Antinuclear antibodies in individuals with COVID-19 reflect underlying disease: Identification of new autoantibodies in systemic sclerosis (CDK9) and malignancy (RNF20, RCC1, TRIP13).

A high prevalence of antinuclear antibodies (ANA) in COVID-19 has been insinuated, but the nature of the target antigens is poorly understood. We studied ANA by indirect immunofluorescence in 229 individuals with COVID-19. The target antigens of high titer ANA (≥1:320) were determined by immunoprecipitation (IP) combined with liquid-chromatography-mass spectrometry (MS).

Anti-Ki/anti-PA28γ autoantibodies contribute to the HEp-2 indirect immunofluorescence nuclear speckled pattern.

Objectives: Antinuclear antibodies (ANAs) are associated with several autoimmune diseases. Indirect immunofluorescence (IIF) on human epithelial type 2 (HEp-2) cells is the golden standard for ANA detection in the clinic. In case of a positive HEp-2 IIF test result, follow-up tests are done to determine autoantibody specificity.

Proteoform Analysis of Matrix Metalloproteinase-9/Gelatinase B and Discovery of Its Citrullination in Rheumatoid Arthritis Synovial Fluids.

Objectives: To explore posttranslational modifications (PTMs), including proteolytic activation, multimerization, complex formation and citrullination of gelatinases, in particular of gelatinase B/MMP-9, and to detect in gelatin-Sepharose affinity-purified synovial fluids, the presence of specific MMP proteoforms in relation to arthritis.

Methods: Latent, activated, complexed and truncated gelatinase-A/MMP-2 and gelatinase B/MMP-9 proteoforms were detected with the use of zymography analysis to compare specific levels, with substrate conversion assays, to test net proteolytic activities and by Western blot analysis to decipher truncation variants. Citrullination was detected with enhanced sensitivity, by the use of a new monoclonal antibody against modified citrullines.

Citrullination as a novel posttranslational modification of matrix metalloproteinases.

Matrix metalloproteinases (MMPs) are enzymes with critical roles in biology and pathology. Glycosylation, nitrosylation and proteolysis are known posttranslational modifications (PTMs) regulating intrinsically the activities of MMPs. We discovered MMP citrullination by peptidyl arginine deiminases (PADs) as a new PTM.

Recombinant Protein-Based Nanoparticles: Elucidating their Inflammatory Effects In Vivo and their Potential as a New Therapeutic Format.

Bacterial inclusion bodies (IBs) are protein-based nanoparticles of a few hundred nanometers formed during recombinant protein production processes in different bacterial hosts. IBs contain active protein in a mechanically stable nanostructured format that has been broadly characterized, showing promising potential in different fields such as tissue engineering, protein replacement therapies, cancer, and biotechnology. For immunomodulatory purposes, however, the interference of the format immunogenic properties-intrinsic to IBs-with the specific effects of the therapeutic protein is still an uncovered gap.

Protease propeptide structures, mechanisms of activation, and functions.

Proteases are a diverse group of hydrolytic enzymes, ranging from single-domain catalytic molecules to sophisticated multi-functional macromolecules. Human proteases are divided into five mechanistic classes: aspartate, cysteine, metallo, serine and threonine proteases, based on the catalytic mechanism of hydrolysis. As a protective mechanism against uncontrolled proteolysis, proteases are often produced and secreted as inactive precursors, called zymogens, containing inhibitory N-terminal propeptides.

The Biological Potential Hidden in Inclusion Bodies.

Inclusion bodies (IBs) are protein nanoclusters obtained during recombinant protein production processes, and several studies have demonstrated their potential as biomaterials for therapeutic protein delivery. Nevertheless, IBs have been, so far, exclusively sifted by their biological activity in vitro to be considered in further protein-based treatments in vivo. Matrix metalloproteinase-9 (MMP-9) protein, which has an important role facilitating the migration of immune cells, was used as model protein.

MMP-9/Gelatinase B Degrades Immune Complexes in Systemic Lupus Erythematosus.

Systemic Lupus Erythematosus (SLE) is a common and devastating autoimmune disease, characterized by a dysregulated adaptive immune response against intracellular antigens, which involves both autoreactive T and B cells. In SLE, mainly intracellular autoantigens generate autoantibodies and these assemble into immune complexes and activate the classical pathway of the complement system enhancing inflammation. Matrix metalloproteinase-9 (MMP-9) levels have been investigated in the serum of SLE patients and in control subjects.

Gelatinase B/Matrix Metalloproteinase-9 as Innate Immune Effector Molecule in Achalasia.

