NF-κB-inducing Kinase Research Articles

NIK Is a Mediator of Inflammation and Intimal Hyperplasia in Endothelial Denudation-Induced Vascular Injury.

Neointimal hyperplasia is the main cause of vascular graft failure in the medium term. NFκB is a key mediator of inflammation that is activated during neointimal hyperplasia following endothelial injury. However, the molecular mechanisms involved in NFκB activation are poorly understood. NFκB may be activated through canonical (transient) and non-canonical (persistent) pathways. NFκB-inducing kinase (NIK, MAP3K14) is the upstream kinase of the non-canonical pathway. We have now explored the impact of NIK deficiency on neointimal hyperplasia following guidewire-induced endothelial cell injury and on local inflammation by comparing NIK activity-deficient alymphoplasia mice (NIKaly/aly) with control wild-type (NIK+/+) mice. Guidewire-induced endothelial cell injury caused neointimal hyperplasia and luminal stenosis and upregulated the local expression of NIK and the NFκB target chemokines monocyte chemoattractant protein-1 (MCP-1/CCL2) and chemokine ligand 5 (RANTES/CCL5). Immunohistochemistry disclosed the infiltration of the media and intima by F4/80 positive macrophages. The intima/media ratio and percentage of stenosis were milder in the NIKaly/aly than in the NIK+/+ mice. Additionally, the gene expression for MCP-1 and RANTES was lower and F4/80+ cell infiltration was milder in the NIKaly/aly than in the NIK+/+ mice. Finally, circulating MCP-1 levels were lower in the NIKaly/aly than in the NIK+/+ mice, reflecting milder systemic inflammation. In conclusion, NIK is a driver of vascular wall inflammation and stenosis following guidewire-induced endothelial cell injury. NIK targeting may be a novel therapeutic approach to limit arterial stenosis following endothelial cell injury.

EDA2R-NIK signaling in cancer cachexia.

Cachexia is a debilitating condition causing weight loss and skeletal muscle wasting that negatively influences treatment and survival of cancer patients. The objective of this review is to describe recent discoveries on the role of a novel signaling pathway involving ectodysplasin A2 receptor (EDA2R) and nuclear factor κB (NFκB)-inducing kinase (NIK) in muscle atrophy. Studies identified tumor-induced upregulation of EDA2R expression in muscle tissues in pre-clinical cachexia models and patients with various cancers. Activation of EDA2R by its ligand promoted atrophy in cultured myotubes and muscle tissue, which depended on NIK activity. The non-canonical NFκB pathway via NIK also stimulated muscle atrophy. Mice lacking EDA2R or NIK were protected from muscle loss due to tumors. Tumor-induced cytokine oncostatin M (OSM) upregulated EDA2R expression in muscles whereas OSM receptor-deficient mice were resistant to muscle wasting. Recent discoveries revealed a mechanism involving EDA2R-NIK signaling and OSM that drives cancer-associated muscle loss, opening up new directions for designing anti-cachexia treatments. The therapeutic potential of targeting this mechanism to prevent muscle loss should be further investigated. Future research should also explore broader implications of the EDA2R-NIK pathway in other muscle wasting diseases and overall muscle health.

Lymphotoxin beta-activated LTBR/NIK/RELB axis drives proliferation in cholangiocarcinoma.

Cholangiocarcinoma (CCA) is an aggressive malignancy arising from the intrahepatic (iCCA) or extrahepatic (eCCA) bile ducts with poor prognosis and limited treatment options. Prior evidence highlighted a significant contribution of the non-canonical NF-κB signalling pathway in initiation and aggressiveness of different tumour types. Lymphotoxin-β (LTβ) stimulates the NF-κB-inducing kinase (NIK), resulting in the activation of the transcription factor RelB. However, the functional contribution of the non-canonical NF-κB signalling pathway via the LTβ/NIK/RelB axis in CCA carcinogenesis and progression has not been established. Human CCA-derived cell lines and organoids were examined to determine the expression of NF-κB pathway components upon activation or inhibition. Proliferation and cell death were analysed using real-time impedance measurement and flow cytometry. Immunoblot, qRT-PCR, RNA sequencing and insitu hybridization were employed to analyse gene and protein expression. Four invivo models of iCCA were used to probe the activation and regulation of the non-canonical NF-κB pathway. Exposure to LTα1/β2 activates the LTβ/NIK/RelB axis and promotes proliferation in CCA. Inhibition of NIK with the small molecule inhibitor B022 efficiently suppresses RelB expression in patient-derived CCA organoids and nuclear co-translocation of RelB and p52 stimulated by LTα1/β2 in CCA cell lines. In murine CCA, RelB expression is significantly increased and LTβ is the predominant ligand of the non-canonical NF-κB signalling pathway. Our study confirms that the non-canonical NF-κB axis LTβ/NIK/RelB drives cholangiocarcinogenesis and represents a candidate therapeutic target.

