Protein Cross-linking Research Articles

Inhibition of Lactate Dehydrogenase-A by Singlet Oxygen and Hypochlorous Acid via Cysteine Oxidation and Irreversible Conformational Changes

Muscle lactate dehydrogenase (LDH-A) catalyzes the reduction of pyruvate to lactate, the end product of anaerobic glycolysis. LDH-A is overexpressed in many cancers prior to and even when tumors receive adequate oxygen, and lactate has multiple cellular roles. We assessed the effect of singlet oxygen and hypochlorous acid (HOCl) on mammalian LDH-A. Oxidants induced distinct patterns of protein crosslinks observed by SDS-PAGE under reducing conditions. LDH-A cysteines were detected using fluorescein-modified maleimide to assess their oxidation and accessibility. Singlet oxygen initially increased cysteine exposure, but higher doses resulted in their oxidation in addition to non-reducible covalent crosslinks. LDH-A cysteines were oxidized by micromolar HOCl (1–10 equivalents over enzyme) but were resistant to millimolar H2O2, chloramines and Angeli’s salt. HOCl oxidation inhibited LDH-A activity and yielded inter-chain disulfides observed by nonreducing SDS-PAGE. Disulfide reduction did not restore LDH-A activity that was lost due to HOCl oxidation. An irreversible conformational change induced by HOCl was detected by native gel electrophoresis and tryptophan fluorescence. In the absence of pyruvate, LDH-A enhanced NADH oxidation resulting in H2O2 formation. Singlet oxygen, but not HOCl, initiated this superoxide-dependent chain reaction. Once damaged by both singlet oxygen or HOCl, LDH-A had decreased NADH oxidation activity.

Optimizing protein crosslinking control: Synergistic quenching effects of glycine, histidine, and lysine on glutaraldehyde reactions

Glutaraldehyde (GA) is a protein crosslinker widely used in biochemical and pharmaceutical research because it can rapidly stabilize and immobilize substrates via amine group interactions. However, controlling GA crosslinking is challenging owing to its swift reactivity and the influence of various solution conditions, such as pH and concentrations of the substrate and crosslinker. Although extensive research has focused on GA cross-linking mechanisms, studies on quenching, which is critical for preventing non-specific aggregation during prolonged storage, remain sparse. This study examines the quenching efficiency of a combined amino acid mixture of glycine, histidine, and lysine, which are commonly used as individual quenchers. Our findings, confirmed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis, demonstrate that this amino acid blend offers superior quenching compared to single amino acids, enhancing quenching activity across a wide pH spectrum. These results provide a novel approach for mitigating the high reactivity of GA with implications for improving sample preservation and stabilization in a range of biochemical applications, including microscopy and cell fixation.

Evaluating protein cross-linking as a therapeutic strategy to stabilize SOD1 variants in a mouse model of familial ALS.

Mutations in the gene encoding Cu-Zn superoxide dismutase 1 (SOD1) cause a subset of familial amyotrophic lateral sclerosis (fALS) cases. A shared effect of these mutations is that SOD1, which is normally a stable dimer, dissociates into toxic monomers that seed toxic aggregates. Considerable research effort has been devoted to developing compounds that stabilize the dimer of fALS SOD1 variants, but unfortunately, this has not yet resulted in a treatment. We hypothesized that cyclic thiosulfinate cross-linkers, which selectively target a rare, 2 cysteine-containing motif, can stabilize fALS-causing SOD1 variants in vivo. We created a library of chemically diverse cyclic thiosulfinates and determined structure-cross-linking-activity relationships. A pre-lead compound, "S-XL6," was selected based upon its cross-linking rate and drug-like properties. Co-crystallographic structure clearly establishes the binding of S-XL6 at Cys 111 bridging the monomers and stabilizing the SOD1 dimer. Biophysical studies reveal that the degree of stabilization afforded by S-XL6 (up to 24°C) is unprecedented for fALS, and to our knowledge, for any protein target of any kinetic stabilizer. Gene silencing and protein degrading therapeutic approaches require careful dose titration to balance the benefit of diminished fALS SOD1 expression with the toxic loss-of-enzymatic function. We show that S-XL6 does not share this liability because it rescues the activity of fALS SOD1 variants. No pharmacological agent has been proven to bind to SOD1 in vivo. Here, using a fALS mouse model, we demonstrate oral bioavailability; rapid engagement of SOD1G93A by S-XL6 that increases SOD1G93A's in vivo half-life; and that S-XL6 crosses the blood-brain barrier. S-XL6 demonstrated a degree of selectivity by avoiding off-target binding to plasma proteins. Taken together, our results indicate that cyclic thiosulfinate-mediated SOD1 stabilization should receive further attention as a potential therapeutic approach for fALS.

Effect of structurally different pectin combined with polyphenol on water distribution, protein oxidation and heterocyclic amines formation in meat-based snacks

This research studied the effect of structurally different pectin combined with polyphenol (PP) on the physicochemical properties, inhibition of protein oxidation and heterocyclic amines (HAs) formation in fish meat snacks. The findings demonstrated that pectin with a high degree of esterification (DE) and average chain length (ACL) significantly reduced cooking loss, improved zeta potential values, and reduced particle size when combined with polyphenols, indicating an inhibition of protein aggregation and crosslinking. The addition of PP decreased the carbonyl formation as compared to individual polyphenol and control. Low Field Nuclear Magnetic Resonance (LF-NMR) analysis publicized that PP combination increased the water binding capacity, which resulted in less free water (T22 and T23) in those samples. Furthermore, LC/MS analysis revealed that PP combination, retained the water inside the macromolecules and prevented the relocation of water to the surface, which resulted in the less HAs formation as compared to control. The scanning electron microscopy (SEM) revealed that combination of PP samples formed a compact structure evidenced by less wide holes while control sample revealed loose structure. The findings suggest that 201 and 601 pectin combined with catechin have potential to improve the physicochemical properties and inhibition of hazardous compound formation.

Improvement of Surimi Gel Quality Using Protein Cross-Linker, Hydrocolloids and Protease Inhibitor

Surimi and its products have been consumed widely, especially in Japan and Southeast Asia. Market value and consumer acceptability are mainly determined by the textural properties of surimi gel. Textural and gelling properties are governed by the type of fish, lipid content, processing conditions, and presence of endogenous protease and transglutaminase. In addition, several additives such as protein cross-linkers, hydrocolloids, protease inhibitors, etc. have been used in surimi to improve their gelling properties. Cross-linking is the promising method for protein modification to obtain desirable surimi-based gel products possessing unique textural properties. Endogenous transglutaminase (TGase) plays a profound role in protein cross-linking during setting in surimi paste. For surimi from fish with low setting phenomenon, microbial TGase (MTGase) has been widely used. Plant polyphenols have also been employed for protein crosslinking to strengthen the surimi gel. Polysaccharides mainly the hydrocolloids also strengthen the gel network and increase water holding capacity (WHC) of gel. During heating, endogenous heatactivated proteases that are firmly attached to muscle proteins are active in hydrolyzing myofibrillar proteins. This results in a weaker gel with low WHC. Protease inhibitors form plants as well animals could impede the protease activity, thus maintaining the myofibrillar proteins responsible for the gelation. In addition to the additives, processing methods such as highpressure technology or ohmic heating in combination with those additives have a positive impact on the gel properties. For this review, the role of various kinds of additives namely protein crosslinkers, hydrocolloids and protease inhibitors in surimi gel improvement was revisited.

Encoding Structure in Intrinsically Disordered Protein Biomaterials.

In nature, proteins range from those with highly ordered secondary and tertiary structures to those that completely lack a well-defined three-dimensional structure, termed intrinsically disordered proteins (IDPs). IDPs are generally characterized by one or more segments that have a compositional bias toward small hydrophilic amino acids and proline residues that promote structural disorder and are called intrinsically disordered regions (IDRs). The combination of IDRs with ordered regions and the interactions between the two determine the phase behavior, structure, and function of IDPs. Nature also diversifies the structure of proteins and thereby their functions by hybridization of the proteins with other moieties such as glycans and lipids; for instance, post-translationally glycosylated and lipidated proteins are important cell membrane components. Additionally, diversity in protein structure and function is achieved in nature through cross-linking proteins within themselves or with other domains to create various topologies. For example, an essential characteristic of the extracellular matrix (ECM) is the cross-linking of its network components, including proteins such as collagen and elastin, as well as polysaccharides such as hyaluronic acid (HA). Inspired by nature, synthetic IDP (SynIDP)-based biomaterials can be designed by employing similar strategies with the goal of introducing structural diversity and hence unique physiochemical properties. This Account describes such materials produced over the past decade and following one or more of the following approaches: (1) incorporating highly ordered domains into SynIDPs, (2) conjugating SynIDPs to other moieties through either genetically encoded post-translational modification or chemical conjugation, and (3) engineering the topology of SynIDPs via chemical modification. These approaches introduce modifications to the primary structure of SynIDPs, which are then translated to unique three-dimensional secondary and tertiary structures. Beginning with completely disordered SynIDPs as the point of origin, structure may be introduced into SynIDPs by each of these three unique approaches individually along orthogonal axes or by combinations of the three, enabling bioinspired designs to theoretically span the entire range of three-dimensional structural possibilities. Furthermore, the resultant structures span a wide range of length scales, from nano- to meso- to micro- and even macrostructures. In this Account, emphasis is placed on the physiochemical properties and structural features of the described materials. Conjugates of SynIDPs to synthetic polymers and materials achieved by simple mixing of components are outside the scope of this Account. Related biomedical applications are described briefly. Finally, we note future directions for the design of functional SynIDP-based biomaterials.

Snake-scale stimulated robust biomimetic composite triboelectric layer for energy harvesting and smart health monitoring

Extreme frictional wear of triboelectric layers severely hinders long-term sustainability of triboelectric nanogenerators (TENGs) for efficient ambient energy harvesting. The state-of-the-art solutions necessitate sophisticated structural designs and complex packaging constraints, impeding the feasibility of TENGs for practical applications. Herein, inspired by the structures and compositions of snake scales, a biomimetic composite (BC) film is fabricated as a wear-resistant triboelectric contact layer. The successful formation of disulfide bond-facilitated cross-linkage of cysteine proteins in the keratin-based BC film promotes excellent mechanical resilience and durability, thereby solving the material abrasion challenges associated with solid-solid triboelectric layers. The BC film, with toothed microstructure, exhibits improved charge transfer, low friction, and consistent long-term stable outputs compared to commercial films. Furthermore, a self-powered BC film-based TENG is integrated into a bicycle for biomechanical energy harvesting, lightening warning signals for safer night ride with stable 300 V electrical output for at least 6 h. Moreover, a smart health monitoring system is further proposed by embedding TENG-based sensors into the bicycle saddle, accurately capturing the rider’s pelvic-saddle interactions on varying terrain slopes. By leveraging the unique biomechanical information, machine learning-assisted user identification and terrain slope data classifications are achieved with high accuracies, laying a foundation for futuristic smart healthcare applications. This work provides a strategy to reduce material abrasion by exploiting the unique characteristics of a biomimetic material, facilitating a basis for efficient environmental energy harvesting and smart sensing.

Formation of pH-responsive hydrogel beads and their gel properties: Soybean protein nanofibers and sodium alginate

Hydrogel beads prepared from protein nanofibers are popular because of their safety, sleek appearance, and protection of biologically active substances. However, extreme external environmental variations, such as pH and temperature, can limit their practical application. To meet the application requirements of hydrogel beads in different environments, non-covalent mixtures of CaCl2 cross-linked soybean protein nanofibers (SNF) and sodium alginate (SA) were used to prepare hydrogel beads. In the present study, the hardness (782.48 g) and elasticity of hydrogel beads formed at SNF/SA = 7:3 and CaCl2 concentration of 0.1 mol/L were the maximum. Furthermore, the water content and pH swelling also reached a peak (98.68 %, 43.85 g/g) due to the best morphology and regular internal network structure. Meanwhile, the pH-responsive hydrogel beads with added anthocyanins were able to respond to the ambient pH under different temperatures and pH conditions and maintained color stability during 96 h of storage (ΔE < 5). In this experiment, a pH-responsive hydrogel bead based on soybean protein nanofiber (SNF) and sodium alginate (SA) was prepared by simple ionic crosslinking. It provides a theoretical and experimental basis for the future application of plant protein nanofibers as pH-responsive hydrogel materials.

Modeling the compaction of bacterial chromosomes by biomolecular crowding and the cross-linking protein H-NS

Cells orchestrate the action of various molecules toward organizing their chromosomes. Using a coarse-grained computational model, we study the compaction of bacterial chromosomes by the cross-linking protein H-NS and cellular crowders. In this work, H-NS, modeled as a mobile “binder,” can bind to a chromosome-like polymer with a characteristic binding energy. The simulation results reported here clarify the relative role of biomolecular crowding and H-NS in condensing a bacterial chromosome in a quantitative manner. In particular, they shed light on the nature and degree of crowder and H-NS synergetics: while the presence of crowders enhances H-NS binding to a chromosome-like polymer, the presence of H-NS makes crowding effects more efficient, suggesting two-way synergetics in chain compaction. Also, the results show how crowding effects promote clustering of bound H-NS. For a sufficiently large concentration of H-NS, the cluster size increases with the volume fraction of crowders.

Expanding limits of artificial enzymes: unprecedented catalysis by an oxidase nanozyme in activating a structural protein for covalent crosslinking and conferring remarkable proteolytic resistance.

Nature has endowed us with some complex enzymes capable of utilizing proteins as their substrates to generate functional proteins through post-translational modification. However, nanozymes' interplay with proteins as substrates is scarce, with their chemistry predominantly established using only small molecule substrates, featuring a significant gap in this area. Due to the huge prospects of nanozymes in biotechnological and therapeutic interventions, studies establishing the unexplored roles of nanozymes in the biological environments and their interplay beyond small molecule substrates warrant immediate attention. In this study, we unveil the unprecedented role of a Mn-based oxidase nanozyme (MnN) in activating a structural protein, collagen, and covalently crosslinking its tyrosine residues with only a trace amount of tannic acid (TA) without compromising its triple-helical structural integrity. While therapeutic applications demand materials prepared from collagen, the current chemical and physical crosslinking of collagen often presents significant challenges such as toxicity, denaturation, or high costs. MnN lucidly accomplishes crosslinking interplay at its 101 facets using oxygen as a co-substrate under mild conditions. This process takes advantage of MnN being active at mild acidic pH where collagen preferentially exists as a soluble triple helix (monomeric form), exposing functionalities and enhancing the crosslinking degree. Importantly, this reaction also confers 100% resistance to collagenase attack on the collagen tendon-derived biological material. The catalyzed TA-tyrosine linkage in the telopeptide region of collagen probably impedes the initial recognition step of collagenase, providing robust protection against its degradative action. Our study not only expands the repertoire of nanozymes' substrates beyond the existing library of small molecules but also establishes a significant step toward designing a gold standard for collagen crosslinking. With biomedical applications demanding biomaterials derived from protein scaffolds with preserved structural integrity, our investigation bridges the gap between nanozymes' chemistry and crosslinking proteins, opening exciting prospects for biomaterial development.

Label-free visualization of unfolding and crosslinking mediated protein aggregation in nonenzymatically glycated proteins.

Nonenzymatic glycation (NEG) unfolds and crosslinks proteins, resulting in aggregation. Label-free evaluation of such structural changes, without disturbing molecular integrity, would be beneficial for understanding the fundamental mechanisms of protein aggregation. The current study demonstrates the assessment of NEG-induced protein aggregation by combining autofluorescence (AF) spectroscopy and imaging. The methylglyoxal (MG) induced protein unfolding and the formation of cross-linking advanced glycation end-products (AGEs) leading to aggregation were evaluated using deep-UV-induced-autofluorescence (dUV-AF) spectroscopy in proteins with distinct structural characteristics. Since the AGEs formed on proteins are fluorescent, the study demonstrated the possibility of autofluorescence imaging of NEG-induced protein aggregates. Autofluorescence spectroscopy can potentially reveal molecular alterations such as protein unfolding and cross-linking. In contrast, AGE-based autofluorescence imaging offers a means to visually explore the structural arrangement of aggregates, regardless of whether they are amyloid or non-amyloid in nature.

The upregulation of TGM2 is associated with poor prognosis and the shaping of the inflammatory tumor microenvironment in lung squamous cell carcinoma.

Tissue transglutaminase (TGM2) is a member of the glutamine transferase superfamily, located within cells and their membranes. When secreted, it catalyzes the cross-linking of extracellular matrix proteins and promotes the formation of extracellular matrix scaffolds. To determine the function of TGM2 in the tumorigenesis of lung squamous cell carcinoma (LUSC), we conducted a comprehensive bioinformatics analysis of TGM2. Our findings indicate that high expression of TGM2 in LUSC was associated with a poorer prognosis. Additionally, we found that high expression of TGM2 is closely related to tumor-promoting inflammation and may increase sensitivity to immunotherapy. We further confirmed the cancer-promoting effect of TGM2 in LUSC through in vitro overexpression and knockdown experiments and showed that TGM2 primarily affects cancer cell proliferation, apoptosis, and invasion. In summary, TGM2 promoted the progression of LUSC, and targeting TGM2 is expected to become a new therapeutic approach for LUSC treatment.

Biocompatibility and Osseointegration of the Biomimetically Coated and Water-Soluble Eggshell Membrane Protein Cross-Linked Ti Alloy Screws.

The surface properties of dental and orthopedic implants are directly related to their osseointegration rate. Coating and/or modifying the implant surface might reduce the time of healing. In this study, we aimed to examine the effects of a hybrid surface consisting of a brushite surface coating and cross-linked water-soluble eggshell membrane protein on the osseointegration of titanium (Ti) screws under in vivo conditions. Twenty Ti alloy screws were implanted monocortically in anteromedial regions of New Zealand rabbit tibiae. Ten screws were untreated and used as controls. The remaining 10 screws were coated with calcium phosphate and following cross-linked with ostrich eggshell membrane protein. All rabbits were sacrificed six weeks after the surgery. Peri-screw tissues were evaluated by micro-computed tomography (µ-CT), histological and histomorphometrical methods. The μ-CT assessments indicated that the experimental group had significantly higher mean bone surface area (BSA) and trabeculae number (TbN) than those of the control group (p ˂ 0.05). Bone surface area (BV), trabecular separation (TbSp), trabecular thickness (TbTh), and bone mineral density (BMD) scores of the control and experimental groups were quite similar (p > 0.05). The vascularization score of the experimental group was significantly higher than the control group (4.29 vs. 0.92%). No sign of the graft-versus-host reaction was observed. Our findings reveal that coating Ti alloy implants with calcium phosphate cross-linked with ostrich eggshell membrane protein increases the osseointegration of Ti alloy screws by increasing the bone surface area, number of trabeculae and vascularization in the implant site.

The role of solvation on the conformational landscape of α-synuclein.

Native ion mobility mass spectrometry has been used extensively to characterize ensembles of intrinsically disordered protein (IDP) conformers, but the extent to which the gaseous measurements provide realistic pictures of the solution conformations for such flexible proteins remains unclear. Therefore, we systematically studied the relationship between the solution and gaseous structural ensembles by measuring electrospray charge state and collision cross section (CCS) distributions for cationic and anionic forms of α-synuclein (αSN), an anionic protein in solution, as well as directly probed gas phase residue to residue distances via ion/ion reactions between gaseous α-synuclein cations and disulfonic acid linkers that form strong electrostatic bonds. We also combined results from in-solution protein crosslinking identified from native tandem mass spectrometry (MS/MS) with an initial αSN ensemble generated computationally by IDPConformerGenerator to generate an experimentally restrained solution ensemble of αSN. CCS distributions were directly calculated for the solution ensembles determined by NMR and compared to predicted gaseous conformers. While charge state and collision cross section distributions are useful for qualitatively describing the relative structural dynamics of proteins and major conformational changes induced by changes to solution states, the predicted and measured gas phase conformers include subpopulations that are significantly different than those expected from completely "freezing" solution conformations and preserving them in the gas phase. However, insights were gained on the various roles of solvent in stabilizing various conformers for extremely dynamic proteins like α-synuclein.

