Mythily Srinivasan

Soluble forms of Toll‐like receptor 4 are present in human saliva and modulate tumour necrosis factor‐α secretion by macrophage‐like cells

In health, mucosal inflammation is prevented by tightly regulated responses via Toll‐like receptors (TLR) that interact with specific microbe associated molecular patterns. Currently, 13 TLRs have been identified. Based on the specificity of ligand recognition, TLR‐2 and TLR‐4 can recognize most oral commensal microorganisms. Recent identification of some soluble TLRs (sTLRs) suggests additional regulatory roles for these receptors. We report here the presence of sTLR‐4 polypeptides in adult human saliva. Functionally, the salivary sTLR‐4 suppressed cytokine secretion by activated macrophages. The sTLR‐4 levels were elevated significantly in oral lichen planus (OLP), a chronic inflammatory condition of the oral mucosa characterized by clinical persistence. In contrast, the epithelial cells in the saliva of OLP subjects expressed significantly reduced TLR‐2 and TLR‐4 mRNA that correlated with fewer bacteria/salivary epithelial cells. Investigating the soluble and cellular components of saliva is useful in identifying potential biomarkers for oral mucosal lesions.

Novel p65 binding glucocorticoid-induced leucine zipper peptide suppresses experimental autoimmune encephalomyelitis.

Background: Glucocorticoid-induced leucine zipper (GILZ), a glucocorticoid inducible molecule, physically binds and inhibits nuclear factor-κB (NF-κB). Results: A peptide mimic derived from the p65 binding carboxyl terminus of GILZ inhibits T cells and prevents autoimmune encephalomyelitis. Conclusion: GILZ-p65 interaction is a potential target for multiple sclerosis. Significance: GILZ-peptide may provide a lead for small molecule NF-κB inhibitors. Multiple sclerosis (MS) is a neurological disease characterized by inflammatory demyelination in the brain and spinal cord. The immune-mediated inflammation involves well orchestrated intermolecular interactions that exhibit rapid binding kinetics. The binding interfaces of transient interactions frequently include proline residues that favor an extended conformation for molecular recognition. Linear interface peptides are excellent lead inhibitors of specific protein-protein interactions because only small segments of the interface contribute to the binding. Glucocorticoid-induced leucine zipper (GILZ), a recently identified molecule exhibits potent anti-inflammatory properties. Mechanistically, a proline-rich segment in the carboxyl terminus of GILZ physically binds the p65 subunit of nuclear factor-κB and inhibits the transactivation of inflammatory cytokines. Integrating knowledge derived from the mechanism of action of GILZ with in silico structure prediction identified an immunomodulatory peptide, the GILZ-P. Treatment with GILZ-P exhibited therapeutic efficacy in experimental autoimmune encephalomyelitis, a model for human MS.

Significance of NF-κB as a pivotal therapeutic target in the neurodegenerative pathologies of Alzheimer's disease and multiple sclerosis.

Introduction: Advances in molecular pathogenesis suggest that the chronic inflammation is a shared mechanism in the initiation and progression of multiple neurodegenerative diseases with diverse clinical manifestations such as Alzheimer’s disease (AD) and Multiple sclerosis (MS). Restricted cell renewal and regenerative capacity make the neural tissues extremely vulnerable to the uncontrolled inflammatory process leading to irreversible tissue damage. Areas covered: A predominant consequence of increased inflammatory signaling is the upregulation of the transcription factor, NF-κB with subsequent neuroprotective or deleterious effects depending on the strength of the signal and the type of NF-κB dimers activated. We discuss the interplay between neuroinflammation and neurodegeneration keeping in focus NF-κB signaling as the point of convergence of multiple pathways associated with the development of the neurodegenerative pathologies, AD and MS. Expert opinion: Considerable interest exists in developing efficient NF-κB inhibitors for neurodegenerative diseases. The review includes an overview of natural compounds and rationally designed agents that inhibit NF-κB and mediate neuroprotection in AD and MS. The key chemical moieties of the natural and the synthetic compounds provide efficient leads for the development of effective small molecule inhibitors that selectively target NF-κB activation; this would result in the desired benefit to risk therapeutic effects.

