Browsing by Author "Li, Ran"
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Item Open Access Abl Kinases Modulate Epithelial Architecture by Regulating Beta1 Integrin and c-Met Signals(2011) Li, RanNormal development and homeostasis require dynamic and tight regulation of epithelial architecture. Abnormal epithelial physiology is associated with various pathological conditions including cancers, and may be induced by changes in epithelial polarity, morphology and/or movement. Among the signaling pathways modulating epithelial physiology are those downstream of integrins and receptor tyrosine kinases (RTKs). Although roles of multiple integrins and RTKs in epithelium homeostasis have been established, the identity of signals regulating the functions of these surface receptors and the pathways connecting them to the regulation of epithelial architecture remain largely unknown. In this dissertation, I have identified the Abl family of non-receptor tyrosine kinases (Abl and Arg) as regulators of beta1 integrin and Met receptor tyrosine kinase signaling.
Abl family kinases are hyper-activated in multiple solid tumors and implicated in epithelial polarity regulation. Dysfunction of beta1 integrin is also associated with carcinoma development. To study the role of the Abl family member Arg in epithelial cell polarity, I have taken advantage of a three-dimensional (3D) cell culture system, where Madin Darby canine kidney type II (MDCKII) cells grown in collagen gels develop into polarized cyst structures. I have found that expression of active Arg kinase results in the formation of cysts with inverted apical polarity and that active Arg modulates epithelial polarity by regulating beta1 integrin and small GTPases pathways. In addition, I have shown that Arg regulates the Rap1-beta1 integrin pathway independently of the Rac1 pathway which promotes basal laminin assembly. I have also found that Abl family kinases function downstream of Met and that Abl kinase hyperactivity correlates with Met activation in a mouse mammary tumor model. Abl kinases are activated by HGF which is the ligand for Met, and active Abl kinases are recruited to the Met receptor and promote its tyrosine phosphorylation. Using fluorescence resonance energy transfer (FRET), I have found that Abl kinases regulate RhoA GTPase activity which contributes to actomyosin contractility induced by Met receptor activation. Further, Abl kinases positively regulate Met-dependent migration and invasion induced by HGF in several breast cancer cell lines.
In conclusion, I have identified novel functions of the Abl kinases in epithelial architecture regulation: modulation of epithelial polarity by targeting beta1 integrin function and promotion of Met signaling required for migration and invasion. This has important implications as it suggests potential roles for Abl kinases in carcinoma initiation mediated by beta1 integrin dysfunction, and development of Abl kinases inhibitors for treatment of cancers driven by hyper-activation of HGF-Met signaling.
Item Open Access Activation of the ATF6 (Activating Transcription Factor 6) Signaling Pathway in Neurons Improves Outcome After Cardiac Arrest in Mice.(Journal of the American Heart Association, 2021-06-11) Shen, Yuntian; Li, Ran; Yu, Shu; Zhao, Qiang; Wang, Zhuoran; Sheng, Huaxin; Yang, WeiBackground Ischemia/reperfusion injury impairs proteostasis, and triggers adaptive cellular responses, such as the unfolded protein response (UPR), which functions to restore endoplasmic reticulum homeostasis. After cardiac arrest (CA) and resuscitation, the UPR is activated in various organs including the brain. However, the role of the UPR in CA has remained largely unknown. Here we aimed to investigate effects of activation of the ATF6 (activating transcription factor 6) UPR branch in CA. Methods and Results Conditional and inducible sATF6-KI (short-form ATF6 knock-in) mice and a selective ATF6 pathway activator 147 were used. CA was induced in mice by KCl injection, followed by cardiopulmonary resuscitation. We first found that neurologic function was significantly improved, and neuronal damage was mitigated after the ATF6 pathway was activated in neurons of sATF6-KI mice subjected to CA/cardiopulmonary resuscitation. Further RNA sequencing analysis indicated that such beneficial effects were likely attributable to increased expression of pro-proteostatic genes regulated by ATF6. Especially, key components of the endoplasmic reticulum-associated degradation process, which clears potentially toxic unfolded/misfolded proteins in the endoplasmic reticulum, were upregulated in the sATF6-KI brain. Accordingly, the CA-induced increase in K48-linked polyubiquitin in the brain was higher in sATF6-KI mice relative to control mice. Finally, CA outcome, including the survival rate, was significantly improved in mice treated with compound 147. Conclusions This is the first experimental study to determine the role of the ATF6 UPR branch in CA outcome. Our data indicate that the ATF6 UPR branch is a prosurvival pathway and may be considered as a therapeutic target for CA.Item Open Access Activation of the XBP1s/O-GlcNAcylation Pathway Improves Functional Outcome After Cardiac Arrest and Resuscitation in Young and Aged Mice.(Shock (Augusta, Ga.), 2021-11) Li, Ran; Shen, Yuntian; Li, Xuan; Lu, Liping; Wang, Zhuoran; Sheng, Huaxin; Hoffmann, Ulrike; Yang, WeiAbstract
After cardiac arrest (CA) and resuscitation, the unfolded protein response (UPR) is activated in various organs including the brain. However, the role of the UPR in CA outcome remains largely unknown. One UPR branch involves spliced X-box-binding protein-1 (XBP1s). Notably, XBP1s, a transcriptional factor, can upregulate expression of specific enzymes related to glucose metabolism, and subsequently boost O-linked β-N-acetylglucosamine modification (O-GlcNAcylation). The current study is focused on effects of the XBP1 UPR branch and its downstream O-GlcNAcylation on CA outcome. Using both loss-of-function and gain-of-function mouse genetic tools, we provide the first evidence that activation of the XBP1 UPR branch in the post-CA brain is neuroprotective. Specifically, neuron-specific Xbp1 knockout mice had worse CA outcome, while mice with neuron-specific expression of Xbp1s in the brain had better CA outcome. Since it has been shown that the protective role of the XBP1s signaling pathway under ischemic conditions is mediated by increasing O-GlcNAcylation, we then treated young mice with glucosamine, and found that functional deficits were mitigated on day 3 post CA. Finally, after confirming that glucosamine can boost O-GlcNAcylation in the aged brain, we subjected aged mice to 8 min CA, and then treated them with glucosamine. We found that glucosamine-treated aged mice performed significantly better in behavioral tests. Together, our data indicate that the XBP1s/O-GlcNAc pathway is a promising target for CA therapy.Item Open Access Bone Marrow Mesenchymal Stem Cell Transplantation Increases GAP-43 Expression via ERK1/2 and PI3K/Akt Pathways in Intracerebral Hemorrhage.(Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology, 2017-01) Cui, Jianzhong; Cui, Changmeng; Cui, Ying; Li, Ran; Sheng, Huaxin; Jiang, Xiaohua; Tian, Yanxia; Wang, Kaijie; Gao, JunlingBackground/aims
Intracerebral hemorrhage (ICH) occurs in hypertensive patients and results in high rates of mortality and disability. This study determined whether bone marrow mesenchymal stem cell (BMSC) transplantation affects axonal regeneration and examined the underlying mechanisms after the administration of PD98059 (p-ERK1/2 inhibitor) or/ and LY294002 (PI3K inhibitor). The hypothesis that was intended to be tested was that BMSC transplantation regulates the expression of growth-associated protein-43 (GAP-43) via the ERK1/2 and PI3K/Akt signaling pathways.Methods
Seventy-five male rats (250-280 g) were subjected to intracerebral blood injection and then randomly received a vehicle, BMSCs, PD98059 or LY294002 treatment. Neurological deficits were evaluated prior to injury and at 1, 3 and 7 days post-injury. The expression of GAP-43, Akt, p-Akt, ERK1/2, and p-ERK1/2 proteins was measured by western blot analysis.Results
BMSC transplantation attenuated neurological deficits 3-7 days post-ICH. The expression of GAP-43 was increased 3 days following BMSC transplantation. However, this increase was inhibited by either PD98059 or LY294002 treatment. Treatment with both PD98059 and LY294002 was more effective than was treatment with an individual compound.Conclusion
BMSC transplantation could attenuate neurological deficits and activate axonal regeneration in this rat ICH model. The protective effects might be associated with increased GAP-43 expression by activating both the ERK1/2 and PI3K/Akt signaling pathways.Item Open Access Cardiac arrest and resuscitation activates the hypothalamic-pituitary-adrenal axis and results in severe immunosuppression.(Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism, 2021-05) Zhao, Qiang; Shen, Yuntian; Li, Ran; Wu, Jiangbo; Lyu, Jingjun; Jiang, Maorong; Lu, Liping; Zhu, Minghua; Wang, Wei; Wang, Zhuoran; Liu, Qiang; Hoffmann, Ulrike; Karhausen, Jörn; Sheng, Huaxin; Zhang, Weiguo; Yang, WeiIn patients who are successfully resuscitated after initial cardiac arrest (CA), mortality and morbidity rates are high, due to ischemia/reperfusion injury to the whole body including the nervous and immune systems. How the interactions between these two critical systems contribute to post-CA outcome remains largely unknown. Using a mouse model of CA and cardiopulmonary resuscitation (CA/CPR), we demonstrate that CA/CPR induced neuroinflammation in the brain, in particular, a marked increase in pro-inflammatory cytokines, which subsequently activated the hypothalamic-pituitary-adrenal (HPA) axis. Importantly, this activation was associated with a severe immunosuppression phenotype after CA. The phenotype was characterized by a striking reduction in size of lymphoid organs accompanied by a massive loss of immune cells and reduced immune function of splenic lymphocytes. The mechanistic link between post-CA immunosuppression and the HPA axis was substantiated, as we discovered that glucocorticoid treatment, which mimics effects of the activated HPA axis, exacerbated post-CA immunosuppression, while RU486 treatment, which suppresses its effects, significantly mitigated lymphopenia and lymphoid organ atrophy and improved CA outcome. Taken together, targeting the HPA axis could be a viable immunomodulatory therapeutic to preserve immune homeostasis after CA/CPR and thus improve prognosis of post-resuscitation CA patients.Item Open Access Development and Evaluation of a Novel Mouse Model of Asphyxial Cardiac Arrest Revealed Severely Impaired Lymphopoiesis After Resuscitation.(J Am Heart Assoc, 2021-05-20) Wang, Wei; Li, Ran; Miao, Wanying; Evans, Cody; Lu, Liping; Lyu, Jingjun; Li, Xuan; Warner, David S; Zhong, Xiaoping; Hoffmann, Ulrike; Sheng, Huaxin; Yang, WeiBackground Animal disease models represent the cornerstone in basic cardiac arrest (CA) research. However, current experimental models of CA and resuscitation in mice are limited. In this study, we aimed to develop a mouse model of asphyxial CA followed by cardiopulmonary resuscitation (CPR), and to characterize the immune response after asphyxial CA/CPR. Methods and Results CA was induced in mice by switching from an O2/N2 mixture to 100% N2 gas for mechanical ventilation under anesthesia. Real-time measurements of blood pressure, brain tissue oxygen, cerebral blood flow, and ECG confirmed asphyxia and ensuing CA. After a defined CA period, mice were resuscitated with intravenous epinephrine administration and chest compression. We subjected young adult and aged mice to this model, and found that after CA/CPR, mice from both groups exhibited significant neurologic deficits compared with sham mice. Analysis of post-CA brain confirmed neuroinflammation. Detailed characterization of the post-CA immune response in the peripheral organs of both young adult and aged mice revealed that at the subacute phase following asphyxial CA/CPR, the immune system was markedly suppressed as manifested by drastic atrophy of the spleen and thymus, and profound lymphopenia. Finally, our data showed that post-CA systemic lymphopenia was accompanied with impaired T and B lymphopoiesis in the thymus and bone marrow, respectively. Conclusions In this study, we established a novel validated asphyxial CA model in mice. Using this new model, we further demonstrated that asphyxial CA/CPR markedly affects both the nervous and immune systems, and notably impairs lymphopoiesis of T and B cells.Item Open Access MCC950, a selective NLPR3 inflammasome inhibitor, improves neurologic function and survival after cardiac arrest and resuscitation.(Journal of neuroinflammation, 2020-08-31) Jiang, Maorong; Li, Ran; Lyu, Jingjun; Li, Xuan; Wang, Wei; Wang, Zhuoran; Sheng, Huaxin; Zhang, Weiguo; Karhausen, Jörn; Yang, WeiBackground
Cardiac arrest (CA) is associated with high morbidity and mortality, even after spontaneous circulation is re-established. This dire situation is partly due to post-CA syndrome for which no specific and effective intervention is available. One key component of post-CA syndrome is sterile inflammation, which affects various organs including the brain. A major effector of sterile inflammation is activated NLRP3 inflammasome, which leads to increased release of interleukin (IL)-1β. However, how NLRP3 inflammasome impacts neuroinflammation and neurologic outcome after CA is largely undefined.Methods
Mice were subjected to a potassium-based murine CA and cardiopulmonary resuscitation (CPR) model. MCC950 was used to suppress activation of NLRP3 inflammasome after CA/CPR. Levels of protein and mRNA were examined by Western blotting and quantitative PCR, respectively. Immunologic changes were assessed by measuring cytokine expression and immune cell compositions. CA outcomes, including neurologic deficits, bacterial load in the lung, and survival rate, were evaluated.Results
Using our CA/CPR model, we found that NLRP3 inflammasome was activated in the post-CA brain, and that pro-inflammatory cytokine levels, including IL-1β, were increased. After treatment with MCC950, a potent and selective NLRP3 inflammasome inhibitor, mice exhibited improved functional recovery and survival rate during the 14-day observational period after CA/CPR. In line with these findings, IL-1β mRNA levels in the post-CA brain were significantly suppressed after MCC950 treatment. Interestingly, we also found that in MCC950- vs. vehicle-treated CA mice, immune homeostasis in the spleen was better preserved and bacterial load in the lung was significantly reduced.Conclusions
Our data demonstrate that activation of NLRP3 inflammasome could be a key event shaping the post-CA immuno- and neuro-pathology, and identify this pathway as a unique and promising therapeutic target to improve outcomes after CA/CPR.Item Open Access Photoacoustic imaging of in vivo hemodynamic responses to sodium nitroprusside.(Journal of biophotonics, 2021-03-26) Zhang, Dong; Li, Ran; Chen, Maomao; Vu, Tri; Sheng, Huaxin; Yang, Wei; Hoffmann, Ulrike; Luo, Jianwen; Yao, JunjieThe in vivo hemodynamic impact of sodium nitroprusside (SNP), a widely used antihypertensive agent, has not been well studied. Here, we applied functional optical-resolution photoacoustic microscopy (OR-PAM) to study the hemodynamic responses to SNP in mice in vivo. As expected, after the application of SNP, the systemic blood pressure (BP) was reduced by 53%. The OR-PAM results show that SNP induced an arterial vasodilation of 24% and 23% in the brain and skin, respectively. A weaker venous vasodilation of 9% and 5% was also observed in the brain and skin, respectively. The results show two different types of blood oxygenation response. In mice with decreased blood oxygenation, the arterial and venous oxygenation was respectively reduced by 6% and 13% in the brain, as well as by 7% and 18% in the skin. In mice with increased blood oxygenation, arterial and venous oxygenation was raised by 4% and 22% in the brain, as well as by 1% and 9% in the skin. We observed venous change clearly lagged the arterial change in the skin, but not in the brain. Our results collectively show a correlation among SNP induced changes in systemic BP, vessel size and blood oxygenation.Item Open Access Single-cell transcriptomic analysis of the immune cell landscape in the aged mouse brain after ischemic stroke.(Journal of neuroinflammation, 2022-04-07) Li, Xuan; Lyu, Jingjun; Li, Ran; Jain, Vaibhav; Shen, Yuntian; Del Águila, Ángela; Hoffmann, Ulrike; Sheng, Huaxin; Yang, WeiBackground
Ischemic stroke is a medical emergency that primarily affects the elderly. A complex immune response in the post-stroke brain constitutes a key component of stroke pathophysiology. This study aimed to determine how stroke affects immune cell populations in the aged brain based on molecular profiles of individual cells.Methods
Single-cell RNA sequencing and a new transient ischemic stroke mouse model with late reperfusion were used.Results
We generated, for the first time, a composite picture of immune cell populations in the stroke aged brain at single-cell resolution. We discovered at least 6 microglial subsets in the stroke aged brain, including a potentially stroke-specific subtype. Moreover, we identified major cell subpopulations formed by infiltrated myeloid cells after stroke, and revealed their unique molecular profiles.Conclusions
This study provided the first scRNA-seq data set for immune cells in the stroke aged brain, and offered novel insights into post-stroke immune cell heterogeneity.Item Open Access Small ubiquitin-like modifier 2 (SUMO2) is critical for memory processes in mice.(FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 2020-11) Yu, Shu; Galeffi, Francesca; Rodriguiz, Ramona M; Wang, Zhuoran; Shen, Yuntian; Lyu, Jingjun; Li, Ran; Bernstock, Joshua D; Johnson, Kory R; Liu, Shuai; Sheng, Huaxin; Turner, Dennis A; Wetsel, William C; Paschen, Wulf; Yang, WeiSmall ubiquitin-like modifier (SUMO1-3) conjugation (SUMOylation), a posttranslational modification, modulates almost all major cellular processes. Mounting evidence indicates that SUMOylation plays a crucial role in maintaining and regulating neural function, and importantly its dysfunction is implicated in cognitive impairment in humans. We have previously shown that simultaneously silencing SUMO1-3 expression in neurons negatively affects cognitive function. However, the roles of the individual SUMOs in modulating cognition and the mechanisms that link SUMOylation to cognitive processes remain unknown. To address these questions, in this study, we have focused on SUMO2 and generated a new conditional Sumo2 knockout mouse line. We found that conditional deletion of Sumo2 predominantly in forebrain neurons resulted in marked impairments in various cognitive tests, including episodic and fear memory. Our data further suggest that these abnormalities are attributable neither to constitutive changes in gene expression nor to alterations in neuronal morphology, but they involve impairment in dynamic SUMOylation processes associated with synaptic plasticity. Finally, we provide evidence that dysfunction on hippocampal-based cognitive tasks was associated with a significant deficit in the maintenance of hippocampal long-term potentiation in Sumo2 knockout mice. Collectively, these data demonstrate that protein conjugation by SUMO2 is critically involved in cognitive processes.