Journal: International journal of molecular sciences

Article Title: Intrarectal Administration of Adelmidrol plus Hyaluronic Acid Gel Ameliorates Experimental Colitis in Mice and Inhibits Pro-Inflammatory Response in Ex Vivo Cultured Biopsies Derived from Ulcerative Colitis-Affected Patients.

doi: 10.3390/ijms25010165

Figure Lengend Snippet: Figure 3. Ade/Ha intrarectal administration reduces endotoxemia and oxidative stress without inter- fering with the PEA level and inhibiting HYAL-1 expression in the mice colon during DNBS-induced colitis. The figure shows the effect of the intrarectal administration of Ade/HA on endotoxemia (A) and IL-1β release (B) in the plasma affected by colitis on the 3rd and 7th day after DNBS chal- lenge; the Figure also shows (C) IL-6 release, (D) TNF-α release, (E) MPO, (F) MDA, (G) and the quantification of PEA. On the right panel, (H) the immunofluorescence analysis of HYAL1 expression on the 3rd and 7th day after the DNBS challenge in the presence of different treatments and (I) the relative quantification expressed as RFU are presented. Magnification: 10×; scale bar: Results are expressed as the mean ± SD of n = 5 experiments **** p < 0.0001 vs. vehicle; *** p < 0.001 vs. vehicle ◦◦◦◦p < 0.0001 vs. DNBS; ◦◦◦p < 0.001 vs. DNBS.

Article Snippet: Both mouse and human sections were blocked with bovine serum albumin and subsequently stained with the rabbit anti-ZO-1 antibody (1:100 dilution v/v; Proteintech, Manchester, UK), rabbit anti-occludin antibody (1:100 dilution v/v; Novus Biologicals, Abingdon, UK) or mouse anti-HYAL1 antibody (1:150 dilution v/v; Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Expressing, Clinical Proteomics, Immunofluorescence, Quantitative Proteomics

Journal: Frontiers in Immunology

Article Title: Dysregulated hyaluronan metabolism drives inflammation and angiogenesis in proliferative diabetic retinopathy

doi: 10.3389/fimmu.2026.1724199

Figure Lengend Snippet: Immunohistochemical analysis of epiretinal fibrovascular membranes that were obtained from 14 patients with proliferative diabetic retinopathy during pars plana vitrectomy. Significant positive correlations between the numbers of blood vessels expressing CD31, reflecting angiogenic activity, and the numbers of blood vessels expressing hyaluronan synthase (HAS)2 (A) , the numbers of stromal cells expressing HAS2 (B) , the numbers of blood vessels expressing hyaluronidase (Hyal)-2 (C) and the numbers of blood vessels expressing CD44 (D) (Pearson’s correlation coefficient). Significant positive correlations between vitreous fluid levels of hyaluronan and the levels of hyaluronidase (Hyal)-1 (E) , heparan sulphate (F) and soluble syndecan-1 (G) (Pearson’s correlation coefficient). Samples from patients with proliferative diabetic retinopathy (PDR) are indicated by open circles, whereas samples from non-diabetic patients with rhegmatogenous retinal detachment (RD) are indicated by closed circles, illustrating sample clustering according to disease entity.

Article Snippet: To determine the presence of Hyal-1, Hyal-2, HAS2, CD44, syndecan-1, heparan sulphate and RHAMM in the vitreous samples, equal volumes (10 μL) of vitreous samples were boiled in Laemmli’s sample buffer (1:1, v/v) under reducing condition for 10 min. Immunodetection was performed with the use of rabbit polyclonal anti-Hyal-1 antibody (1:1000, NBP2-16906, Novus Biologicals), mouse polyclonal anti-Hyal-2 antibody (1:1000, H00008692-B02P, Novus Biologicals), mouse monoclonal anti-HAS2 antibody (1:1000, ab140671, Abcam), rabbit monoclonal anti-CD44 antibody (1:1000, ab189524, Abcam), rabbit monoclonal anti-RHAMM antibody (1:1000, ab124729, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho-NF-κB) (1:1000, NB100-82086, Novus Biologicals), rabbit polyclonal anti-high-mobility group box1 (HMGB1) (1:1000, Cat. no. ab18256, Abcam), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), and mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.).

