antihuman cd31 pecam 1 dylight 550  (Novus Biologicals)

Journal: Science Advances

Article Title: Restoring brain functions in the Mo-blo mouse model of Menkes disease with copper nanoclusters

doi: 10.1126/sciadv.aeg6972

Figure Lengend Snippet: ( A ) Representative images showing a Mo-blo mouse with light coat color (right) compared with a wild-type littermate with black fur (left) [at postnatal day 15 (PND15)]. ( B ) Mo-blo mice display abnormally coiled whiskers (➔). ( C ) Relative expression of Atp7a mRNA in brain tissue from Mo-blo versus wild-type mice, as measured by reverse transcription quantitative polymerase chain reaction (RT-qPCR; control, n = 13; Mo-blo, n = 20). ( D and E ) Ceruloplasmin protein levels and ferroxidase activity in plasma from PND21 mice assessed by Western blot (control, n = 24; and Mo-blo, n = 9) and colorimetric assay (control, n = 18; and Mo-blo, n = 8), respectively. ( F and G ) TYR protein levels (control, n = 20; and Mo-blo, n = 9) and melanin (via PTCA quantification) content (control, n = 26; and Mo-blo, n = 10) in skin and hair from PND21 mice, analyzed by Simple Western and HPLC. ( H ) Macroscopic image of a PND21 Mo-blo mouse after death due to an aortic aneurysm. ( I ) Kaplan-Meier survival curve showing increased mortality in Mo-blo mice (control, n = 20; and Mo-blo, n = 14). ( J and K ) Tissue (heart) (control, n = 22; and Mo-blo, n = 7) and circulating (plasma) (control, n = 17; and Mo-blo, n = 8) LOX protein levels in PND21 mice analyzed by Simple Western and Western blot, respectively. ( L and M ) COX protein levels (control, n = 16; and Mo-blo, n = 9) and complex IV activity (control, n = 14; and Mo-blo, n = 9) in the brain of PND21 mice, assessed by Western blot and high-resolution respirometry (or oxygraphy). Data are presented as means ± SEM. Statistical significance was determined by unpaired t test and is indicated as follows: * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001. n.s., not significant.

Article Snippet: The primary antibodies used (LOX, Novus NB100-25-27, 1:250; TYR, Novus NBP2-67233, 1:250) were diluted in a dilution solution supplied in the kit at specific concentrations depending on the development.

Techniques: Expressing, Tissue, Reverse Transcription, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Control, Activity Assay, Clinical Proteomics, Western Blot, Colorimetric Assay, Simple Western

Journal: Science Advances

Article Title: Restoring brain functions in the Mo-blo mouse model of Menkes disease with copper nanoclusters

doi: 10.1126/sciadv.aeg6972

Figure Lengend Snippet: ( A ) Phenotypic observations of coat and ( B ) whiskers (➔) of Mo-blo mice during treatment (CuNC) about the activity of the cupro-dependent proteins: HEPHL1, LOX, and TYR, respectively. Same picture as used for comparison with CuNC-treated mice. ( C ) Quantification of TYR by Simple Western and ( D ) microscopic observations of melanin pigment production in the hairs (×40) in correlation with ( E ) the analysis of PTCA pigment, the result of enzymatic TYR activity. ( F and G ) Quantifications of LOX expression (intracellular and secreted forms) were performed by Simple Western and Western blot, respectively. ( H ) Quantifications of ceruloplasmin activity were performed by colorimetric assay. The shaded values for untreated group refer to the results shown in . TYR and PTCA analyses were carried out on skin and hairs samples using the following numbers: control, n = 20; CuNCs, n = 20; and control, n = 26; CuNCs, n = 26, respectively. Data are presented as means ± SEM. Statistical significance was calculated by one-way ANOVA with Tukey’s test and ANOVA Welch test following Games-Howell’s test. n.s., not significant ( P > 0.05); *** P < 0.001; **** P < 0.0001. For LOX, analyses were carried out on heart (intracellular form) and plasma (secreted form), according to the following numbers: control, n = 25; CuNCs, n = 15; and control, n = 15; CuNCs, n = 15, respectively (biologically independent samples). Data are presented as means ± SEM. Statistical significance was calculated by ANOVA Welch test following Games-Howell’s and Kruskal-Wallis test with Dunn’s test. n.s., not significant ( P > 0.05). Ceruloplasmin activity analyses were carried out on plasma samples using the following numbers: control, n = 18; and CuNCs, n = 14. Data are presented as means ± SEM. Statistical significance was calculated by one-way ANOVA with Tukey’s test and ANOVA Welch test following Games-Howell’s test. n.s., not significant ( P > 0.05); ** P < 0.01; *** P < 0.001; **** P < 0.0001.