Objectives: Achalasia is a primary esophageal motility disorder resulting from selective loss of inhibitory neurons in the esophageal myenteric plexus, likely due to an autoimmune response with involvement of the adaptive immune system. Innate immune processes of the host constitute the bridge between environmental etiological factors and the adaptive immune system. Although these remain poorly investigated, they might be of diagnostic and therapeutic relevance.

Propeptide glycosylation and galectin-3 binding decrease proteolytic activation of human proMMP-9/progelatinase B.

Matrix metalloproteinases (MMPs) are secreted as proenzymes, containing propeptides that interact with the catalytic zinc, thereby controlling MMP activation. The MMP-9 propeptide is unique in the MMP family because of its post-translational modification with an N-linked oligosaccharide. ProMMP-9 activation by MMP-3 occurs stepwise by cleavage of the propeptide in an aminoterminal (pro-AT) and carboxyterminal (pro-CT) peptide.

EDTA/gelatin zymography method to identify C1s versus activated MMP-9 in plasma and immune complexes of patients with systemic lupus erythematosus.

Gelatin zymography analysis is a sensitive method and commonly used to characterize and quantify the presence of the gelatinases (MMP-2 and MMP-9) in biological samples. In human plasma samples from healthy controls and systemic lupus erythematosus (SLE) patients, we observed a gelatinolytic molecule at 80 kDa, suggestive for activated human MMP-9. However, by developing and using the EDTA/gelatin zymography method and after purification of the 80 kDa entity, we proved that this molecule was the C1s subunit of the complement system.

Gelatinase B/matrix metalloproteinase-9 is a phase-specific effector molecule, independent from Fas, in experimental autoimmune encephalomyelitis.

Gelatinase B/matrix metalloproteinase-9 (MMP-9) triggers multiple sclerosis (MS) and the animal model of experimental autoimmune encephalomyelitis (EAE) by the breakdown of the blood-brain barrier. Interestingly, MMP-9 is beneficial in systemic autoimmunity caused by Fas-deficiency. Fas-deficient (faslpr) and Fas-ligand-deficient mice are protected against EAE.

Neutrophils and Activated Macrophages Control Mucosal Immunity by Proteolytic Cleavage of Antileukoproteinase.

Antileukoproteinase or secretory leukocyte peptidase inhibitor is a small protein which protects the mucosal linings against excessive proteolysis, inflammation, and microbial infection. We discovered that gelatinase B or matrix metalloproteinase (MMP)-9, a secreted zinc-dependent endopeptidase typically found at sites of inflammation, destroys antileukoproteinase by cleavages within both of its two functional domains: the anti-microbial N-terminal and the anti-proteolytic C-terminal domains. Cleaved antileukoproteinase possessed a significantly lower ability to bind lipopolysaccharides (LPS) and a reduced capacity to inhibit neutrophil elastase (NE) activity.

Endotoxemia shifts neutrophils with TIMP-free gelatinase B/MMP-9 from bone marrow to the periphery and induces systematic upregulation of TIMP-1.

Lipopolysaccharides or endotoxins elicit an excessive host inflammatory response and lead to life-threatening conditions such as endotoxemia and septic shock. Lipopolysaccharides trigger mobilization and stimulation of leukocytes and exaggerated production of pro-inflammatory molecules including cytokines and proteolytic enzymes. Matrix metalloproteinase-9 (MMP-9) or gelatinase B, a protease stored in the tertiary granules of polymorphonuclear leukocytes, has been implicated in such inflammatory reactions.

Differential inhibition of activity, activation and gene expression of MMP-9 in THP-1 cells by azithromycin and minocycline versus bortezomib: A comparative study.

Gelatinase B or matrix metalloproteinase-9 (MMP-9) (EC 3.4.24.

Glycosylation of matrix metalloproteases and tissue inhibitors: present state, challenges and opportunities.

Matrix metalloproteases (MMPs) are crucial components of a complex and dynamic network of proteases. With a wide range of potential substrates, their production and activity are tightly controlled by a combination of signalling events, zymogen activation, post-translational modifications and extracellular inhibition. Slight imbalances may result in the initiation or progression of specific disease states, such as cancer and pathological inflammation.

PHD1 regulates p53-mediated colorectal cancer chemoresistance.

Overcoming resistance to chemotherapy is a major challenge in colorectal cancer (CRC) treatment, especially since the underlying molecular mechanisms remain unclear. We show that silencing of the prolyl hydroxylase domain protein PHD1, but not PHD2 or PHD3, prevents p53 activation upon chemotherapy in different CRC cell lines, thereby inhibiting DNA repair and favoring cell death. Mechanistically, PHD1 activity reinforces p53 binding to p38α kinase in a hydroxylation-dependent manner.