of macrophages after ICH. Patients with higher circulating soluble AXL had poor 1-year outcomes after ICH onset, suggesting that therapeutically augmenting efferocytosis may improve functional outcomes by both reducing tissue injury and promoting the development of reparative macrophage responses. Thus, our results identify the efferocytosis of eryptotic erythrocytes through AXL/MERTK as a critical mechanism modulating macrophage phenotype and contributing to recovery from ICH.
Gliomas maintain an acidic extracellular pH (pH
), which promotes tumor growth and builds resistance to therapy. Given evidence that acidic pH
beyond the tumor core indicates infiltration, we ...hypothesized that imaging the intratumoral pH
in relation to the peritumoral pH
can provide a novel readout of therapeutic influence on the tumor microenvironment. We used Biosensor Imaging of Redundant Deviation in Shifts (BIRDS), which utilizes chemical shifts of non-exchangeable protons from macrocyclic chelates (e.g., DOTP
) complexed with paramagnetic thulium (Tm
), to generate pH
maps in rat brains bearing U251 tumors. Following TmDOTP
infusion, T
-weighted MRI provided delineation of the tumor boundary and BIRDS was used to image the pH
gradient between intratumoral and peritumoral regions (ΔpH
) in both untreated and temozolomide treated (40 mg/kg) rats bearing U251 tumors. Treated rats had reduced tumor volume (p < 0.01), reduced proliferation (Ki-67 staining; p < 0.03) and apoptosis induction (cleaved Caspase-3 staining; p < 0.001) when compared to untreated rats. The ΔpH
was significantly higher in untreated compared to treated rats (p < 0.002), suggesting that temozolomide, which induces apoptosis and hinders proliferation, also normalizes intratumoral pH
. Thus, BIRDS can be used to map the ΔpH
in gliomas and provide a physiological readout of the therapeutic response on the tumor microenvironment.
Neural tube defects are a common congenital anomaly involving incomplete closure of the spinal cord. Myelomeningocele (MMC) is a severe form in which there is complete exposure of neural tissue with ...a lack of skin, soft tissue, or bony covering to protect the spinal cord. The all-trans retinoic acid (ATRA) induced rat model of (MMC) is a reproducible, cost-effective means of studying this disease; however, there are limited modalities to objectively quantify disease severity, or potential benefits from experimental therapies. We sought to determine the feasibility of detecting differences between MMC and wild type (WT) rat fetuses using diffusion magnetic resonance imaging techniques (MRI). Rat dams were gavage-fed ATRA to produce MMC defects in fetuses, which were surgically delivered prior to term. Average diffusion coefficient (ADC) and fractional anisotropy (FA) maps were obtained for each fetus. Brain volumes and two anatomically defined brain length measurements (D1 and D2) were significantly decreased in MMC compared to WT. Mean ADC signal was significantly increased in MMC compared to WT, but no difference was found for FA signal. In summary, ADC and brain measurements were significantly different between WT and MMC rat fetuses. ADC could be a useful complementary imaging biomarker to current histopathologic analysis of MMC models, and potentially expedite therapeutic research for this disease.
Purpose
To demonstrate feasibility of developing a noninvasive extracellular pH (pHe) mapping method on a clinical MRI scanner for molecular imaging of liver cancer.
Methods
In vivo pHe mapping has ...been demonstrated on preclinical scanners (e.g., 9.4T, 11.7T) with Biosensor Imaging of Redundant Deviation in Shifts (BIRDS), where the pHe readout by 3D chemical shift imaging (CSI) depends on hyperfine shifts emanating from paramagnetic macrocyclic chelates like TmDOTP5− which upon extravasation from blood resides in the extracellular space. We implemented BIRDS‐based pHe mapping on a clinical 3T Siemens scanner, where typically diamagnetic 1H signals are detected using millisecond‐long radiofrequency (RF) pulses, and 1H shifts span over ±10 ppm with long transverse (T2, 102 ms) and longitudinal (T1, 103 ms) relaxation times. We modified this 3D‐CSI method for ultra‐fast acquisition with microsecond‐long RF pulses, because even at 3T the paramagnetic 1H shifts of TmDOTP5− have millisecond‐long T2 and T1 and ultra‐wide chemical shifts (±200 ppm) as previously observed in ultra‐high magnetic fields.
Results
We validated BIRDS‐based pH in vitro with a pH electrode. We measured pHe in a rabbit model for liver cancer using VX2 tumors, which are highly vascularized and hyperglycolytic. Compared to intratumoral pHe (6.8 ± 0.1; P < 10−9) and tumor's edge pHe (6.9 ± 0.1; P < 10−7), liver parenchyma pHe was significantly higher (7.2 ± 0.1). Tumor localization was confirmed with histopathological markers of necrosis (hematoxylin and eosin), glucose uptake (glucose transporter 1), and tissue acidosis (lysosome‐associated membrane protein 2).
Conclusion
This work demonstrates feasibility and potential clinical translatability of high‐resolution pHe mapping to monitor tumor aggressiveness and therapeutic outcome, all to improve personalized cancer treatment planning.
Under normal conditions, high sodium (Na
) in extracellular (Na
) and blood (Na
) compartments and low Na
in intracellular milieu (Na
) produce strong transmembrane (ΔNa
) and weak transendothelial ...(ΔNa
) gradients respectively, and these manifest the cell membrane potential (V
) as well as blood-brain barrier (BBB) integrity. We developed a sodium (
Na) magnetic resonance spectroscopic imaging (MRSI) method using an intravenously-administered paramagnetic polyanionic agent to measure ΔNa
and ΔNa
. In vitro
Na-MRSI established that the
Na signal is intensely shifted by the agent compared to other biological factors (e.g., pH and temperature). In vivo
Na-MRSI showed Na
remained unshifted and Na
was more shifted than Na
, and these together revealed weakened ΔNa
and enhanced ΔNa
in rat gliomas (vs. normal tissue). Compared to normal tissue, RG2 and U87 tumors maintained weakened ΔNa
(i.e., depolarized V
) implying an aggressive state for proliferation, whereas RG2 tumors displayed elevated ∆Na
suggesting altered BBB integrity. We anticipate that
Na-MRSI will allow biomedical explorations of perturbed Na
homeostasis in vivo.
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