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syntaxin12  (Proteintech)


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    Proteintech syntaxin12
    Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). <t>Syntaxin12</t> (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .
    Syntaxin12, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/syntaxin12/product/Proteintech
    Average 93 stars, based on 13 article reviews
    syntaxin12 - by Bioz Stars, 2026-05
    93/100 stars

    Images

    1) Product Images from "Therapeutic Modulation of the Nox2–Hv1–ROS Axis by Botulinum Neurotoxin A Confers Protection Against CoCl 2 -Induced Retinal Hypoxic Injury"

    Article Title: Therapeutic Modulation of the Nox2–Hv1–ROS Axis by Botulinum Neurotoxin A Confers Protection Against CoCl 2 -Induced Retinal Hypoxic Injury

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms262110806

    Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). Syntaxin12 (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .
    Figure Legend Snippet: Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). Syntaxin12 (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .

    Techniques Used: CCK-8 Assay, Control, Expressing, Immunocytochemistry, Activity Assay, Staining, Translocation Assay, Disruption

    BoNT/A protects retinal structure and reduces retinal ganglion cell loss in an ex vivo model. ( A ) Experimental scheme of the ex vivo retina model. Rat retinas were cultured ex vivo, pretreated with BoNT/A (2 h), and subsequently exposed to CoCl 2 (300 μM). Samples were collected at 3h, 4D (days), and 8D. ( B ) Representative hematoxylin–eosin (H&E) staining images of paraffin-embedded retinal cross-sections at the indicated time points. Retinal thickness was quantified for the whole retina as well as for specific layers, including the inner plexiform layer (IPL), inner nuclear layer (INL), and outer nuclear layer (ONL). CoCl 2 treatment caused progressive thinning of the retina and individual layers, whereas BoNT/A pretreatment preserved retinal thickness at levels comparable to controls. Scale bar: 50 μm. ( C ) Immunofluorescence staining of cleaved-SNAP25 confirmed BoNT/A enzymatic activity in retinal tissues. Robust cleaved-SNAP25 signals were observed primarily in the IPL of BoNT/A-pretreated retinas but were absent in control and CoCl 2 -only groups. Enlarged views highlight localization within the IPL and adjacent INL. Quantitative fluorescence analyses are shown in the adjacent graphs. Scale bars: 50 μm (overview); 25 μm (insets). ( D ) Brn3a immunostaining demonstrated that BoNT/A pretreatment preserved retinal ganglion cell labeling compared with CoCl 2 -treated retinas. Scale bar: 50 μm. ( E ) TUNEL assay revealed abundant apoptotic nuclei (green) in CoCl 2 -treated retinas, which were markedly reduced in BoNT/A-pretreated samples, indicating protection against hypoxia-induced apoptosis. Scale bar: 50 μm. ( F ) IBA1 (red) and GFAP (green) were strongly upregulated in CoCl 2 -treated retinas, confirming successful induction of hypoxia, but were reduced by BoNT/A treatment, indicating suppression of glial activation. Scale bar: 50 μm. ( G ) Immunostaining for Hv1 (red) and Nox2 (green) revealed increased expression in CoCl 2 -treated retinas, which was reduced following BoNT/A pretreatment. Scale bar: 50 μm. ( H ) Western blot analyses using the same antibodies as in R28 cells showed that Nox2, Hv1, COX2, NLRP3, and TNF-α were elevated by CoCl 2 and downregulated by BoNT/A. In contrast, SOCS3, GAP43, and Syntaxin12 were increased in BoNT/A-treated samples, suggesting enhanced anti-inflammatory and neuroprotective signaling. Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. Control; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. CoCl 2 . + p < 0.05, ++ p < 0.01, +++ p < 0.001 BoNT/A 0.25 IU vs. 0.5 IU. Scale bar: 50 μm.
    Figure Legend Snippet: BoNT/A protects retinal structure and reduces retinal ganglion cell loss in an ex vivo model. ( A ) Experimental scheme of the ex vivo retina model. Rat retinas were cultured ex vivo, pretreated with BoNT/A (2 h), and subsequently exposed to CoCl 2 (300 μM). Samples were collected at 3h, 4D (days), and 8D. ( B ) Representative hematoxylin–eosin (H&E) staining images of paraffin-embedded retinal cross-sections at the indicated time points. Retinal thickness was quantified for the whole retina as well as for specific layers, including the inner plexiform layer (IPL), inner nuclear layer (INL), and outer nuclear layer (ONL). CoCl 2 treatment caused progressive thinning of the retina and individual layers, whereas BoNT/A pretreatment preserved retinal thickness at levels comparable to controls. Scale bar: 50 μm. ( C ) Immunofluorescence staining of cleaved-SNAP25 confirmed BoNT/A enzymatic activity in retinal tissues. Robust cleaved-SNAP25 signals were observed primarily in the IPL of BoNT/A-pretreated retinas but were absent in control and CoCl 2 -only groups. Enlarged views highlight localization within the IPL and adjacent INL. Quantitative fluorescence analyses are shown in the adjacent graphs. Scale bars: 50 μm (overview); 25 μm (insets). ( D ) Brn3a immunostaining demonstrated that BoNT/A pretreatment preserved retinal ganglion cell labeling compared with CoCl 2 -treated retinas. Scale bar: 50 μm. ( E ) TUNEL assay revealed abundant apoptotic nuclei (green) in CoCl 2 -treated retinas, which were markedly reduced in BoNT/A-pretreated samples, indicating protection against hypoxia-induced apoptosis. Scale bar: 50 μm. ( F ) IBA1 (red) and GFAP (green) were strongly upregulated in CoCl 2 -treated retinas, confirming successful induction of hypoxia, but were reduced by BoNT/A treatment, indicating suppression of glial activation. Scale bar: 50 μm. ( G ) Immunostaining for Hv1 (red) and Nox2 (green) revealed increased expression in CoCl 2 -treated retinas, which was reduced following BoNT/A pretreatment. Scale bar: 50 μm. ( H ) Western blot analyses using the same antibodies as in R28 cells showed that Nox2, Hv1, COX2, NLRP3, and TNF-α were elevated by CoCl 2 and downregulated by BoNT/A. In contrast, SOCS3, GAP43, and Syntaxin12 were increased in BoNT/A-treated samples, suggesting enhanced anti-inflammatory and neuroprotective signaling. Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. Control; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. CoCl 2 . + p < 0.05, ++ p < 0.01, +++ p < 0.001 BoNT/A 0.25 IU vs. 0.5 IU. Scale bar: 50 μm.