Suppression of colon inflammation by CD80 blockade: Evaluation in two murine models of inflammatory bowel disease

Background: Human inflammatory bowel disease (IBD) is a chronic condition mediated by aberrant immune responses to the luminal antigens by activated CD4+ T cells. The CD80/CD86:CD28/CD152 costimulatory pathways transmit signals critical for T cell activation and suppression. Macrophages and epithelial cells are the chief antigen‐presenting cells in the gut. Macrophages from the IBD colon express significantly elevated levels of CD80 and CD86 costimulatory molecules. The CD28‐CD80 interaction primarily participates in breaking the tolerance and inducing the immune response in murine models of colitis. Blockade of CD80‐costimulatory axis is an attractive strategy in the treatment of IBD. Methods: Incorporating the structural information of the CD80:CD152 complex together with the preferences of interface residues to form polyproline type II helix, we designed novel peptide agents that selectively blocked CD80 receptor interactions. Results: Administration of CD80 blocking agent at the time of adoptive transfer prevented the SCID mice from CD4+CD45Rbhigh T‐cell mediated colitis. Significantly, CD80‐CAP (competitive antagonist peptide) treatment suppressed established inflammation in TNBS‐induced colitis, a model for Th1‐mediated Crohns disease. The colons of the mice receiving the CD80 blocking agent appeared unaffected macroscopically and exhibited negligible microscopic inflammation. The CD80‐CAP treatment was associated with significantly reduced Th1 cytokines in the colon. Conclusions: The CD80 blocking peptide appeared to mediate protection against colitis by inducing Th2 skewing of the cytokine response.

CD80 Binding Polyproline Helical Peptide Inhibits T Cell Activation

The critical role played by the CD28/CD152-CD80/CD86 costimulatory molecules in mediating T cell activation and suppression provides attractive targets for therapeutic strategies. CD28 and CD152 share a conserved polyproline motif in the ligand-binding region. Similar proline-rich regions in globular domains preferentially adopt a polyproline type II (PP) helical conformation and are involved in transient IIprotein-protein interactions. Interestingly, in the human CD80-CD152 complex, Pro102 of CD152 restricts the preceding proline to PPII helix in the binding orientation in relation to the shallow binding pocket of CD80. Peptide agents derived from binding sites of receptors that mimic the bioactive conformation have been shown to block receptor-ligand interactions. Contact preferences of the interface amino acids at the protein-protein interaction sites and the propensity of each residue to form PPII helix were integrated in the design of novel peptide agents referred to as CD80 competitive antagonist peptides. Structural and functional studies suggest potential therapeutic value for select CD80 competitive antagonist peptides.

Proline Rich Motifs as Drug Targets in Immune Mediated Disorders

The current version of the human immunome network consists of nearly 1400 interactions involving approximately 600 proteins. Intermolecular interactions mediated by proline-rich motifs (PRMs) are observed in many facets of the immune response. The proline-rich regions are known to preferentially adopt a polyproline type II helical conformation, an extended structure that facilitates transient intermolecular interactions such as signal transduction, antigen recognition, cell-cell communication and cytoskeletal organization. The propensity of both the side chain and the backbone carbonyls of the polyproline type II helix to participate in the interface interaction makes it an excellent recognition motif. An advantage of such distinct chemical features is that the interactions can be discriminatory even in the absence of high affinities. Indeed, the immune response is mediated by well-orchestrated low-affinity short-duration intermolecular interactions. The proline-rich regions are predominantly localized in the solvent-exposed regions such as the loops, intrinsically disordered regions, or between domains that constitute the intermolecular interface. Peptide mimics of the PRM have been suggested as potential antagonists of intermolecular interactions. In this paper, we discuss novel PRM-mediated interactions in the human immunome that potentially serve as attractive targets for immunomodulation and drug development for inflammatory and autoimmune pathologies.

Literature-Based Discovery of Salivary Biomarkers for Type 2 Diabetes Mellitus

The alarming increase in type 2 diabetes mellitus (T2DM) underscores the need for efficient screening and preventive strategies. Select protein biomarker profiles emerge over time during T2DM development. Periodic evaluation of these markers will increase the predictive ability of diabetes risk scores. Noninvasive methods for frequent measurements of biomarkers are increasingly being investigated. Application of salivary diagnostics has gained importance with the establishment of significant similarities between the salivary and serum proteomes. The objective of this study is to identify T2DM–specific salivary biomarkers by literature–based discovery. A serial interrogation of the PubMed database was performed using MeSH terms of specific T2DM pathological processes in primary and secondary iterations to compile cohorts of T2DM–specific serum markers. Subsequent search consisted of mining for the identified serum markers in human saliva. More than 60% of T2DM–associated serum proteins have been measured in saliva. Nearly half of these proteins have been reported in diabetic saliva. Measurements of salivary lipids and oxidative stress markers that can exhibit correlated saliva plasma ratio could constitute reliable factors for T2DM risk assessment. We conclude that a high percentage of T2DM–associated serum proteins can be measured in saliva, which offers an attractive and economical strategy for T2DM screening.

