Multiomics Integration in Skin Diseases with Alterations in Notch Signaling Pathway: PlatOMICs Phase 1 Deployment

The high volume of information produced in the age of omics was and still is an important step to understanding several pathological processes, providing the enlightenment of complex molecular networks and the identification of molecular targets associated with many diseases. Despite these remarkable scientific advances, the majority of the results are disconnected and divergent, making their use limited. Skin diseases with alterations in the Notch signaling pathway were extensively studied during the omics era. In the GWAS Catalog, considering only studies on genomics association (GWAS), several works were deposited, some of which with divergent results. In addition, there are thousands of scientific articles available about these skin diseases. In our study, we focused our attention on skin diseases characterized by the impairment of Notch signaling, this pathway being of pivotal importance in the context of epithelial disorders. We considered the pathologies of five human skin diseases, Hidradenitis Suppurativa, Dowling Degos Disease, Adams–Oliver Syndrome, Psoriasis, and Atopic Dermatitis, in which the molecular alterations in the Notch signaling pathway have been reported. To this end, we started developing a new multiomics platform, PlatOMICs, to integrate and re-analyze omics information, searching for the molecular interactions involved in the pathogenesis of skin diseases with alterations in the Notch signaling pathway.     

Signaling Involved in Hair Follicle Morphogenesis and Development

Hair follicle morphogenesis depends on Wnt, Shh, Notch, BMP and other signaling pathways interplay between epithelial and mesenchymal cells. The Wnt pathway plays an essential role during hair follicle induction, Shh is involved in morphogenesis and late stage differentiation, Notch signaling determines stem cell fate while BMP is involved in cellular differentiation. The Wnt pathway is considered to be the master regulator during hair follicle morphogenesis. Wnt signaling proceeds through EDA/EDAR/NF-κB signaling. NF-κB regulates the Wnt pathway and acts as a signal mediator by upregulating the expression of Shh ligand. Signal crosstalk between epithelial and mesenchymal cells takes place mainly through primary cilia. Primary cilia formation is initiated with epithelial laminin-511 interaction with dermal β-1 integrin, which also upregulates expression of downstream effectors of Shh pathway in dermal lineage. PDGF signal transduction essential for crosstalk is mediated through epithelial PDGF-A and PDGFRα expressed on the primary cilia. Dermal Shh and PDGF signaling up-regulates dermal noggin expression; noggin is a potent inhibitor of BMP signaling which helps in counteracting BMP mediated β-catenin inhibition. This interplay of signaling between the epithelial and dermal lineage helps in epithelial Shh signal amplification. The dermal Wnt pathway helps in upregulation of epithelial Notch expression. Dysregulation of these pathways leads to certain abnormalities and in some cases even tumor outgrowth.     

Enhanced Osteogenic Differentiation of Pluripotent Stem Cells via γ-Secretase Inhibition

Bone healing is a complex, well-organized process. Multiple factors regulate this process, including growth factors, hormones, cytokines, mechanical stimulation, and aging. One of the most important signaling pathways that affect bone healing is the Notch signaling pathway. It has a significant role in controlling the differentiation of bone mesenchymal stem cells and forming new bone. Interventions to enhance the healing of critical-sized bone defects are of great importance, and stem cell transplantations are eminent candidates for treating such defects. Understanding how Notch signaling impacts pluripotent stem cell differentiation can significantly enhance osteogenesis and improve the overall healing process upon transplantation. In Rancourt’s lab, mouse embryonic stem cells (ESC) have been successfully differentiated to the osteogenic cell lineage. This study investigates the role of Notch signaling inhibition in the osteogenic differentiation of mouse embryonic and induced pluripotent stem cells (iPS). Our data showed that Notch inhibition greatly enhanced the differentiation of both mouse embryonic and induced pluripotent stem cells.     

Monogenic Autoinflammatory Diseases: State of the Art and Future Perspectives

Systemic autoinflammatory diseases are a heterogeneous family of disorders characterized by a dysregulation of the innate immune system, in which sterile inflammation primarily develops through antigen-independent hyperactivation of immune pathways. In most cases, they have a strong genetic background, with mutations in single genes involved in inflammation. Therefore, they can derive from different pathogenic mechanisms at any level, such as dysregulated inflammasome-mediated production of cytokines, intracellular stress, defective regulatory pathways, altered protein folding, enhanced NF-kappaB signalling, ubiquitination disorders, interferon pathway upregulation and complement activation. Since the discover of pathogenic mutations of the pyrin-encoding gene MEFV in Familial Mediterranean Fever, more than 50 monogenic autoinflammatory diseases have been discovered thanks to the advances in genetic sequencing: the advent of new genetic analysis techniques and the discovery of genes involved in autoinflammatory diseases have allowed a better understanding of the underlying innate immunologic pathways and pathogenetic mechanisms, thus opening new perspectives in targeted therapies. Moreover, this field of research has become of great interest, since more than a hundred clinical trials for autoinflammatory diseases are currently active or recently concluded, allowing us to hope for considerable acquisitions for the next few years. General paediatricians need to be aware of the importance of this group of diseases and they should consider autoinflammatory diseases in patients with clinical hallmarks, in order to guide further examinations and refer the patient to a specialist rheumatologist. Here we resume the pathogenesis, clinical aspects and diagnosis of the most important autoinflammatory diseases in children.     

Deficiency of Lipin2 Results in Enhanced NF-κB Signaling and Osteoclast Formation in RAW-D Murine Macrophages

Lipin2 is a phosphatidate phosphatase that plays critical roles in fat homeostasis. Alterations in Lpin2, which encodes lipin2, cause the autoinflammatory bone disorder Majeed syndrome. Lipin2 limits lipopolysaccharide (LPS)-induced inflammatory responses in macrophages. However, little is known about the precise molecular mechanisms underlying its anti-inflammatory function. In this study, we attempted to elucidate the molecular link between the loss of lipin2 function and autoinflammatory bone disorder. Using a Lpin2 knockout murine macrophage cell line, we showed that lipin2 deficiency enhances innate immune responses to LPS stimulation through excessive activation of the NF-κB signaling pathway, partly because of TAK1 signaling upregulation. Lipin2 depletion also enhanced RANKL-mediated osteoclastogenesis and osteoclastic resorption activity accompanied by NFATc1 dephosphorylation and increased nuclear accumulation. These results suggest that lipin2 suppresses the development of autoinflammatory bone disorder by fine-tuning proinflammatory responses and osteoclastogenesis in macrophages. Therefore, this study provides insights into the molecular pathogenesis of monogenic autoinflammatory bone disorders and presents a potential therapeutic intervention.     

Advances in Understanding Activation and Function of the NLRC4 Inflammasome

Innate immune receptors initiate a host immune response, or inflammatory response, upon detecting pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs). Among the innate immune receptors, nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) play a pivotal role in detecting cytosolic PAMPs and DAMPs. Some NLRs can form a multiprotein cytosolic complex known as the inflammasome. Inflammasome activation triggers caspase-1–mediated cleavage of the pore-forming protein gasdermin D (GSDMD), which drives a form of inflammatory cell death called pyroptosis. Parallelly, activated caspase-1 cleaves immature cytokines pro–IL-1β and pro–IL-18 into their active forms, which can be released via GSDMD membrane pores. The NLR family apoptosis inhibitory proteins (NAIP)-NLR family caspase-associated recruitment domain-containing protein 4 (NLRC4) inflammasome is important for mounting an immune response against Gram-negative bacteria. NLRC4 is activated through NAIPs sensing type 3 secretion system (T3SS) proteins from Gram-negative bacteria, such as Salmonella Typhimurium. Mutations in NAIPs and NLRC4 are linked to autoinflammatory disorders in humans. In this review, we highlight the role of the NAIP/NLRC4 inflammasome in host defense, autoinflammatory diseases, cancer, and cell death. We also discuss evidence pointing to a role of NLRC4 in PANoptosis, which was recently identified as a unique inflammatory programmed cell death pathway with important physiological relevance in a range of diseases. Improved understanding of the NLRC4 inflammasome and its potential roles in PANoptosis paves the way for identifying new therapeutic strategies to target disease.     

How to Build and to Protect the Neuromuscular Junction: The Role of the Glial Cell Line-Derived Neurotrophic Factor