    Techniques Used: Ex Vivo, Cell Culture, Staining, Immunofluorescence, Activity Assay, Control, Fluorescence, Immunostaining, Labeling, TUNEL Assay, Activation Assay, Expressing, Western Blot

    Suppression of the Nox2–Hv1 Axis and Enhancement of Neuroprotective Signaling by BoNT/A in hypoxic injury. Cobalt chloride (CoCl 2 )–induced hypoxia stabilizes HIF-1α and triggers excessive ROS production, leading to metabolic stress within cells. This condition further drives inflammation (Hv1, Nox2, NLRP3, COX2, TNF-α), retinal thinning, apoptosis, and glial activation. Pretreatment with Botulinum Toxin A (BoNT/A) suppresses the Nox2–Hv1 axis, reduces ROS and inflammatory signaling, preserves retinal ganglion cells (Brn3a), and attenuates gliosis (Iba1, GFAP). Moreover, BoNT/A enhances protective mediators (SOCS3, GAP43, Syntaxin12), thereby shifting the retinal microenvironment from a degenerative toward a neuroprotective state.
    Figure Legend Snippet: Suppression of the Nox2–Hv1 Axis and Enhancement of Neuroprotective Signaling by BoNT/A in hypoxic injury. Cobalt chloride (CoCl 2 )–induced hypoxia stabilizes HIF-1α and triggers excessive ROS production, leading to metabolic stress within cells. This condition further drives inflammation (Hv1, Nox2, NLRP3, COX2, TNF-α), retinal thinning, apoptosis, and glial activation. Pretreatment with Botulinum Toxin A (BoNT/A) suppresses the Nox2–Hv1 axis, reduces ROS and inflammatory signaling, preserves retinal ganglion cells (Brn3a), and attenuates gliosis (Iba1, GFAP). Moreover, BoNT/A enhances protective mediators (SOCS3, GAP43, Syntaxin12), thereby shifting the retinal microenvironment from a degenerative toward a neuroprotective state.

