Hsiao Jung Wang

Enhancement of neuroplasticity through upregulation of β1-integrin in human umbilical cord-derived stromal cell implanted stroke model

Neuroplasticity subsequent to functional angiogenesis is an important goal for cell-based therapy of ischemic neural tissues. At present, the cellular and molecular mechanisms involved are still not well understood. In this study, we isolated mesenchymal stem cells (MSCs) from Whartons jelly (WJ) to obtain clonally expanded human umbilical cord-derived mesenchymal stem cells (HUCMSCs) with multilineage differentiation potential. Experimental rats receiving intracerebral HUCMSC transplantation showed significantly improved neurological function compared to vehicle-treated control rats. Cortical neuronal activity, as evaluated by proton MR spectroscopy (1H-MRS), also increased considerably in the transplantation group. Transplanted HUCMSCs migrated towards the ischemic boundary zone and differentiated into glial, neuronal, doublecortin+, CXCR4+, and vascular endothelial cells to enhance neuroplasticity in the ischemic brain. In addition, HUCMSC transplantation promoted the formation of new vessels to increase local cortical blood flow in the ischemic hemisphere. Modulation by stem cell-derived macrophage/microglial interactions, and increased beta1-integrin expression, might enhance this angiogenic architecture within the ischemic brain. Inhibition of beta1-integrin expression blocked local angiogenesis and reduced recovery from neurological deficit. In addition, significantly increased modulation of neurotrophic factor expression was also found in the HUCMSC transplantation group. In summary, regulation of beta1-integrin expression plays a critical role in the plasticity of the ischemic brain after the implantation of HUCMSCs.

Stromal cell-derived factor-1α promotes neuroprotection, angiogenesis, and mobilization/homing of bone marrow-derived cells in stroke rats

Stromal cell-derived factor (SDF)-1α is involved in the trafficking of hematopoietic stem cells from bone marrow to peripheral blood, and its expression is increased in the penumbra of the ischemic brain. In this study, SDF-1α was found to exert neuroprotective effects that rescued primary cortical cultures from H2O2 neurotoxicity, and to modulate neurotrophic factor expression. Rats receiving intracerebral administration of SDF-1α showed less cerebral infarction due to up-regulation of antiapoptotic proteins, and they had improved motor performance. SDF-1α injection enhanced the targeting of bone marrow (BM)-derived cells to the injured brain, as demonstrated in green fluorescent protein-chimeric mice with cerebral ischemia. In addition, increased vascular density in the ischemic cortex of SDF-1α-treated rats enhanced functional local cerebral blood flow. In summary, intracerebral administration of SDF-1α resulted in neuroprotection against neurotoxic insult, and it induced increased BM-derived cell targeting to the ischemic brain, thereby reducing the volume of cerebral infarction and improving neural plasticity.

Secretoneurin promotes neuroprotection and neuronal plasticity via the Jak2/Stat3 pathway in murine models of stroke

Secretoneurin (SN), a neuropeptide derived from secretogranin II, promotes neurite outgrowth of immature cerebellar granule cells. SN also aids in the growth and repair of neuronal tissue, although the precise mechanisms underlying the promotion of brain tissue neuroprotection and plasticity by SN are not understood. Here, in a rat model of stroke and in ischemic human brain tissue, SN was markedly upregulated in both neurons and endothelial cells. SN-mediated neuroprotection rescued primary cortical cell cultures from oxygen/glucose deprivation. SN also induced expression of the antiapoptotic proteins Bcl-2 and Bcl-xL through the Jak2/Stat3 pathway and inhibited apoptosis by blocking caspase-3 activation. In addition, rats with occluded right middle cerebral arteries showed less cerebral infarction, improved motor performance, and increased brain metabolic activity following i.v. administration of SN. Furthermore, SN injection enhanced stem cell targeting to the injured brain in mice and promoted the formation of new blood vessels to increase local cortical blood flow in the ischemic hemisphere. Both in vitro and in vivo, SN not only promoted neuroprotection, but also enhanced neurogenesis and angiogenesis. Our results demonstrate that SN acts directly on neurons after hypoxia and ischemic insult to further their survival by activating the Jak2/Stat3 pathway.

Implantation of olfactory ensheathing cells promotes neuroplasticity in murine models of stroke

Murine olfactory ensheathing cells (OECs) promote central nervous system axonal regeneration in models of spinal cord injury. We investigated whether OECs could induce a neuroplastic effect to improve the neurological dysfunction caused by hypoxic/ischemic stress. In this study, human OECs/olfactory nerve fibroblasts (hOECs/ONFs) specifically secreted trophic factors including stromal cell-derived factor-1alpha (SDF-1alpha). Rats with intracerebral hOEC/ONF implantation showed more improvement on behavioral measures of neurological deficit following stroke than control rats. [18F]fluoro-2-deoxyglucose PET (FDG-PET) showed increased glucose metabolic activity in the hOEC/ONF-treated group compared with controls. In mice, transplanted hOECs/ONFs and endogenous homing stem cells including intrinsic neural progenitor cells and bone marrow stem cells colocalized with specific neural and vascular markers, indicating stem cell fusion. Both hOECs/ONFs and endogenous homing stem cells enhanced neuroplasticity in the rat and mouse ischemic brain. Upregulation of SDF-1alpha and CXCR4 in hOECs/ONFs promoted neurite outgrowth of cocultured primary cortical neurons under oxygen glucose deprivation conditions and in stroke animals through upregulation of cellular prion protein (PrP C) expression. Therefore, the upregulation of SDF-1alpha and the enhancement of CXCR4 and PrP C interaction induced by hOEC/ONF implantation mediated neuroplastic signals in response to hypoxia and ischemia.

Granulocyte Colony-Stimulating Factor Activating HIF-1α Acts Synergistically with Erythropoietin to Promote Tissue Plasticity

Stroke and peripheral limb ischemia are serious clinical problems with poor prognosis and limited treatment. The cytokines erythropoietin (EPO) and granulocyte-colony stimulating factor (G-CSF) have been used to induce endogenous cell repair and angiogenesis. Here, we demonstrated that the combination therapy of EPO and G-CSF exerted synergistic effects on cell survival and functional recovery from cerebral and peripheral limbs ischemia. We observed that even under normoxic conditions, G-CSF activates hypoxia-inducible factor-1α (HIF-1α), which then binds to the EPO promoter and enhances EPO expression. Serum EPO level was significantly increased by G-CSF injection, with the exception of Tg-HIF-1α+f/+f mice. The neuroplastic mechanisms exerted by EPO combined with G-CSF included enhanced expression of the antiapoptotic protein of Bcl-2, augmented neurotrophic factors synthesis, and promoted neovascularization. Further, the combination therapy significantly increased homing and differentiation of bone marrow stem cells (BMSCs) and intrinsic neural progenitor cells (INPCs) into the ischemic area. In summary, EPO in combination with G-CSF synergistically enhanced angiogenesis and tissue plasticity in ischemic animal models, leading to greater functional recovery than either agent alone.

Nonsenescent Hsp27-upregulated MSCs implantation promotes neuroplasticity in stroke model.

Cellular senescence induces changes in cellular physiology, morphology, proliferative capacity, and gene expression. Stem cell senescence might be one of the major issues of limited efficacy of stem cell transplantation. In this study, we demonstrated that implantation of human umbilical cord mesenchymal stem cells (hUCMSCs) cultured in human umbilical cord serum (hUCS) significantly enhanced neuroplasticity and angiogenesis in stroke and ischemic limb models. Immunophenotypic analysis indicated that hUCMSCs cultured in hUCS had more small and rapidly self-renewing cells than those expanded in FCS. The main cause of greater senescence in FCS-cultured cells was increased generation of reactive oxygen species (ROS). Proteome profiling showed significantly more senescence-associated vimentin in FCS-cultured hUCMSCs than in hUCS-cultured hUCMSCs. In contrast, there was significant upregulation of heat shock protein 27 (Hsp27) in the hUCS-cultured hUCMSCs. By gene targeting, we found that overexpression of Hsp27 may downregulate vimentin expression through inhibition of the nuclear translocation of p65 (NF-κB signaling). Thus, an interaction between Hsp27 and vimentin may modulate the degree of senescence in hUCS- and FCS-cultured hUCMSCs. In summary, hUCMSCs exhibiting senescence are detrimental to cell engraftment and differentiation in animal models via activation of NF-κB pathway. Human stem cells incubated in hUCS might reduce the senescent process through upregulation of Hsp27 to increase implantation efficiency.

