• KSBB Journal
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• v.32 no.2
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• pp.90-95
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• 2017

Mammalian cell cultures have been used extensively to produce proteins for therapeutic agent because of their ability to perform post-translational modification including glycosylation. To produce recombinant protein, many factors and parameter are considered such as media composition, host cell type, and culture process. In this study, recombinant human erythropoietin (rhEPO) producing cell line was established by using glutamine synthetase system. To reduce serum concentration in media, we compared direct adaptation with step adaptation. Cell growth was faster in step adaptation. In low-level serum media, there were insufficient glucose for cell growth. Thus, we added glucose in low-level serum media from 2 g/L to 4.5 g/L. Titer of rhEPO was higher than other conditions at 4.5 g/L of glucose. Additionally, N-methyl-D-aspartate (NMDA), 13-cis-retinal, and pluronic F-68 (PF-68) were added to enhance productivity in CHO cell cultures. In conclusion, we applied CHO cell producing rhEPO to low-level of serum in media using step-adaptation. Also, we confirmed positive effect of NMDA, 13-cis-retinal, and PF-68.

• Journal of Biomedical Engineering Research
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• v.28 no.2
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• pp.212-217
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• 2007

As a preliminary study for the development of electrical stimulation strategy of artificial retina, we set up a method fur the reconstruction of input intensity variation from retinal ganglion cell(RGC) responses. In order to estimate light intensity variation, we used an optimal linear filter trained from given stimulus intensity variation and multiple single unit spike trains from RGCs. By applying ON/OFF stimulation(ON duration: 2 sec, OFF duration: 5 sec) repetitively, we identified three functional types of ganglion cells according to when they respond to the ON/OFF stimulus actively: ON cell, OFF cell, and ON-OFF cell. Experiments were also performed using a Gaussian random stimulus and a binary random stimulus. The input intensity was updated once every 90 msec(i. e. 11 Hz) to present the stimulus. The result of reconstructing 11 Hz Gaussian and binary random stimulus was not satisfactory and showed low correlation between the original and reconstructed stimulus. In the case of ON/OFF stimulus in which temporal variation is slow, successful reconstruction was achieved and the correlation coefficient was as high as 0.8.

• Molecules and Cells
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• v.46 no.7
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• pp.441-450
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• 2023

β-Catenin (Ctnnb1) has been shown to play critical roles in the development and maintenance of epithelial cells, including the retinal pigment epithelium (RPE). Ctnnb1 is not only a component of intercellular junctions in the epithelium, it also functions as a transcriptional regulator in the Wnt signaling pathway. To identify which of its functional modalities is critically involved in mouse RPE development and maintenance, we varied Ctnnb1 gene content and activity in mouse RPE lineage cells and tested their impacts on mouse eye development. We found that a Ctnnb1 double mutant (Ctnnb1^dm), which exhibits impaired transcriptional activity, could not replace Ctnnb1 in the RPE, whereas Ctnnb1^Y654E, which has reduced affinity for the junctions, could do so. Expression of the constitutively active Ctnnb1^∆ex3 mutant also suppressed the development of RPE, instead facilitating a ciliary cell fate. However, the post-mitotic or mature RPE was insensitive to the loss, inactivation, or constitutive activation of Ctnnb1. Collectively, our results suggest that Ctnnb1 should be maintained within an optimal range to specify RPE through transcriptional regulation of Wnt target genes in the optic neuroepithelium.