    Techniques Used: Activation Assay



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    Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). <t>Syntaxin12</t> (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .
    Syntaxin12, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). <t>Syntaxin12</t> (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .
    Syntaxin12/13 Antibody, supplied by Synaptic Systems, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). Syntaxin12 (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .

    Journal: International Journal of Molecular Sciences

    Article Title: Therapeutic Modulation of the Nox2–Hv1–ROS Axis by Botulinum Neurotoxin A Confers Protection Against CoCl 2 -Induced Retinal Hypoxic Injury

    doi: 10.3390/ijms262110806

    Figure Lengend Snippet: Protective effects of BoNT/A in R28 cells under hypoxic stress. ( A ) Schematic illustration of the experimental timeline. R28 cells were pretreated with BoNT/A for 2 h, followed by CoCl 2 (200 μM) exposure, and samples were collected at 3 h and 24 h. ( B ) Cell viability was tested using the CCK-8 assay. Data are presented as mean ± SEM (** p < 0.01 vs. control; # p < 0.05 vs. CoCl 2 ). ( C ) Expression levels of target proteins in R28 cells exposed to CoCl 2 for 3 h. ( D ) Immunocytochemistry analyses revealed multiple BoNT/A-related effects. The first panel confirmed BoNT/A activity by cleaved-SNAP25 staining (green). The second panel showed increased SOCS3 expression after BoNT/A treatment. The third panel demonstrated that BoNT/A suppressed the CoCl 2 -induced upregulation of Hv1 and Nox2. The fourth panel revealed that CoCl 2 promoted NF-κB nuclear translocation, which was reduced by BoNT/A pretreatment. The fifth panel showed that Brn3a expression, markedly decreased in CoCl 2 -treated cells, was preserved in the CoCl 2 + BoNT/A group, indicating protection of retinal ganglion cell integrity. Scale bars: 50 μm. ( E ) VDAC1 (red) displayed pronounced perinuclear clustering in CoCl 2 -treated cells, consistent with mitochondrial aggregation under stress, whereas this effect was attenuated by BoNT/A pretreatment (arrows). Syntaxin12 (red), normally extended in filamentous structures, exhibited fragmentation in CoCl 2 + BoNT/A samples at 24 h, suggesting cleavage or disruption of its morphology. The yellow boxes indicate the regions that were magnified and shown below to highlight these structural changes. Scale bars: 50 μm (overview); 25 μm (insets). Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control; # p < 0.05, ### p < 0.001 vs. CoCl 2 .

    Article Snippet: Syntaxin12 , Rabbit , Proteintech, Rosemont, IL, USA , 14259-1-AP , 1:200 (ICC), 1:1000 (WB).

    Techniques: CCK-8 Assay, Control, Expressing, Immunocytochemistry, Activity Assay, Staining, Translocation Assay, Disruption