PUM1-TRAF3 fusion protein activates non-canonical NF-κB signaling via rescued NIK in biliary tract cancer

Discovery and verification of diagnostic or therapeutic biomarkers for biliary tract cancer (BTC) is challenging owing to the low prevalence of the disease. Here, we identified and investigated the clinical impact of a fusion gene, Pumilio1-tumor necrosis factor receptor-associated factor 3 (PUM1-TRAF3), caused by 1;14 chromosomal translocation in BTC. PUM1-TRAF3 was initially identified in the RNA-sequencing of five BTC surgical tissues and confirmed by fluorescence in situ hybridization. Expression of the fusion gene was validated in an expanded cohort (5/55, 9.1%). Establishment and molecular assessment of PUM1-TRAF3 expressing BTC cells revealed that PUM1-TRAF3 activates non-canonical NF-κB signaling via NF-κB-inducing kinase (NIK). Abnormal TRAF3 activity, driven by competitive binding of PUM1-TRAF3 and TRAF3 to NIK, led to NIK rescue followed by P52/RelB nuclear translocation, all of which were reverted by an NIK inhibitor. The elevated expression of NIK and activated NF-κB signaling was observed in the PUM1-TRAF3-expressing regions of patient tissues. Expression of the PUM1-TRAF3 fusion was significantly correlated with strong NIK expression, which is associated with a poorer prognosis for patients with BTC. Overall, our study identifies a new fusion gene, PUM1-TRAF3, that activates NIK and non-canonical NF-κB signaling, which may be beneficial for developing precise treatment strategies for BTC.

Lipid Nanoparticle-mRNA Engineered Dendritic Cell Based Adoptive Cell Therapy Enhances Cancer Immune Response.

Lipid nanoparticles encapsulating mRNA (LNP-mRNA) revolutionized medicine over the past several years. While clinically approved indications currently focus on infectious disease vaccination, LNP-mRNA based treatments also hold promise for cancer immunotherapy. However, the route of dosing may impact treatment efficacy, safety, and dose. To minimize adverse effects, it is hypothesized that LNP-mRNA can be used to activate and engineer dendritic cells (DC) ex vivo before re-administration of these cells. Here, it is shown that LNP-mRNA engineered DCs can indeed vaccinate recipient mice. Vaccinated mice showed strong anti-tumor T cell responses, rejected tumor challenge, and displayed no evidence of toxicity. Further, it is found that DC specific ablation of the immune activating kinase NFkB inducing kinase (NIK) abrogated vaccination efficacy, demonstrating that adoptively transferred DCs can be functionally modified in addition to their antigen presentation capacity. Collectively, these studies show that ex vivo LNP-mRNA engineering of DCs is a feasible and robust therapeutic strategy for cancer.

Grass carp (Ctenopharyngodon idella) NIK up-regulates the expression of IL-8 by activating the NF-κB canonical pathway

NIK (NF-κB inducing kinase) belongs to the mitogen-activated protein kinase family, which activates NF-κB and plays a vital role in immunology, inflammation, apoptosis, and a series of pathological responses. In NF-κB noncanonical pathway, NIK and IKKα have been often studied in mammals and zebrafish. However, few have explored the relationship between NIK and other subunits of the IKK complex. As a classic kinase in the NF-κB canonical pathway, IKKβ has never been researched with NIK in fish. In this paper, the full-length cDNA sequence of grass carp (Ctenopharyngodon idella) NIK (CiNIK) was first cloned and identified. The expression level of CiNIK in grass carp cells was increased under GCRV stimuli. Under the stimulation of GCRV, poly (I:C), and LPS, the expression of NIK in various tissues of grass carp was also increased. This suggests that CiNIK responds to viral stimuli. To study the relationship between CiNIK and CiIKKβ, we co-transfected CiNIK-FLAG and CiIKKB-GFP into grass carp cells in coimmunoprecipitation and immunofluorescence experiments. The results revealed that CiNIK interacts with CiIKKβ. Besides, the degree of autophosphorylation of CiNIK was enhanced under poly (I:C) stimulation. CiIKKβ was phosphorylated by CiNIK and then activated the activity of p65. The activity change of p65 indicates that NF-κB downstream inflammatory genes will be functioning. CiNIK or CiIKKβ up-regulated the expression of IL-8. It got higher when CiNIK and CiIKKβ coexisted. This paper revealed that NF-κB canonical pathway and noncanonical pathway are not completely separated in generating benefits.

Paricalcitol Has a Potent Anti-Inflammatory Effect in Rat Endothelial Denudation-Induced Intimal Hyperplasia.

Neointimal hyperplasia is the main cause of vascular graft failure in the medium term. Vitamin D receptor activation modulates the biology of vascular smooth muscle cells and has been reported to protect from neointimal hyperplasia following endothelial injury. However, the molecular mechanisms are poorly understood. We have now explored the impact of the selective vitamin D receptor activator, paricalcitol, on neointimal hyperplasia, following guidewire-induced endothelial cell injury in rats, and we have assessed the impact of paricalcitol or vehicle on the expression of key cell stress factors. Guidewire-induced endothelial cell injury caused neointimal hyperplasia and luminal stenosis and upregulated the expression of the growth factor growth/differentiation factor-15 (GDF-15), the cytokine receptor CD74, NFκB-inducing kinase (NIK, an upstream regulator of the proinflammatory transcription factor NFκB) and the chemokine monocyte chemoattractant protein-1 (MCP-1/CCL2). Immunohistochemistry confirmed the increased expression of the cellular proteins CD74 and NIK. Paricalcitol (administered in doses of 750 ng/kg of body weight, every other day) had a non-significant impact on neointimal hyperplasia and luminal stenosis. However, it significantly decreased GDF-15, CD74, NIK and MCP-1/CCL2 mRNA expression, which in paricalcitol-injured arteries remained within the levels found in control vehicle sham arteries. In conclusion, paricalcitol had a dramatic effect, suppressing the stress response to guidewire-induced endothelial cell injury, despite a limited impact on neointimal hyperplasia and luminal stenosis. This observation identifies novel molecular targets of paricalcitol in the vascular system, whose differential expression cannot be justified as a consequence of improved tissue injury.

Broad spectrum antimicrobial, anti-inflammatory and peripheral analgesic activities of heteroaryl nitazoxanide analogs

Background The antiparasitic drug nitazoxanide possesses diverse biological activity. However, very few investigation was accomplished with nitazoxanide analogs. Therefore, herein we focused on the screening of bioactivities using some nitazoxanide-like synthesized molecules. Objectives Four heteroaryl nitazoxanide analogs synthesized in our laboratory were investigated for antimicrobial, anti-inflammatory, and analgesic activity. Materials and methods Disc diffusion method was used for assessing antimicrobial potency against several Gram-positive bacteria, Gram-negative bacteria, and fungi. Carrageenan-induced rat paw edema model was performed to evaluate anti-inflammatory activity. The analgesic property was evaluated using the acetic acid-induced writing inhibition method in the mice model. Molecular docking simulations against cyclooxygenase-1, cyclooxygenase-2, phospholipase A2, NF-κB inducing kinase, and interleukin-1 receptor-associated kinase 4 were also performed. Results and conclusion All the synthesized compounds exhibited broad spectrum antimicrobial property against a number of Gram-positive, Gram-negative species and unicellular fungi. Compound 4 or N-(5-nitrothiazol-2-yl)-furan-3-carboxamide emerged as the most prominent antimicrobial agent exhibiting zone of inhibition ranging in 14–22 mm. These zone diameters are sometimes greater than that displayed by nitazoxanide. Compounds 2 and 3 also showed remarkable broad-spectrum antimicrobial activity with a zone of inhibition 10–20 mm and 12–20 mm, respectively. Compound 4 also displayed potential anti-inflammatory activity which is comparable to standard aceclofenac. Compound 4 also showed mild analgesic effects. The compounds also exhibited moderate binding affinities against the selected target receptors and enzymes during in silico molecular docking. Heteroaryl nitazoxanide analogs showed prominent broad-spectrum antimicrobial, anti-inflammatory, and mild analgesic properties. This study indicates that heteroaryl nitazoxanide analogs might be interesting candidates for new drug discovery.

