Journal: Biofabrication

Article Title: Collagen hydrogel tube microbioreactors for cell and tissue manufacturing

doi: 10.1088/1758-5090/ae2718

Figure Lengend Snippet: Culturing human embryonic stem cells in ColTubes. (A) Phase-contrast images of H9 hESCs cultured in ColTubes at days 0, 1, 3, 5, and 7. (B) Live/Dead staining of H9 cells inside the tubes and released from the tubes on day 7. (C) Flow cytometry analysis shows that 99.6% of cells harvested on day 7 are Calcein AM-positive (live cells). (D) Flow cytometry analysis confirms that majority cells on day 7 express pluripotency markers OCT4 and Nanog.

Article Snippet: For a typical cell culture, H9 human embryonic stem cells (hESCs) (WA09, WiCell) loaded in 20 μ l ColTubes were suspended in 2 ml Essential 8 medium (Gibco) supplemented with 10 μ M Y-27632 (Sigma) in a 6-well plate and incubated at 37 °C with 5% CO2 and 21% O2.

Techniques: Cell Culture, Staining, Flow Cytometry

Journal: Biofabrication

Article Title: Collagen hydrogel tube microbioreactors for cell and tissue manufacturing

doi: 10.1088/1758-5090/ae2718

Figure Lengend Snippet: Differentiating H9 hESCs into Cardiomyocytes in ColTubes. (A) The cardiomyocyte production protocol. H9 hESCs are processed into ColTubes and expanded in E8 medium, followed by mesoderm induction for 1 d, cardiomyocyte differentiation from days 2–11, and metabolic enrichment from days 11–18. (B) Phase-contrast and fluorescent images of cells in ColTubes on days 0, 1, 3, 5, 7, 11, 15, and 18. Cardiomyocytes are cTnT-positive.

Article Snippet: For a typical cell culture, H9 human embryonic stem cells (hESCs) (WA09, WiCell) loaded in 20 μ l ColTubes were suspended in 2 ml Essential 8 medium (Gibco) supplemented with 10 μ M Y-27632 (Sigma) in a 6-well plate and incubated at 37 °C with 5% CO2 and 21% O2.

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: Generation of FHOD3-deficient hESC model. A Diagram of the FHOD3 knockout pattern, showing the gene editing position and deleted base pair. B Immunofluorescence staining of the human pluripotency markers SSEA4 and OCT4 in WT and FHOD3 KO hESCs. Scale bar, 75 μm. C QPCR analysis of SSEA4 and OCT4 expression in WT and FHOD3 KO hESCs, n = 3 per group. D Diagram of cardiomyocytes differentiation from hESCs by using small molecule-based methods. E Verification of FHDO3 knockout by WB, Full-length blots are presented in Supplementary Figure S3. F Flow cytometry and quantification of the myocardial-specific marker cardiac Troponin T (TNNT2) in WT and FHOD3 KO cardiomyocytes after 15days of differentiation, n = 3 per group. Data are represented as mean ± SEM; * P < 0.05, ** P < 0.01, *** P < 0.001. Data were analyzed via two-sample t test followed by the Bonferroni post hoc test

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques: Knock-Out, Immunofluorescence, Staining, Expressing, Flow Cytometry, Marker

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: FHOD3 deficiency results in cardiomyocytes abnormality and sarcomere disassembly. A Immunofluorescence staining of sarcomeric proteins cTNT and α-actinin to showed significant larger percentage of disrupted and disorganized sarcomeres in FHOD3 KO hESC-CMs (30 days of differentiation). Scale bar, 10 μm. B Transmission electron microscopy images of sarcomere structures in WT and FHOD3 KO hESC-CMs (30 days of differentiation) and quantification of abnormal sarcomeres. Scale bar, 100 nm. C , D Calibration of forward scatter (FSC) in flow cytometry (FSC;10000 cells/ sample) to shows the volume of cardiomyocytes (30 days of differentiation), n = 3 per group. E , F Immunofluorescence of phalloidin in WT and FHOD3 KO hESC-CMs (30 days of differentiation) to show the mean cell area, Scale bar, 50 μm. Data are represented as mean ± SEM, * P < 0.05, ** P < 0.01, *** P < 0.001. Data were analyzed via two-sample t test followed by the Bonferroni post hoc test

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques: Immunofluorescence, Staining, Transmission Assay, Electron Microscopy, Flow Cytometry

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: FHOD3 knockout cardiomyocytes exhibit compromised contractile functions. A Schematic diagram of contractility measurement in WT and FHOD3 KO hESC-CMs (30 days of differentiation). B The curve of contraction versus time detected by ‘MUSCLEMOTION’. C Quantification of contraction amplitude, time to peak, relaxation time and 90% contraction duration, n = 6 per group. D Representative Ca 2+ transient signals in WT and FHOD3 KO hESC-CMs (30 days of differentiation) measured by using Fluo-4 AM. E Waveform diagram of calcium transient. F Schematic diagram of calcium transient measurement indicators. G , H Ca 2+ transient induced by 10 mmol/L caffeine and waveform diagram. I Quantification of amplitude, time to peak value and 50% decay time of calcium transient in (D), n = 10 per group. J Quantification of amplitude, time to peak value and 50% decay time of calcium transient induced by 10 mmol/L caffeine in (G), n = 4 per group. Data are represented as mean ± SEM; * P < 0.05, ** P < 0.01, *** P < 0.001. Data were analyzed via two-sample t test followed by the Bonferroni post hoc test

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques: Knock-Out

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: Different transcriptome between WT and KO hESC-CMs (30 days of differentiation). A The volcano plot shows differentially expressed genes between two groups ( n = 3, respectively). Red denotes up-regulated genes, whereas green represents down-regulated genes. B KEGG Enrichment Scatter Plot dispalys the top 20 significant pathways. C KEGG Enrichment Chord Diagram displays the 10 Genes with the highest fold change (left) in the top 9 significant pathways (right). D The scatter plot of GO enrichment for differentially expressed genes. E QPCR analysis of the genes involved sarcomere structure and calcium handling pathways, normalized to IPO8, n = 3 per group

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: FHOD3 knockout hESC-CMs develop mitochondrial dysfunction. A Seahorse Mito Stress Test evaluating mitochondiral respiration in WT and KO hESC-CMs (30 days of differentiation), Left: real-time OCR profiles; Right: quantification of mitochondrial respiration parameters, n = 6 per group. B ATP production detection in WT and KO hESC-CMs (30 days of differentiation), n = 6 per group. C Representative fluorescence images of ROS levels in WT and KO hESC-CMs (30 days of differentiation) and quantification analysis, n = 7 per group. Enzymatic activity of individual ETC complexes I, IV and V assessed by using commercial assay kits (30 days of differentiation), n = 6 per group. Data are represented as mean ± SEM; * P < 0.05, ** P < 0.01, *** P < 0.001. Data were analyzed via two-sample t test followed by the Bonferroni post hoc test

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques: Knock-Out, Fluorescence, Activity Assay

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: FHOD3 deficiency activates calcium signaling pathway to promote heart failure progression. A Representative Western blot showing sarcomere structure proteins in WT and FHOD3 KO hESC-CMs (30 days of differentiation), Quantification shown below, Full-length blots are presented in Supplementary Figure S5. B Representative Western blot showing calcium related proteins in WT and FHOD3 KO hESC-CMs (30 days of differentiation), Quantification shown below, Full-length blots are presented in Supplementary Figure S6. C QPCR analysis of cardiac remodeling markers reglated by the CAMKII downstream effectors in WT and FHOD3 KO hESC-CMs (30 days of differentiation), n = 3 per group.Data are represented as mean ± SEM; * P < 0.05, ** P < 0.01, *** P < 0.001. Data were analyzed via two-sample t test followed by the Bonferroni post hoc test

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques: Western Blot

Journal: Stem Cell Research & Therapy

Article Title: FHOD3 deficiency disrupts sarcomere organization and activates caMKII signaling in human stem cell-derived cardiomyocytes

doi: 10.1186/s13287-026-04902-z

Figure Lengend Snippet: Candidate myosin activator OM rescues myocardial contractile dysfunction caused by FHOD3 deficiency. A The curve of contraction versus time detected by ‘MUSCLEMOTION’. B Quantification of contraction amplitude, time to peak, relaxation time and 90% contraction duration in WT and FHOD3 KO hESC-CMs (30 days of differentiation), n = 8 per group. C Representative Ca 2+ transient signals in WT and FHOD3 KO hESC-CMs (30 days of differentiation) measured by using Fluo-4 AM, and waveform diagram of calcium transient. D Quantification of amplitude, time to peak value and 50% decay time of calcium transient in (C), n = 10 per group. Data are represented as mean ± SEM; * P < 0.05, ** P < 0.01, *** P < 0.001. Data were analyzed via one-way ANOVA followed by the Bonferroni post hoc test

Article Snippet: Human embryonic stem cell (hESC) line H9 (WA09, WiCell) was used in this study.

Techniques: