Childrens Hospital Research Foundation

Cincinnati, OH, United States

Childrens Hospital Research Foundation

Cincinnati, OH, United States
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Li J.,Southern Medical University | Liu N.,Southern Medical University | Lu K.,Southern Medical University | Zhang L.,Southern Medical University | And 3 more authors.
Neuroscience Letters | Year: 2012

Repeated exposure to cocaine can induce persistent alterations in the brain's reward system, including increases in the number of dendrites and spine density on medium-sized spiny neurons (MSNs) in the nucleus accumbens (NAc). The structural remodeling of dendrites and spines in the NAc is thought to play a critical role in cocaine addiction. MSNs in the NAc can be classified by expression of either D1 or D2 dopamine receptors, which are localized to the direct and indirect pathway, respectively. It is unknown whether the dendritic changes induced by repeated cocaine treatment occur in MSNs of the direct or indirect pathway. Because the traditional Golgi-Cox impregnation of neurons precludes identifying particular subpopulations of MSNs, we performed dendritic morphology analysis after biocytin-labeling and Golgi-Cox impregnation. We found that the biocytin staining MSNs showed higher dendritic spine density and higher number of dendrites than that in Golgi impregnation group. In addition, we found that the increasing spine density induced by repeated cocaine treatment in female mice was higher than that in male mice. Next we used biocytin staining and dynorphin/D2 receptor colocalization to determine which cell type(s) displayed dendritic changes after repeated cocaine treatment. We found that cocaine-induced changes in dendritic parameters occurred in MSNs of both the direct (D1-expressing) and indirect (D2-expressing) pathways. © 2012 Elsevier Ireland Ltd.

Zhang S.,Childrens Hospital Research Foundation | Konstantinidis D.G.,Childrens Hospital Research Foundation | Yang J.-Q.,Childrens Hospital Research Foundation | Mizukawa B.,Childrens Hospital Research Foundation | And 5 more authors.
Journal of Immunology | Year: 2014

Thymocyte development is regulated by complex signaling pathways. How these signaling cascades are coordinated remains elusive. RhoA of the Rho family small GTPases plays an important role in actin cytoskeleton organization, cell adhesion, migration, proliferation, and survival. Nonetheless, the physiological function of RhoA in thymocyte development is not clear. By characterizing a conditional gene targeting mouse model bearing T cell deletion of RhoA, we show that RhoA critically regulates thymocyte development by coordinating multiple developmental events. RhoA gene disruption caused a strong developmental block at the pre-TCR checkpoint and during positive selection. Ablation of RhoA led to reduced DNA synthesis in CD4- CD8-, CD4+CD8-, and CD4-CD8+ thymocytes but not in CD4+CD8+ thymocytes. Instead, RhoA-deficient CD4+CD8+ thymocytes showed an impaired mitosis. Furthermore, we found that abrogation of RhoA led to an increased apoptosis in all thymocyte subpopulations. Importantly, we show that the increased apoptosis was resulted from reduced pre-TCR expression and increased production of reactive oxygen species (ROS), which may be because of an enhanced mitochondrial function, as manifested by increased oxidative phosphorylation, glycolysis, mitochondrial membrane potential, and mitochondrial biogenesis in RhoA-deficient thymocytes. Restoration of pre-TCR expression or treatment of RhoA-deficient mice with a ROS scavenger N-acetylcysteine partially restored thymocyte development. These results suggest that RhoA is required for thymocyte development and indicate, to our knowledge, for the first time that fine-tuning of ROS production by RhoA, through a delicate control of metabolic circuit, may contribute to thymopoiesis. Copyright © 2014 by The American Association of Immunologists, Inc.

