anti tmbim6  (Proteintech)

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A In silico TMBIM6 mRNA expression in human CNS from THPA database. B Tmbim6 mRNA expression on N2a cells after 18 h of exposure to 25 μM 6-OHDA or 50 μM rotenone. C Tmbim6 mRNA levels over time in PCNs exposed to aSyn for 96 h. D Changes of expression of Tmbim6 , BcL2 , and Bax over time in PCNs exposed to aSyn for 96 h. E Representative Western blots of total protein extracts from postmortem human SN from neurologically healthy controls and PD patients, probed for TMBIM6 and GAPDH (loading control). Full, uncropped blots are provided in Supplementary Material. F Densitometric quantification of TMBIM6 from blots in ( E ). Band intensities were normalized to GAPDH for each lane; individual data points are shown with mean ± SEM (n = 9–10 per group). For experiments involving more than two groups, a two-way ANOVA followed by Tukey’s multiple comparison test was performed. Pairwise comparisons between two groups were analyzed using unpaired t-test ( B ) or the Mann–Whitney U test ( F ). All bars represent mean ± SEM. Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; **=p < 0.01; ***=p < 0.001; ****=p < 0.0001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: In Silico, Expressing, Western Blot, Control, Comparison, MANN-WHITNEY

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A Graph shows validation of decreased Tmbim6 mRNA levels after 48 h of siRNA transfection in SN4741. B , C Representative immunoblots and quantification show mTmbim6 KD cells after 48 h. D Cytotoxicity assay shows cell death induced by 10 μM Tunicamycin after 24 h in KD cells. Results are expressed as % of LDH release. E Cytotoxicity assay shows cell death induced by 50 μM 6-OHDA after 24 h in KD cells. F Retention of DiOC6(3) assay shows the effect of aSyn on ΔΨm in KD cells after 18 h. Results are expressed as % of DiOC6(3) retention. G MTT assay shows mitochondrial-dependent cell death induced by aSyn in KD cells after 24 h. Results are expressed as % of MTT. H DEVD-AMC fluorescent assay shows the effect of aSyn on Caspase-3 activity in Tmbim6 KD cells after 24 h. Results are expressed as fold change of DEVD-AMC fluorescence intensity relative to vehicle. I Cytotoxicity assay shows the KD cell death induced by aSyn after 24 h. J Representative immunoblot of high–molecular-weight (HMW) aSyn species in SN4741 cells transfected with si Cntrl or si Tmbim6 and treated with 10 µM aSyn PFFs for 24 h; Tubulin was used as a loading control. K Densitometric quantification of HMW aSyn bands from the experiment described in ( J ) (integrated density normalized to Tubulin). All bars represent mean ± SEM. For experiments involving more than two groups, a two-way ANOVA followed by Tukey’s multiple comparison test was performed. Pairwise comparisons between two groups were analyzed using unpaired t-test ( A , C ) or the Mann–Whitney U test ( K ). Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; **=p < 0.01; ***=p < 0.001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: Biomarker Discovery, Transfection, Western Blot, Cytotoxicity Assay, MTT Assay, Fluorescence, Activity Assay, High Molecular Weight, Control, Comparison, MANN-WHITNEY

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A d Tmbim6 mRNA expression on homogenized flies’ heads. Results are expressed as fold change of mRNA expression. The bars represent mean ± SEM. B Optic image of eye integrity in RNAi-dTmbim6 flies incubated at 25 °C. C Quantification of eye integrity score in RNAi- dTmbim6 flies incubated at 25 °C (n = 25 per group). D Immunostaining of TH+ neurons in the lamina of RNAi-dTmbim6 flies incubated at 25 °C (n = 6 per group). E Quantification of the number of TH+ neurons in the lamina of RNAi-dTmbim6 flies incubated at 25 °C (n = 6 per group). F Schematic representation of rotenone-induced PD model in D. mel . G Spontaneous activity in DAergic RNAi-dTmbim6 flies after exposition to rotenone 300 μM for 7 days (n = 6 populations of 12 flies). H Climbing assay showed the motor ability of DAergic RNAi-dTmbim6 flies exposed to rotenone 300 μM for 7 days (n = 12 per group). I Representative confocal images of IF assay showed TH+ cells from DAergic RNAi-dTmbim6 flies exposed to rotenone 300 μM for 7 days. J The numbers of TH+ cells were quantified in each DAergic cluster, and K the somal size was analyzed (n = 7 per group). All bars represent mean ± SEM. For experiments involving more than two groups, a two-way ANOVA followed by Tukey’s multiple comparison test was performed. Pairwise comparisons between two groups were analyzed using unpaired t-test ( A , C ) or the Mann–Whitney U test ( E ). Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; ** = p < 0.01; ***= p < 0.001; ****= p < 0.0001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: Expressing, Incubation, Immunostaining, Activity Assay, Climbing Assay, Comparison, MANN-WHITNEY