Abstract B049: Uncovering sex-biased immunometabolic roles for NIK in the GBM tumor microenvironment

Abstract The interconnected relationship between the central nervous system (CNS) and the immune system is crucial for brain function. Astrocytes, microglia and infiltrating macrophages perform diverse functions for the maintenance of CNS homeostasis, including inflammatory responses to injury or infection, tissue repair and immune surveillance of cancers such as glioblastoma (GBM). Indeed, these cells comprise up to 50% of GBM tumor mass, yet the impact of their interactions in regulating neuroinflammation and immunosuppression in the GBM tumor microenvironment is not well understood. We have recently found that NF-κB-Inducing Kinase (NIK), an essential activator of the noncanonical NF-κB pathway, governs the mitochondrial metabolic reprogramming that is required for the acquisition of pro-repair bone-marrow-derived macrophage (BMDM) immune effector functions. Although NIK is highly expressed in brain immune cells, including resident macrophages/microglia and astrocytes, little is known regarding NIK-dependent regulatory pathways controlling their immune cell effector functions in brain health and systemic responses to disease. Our preliminary data demonstrate that similar to BMDMs, NIK knockout (KO) microglia and astrocytes have impaired mitochondrial oxidative metabolism and skew towards pro-inflammatory phenotypes. Moreover, NIK KO mice harboring orthotopic syngeneic brain tumors exhibit improved survival and reduced microglia and macrophage tumor infiltration, suggesting that NIK functions in the TME to support immune cell interactions that promote GBM growth. Notably these effects were sex-biased, with male NIK KO mice having a more pronounced survival benefit. Additionally, we observe whole-body metabolic dysfunction in myeloid- and astrocyte-specific conditional NIK knockout mice, implying a potential NIK-dependent feedback relationship between CNS immune/glial cell functions and control of systemic metabolism. Overall, these results are consistent with poorer survival of patients with elevated NIK expression and provides a possible rationale for the higher incidence of GBM in males. Coupled with our previous work demonstrating that NIK has several GBM cell-intrinsic roles to promote tumor invasion and adaptive growth through control of mitochondrial fission and cellular metabolism, these new findings highlight the importance of understanding NIK-dependent immunometabolic functions in the brain. Therapeutic strategies targeting NIK may not only attenuate GBM cell invasion and render them vulnerable to metabolic stress, but also modulate metabolic rewiring of immune cells in the tumor microenvironment to improve patient outcomes. Citation Format: Justin N. Keeney, Caren Stuebe, Kathryn M. Pflug, L. Gerard Toussaint, Raquel Sitcheran. Uncovering sex-biased immunometabolic roles for NIK in the GBM tumor microenvironment [abstract]. In: Proceedings of the AACR Special Conference on Brain Cancer; 2023 Oct 19-22; Minneapolis, Minnesota. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_1):Abstract nr B049.

NF-κB Inducing Kinase Attenuates Colorectal Cancer by Regulating Noncanonical NF-κB Mediated Colonic Epithelial Cell Regeneration

