Mechanisms of NF-κB p65 and strategies for therapeutic manipulation

The transcription factor NF-κB is a critical regulator of immune and inflammatory responses. In mammals, the NF-κB/Rel family comprises five members: p50, p52, p65 (Rel-A), c-Rel, and Rel-B proteins, which form homo- or heterodimers and remain as an inactive complex with the inhibitory molecules called IκB proteins in resting cells. Two distinct NF-κB signaling pathways have been described: 1) the canonical pathway primarily activated by pathogens and inflammatory mediators, and 2) the noncanonical pathway mostly activated by developmental cues. The most abundant form of NF-κB activated by pathologic stimuli via the canonical pathway is the p65:p50 heterodimer. Disproportionate increase in activated p65 and subsequent transactivation of effector molecules is integral to the pathogenesis of many chronic diseases such as the rheumatoid arthritis, inflammatory bowel disease, multiple sclerosis, and even neurodegenerative pathologies. Hence, the NF-κB p65 signaling pathway has been a pivotal point for intense drug discovery and development. This review begins with an overview of p65-mediated signaling followed by discussion of strategies that directly target NF-κB p65 in the context of chronic inflammation

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

Psychometric Characteristics of Oral Pathology Test Items in the Dental Hygiene Curriculum—A Longitudinal Analysis

As the landscape of oral healthcare and the delivery of services continue to undergo change, the dental hygienist plays an increasing role in assisting dentists with oral diagnosis and preventive strategies. Hence, the dental hygiene curriculum standards require biomedical science instructions, including general and oral pathology. Student learning and cognitive competencies are often measured using multiple-choice questions (MCQs). The objectives of this study were to perform a longitudinal analysis of test items and to evaluate their relation to the absolute grades of the oral pathology course in the dental hygiene curriculum. A total of 1033 MCQs covering different concepts of oral pathology administered from 2015 through 2019 were analyzed for difficulty and discriminatory indices, and the differences between the years were determined by one-way ANOVA. Test reliability as determined by the average KR-20 value was 0.7 or higher for each exam. The mean difficulty index for all exams was 0.73 +/− 0.05, and that of the discriminatory index was 0.33 +/− 0.05. Wide variations were observed in the discriminatory indices of test items with approximately the same difficulty index, as well as in the grade distribution in each cohort. Furthermore, longitudinal data analyses identified low achieving cohorts amongst the groups evaluated for the same knowledge domain, taught with the same instruction, and using similar test tools. This suggest that comparative analyses of tests could offer feedback not only on student learning attributes, but also potentially on the admission processes to the dental hygiene program

Significance of NF-κΒ 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

Functional characterization of a competitive peptide antagonist of p65 in human macrophage- like cells suggests therapeutic potential for chronic inflammation

Glucocorticoid-induced leucine zipper (GILZ) is a glucocorticoid responsive protein that links the nuclear factor-kappa B (NFκB) and the glucocorticoid signaling pathways. Functional and binding studies suggest that the proline-rich region at the carboxy terminus of GILZ binds the p65 subunit of NFκB and suppresses the immunoinflammatory response. A widely-used strategy in the discovery of peptide drugs involves exploitation of the complementary surfaces of naturally occurring binding partners. Previously, we observed that a synthetic peptide (GILZ-P) derived from the proline-rich region of GILZ bound activated p65 and ameliorated experimental encephalomyelitis. Here we characterize the secondary structure of GILZ-P by circular dichroic analysis. GILZ-P adopts an extended polyproline type II helical conformation consistent with the structural conformation commonly observed in interfaces of transient intermolecular interactions. To determine the potential application of GILZ-P in humans, we evaluated the toxicity and efficacy of the peptide drug in mature human macrophage-like THP-1 cells. Treatment with GILZ-P at a wide range of concentrations commonly used for peptide drugs was nontoxic as determined by cell viability and apoptosis assays. Functionally, GILZ-P suppressed proliferation and glutamate secretion by activated macrophages by inhibiting nuclear translocation of p65. Collectively, our data suggest that the GILZ-P has therapeutic potential in chronic CNS diseases where persistent inflammation leads to neurodegeneration such as multiple sclerosis and Alzheimer’s disease

Soluble toll like receptor 2 (TLR-2) is increased in saliva of children with dental caries