Techniques: Immunohistochemical staining, Expressing, Activity Assay

Journal: Frontiers in Immunology

Article Title: Dysregulated hyaluronan metabolism drives inflammation and angiogenesis in proliferative diabetic retinopathy

doi: 10.3389/fimmu.2026.1724199

Figure Lengend Snippet: Determination of hyaluronan synthase (HAS)2 (A) , hyaluronidase (Hyal)-1 (B) , Hyal-2 (C) , soluble CD44 (D) , syndecan-1 (E) and heparan sulphate (F) levels in vitreous fluid samples. Equal volumes (15 µL) of vitreous fluid from patients with proliferative diabetic retinopathy (PDR; n=10) and from non-diabetic patients with rhegmatogenous retinal detachment (RD; n=10) were subjected to gel electrophoresis and the presence of HAS2, Hyal-1, Hyal-2, soluble CD44, syndecan-1, and heparan sulphate were detected by Western blot analysis. Representative sets of samples are shown. The intensity of the protein bands was determined in all samples and band intensities were compared between RD and PDR patients. Results are expressed as mean ± standard deviation or standard error of mean (*p < 0.05, independent t-test).

Article Snippet: To determine the presence of Hyal-1, Hyal-2, HAS2, CD44, syndecan-1, heparan sulphate and RHAMM in the vitreous samples, equal volumes (10 μL) of vitreous samples were boiled in Laemmli’s sample buffer (1:1, v/v) under reducing condition for 10 min. Immunodetection was performed with the use of rabbit polyclonal anti-Hyal-1 antibody (1:1000, NBP2-16906, Novus Biologicals), mouse polyclonal anti-Hyal-2 antibody (1:1000, H00008692-B02P, Novus Biologicals), mouse monoclonal anti-HAS2 antibody (1:1000, ab140671, Abcam), rabbit monoclonal anti-CD44 antibody (1:1000, ab189524, Abcam), rabbit monoclonal anti-RHAMM antibody (1:1000, ab124729, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho-NF-κB) (1:1000, NB100-82086, Novus Biologicals), rabbit polyclonal anti-high-mobility group box1 (HMGB1) (1:1000, Cat. no. ab18256, Abcam), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), and mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.).

Techniques: Nucleic Acid Electrophoresis, Western Blot, Standard Deviation

Journal: Frontiers in Immunology

Article Title: Dysregulated hyaluronan metabolism drives inflammation and angiogenesis in proliferative diabetic retinopathy

doi: 10.3389/fimmu.2026.1724199

Figure Lengend Snippet: Expression levels of hyaluronidase (Hyal)-1 (A) , CD44 (B) and receptor for hyaluronan-mediated motility (RHAMM) (C) in the retinal lysates of non-diabetic control rats (C) (n=12) and diabetic rats (D) (n=12) were determined by Western blot analysis. After determination of the intensity of the protein bands, intensities were adjusted to those of β-actin in the samples. Oxidative stress was monitored with the use of 2’,7’-Dichlorofluorescein (DCF) fluorescence intensity analysis (D) . Results are expressed as mean ± standard deviation. Ultra-Low molecular weight hyaluronan (ULMW-HA) induces breakdown of blood-retinal barrier (E) . ULMW-HA was injected intravitreally at the dose of 50 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of normal rats. The BRB was quantified with the fluorescein isothiocyanate-conjugated dextran technique. Results are expressed as mean ± standard deviation of 12 rats. *p < 0.05 compared to the values obtained from PBS-injected eyes. (independent t-test). Western blot analysis of retinas demonstrated that intravitreal injection of ULMW-HA induced significant upregulation of the expression of phospho-NF-κB (F) , phospho-ERK1/2 (G) , vascular endothelial growth factor (VEGF) (H) , intercellular adhesion molecule-1 (ICAM-1) (I) , vascular cell adhesion molecule-1 (VCAM-1) (J) and high-mobility group box-1 (HMGB1) (K) . Results are expressed as mean ± standard deviation or standard error of mean of 8–10 rats in each group (*p < 0.05; independent t-test).

Article Snippet: To determine the presence of Hyal-1, Hyal-2, HAS2, CD44, syndecan-1, heparan sulphate and RHAMM in the vitreous samples, equal volumes (10 μL) of vitreous samples were boiled in Laemmli’s sample buffer (1:1, v/v) under reducing condition for 10 min. Immunodetection was performed with the use of rabbit polyclonal anti-Hyal-1 antibody (1:1000, NBP2-16906, Novus Biologicals), mouse polyclonal anti-Hyal-2 antibody (1:1000, H00008692-B02P, Novus Biologicals), mouse monoclonal anti-HAS2 antibody (1:1000, ab140671, Abcam), rabbit monoclonal anti-CD44 antibody (1:1000, ab189524, Abcam), rabbit monoclonal anti-RHAMM antibody (1:1000, ab124729, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho-NF-κB) (1:1000, NB100-82086, Novus Biologicals), rabbit polyclonal anti-high-mobility group box1 (HMGB1) (1:1000, Cat. no. ab18256, Abcam), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), and mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.).

Techniques: Expressing, Control, Western Blot, Fluorescence, Standard Deviation, Molecular Weight, Injection, Saline

Journal: Frontiers in Immunology

Article Title: Dysregulated hyaluronan metabolism drives inflammation and angiogenesis in proliferative diabetic retinopathy

doi: 10.3389/fimmu.2026.1724199

Figure Lengend Snippet: Human retinal Müller glial cells were left untreated or treated with high-glucose (HG) (25 mM) for 24 (h). For HG treatment, cultures treated with mannitol (25 mM) were used as a control. Protein expression of hyaluronan synthase (HAS)2 (A) , hyaluronidase (Hyal)-1 (B) , Hyal-2 (C) , CD44 (D) and receptor for hyaluronan-mediated motility (RHAMM) (E) in cell lysate was determined by Western blot analysis. Human retinal Müller glial cells were left untreated or treated with HG (25 mM), cobalt chloride (CoCl 2 ) (300 µM), tumor necrosis factor-α (TNFα) (5 ng/mL) or hydrogen peroxide (H 2 O 2 ) (10 mM) for 24 (h) Levels of hyaluronidase (Hyal)-1 were quantified in the culture media by ELISA. (F) Results are expressed as mean ± standard deviation or standard error of mean from three different experiments each performed in triplicate (*p < 0.05; independent t-test).

Article Snippet: To determine the presence of Hyal-1, Hyal-2, HAS2, CD44, syndecan-1, heparan sulphate and RHAMM in the vitreous samples, equal volumes (10 μL) of vitreous samples were boiled in Laemmli’s sample buffer (1:1, v/v) under reducing condition for 10 min. Immunodetection was performed with the use of rabbit polyclonal anti-Hyal-1 antibody (1:1000, NBP2-16906, Novus Biologicals), mouse polyclonal anti-Hyal-2 antibody (1:1000, H00008692-B02P, Novus Biologicals), mouse monoclonal anti-HAS2 antibody (1:1000, ab140671, Abcam), rabbit monoclonal anti-CD44 antibody (1:1000, ab189524, Abcam), rabbit monoclonal anti-RHAMM antibody (1:1000, ab124729, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho-NF-κB) (1:1000, NB100-82086, Novus Biologicals), rabbit polyclonal anti-high-mobility group box1 (HMGB1) (1:1000, Cat. no. ab18256, Abcam), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), and mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.).

Techniques: Control, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Frontiers in Immunology

Article Title: Dysregulated hyaluronan metabolism drives inflammation and angiogenesis in proliferative diabetic retinopathy

doi: 10.3389/fimmu.2026.1724199

Figure Lengend Snippet: Human retinal microvascular endothelial cells (HRMECs) were left untreated or treated with high-glucose (HG) (25 mM) for 24h. For HG treatment, cultures treated with mannitol (25 mM) were used as a control. Protein expression of hyaluronan synthase (HAS)2 (A) , hyaluronidase (Hyal)-1 (B) , Hyal-2 (C) and CD44 (D) in cell lysate was determined by Western blot analysis. (E) HRMECs were left untreated or treated with tumor necrosis factor–α (TNF-α) (5ng/ml) or hydrogen peroxide (H 2 O 2 ) (10mM) for 24h. Levels of Hyal-1 were quantified in the culture media by ELISA. (F) HRMECs and human retinal Müller glial cells were compared after treatment with mannitol (25 mM) or HG (25 mM) for 24h. Levels of hyaluronan (HA) were quantified in the culture media by ELISA. Results are expressed as mean ± standard deviation from three different experiments each performed in triplicate (*p < 0.05; independent t-test).

Article Snippet: To determine the presence of Hyal-1, Hyal-2, HAS2, CD44, syndecan-1, heparan sulphate and RHAMM in the vitreous samples, equal volumes (10 μL) of vitreous samples were boiled in Laemmli’s sample buffer (1:1, v/v) under reducing condition for 10 min. Immunodetection was performed with the use of rabbit polyclonal anti-Hyal-1 antibody (1:1000, NBP2-16906, Novus Biologicals), mouse polyclonal anti-Hyal-2 antibody (1:1000, H00008692-B02P, Novus Biologicals), mouse monoclonal anti-HAS2 antibody (1:1000, ab140671, Abcam), rabbit monoclonal anti-CD44 antibody (1:1000, ab189524, Abcam), rabbit monoclonal anti-RHAMM antibody (1:1000, ab124729, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho-NF-κB) (1:1000, NB100-82086, Novus Biologicals), rabbit polyclonal anti-high-mobility group box1 (HMGB1) (1:1000, Cat. no. ab18256, Abcam), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM-1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), and mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.).

Techniques: Control, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Journal for Immunotherapy of Cancer

Article Title: An oncolytic vaccinia virus encoding hyaluronidase reshapes the extracellular matrix to enhance cancer chemotherapy and immunotherapy

doi: 10.1136/jitc-2023-008431

Figure Lengend Snippet: Characterization of oncolytic vaccinia virus encoding the hyaluronidase (OVV-Hyal1) and intratumoral injections of OVV-Hyal1 enhances antitumor efficacy in subcutaneous tumor models. (A) A schematic diagram of homologous recombination. To generate OVV-Hyal1, a shuttle plasmid pVV-Hyal1 was used for homologous recombination with a western reserve (WR) strain of VV by using the left (L) and right (R) flanking sequences of thymidine kinase (TK). (B) TCID50 method was used to detect viral replication in murine tumor cells. (C) Western blot analysis of OVV-Hyal1-induced HYAL1 overexpression in three murine tumor cell lines. (D) MTT assay was used to detect the oncolytic ability of OVV-Hyal1 and OVV-Ctrl against murine tumor cells. (E) 5×10 5 Panc02, 1×10 6 KPC, 5×10 5 4T1, and 1×10 6 CT26 were inoculated into the right flank of C57BL/6 or BALB/c mice. When tumor volume reached approximately 50 to 100 mm 3 , mice were administered intratumorally (IT) with either 2×10 7 pfu OVV-Hyal1 or equal dose of OVV-Ctrl every other day for a total three times. Mice received PBS intratumoral injections were used as untreated controls. (F) Tumor volume were measured every two days. Error bars represent SD. Once the tumor volume exceeded 2000 mm 3 , the mouse was considered dead. (G) Kaplan-Meier survival analysis of tumor-bearing mice treated with PBS (control), OVV-Ctrl, or OVV-Hyal1. Error bars represent SD. ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Subsequently, the PVDF membrane was subjected to an overnight incubation at 4°C with a mouse anti-Hyal1 antibody (25 179-1-AP, Proteintech, Wuhan, China) and a mouse anti-GAPDH antibody (60 004-1-Ig, Proteintech).

Techniques: Virus, Homologous Recombination, Plasmid Preparation, Western Blot, Over Expression, MTT Assay, Control

Journal: Journal for Immunotherapy of Cancer

Article Title: An oncolytic vaccinia virus encoding hyaluronidase reshapes the extracellular matrix to enhance cancer chemotherapy and immunotherapy

doi: 10.1136/jitc-2023-008431

Figure Lengend Snippet: OVV-Hyal1 induces extracellular matrix (ECM) degradation and enhances OVVs replication in vivo. (A, B) 5×10 5 Panc02 injected into the right flank of C57BL/6 mice. When the tumor reached approximately 50–100 mm 3 , mice were administered intratumorally (IT) with either 2×10 7 pfu OVV-Hyal1 or the equal dose of OVV-Ctrl. Mice received PBS IT was used as untreated control. Tumors were isolated 48 hours after last viral injection and tissue slices were then stained by (A) H&E, Masson, Van Gieson (VG) and picrosirius red staining, and (B) immunohistochemical staining for collagen I, collagen III, elastin, fibronectin. Microscopic images were analyzed semi-quantitatively, and the expression levels of ECM components were expressed as fold change relative to untreated controls. Data are presented as means±SD. Scale bars are equal to 625 µm for H&E and 100 µm for Masson, VG, picrosirius red staining, and immunohistochemistry. (C) The virus titers in tumor tissues or sera of Panc02, KPC, 4T1 and CT26 mice were quantified by the TCID50 method. Error bars represent SD. (D) The subcutaneous tumor models of 4T1 Hyal1 and parental 4T1 were established by inoculation of same number of cells (5×10 5 cells) on the right flank of BALB/c mice. At the end of the mouse experiment, take the mouse lungs and count the number of metastatic lesions. Data are presented as means±SD. (E) Cell scratch assay was used to detect the migration ability of 4T1 Hyal1 cells. Scale bars are equal to 100 µm. (F) Transwell experiment was used to detect the metastation ability of 4T1 Hyal1 cells. Scale bars are equal to 100 µm. Data are presented as means±SD. ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Subsequently, the PVDF membrane was subjected to an overnight incubation at 4°C with a mouse anti-Hyal1 antibody (25 179-1-AP, Proteintech, Wuhan, China) and a mouse anti-GAPDH antibody (60 004-1-Ig, Proteintech).

Techniques: In Vivo, Injection, Control, Isolation, Staining, Immunohistochemical staining, Expressing, Immunohistochemistry, Virus, Wound Healing Assay, Migration

Journal: Journal for Immunotherapy of Cancer

Article Title: An oncolytic vaccinia virus encoding hyaluronidase reshapes the extracellular matrix to enhance cancer chemotherapy and immunotherapy

doi: 10.1136/jitc-2023-008431

Figure Lengend Snippet: Oncolytic vaccinia virus encoding the hyaluronidase (OVV-Hyal1) significantly improves chemotherapy and GLP1-mediated antitumor efficacy. (A) Treatment scheme of combination OVV-Hyal1 and doxorubicin or gemcitabine in Panc02 subcutaneous tumor model. (B) Tumor single-cell suspensions were preprepared 1 day after last doxorubicin injection and analyzed by flow cytometry. Representative diagram of cell with red fluorescence in tumor tissues. (C) Mean tumor volume of combination therapy of OVV-Hyal1 and doxorubicin. (D) Mean tumor volume of combination therapy of OVV-Hyal1 and gemcitabine. (E) Individual tumor growth curve of combination therapy of OVV-Hyal1 and gemcitabine. (F) Kaplan-Meier survival curves of tumor-bearing mice. (G) Body weight of mice was measured every 3 days. (H) Induction of apoptosis in tumor tissues was assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Microscopic images were analyzed semi-quantitatively. Scale bars are equal to 50 µm. (I) Treatment scheme of combination OVV-Hyal1 and liraglutide in Panc02 subcutaneous tumor model. (J) Mean tumor volume of combination therapy of OVV-Hyal1 and liraglutide. (K) Individual tumor growth curve of combination therapy of OVV-Hyal1 and liraglutide. (L) Kaplan-Meier survival curves of tumor-bearing mice. (M) Body weight of mice was measured every 3 days. Error bars represent SD. ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Subsequently, the PVDF membrane was subjected to an overnight incubation at 4°C with a mouse anti-Hyal1 antibody (25 179-1-AP, Proteintech, Wuhan, China) and a mouse anti-GAPDH antibody (60 004-1-Ig, Proteintech).

Techniques: Virus, Injection, Flow Cytometry, Fluorescence, TUNEL Assay

Journal: Journal for Immunotherapy of Cancer

Article Title: An oncolytic vaccinia virus encoding hyaluronidase reshapes the extracellular matrix to enhance cancer chemotherapy and immunotherapy

doi: 10.1136/jitc-2023-008431

Figure Lengend Snippet: Oncolytic vaccinia virus encoding the hyaluronidase (OVV-Hyal1) enhances infiltration of immune cells in tumor microenvironment (TME) and sensitizes pancreatic cancer to CAR-T cell therapy. (A–C) Panc02, KPC and 4T1 models were established as described in and the single-cell suspensions were preprepared two days after last viral injection. (A) Flow cytometric analysis of the proportion of lymphocytes, CD3 + T cells, CD8 + T cells, CD4 + T cells, and NK cells in Panc02 tumor tissues. (B) Flow cytometric analysis of the proportions of CD3 + T cells, CD8 + T cells, CD4 + T cells in KPC tumors. (C) Flow cytometric analysis of the proportion of CD8 + T cells, CD4 + T cells and NK cells in 4T1 tumor tissues. (D) Treatment scheme of combination OVV-Hyal1 and CD19 CAR-T cells in Panc02/CD19 subcutaneous tumor model. (E) Mean tumor volume of combination therapy of OVV-Hyal1 and CD19 CAR-T cells. (F) Individual tumor growth curve of combination therapy of OVV-Hyal1 and CD19 CAR-T cells. (G) Kaplan-Meier survival curves of tumor-bearing mice. (H) Body weight of mice was measured every three days. Error bars represent SD. ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Subsequently, the PVDF membrane was subjected to an overnight incubation at 4°C with a mouse anti-Hyal1 antibody (25 179-1-AP, Proteintech, Wuhan, China) and a mouse anti-GAPDH antibody (60 004-1-Ig, Proteintech).

Techniques: Virus, Injection

Journal: Journal for Immunotherapy of Cancer

Article Title: An oncolytic vaccinia virus encoding hyaluronidase reshapes the extracellular matrix to enhance cancer chemotherapy and immunotherapy

doi: 10.1136/jitc-2023-008431

Figure Lengend Snippet: Oncolytic vaccinia virus encoding the hyaluronidase (OVV-Hyal1) augments CD8 + T cells responses and sensitizes pancreatic cancer to anti-PD-1 therapy. (A, B) Panc02 were established as described in and the single-cell suspensions were preprepared two days after last viral injection. (A) Flow cytometric analysis of the expression of granzyme B, IFN-γ and CD107A on CD8 + T cells in Panc02 tumor tissues. (B) Flow cytometric analysis of the expression of immune checkpoints PD-1, CTLA-4, TIM-3, and LAG-3 on CD8 + T cells. (C) After a 24 hours incubation of OVVs at an MOI of 0.5, tumor cells were harvested and the expression of PD-L1 and CD47 on these cells was detected by flow cytometry using anti-PD-L1 antibody or anti-CD47 antibody. Data are presented as means±SD. (D–G) Combined therapy of OVV-Hyal1 and anti-PD-1 antibody in Panc02 subcutaneous tumor model. (D) Treatment scheme of combination OVV-Hyal1 and anti-PD-1 antibody. (E) Mean tumor volume of combination therapy of OVV-Hyal1 and anti-PD-1 antibody. (F) Individual tumor growth curve of combination therapy of OVV-Hyal1 and anti-PD-1 antibody. (G) Kaplan-Meier survival curves of tumor-bearing mice. (H–K) Combined therapy of OVV-Hyal1 and anti-PD-1 antibody in KPC subcutaneous tumor model. (H) Treatment scheme of combination OVV-Hyal1 and anti-PD-1 antibody. (I) Mean tumor volume of combination therapy of OVV-Hyal1 and anti-PD-1 antibody. (J) Individual tumor growth curve of combination therapy of OVV-Hyal1 and anti-PD-1 antibody. (K) Kaplan-Meier survival curves of tumor-bearing mice. (L−N) Treatment of mice with combination OVV-Hyal1 and anti-PD-1 antibody established long-term tumor-specific immunological memory in Panc02 subcutaneous tumor model. (L–N) Combined treatment of OVV-Hyal1 and anti-PD-1 established long-term tumor-specific immunological memory. (L) The cured mice previously challenged with Panc02 and naïve mice were subcutaneously injected with Panc02 cells, and tumor volumes were measured. (M) By the end of the rechallenging experiment, spleens were obtained from either previously cured mice or naïve mice, single cell suspensions were stained and subjected to flow cytometry to detect the proportion of CD8 + T cells and CD4 + T cells, naïve (CD62L + CD44 − ), central memory (CD62L + CD44 + ) and effector memory (CD62L − CD44 + ) CD8 + T cells and CD4 + T cells. Data are presented as means±SD. (N) Splenocytes from either previously cured mice or naïve mice were co-cultured with Panc02 cells or other mouse cancer cell lines as depicted. 72 hours later, the supernatants were harvested and the cytokines including interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α) and interleukin 2 (IL-2) were detected by ELISA. Data are presented as means±SD. ns, not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Subsequently, the PVDF membrane was subjected to an overnight incubation at 4°C with a mouse anti-Hyal1 antibody (25 179-1-AP, Proteintech, Wuhan, China) and a mouse anti-GAPDH antibody (60 004-1-Ig, Proteintech).

Techniques: Virus, Injection, Expressing, Incubation, Flow Cytometry, Staining, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: Journal for Immunotherapy of Cancer

Article Title: An oncolytic vaccinia virus encoding hyaluronidase reshapes the extracellular matrix to enhance cancer chemotherapy and immunotherapy

doi: 10.1136/jitc-2023-008431

Figure Lengend Snippet: Oncolytic vaccinia virus encoding the hyaluronidase (OVV-Hyal1) promotes macrophages infiltration and M1 polarization and sensitizes breast cancer to anti-CD47 therapy. (A, C, D) Panc02 and 4T1 tumor models were established as described in and the single-cell suspensions were preprepared two days post the last viral injection. (A) Flow cytometric analysis of the proportions of F4/80 + CD11b + macrophages in Panc02 model. Data are presented as means±SD. (B) Immunohistochemistry for CD68 in Panc02 model. The establishment of the Panc02 model and the treatment scheme was similar to . Immunohistochemical staining was used to detect the infiltration of CD68 + T cells in the tumor tissue of the Panc02 subcutaneous model. Scale bar represents 50 µm. (C) Flow cytometric analysis of the proportions of CD80 + CD206 − M1 macrophages and CD80 + CD206 + M2 macrophages in Panc02 model. Data are presented as means±SD. (D) Flow cytometric analysis of the proportions of F4/80 + CD11b + macrophages in 4T1 model. Data are presented as means±SD. (E) Treatment scheme of combination OVV-Hyal1 and anti-CD47 antibody in 4T1 subcutaneous tumor model. (F) Mean tumor volume of combination therapy of OVV-Hyal1 and anti-CD47 antibody. (G) Individual tumor growth curve of combination therapy of OVV-Hyal1 and anti-CD47 antibody. (H) Kaplan-Meier survival curves of tumor-bearing mice. (I) Body weight of mice was measured every three days. Error bars represent SD. ns, not significant; *p<0.05; ***p<0.001; ****p<0.0001.

Article Snippet: Subsequently, the PVDF membrane was subjected to an overnight incubation at 4°C with a mouse anti-Hyal1 antibody (25 179-1-AP, Proteintech, Wuhan, China) and a mouse anti-GAPDH antibody (60 004-1-Ig, Proteintech).

Techniques: Virus, Injection, Immunohistochemistry, Immunohistochemical staining, Staining

Journal: Cancer Microenvironment

Article Title: Targeting Hyaluronan Interactions for Glioblastoma Stem Cell Therapy

doi: 10.1007/s12307-019-00224-2

Figure Lengend Snippet: At baseline, GSC markers, HA receptors, and drug resistance genes are highly co-expressed in GBM, with cell-line dependence. a Percent of population expressing SOX2 and CD44 within U87-MG and D456 cells treated in NBE with varying concentrations of HA for a full passage length, measured via flow cytometry (mean ± SE; n = 3). b Confocal microscopy was performed on D456 cells grown in NBE for NANOG, SOX2, and HYAL1 (scale bars = 200 μm). c qRT-PCR gene expression of (left to right) GSC markers, HA-related genes, and drug resistance genes in U87-MG and D456 cells. Reported as relative expression compared to housekeeping gene GAPDH with log10 transformation and significance noted between cell lines. HYAL1 and MDR1 were lower than detection level (n.d.) in D456 (mean ± SE; n = 3; * p < 0.05, † p < 0.01, and ‡ p < 0.001

Article Snippet: Cells were stained with mouse anti-human IgG 1,κ NANOG PE conjugated antibody (BD), mouse anti-SOX2 IgG 1,κ Alexa Fluor 647 conjugated antibody (BD), or mouse anti-HYAL1 IgG 1 monoclonal primary antibody (Santa Cruz Biotechnology) and rat anti-mouse IgG 1 APC secondary antibody (BD).

Techniques: Expressing, Flow Cytometry, Confocal Microscopy, Quantitative RT-PCR, Transformation Assay

Journal: Cancer Microenvironment

Article Title: Targeting Hyaluronan Interactions for Glioblastoma Stem Cell Therapy

doi: 10.1007/s12307-019-00224-2

Figure Lengend Snippet: HAase decreases stemness through involvement of CD44 signaling. qRT-PCR gene expression of U87-MG and D456 cells treated with HAase for a full passage length. a GSC markers CD133, SOX2, NES, and NANOG. b HA-related genes HAS2, RHAMM, and CD44. c Drug resistance genes STAT3, EGFR, and MDR1. MDR1 was not detected in D456 cells. All genes reported as relative expression compared to housekeeping gene GAPDH (mean ± SE; n = 3;* p < 0.05, † p < 0.01)

Article Snippet: Cells were stained with mouse anti-human IgG 1,κ NANOG PE conjugated antibody (BD), mouse anti-SOX2 IgG 1,κ Alexa Fluor 647 conjugated antibody (BD), or mouse anti-HYAL1 IgG 1 monoclonal primary antibody (Santa Cruz Biotechnology) and rat anti-mouse IgG 1 APC secondary antibody (BD).

Techniques: Quantitative RT-PCR, Expressing