anti rtn4 nogo b  (Proteintech)

Journal: Cell Death & Disease

Article Title: Identification of Nogo-B as a potential therapeutic target of osteosarcoma via stereochemically selective covalent probes

doi: 10.1038/s41419-025-07765-z

Figure Lengend Snippet: a Nogo-B knockdown to verify experiment. b Nogo-B overexpression to verify experiment. c Cells migration of shControl, shNogo-B2 and shNogo-B3 group were tested by cell wound healing assay. d Quantitative results of wound healing assay. e Cells migration of NC and OE-Nogo-B group were tested by cell wound healing assay. f Quantitative results of wound healing assay. g Cells proliferation of shControl, shNogo-B2 and shNogo-B3 group were tested by colony formation assay. h Quantitative results of colony formation assay. i Cells proliferation of NC and OE-Nogo-B group were tested by colony formation assay. j Quantitative results of colony formation assay. k The effect of Nogo-B knockdown and overexpression on 143B cell activity was detected by CCK-8 assay. l Analysis of the cytotoxicity of treatment with ( S , R )- 4v in shNogo-B3 and OE-Nogo-B 143B cells. m - p Examination on tumors formed by 143B and shNogo-B3 cells in Balb/c mice: m Representative picture of tumors derived from mice. n Tumor weight. o Tumor volume. p Average body weights. The data are reported as the mean ± SD. All data are representative of three independent experiments. Ns=no significant, ** P < 0.01; *** P < 0.005; **** P < 0.001.

Article Snippet: Human Nogo-B overexpression plasmid was designed and constructed by Genechem Company (Shanghai, China).

Techniques: Knockdown, Over Expression, Migration, Wound Healing Assay, Colony Assay, Activity Assay, CCK-8 Assay, Derivative Assay

Journal: Nature Communications

Article Title: Intestinal Nogo-B reduces GLP1 levels by binding to proglucagon on the endoplasmic reticulum to inhibit PCSK1 cleavage

doi: 10.1038/s41467-024-51352-3

Figure Lengend Snippet: Volcano plot depicting differential transcriptome in the liver ( a ), pancreas ( b ), and small intestine ( c ) of db/db mice. Red dots represent up-regulated genes, while black dots represent down-regulated genes. d Identification of candidate genes upregulated in the liver, pancreas, and small intestine of T2DM mice. e Experimental design for siRNA injection in vivo. db/db mice were injected with scrambled siRNA or Nogo-B siRNA, and db/m mice were injected with 0.9% normal saline as the healthy control. f Body weight of mice (n = 6 mice per group). g Fasted glucose levels of mice (n = 6 mice per group). h – j Serum levels of insulin, glucagon and GLP1 of mice (n = 6 mice per group). k Oral glucose tolerance test (OGGT) and area under the curve (AUC) analysis of mice (n = 6). l Insulin tolerance test (ITT) and AUC analysis of mice (n = 6 mice per group). m Glucose stimulated insulin secretion (GSIS) test and AUC analysis of mice (n = 6 mice per group). n pINSR, INSR, pIRS1, IRS1, pIRS2, IRS2, pAKT and AKT protein levels in mouse liver (n = 6 mice per group). o Hepatic glucose production (HGP) under basal and clamp conditions of mice (n = 3 mice per group). HGP suppression ( p ), glucose infusion rate (GIR, q ),and glucose disposal rate (GDR, r ) of mice (n = 3 mice per group). Data are expressed as the mean ± SEM. The p values of ( a – c ) were calculated by two-sided test and adjusted for p values by the Benjamini-Hochberg method to control for false positives due to multiple comparisons. The p values of ( f – l ) were calculated by one-way ANOVAs. The p values of ( m , o – r ) were calculated by two-tailed Student’s t test. The p values of ( f , g , k , l ) are indicated as db/db-KD group vs. db/db group. Source data are provided as a file.

Article Snippet: Both scrambled siRNA and Nogo-B siRNA were synthesized by GENEWIZ (Suzhou, China).

Techniques: Injection, In Vivo, Saline, Control, Two Tailed Test

Journal: Nature Communications

Article Title: Intestinal Nogo-B reduces GLP1 levels by binding to proglucagon on the endoplasmic reticulum to inhibit PCSK1 cleavage

doi: 10.1038/s41467-024-51352-3

Figure Lengend Snippet: a Tissue-specific post-translational modifications of glucagon and GLP1, and the strategy for the co-IP experiments. h in “h-Nogo-B, h-proGCG and h-insulin” means human-derived. b Immunoprecipitation assay with Nogo-B and exogenous insulin in HEK293T cells (n = 3 biological replicates). c , d Immunoprecipitation assay with Nogo-B and exogenous proGCG in HEK293T cells (n = 3 biological replicates). e Construction of EGFP-tagged expression vectors for proGCG-sheared basic short peptides (GRPP, glucagon, GLP1, and GLP2). f – i Immunoprecipitation assay with Nogo-B and proGCG-sheared basic short peptides (GRPP, glucagon, GLP1, and GLP2) (n = 3 biological replicates). j Construction of EGFP-tagged expression vectors for glicentin (GLI) and major proglucagon fragment (MPGF). k , l Immunoprecipitation assay with Nogo-B and GLI and MPGF (n = 3 biological replicates). m Mass spectrometric detection of the peptide bound to MPGF by Nogo-B, where the peptide with the highest secondary structure similarity to the profile was in the IP2 region. Construction of EGFP-tagged expression vectors for the MPFG with mutations in the hydrophobic amino acid site. n – p Immunoprecipitation assay with Nogo-B and MPFG with mutations in the hydrophobic amino acid site (n = 3 biological replicates). q ELISA for the binding of Nogo-B to proGCG. r Non-linear fit analysis of ELISA results (n = 6 biological replicates). s Affinity constants of Nogo-B and proGCG detected by SPR. All experiments of ( b – d , f – I , k , l , n – p ) were repeated at least three times Data are expressed as the mean ± SEM. Source data are provided as a file.

