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BioRxiv : the Preprint Server For... Jun 2024An imbalance in matrix metalloproteinase-9 (MMP-9) regulation can lead to numerous diseases, including neurological disorders, cancer, and pre-term labor. Engineering...
An imbalance in matrix metalloproteinase-9 (MMP-9) regulation can lead to numerous diseases, including neurological disorders, cancer, and pre-term labor. Engineering single-chain antibody fragments (scFvs) Targeting MMP-9 to develop novel therapeutics for such diseases is desirable. We screened a synthetic scFv antibody library displayed on the yeast surface for binding improvement to MMP-9 using FACS (fluorescent-activated cell sorting). The scFv antibody clones isolated after FACS showed improvement in binding to MMP-9 compared to the endogenous inhibitor. To understand molecular determinants of binding between engineered scFv antibody variants and MMP-9, next-generation DNA sequencing, and computational protein structure analysis were used. Additionally, a deep-learning language model was trained on the synthetic library to predict the binding of scFv variants using their CDR-H3 sequences.
PubMed: 38895413
DOI: 10.1101/2024.06.04.597476 -
BioRxiv : the Preprint Server For... Jun 2024The formation of functional epithelial tubules is a central feature of many organ systems. Although the process of tubule formation by epithelial cells is well-studied,...
The formation of functional epithelial tubules is a central feature of many organ systems. Although the process of tubule formation by epithelial cells is well-studied, the way in which tubules connect with each other (i.e. anastomose) to form functional networks both and is not well understood. A key, unanswered question in the kidney is how the renal vesicles of the embryonic kidney connect with the nascent collecting ducts to form a continuous urinary system. We performed a ligand-receptor pair analysis on single cell RNA-seq data from embryonic mouse kidney tubules undergoing anastomosis to select candidates that might mediate this process . This analysis identified hepatocyte growth factor (HGF), which has known roles in cell proliferation, migration, and tubulogenesis, as one of several possible candidates. To test this possibility, we designed a novel assay to quantitatively examine epithelial tubule anastomosis using epithelial spheroids with fluorescently-tagged apical surfaces to enable direct visualization of anastomosis. This revealed that HGF is a potent inducer of tubule anastomosis. Tubule anastomosis occurs through a proliferation-independent mechanism that acts through the MAPK signaling cascade and matrix metalloproteinases (MMPs), the latter suggestive of a role in extracellular matrix turnover. Accordingly, treatment of explanted embryonic mouse kidneys with HGF and collagenase was sufficient to induce kidney tubule anastomosis. These results lay the groundwork for investigating how to promote functional interconnections between tubular epithelia, which have important clinical implications for utilizing grown kidney tissue in transplant medicine.
PubMed: 38895378
DOI: 10.1101/2024.06.03.597185 -
Diagnostics (Basel, Switzerland) May 2024Peritoneal dialysis-related peritonitis (PDRP) is the most common complication of peritoneal dialysis (PD), which can lead to poor outcomes if not diagnosed and treated...
BACKGROUND
Peritoneal dialysis-related peritonitis (PDRP) is the most common complication of peritoneal dialysis (PD), which can lead to poor outcomes if not diagnosed and treated early. We aimed to investigate the diagnostic accuracy of MMP-8 and IL-6-based point-of-care tests (POCTs) in diagnosing PDRP in PD patients.
METHODS
This retrospective chart review study was conducted at a comprehensive kidney center in Qatar. It involved all adult PD patients who underwent PDRP from July 2018 to October 2019 and for whom MMP-8 and IL-6-based POCTs were used to diagnose presumptive peritonitis. Measures of diagnostic accuracy were computed. Peritoneal fluid effluent analysis was the reference standard.
RESULTS
We included 120 patients (68 [56.7%] females, ages 55.6 ± 15.6 years, treatment duration 39.5 ± 30.4 months [range: 5-142 months]). In this population, MMP-8 and IL-6-based POCTs yielded 100% in all dimensions of diagnostic accuracy (sensitivity, specificity, positive and negative predictive values).
CONCLUSIONS
MMP-8 and IL-6-based POCTs might be helpful in the early detection of PDRP. This monocentric observation requires further confirmation in a prospective multicentric setting.
PubMed: 38893639
DOI: 10.3390/diagnostics14111113 -
Cancers May 2024Glioblastoma (GBM) cells are highly invasive, infiltrating the surrounding normal brain tissue, thereby limiting the efficacy of surgical resection and focal...
Glioblastoma (GBM) cells are highly invasive, infiltrating the surrounding normal brain tissue, thereby limiting the efficacy of surgical resection and focal radiotherapy. Cysteamine, a small aminothiol molecule that is orally bioavailable and approved for cystinosis, has potential as a cancer treatment by inhibiting tumor cell invasion and metastasis. Here we demonstrate that these potential therapeutic effects of cysteamine are likely due to the inhibition of matrix metalloproteinases (MMPs) in GBM. In vitro assays confirmed that micromolar concentrations of cysteamine were not cytotoxic, enabling the interrogation of the cellular effects without confounding tumor cell loss. Cysteamine's inhibition of MMP activity, especially the targeting of MMP2, MMP9, and MMP14, was observed at micromolar concentrations, suggesting the mechanism of action in suppressing invasion and cell migration is by inhibition of these MMPs. These findings suggest that achievable micromolar concentrations of cysteamine effectively inhibit cancer cell invasion and migration in GBM, supporting the potential for use as an adjunct cancer treatment.
PubMed: 38893149
DOI: 10.3390/cancers16112029 -
International Journal of Molecular... Jun 2024Ovarian cancer (OC) has an unfavorable prognosis. Due to the lack of effective screening tests, new diagnostic methods are being sought to detect OC earlier. The aim of...
Ovarian cancer (OC) has an unfavorable prognosis. Due to the lack of effective screening tests, new diagnostic methods are being sought to detect OC earlier. The aim of this study was to evaluate the concentration and diagnostic utility of selected matrix metalloproteinases (MMPs) as OC markers in comparison with HE4, CA125 and the ROMA algorithm. The study group consisted of 120 patients with OC; the comparison group consisted of 70 patients with benign lesions and 50 healthy women. MMPs were determined via the ELISA method, HE4 and CA125 by CMIA. Patients with OC had elevated levels of MMP-3 and MMP-11, similar to HE4, CA125 and ROMA values. The highest SE, SP, NPV and PPV values were found for MMP-26, CA125 and ROMA in OC patients. Performing combined analyses of ROMA with selected MMPs increased the values of diagnostic parameters. The topmost diagnostic power of the test was obtained for MMP-26, CA125, HE4 and ROMA and performing combined analyses of MMPs and ROMA enhanced the diagnostic power of the test. The obtained results indicate that the tested MMPs do not show potential as stand-alone OC biomarkers, but can be considered as additional tests to raise the diagnostic utility of the ROMA algorithm.
Topics: Humans; Female; Ovarian Neoplasms; CA-125 Antigen; WAP Four-Disulfide Core Domain Protein 2; Middle Aged; Algorithms; Biomarkers, Tumor; Adult; Aged; Matrix Metalloproteinase 2; Proteins; Matrix Metalloproteinases; Matrix Metalloproteinase 3; Membrane Proteins; Case-Control Studies; ROC Curve; Matrix Metalloproteinase 11
PubMed: 38892452
DOI: 10.3390/ijms25116265 -
International Journal of Molecular... Jun 2024Triple-negative breast cancer (TNBC) accounts for approximately 15-20% of all breast cancer types, indicating a poor survival prognosis with a more aggressive biology of...
Triple-negative breast cancer (TNBC) accounts for approximately 15-20% of all breast cancer types, indicating a poor survival prognosis with a more aggressive biology of metastasis to the lung and a short response duration to available therapies. Ibulocydine (IB) is a novel (cyclin-dependent kinase) CDK7/9 inhibitor prodrug displaying potent anti-cancer effects against various cancer cell types. We performed in vitro and in vivo experiments to determine whether IB inhibits metastasis and eventually overcomes the poor drug response in TNBC. The result showed that IB inhibited the growth of TNBC cells by inducing caspase-mediated apoptosis and blocking metastasis by reducing MMP-9 expression in vitro. Concurrently, in vivo experiments using the metastasis model showed that IB inhibited metastasis of MDA-MB-231-Luc cells to the lung. Collectively, these results demonstrate that IB inhibited the growth of TNBC cells and blocked metastasis by regulating MMP-9 expression, suggesting a novel therapeutic agent for metastatic TNBC.
