Detection of Matrix Metalloproteinases by Zymography

Tajhya, Rajeev B.; Patel, Rutvik S.; Beeton, Christine

doi:10.1007/978-1-4939-6863-3_12

Cited by 43 publications

(30 citation statements)

References 17 publications

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“…Apparent bands formed, revealing that the purified aNtMMP-9 bounded to the anti-His Tag antibody via Western Blotting ( Figure S5B). The activity levels of MMP-9 were also assessed by zymography [3,42]. A white band of aNtMMP-9 was also found via gelatin zymography with a molecular weight of approximate 63.4 kDa ( Figure S5C), confirming the protease activity of NtMMP-9 in the biological function.…”

Section: Heterologous Expression and The Enzyme Activity Of Ntmmp-9mentioning

confidence: 63%

Characterization of Matrix Metalloprotease-9 Gene from Nile tilapia (Oreochromis niloticus) and Its High-Level Expression Induced by the Streptococcus agalactiae Challenge

et al. 2020

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The bacterial diseases of tilapia caused by Streptococcus agalactiae have resulted in the high mortality and huge economic loss in the tilapia industry. Matrix metalloproteinase-9 (MMP-9) may play an important role in fighting infection. However, the role of MMP-9 in Nile tilapia against S. agalactiae is still unclear. In this work, MMP-9 cDNA of Nile tilapia (NtMMP-9) has been cloned and characterized. NtMMP-9 has 2043 bp and encodes a putative protein of 680 amino acids. NtMMP-9 contains the conserved domains interacting with decorin and inhibitors via binding forces compared to those in other teleosts. Quantitative real-time-polymerase chain reaction (qPCR) analysis reveals that NtMMP-9 distinctly upregulated following S. agalactiae infection in a tissue- and time-dependent response pattern, and the tissues, including liver, spleen, and intestines, are the major organs against a S. agalactiae infection. Besides, the proteolytic activity of NtMMP-9 is also confirmed by heterologous expression and zymography, which proves the active function of NtMMP-9 interacting with other factors. The findings indicate that NtMMP-9 was involved in immune responses against the bacterial challenge at the transcriptional level. Further work will focus on the molecular mechanisms of NtMMP-9 to respond and modulate the signaling pathways in Nile tilapia against S. agalactiae invasion and the development of NtMMP-9-related predictive biomarkers or vaccines for preventing bacterial infection in the tilapia industry.

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Section: Heterologous Expression and The Enzyme Activity Of Ntmmp-9mentioning

confidence: 63%

Characterization of Matrix Metalloprotease-9 Gene from Nile tilapia (Oreochromis niloticus) and Its High-Level Expression Induced by the Streptococcus agalactiae Challenge

et al. 2020

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“…The activity of MMPs (MMP‐1, MMP‐2, and MMP‐9) in samples was determined on Days 1, 3, and 7 by the zymography method described elsewhere 33,34 . Gelatin and casein were used as substrates to evaluate the activity of gelatinases (MMP‐2, MMP‐9) and collagenases (MMP‐1), respectively.…”

Section: Methodsmentioning

confidence: 99%

“…The supernatant was immediately used for analysis. For standardization of samples the protein content in supernatant was determined by the Bradford assay.The activity of MMPs (MMP-1, MMP-2, and MMP-9) in samples was determined on Days 1, 3, and 7 by the zymography method described elsewhere 33,34. Gelatin and casein were used as substrates to evaluate the activity of gelatinases (MMP-2, MMP-9) and collagenases (MMP-1), respectively.…”

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confidence: 99%

Protecting the skin‐implant interface with transcutaneous silver‐coated skin‐and‐bone‐integrated pylon in pig and rabbit dorsum models

et al. 2020

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Implant‐associated soft tissue infections at the skin‐implant interface represent the most frequent complications in reconstructive surgery and lead to implant failures and revisions. Titanium implants with deep porosity, called skin‐and‐bone‐integrated‐pylons (SBIP), allow for skin ingrowth in the morphologically natural direction, thus restoring a reliable dermal barrier and reducing the risk of infection. Silver coating of the SBIP implant surface using physical vapor deposition technique offers the possibility of preventing biofilm formation and exerting a direct antimicrobial effect during the wound healing phase. In vivo studies employing pig and rabbit dorsum models for assessment of skin ingrowth into the pores of the pylon demonstrated the safety of transcutaneous implantation of the SBIP system. No postoperative complications were reported at the end of the follow‐up period of 6 months. Histological analysis proved skin ingrowth in the minipig model without signs of silver toxicity. Analysis of silver release (using energy dispersive X‐ray spectroscopy) in the model of intramedullary‐inserted silver‐coated SBIP in New Zealand rabbits demonstrated trace amounts of silver after 3 months of in‐bone implantation. In conclusion, selected temporary silver coating of the SBIP implant surface is powerful at preventing the periprosthetic infections without imparing skin ingrowth and can be considered for clinical application.

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“…Activity of metaloproteinases (MMPs) was analyzed by a gelatin zymography assay as previously described [ 19 ]. Cells were seeded in 6-well plates and grown for 48 h in medium containing 10 % FBS.…”

Section: Methodsmentioning

confidence: 99%

Anti-metastatic action of an N4-aryl substituted thiosemicarbazone on advanced triple negative breast cancer.

Sólimo

Santacruz

Vanzulli

et al. 2020

Heliyon

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Purpose Advanced triple negative breast cancer (ATNBC) is defined by a lack of expression of hormones receptors as well as HER2/neu and its high probability of visceral metastasis. This pathology is associated with a poor prognosis. Previously, we found that T2, an N 4 -arylsubstituted thiosemicarbazone ( N 4 -TSC), had cytotoxic effect on human breast cancer cells lines. Hence, in this study, we investigated the anti-metastasic action of T2 on ATNBC. Methods In order to deepen T2 action mode on ATNBC, we first confirmed T2 cytotoxicity on a panel of TNBC cells and then continued studying T2 effects in vitro an in vivo on the syngeneic 4T1 mouse model. Results We found that T2 had a cytotoxic effect comparable to chemotherapeutics used in present treatment schemes for ATNBC. T2 treatment not only induced apoptosis, but it also down-modulated 4T1 invasive and metastatic-associated capacities, such as clonogenicity, migration and metallo-proteases activity. Moreover, this agent reduced the number of 4T1 cancer stem cells. Finally, T2 treatment induced a more differentiated cell phenotype and the overexpression of the metastasis suppressor gene NDRG-1. In vivo assays showed that T2 reduced tumor burden, down modulated local tumor invasion and significantly reduced the number of lung metastases in the 4T1 advanced TNBC murine model, while the compound did not exhibit intolerable toxicity. Conclusion This study provided evidence that T2 not only exerted an anti-tumor activity but it also showed anti-invasive and anti-metastatic actions on ATNBC in vivo and in vitro , suggesting that T2 could be considered as a promising therapy that deserves further analysis.

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Detection of Matrix Metalloproteinases by Zymography

Cited by 43 publications

References 17 publications

Characterization of Matrix Metalloprotease-9 Gene from Nile tilapia (Oreochromis niloticus) and Its High-Level Expression Induced by the Streptococcus agalactiae Challenge

Characterization of Matrix Metalloprotease-9 Gene from Nile tilapia (Oreochromis niloticus) and Its High-Level Expression Induced by the Streptococcus agalactiae Challenge

Protecting the skin‐implant interface with transcutaneous silver‐coated skin‐and‐bone‐integrated pylon in pig and rabbit dorsum models

Anti-metastatic action of an N4-aryl substituted thiosemicarbazone on advanced triple negative breast cancer.

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