• Journal of Life Science
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• v.17 no.8 s.88
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• pp.1068-1074
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• 2007

Parvalbumin occurs in various types of cells in the retina. We previously reported parvalbumin distribution in the inner nuclear layer of bat retina. In the present study, we identified the parvalbumin-immunoreactive neurons in the ganglion cell layer of the retina of a bat, Rhinolophus ferrumequinum, and investigated the distribution pattern of the labeled neurons. Parvalbumin immunoreactivity was found in numerous cell bodies in the ganglion cell layer. Quantitative analysis showed that these cells had medium to large-sized somas. The soma diameter of the parvalbumin-immunoreactive cells in the ganglion cell layer ranged from 12.35 to 19.12 ${\mu}m$ (n=166). As the fibers in the nerve fiber layer were also stained, the majority of parvalbumin-immunoreactive cells in the ganglion cell layer should be medium to large-sized retinal ganglion cells. The mean nearest neighbor distance of the parvalbumin-immunoreactive cells in the ganglion cell layer of the bat retina ranged from 59.57 to 62.45 ${\mu}m$ and the average regularity index was 2.95 ${\pm}$ 0.3 (n=4). The present results demonstrate that parvalbu-min is expressed in medium to large-sized retinal ganglion cells in bat retina, and they have a well-or-ganized distributional pattern with regular mosaics. These results should be important as they are applicable to a better understanding of the unsolved issue of a bat vision. This data will help to provide fundamental knowledge for the better understanding of the unique behavioral aspects of bat flight maneuverability.

• Journal of Nutrition and Health
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• v.38 no.2
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• pp.96-103
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• 2005

It has been suggested that the elevated plasma homocysteine may lead to retinal dysfunction. We investigated the effects of plasma levels of homocysteine and folate on the retinal glial cells' injuries. Male Sprague-Dawley rats were raised either on a control diet or on an experimental diet containing 3.0 g/kg homocystine without folic acid for 10 weeks. Plasma homocysteine concentrations were measured by a HPLC-fluorescence detection method. Plasma folate and vitamin B/sub 12/ levels were analyzed by a radioimmunoassay. The response of Muller cells which are the principal glial cells of the retina was immunohistochemically examined using an antibody for vimentin, a cytoskeletal protein belonging to the family of intermediate filament. At 2 weeks, the homocystine diet induced a twofold increase in plasma homocysteine, and a concomitant increase in the expression of vimentin in the Muller cells' processes spanning from the inner to outer membranes of the retina indicating arterial degeneration. At 10 weeks, the homocystine diet induced a fourfold increase in plasma homocystine, but vimentin immunoreactivity in the retinas was similar in both groups. In conclusion, increased plasma homocysteine levels have influence on morphological and functional changes of Muller cells in the retina. (Korean J Nutrition 38(2): 96～103, 2005)

• BMB Reports
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• v.56 no.9
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• pp.502-507
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• 2023

Photobiomodulation therapy has been proposed as a promising therapeutic approach for retinal degenerative diseases. However, its effect on the regenerative capacity in mammalian retina and its intracellular signalling mechanisms remain unknown. Here, we show that photobiomodulation with 670 nm light stimulates Müller glia cell cycle re-entry and dedifferentiation into a progenitor-like state in both the uninjured and injured retina. We also find that 670 nm light treatment inhibits the Hippo pathway, which is activated in Müller glia following NaIO₃-induced retinal injury. YAP, a major downstream effector of the Hippo signalling pathway was translocated into the nucleus of Müller glia along with YAP dephosphorylation in retina treated with 670 nm light. Deficiency of YAP attenuated Müller glia cell cycle re-entry and dedifferentiation. Our data reveal that the Hippo-YAP signalling pathway is associated with the photostimulatory effect on regenerative response in mammalian retina, and suggest a potential therapeutic strategy for retinal degenerative diseases.

• Journal of Biomedical Engineering Research
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• v.30 no.4
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• pp.347-354
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• 2009