Modulation of Aβ 16-22 aggregation by glucose.

The self-assembly of amyloid-beta (Aβ) peptides into fibrillar structures in the brain is a signature of Alzheimer's disease. Recent studies have reported correlations between Alzheimer's disease and type-2 diabetes. Structurally, hyperglycemia induces covalent protein crosslinkings by advanced glycation end products (AGE), which can affect the stability of Aβ oligomers. In this work, we leverage physics-based coarse-grained molecular simulations to probe alternate thermodynamic pathways that affect peptide aggregation propensities at varying concentrations of glucose molecules. Similar to previous experimental reports, our simulations show a glucose concentration-dependent increase in Aβ aggregation rates, without changes in the overall secondary structure content. We discovered that glucose molecules prefer partitioning onto the aggregate-water interface at a specific orientation, resulting in a loss of molecular rotational entropy. This effectively hastens the aggregation rates, as peptide self-assembly can reduce the available surface area for peptide-glucose interactions. This work introduces a new thermodynamic-driven pathway, beyond chemical cross-linking, that can modulate Aβ aggregation.

Glycation-induced Amyloid Formation in Proteins: An Emerging Perspective to Explore Diabetes Associated Onset of Neurodegenerative Symptoms

Abstract: Non-enzymatic protein glycation occurs spontaneously via the formation of sugarprotein Schiff adducts. The end products of this pathway are terminally misfolded proteins popularly known as Advanced Glycation End (AGE) Products. Glycated proteins account for a diverse spectrum of physiological maladies including arteriosclerosis, renal failure, diabetic complications, obesity, and neurological disorders. AGEs not only jeopardise the functionality of modified proteins but also induce the formation of Covalent protein cross-links. Glycation has the potential to induce the unfolding and refolding of globular proteins into cross-β structures thus resembling many amyloid deposits like amyloid beta, tau protein, and prions. However, glycation-induced amyloid formation is not a generic property of proteins; instead, it is guided by the nature and conformation of the protein, the type of glycation agent as well as the solution conditions governing the glycation reaction. The half-lives of AGE adducts are prolonged by their impaired proteasomal clearance since glycation modifies the lysine residues and renders them unavailable for ubiquitination. AGEs are cleared via sequestration with specific cell surface receptors (RAGE); subsequently, downstream signalling events involving MAPK and NF-κB result in the activation of inflammatory response or the apoptotic pathway. This perspective article discusses the current developments in understanding the various facets of glycationassociated protein aggregation and the corresponding development of amyloid-like entities.

Enhanced thermostability of Streptomyces mobaraensis transglutaminase via computation-aided site-directed mutations and structural analysis

Streptomyces mobaraensis transglutaminase (smTG) has been widely used in the food processing industry for protein crosslinking.

Cyclosporine A Protects Podocytes by Regulating Transgelin in Adriamycin-Induced Podocyte Injury.

The calcineurin inhibitor cyclosporine A (CsA) has been shown to effectively reduce proteinuria. However, its precise mechanism is still not fully understood. Our previous study showed that CsA reduced proteinuria by directly stabilizing the foot process (FP) cytoskeletal structure via cofilin-1, suggesting that synaptopodin, a podocyte-specific actin protein, is not the sole target of CsA in podocytes. In this study, we established an adriamycin (ADR)-induced nephropathy rat model and a cultured podocyte injury model. We employed Western blotting and immunofluorescence techniques to assess the expression and distribution of transgelin, Krüppel-like factor-4 (KLF-4), nephrin, and synaptopodin. We observed a significant increase in proteinuria levels accompanied by loss of normal FP structure in the ADR-induced nephropathy rat model. The levels of the actin cross-linking protein transgelin were increased significantly, while those of the podocyte-specific molecules nephrin and synaptopodin were decreased in vivo. Treatment with CsA effectively reduced proteinuria while restoring FP effacement stability in ADR-induced nephropathy models and restoring the expression of transgelin, nephrin, and synaptopodin both in vivo and in vitro. Furthermore, CsA treatment dose-dependently decreased transgelin levels while significantly increasing KLF-4 expression in injured podocytes. In addition, CsA failed to downregulate transgelin when KLF-4 was specifically knocked down. Our findings suggest that CsA protects against podocyte injury by downregulating abnormally high levels of transgelin via upregulation of KLF-4 expression.