CD80 Blockade Enhance Glucocorticoid-Induced Leucine Zipper Expression and Suppress Experimental Autoimmune Encephalomyelitis

Designing mimetic of the interface functional groups of known receptor-ligand complexes is an attractive strategy for developing potential therapeutic agents that interfere with target protein-protein interactions. The CD80/CD86-CD28/CD152 costimulatory interactions transmit signals for CD4+ T cell activation and suppression and are critically involved in the initiation, progression, and reactivation of the immunopathology in multiple sclerosis. Differences in the pattern, levels, and kinetics of expression of CD80/CD86 molecules in conjunction with differences in the strength of the signals delivered upon binding CD28 or CD152 determine the outcome of the immune response. A temporal up-regulation of surface expression of CD80 relative to CD86 on APCs and CNS-infiltrating cells has been shown to correlate with disease progression in experimental autoimmune encephalomyelitis an animal model for multiple sclerosis. Hence blockade of the CD80 costimulatory axis has therapeutic potential in multiple sclerosis. In this study, we report the efficacy of a novel CD80-blocking agent CD80-competitive antagonist peptide (CD80-CAP) in suppressing clinical disease and relapse in experimental autoimmune encephalomyelitis. The CD80-CAP mediates protection by inhibiting proinflammatory cytokines and skewing toward anti-inflammatory response presumably by enhancing the expression of glucocorticoid-induced leucine zipper in activated CD4+ T cells.

Glucocorticoid-Induced Leucine Zipper in Central Nervous System Health and Disease

The central nervous system (CNS) is a large network of intercommunicating cells that function to maintain tissue health and homeostasis. Considerable evidence suggests that glucocorticoids exert both neuroprotective and neurodegenerative effects on the CNS. Glucocorticoids act by binding two related receptors in the cytoplasm, the mineralocorticoid receptor (MR) and the glucocorticoid receptor (GR). The glucocorticoid receptor complex mediates cellular responses by transactivating target genes and by protein: protein interactions. The paradoxical effects of glucocorticoids on neuronal survival and death have been attributed to the concentration and the ratio of mineralocorticoid to glucocorticoid receptor activation. Glucocorticoid-induced leucine zipper (GILZ) is a recently identified protein transcriptionally upregulated by glucocorticoids. Constitutively, expressed in many tissues including brain, GILZ mediates many of the actions of glucocorticoids. It mimics the anti-inflammatory and anti-proliferative effects of glucocorticoids but exerts differential effects on stem cell differentiation and lineage development. Recent experimental data on the effects of GILZ following induced stress or trauma suggest potential roles in CNS diseases. Here, we provide a short overview of the role of GILZ in CNS health and discuss three potential rationales for the role of GILZ in Alzheimer’s disease pathogenesis.

Epithelial expression of keratinocytes growth factor in oral precancer lesions

Background: Keratinocyte growth factor (KGF) is a potent epithelial mitogen that acts by binding the KGF receptors (KGFRs) expressed on epithelial cells and regulates proliferation and differentiation. The objective of this study was to investigate the expression of KGF in the epithelium in oral precancer. Materials and Methods: Archival tissues of oral submucous fibrosis (SMF) and leukoplakia were assessed for epithelial KGF expression by immunohistochemistry and real-time quantitative polymerase chain reaction. Results: KGF was predominantly expressed in the basal and parabasal cells in the epithelium of SMF tissues. KGF transcript in the epithelial cells increased with increasing severity of epithelial dysplasia in oral leukoplakia. Conclusion: Although widely reported as a product secreted by the mesenchymal cells, our data suggest that the KGF is also expressed in oral epithelial cells much like the expression in ovarian epithelial cells. Based on the localization of KGF in cells at the epithelial mesenchymal junction and that of the reported presence of KGFR in oral keratinocytes, a potential mechanism involving paracrine and autocrine interactions of KGF and KGFR in early stages of oral precancer is postulated.