Article Snippet: The primary antibodies used (LOX, Novus NB100-25-27, 1:250; TYR, Novus NBP2-67233, 1:250) were diluted in a dilution solution supplied in the kit at specific concentrations depending on the development.

Techniques: Activity Assay, Comparison, Simple Western, Analysis, Expressing, Western Blot, Colorimetric Assay, Control, Clinical Proteomics

Journal: PLOS One

Article Title: AZP2006 in Parkinson’s disease models: A promising new neuroprotective treatment involving lysosomal homeostasis and the Progranulin/Prosaposin complex

doi: 10.1371/journal.pone.0359269

Figure Lengend Snippet: Body mass evolution during 35 days of daily treatment with vehicle or AZP2006 post-lesion (A). Representative IHC images TH (red)/α-syn (green)/Iba1 (purple) (B). IHC analysis shows the total number of TH(+) neurons (C), area of α-syn accumulation in TH(+) neurons (D) and the number of Iba1(+) microglia in the SNpc (E) expressed as % of sham. Raw values for sham (i.e., 100%); TH(+) neurons = 65 ± 1.6; α-syn area/TH(+) neuron = 168 ± 9.4 µm 2 ; Iba1(+) cells = 53 ± 2.8. Each bar represents the mean ± SEM, one-way ANOVA followed by Fisher’s test. ## p < 0.01 or #### p < 0.0001 from sham; **p < 0.01; ***p < 0.001 and ****p < 0.0001 from α-syn+vehicle injury, (n = 7 mice / group).

Article Snippet: Iba1 , Rabbit , Polyclonal , Novus , NBP2–19019 , AB_3073939 , .

Techniques: Analysis

Journal: PLOS One

Article Title: AZP2006 in Parkinson’s disease models: A promising new neuroprotective treatment involving lysosomal homeostasis and the Progranulin/Prosaposin complex

doi: 10.1371/journal.pone.0359269

Figure Lengend Snippet: Antibodies used for in vivo investigations.

Article Snippet: Iba1 , Rabbit , Polyclonal , Novus , NBP2–19019 , AB_3073939 , .

Techniques: In Vivo, Cell-Signaling

Journal: Aging Cell

Article Title: IGF ‐1/ SLC25A33 Maintains Blood‐Labyrinth Barrier Integrity by Suppressing TGF ‐β/Smad2‐Mediated Pericyte Activation in Age‐Related Hearing Loss

doi: 10.1111/acel.70712

Figure Lengend Snippet: Transcriptomic analysis identifies IGF‐1/SLC25A33 dysregulation and TGF‐β pathway activation in aged pericytes, which may be associated with ARHL. (A) Representative click‐evoked ABR waveforms showing a threshold of 30 dB SPL (red trace, left) in a young mouse versus 60 dB SPL (red trace, right) in an aged mouse. (B) Click‐evoked ABR thresholds are significantly elevated in old mice compared to young controls. (C) Frequency‐specific ABR thresholds are markedly increased across all tested frequencies (4–32 kHz) in aged mice. (D) DPOAE thresholds are significantly higher in old mice at F 2 = 17.44 kHz ( n = 6 mice). (E) RNA sequencing of FACS‐purified pericytes from young and aged stria vascularis reveals significant downregulation of Igf1 and Slc25a33 mRNA in aged mice ( n = 6 mice). (F) The most enriched DO terms for the differentially expressed genes when old vs. young. (G) The most enriched GO terms for the downregulated genes when old versus young. (H) The most enriched Reactome terms for the downregulated genes when old versus young. (I) KEGG pathway enrichment analysis highlights the TGF‐β signaling pathway as one of the most differentially activated pathways. (J) Gene–gene interaction network for the downregulated genes when old versus young.

