Journal: Biomedicines

Article Title: Functional Characterization of the SCN5A p.D372H Variant Associated with Brugada Syndrome

doi: 10.3390/biomedicines14030582

Figure Lengend Snippet: The SCN5A p.D372H variant reduces Nav1.5 expression through decreased mRNA abundance and stability, independent of plasmid DNA levels. ( A ) Relative SCN5A mRNA expression measured by RT-qPCR. Transcript levels were significantly reduced in the D372H group compared with WT (**** p < 0.0001). Co-expression of WT and D372H (WT+MUT) also resulted in a significant decrease in SCN5A mRNA expression compared with WT (**** p < 0.0001). Data were normalized to WT. ( B ) Quantification of intracellular SCN5A plasmid DNA levels. No significant differences were observed among WT, WT+MUT, and MUT groups (ns), indicating comparable transfection efficiency. ( C ) SCN5A mRNA decay following transcriptional inhibition with actinomycin D. Relative mRNA remaining was normalized to the 0-h time point. The D372H group exhibited accelerated mRNA decay compared with WT, indicating reduced transcript stability. ( D ) Representative Western blot images showing total Nav1.5 protein expression in BLANK, CONTROL, WT, WT+MUT, and MUT groups. Molecular weight markers are indicated in kilodaltons (kDa). The predicted molecular weight of native Nav1.5 is approximately 227 kDa; however, in this study SCN5A was expressed as a GFP-tagged fusion protein, resulting in an apparent band migrating above 245 kDa. Tubulin (50 kDa) was used as a loading control. ( E ) Quantitative analysis of Nav1.5 protein expression normalized to Tubulin. Protein levels were significantly reduced in the D372H group compared with WT (* p < 0.05). The WT+MUT group also showed decreased expression compared with WT (** p < 0.01). Data are presented as mean ± SD from at least three independent experiments.

Article Snippet: The membrane was then incubated with primary antibodies against Nav1.5 (1:1000, Cat# 23016-1-AP, Proteintech, Wuhan, China) and Tubulin (1:000, Cat# A03989-1, Boster, Wuhan, China) to detect the target protein and serve as a loading control, respectively.

Techniques: Variant Assay, Expressing, Plasmid Preparation, Quantitative RT-PCR, Transfection, Inhibition, Western Blot, Control, Molecular Weight

Journal: International Journal of Molecular Sciences

Article Title: The Protective Role of Intermedin in Contrast-Induced Acute Kidney Injury: Enhancing Peritubular Capillary Endothelial Cell Adhesion and Integrity Through the cAMP/Rac1 Pathway

doi: 10.3390/ijms252011110

Figure Lengend Snippet: IMD activates the cAMP/Rac1 pathway and alleviates renal peritubular capillary injury in CIAKI rats. ( A , B ): The renal lysates of rats were collected and the expression of Rac1, VE-cadherin, VEGFR2, and GAPDH were detected using Western blotting. ( C ) Kidney sections were stained with CD34 immunohistochemical staining (magnification, ×400, scale 30 μm). ( D ) Kidney sections were stained with CD34 and ICAM1 immunofluorescence staining (magnification, ×200, scale 50 μm). ( E ) The average optical density of CD34 positive staining in each group was quantified ( n = 6). ( E ) Quantification of the average optical density of ICAM1 immunofluorescence ( n = 6). The data were analyzed using ANOVA, followed by Tukey’s multiple comparisons test for subgroup analysis. All data are expressed as mean ± SD, * p < 0.05 compared to the control group, # p < 0.05. For comparison between groups, ns: not significant.

Article Snippet: The antibodies used in this study were cd34 (BM4082, Boster, Wuhan, China) and ICAM1 (A00171, Boster, Wuhan, China).

Techniques: Expressing, Western Blot, Staining, Immunohistochemical staining, Immunofluorescence, Control, Comparison