Role of HIF-1α-activated Epac1 on HSC-mediated neuroplasticity in stroke model

Exchange protein activated by cAMP-1 (Epac1) plays an important role in cell proliferation, cell survival and neuronal signaling, and activation of Epac1 in endothelial progenitor cells increases their homing to ischemic muscles and promotes neovascularization in a model of hind limb ischemia. Moreover, upregulation of Epac1 occurs during organ development and in diseases such as myocardial hypertrophy, diabetes, and Alzheimers disease. We report here that hypoxia upregulated Epac1 through HIF-1α induction in the CD34-immunosorted human umbilical cord blood hematopoietic stem cells (hUCB(34)). Importantly, implantation of hUCB(34) subjected to hypoxia-preconditioning (HP-hUCB(34)) improved stroke outcome, more than did implantation of untreated hUCB(34), in rodents subjected to cerebral ischemia, and this required Epac1-to-matrix metalloprotease (MMP) signaling. This improved therapeutic efficacy correlated with better engraftment and differentiation of these cells in the ischemic host brain. In addition, more than did implantation of untreated HP-hUCB(34), implantation of HP-hUCB(34) improved cerebral blood flow into the ischemic brain via induction of angiogenesis, facilitated proliferation/recruitment of endogenous neural progenitor cells in the ischemic brain, and promoted neurite outgrowth following cerebral ischemia. Consistent with our proposed role of Epac1-to-MMP signaling in hypoxia-preconditioning, the above mentioned effects of implanting HP-hUCB(34) could be abolished by pharmacological inhibition and genetic disruption/deletion of Epac1 or MMPs. We have discovered a HIF-1α-to-Epac1-to-MMP signaling pathway that is required for the improved therapeutic efficacy resulting from hypoxia preconditioning of hUCB(34) in vitro prior to their implantation into the host brain in vivo.

New molecular insights into cellular survival and stress responses: neuroprotective role of cellular prion protein (PrPC).

Knowledge of the physiological function of cellular prion protein has been acquired from prion diseases such as Creutzfeldt–Jakob disease, as well as PRNP knock out and transgenic mice. Recent progress in neurobiology has further delineated the neuroprotective role played by cellular prion protein. In this paper, we review the role of cellular prion protein in cell survival including its antiapoptotic effect on Bax-mediated cell death and its responses to various environmental stresses including oxidative stress, and ischemia. Finally, we discuss the significance of cellular prion protein in different neurodegenerative diseases and the possible development of future therapies.

Role of stress‐inducible protein‐1 in recruitment of bone marrow derived cells into the ischemic brains

Stress‐inducible protein‐1 (STI‐1) is the proposed ligand for the cellular prion protein (PrPC), which is thought to facilitate recovery following stroke. Whether STI‐1 expression is affected by stroke and how its signalling facilitates recovery remain elusive. Brain slices from patients that died of ischemic stroke were collected for STI‐1 immunohistochemistry. These findings were compared to results from cell cultures, mice with or without the PrPC knockout, and rats. Based on these findings, molecular and pharmacological interventions were administered to investigate the underlying mechanisms and to test the possibility for therapy in experimental stroke models. STI‐1 was upregulated in the ischemic brains from humans and rodents. The increase in STI‐1 expression in vivo was not cell‐type specific, as it was found in neurons, glia and endothelial cells. Likewise, this increase in STI‐1 expression can be mimicked by sublethal hypoxia in primary cortical cultures (PCCs) in vitro, and appear to have resulted from the direct binding of the hypoxia inducible factor‐1α (HIF‐1α) to the STI‐1 promoter. Importantly, this STI‐1 signalling promoted bone marrow derived cells (BMDCs) proliferation and migration in vitro and recruitment to the ischemic brain in vivo, and augmenting its signalling facilitated neurological recovery in part by recruiting BMDCs to the ischemic brain. Our results thus identified a novel mechanism by which ischemic insults can trigger a self‐protective mechanism to facilitate recovery.

Induction of GAP-43 modulates neuroplasticity in PBSC (CD34+) implanted-Parkinson's model.

As a result of the progressive decrease in efficacy of drugs used to treat Parkinsons disease (PD) and the rapid development of motor complications, effective alternative treatments for PD are required. In a 6‐hydroxydopamine (6‐OHDA)‐induced Parkinsons rat model, intracerebral peripheral blood stem cell (CD34+) (PBSC) transplantation significantly protected dopaminergic neurons from 6‐OHDA‐induced neurotoxicity, enhanced neural repair of tyrosine hydroxylase neurons through up‐regulation of Bcl‐2, facilitated stem cell plasticity, and attenuated activation of microglia, in comparison with vehicle‐control rats. The 6‐OHDA‐lesioned hemi‐Parkinsonian rats receiving intrastriatal transplantation of PBSCs also showed: 1) enhanced glucose metabolism in the lesioned striatum and thalamus, demonstrated by [18F]fluoro‐2‐deoxyglucose positron emission tomography (FDG‐PET), 2) improved neurochemical activity as shown by proton magnetic resonance spectroscopy (1H‐MRS), and 3) significantly reduced rotational behavior in comparison with control lesioned rats. These observations might be explained by an up‐regulation of growth‐associated protein 43 (GAP‐43) expression because improvements in neurological dysfunction were blocked by injection of MK‐801 in the PBSC‐treated group. In addition, a significant increase in neurotrophic factor expression was found in the ipsilateral hemisphere of the PBSC‐treated group. In summary, this protocol may be a useful strategy for the treatment of clinical PD.