Functions and Clinical Significance of Myocardial Cell-Derived Immunoglobulins.

Immunoglobulins (Igs) have been widely accepted to be exclusively expressedby B cells. Nonetheless, this theory is challenged by mounting evidence which suggests thatIgs can also be generated by nonB cells (nonB-Ig), including cardiomyocytes (CM). NonB-Ig exhibits unique physical and chemical characteristics, unique variable region sequences and functions, which diverge from those of B-Ig. For instance, nonB-Ig demonstrates hydrophobicity, limited diversity in the variable region, and extracellular matrix protein activity. Likewise, cardiomyocytes can express different classes of Igs, including IgM, IgG, and free Igκ light chains (cardiomyocyte derived-Igs, CM-Igs). In particular, CM-Igs can be secreted into the extracellular space in various cardiovascular diseases, such as myocardial ischaemia and myocardial fibrosiswhere they might be involved in complement activationand direct damage to cardiomyocytes. Nevertheless, the precise pathological activity of CM-Igs remains unclear. Recently, Zhu et al. focused on studying the sequence characteristics and functions of CM-Igκ; they discovered that the CM-Igκ exhibits a unique VJ recombination pattern, high hydrophobicity, and isprincipally located on the intercalated discs and cross striations of the cardiomyocytes. Interestingly, loss of Igκ in cardiomyocytes results in structural disorders in intercalated discs and dysfunction in myocardial contraction and conduction. Mechanically, Igκ promotes the stabilisation of plectin, a cytoskeleton cross-linker protein that connects desmin to desomsome, to maintain the normal structure of the intercalated disc. This finding indicatesthatCM-Igκ plays an integral role in maintaining cytoskeleton structure. Consequently, it is imperative to reveal the physiological functions and mechanisms of pathological injury associated with CM-Igs.

Linking inflammation and angiogenesis with fibrogenesis: Expression of FXIIIA, MMP-9, and VEGF in oral submucous fibrosis

ObjectivesInterplay of Factor XIIIa (FXIIIa), a transglutaminase, responsible for cross-linking of matrix proteins, Matrix Metalloproteinase-9 (MMP-9), a gelatinase, and Vascular Endothelial Growth Factor (VEGF), an angiogenic inducer, were studied in relation to fibrogenesis and disease progression in oral submucous fibrosis (OSMF). Material and methodsImmunohistochemical expression of markers was studied in 60 formalin-fixed paraffin-embedded tissue blocks of OSMF and 20 normal oral mucosal tissues. FXIIIa was studied quantitatively while MMP-9 and VEGF were assessed semi-quantitatively. Expression was compared with histopathological grades of OSMF. ResultsFXIIIa expression significantly increased in OSMF (p-value 0.000). However, expression decreased and cells became quiescent with increasing grades (p-value 0.000). MMP-9 (p-value epithelium 0.011, p-value connective tissue 0.000) and VEGF expression (p-value epithelium 0.000, connective tissue 0.000) increased in OSMF. A negative correlation between FXIIIa and MMP-9 (−0.653) in early grade (p-value of 0.021) and a positive correlation between FXIIIa and VEGF (0.595) (p-value of 0.032) was found in the moderate grade OSMF. Regression analysis showed a significant association (p<0.01) of FXIIIa in OSMF and with increasing grades of OSMF. ConclusionFXIIIa may play a crucial role in initiation of fibrosis in OSMF. MMP-9 may have a diverse role to play in OSMF as a regulator of fibrosis. VEGF may show an angio-fibrotic switch and contribute to fibrosis in OSMF. These cytokines may show altered function and can contribute to fibrosis and chronicity of disease due to changes in the microenvironment. Tissue stiffness in OSMF itself creates an environment that enhances the chronicity of the disease.