Dysregulated colonic epithelial cell (CEC) proliferation is a critical feature in the development of colorectal cancer. We show that NF-κB-inducing kinase (NIK) attenuates colorectal cancer through coordinating CEC regeneration/differentiation via noncanonical NF-κB signaling that is unique from canonical NF-kB signaling. Initial studies evaluated crypt morphology/functionality, organoid generation, transcriptome profiles, and the microbiome. Inflammation and inflammation-induced tumorigenesis were initiated in whole-body NIK knockout mice (Nik-/-) and conditional-knockout mice following administration of azoxymethane and dextran sulfate sodium. Human transcriptomic data revealed dysregulated noncanonical NF-kB signaling. Invitro studies evaluating Nik-/- crypts and organoids derived from mature, nondividing CECs, and colonic stem cells exhibited increased accumulation and stunted growth, respectively. Transcriptomic analysis of Nik-/- cells revealed gene expression signatures associated with altered differentiation-regeneration. When assessed invivo, Nik-/- mice exhibited more severe colitis with dextran sulfate sodium administration and an altered microbiome characterized by increased colitogenic microbiota. In the inflammation-induced tumorigenesis model, we observed both increased tumor burdens and inflammation in mice where NIK is knocked out in CECs (NikΔCEC). Interestingly, this was not recapitulated when NIK was conditionally knocked out in myeloid cells (NikΔMYE). Surprisingly, conditional knockout of the canonical pathway in myeloid cells (RelAΔMYE) revealed decreased tumor burden and inflammation and no significant changes when conditionally knocked out in CECs (RelAΔCEC). Dysregulated noncanonical NF-κB signaling is associated with the development of colorectal cancer in a tissue-dependent manner and defines a critical role for NIK in regulating gastrointestinal inflammation and regeneration associated with colorectal cancer.

NF-κB signaling activation and roles in thyroid cancers: implication of MAP3K14/NIK

Among follicular-derived thyroid cancers (TC), those with aggressive behavior and resistance to current treatments display poor prognosis. NF-κB signaling pathways are involved in tumor progression of various cancers. Here, we finely characterize the NF-κB pathways and their involvement in TC. By using immunoblot and gel shift assays, we demonstrated that both classical and alternative NF-κB pathways are activated in ten TC-derived cell lines, leading to activated RelA/p50 and RelB/p50 NF-κB dimers. By analyzing the RNAseq data of the large papillary thyroid carcinoma (PTC) cohort from The Cancer Genome Atlas (TCGA) project, we identified a tumor progression-related NF-κB signature in BRAFV600E mutated-PTCs. That corroborated with the role of RelA and RelB in cell migration and invasion processes that we demonstrated specifically in BRAFV600E mutated-cell lines, together with their role in the control of expression of genes implicated in invasiveness (MMP1, PLAU, LCN2 and LGALS3). We also identified NF-κB-inducing kinase (NIK) as a novel actor of the constitutive activation of the NF-κB pathways in TC-derived cell lines. Finally, its implication in invasiveness and its overexpression in PTC samples make NIK a potential therapeutic target for advanced TC treatment.

Targeting NF-κB signaling in B cells as a potential new treatment modality for ANCA-associated vasculitis

B lineage cells are critically involved in ANCA-associated vasculitis (AAV), evidenced by alterations in circulating B cell subsets and beneficial clinical effects of rituximab (anti-CD20) therapy. This treatment renders a long-term, peripheral B cell depletion, but allows for the survival of long-lived plasma cells. Therefore, there is an unmet need for more reversible and full B lineage cell targeting approaches. To find potential novel therapeutic targets, RNA sequencing of CD27+ memory B cells of patients with active AAV was performed, revealing an upregulated NF-κB-associated gene signature. NF-κB signaling pathways act downstream of various B cell surface receptors, including the BCR, CD40, BAFFR and TLRs, and are essential for B cell responses. Here we demonstrate that novel pharmacological inhibitors of NF-κB inducing kinase (NIK, non-canonical NF-κB signaling) and inhibitor-of-κB-kinase-β (IKKβ, canonical NF-κB signaling) can effectively inhibit NF-κB signaling in B cells, whereas T cell responses were largely unaffected. Moreover, both inhibitors significantly reduced B cell proliferation, differentiation and production of antibodies, including proteinase-3 (PR3) autoantibodies, in B lineage cells of AAV patients. These findings indicate that targeting NF-κB, particularly NIK, may be an effective, novel B lineage cell targeted therapy for AAV and other autoimmune diseases with prominent B cell involvement.

Development of Allosteric NIK Ligands from Fragment-Based NMR Screening.