Article Snippet: Sections were blocked in 10% goat serum + 0.3% Triton X‐100 in PBS for 1 h. Primary antibodies were applied overnight at 4°C: IGF‐1 (NOVUS, SPM406, 1:200), SLC25A33 (Bioss, bs‐21231R, 1:200), PDGFRβ (CST, #3169, 1:200), E‐cadherin (CST, #14472, 1:200), α‐SMA (Bioss, bs‐10196R, 1:200), TGFβ1 (Bioworld, BS80030, 1:50), p‐Smad2 (Bioworld, bs‐3419R, 1:500).

Techniques: Analysis, Activation Assay, RNA Sequencing, Purification

Journal: Aging Cell

Article Title: IGF ‐1/ SLC25A33 Maintains Blood‐Labyrinth Barrier Integrity by Suppressing TGF ‐β/Smad2‐Mediated Pericyte Activation in Age‐Related Hearing Loss

doi: 10.1111/acel.70712

Figure Lengend Snippet: Downregulation of IGF‐1 and SLC25A33 are confirmed in the aged stria vascularis. In vitro studies reveal IGF‐1 regulates SLC25A33 expression. (A, B) qPCR shows downregulation of Igf1 and Slc25a33 mRNA expression in aged pericytes ( n = 3 mice); (C) Immunofluorescence staining confirms colocalization of IGF‐1 (green), SLC25A33 (red) and PDGFRβ (purple, pericyte marker) in the stria vascularis. Age‐dependent alteration of IGF‐1 and SLC25A33 were markedly occurred in pericytes, as indicated by arrows. (D‐E) Quantitative analysis reveals significant downregulation of both IGF‐1 and SLC25A33 fluorescence intensity in aged mice relative to young controls ( n = 5, five typical sections from five individual mice were analyzed per group). (F) qPCR shows that inhibition of IGF‐1 signaling (via NVP‐AEW541) significantly reduces Slc25a33 mRNA levels. (G) Representative Western blot of SLC25A33 protein expression. (H) Densitometric quantification demonstrates reduced SLC25A33 protein upon IGF‐1 inhibition. Control: HBVPs without treatment; IGF‐1 (−): HBVPs treated with NVP‐AEW541; IGF‐1 (+): Cells treated with recombinant IGF‐1. Data are mean ± SEM; n = 3 mice.

Article Snippet: Sections were blocked in 10% goat serum + 0.3% Triton X‐100 in PBS for 1 h. Primary antibodies were applied overnight at 4°C: IGF‐1 (NOVUS, SPM406, 1:200), SLC25A33 (Bioss, bs‐21231R, 1:200), PDGFRβ (CST, #3169, 1:200), E‐cadherin (CST, #14472, 1:200), α‐SMA (Bioss, bs‐10196R, 1:200), TGFβ1 (Bioworld, BS80030, 1:50), p‐Smad2 (Bioworld, bs‐3419R, 1:500).

Techniques: In Vitro, Expressing, Immunofluorescence, Staining, Marker, Analysis, Fluorescence, Inhibition, Western Blot, Control, Recombinant