Topics: Matrix Metalloproteinase 9; Humans; Triple Negative Breast Neoplasms; Cell Movement; Female; Cell Line, Tumor; Animals; Mice; Apoptosis; Cell Proliferation; Neoplasm Invasiveness; Xenograft Model Antitumor Assays; Gene Expression Regulation, Neoplastic; Antineoplastic Agents; Lung Neoplasms; Mice, Nude
PubMed: 38892310
DOI: 10.3390/ijms25116123 -
International Journal of Molecular... May 2024Desmoplasia is a common feature of aggressive cancers, driven by a complex interplay of protein production and degradation. Basigin is a type 1 integral membrane...
Desmoplasia is a common feature of aggressive cancers, driven by a complex interplay of protein production and degradation. Basigin is a type 1 integral membrane receptor secreted in exosomes or released by ectodomain shedding from the cell surface. Given that soluble basigin is increased in the circulation of patients with a poor cancer prognosis, we explored the putative role of the ADAM12-generated basigin ectodomain in cancer progression. We show that recombinant basigin ectodomain binds β1 integrin and stimulates gelatin degradation and the migration of cancer cells in a matrix metalloproteinase (MMP)- and β1-integrin-dependent manner. Subsequent in vitro and in vivo experiments demonstrated the altered expression of extracellular matrix proteins, including fibronectin and collagen type 5. Thus, we found increased deposits of collagen type 5 in the stroma of nude mice tumors of the human tumor cell line MCF7 expressing ADAM12-mimicking the desmoplastic response seen in human cancer. Our findings indicate a feedback loop between ADAM12 expression, basigin shedding, TGFβ signaling, and extracellular matrix (ECM) remodeling, which could be a mechanism by which ADAM12-generated basigin ectodomain contributes to the regulation of desmoplasia, a key feature in human cancer progression.
Topics: Humans; Animals; ADAM12 Protein; Mice; Extracellular Matrix Proteins; Basigin; Protein Binding; Mice, Nude; Protein Domains; Cell Movement; MCF-7 Cells; Gene Expression Regulation, Neoplastic; Neoplasms; Female; Cell Line, Tumor; Extracellular Matrix
PubMed: 38892056
DOI: 10.3390/ijms25115871 -
International Journal of Molecular... May 2024Keloids, marked by abnormal cellular proliferation and excessive extracellular matrix (ECM) accumulation, pose significant therapeutic challenges. Ethyl pyruvate (EP),...
Keloids, marked by abnormal cellular proliferation and excessive extracellular matrix (ECM) accumulation, pose significant therapeutic challenges. Ethyl pyruvate (EP), an inhibitor of the high-mobility group box 1 (HMGB1) and TGF-β1 pathways, has emerged as a potential anti-fibrotic agent. Our research evaluated EP's effects on keloid fibroblast (KF) proliferation and ECM production, employing both in vitro cell cultures and ex vivo patient-derived keloid spheroids. We also analyzed the expression levels of ECM components in keloid tissue spheroids treated with EP through immunohistochemistry. Findings revealed that EP treatment impedes the nuclear translocation of HMGB1 and diminishes KF proliferation. Additionally, EP significantly lowered mRNA and protein levels of collagen I and III by attenuating TGF-β1 and pSmad2/3 complex expression in both human dermal fibroblasts and KFs. Moreover, metalloproteinase I (MMP-1) and MMP-3 mRNA levels saw a notable increase following EP administration. In keloid spheroids, EP induced a dose-dependent reduction in ECM component expression. Immunohistochemical and western blot analyses confirmed significant declines in collagen I, collagen III, fibronectin, elastin, TGF-β, AKT, and ERK 1/2 expression levels. These outcomes underscore EP's antifibrotic potential, suggesting its viability as a therapeutic approach for keloids.