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A Representative immunoblot shows a stable expression of TMBIM6 HA in SN4741 cells. B Cytotoxicity assay shows the cell death induced by Tunicamycin in SN4741 TMBIM6 HA cells after 24 h. C , D MTT assay shows cell death induced by 6-OHDA or rotenone in SN4741 TMBIM6 HA cells after 24 h. Results are expressed as % of MTT. E Retention of DiOC6(3) assay shows the effect of aSyn on ΔΨm in SN4741 hTMBIM6 HA cells after 18 h. Results are expressed as % of DiOC6(3) retention. F MTT assay shows cell death induced by aSyn in SN4741 TMBIM6 HA cells after 24 h. G DEVD-AMC fluorescent assay shows the effect of aSyn on Caspase-3 activity in SN4741 TMBIM6 HA cells after 24 h. H Cytotoxicity assay shows the cell death induced by aSyn in SN4741 TMBIM6 HA cells after 24 h. I Representative immunoblot of HMW aSyn species in Mock or TMBIM6 HA cells treated with aSyn PFFs for 24 h; TCE staining was used as a loading control. J Densitometric quantification of HMW aSyn bands from the experiment described in J (integrated density normalized to TCE). K Representative immunoblot shows expression of TMBIM6 HA and TMBIM6 D213A/HA in SN4741 cells. L MTT assay shows cell death induced by Tunicamycin and Thapsigargin in SN4741 Mock, TMBIM6 HA , and TMBIM6 D213A/HA cells after 24 h. Results are expressed as % of MTT. M Cytotoxicity assay shows the cell death induced by aSyn in SN4741 TMBIM6 HA and TMBIM6 D213A/HA cells after 24 h. N Cytotoxicity assay shows the cell death induced by aSyn after 10 days in PCNs transfected with Mock, TMBIM6 HA and TMBIM6 D213A/HA constructs. All bars represent mean ± SEM. For experiments involving more than two groups, a two-way ANOVA followed by Tukey’s multiple comparison test was performed. Pairwise comparisons between two groups were analyzed using the Mann–Whitney U test. Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; **=p < 0.01; ***=p < 0.001; ****=p < 0.0001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: Western Blot, Expressing, Cytotoxicity Assay, MTT Assay, Fluorescence, Activity Assay, Staining, Control, Transfection, Construct, Comparison, MANN-WHITNEY

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A An in-silico assay using Ingenuity Pathway Analysis (IPA) software shows the canonical pathways significantly associated with TMBIM6 interactors. B , C UMAP visualizations of the snRNA-seq dataset (GEO: GSE178265 ) from human postmortem substantia nigra. B shows TMBIM6 expression across all nuclei, while C distinguishes nuclei from healthy and PD donors. D Dot plot comparing the expression of TMBIM6 and UPR-related genes ( HSPA5, ERN1, XBP1, BLOC1S1 ) between healthy and PD conditions across all nuclei. E UMAP plot identifying resistant and vulnerable DAergic neuron populations within the dataset. F Dot plot comparing gene expression between resistant and vulnerable DAergic neurons within the PD cohort. For dot plots ( D , F ), dot size represents the percentage of cells expressing the gene, and color intensity indicates the mean expression level. Statistical significance for the differential expression shown in ( D , F ) was determined using the Model-based Analysis of Single-cell Transcriptomics (MAST) test. Full statistical details, including FDR-adjusted p-values, are provided in Supplementary Fig. .

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: In Silico, Software, Expressing, Gene Expression, Quantitative Proteomics, Single-cell Transcriptomics

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A Representative images show red fluorescent dots of PLA assay to TMBIM6 HA /IRE1a in stable SN4741 TMBIM6 HA cells exposed to aSyn. B Quantification of PLA dots per cell. Kruskal-Wallis followed by Dunn’s multiple comparison test. C In SN4741 siRNA- mTMBIM6 cells, an RT-qPCR assay shows the effect of aSyn on mRNA levels of mouse XBP1s ( mXbp1s ), D mouse BLOCS1 (mBlocs1) , and E mouse BIP (mBip) . F In SN4741 TMBIM6 HA cells, RT-qPCR assay shows the effect of aSyn in mXbp1s , G mBlocs1 , and H mBip , mRNA levels. Results are expressed as fold change, and bars represent mean ± SEM. All bars represent mean ± SEM. In ( B ), a Kruskal–Wallis test was performed followed by Dunn’s multiple comparison test. For experiments involving more than two groups, a two-way ANOVA followed by Tukey’s multiple comparison test was performed ( C – H ). Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; ***=p < 0.001; ****=p < 0.0001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: Comparison, Quantitative RT-PCR