Li J.,Southern Medical University | Gu J.,Southern Medical University | Wang B.,Southern Medical University | Xie M.,Southern Medical University | And 5 more authors.
Molecular Neurobiology | Year: 2015

Dopamine (DA) is an important regulator of neuronal plasticity in the prefrontal cortex (PFC) and plays a critical role in addiction-related neuroadaptation. The Rho GTPases, including Rac1, RhoA and Cdc42, are key regulators of actin cytoskeleton rearrangement that play important roles in dendritic morphogenesis. The goal of the current study was to use cultures of primary PFC neurons to gain a better understanding of the molecular mechanisms underlying DA-induced dendritic morphogenesis, a phenomenon that mimics the increase in DA synaptic transmission observed in the PFC of in vivo cocaine administration. We investigated the effects of repeated DA treatments on dendritic morphology changes in PFC neurons, and identified Rac1 and RhoA as downstream effectors of D1 receptors during the regulation of dendritic morphogenesis. Importantly, we found that D1 receptor-regulated Rac1 and RhoA have distinct roles in the regulation of dendritic morphogenesis after repeated DA treatments. Our data provide the first evidence that Rac1 and RhoA are effectors of D1 receptor signaling during dendritic morphogenesis and represent new signaling molecules involved in long-lasting neuroadaptation in the PFC. © 2014, Springer Science+Business Media New York.

Karunanayaka P.,Childrens Hospital Research Foundation | Schmithorst V.J.,Childrens Hospital Research Foundation | Vannest J.,Childrens Hospital Research Foundation | Szaflarski J.P.,University of Cincinnati | And 2 more authors.
NeuroImage | Year: 2010

Semantic language skills are an integral part of early childhood language development. The semantic association between verbs and nouns constitutes an important building block for the construction of sentences. In this large-scale functional magnetic resonance imaging (fMRI) study, involving 336 subjects between the ages of 5 and 18years, we investigated the neural correlates of covert verb generation in children. Using group independent component analysis (ICA), seven task-related components were identified including the mid-superior temporal gyrus, the most posterior aspect of the superior temporal gyrus, the parahippocampal gyrus, the inferior frontal gyrus, the angular gyrus, and medial aspect of the parietal lobule (precuneus/posterior cingulate). A highly left-lateralized component was found including the medial temporal gyrus, the frontal gyrus, the inferior frontal gyrus, and the angular gyrus. The associated independent component (IC) time courses were analyzed to investigate developmental changes in the neural elements supporting covert verb generation. Observed age effects may either reflect specific local neuroplastic changes in the neural substrates supporting language or a more global transformation of neuroplasticity in the developing brain. The results are analyzed and presented in the framework of two theoretical models for neurocognitive brain development. In this context, group ICA of fMRI data from our large sample of children aged 5-18years provides strong evidence in support of the regionally weighted model for cognitive neurodevelopment of language networks. © 2010 Elsevier Inc.

Raghu H.,Childrens Hospital Research Foundation | Cruz C.,Childrens Hospital Research Foundation | Rewerts C.L.,Childrens Hospital Research Foundation | Frederick M.D.,Childrens Hospital Research Foundation | And 5 more authors.
Blood | Year: 2015

Rheumatoid arthritis is a chronic inflammatory disease characterized by synovial hyperplasia, inflammatory cell infiltration, irreversible cartilage and bone destruction, and exuberant coagulation system activity within joint tissue. Here, we demonstrate that the coagulation transglutaminase, factor XIII (fXIII), drives arthritis pathogenesis by promoting local inflammatory and tissue degradative and remodeling events. All pathological features of collagen-induced arthritis (CIA) were significantly reduced in fXIII-deficient mice. However, the most striking difference in outcome was the preservation of cartilage and bone in fXIIIA-/- mice concurrent with reduced osteoclast numbers and activity. The local expression of osteoclast effectors receptor activator ofnuclear factor-κB ligand (RANKL) and tartrate resistant acid phosphatase were significantly diminished in CIA-challenged and even unchallenged fXIIIA-/- mice relative to wild-type animals, but were similar in wild-typeandfibrinogen-deficientmice. Impairedosteoclast formationin fXIIIA-/- mice was not due to an inherent deficiency of monocyte precursors, but it was linked to reduced RANKL-driven osteoclast formation. Furthermore, treatment ofmice with the pan-transglutaminase inhibitor cystamine resulted in significantly diminishedCIA pathology and local markers of osteoclastogenesis. Thus, eliminating fXIIIA limits inflammatory arthritis and protects from cartilage and bone destruction in part through mechanisms linked to reduced RANKL-mediated osteoclastogenesis. In summary, therapeutic strategies targeting fXIII activity may prove beneficial in limiting arthropathies and other degenerative bone diseases. © 2015 by The American Society of Hematology.

