Browsing by Author "Hsu, Chih-Chien"

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Carboxylated nanodiamond-mediated CRISPR-Cas9 delivery of human retinoschisis mutation into human iPSCs and mouse retina
(2020-01-01)
Yang, Tien-Chun
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Chang, Chia-Yu
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Yarmishyn, Aliaksandr A.
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Mao, Yen-Shiang
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Yang, Yi-Ping
Nanodiamonds (NDs) are considered to be relatively safe carbon nanomaterials used for the transmission of DNA, proteins and drugs. The feasibility of utilizing the NDs to deliver CRISPR-Cas9 system for gene editing has not been clearly studied. Therefore, in this study, we aimed to use NDs as the carriers of CRISPR-Cas9 components designed to introduce the mutation in RS1 gene associated with Xlinked retinoschisis (XLRS). ND particles with a diameter of 3 nm were functionalized by carboxylation of the surface and covalently conjugated with fluorescent mCherry protein. Two linear DNA constructs were attached to the conjugated mCherry: one encoded Cas9 endonuclease and GFP reporter, another encoded sgRNA and contained insert of HDR template designed to introduce RS1 c.625C>T mutation. Such nanoparticles were successfully delivered and internalized by human iPSCs and mouse retinas, the efficiency of internalization was significantly improved by mixing with BSA. The delivery of ND particles led to introduction of RS1 c.625C>T mutation in both human iPSCs and mouse retinas. Rs1 gene editing in mouse retinas resulted in several pathological features typical for XLRS, such as aberrant photoreceptor structure. To conclude, our ND-based CRISPR-Cas9 delivery system can be utilized as a tool for creating in vitro and in vivo disease models of XLRS. Statement of significance X-linked retinoschisis (XLRS) is a prevalent hereditary retinal disease, which is caused by mutations in RS1 gene, whose product is important for structural organization of the retina. The recent development of genome editing techniques such as CRISPR-Cas9 significantly improved the prospects for better understanding the pathology and development of treatment for this disease. Firstly, gene editing can allow development of appropriate in vitro and in vivo disease models; secondly, CRISPR-Cas9 can be applied for gene therapy by removing the disease-causative mutation in vivo. The major prerequisite for these approaches is to develop safe and efficient CRISPR-Cas9 delivery system. In this study, we tested specifically modified nanodiamonds for such a delivery system. We were able to introduce Rs1 mutation into the mouse retina and, importantly, observed several XLRS-specific effects. (C) 2019 Acta Materialia Inc. Published by Elsevier Ltd.
Dynamic load balancing scheme for VoIP over WLANs
(2008-01-01)
Tsao, Shiao-Li
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Hsu, Chih-Chien
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資訊工程學系
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Department of Computer Science
Coverage areas of WLAN access points (APs) are usually overlapped so a WLAN station (STA) might be able to find several APs to attach in a WLAN hotspot. Experimental results indicate that a WLAN STA normally associates with an AP with the maximal signal strength and requests the bandwidth of the AP for establishing network connections. However, this kind of STA-centric association and bandwidth request policy may introduce unbalance loads of APs, and the bandwidths of APs in a WLAN hotspot cannot be fully utilized. This unbalance load problem is a critical issue for the commercial deployment of voice over IP (VOlP) over WLAN (VoWLAN) service which has to maximize the number of concurrent VoWLAN sessions with Quality of Service (QoS) guarantees. In this paper, a novel dynamic load balancing scheme is proposed for a VoWLAN system. The network-assisted association policy first advices an STA to request a VoWLAN session through an AP with the minimal load. In case of the AN which the STA can attach are all overloaded, the proposed load balancing scheme further rearranges the serving VoWLAN STAB between APs in order to spare enough resources for accommodating that new request. Simulation results demonstrate that the reject rate of service requests for a VoWLAN system can be considerably reduced by employing the proposed scheme.
Glutamate Stimulation Dysregulates AMPA Receptors-Induced Signal Transduction Pathway in Leber's Inherited Optic Neuropathy Patient-Specific hiPSC-Derived Retinal Ganglion Cells
(2019-06-01)
Yang, Yi-Ping
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Phan Nguyen Nhi Nguyen
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Lin, Tai-Chi
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Yarmishyn, Aliaksandr A.
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Chen, Wun-Syuan
The mitochondrial genetic disorder, Leber's hereditary optic neuropathy (LHON), is caused by a mutation in MT-ND4 gene, encoding NADH dehydrogenase subunit 4. It leads to the progressive death of retinal ganglion cells (RGCs) and causes visual impairment or even blindness. However, the precise mechanisms of LHON disease penetrance and progression are not completely elucidated. Human-induced pluripotent stem cells (hiPSCs) offer unique opportunities to investigate disease-relevant phenotypes and regulatory mechanisms underlying LHON pathogenesis at the cellular level. In this study, we successfully generated RGCs by differentiation of LHON patient-specific hiPSCs. We modified the protocol of differentiation to obtain a more enriched population of single-cell RGCs for LHON study. Based on assessing morphology, expression of specific markers and electrophysiological activity, we found that LHON-specific hiPSC-derived were more defective in comparison with normal wild-type RGCs. Based on our previous study, whereby by using microarray analysis we identified that the components of glutamatergic synapse signaling pathway were significantly downregulated in LHON-specific RGCs, we focused our study on glutamate-associated alpha -amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors. We found that the protein expression levels of the subunits of the AMPA receptor, GluR1 and GluR2, and their associated scaffold proteins were decreased in LHON-RGCs. By performing the co-immunoprecipitation assay, we found several differences in the efficiencies of interaction between AMPA subunits and scaffold proteins between normal and LHON-specific RGCs.
Load Balancing Apparatus And Method In Wireless Network Hotspots
(2008-04-10)
Tsao, Shiao-Li
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Hsu, Chih-Chien
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Yu, Tai-Xing
Disclosed is a load balancing apparatus and method in wireless network hotspots, which comprises a resource allocation module and a load balancer. The resources reallocation module establishes the resources module and the relationship between access points (APs) and STAs in the wireless network hotspots, and seeks possible load balance shift paths (LBSPs). From these possible LBSPs, an LBSP is selected. Based on the selected LBSP, the load balancer reallocates network resources and dynamically arranges the load among the APs in the wireless network hotspots. This invention can be applicable to a centralized or a decentralized wireless communication system.