Journal: Aging Cell

Article Title: IGF ‐1/ SLC25A33 Maintains Blood‐Labyrinth Barrier Integrity by Suppressing TGF ‐β/Smad2‐Mediated Pericyte Activation in Age‐Related Hearing Loss

doi: 10.1111/acel.70712

Figure Lengend Snippet: IGF‐1 upregulation promotes SLC25A33 expression in the stria vascularis of aged mice. (A) Immunofluorescence images show IGF‐1 (green) and SLC25A33 (red) expression in the stria vascularis. (B, C) Quantitative analysis reveals that exogenous IGF‐1 supplementation increases both IGF‐1 and SLC25A33 fluorescence intensity, while IGF‐1 inhibition further reduces their expression ( n = 5, five typical sections from five individual mice were analyzed per group). (D) qPCR confirms exogenous IGF‐1 supplementation upregulates Slc25a33 mRNA levels, whereas IGF‐1 inhibition decreases its expression ( n = 3 mice). Control: wild‐type C57BL/6J mice without treatment; IGF‐1 (−): mice treated with NVP‐AEW541; IGF‐1 (+): mice treated with recombinant IGF‐1. Data are mean ± SEM.

Article Snippet: Sections were blocked in 10% goat serum + 0.3% Triton X‐100 in PBS for 1 h. Primary antibodies were applied overnight at 4°C: IGF‐1 (NOVUS, SPM406, 1:200), SLC25A33 (Bioss, bs‐21231R, 1:200), PDGFRβ (CST, #3169, 1:200), E‐cadherin (CST, #14472, 1:200), α‐SMA (Bioss, bs‐10196R, 1:200), TGFβ1 (Bioworld, BS80030, 1:50), p‐Smad2 (Bioworld, bs‐3419R, 1:500).

Techniques: Expressing, Immunofluorescence, Analysis, Fluorescence, Inhibition, Control, Recombinant

Journal: Aging Cell

Article Title: IGF ‐1/ SLC25A33 Maintains Blood‐Labyrinth Barrier Integrity by Suppressing TGF ‐β/Smad2‐Mediated Pericyte Activation in Age‐Related Hearing Loss

doi: 10.1111/acel.70712

Figure Lengend Snippet: Exogenous IGF‐1 rescues blood‐labyrinth barrier integrity and improves hearing function in aged mice. (A) TEM images reveal ultrastructural changes in the stria vascularis. In the IGF‐1‐inhibited group, pericytes exhibit abnormal morphology, loose association with endothelial cells, and widened tight junctions (yellow arrows). These defects are ameliorated by exogenous IGF‐1 administration. (B) Click‐evoked ABR thresholds are significantly elevated in IGF‐1‐suppressed mice but partially rescued by IGF‐1 supplementation. (C–E) At 12 kHz, ABR thresholds are significantly lower in the IGF‐1‐treated group compared to control and suppressed groups, indicating improved auditory sensitivity. (F) Proposed model of IGF‐1/SLC25A33‐mediated regulation of pericyte phenotype and BLB integrity in age‐related hearing loss. Under physiological conditions, pericytes maintain a quiescent state characterized by low α‐SMA and high E‐cadherin expression, contributing to BLB stability. With aging, declining IGF‐1 levels lead to reduced SLC25A33 expression, resulting in mtROS accumulation and activation of the TGF‐β/Smad2 signaling pathway. This drives pericyte transition toward a contractile, pro‐fibrotic phenotype (high α‐SMA, low E‐cadherin), disrupting BLB integrity and promoting cochlear dysfunction. Targeting this axis may offer therapeutic potential for ARHL. Control: wild‐type C57BL/6J mice without treatment; IGF‐1 (−): mice treated with NVP‐AEW541; IGF‐1 (+): mice treated with recombinant IGF‐1. Data are mean ± SEM; n = 6 mice.

Article Snippet: Sections were blocked in 10% goat serum + 0.3% Triton X‐100 in PBS for 1 h. Primary antibodies were applied overnight at 4°C: IGF‐1 (NOVUS, SPM406, 1:200), SLC25A33 (Bioss, bs‐21231R, 1:200), PDGFRβ (CST, #3169, 1:200), E‐cadherin (CST, #14472, 1:200), α‐SMA (Bioss, bs‐10196R, 1:200), TGFβ1 (Bioworld, BS80030, 1:50), p‐Smad2 (Bioworld, bs‐3419R, 1:500).

Techniques: Morphology, Control, Sensitivity, Expressing, Stability, Activation Assay, Recombinant