For the reliable transmission of meaningful visual information using prosthetic electrical stimulation, it is required to develop an effective stimulation strategy for the generation of electrical pulse trains based on input visual information. The characteristics of neuronal activities of retinal ganglion cells (RGCs) evoked by electrical stimulation should be understood for this purpose. In this study, for the development of an optimal stimulation strategy for visual prosthesis, we analyzed the neuronal responses of RGCs in rd1 mouse, photoreceptor-degenerated retina of animal model of retinal diseases (retinitis pigmentosa). Based on the in-vitro model of epiretinal prosthesis which consists of planar multielectrode array (MEA) and retinal patch, we recorded and analyzed multiunit RGC activities evoked by amplitude-modulated electrical pulse trains. Two modes of responses were observed. Short-latency responses occurring at 3 ms after the stimulation were estimated to be from direct stimulation of RGCs. Long-latency responses were also observed mainly at 2 - 100 ms after stimulation and showed rhythmic firing with same frequency as the oscillatory background field potential. The long-latency responses could be modulated by pulse amplitude and duration. From the results, we expect that optimal stimulation conditions such as pulse amplitude and pulse duration can be determined for the successful transmission of visual information by electrical stimulation.

• Progress in Medical Physics
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• v.18 no.1
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• pp.20-26
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• 2007

It is expected that synaptic construction and electrical characteristics In degenerate retina might be different from those In normal retina. Therefore, we analyzed the retinal waveform recorded with multielectrode array in normal and degenerate retina using principal component analysis (PCA) and Independent component analysis (ICA) and compared the results. PCA Is a well established method for retinal waveform while ICA has not tried for retinal waveform analysis. We programmed ICA toolbox for spatiotemporal analysis of retinal waveform. In normal mouse, the MEA spatial map shows a single hot spot perfectly matched with PCA-derived ON or OFF ganglion cell response. However In rd/rd mouse, the MEA spatial map shows numerous hot and cold spots whose underlying interactions and mechanisms need further Investigation for better understanding.

• Biomolecules & Therapeutics
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• v.17 no.2
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• pp.175-180
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• 2009

Taurine is the most abundant free amino acid in the retina and transported into retina via taurine transporter (TauT) at the inner blood-retinal barrier (iBRB). In the present study, we investigated whether the taurine transport at the iBRB is regulated by oxidative stress or disease-like state in a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB) used as an in vitro model of iBRB. First, [$^3H$]taurine uptake and efflux by TR-iBRB were regulated in the presence of extracellular $Ca^{2+}$. [$^3H$]Taurine uptake was inhibited and efflux was enhanced under $Ca^{2+}$ free condition in the cells. In addition, oxidative stress inducing agents such as tumor necrosis factor-$\alpha$ (TNF-$\alpha$), lipopolysaccharide (LPS), diethyl maleate (DEM) and glutamate increased [$^3H$]taurine uptake and decreased [$^3H$]taurine efflux in TR-iBRB cells. Whereas, 3-morpholinosydnonimine (SIN-1), which is known to NO donor decreased [$^3H$]taurine uptake. Lastly, TR-iBRB cells exposed to high glucose (25 mM) medium and the [$^3H$]taurine uptake was reduced about 20% at the condition. Also, [$^3H$]taurine uptake was decreased by cytochalasin B, which is known to glucose transport inhibitor. In conclusion, taurine transport in TR-iBRB cells is regulated diversely at extracellular $Ca^{2+}$, oxidative stress and hyperglycemic condition. It suggested that taurine would play a role as a retinal protector in diverse disease states.

• BMB Reports
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• v.42 no.7
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• pp.456-461
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• 2009

Proteomic analyses of differentially expressed proteins in rat retinal ganglion cells (RGC-5) following S-nitrosoglutathione (GSNO), an NO donor, treatment were conducted. Of the approximately 314 protein spots that were detected, 19 were differentially expressed in response to treatment with GSNO. Of these, 14 proteins were up-regulated and 5 were down- regulated. Notably, an increase in GAPDH expression following GSNO treatment was detected in RGC-5 cells through Western blotting as well as proteomics. The increased GAPDH expression in response to GSNO treatment was accompanied by an increase in Herc6 protein, an E3 ubiquitin ligase. Moreover, GSNO treatment resulted in the translocation of GADPH from the cytosol to the nucleus and its subsequent accumulation. These results suggest that NO stress-induced apoptosis may be associated with the nuclear translocation and accumulation of GAPDH in RGC-5 cells.