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A , B Cytotoxicity assay shows the effect of IRE1a inhibition using MKC or 4 μ 8c on cell death induced by aSyn in Tmbim6 KD cells after 24 h. Results are expressed as % of LDH release. C Cytotoxicity assay shows the effect of PERK inhibitor on cell death induced by aSyn in mTmbim6 KD cells after 24 h. D A RT-qPCR assay shows effective double knockdown of both mTmbim6 (left panel) and mIre1 a (right panel) in SN4741 cells. Results are expressed as fold change, and bars represent mean ± SEM. One way ANOVA, Dunnett´s multiple comparation test. E Cytotoxicity assay shows downregulation of mIRE1a over cell death induced by aSyn in mTmbim6 KD cells after 24 h. Results are expressed as % of LDH release. F Cytotoxicity assay shows the JNK inhibitor AS60125 over cell death induced by aSyn in mTmbim6 KD cells after 24 h. G Cytotoxicity assay shows the BAX inhibitor BAI-1 over cell death induced by aSyn in mTmbim6 KD cells after 24 h. H Cytotoxicity assay shows the pan-caspase inhibitor ZVAD-FMK (casp-inh) over cell death induced by aSyn in mTmbim6 KD cells after 24 h. All bars represent mean ± SEM. In ( D ), a one-way ANOVA followed by Dunnett’s multiple comparison test was performed, whereas in ( E , F , G , H ), a two-way ANOVA with Tukey’s multiple comparisons test was conducted. Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; **=p < 0.01; ***=p < 0.001; ****=p < 0.0001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: Cytotoxicity Assay, Inhibition, Quantitative RT-PCR, Knockdown, Comparison

Journal: Cell Death & Disease

Article Title: TMBIM6 enhances dopaminergic neuron survival by modulating the IRE1a pathway in Parkinson’s disease

doi: 10.1038/s41419-025-08391-5

Figure Lengend Snippet: A Effect of two 6-OHDA doses on motor performance of mice, as a pharmacologic in vivo PD model (n = 4 per condition). Results are expressed as a percentage of contralateral forelimb use. B Timeline of in vivo transduction of the AAV-TMBIM6 HA/GFP and AAV-Mock GFP in SN and motor performance measurements in mice wild-type lesioned with 6-OHDA in CPu. C Cylinder test shows the effect of AAV-TMBIM6 HA/GFP and AAV-Mock GFP expression in SN over forelimb use after 2-, 3-, 4-, and 5-weeks post-injection (wpi). The colored area shows treatment with 6-OHDA injuries in the CPu. Results are expressed as a percentage of contralateral forelimb use (n Mockl = 4 and n TMBIM6 = 5). D Beam test shows the effect of the AAV-TMBIM6 HA/GFP and AAV-Mock GFP expression in SN on balance and coordination after 2, 3, 4, and 5 weeks after injection. The colored area shows treatment with 6-OHDA injury in the CPu. Results are expressed as the number of paws slips (n Mockl = 4 and n TMBIM6 = 5). E Effect of the AAV-TMBIM6 HA/GFP and AAV-Mock GFP expression over time, animals used to complete the Beam test during 2-, 3-, 4-, and 5-wpi. The colored area shows treatment with 6-OHDA injury in the CPu. Results are expressed as the time in seconds to complete the test (n Mockl = 4 and n TMBIM6 = 5). All bars represent mean ± SEM. In all tests, two-way ANOVA followed by Tukey’s multiple comparison test was performed. Statistical significance (p < 0.05) between samples is indicated in the figures as follows: *=p < 0.05; **=p < 0.01; ***=p < 0.001; ****=p < 0.0001.

Article Snippet: Using data on TMBIM6 mRNA expression levels from The Human Protein Atlas database (THPA) [ ], we confirmed through in silico analysis that TMBIM6 is expressed in various cortical regions of the human brain, with notably high expression in the midbrain and retina, two areas rich in DAergic cells [ , ] (Fig. ).

Techniques: In Vivo, Transduction, Expressing, Injection, Comparison