Obesity gives rise to metabolic complications by mechanisms that are poorly understood. Although chronic inflammatory signaling in adipose tissue is typically associated with metabolic deficiencies ...linked to excessive weight gain, we identified a subset of neuropilin-1 (NRP1)-expressing myeloid cells that accumulate in adipose tissue and protect against obesity and metabolic syndrome. Ablation of NRP1 in macrophages compromised lipid uptake in these cells, which reduced substrates for fatty acid β-oxidation and shifted energy metabolism of these macrophages toward a more inflammatory glycolytic metabolism. Conditional deletion of NRP1 in LysM Cre-expressing cells leads to inadequate adipose vascularization, accelerated weight gain, and reduced insulin sensitivity even independent of weight gain. Transfer of NRP1
hematopoietic cells improved glucose homeostasis, resulting in the reversal of a prediabetic phenotype. Our findings suggest a pivotal role for adipose tissue-resident NRP1
-expressing macrophages in driving healthy weight gain and maintaining glucose tolerance.
Glaucoma is the leading cause of irreversible blindness worldwide. At present, there is no cure for glaucoma, and current therapies are often inadequate. Loss of vision in glaucoma results from the ...death of retinal ganglion cells, the neurons that send visual information from the retina to the brain. The principal mechanism leading to retinal ganglion cell damage during glaucoma is not well understood, however, putative culprits have been proposed including excitotoxicity, neurotrophin deprivation, mechanical compression, ischemia, reactive astrocytes and oxidative stress. It is well established that retinal ganglion cell loss during glaucoma is caused by apoptotic programmed cell death, however, the precise mechanisms that lead to apoptotic death of adult retinal ganglion cells are poorly defined. To address this point, I put forth the central hypothesis that the identification of signaling pathways involved in apoptotic retinal ganglion cell death would offer therapeutic avenues to slow or prevent retinal ganglion cell death during ocular neuropathies such as glaucoma. In the first part of my thesis, I characterised the role of Apoptosis Stimulating Protein of p53 family (ASPP) proteins, which are regulators of p53, in the apoptotic death of retinal ganglion cells. p53 is a nuclear transcription factor implicated in cellular functions ranging from transcription to apoptosis. ASPP family members ASPP1, ASPP2 and iASPP are p53 binding proteins that belong to a family of protein regulators of p53-dependent apoptotic death. However, the role of ASPP family members in retinal ganglion cell death is unknown. As ASPP1 and ASPP2 are pro-apoptotic, the hypothesis of our first study was that the knockdown of ASPP1 and ASPP2 gene expression would lead to retinal ganglion cell survival after an optic nerve lesion. We used a well-characterized experimental model of neuronal apoptosis induced by optic nerve axotomy in Sprague Dawley rats. The results of this study (Wilson et al. Journal of Neuroscience, 2013) demonstrated that p53 is implicated in retinal ganglion cell death, and that targeted knockdown of ASPP1 and ASPP2 by short interference ribonucleic acid promotes retinal ganglion cell survival. The knockdown of ASPP2 correlates with a reduction in the levels of pro-apoptotic p53 regulated targets PUMA and Fas/CD95. In the second part of my thesis, I characterized the role of the anti-apoptotic member of the ASPP family, iASPP, in the apoptotic death of retinal ganglion cells. The hypothesis of this second study is that the overexpression of iASPP would promote retinal ganglion cell survival after axotomy. The data (Wilson et al. PLoS ONE, 2014) demonstrate that the targeted knockdown of iASPP by short interference ribonucleic acid exacerbates retinal ganglion cell death, and that the overexpression of iASPP by adeno-associated virus promotes retinal ganglion cell survival. The overexpression of iASPP correlates with a reduction in protein levels of PUMA and Fas/CD95. In conclusion, the findings presented in this thesis contribute to a better understanding of the pathological mechanisms underlying retinal ganglion cell loss by apoptosis and might provide insights into the design of novel neuroprotective treatments for neurodegenerative diseases such as glaucoma. Key words: retinal ganglion cell, neuronal death, apoptosis-stimulating protein of p53, axotomy.
This study describes the initial contact of 255 potential participants, recruitment of 63 participants, and retention of 57 participants from three geriatric Hoarding Disorder (HD) studies. Patients ...with HD were easily recruited from the community, primarily through the use of clinician referrals and posted flyers, as evidenced by steady patient flow despite lack of compensation for participation. Contrary to treatment outcomes of late life mood and anxiety disorders, geriatric HD patients are largely retained in clinical research treatment studies. Results demonstrate that older adults with HD can be engaged in treatment. Participants often needed the study staff to provide substantial support, including informal motivational interviewing and problem solving, in scheduling and attending the initial visit.
Abstract
Although they have distinct functions, the signaling of dopamine‐D
2
receptor short and long isoforms (D
2
S and D
2
L) is virtually identical. We compared inhibitory regulation of ...extracellular signal‐regulated kinases (ERK1/2) in GH4 pituitary cells separately transfected with these isoforms. Activation of rat or human dopamine‐D
2
S, muscarinic or somatostatin receptors inhibited thyrotropin‐releasing hormone‐induced ERK1/2 phosphorylation, while the D
2
L receptor failed to inhibit this response. In order to address the structural basis for the differential signaling of D
2
S and D
2
L receptors, we examined the D
2
L‐SS mutant, in which a protein kinase C (PKC) pseudosubstrate site that is present in the D
2
L but not D
2
S receptor was converted to a consensus PKC site. In transfected GH4 cells, the D
2
L‐SS mutant inhibited thyrotropin‐releasing hormone‐induced ERK1/2 phosphorylation almost as strongly as the D
2
S receptor. A D
2
S‐triple mutant that eliminates PKC sites involved in D
2
S receptor desensitization also inhibited ERK1/2 activation. Similarly, in striatal cultures, the D
2
‐selective agonist quinpirole inhibited potassium‐stimulated ERK1/2 phosphorylation, indicating the presence of this pathway in neurons. In conclusion, the D
2
S and D
2
L receptors differ in inhibitory signaling to ERK1/2 due to specific residues in the D
2
L receptor alternatively spliced domain, which may account for differences in their function
in vivo
.
Although they have distinct functions, the signaling of dopamine-D sub(2) receptor short and long isoforms (D sub(2)S and D sub(2)L) is virtually identical. We compared inhibitory regulation of ...extracellular signal-regulated kinases (ERK12) in GH4 pituitary cells separately transfected with these isoforms. Activation of rat or human dopamine-D sub(2)S, muscarinic or somatostatin receptors inhibited thyrotropin-releasing hormone-induced ERK12 phosphorylation, while the D sub(2)L receptor failed to inhibit this response. In order to address the structural basis for the differential signaling of D sub(2)S and D sub(2)L receptors, we examined the D sub(2)L-SS mutant, in which a protein kinase C (PKC) pseudosubstrate site that is present in the D sub(2)L but not D sub(2)S receptor was converted to a consensus PKC site. In transfected GH4 cells, the D sub(2)L-SS mutant inhibited thyrotropin-releasing hormone-induced ERK12 phosphorylation almost as strongly as the D sub(2)S receptor. A D sub(2)S-triple mutant that eliminates PKC sites involved in D sub(2)S receptor desensitization also inhibited ERK12 activation. Similarly, in striatal cultures, the D sub(2)-selective agonist quinpirole inhibited potassium-stimulated ERK12 phosphorylation, indicating the presence of this pathway in neurons. In conclusion, the D sub(2)S and D sub(2)L receptors differ in inhibitory signaling to ERK12 due to specific residues in the D sub(2)L receptor alternatively spliced domain, which may account for differences in their function in vivo.
Although they have distinct functions, the signaling of dopamine-D sub(2) receptor short and long isoforms (D sub(2)S and D sub(2)L) is virtually identical. We compared inhibitory regulation of ...extracellular signal-regulated kinases (ERK1/2) in GH4 pituitary cells separately transfected with these isoforms. Activation of rat or human dopamine-D sub(2)S, muscarinic or somatostatin receptors inhibited thyrotropin-releasing hormone-induced ERK1/2 phosphorylation, while the D sub(2)L receptor failed to inhibit this response. In order to address the structural basis for the differential signaling of D sub(2)S and D sub(2)L receptors, we examined the D sub(2)L-SS mutant, in which a protein kinase C (PKC) pseudosubstrate site that is present in the D sub(2)L but not D sub(2)S receptor was converted to a consensus PKC site. In transfected GH4 cells, the D sub(2)L-SS mutant inhibited thyrotropin-releasing hormone-induced ERK1/2 phosphorylation almost as strongly as the D sub(2)S receptor. A D sub(2)S-triple mutant that eliminates PKC sites involved in D sub(2)S receptor desensitization also inhibited ERK1/2 activation. Similarly, in striatal cultures, the D sub(2)-selective agonist quinpirole inhibited potassium-stimulated ERK1/2 phosphorylation, indicating the presence of this pathway in neurons. In conclusion, the D sub(2)S and D sub(2)L receptors differ in inhibitory signaling to ERK1/2 due to specific residues in the D sub(2)L receptor alternatively spliced domain, which may account for differences in their function in vivo.
To address their role in 5‐HT1A receptor–G protein coupling, over sixty random point mutant 5‐HT1A receptors in the second intracellular loop C‐terminal domain (Ci2) (143DYVNKRTPRR152) were generated ...and most retained agonist binding. Mutants were tested for Gβγ signaling to ACII or PLC, and Gαi coupling. Most Ci2 mutations blocked 5‐HT1A signaling to Gβγ, but several also affected Gαi signaling. Ci2 residues K147, R148/151/152 and P150 were essential for coupling, consistent with an obligatory structural role. Polar residues (T149, N146) were required for Gβγ but not Gαi coupling, suggesting a selective interface with Gβγ. The Y144 residue directed specificity for both Gβγ and Gαi pathways and is predicted to form hydrogen bonds with D133/R134 (Ni2 DRY motif) and E340 (Ci3) to stabilize the G‐protein coupling domain. Consistent with a primary role of Ci3 in Gαi coupling, E340 mutants preferentially abrogated Gαi signaling, but one mutant partly blocked Gβγ signaling as well. Thus, the 5‐HT1A receptor Ci2 domain determines Gβγ specificity and stabilizes Gαi‐mediated signaling, while the Ci3 domain primarily directs Gαi coupling and stabilizes Gβγ coupling.
Supported by CIHR.
Millimeter-wave imaging has gathered attention in recent years for its ability to penetrate clothing, thin layers of soils, and certain construction materials. However, image quality remains a ...challenge that needs to be addressed. One way of improving image quality is by increasing the dimensions of the collecting aperture. A sparse array can be used to synthesize a larger aperture with a limited set of relatively small detectors. In this research we design, build, and test a test-bed having an active source at 94 GHz and an array of coherent detectors, mounted on arms that extend radially on a rotary table. Using this test bed a circular area with a maximum diameter of 900 mm can be scanned. The signal is down-converted using heterodyne receivers with digital in-phase and quadrature detection. Signal correlation is performed using the digitized data, which is stored for post-processing, electronic focusing, and image reconstruction. Near-field imaging using interferometric reconstructions is achieved using electronic focusing. Imaging tests show the ability of the system to generate imagery of concealed and unconcealed objects at distances between 400 and 700 mm. A study of the effects of redundant and nonredundant configurations on image quality for 4 common detector configurations is presented. In this document we show that an active sparse-aperture imaging system using digital correlators is a viable way to generate millimeter-wave images.
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