Journal: PLoS ONE

Article Title: CXC-Type Chemokines Promote Myofibroblast Phenoconversion and Prostatic Fibrosis

doi: 10.1371/journal.pone.0049278

Figure Lengend Snippet: N1 immortalized prostate fibroblast cells were grown to 70% confluence and treated with 0 (blue diamonds), 2 (red squares), or 4 (green triangles) ng/ml TGF-β1, or 0 (blue diamonds), 10 pM (red squares), 100 pM (green triangles), or 1 nM (purple diamonds) CXCL12, CXCL8 or CXCL5 and assessed for transcription of the RPLPO (housekeeping gene), COL1, αSMA, or COL3 genes at 2–4 hr intervals over a 12 or 24 hour period. Cycle numbers to threshold were calculated by subtracting averaged untreated from averaged treated values and normalized to those of RPLPO. Transcript levels are expressed as 2ddCt (T-UT). All cells tested demonstrated significantly increased TGF-β1, COL1, αSMA, and COL3, and transcript levels above basal levels upon treatment with TGF-β1 or CXC-type chemokines.

Article Snippet: Gene-specific assays were Hs0016400_m1 for COL1, Hs00909449_m1 for αSMA, Hs00943809_ml for COL3, Hs00998130 for_m1 for TGF-β1, and Hs99999902_m1 for RPLPO (Applied Biosystems, Carlsbad, CA).

Techniques:

Journal: PLoS ONE

Article Title: CXC-Type Chemokines Promote Myofibroblast Phenoconversion and Prostatic Fibrosis

doi: 10.1371/journal.pone.0049278

Figure Lengend Snippet: Primary prostate stromal fibroblasts cultured from patient 0912 were grown to 70% confluence and treated with 0 (blue diamonds), 2 (red squares), or 4 (green triangles) ng/ml TGF-β1, or 0 (blue diamonds), 10 pM (red squares), 100 pM (green triangles), or 1 nM (purple diamonds) CXCL12, CXCL8 or CXCL5 and assessed for transcription of the RPLPO (housekeeping gene), COL1, αSMA, or COL3 genes at 2–4 hr intervals over a 12 or 24 hour period. Cycle numbers to threshold were calculated by subtracting averaged untreated from averaged treated values and normalized to those of RPLPO. Transcript levels are expressed as 2ddCt(T-UT). All cells tested demonstrated significantly increased TGF-β1, COL1, αSMA, and COL3, and transcript levels above basal levels upon treatment with TGF-β1 or CXC-type chemokines.

Article Snippet: Gene-specific assays were Hs0016400_m1 for COL1, Hs00909449_m1 for αSMA, Hs00943809_ml for COL3, Hs00998130 for_m1 for TGF-β1, and Hs99999902_m1 for RPLPO (Applied Biosystems, Carlsbad, CA).

Techniques: Cell Culture

Journal: Advanced Science

Article Title: Tough Gelatin Hydrogel for Tissue Engineering

doi: 10.1002/advs.202301665

Figure Lengend Snippet: a) Introduction to in vivo tendon repair experiments. b) Intraoperative images. c) H&E and Masson staining of tendon tissues at four and eight weeks. Blue arrows indicate curved fibers, yellow arrows indicate densely aligned fibers, n = 3. d) Pathological scoring based on H&E and Masson staining. * p < 0.05; * *p < 0.01; ** *p < 0.001, n = 3. e) Tensile strength of the repaired tendon at four and eight weeks. f) Expression of gene of col1a1, tnmd, mkx , mmp3 , and mmp13 in tendon tissue at eight weeks. *p = 0.0107; * *p < 0.01; ** *p < 0.001, n = 3. g,h) Polarized observation results and statistics of Sirius red staining. *p = 0.0313; * *p < 0.01; ** *p < 0.001. n = 3. i) Tendon immunofluorescence staining of COL3 (red), COL1 (green), and the nucleus (blue). j) Tendon immunofluorescence staining of YAP (green), TNMD (red), and the nucleus (blue). k) Healing efficiency of training and initial hydrogel (vs NC). * p < 0.05, n = 3. l) Statistics of tendon immunofluorescence staining of YAP and TNMD. * p < 0.05; ** *p < 0.001, n = 9. m) Different stages of tendon reconstruction. All scale bars indicate 200 µm.

Article Snippet: The tendon tissue was immunohistochemistry stained with COL1 (Proteintech), COL3 (Proteintech), YAP (BIOSS) and TNMD (BIOSS) to evaluate the tendon differentiation and collagen regeneration.

