Table_1_Effect of gait training using Welwalk on gait pattern in individuals with hemiparetic stroke: a cross-sectional study.DOCX

IntroductionWe aimed to explore the effect of gait training using Welwalk on gait patterns by comparing differences in gait patterns between robotic-assisted gait training using Welwalk and gait training using an orthosis in individuals with hemiparetic stroke.MethodsThis study included 23 individuals with hemiparetic stroke who underwent gait training with Welwalk combined with overground gait training using an orthosis. Three-dimensional motion analysis on a treadmill was performed under two conditions for each participant: during gait training with Welwalk and with the ankle-foot orthosis. The spatiotemporal parameters and gait patterns were compared between the two conditions.ResultsThe affected step length was significantly longer, the step width was significantly wider, and the affected single support phase ratio was significantly higher in the Welwalk condition than in the orthosis condition. The index values of abnormal gait patterns were significantly lower while using Welwalk than in the orthosis condition. The following four indices were lower in the Welwalk condition: contralateral vaulting, insufficient knee flexion, excessive hip external rotation during the paretic swing phase, and paretic forefoot contact.DiscussionGait training using Welwalk increased the affected step length, step width, and single support phase while suppressing abnormal gait patterns as compared to gait training using the ankle-foot orthosis. This study suggests that gait training using Welwalk may promote a more efficient gait pattern reacquisition that suppresses abnormal gait patterns.Trial registrationProspectively registered in the Japan Registry of Clinical Trials (https://jrct.niph.go.jp; jRCTs042180152).</p

Differing susceptibility to autophagic degradation of two LC3-binding proteins: SQSTM1/p62 and TBC1D25/OATL1

<p>MAP1LC3/LC3 (a mammalian ortholog family of yeast Atg8) is a ubiquitin-like protein that is essential for autophagosome formation. LC3 is conjugated to phosphatidylethanolamine on phagophores and ends up distributed both inside and outside the autophagosome membrane. One of the well-known functions of LC3 is as a binding partner for receptor proteins, which target polyubiquitinated organelles and proteins to the phagophore through direct interaction with LC3 in selective autophagy, and their LC3-binding ability is essential for degradation of the polyubiquitinated substances. Although a number of LC3-binding proteins have been identified, it is unknown whether they are substrates of autophagy or how their interaction with LC3 is regulated. We previously showed that one LC3-binding protein, TBC1D25/OATL1, plays an inhibitory role in the maturation step of autophagosomes and that this function depends on its binding to LC3. Interestingly, TBC1D25 seems not to be a substrate of autophagy, despite being present on the phagophore. In this study we investigated the molecular basis for the escape of TBC1D25 from autophagic degradation by performing a chimeric analysis between TBC1D25 and SQSTM1/p62 (sequestosome 1), and the results showed that mutant TBC1D25 with an intact LC3-binding site can become an autophagic substrate when TBC1D25 is forcibly oligomerized. In addition, an ultrastructural analysis showed that TBC1D25 is mainly localized outside autophagosomes, whereas an oligomerized TBC1D25 mutant rather uniformly resides both inside and outside the autophagosomes. Our findings indicate that oligomerization is a key factor in the degradation of LC3-binding proteins and suggest that lack of oligomerization ability of TBC1D25 results in its asymmetric localization at the outer autophagosome membrane.</p

Effects of eye-drop applications of Tat-µCL on ERG in P23H rats.

<p>A) Representative ERG traces. Eye-drops containing vehicle (PBS) or 1 mM Tat-µCL in saline were administered to P23H rats from PN 14 to 89 days. Scotopic ERGs were recorded at PN 30, 70, or 90 days. B) Mean amplitudes of photoreceptor-derived a-waves. C) Mean amplitudes of Müller cells-derived b-waves. Data are expressed as means ± standard deviation (n = 8 eyes (8 rats) per group). *<i>P</i><0.05 and **<i>P</i><0.01 versus the vehicle-treated group (<i>t</i>-test).</p

Effects of an intravitreal injection or eye drop applications of Tat-µCL on ERG in S334ter rats.

<p>S334ter rats received an intravitreal injection of 2 µl of 20 mM Tat-µCL at PN 15 days (▪). Another group of S334ter rats received eye-drops containing 20 mM Tat-µCL from PN 13 to 55 days (•). Scotopic ERGs were recorded at PN 18, 21, 24, 28, 35, 42, 49, and 56 days. A) Mean amplitudes of photoreceptor-derived a-waves. B) Mean amplitudes of Müller cells-derived b-waves. Data are expressed as means ± standard deviation (n = 8 eyes (8 rats) per group). *<i>P</i><0.05 and **<i>P</i><0.01 versus the none-treated group (○) (<i>t</i>-test).</p

Determinations of photoreceptor cell death in S334ter rat retinas.

<p><b>A)</b> TUNEL assay of retinal sections of S334ter rats. Eyes were enucleated at PN 13, 14, 15, 18, 21, 24, 27, or 30 days. Retinal sections were stained with TUNEL (green) and DAPI (blue). B) Quantitative analysis of the number of TUNEL-positive cells in the ONL. Data are expressed as means ± standard deviation (n = 12 eyes (6 rats) per group). Abbreviations: ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer.</p

Effects of intravitreal injection of Tat-µCL on photoreceptor cell death in S334ter rats.

<p>A) TUNEL staining of retinal sections of S334ter rats treated with Tat-µCL. S334ter rats received intravitreal injection of 2 µl of vehicle (PBS), 4 mM PD150606, or 20 mM Tat-µCL at PN 15 days. Eyes were enucleated at PN 18 days. Retinal sections were stained with TUNEL (green) and DAPI (blue). B) Quantitative analysis of the number of TUNEL-positive cells in the ONL at PN 18 days. Data are expressed as means ± standard deviation (n = 12 eyes (6 rats) per group). ***<i>P</i><0.001 versus vehicle (<i>t</i>-test). Abbreviations: ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer.</p

Effects of eye-drop applications of Tat-µCL on photoreceptor cell death in S334ter rats.

<p>A) TUNEL staining of retinal sections of S334ter rats treated with eye-drops containing Tat-µCL. Eye-drops containing vehicle (PBS) or 20 mM Tat-µCL were administered from PN 13 to 17 days. Eyes were enucleated at PN 18 days. Retinal sections were stained with TUNEL (green) and DAPI (blue). B) Quantitative analysis of the number of TUNEL-positive cells in the ONL at PN 18 days. Data are expressed as means ± standard deviation (n = 12 eyes (6 rats) per group). ***<i>P</i><0.001 versus vehicle (<i>t</i>-test). Abbreviations: ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer.</p

Effects of eye drop applications of Tat-µCL on photoreceptor cell death in P23H rats.

<p>A) TUNEL of retinal sections of P23H rats treated with eye-drops containing Tat-µCL. Eye-drops containing saline (PBS), 1 mM Tat-µCL in saline, or 1 mM Tat-µCL in 0.1% HA were administered from PN 14 to 49 days. Eyes were enucleated at PN 30, 40, or 50 days. Retinal sections were stained with TUNEL (green) and DAPI (blue). B) Quantitative analysis of the number of TUNEL-positive cells in the ONL at PN 30, 40, and 50 days. Data are expressed as means ± standard deviation (n = 12 eyes (6 rats) per group). *<i>P</i><0.05 and **<i>P</i><0.01 versus the saline-treated group (<i>t</i>-test). Abbreviations: ONL, outer nuclear layer.</p

Determination of nuclear translocation of AIF in P23H rat retinas.

<p>A) Eyes were enucleated at PN 40 days, and retinal sections were stained with AIF (red), TUNEL (green) and DAPI (blue). AIF was detected in photoreceptor cell nuclei. Arrows indicate localization of AIF in TUNEL-positive photoreceptor nuclei. B) Effects of eye-drop applications of Tat-µCL on nuclear translocation of AIF in P23H rats. Eye-drops containing saline (PBS), 1 mM Tat-µCL in saline, or 1 mM Tat-µCL in 0.1% HA were administered from PN 14 to 39 days. Eyes were enucleated at PN 40 days. Retinal sections were stained with AIF (red) and DAPI (blue). White circles indicate translocation of AIF inside photoreceptor nuclei (shown by pink color). Abbreviations: OS, photoreceptor outer segment; IS, photoreceptor inner segment; ONL, outer nuclear layer; INL, inner nuclear layer; GCL, ganglion cell layer.</p

ATP-Binding Cassette Transporter A Subfamily 8 Is a Sinusoidal Efflux Transporter for Cholesterol and Taurocholate in Mouse and Human Liver

The ATP-binding cassette (ABC) transporter A subfamily 8 (ABCA8) belongs to the ABCA6-like transporters subgroup, which is distinct from the ABCA1-like subgroup in the ABCA family. The expression and function of the short-size human ABCA8 lacking one of the two ATP-binding domains for ATP hydrolysis, which are regularly present in the other ABCA transporters, have been reported. However, the functional differences between the short-size human ABCA8 and full-size human ABCA8, which has the two ATP-binding domains, remain unknown. The purpose of the present study was to clarify the tissue expression profiles of ABCA6-like and ABCA1-like subgroup transporters and the functional characteristics of ABCA8 in mouse and human. The tissue distribution of mouse ABCA (mABCA) transporter protein and the changes in mABCA8 protein expression levels in a mouse model of obstructive cholestasis were elucidated by means of quantitative targeted absolute proteomics (QTAP). The transport characteristics were clarified in a HEK293 cell line overexpressing full-size ABCA8 protein. QTAP and immunohistochemical analyses revealed that mABCA transporters exhibited the distinct protein expression patterns in the tissues, and mABCA8b, its mouse orthologue, was abundant in the liver and predominantly distributed in sinusoidal membranes of the hepatocytes. Further, protein expression of mABCA8b was decreased in the mouse cholestasis liver. Changes of mABCA8b expression level in cholestasis were similar to those of mABCA1, a sinusoidal cholesterol efflux transporter. Uptake and efflux assays showed that ABCA8 mediates efflux of [³H]­cholesterol and [³H]­taurocholate, while it showed no significant efflux activity for [³H]­estrone sulfate, [³H]­digoxin, [³H]­vinblastine, [³H]<i>para</i>-aminohippuric acid, [³H]­oleic acid, [¹⁴C]­nicotine, or [³H]­methotrexate. [³H]­Cholesterol efflux was increased by extracellularly applied taurocholate. These results suggest that mABCA8b/ABCA8 functions as a sinusoidal efflux transporter for at least cholesterol and taurocholate in mouse and human liver