The neuromuscular junction (NMJ) is at the crossroad between the nervous system (NS) and the muscle. Following neurotransmitter release from the motor neurons (MNs), muscle contraction occurs and movement is generated. Besides eliciting muscle contraction, the NMJ represents a site of chemical bidirectional interplay between nerve and muscle with the active participation of Schwann cells. Indeed, signals originating from the muscle play an important role in synapse formation, stabilization, maintenance and function, both in development and adulthood. We focus here on the contribution of the Glial cell line-Derived Neurotrophic Factor (GDNF) to these processes and to its potential role in the protection of the NMJ during neurodegeneration. Historically related to the maintenance and survival of dopaminergic neurons of the substantia nigra, GDNF also plays a fundamental role in the peripheral NS (PNS). At this level, it promotes muscle trophism and it participates to the functionality of synapses. Moreover, compared to the other neurotrophic factors, GDNF shows unique peculiarities, which make its contribution essential in neurodegenerative disorders. While describing the known structural and functional changes occurring at the NMJ during neurodegeneration, we highlight the role of GDNF in the NMJ–muscle cross-talk and we review its therapeutic potential in counteracting the degenerative process occurring in the PNS in progressive and severe diseases such as Alzheimer’s disease (AD), Amyotrophic Lateral Sclerosis (ALS) and Spinal Muscular Atrophy (SMA). We also describe functional 3D neuromuscular co-culture systems that have been recently developed as a model for studying both NMJ formation in vitro and its involvement in neuromuscular disorders.     

Generalized Pustular Psoriasis: Divergence of Innate and Adaptive Immunity

Generalized pustular psoriasis (GPP) is a severe, relapsing, immune-mediated disease characterized by the presence of multiple sterile pustules all over the body. The exact pathomechanisms behind GPP remain elusive, although increased interest in the genetic basis and immunological disturbances have provided some revealing insights into the underlying signaling pathways and their mutual interaction. The genetic background of GPP has been thoroughly investigated over the past few years. The conducted studies have identified genetic variants that predispose to pustular forms of psoriasis. The loss-of-function mutation of the interleukin 36 receptor antagonist gene, along with rare gain-of-function mutations in the gene that encodes the keratinocyte signaling molecule (CARD14), are examples of the uncovered abnormalities. Interleukin 36 (IL-36), along with neutrophils, is now considered a central cytokine in GPP pathogenesis, with IL-36 signaling providing a link between innate and adaptive immune responses. More recently, a new concept of inflammation, caused by a predominantly genetically determined abnormal activation of innate immune response and leading to inflammatory keratinization, has arisen. GPP is currently considered a representative of this novel group of skin conditions, called autoinflammatory keratinization diseases. As no therapeutic agents have been approved for GPP to date in the United States and Europe, the novel anti-IL-36R antibodies are particularly promising and may revolutionize management of the disease.     

The Role of Gut Microbiota and Gut–Brain Interplay in Selected Diseases of the Central Nervous System

The gut microbiome has attracted increasing attention from researchers in recent years. The microbiota can have a specific and complex cross-talk with the host, particularly with the central nervous system (CNS), creating the so-called “gut–brain axis”. Communication between the gut, intestinal microbiota, and the brain involves the secretion of various metabolites such as short-chain fatty acids (SCFAs), structural components of bacteria, and signaling molecules. Moreover, an imbalance in the gut microbiota composition modulates the immune system and function of tissue barriers such as the blood–brain barrier (BBB). Therefore, the aim of this literature review is to describe how the gut–brain interplay may contribute to the development of various neurological disorders, combining the fields of gastroenterology and neuroscience. We present recent findings concerning the effect of the altered microbiota on neurodegeneration and neuroinflammation, including Alzheimer’s and Parkinson’s diseases, as well as multiple sclerosis. Moreover, the impact of the pathological shift in the microbiome on selected neuropsychological disorders, i.e., major depressive disorders (MDD) and autism spectrum disorder (ASD), is also discussed. Future research on the effect of balanced gut microbiota composition on the gut–brain axis would help to identify new potential opportunities for therapeutic interventions in the presented diseases.     

Nanoparticle-Mediated Pulmonary Drug Delivery: A Review

Colloidal drug delivery systems have been extensively investigated as drug carriers for the application of different drugs via different routes of administration. Systems, such as solid lipid nanoparticles, polymeric nanoparticles and liposomes, have been investigated for a long time for the treatment of various lung diseases. The pulmonary route, owing to a noninvasive method of drug administration, for both local and systemic delivery of an active pharmaceutical ingredient (API) forms an ideal environment for APIs acting on pulmonary diseases and disorders. Additionally, this route offers many advantages, such as a high surface area with rapid absorption due to high vascularization and circumvention of the first pass effect. Aerosolization or inhalation of colloidal systems is currently being extensively studied and has huge potential for targeted drug delivery in the treatment of various diseases. Furthermore, the surfactant-associated proteins present at the interface enhance the effect of these formulations by decreasing the surface tension and allowing the maximum effect. The most challenging part of developing a colloidal system for nebulization is to maintain the critical physicochemical parameters for successful inhalation. The following review focuses on the current status of different colloidal systems available for the treatment of various lung disorders along with their characterization. Additionally, different in vitro, ex vivo and in vivo cell models developed for the testing of these systems with studies involving cell culture analysis are also discussed.     