Techniques: In Vivo, Staining, Expressing, Immunofluorescence

Journal: Research Square

Article Title: Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications

doi: 10.21203/rs.3.rs-2631314/v1

Figure Lengend Snippet: Human skin fibroblasts were transfected with non-targeting siRNA (siCtrl) or Col3 siRNA (siCol3). After 96h, the efficiency and specificity of knockdown were confirmed by qPCR for col3a1 and col1a1 (A and B). Representative Second Harmonic Generation (SHG) images of collagen matrices from human fibroblast-derived matrices (FDM) prepared from fibroblast/ECM units treated with control and Col3-targeted siRNA (C and D). Collagen matrices were analyzed for total collagen amount (E) and collagen alignment (F). CT-FIRE was used to quantify collagen fiber number (G) straightness (H), width (I), and length (J) from SHG images. Data from representative experiments are presented (5 individual experiments performed with similar results). Error bars show mean ± SD (n = 5). Student unpaired t-tests: (*,p< 0.05; **,p < 0.01; ****, p<0.0001). Scale bar = 25 μm.

Article Snippet: Due to common cross-reactivity of collagen antibodies with other collagen types, the Col3 antibody (1330-01; Southern Biotech, Birmingham, Al) utilized was determined to be specific for Col3 based on immunoreactivity in young adult mouse (Col3 +/+ and Col3 −/− , 4 weeks old) dermis sections following the same protocol as human biopsies (see above).

Techniques: Transfection, Knockdown, Derivative Assay, Control

Journal: Research Square

Article Title: Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications

doi: 10.21203/rs.3.rs-2631314/v1

Figure Lengend Snippet: Three breast cancer cell lines, MCF-7, MDA-MB-453 and MDA-MB-231 were cultured on collagen matrices produced by control fibroblasts (siCtrl) or Col3-deficient fibroblasts (siCol3). Cells were fixed and stained with Ki67, a cell proliferation marker or active caspase 3, a cell apoptosis marker. Quantification of the percentages of Ki67 (A, B, C), and active caspase 3 positive cells (D, E, F). Error bars show mean±SD (n = 3 for ki67, n = 4 for active caspase 3). Paired Student t-tests: (*,p<0.05; **, p<0.01; ***,p<0.001).

Article Snippet: Due to common cross-reactivity of collagen antibodies with other collagen types, the Col3 antibody (1330-01; Southern Biotech, Birmingham, Al) utilized was determined to be specific for Col3 based on immunoreactivity in young adult mouse (Col3 +/+ and Col3 −/− , 4 weeks old) dermis sections following the same protocol as human biopsies (see above).

Techniques: Cell Culture, Produced, Control, Staining, Marker

Journal: Research Square

Article Title: Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications

doi: 10.21203/rs.3.rs-2631314/v1

Figure Lengend Snippet: Hematoxylin-Eosin images were used to identify invasive and noninvasive regions within the same human triple-negative breast cancer (TNBC) biopsy. (A-B) A representative image of a human TNBC biopsy was immunolabeled for Col1 (Red), Col3 (Green), and stained with the nuclear stain DRAQ5 (Blue) (B). Second harmonic generation microscopy (SHG, white; Cytokeratin, magenta; DRAQ5/nuclei, blue; C-D) images were combined with confocal fluorescent imaging (Col1, red; Col3, green; E-F). SHG images were used to measure fiber alignment (FFT aspect ratio, G). The positive immunoreactivity of Col1 and Col3 integrated density of in both invasive and noninvasive regions, and their ratio, was analyzed using ImageJ (H-J). N=23 tumors. Paired Student t-tests: (**p<0.01, ****p<0.0001). Scale bar = 4mm.

Article Snippet: Due to common cross-reactivity of collagen antibodies with other collagen types, the Col3 antibody (1330-01; Southern Biotech, Birmingham, Al) utilized was determined to be specific for Col3 based on immunoreactivity in young adult mouse (Col3 +/+ and Col3 −/− , 4 weeks old) dermis sections following the same protocol as human biopsies (see above).

Techniques: Immunolabeling, Staining, Microscopy, Imaging

Journal: Research Square

Article Title: Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications

doi: 10.21203/rs.3.rs-2631314/v1

Figure Lengend Snippet: A. Kaplan-Meier survival curves showing that Col3:Col1 high patients have significantly better prognosis than Col1:Col3 high patients in terms of DFS and PFS across the entire TCGA BRCA patient cohort (p<0.05). B. Patients with Col3:Col1 high basal tumors demonstrated significantly better DFS, PFS, DSS, and OS compared to those with Col1:Col3 high basal tumors (p<0.05).

Article Snippet: Due to common cross-reactivity of collagen antibodies with other collagen types, the Col3 antibody (1330-01; Southern Biotech, Birmingham, Al) utilized was determined to be specific for Col3 based on immunoreactivity in young adult mouse (Col3 +/+ and Col3 −/− , 4 weeks old) dermis sections following the same protocol as human biopsies (see above).

