Journal: International Journal of Molecular Sciences

Article Title: Identification of SCN5a p.C335R Variant in a Large Family with Dilated Cardiomyopathy and Conduction Disease

doi: 10.3390/ijms222312990

Figure Lengend Snippet: Identifying SCN5a p.C335R and TTN truncating variants by using precision diagnostic methods. ( A ) Patient’s pedigree showing co-segregation of an SCN5a C335R variant. The index patient (III. 3) was diagnosed with DCM and sick sinus syndrome at age 59 and underwent CRT-D implantation. She only carried the SCN5a C335R variant. Patient IV.1 was diagnosed with DCM and conduction disease at age 24. He carried both SCN5a and TTN truncating variants. ( B ) Masson Trichrome stained left ventricular endomyocardial biopsy of patient IV.1 with SCN5a and TTN truncating variants. Histopathological examination demonstrated extensive cardiac fibrosis in this patient with DCM and conduction disease. ( C ) Schematic representation of the Nav1.5 cardiac sodium channel. The variant is located in Ploop between S5 and S6 of Domain-I.

Article Snippet: Primary anti-human NaV1.5 antibodies (Alomone Labs, Jerusalem, Israel) and anti-Rad50 as a reference (Bethyl Laboratories Inc., Montgomery, AL, USA) were diluted in the same blocking solution and incubated with the membranes overnight.

Techniques: Diagnostic Assay, Variant Assay, Staining

Journal: International Journal of Molecular Sciences

Article Title: Identification of SCN5a p.C335R Variant in a Large Family with Dilated Cardiomyopathy and Conduction Disease

doi: 10.3390/ijms222312990

Figure Lengend Snippet: SCN5a p.C335R carriers show conduction disease and reduced left ventricular ejection fraction (LV-EF). All family members with SCN5a p.C335R variant showed significantly longer PQ ( A ) and QRS intervals ( B ), lower left ventricular EF ( C ), and higher atrial fibrillation (AF) rates ( D ) than other family members without this variant. All 4 family members carrying SCN5a p.C335R who received CRT-D were non-responders ( E , F ). T-Test; * = p ≤ 0.05, ** = p ≤ 0.01. Data are presented as mean ± SD.

Article Snippet: Primary anti-human NaV1.5 antibodies (Alomone Labs, Jerusalem, Israel) and anti-Rad50 as a reference (Bethyl Laboratories Inc., Montgomery, AL, USA) were diluted in the same blocking solution and incubated with the membranes overnight.

Techniques: Variant Assay

Journal: International Journal of Molecular Sciences

Article Title: Identification of SCN5a p.C335R Variant in a Large Family with Dilated Cardiomyopathy and Conduction Disease

doi: 10.3390/ijms222312990

Figure Lengend Snippet: Functional analysis of SCN5a p.C335R variant. ( A ) Na + current traces from Nav1.5 wild-type and p.C335R transfected Xenopus oocytes. This showed a loss of function in Nav1.5 p. C335R. Sodium channel currents (INa) were evoked from −80 mV to 70 mV. Western blot analysis confirmed that the wild-type and mutated Nav1.5 were expressed and that loss of Nav1.5 function was not due to absent expression of the protein. Anti-Rad50 used as a reference. ( B , C ) Representative traces of INa in iPSC-CMs with wild-type and SCN5a p.C335R variant. Sodium channel currents (n = 9) were reduced in iPSC-CMs of the DCM patients.

Article Snippet: Primary anti-human NaV1.5 antibodies (Alomone Labs, Jerusalem, Israel) and anti-Rad50 as a reference (Bethyl Laboratories Inc., Montgomery, AL, USA) were diluted in the same blocking solution and incubated with the membranes overnight.