    BoNT/A protects retinal structure and reduces retinal ganglion cell loss in an ex vivo model. ( A ) Experimental scheme of the ex vivo retina model. Rat retinas were cultured ex vivo, pretreated with BoNT/A (2 h), and subsequently exposed to CoCl 2 (300 μM). Samples were collected at 3h, 4D (days), and 8D. ( B ) Representative hematoxylin–eosin (H&E) staining images of paraffin-embedded retinal cross-sections at the indicated time points. Retinal thickness was quantified for the whole retina as well as for specific layers, including the inner plexiform layer (IPL), inner nuclear layer (INL), and outer nuclear layer (ONL). CoCl 2 treatment caused progressive thinning of the retina and individual layers, whereas BoNT/A pretreatment preserved retinal thickness at levels comparable to controls. Scale bar: 50 μm. ( C ) Immunofluorescence staining of cleaved-SNAP25 confirmed BoNT/A enzymatic activity in retinal tissues. Robust cleaved-SNAP25 signals were observed primarily in the IPL of BoNT/A-pretreated retinas but were absent in control and CoCl 2 -only groups. Enlarged views highlight localization within the IPL and adjacent INL. Quantitative fluorescence analyses are shown in the adjacent graphs. Scale bars: 50 μm (overview); 25 μm (insets). ( D ) Brn3a immunostaining demonstrated that BoNT/A pretreatment preserved retinal ganglion cell labeling compared with CoCl 2 -treated retinas. Scale bar: 50 μm. ( E ) TUNEL assay revealed abundant apoptotic nuclei (green) in CoCl 2 -treated retinas, which were markedly reduced in BoNT/A-pretreated samples, indicating protection against hypoxia-induced apoptosis. Scale bar: 50 μm. ( F ) IBA1 (red) and GFAP (green) were strongly upregulated in CoCl 2 -treated retinas, confirming successful induction of hypoxia, but were reduced by BoNT/A treatment, indicating suppression of glial activation. Scale bar: 50 μm. ( G ) Immunostaining for Hv1 (red) and Nox2 (green) revealed increased expression in CoCl 2 -treated retinas, which was reduced following BoNT/A pretreatment. Scale bar: 50 μm. ( H ) Western blot analyses using the same antibodies as in R28 cells showed that Nox2, Hv1, COX2, NLRP3, and TNF-α were elevated by CoCl 2 and downregulated by BoNT/A. In contrast, SOCS3, GAP43, and Syntaxin12 were increased in BoNT/A-treated samples, suggesting enhanced anti-inflammatory and neuroprotective signaling. Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. Control; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. CoCl 2 . + p < 0.05, ++ p < 0.01, +++ p < 0.001 BoNT/A 0.25 IU vs. 0.5 IU. Scale bar: 50 μm.

    Journal: International Journal of Molecular Sciences

    Article Title: Therapeutic Modulation of the Nox2–Hv1–ROS Axis by Botulinum Neurotoxin A Confers Protection Against CoCl 2 -Induced Retinal Hypoxic Injury

    doi: 10.3390/ijms262110806

    Figure Lengend Snippet: BoNT/A protects retinal structure and reduces retinal ganglion cell loss in an ex vivo model. ( A ) Experimental scheme of the ex vivo retina model. Rat retinas were cultured ex vivo, pretreated with BoNT/A (2 h), and subsequently exposed to CoCl 2 (300 μM). Samples were collected at 3h, 4D (days), and 8D. ( B ) Representative hematoxylin–eosin (H&E) staining images of paraffin-embedded retinal cross-sections at the indicated time points. Retinal thickness was quantified for the whole retina as well as for specific layers, including the inner plexiform layer (IPL), inner nuclear layer (INL), and outer nuclear layer (ONL). CoCl 2 treatment caused progressive thinning of the retina and individual layers, whereas BoNT/A pretreatment preserved retinal thickness at levels comparable to controls. Scale bar: 50 μm. ( C ) Immunofluorescence staining of cleaved-SNAP25 confirmed BoNT/A enzymatic activity in retinal tissues. Robust cleaved-SNAP25 signals were observed primarily in the IPL of BoNT/A-pretreated retinas but were absent in control and CoCl 2 -only groups. Enlarged views highlight localization within the IPL and adjacent INL. Quantitative fluorescence analyses are shown in the adjacent graphs. Scale bars: 50 μm (overview); 25 μm (insets). ( D ) Brn3a immunostaining demonstrated that BoNT/A pretreatment preserved retinal ganglion cell labeling compared with CoCl 2 -treated retinas. Scale bar: 50 μm. ( E ) TUNEL assay revealed abundant apoptotic nuclei (green) in CoCl 2 -treated retinas, which were markedly reduced in BoNT/A-pretreated samples, indicating protection against hypoxia-induced apoptosis. Scale bar: 50 μm. ( F ) IBA1 (red) and GFAP (green) were strongly upregulated in CoCl 2 -treated retinas, confirming successful induction of hypoxia, but were reduced by BoNT/A treatment, indicating suppression of glial activation. Scale bar: 50 μm. ( G ) Immunostaining for Hv1 (red) and Nox2 (green) revealed increased expression in CoCl 2 -treated retinas, which was reduced following BoNT/A pretreatment. Scale bar: 50 μm. ( H ) Western blot analyses using the same antibodies as in R28 cells showed that Nox2, Hv1, COX2, NLRP3, and TNF-α were elevated by CoCl 2 and downregulated by BoNT/A. In contrast, SOCS3, GAP43, and Syntaxin12 were increased in BoNT/A-treated samples, suggesting enhanced anti-inflammatory and neuroprotective signaling. Statistical significance was indicated as * p < 0.05, ** p < 0.01, *** p < 0.001 vs. Control; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. CoCl 2 . + p < 0.05, ++ p < 0.01, +++ p < 0.001 BoNT/A 0.25 IU vs. 0.5 IU. Scale bar: 50 μm.