Topics: Humans; Keloid; Fibroblasts; Pyruvates; Spheroids, Cellular; Matrix Metalloproteinase 1; Transforming Growth Factor beta1; HMGB1 Protein; Collagen; Cell Proliferation; Cells, Cultured; Matrix Metalloproteinase 3; Extracellular Matrix; Collagen Type I; Smad2 Protein; Smad3 Protein; Up-Regulation; Male
PubMed: 38892032
DOI: 10.3390/ijms25115844 -
International Journal of Molecular... May 2024Human abdominal aortic aneurysms (AAAs) are characterized by increased activity of matrix metalloproteinases (MMP), including MMP-12, alongside macrophage accumulation...
Human abdominal aortic aneurysms (AAAs) are characterized by increased activity of matrix metalloproteinases (MMP), including MMP-12, alongside macrophage accumulation and elastin degradation, in conjunction with superimposed atherosclerosis. Previous genetic ablation studies have proposed contradictory roles for MMP-12 in AAA development. In this study, we aimed to elucidate if pharmacological inhibition of MMP-12 activity with a phosphinic peptide inhibitor protects from AAA formation and progression in angiotensin (Ang) II-infused Apoe mice. Complimentary studies were conducted in a human ex vivo model of early aneurysm development. Administration of an MMP-12 inhibitor (RXP470.1) protected hypercholesterolemia Apoe mice from Ang II-induced AAA formation and rupture-related death, associated with diminished medial thinning and elastin fragmentation alongside increased collagen deposition. Proteomic analyses confirmed a beneficial effect of MMP-12 inhibition on extracellular matrix remodeling proteins combined with inflammatory pathways. Furthermore, RXP470.1 treatment of mice with pre-existing AAAs exerted beneficial effects as observed through suppressed aortic dilation and rupture, medial thinning, and elastin destruction. Our findings indicate that pharmacological inhibition of MMP-12 activity retards AAA progression and improves survival in mice providing proof-of-concept evidence to motivate translational work for MMP-12 inhibitor therapy in humans.
Topics: Animals; Aortic Aneurysm, Abdominal; Angiotensin II; Matrix Metalloproteinase 12; Mice; Apolipoproteins E; Humans; Matrix Metalloproteinase Inhibitors; Male; Disease Models, Animal; Mice, Knockout; Mice, Inbred C57BL; Elastin; Proteomics
PubMed: 38891996
DOI: 10.3390/ijms25115809 -
International Journal of Molecular... May 2024Childhood glaucoma encompasses congenital and juvenile primary glaucoma, which are heterogeneous, uncommon, and irreversible optic neuropathies leading to visual...
Childhood glaucoma encompasses congenital and juvenile primary glaucoma, which are heterogeneous, uncommon, and irreversible optic neuropathies leading to visual impairment with a poorly understood genetic basis. Our goal was to identify gene variants associated with these glaucoma types by assessing the mutational burden in 76 matrix metalloproteinase-related genes. We studied 101 childhood glaucoma patients with no identified monogenic alterations using next-generation sequencing. Gene expression was assessed through immunohistochemistry. Functional analysis of selected gene variants was conducted in cultured cells and in zebrafish. Patients presented a higher proportion of rare variants in four metalloproteinase-related genes, including and , compared to controls. ADAMTSL4 protein expression was observed in the anterior segment of both the adult human and zebrafish larvae's eye, including tissues associated with glaucoma. In HEK-293T cells, expression of four ADAMTSL4 variants identified in this study showed that two variants (p.Arg774Trp and p.Arg98Trp) accumulated intracellularly, inducing endoplasmic reticulum stress. Additionally, overexpressing these ADAMTSL4 variants in zebrafish embryos confirmed partial loss-of-function effects for p.Ser719Leu and p.Arg1083His. Double heterozygous functional suppression of and zebrafish orthologs resulted in reduced volume of both the anterior eye chamber and lens within the chamber, supporting a genetic interaction between these genes. Our findings suggest that accumulation of partial functional defects in matrix metalloproteinase-related genes may contribute to increased susceptibility to early-onset glaucoma and provide further evidence supporting the notion of a complex genetic inheritance pattern underlying the disease.
Topics: Humans; Animals; Zebrafish; Glaucoma; Child; Male; Female; Child, Preschool; HEK293 Cells; Genetic Predisposition to Disease; Mutation; Matrix Metalloproteinases; ADAMTS Proteins; Adolescent; Infant; Zebrafish Proteins; Endoplasmic Reticulum Stress
PubMed: 38891949
DOI: 10.3390/ijms25115757