Techniques: Functional Assay, Variant Assay, Transfection, Western Blot, Expressing

Journal: Cells

Article Title: Short-Term Autophagy Preconditioning Upregulates the Expression of COX2 and PGE2 and Alters the Immune Phenotype of Human Adipose-Derived Stem Cells In Vitro

doi: 10.3390/cells11091376

Figure Lengend Snippet: Autophagy preconditioning in hASCs does not alter self-renewal capacity or surface expression of MSC markers. ( A ) Representative images of hASCs morphology in culture after both short= and long-term exposure to autophagy preconditioning agents. Scale bar = 300 µm. Immunophenotype of Rapa-ASCs ( B ) and 3MA-ASCs ( C ) by flow cytometric analysis of surface proteins. Data are presented as mean ± SEM of 3 independent experiments. Quantifaction and images of colony-forming units by Rapa-ASCs ( D ) and 3MA-ASCs ( E ). Data are presented as mean ± SEM of 4 independent experiments. All data comparing 3 groups are analyzed by using one-way analysis of variance (ANOVA) with Tukey’s post-hoc multiple comparisons, and CFU-F data are analyzed with unpaired student’s t -test. Abbreviations: Rapa, Rapamycin; 3-MA, 3-methyladenine; CFU-F, colony forming units-fibroblasts. ** p < 0.01.

Article Snippet: Primary human ASCs (hASCs) were purchased from LaCell LLC (New Orleans, LA, USA).

Techniques: Expressing

Journal: Cells

Article Title: Short-Term Autophagy Preconditioning Upregulates the Expression of COX2 and PGE2 and Alters the Immune Phenotype of Human Adipose-Derived Stem Cells In Vitro

doi: 10.3390/cells11091376

Figure Lengend Snippet: Autophagy preconditioning in hASCs alters expression of both anti-inflammatory and pro-inflammatory mediators. Rapa-ASCs (left) and 3MA-ASCs (right) were treated for 4 or 24 h, then relative expression levels of anti-inflammatory ( A ) and pro-inflammatory ( B ) genes were measured via RT-qPCR by using the ΔΔCt method. Data are presented as mean relative fold-change ± SEM of 4 independent experiments. ( C ) Secreted PGE2 levels were measured via ELISA in 24 h CM from control and autophagy preconditioned hASCs. Data are presented as means ± SEM of 3 independent experiments. Statistical analysis was performed by using one-way analysis of variance (ANOVA), and differences between the means are indicated with * p < 0.05, ** p < 0.01, *** p < 0.001. Abbreviations: TGF- β , transforming growth factor-beta; IDO , indoleamine 2,3-dioxygenase; TSG-6 , TNF Alpha Induced Protein 6, COX2 , cyclooxygenase 2; IL-6 , interleukin-6; IL-1 β , interleukin-1 beta; PGE2, prostaglandin E2.

Article Snippet: Primary human ASCs (hASCs) were purchased from LaCell LLC (New Orleans, LA, USA).

Techniques: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Control

Journal: Cells

Article Title: Short-Term Autophagy Preconditioning Upregulates the Expression of COX2 and PGE2 and Alters the Immune Phenotype of Human Adipose-Derived Stem Cells In Vitro

doi: 10.3390/cells11091376

Figure Lengend Snippet: Autophagy preconditioning in hASCs alters their transcriptional and secretory response to pro-inflammatory stimulation with hIFNγ. Rapa-ASCs (left) and 3MA-ASCs (right) were preconditioned for 4 or 24 h with their respective autophagy compounds, then stimulated with hIFNγ (5 ng/mL) for 24 h. Following stimulation, anti-inflammatory ( A ) and pro-inflammatory ( B ) genes were measured by using RT-qPCR. Data are presented as means ± SEM of 4 independent experiments and normalized to controls that were not treated with hIFNγ. ( C ) Secreted PGE2 levels were measured in 24 h CM from autophagy-preconditioned and hIFNγ-stimulated hASCs. Data are presented as means ± SEM of 3 independent experiments. All statistical significance is determined by using one-way analysis of variance (ANOVA). Statistical differences between the means are indicated with * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Abbreviations: TGF-β, transforming growth factor-beta; IDO, indoleamine 2,3-dioxygenase; TSG-6, TNF Alpha Induced Protein 6, COX2, cyclooxygenase 2; IL-6, IL-1β, interleukin-6; interleukin-1 beta; PGE2, prostaglandin E2.

Article Snippet: Primary human ASCs (hASCs) were purchased from LaCell LLC (New Orleans, LA, USA).

Techniques: Quantitative RT-PCR