Regulation of the Expression,Oligomerisation and Signaling of the Inhibitory Receptor CLEC12A by Cysteine Residues in the Stalk Region

CLEC12A is a myeloid inhibitory receptor that negatively regulates inflammation in mouse models of autoimmune and autoinflammatory arthritis. Reduced CLEC12A expression enhances myeloid cell activation and inflammation in CLEC12A knock-out mice with collagen antibody-induced or gout-like arthritis. Similarly to other C-type lectin receptors, CLEC12A harbours a stalk domain between its ligand binding and transmembrane domains. While it is presumed that the cysteines in the stalk domain have multimerisation properties, their role in CLEC12A expression and/or signaling remain unknown. We thus used site-directed mutagenesis to determine whether the stalk domain cysteines play a role in CLEC12A expression, internalisation, oligomerisation, and/or signaling. Mutation of C118 blocks CLEC12A transport through the secretory pathway diminishing its cell-surface expression. In contrast, mutating C130 does not affect CLEC12A cell-surface expression but increases its oligomerisation, inducing ligand-independent phosphorylation of the receptor. Moreover, we provide evidence that CLEC12A dimerisation is regulated in a redox-dependent manner. We also show that antibody-induced CLEC12A cross-linking induces flotillin oligomerisation in insoluble membrane domains in which CLEC12A signals. Taken together, these data indicate that the stalk cysteines in CLEC12A differentially modulate this inhibitory receptor’s expression, oligomerisation and signaling, suggestive of the regulation of CLEC12A in a redox-dependent manner during inflammation.     

Osteoblast Dysfunction in Non-Hereditary Sclerosing Bone Diseases

A review of the available literature was performed in order to summarize the existing evidence between osteoblast dysfunction and clinical features in non-hereditary sclerosing bone diseases. It has been known that proliferation and migration of osteoblasts are concerted by soluble factors such as fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), transforming growth factor (TGF), bone morphogenetic protein (BMP) but also by signal transduction cascades such as Wnt signaling pathway. Protein kinases play also a leading role in triggering the activation of osteoblasts in this group of diseases. Post-zygotic changes in mitogen-activated protein kinase (MAPK) have been shown to be associated with sporadic cases of Melorheostosis. Serum levels of FGF and PDGF have been shown to be increased in myelofibrosis, although studies focusing on Sphingosine-1-phosphate receptor was shown to be strongly expressed in Paget disease of the bone, which may partially explain the osteoblastic hyperactivity during this condition. Pathophysiological mechanisms of osteoblasts in osteoblastic metastases have been studied much more thoroughly than in rare sclerosing syndromes: striking cellular mechanisms such as osteomimicry or complex intercellular signaling alterations have been described. Further research is needed to describe pathological mechanisms by which rare sclerosing non hereditary diseases lead to osteoblast dysfunction.     

Low-Dose Interleukin-2 Therapy: A Driver of an Imbalance between Immune Tolerance and Autoimmunity

For many years, the role of interleukin-2 (IL-2) in autoimmune responses was established as a cytokine possessing strong pro-inflammatory activity. Studies of the past few years have changed our knowledge on IL-2 in autoimmune chronic inflammation, suggesting its protective role, when administered at low-doses. The disrupted balance between regulatory and effector T cells (Tregs and Teffs, respectively) is a characteristic of autoimmune diseases, and is dependent on homeostatic cytokines, including IL-2. Actually, inherent defects in the IL-2 signaling pathway and/or levels leading to Treg compromised function and numbers as well as Th17 expansion have been attributed to autoimmune disorders. In this review, we discuss the role of IL-2 in the pathogenesis of autoimmune diseases. In particular, we highlight the impact of the dysregulated IL-2 pathway on disruption of the Treg/Th17 balance, reversal of which appears to be a possible mechanism of the low-dose IL-2 treatment. The negative effects of IL-2 on the differentiation of follicular helper T cells (Tfh) and pathogenic Th17 cells, both of which contribute to autoimmunity, is emphasized in the paper as well. We also compare the current IL-2-based therapies of animal and human subjects with immune-mediated diseases aimed at boosting the Treg population, which is the most IL-2-dependent cell subset desirable for sufficient control of autoimmunity. New perspectives of therapeutic approaches focused on selective delivery of IL-2 to inflamed tissues, thus allowing local activity of IL-2 to be combined with its reduced systemic and pleiotropic toxicity, are also proposed in this paper.     

GSK-3β, FYN,and DYRK1A: Master Regulators in Neurodegenerative Pathways

Protein kinases (PKs) have been recognized as central nervous system (CNS)-disease-relevant targets due to their master regulatory role in different signal transduction cascades in the neuroscience space. Among them, GSK-3β, FYN, and DYRK1A play a crucial role in the neurodegeneration context, and the deregulation of all three PKs has been linked to different CNS disorders with unmet medical needs, including Alzheimer’s disease (AD), Parkinson’s disease (PD), frontotemporal lobar degeneration (FTLD), and several neuromuscular disorders. The multifactorial nature of these diseases, along with the failure of many advanced CNS clinical trials, and the lengthy approval process of a novel CNS drug have strongly limited the CNS drug discovery. However, in the near-decade from 2010 to 2020, several computer-assisted drug design strategies have been combined with synthetic efforts to develop potent and selective GSK-3β, FYN, and DYRK1A inhibitors as disease-modifying agents. In this review, we described both structural and functional aspects of GSK-3β, FYN, and DYRK1A and their involvement and crosstalk in different CNS pathological signaling pathways. Moreover, we outlined attractive medicinal chemistry approaches including multi-target drug design strategies applied to overcome some limitations of known PKs inhibitors and discover improved modulators with suitable blood–brain barrier (BBB) permeability and drug-like properties.     

The Impact of the ‘Mis-Peptidome’ on HLA Class I-Mediated Diseases: Contribution of ERAP1 and ERAP2 and Effects on the Immune Response

The strong association with the Major Histocompatibility Complex (MHC) class I genes represents a shared trait for a group of autoimmune/autoinflammatory disorders having in common immunopathogenetic basis as well as clinical features. Accordingly, the main risk factors for Ankylosing Spondylitis (AS), prototype of the Spondyloarthropathies (SpA), the Behçet’s disease (BD), the Psoriasis (Ps) and the Birdshot Chorioretinopathy (BSCR) are HLA-B*27, HLA-B*51, HLA-C*06:02 and HLA-A*29:02, respectively. Despite the strength of the association, the HLA pathogenetic role in these diseases is far from being thoroughly understood. Furthermore, Genome-Wide Association Studies (GWAS) have highlighted other important susceptibility factors such as Endoplasmic Reticulum Aminopeptidase (ERAP) 1 and, less frequently, ERAP2 that refine the peptidome presented by HLA class I molecules to CD8⁺ T cells. Mass spectrometry analysis provided considerable knowledge of HLA-B*27, HLA-B*51, HLA-C*06:02 and HLA-A*29:02 immunopeptidome. However, the combined effect of several ERAP1 and ERAP2 allelic variants could generate an altered pool of peptides accounting for the “mis-immunopeptidome” that ranges from suboptimal to pathogenetic/harmful peptides able to induce non-canonical or autoreactive CD8⁺ T responses, activation of NK cells and/or garbling the classical functions of the HLA class I molecules. This review will focus on this class of epitopes as possible elicitors of atypical/harmful immune responses which can contribute to the pathogenesis of chronic inflammatory diseases.     

Female Germ Cell Development,Functioning and Associated Adversities under Unfavorable Circumstances

In the present era, infertility is one of the major issues which restricts many couples to have their own children. Infertility is the inability to achieve a clinical pregnancy after regular unprotected sexual intercourse for the period of one year or more. Various factors including defective male or female germ cell development, unhealthy and improper lifestyles, diseases like cancer and associated chemo-or-radiation therapies, congenital disorders, etc., may be responsible for infertility. Therefore, it is highly important to understand the basic concepts of germ cell development including primordial germ cell (PGC) formation, specification, migration, entry to genital ridges and their molecular mechanisms, activated pathways, paracrine and autocrine signaling, along with possible alteration which can hamper germ cell development and can cause adversities like cancer progression and infertility. Knowing all these aspects in a proper way can be very much helpful in improving our understanding about gametogenesis and finding possible ways to cure related disorders. Here in this review, various aspects of gametogenesis especially female gametes and relevant factors causing functional impairment have been thoroughly discussed.     

From Channels to Canonical Wnt Signaling: A Pathological Perspective

Wnt signaling is an important pathway mainly active during embryonic development and controlling cell proliferation. This regulatory pathway is aberrantly activated in several human diseases. Ion channels are known modulators of several important cellular functions ranging from the tuning of the membrane potential to modulation of intracellular pathways, in particular the influence of ion channels in Wnt signaling regulation has been widely investigated. This review will discuss the known links between ion channels and canonical Wnt signaling, focusing on their possible roles in human metabolic diseases, neurological disorders, and cancer.