Techniques:

Journal: Research Square

Article Title: Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications

doi: 10.21203/rs.3.rs-2631314/v1

Figure Lengend Snippet: MCF-7 cells were grown in 3D-culture consisting of Matrigel supplemented with human recombinant Col1, Col3 or 50:50 Col1+Col3 for 10 days. Images were taken with confocal microscopy of nuclei (DRAQ5, magenta), E-cadherin (cyan, shown in inset only) and fibrillar collagen (SHG, white) on max projection images. Mid-sections of the images showed the apparition of lumens (insets) (A-C). Spheroids (green outlines) and single cells (red outlines) were identified using ImageJ (D-F). Quantification of the area of single cells normalized to the total nuclei area (G). Integrated density of collagen fibers was analyzed by ImageJ and normalized to the total nuclei area (H). Lumens were counted and normalized total spheroids (I). Error bars show mean ± SD (n = 3). Data from three independent experiments. One-way AVONAs followed by Tukey post-hoc tests: (*, p<0.05; **,p<0.01,). Scale bar= 100μm.

Article Snippet: Due to common cross-reactivity of collagen antibodies with other collagen types, the Col3 antibody (1330-01; Southern Biotech, Birmingham, Al) utilized was determined to be specific for Col3 based on immunoreactivity in young adult mouse (Col3 +/+ and Col3 −/− , 4 weeks old) dermis sections following the same protocol as human biopsies (see above).

Techniques: Recombinant, Confocal Microscopy

Journal: Research Square

Article Title: Tumor-restrictive type III collagen in the breast cancer microenvironment: prognostic and therapeutic implications

doi: 10.21203/rs.3.rs-2631314/v1

Figure Lengend Snippet: RNA was collected from 4T1 tumors in Col3 +/+ and Col3 +/− mice and analyzed for Col1 and Col3 expression. Fold changes to Col3 +/+ D0 samples were used to generate Col1:Col3 ratios (A). Tumor volume 14 days after marginal resection in Col3 +/+ and Col3 +/− mice (B). Hydrogels were prepared with murine TNBC-like breast cancer cells (4T1) and supplemented with 100 μg of human recombinant Col3 or acetic acid (Ctrl) prior to injection into mammary fat pads of BalbC mice and harvested after 28 days (C). Tumor sections were stained for DAPI (nuclei, blue) and Ki67 (proliferating nuclei, red, D) or active caspase 3 (apoptotic cells, red, E). Scale bar = 50 μm. Staining was quantified as % Ki67 positive nuclei (D) or total active caspase 3 staining (integrated density, E). Lung sections were stained with H&E (F-G) and gross pulmonary metastasis was quantified (H). Scale bar = 3mm. For tumor growth, 2-way ANOVA followed by Sidak’s post-hoc test; for Ki67, active caspase 3, and metastasis, unpaired student t-tests: (*,p<0.05; **,p<0.01).

Article Snippet: Due to common cross-reactivity of collagen antibodies with other collagen types, the Col3 antibody (1330-01; Southern Biotech, Birmingham, Al) utilized was determined to be specific for Col3 based on immunoreactivity in young adult mouse (Col3 +/+ and Col3 −/− , 4 weeks old) dermis sections following the same protocol as human biopsies (see above).

Techniques: Expressing, Recombinant, Injection, Staining

Journal: Journal of Nanobiotechnology

Article Title: High-throughput screening-based design of multifunctional natural polyphenol nano-vesicles to accelerate diabetic wound healing

doi: 10.1186/s12951-024-02950-2

Figure Lengend Snippet: Evaluation of the therapeutic efficacy after BPPNs treatment. ( A ) Effects of different preparations on SOD and MDA level on day 14. ( B ) Immunofluorescence staining of neovascularizationat the wound tissue with different treatments on day 14. α-SMA (green), CD31 (red), and DAPI (blue). Scale bar: 100 μm. ( C ) Quantitative analysis of the blood vessel density on day 14. ( D ) Corresponding immunohistochemistry staining of COX2, HO-1, Cyto C, VEGF and COL3 on day 14. Scale bar: 200 μm. E - I . Immunohistochemical quantification of COX2, HO-1, HIF-1α, VEGF and COL3, respectively. n = 5, * P < 0.05, ** P < 0.01, *** P < 0.005

Article Snippet: COL3 , IHC , #ER1906-50, Huabio, China.

Techniques: Drug discovery, Immunofluorescence, Staining, Immunohistochemistry, Immunohistochemical staining

Journal: Journal of Nanobiotechnology

Article Title: High-throughput screening-based design of multifunctional natural polyphenol nano-vesicles to accelerate diabetic wound healing

doi: 10.1186/s12951-024-02950-2

Figure Lengend Snippet: Primary antibodies were used in the study

Article Snippet: COL3 , IHC , #ER1906-50, Huabio, China.

Techniques: