Wild-type female C57BL/6 mice (6 weeks old) were obtained from the Laboratory Animal Center of Southern Medical University. Mice were randomly divided into four groups (n = 4/group) as follows: (1) intraperitoneal saline + DMSO, (2) intraperitoneal saline + SB216763 (20 mg/kg) (Selleck, Houston, TX, USA) dissolved in DMSO, (3) intraperitoneal saline + SB3CT (20 mg/kg) (Selleck, Houston, TX, USA) dissolved in DMSO, and (4) intraperitoneal saline + SB3CT (20 mg/kg) + SB216763 (20 mg/kg) dissolved in DMSO. To establish a pulmonary tuberculosis model, mice were injected H37Rv intraperitoneally with 1 × 106 CFU/mouse on day 0. SB216763-treated mice were administered with saline + SB216763 dissolved in DMSO intravenously at day 0 and then intraperitoneally twice a week until day 28. SB3CT-treated mice were intraperitoneally injected with saline + SB3CT dissolved in DMSO daily until day 28. The control group was injected with saline + DMSO as vehicle control. The mice were sacrificed on days 7 and 28, and lung and spleen tissues were grinded for CFU assay. The supernatant of lung and spleen tissue grinding was used to detect the expression of MMP-9 by ELISA assay.
Sb 3ct
Manufactured by Selleck Chemicals
Sourced in United States
SB-3CT is a selective gelatinase inhibitor. It functions as a potent and selective inhibitor of the gelatinase family of matrix metalloproteinases, including MMP-2 and MMP-9.
Automatically generated - may contain errors
Lab products found in correlation
Minocycline, by Selleck Chemicals (1 mentions)
Ab92536, by Abcam (1 mentions)
Ag rnaex pro reagent, by Accurate Biology (1 mentions)
Pro taq hs qpcr kit, by Accurate Biology (1 mentions)
Sybr green premix, by Accurate Biology (1 mentions)
Mmp assay kit, by Abcam (1 mentions)
Ecl reagent, by Sartorius (1 mentions)
Plx3397, by Selleck Chemicals (1 mentions)
Evans blue, by Merck Group (1 mentions)
Rotenone, by Merck Group (1 mentions)
Ab38898, by Abcam (1 mentions)
Trizol reagent, by Takara Bio (1 mentions)
P shp2, by Cell Signaling Technology (1 mentions)
Phenylhydrazonopyrazolone sulfonate 1 phps1, by Merck Group (1 mentions)
Shp2 primary antibody, by Santa Cruz Biotechnology (1 mentions)
α sma, by Cell Signaling Technology (1 mentions)
Smad2, by Cell Signaling Technology (1 mentions)
P smad2, by Cell Signaling Technology (1 mentions)
Smad3, by Cell Signaling Technology (1 mentions)
P smad3, by Cell Signaling Technology (1 mentions)
8 protocols using sb 3ct
1
Regulation of Tuberculosis Pathogenesis
2
Neuroinflammation and BBB Dysfunction
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Rotenone (#R8875) and Evans blue (#E2129) were provided by Sigma-Aldrich, Inc. (St. Louis, MO, USA). The AG RNAex Pro Reagent (#AG21102), Pro Taq HS qPCR Kit, and SYBR Green Premix (#AG11720) were provided by Accurate Biotechnology (Hunan, China). The PLX3397 (#S7818), minocycline (#S4226), and SB-3CT (#S7430) were provided by Selleck (Shanghai, China). Antibodies against tyrosine hydroxylase (TH, #AB152) and ionized calcium binding adaptor molecule-1 (Iba-1, #019-19741) antibodies were provided by EMD Millipore (Temecula, CA, USA) and Wako Chemicals (Richmond, VA, USA), respectively. Antibodies against fibrinogen (#ab34269), zonula occludens-1 (ZO-1, #ab96587), claudin-5 (#ab131259), occludin (#ab167161), MMP-2 (#ab92536), MMP-9 (#ab38898), GAPDH (#ab181602), and the MMP assay kit (#ab112146) were purchased from Abcam (Cambridge, MA, USA). The ECL reagents (#20-500-120) were provided by Biological Industries (Cromwell, CT, USA).
3
Molecular Mechanisms of PHPS1-Mediated Regulation
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Phenylhydrazonopyrazolone sulfonate 1 (PHPS1) was obtained from Sigma-Aldrich (St. Louis, MO). SB-3CT was obtained from Selleckchem (Houston, TX). RPMI 1640, FBS, penicillin, and streptomycin were obtained from HyClone (Logan, UT). TRIzol reagents were purchased from Takara Bio, Dalian, China. All PCR primers were purchased from Shanghai Bioengineering (Shanghai, China). ERK, p-ERK, Shp2, p-Shp2, Smad2, p-Smad2, Smad3, p-Smad3, JNK, P-JNK, P38, p-P38, MMP-9, MMP-2, E-cadherin, vimentin, α-SMA and β-actin primary antibody were from Cell Signaling Technology (Danvers, MA), and Shp2 primary antibody was from Santa Cruz Biotechnology (Santa Cruz, CA). MMP-9 and TGF-β1 ELISA kits were from Boster (Wuhang, China). Lipofectamine LTX was from Invitrogen (Carlsbad, CA).
4
Invasion Assay with MMP-2 Inhibitor
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HCC cells were seeded in complete medium as described in Section 2.4.1 . On the 2nd and 3rd days after seeding, cells were treated with 0.1μM MMP-2 inhibitor (SB-3CT; Selleckchem, Houston, TX, USA). Simultaneously with the second SB-3CT treatment, scratching was performed, and cells were grown in medium enriched with only 1% FBS to minimize cell proliferation (protocol 9, Figure S1 ). Images were acquired as in Section 2.4.4 .
5
Immunohistochemical Analysis of Immune Checkpoint Proteins
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Human anti-PD-L1-Rb (ab213524) and mouse anti-PD-L1-Rb (ab213480) were purchased from Abcam; anti-CD8α-Rb (GB11068 and GB11068-1) was purchased from Servicebio; anti-PD-1-Rb (84651) was purchased from Cell Signaling Technology; the antibodies specific for human anti-MMP2-Rb (10375-2-AP), anti-MMP9-Rb (10373-2-AP), anti-MMP9-Rb (10373-2-AP), anti-GAPDH-M (60004-1-Ig), and anti-PD-L1-M (66248) were purchased from Proteintech; SB-3CT was purchased from Selleck(S7430); and in vivo mAb anti-PD-1-M (BE0146), anti-CTLA-4-M (BE0164), and IgG isotype control (BE0086, BE0089) were purchased from Bioxcell.
6
CRISPR-Engineered Fibrosis Mouse Model
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Animal maintenance and intervention were approved by the Animal Care Committee of Ruijin Hospital, Shanghai Jiao Tong University School of Medicine. The genetic background of transgenic mice was 129. The animal model constructed by CRISPR/Cas9 technology was Col4a3 C1615Y transgenic mice. This missense mutation corresponds to amino acid number 1616 in humans and was first found in an FSGS family receiving renal biopsy in our department. Protein sequence alignment verified the corresponding mutation to C1615Y in model mice. There were 4 mice in each group. Mice were bred with standard rodent chow food. Male mice were used in the experiments, and the weight difference was less than 10%. All mice received treatment at 8 weeks. MMP-2 inhibitor, SB-3CT (Selleckchem, USA) was administrated intraperitoneally for 6 days before sacrifice at the dosage of 50 mg/kg; NOX4 inhibitor, GKT137831 (Selleckchem, USA) was administrated gavage for 28 days before sacrifice at the dosage of 60 mg/kg. All animal experiments were approved by the Ethics Committee of Ruijin Hospital.
7
Sorafenib and SB-3CT Combination Therapy
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Sorafenib (Nexavar®) and SB-3CT (MMP-2 inhibitor) were purchased from Selleck Chemicals (Houston, TX, USA). LY294002 (PI3K inhibitor), MK-2206 (AKT inhibitor), rapamycin (mTOR inhibitor), RG7204 (RAF inhibitor), U0126 (MEK inhibitor), and PD98059 (ERK inhibitor) were obtained from Selleck Chemicals. For in vitro experiments, these reagents were dissolved in dimethyl sulfoxide (DMSO) for long-term storage and then diluted to the working concentration with DMEM. The final DMSO concentration was less than 0.1%. For in vivo experiments, SB-3CT was dissolved in 10% DMSO, and sorafenib was dissolved in a mixture of Cremophor EL/ethanol/ddH2O (1:1:6). Primary antibodies for p/t-PI3K, p/t-AKT, p/t-mTOR, p/t-RAF, p/t-MEK, and p/t-ERK1/2 were purchased from Cell Signaling Technology (Danvers, MA, USA). Anti-MMP-2 antibody was purchased from Abcam (Cambridge, UK). The antibody against glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and horseradish peroxidase (HRP)-labeled anti-rabbit secondary antibodies were obtained from CWBIO (Beijing, P.R. China).
8
Inhibition of MMP-2/9 in Pancreatic Cancer
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In addition, WT (n=7), STZ (n=9) and db/db (n=9) mice with pancreatic cancer were given an i.p. injection of the selective MMP-2/9 inhibitor SB-3CT (Selleck, USA) diluted in 10% DMSO at a dosage of 25 mg/kg daily for 20 days (44 (link)). The control groups for WT (n=9), STZ (n=9) and db/db (n=11) mice were administered an equal volume of vehicle (10% DMSO). Tumor growth was monitored using T2-weighted MRI. Metastatic lesions were counted in bioluminescence images. Western blotting was conducted to evaluate MMP-2 and MMP-9 expression levels.
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