Shang X.,Childrens Hospital Research Foundation | Marchioni F.,Childrens Hospital Research Foundation | Sipes N.,Childrens Hospital Research Foundation | Evelyn C.R.,Childrens Hospital Research Foundation | And 5 more authors.
Chemistry and Biology | Year: 2012

Rho GTPases have been implicated in diverse cellular functions and are potential therapeutic targets. By virtual screening, we have identified a Rho-specific inhibitor, Rhosin. Rhosin contains two aromatic rings tethered by a linker, and it binds to the surface area sandwiching Trp58 of RhoA with a submicromolar Kd and effectively inhibits GEF-catalyzed RhoA activation. In cells, Rhosin specifically inhibited RhoA activity and RhoA-mediated cellular function without affecting Cdc42 or Rac1 signaling activities. By suppressing RhoA or RhoC activity, Rhosin could inhibit mammary sphere formation by breast cancer cells, suppress invasion of mammary epithelial cells, and induce neurite outgrowth of PC12 cells in synergy with NGF. Thus, the rational designed RhoA subfamily-specific small molecule inhibitor is useful for studying the physiological and pathologic roles of Rho GTPase. © 2012 Elsevier Ltd All rights reserved.

Plageman T.F.,Childrens Hospital Research Foundation | Zacharias A.L.,University of Michigan | Gage P.J.,University of Michigan | Lang R.A.,Childrens Hospital Research Foundation | Lang R.A.,University of Cincinnati
Developmental Biology | Year: 2011

The cytoskeletal protein Shroom3 is a potent inducer of epithelial cell shape change and is required for lens and neural plate morphogenesis. Analysis of gut morphogenesis in Shroom3 deficient mouse embryos revealed that the direction of gut rotation is also disrupted. It was recently established that Pitx2-dependent, asymmetrical cellular behaviors in the dorsal mesentery (DM) of the early mid-gut, a structure connecting the gut-tube to the rest of the embryo, contribute to the direction of gut rotation in chicken embryos by influencing the direction of the dorsal mesenteric tilt. Asymmetric cell shapes in the DM epithelium are hypothesized to contribute to the tilt, however, it is unclear what lies downstream of Pitx2 to alter epithelial cell shape. The cells of the left DM epithelium in either Pitx2 or Shroom3 deficient embryos are shorter and wider than those in control embryos and resemble the shape of those on the right, demonstrating that like Pitx2, Shroom3 is required for cell shape asymmetry and the leftward DM tilt. Because N-cadherin expression is specific to the left side and is Pitx2 dependent, we determined whether Shroom3 and N-cadherin function together to regulate cell shape in the left DM epithelium. Analysis of mouse embryos lacking one allele of both Shroom3 and N-cadherin revealed that they possess shorter and wider left epithelial DM cells when compared with Shroom3 or N-cadherin heterozygous embryos. This indicates a genetic interaction. Together these data provide evidence that Shroom3 and N-cadherin function cooperatively downstream of Pitx2 to directly regulate cell shape changes necessary for early gut tube morphogenesis. © 2011 Elsevier Inc.

McLendon P.M.,Childrens Hospital Research Foundation | Robbins J.,Childrens Hospital Research Foundation
Circulation Research | Year: 2015

Baseline physiological function of The mammalian heart is under The constant threat of environmental or intrinsic pathological insults. Cardiomyocyte proteins are thus subject to unremitting pressure to function optimally, and this depends on them assuming and maintaining proper conformation. This review explores The multiple defenses a cell may use for its proteins to assume and maintain correct protein folding and conformation. There are multiple quality control mechanisms to ensure that nascent polypeptides are properly folded and mature proteins maintain their functional conformation. When proteins do misfold, either in The face of normal or pathological stimuli or because of intrinsic mutations or post-translational modifications, they must either be refolded correctly or recycled. In The absence of these corrective processes, they may become toxic to The cell. Herein, we explore some of The underlying mechanisms that lead to proteotoxicity. The continued presence and chronic accumulation of misfolded or unfolded proteins can be disastrous in cardiomyocytes because these misfolded proteins can lead to aggregation or The formation of soluble peptides that are proteotoxic. This in turn leads to compromised protein quality control and precipitating a downward spiral of The cell's ability to maintain protein homeostasis. Some underlying mechanisms are discussed and The therapeutic potential of interfering with proteotoxicity in The heart is explored. © 2015 American Heart Association, Inc.

Zhang S.,Childrens Hospital Research Foundation | Zhou X.,Childrens Hospital Research Foundation | Lang R.A.,Childrens Hospital Research Foundation | Guo F.,Childrens Hospital Research Foundation
PLoS ONE | Year: 2012

RhoA is a member of the Rho family small GTPases that are implicated in various cell functions including proliferation and survival. However, the physiological role of RhoA in vivo remains largely unknown. Here, we deleted RhoA in the B cell and hematopoietic stem cell (HSC) populations in RhoA flox/flox mice with CD19 and Mx promoter-driven Cre expression, respectively. Deletion of RhoA by CD19 Cre/+ significantly blocked B cell development in spleen, leading to a marked reduction in the number of transitional, marginal zone, and follicular B cells. Surprisingly, neither B cell proliferation in response to either LPS or B cell receptor (BCR) engagement nor B cell survival rate in vivo was affected by RhoA deletion. Furthermore, RhoA -/- B cells, like control cells, were rescued from apoptosis by BCR crosslinking in vitro. In contrast, RhoA deficiency led to a defect in B cell activating factor (BAFF)-mediated B cell survival that was associated with a dampened expression of BAFF receptor and a loss of BAFF-mediated Akt activation. Finally, HSC deletion of RhoA by Mx-Cre severely reduced proB/preB and immature B cell populations in bone marrow while common lymphoid progenitors were increased, indicating that RhoA is also required for B cell progenitor/precursor differentiation. Taken together, our results uncover an important role for RhoA at multiple stages of B cell development. © 2012 Zhang et al.

Raghu H.,Childrens Hospital Research Foundation | Flick M.J.,Childrens Hospital Research Foundation
Current Pharmaceutical Biotechnology | Year: 2011

Fibrinogen is a provisional matrix protein of the coagulation system that following proteolytic cleavage by the protease thrombin polymerizes to form fibrin, the structural basis of the blood clot. Fibrin polymer formation at sites of vessel injury is critical to normal hemostasis. However, fibrin deposition within damaged tissues is also a common pathological feature of inflammatory diseases, including rheumatoid arthritis. Fibrin deposition has been readily detected along articular surfaces, within inflamed hyperplastic synovial tissue, and as a component of insoluble "rice bodies" within the synovial fluid of arthritic joints. Recent data has suggested that fibrin deposition within inflamed tissues is not simply a reflection of a disease process but rather actively contributes to disease pathogenesis. One mechanism that has been demonstrated to directly link fibrin(ogen) to the regulation of inflammation is the ability of fibrin(ogen) to serve as a ligand for cell-surface receptors, particularly integrins. Indeed, engagement of fibrin(ogen) by the leukocyte integrin receptor α Mβ 2 appears to be a common and fundamental event driving local inflammation. Recent studies have demonstrated that eliminating fibrin(ogen)- α Mβ 2 interactions can significantly limit the progression of multiple inflammatory diseases, including arthritis, without compromising the ability of fibrinogen to function in coagulation. These exciting findings have opened the door to new opportunities for targeting fibrinogen as an inflammatory mediator while leaving intact its hemostatic properties. © 2011 Bentham Science Publishers.

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