Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 1. Transgenic overexpression of human thymosin beta 4 in rat hearts for three months. (a–c) The micrographs are low power images from normal control (left), DsRed control (middle) and TB4 treated (right) (scale bar: 150 mm). There is sparse TB4 signal in the epicardium of controls, with robust signal in the TB4-treated group. (d–f) The middle panels are higher power images (scale bar: 20 mm) from a TB4-treated rat showing antibodies against TB4 (left panel, red). (g) The graphs at the left bottom of the figure show the results of qRT–PCR for human TB4 gene mRNA at 1, 2, 3, 4 and 12 weeks after UTMD treatment, **Po0.001 vs normal control or UTMD-DsRed; Error bars represent mean±s.e.m. Number of hearts analyzed for each group ¼ 6. (h) The graphs at the right bottom of the figure show the results of RT–PCR for human TB4 and rat TB4 mRNA. Overexpression of exogenous human TB4 mRNA do not have any crossing reaction with endogenous rat TB4 mRNA.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Transgenic Assay, Over Expression, Control, Quantitative RT-PCR, Reverse Transcription Polymerase Chain Reaction

Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 2. Exogenous TB4 reprogrammed epicardium layer cells into WT1-positive adult cardiac progenitor cells and then formed new cardiac muscle cells. (a) Negative control for 12 weeks post-UTMD-DsRed. (b–f) The panels are high power images (scale bar: 50 mm) from TB4-treated rats killed at 1, 2, 3, 4 and 12 weeks after UTMD. WT1 signal from nucleus of epicardium layer cells at 1–3 weeks after UTMD-TB4 treatment and then migrated into myocardium layer to form new cardiac muscle cells (WT1 signal from nucleus of cTnT-positive cells, arrows) at 4 or 12 weeks after UTMD-TB4 treatment.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Negative Control

Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 3. Microscopic images showing evidence of angiogenesis after TB4 gene therapy by UTMD (scale bar ¼ 150 mm). (a–c): The top panels are stained antibodies against smooth muscle a-actin (SMaA) (red) and nucleus (blue). Left is normal control, middle is DsRed control, and right is TB4 treated. There is an increase in SMaA in the treatment group, indicating coronary arteriogenesis. (d–f) The middle panels are similar except the antibody is against CD31. (g,h):The graphs at the bottom show quantitative assay for SMaA-positive arteriolar density (left) and CD31-positive capillary density(right), *Po0.01 vs normal control or UTMD-DsRed; Error bars represent mean±s.e.m. Number of hearts analyzed for each group: N ¼ 6.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Staining, Control

Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 4. Images stained with antibodies against NKX2.5 (red), cTnT (green) and nucleus (blue). (a–c) The top panels are high power images from normal controls (left), DsRed controls (middle) and TB4-treated rats (right) (scale bar ¼ 10 mm). There is colocalization of NKX2.5 and Dapi in cTnT-positive cells in the rats treated with TB4 by UTMD, suggesting that these are cardiomyocyte progenitors (arrows). (d–f) The middle panels are high power images (scale bar ¼ 10 mm) from the TB4 treatment group showing anti-NKX2.5 (red, left), anti-nuclear DAPI (blue, middle), and colocalization (right) within the nucleus. The right panel is also stained with anti-cTnT(arrows). (g): The graphs at bottom show qRT–PCR for rat NKX2.5 gene mRNA, expressed as fold increase (left) at 1, 2, 3, 4 and 12 weeks after UTMD treatment. (h) Percentage of NKX2.5-positive cells in cTnT-positive cells. **Po0.001 vs normal control or UTMD-DsRed; Error bars represent mean±s.e.m. Number of hearts analyzed for each group: N ¼ 6.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Staining, Quantitative RT-PCR, Control

Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 5. Images stained with antibodies against ISL-1 (red), cTnT (green) and nucleus (blue). (a–c) The top panels are high power images from normal controls (left), DsRed controls (middle) and TB4-treated rats (right). There is colocalization of ISL-1 and Dapi in cTnT-positive cells in the rats treated with TB4 by UTMD, suggesting that these are cardiomyocyte progenitors (arrows). (d–f): The middle panels are high power images (scale bar ¼ 10 mm) from the TB4 treatment group showing anti-ISL-1 (red, left), anti-nuclear DAPI (blue, middle) and colocalization (right) within the nucleus. The right panel is also stained with anti-cTnT. (g) The graphs at bottom show qRT–PCR for rat ISL-1 gene mRNA, expressed as fold increase (left) at 1, 2, 3, 4 and 12 weeks after UTMD treatment. (h) Percentage of ISL-1-positive cells in cTnT-positive cells. **Po0.001 vs normal control or UTMD-DsRed; Error bars represent mean±s.e.m. Number of hearts analyzed for each group: N ¼ 6.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Staining, Quantitative RT-PCR, Control

Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 6. Adult cardiac progenitor cells or new formed cardiac muscle cells in the proliferation. (a–c) The top panels are high power images from normal controls (left), DsRed controls (middle) and TB4-treated rats (right) (scale bar ¼ 10 mm). There is colocalization of BrDu and Dapi in cTnT-positive cells in the rats treated with TB4 by UTMD, suggesting that these cells are proliferating(arrows). (d–f) The panels are high power images from normal controls (left), DsRed controls (middle) and TB4-treated rats (right). There is colocalization of Ki-67 and Dapi in cTnT- positive cells in the rats treated with TB4 by UTMD, suggesting that these cells are proliferating (arrows). (g–h) The graphs at bottom show percentage of BrDU-positive cells in cTnT-positive cells(left) and percentage of Ki-67-positive cells in cTnT-positive cells(right) at 1, 2, 3, 4 and 12 wks after UTMD treatment. **Po0.001 vs normal control or UTMD-DsRed; Error bars represent mean±s.e.m. Number of hearts analyzed for each group: N ¼ 6.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Control

Journal: Gene therapy

Article Title: Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery.

doi: 10.1038/gt.2012.89

Figure Lengend Snippet: Figure 7. These new formed cardiac muscle cells in mitosis condition. (a–c) Images stained with antibodies against the cell mitosis marker PHH3 (red), cTnT (green) and nucleus (blue). The top panels are high power images (scale bar ¼ 10 mm) from normal controls (left), DsRed controls (middle) and TB4-treated rats (right). There is colocalization of PHH3 and Dapi in cTnT-positive cells in the rats treated with TB4 by UTMD, suggesting that these cells are in mitotic condition (arrows). (d–f) The middle panels are high power images (scale bar ¼ 10 mm) showing there is colocalization of aurora B and Dapi in cTnT-positive cells in the rats treated with TB4 by UTMD, suggesting that these cardiomyocytes are undergoing cytokinesis (arrows). (g–k) A larger magnification view of aurora B-positive cells displayed these new formed cardiomyocytes are undergoing mitosis (arrows). (l,m) The graphs at bottom show percentage of PHH3-positive cells in cTnT-positive cells(left) and percentage of aurora B-positive cells in cTnT-positive cells(right) at 1, 2, 3, 4 and 12 weeks after UTMD treatment. **Po0.001 vs normal control or UTMD-DsRed; Error bars represent mean±s.e.m. Number of hearts analyzed for each group: N ¼ 6.

Article Snippet: Human TB4 cDNA was purchased from Origene technologies (Rockville, MD, USA).

Techniques: Staining, Marker, Control

Journal: eLife

Article Title: Human embryonic lung epithelial tips are multipotent progenitors that can be expanded in vitro as long-term self-renewing organoids

doi: 10.7554/eLife.26575

Figure Lengend Snippet: ( A ) E13.5 mouse lung staining illustrating absence of SOX2 in SOX9 + tip cells. Green: SOX9; red: SOX2; white: ECAD. ( B ) Whole-mount staining of a 5 pcw human embryonic lung showing the primary branches and SOX2/SOX9 co-expression in the tips (boxed area in B is magnified in B’). Green: SOX9; red: SOX2; white: ECAD. ( C ) Confocal image of a 8 pcw human embryonic lung illustrating SOX2 expression in SOX9 + tip cells. Green: SOX9; red: SOX2. ( D ) qRT-PCR of microdissected tip and stalk compared to whole lung: SOX9 and SOX2 data confirm immunostainings. TBX4 is a mesenchymal marker. ID2 is another well-characterised mouse tip gene. Scale bars = 50 μm ( A, C ); 200 μm ( B ); 100 μm ( B’ ). DOI: http://dx.doi.org/10.7554/eLife.26575.010

Article Snippet: TBX4 , Thermo Fisher Scientific , # 4331182 , Hs00218515_m1.

Techniques: Staining, Expressing, Quantitative RT-PCR, Marker

Journal: eLife

Article Title: Human embryonic lung epithelial tips are multipotent progenitors that can be expanded in vitro as long-term self-renewing organoids

doi: 10.7554/eLife.26575

Figure Lengend Snippet: RT-PCR primers DOI: http://dx.doi.org/10.7554/eLife.26575.043

Article Snippet: TBX4 , Thermo Fisher Scientific , # 4331182 , Hs00218515_m1.

Techniques:

Journal: STAR Protocols

Article Title: Protocol for minicircle production for gene therapy without subsequent cleanup steps

doi: 10.1016/j.xpro.2025.103982

Figure Lengend Snippet: Gel electrophoresis of induction of mCherry-P2A- TBX4 minicircles with corresponding nanodrops Gel showing parental plasmid and two separate elutions of minicircle post-induction of mCherry-P2A-TBX4 showing clean productions. (A) shows gel electrophoresis of DNA ladder (Ladder), Parental Plasmid (PP) from preinduction miniprep and two separate elutions of minicircle (MC) from the GeneJet Maxiprep which correspond to minicircle 1 (MC1) and minicircle 2 (MC2). The minicircles are lower in size which correspond to successful induction and lack parental plasmid band noted demonstrating lack of contamination. (B) shows nanodrop quantifications from the two minicircle elutions. The minicircle elution corresponds the number on the nanodrop.

Article Snippet: TaqMan mouse Tbx4 probes , Thermo Scientific , Mm01299756_g1.

Techniques: Nucleic Acid Electrophoresis, Plasmid Preparation

Journal: STAR Protocols

Article Title: Protocol for minicircle production for gene therapy without subsequent cleanup steps

doi: 10.1016/j.xpro.2025.103982

Figure Lengend Snippet: qPCR results comparing mouse and human expression of TBX4 in NIH/3T3 cells NIH/3T3 cells were transfected with either Empty or mCherry-P2A-TBX4 minicircles and incubated for 72 h before RNA extraction and reverse transcription. Mouse and human TBX4 was analyzed in both Empty and mCherry-P2A-TBX4 minicircles compared to mouse beta-actin expression. Values above the line are p-values between compared conditions. Data are represented as mean ± SD. p -values were calculated using a two-tailed T-test. p < 0.05 was deemed significant.

Article Snippet: TaqMan mouse Tbx4 probes , Thermo Scientific , Mm01299756_g1.

Techniques: Expressing, Transfection, Incubation, RNA Extraction, Reverse Transcription, Two Tailed Test