Article Snippet: Both scrambled siRNA and Nogo-B siRNA were synthesized by GENEWIZ (Suzhou, China).

Techniques: Co-Immunoprecipitation Assay, Derivative Assay, Immunoprecipitation, Expressing, Enzyme-linked Immunosorbent Assay, Binding Assay

Journal: Nature Communications

Article Title: Intestinal Nogo-B reduces GLP1 levels by binding to proglucagon on the endoplasmic reticulum to inhibit PCSK1 cleavage

doi: 10.1038/s41467-024-51352-3

Figure Lengend Snippet: a Representative images of immunofluorescence staining (ER, green; proGCG, red; DAPI, blue) of HEK293T cells transfected with human-derived proGCG and PCSK1. Overexpression of Nogo-B detects the localization of proGCG on the ER. I-II: The intensity distribution of the marker scans in the boxed area shows the spatial distribution and fluorescence intensity of the different markers. Enhanced co-localization is indicated by the resemblance in shape of the intensity distribution curves and the increased number of overlaps. The experiment was conducted independently on three occasions. b Detection of Nogo-B overexpression on exogenous PCSK1 cleavage of exogenous proGCG by Western blot in HEK293T cells (n = 3 biological replicates). c Endogenous co-IP assay with Nogo-B and proGCG in STC-1 cells (n = 3 biological replicates). d Representative images of immunofluorescence staining (PCSK1, green; proGCG, red; DAPI, blue) of STC-1 cells transfected with scrambled siRNA (si-Ctrl) or Nogo-B siRNA (si-Nogo-B) to detect co-localization of proGCG and PCSK1. I-II: The intensity distribution of the marker scans in the boxed area shows the spatial distribution and fluorescence intensity of the different markers. Enhanced co-localization is indicated by the resemblance in shape of the intensity distribution curves and the increased number of overlaps. The experiment was conducted independently on three occasions. e GLP1 levels in the culture medium of STC-1 cells transfected with si-Ctrl or si-Nogo-B (n = 5 biological replicates). f PCSK1, proGCG, GLP1 and Nogo-B protein levels in STC-1 cells transfected with si-Ctrl or si-Nogo-B (n = 6 biological replicates). g Nogo-B, PCSK1, PCSK2 and proGCG mRNA levels in STC-1 cells transfected with si-Ctrl or si-Nogo-B (n = 5 biological replicates). All experiments were repeated at least three times. Data are expressed as the mean ± SEM. The p values were calculated by two-tailed Student’s t test. Source data are provided as a file.

Article Snippet: Both scrambled siRNA and Nogo-B siRNA were synthesized by GENEWIZ (Suzhou, China).

Techniques: Immunofluorescence, Staining, Transfection, Derivative Assay, Over Expression, Marker, Fluorescence, Western Blot, Co-Immunoprecipitation Assay, Two Tailed Test

Journal: Nature Communications

Article Title: Intestinal Nogo-B reduces GLP1 levels by binding to proglucagon on the endoplasmic reticulum to inhibit PCSK1 cleavage

doi: 10.1038/s41467-024-51352-3

Figure Lengend Snippet: a Representative images of immunohistochemical staining of Nogo-B in small intestine from patients with or without T2DM. Quantification of Nogo-B density from 5 individual images (n = 5 patients per group). b Representative images of immunofluorescence staining (PCSK1, green; proGCG, red; DAPI, blue) of small intestine from patients with or without T2DM. a – f The intensity distribution of the marker scans in the boxed area shows the spatial distribution and fluorescence intensity of the different markers. Enhanced co-localization is indicated by the resemblance in shape of the intensity distribution curves and the increased number of overlaps. The experiment was conducted independently on three occasions. c working model of Nogo-B affecting GLP1 levels. Data are expressed as the mean ± SEM. The p values were calculated by two-tailed Student’s t test. Source data are provided as a file.

Article Snippet: Both scrambled siRNA and Nogo-B siRNA were synthesized by GENEWIZ (Suzhou, China).

Techniques: Immunohistochemical staining, Staining, Immunofluorescence, Marker, Fluorescence, Two Tailed Test

Journal: Nature Communications

Article Title: Intestinal Nogo-B reduces GLP1 levels by binding to proglucagon on the endoplasmic reticulum to inhibit PCSK1 cleavage

doi: 10.1038/s41467-024-51352-3

Figure Lengend Snippet: This study demonstrates that the ER-resident protein, Nogo-B, interacts with the MPGF of proGCG to retain proglucagon within the ER, thereby inhibiting the PCSK1-mediated cleavage of proGCG in the Golgi. The knockout of intestinal Nogo-B in mice with T2DM results in elevated levels of GLP1 and insulin, while reducing glucagon levels. Consequently, this alleviates pancreatic injury and improves insulin resistance.

Article Snippet: Both scrambled siRNA and Nogo-B siRNA were synthesized by GENEWIZ (Suzhou, China).

Techniques: Knock-Out