Background Dental caries is the most common microbial disease affecting mankind. Caries risk assessment methods, identification of biomarkers and vaccine development strategies are being emphasized to control the incidence of the largely preventable disease. Pattern recognition receptors such as the toll like receptors (TLR) have been implicated as modulators of host-microbial interactions. Soluble TLR-2 and its co-receptor, CD14 identified in saliva can bind the cell wall components of cariogenic bacteria and modulate the disease process. The objective of this study is to determine the potential of salivary sTLR-2 and sCD14 as biomarkers of caries activity and indirect measures of the cariogenic bacterial burden. Methods Unstimulated whole saliva was collected from twenty caries free and twenty caries active children between the ages of 5 and 13 years. The concentration of sCD14 and sTLR-2 together with that of the cytokine IL-8 reported to be increased in dental caries was assessed by the enzyme linked immunosorbent assay. Results While the level of sCD14 and that of IL-8 was equivocal between the two groups, the sTLR-2 concentration in caries active saliva was significantly higher than that in caries free saliva. Conclusions The sTLR-2 in saliva could serve as a potential biomarker for caries activity

Oral epithelial expression of angiotensin converting enzyme-2: Implications for COVID-19 diagnosis and prognosis

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) uses the angiotensin converting enzyme (ACE)-2 as the host receptor for target cell entry. The extent and distribution of ACE-2 has been associated with the clinical symptoms of coronavirus disease (COVID)-19. Here we show by immunofluorescence analysis that the ACE2 is abundantly expressed in oral mucosa, particularly in the surface epithelial cells suggesting that these cells could represent sites of entry for SARS-CoV-2. Further, together with the reports on ACE2 ectodomain shedding, we discuss the rationale for the hypothesis that the ACE-2 measurement in saliva could be a marker for COVID-19 infection during early phase following SARS-CoV-2 exposure

Novel Nuclear Factor-KappaB Targeting Peptide Suppresses β-Amyloid Induced Inflammatory and Apoptotic Responses in Neuronal Cells

In the central nervous system (CNS), activation of the transcription factor nuclear factor-kappa B (NF-κβ) is associated with both neuronal survival and increased vulnerability to apoptosis. The mechanisms underlying these dichotomous effects are attributed to the composition of NF-κΒ dimers. In Alzheimer’s disease (AD), β-amyloid (Aβ) and other aggregates upregulate activation of p65:p50 dimers in CNS cells and enhance transactivation of pathological mediators that cause neuroinflammation and neurodegeneration. Hence selective targeting of activated p65 is an attractive therapeutic strategy for AD. Here we report the design, structural and functional characterization of peptide analogs of a p65 interacting protein, the glucocorticoid induced leucine zipper (GILZ). By virtue of binding the transactivation domain of p65 exposed after release from the inhibitory IκΒ proteins in activated cells, the GILZ analogs can act as highly selective inhibitors of activated p65 with minimal potential for off-target effects

A nuclear factor-kappa B inhibiting peptide suppresses innate immune receptors and gliosis in a transgenic mouse model of Alzheimer’s disease

A disproportionate increase in activated nuclear factor-kappa B (NF-κB) has been shown to drive the Aβ deposition, neuroinflammation and neurodegeneration in Alzheimer’s disease (AD). Hence, selective targeting of activated p65 represents an attractive therapeutic approach for AD. Glucocorticoid induced leucine zipper (GILZ) is a NF-κB interactant that binds and sequesters the activated p65 in the cytoplasm. The p65 binding domain of GILZ adopts a polyproline type II helical conformation, a motif that acts as an adaptable glove in the interface with the binding partner and constitutes an excellent template for drug design. Previously, peptide analogs of the p65 binding domain of GILZ, referred to as GA have been shown to suppress pathology in the lipopolysaccharide induced model of neuroinflammation. In this study, we investigated the CNS delivery of labeled GA administered intraperitoneally in adult mice for a period of upto 24 h. Further, we evaluated the suppressive potential of GA in 5xFAD mice, an aggressive model with five genetic mutations closely associated with human AD. Groups of 5xFAD mice administered GA or control peptide intraperitoneally on alternate days for six weeks were evaluated for Aβ deposition, microglia, inflammation and innate immune responses by immunohistochemistry and real time polymerase reaction. GA was observed in proximity with NeuN positive neurons suggesting that the compound crossed the blood brain barrier to reach the brain parenchyma. Further, GA treatment decreased Aβ load, reduced Iba1 + microglia and glial fibrillary acidic protein (GFAP)+ astrocytes, inhibited inflammatory cytokines and suppressed toll like receptor (TLR-2, TLR-4) expressions in 5xFAD mice