    Article Snippet: Syntaxin12 , Rabbit , Proteintech, Rosemont, IL, USA , 14259-1-AP , 1:200 (ICC), 1:1000 (WB).

    Techniques: Ex Vivo, Cell Culture, Staining, Immunofluorescence, Activity Assay, Control, Fluorescence, Immunostaining, Labeling, TUNEL Assay, Activation Assay, Expressing, Western Blot

    Suppression of the Nox2–Hv1 Axis and Enhancement of Neuroprotective Signaling by BoNT/A in hypoxic injury. Cobalt chloride (CoCl 2 )–induced hypoxia stabilizes HIF-1α and triggers excessive ROS production, leading to metabolic stress within cells. This condition further drives inflammation (Hv1, Nox2, NLRP3, COX2, TNF-α), retinal thinning, apoptosis, and glial activation. Pretreatment with Botulinum Toxin A (BoNT/A) suppresses the Nox2–Hv1 axis, reduces ROS and inflammatory signaling, preserves retinal ganglion cells (Brn3a), and attenuates gliosis (Iba1, GFAP). Moreover, BoNT/A enhances protective mediators (SOCS3, GAP43, Syntaxin12), thereby shifting the retinal microenvironment from a degenerative toward a neuroprotective state.

    Journal: International Journal of Molecular Sciences

    Article Title: Therapeutic Modulation of the Nox2–Hv1–ROS Axis by Botulinum Neurotoxin A Confers Protection Against CoCl 2 -Induced Retinal Hypoxic Injury

    doi: 10.3390/ijms262110806

    Figure Lengend Snippet: Suppression of the Nox2–Hv1 Axis and Enhancement of Neuroprotective Signaling by BoNT/A in hypoxic injury. Cobalt chloride (CoCl 2 )–induced hypoxia stabilizes HIF-1α and triggers excessive ROS production, leading to metabolic stress within cells. This condition further drives inflammation (Hv1, Nox2, NLRP3, COX2, TNF-α), retinal thinning, apoptosis, and glial activation. Pretreatment with Botulinum Toxin A (BoNT/A) suppresses the Nox2–Hv1 axis, reduces ROS and inflammatory signaling, preserves retinal ganglion cells (Brn3a), and attenuates gliosis (Iba1, GFAP). Moreover, BoNT/A enhances protective mediators (SOCS3, GAP43, Syntaxin12), thereby shifting the retinal microenvironment from a degenerative toward a neuroprotective state.

    Article Snippet: Syntaxin12 , Rabbit , Proteintech, Rosemont, IL, USA , 14259-1-AP , 1:200 (ICC), 1:1000 (WB).

    Techniques: Activation Assay