NF-κB inducing kinase (NIK) is vital for the induction of many immune responses, and as such, NIK dysregulation has been implicated in various inflammatory diseases and cancers. NIK has been pursued as a potential therapeutic target, and small-molecule inhibitors that bind the orthosteric site on NIK have been reported. However, despite the established chemical matter, NIK inhibitors have not yet reached the clinic. With the goal of developing allosteric NIK ligands using a fragment-based NMR screening approach, we report the identification and development of a series of allosteric, fragment-sized NIK ligands that bind with micromolar potency and good ligand efficiency.

Structure-based optimization of indoline-containing compounds as second generation inhibitors of NF-κB-inducing kinase (NIK)

Overactivation of NIK led to increased transcriptional activity of NF-κB2, which was associated with human metabolic diseases, immunological disorders, and cancer. Therefore, NIK inhibitors were pursued as a potential treatment of NIK-driven human diseases. The first generation of NIK inhibitors features propargylic alcohol moieties, which were metabolic soft spots that led to the discovery of a second generation of NIK inhibitors without a propargylic alcohol. In this study, a series of compounds were designed, synthesized, and evaluated as novel NIK inhibitors using a NIK kinase assay. To evaluate selectivity profiles of these inhibitors, the most potent compound 50 was tested over COT and other two panels of 31 kinases. In addition, 50 inhibited NIK-driven upregulation of CCL5 gene in primary hepatocytes and showed low cytotoxicity against normal kidney cells HEK-293. These attributes make 50 a promising candidate for future optimization aimed to increase potency and selectivity against NIK and NF-κB2 pathway.

Transcriptional induction of NF-κB-inducing kinase by E2F4/5 facilitates collective invasion of GBM cells

The prognosis of high-grade gliomas, such as glioblastoma multiforme (GBM), is extremely poor due to the highly invasive nature of these aggressive cancers. Previous work has demonstrated that TNF-weak like factor (TWEAK) induction of the noncanonical NF-κB pathway promotes the invasiveness of GBM cells in an NF-κB-inducing kinase (NIK)-dependent manner. While NIK activity is predominantly regulated at the posttranslational level, we show here that NIK (MAP3K14) is upregulated at the transcriptional level in invading cell populations, with the highest NIK expression observed in the most invasive cells. GBM cells with high induction of NIK gene expression demonstrate characteristics of collective invasion, facilitating invasion of neighboring cells. Furthermore, we demonstrate that the E2F transcription factors E2F4 and E2F5 directly regulate NIK transcription and are required to promote GBM cell invasion in response to TWEAK. Overall, our findings demonstrate that transcriptional induction of NIK facilitates collective cell migration and invasion, thereby promoting GBM pathogenesis.

NIK-mediated reactivation of SIX2 enhanced the CSC-like traits of hepatocellular carcinoma cells through suppressing ubiquitin-proteasome system.

The critical roles of NF-κB Inducing Kinase (NIK) in tumor progression have been elucidated in various tumors; however, its effects on hepatocellular carcinoma (HCC) progression are still confusing. Here, we found that NIK level was upregulated in HCC tissues compared to that of normal tissues, and positively correlated with the levels of cancer stem cell (CSC) markers. Then we established HCC cells with NIK-stable knockdown and found that NIK knockdown suppressed the CSC-like traits of HCC cells through in vivo and in vitro experiments. Mechanistically, we revealed that SIX2 protein level, but not its mRNA level, was significantly reduced in HCC cells with NIK knockdown, which was rescued by MG132 treatment. Furthermore, NIK knockdown promoted the ubiquitination level of SIX2 and decreased its protein stability. Moreover, Six2 overexpression partially reversed the inhibition of NIK knockdown on the CSC-like traits of HCC cells. This study identified a novel NIK/SIX2 axis conferring HCC stemness.

High expression of NF-κB inducing kinase in the bulge region of hair follicle induces tumor

The bulge region, a reservoir of multipotent stem cells, is possibly responsible for tumorigenesis. NF-κB-inducing kinase (NIK) is a kinase involved in the activation of the noncanonical NF-κB pathway and exhibits positive staining in tumor cells. However, whether high expression of NIK can result in tumorigenesis has not been reported in published papers. By establishing Nik-coe (Nik-stopF/F crossed with Chat-cre) and Nik-soe (Nik-stopF/F crossed with Sox9-cre) mice, we found that overexpression of Nik in the bulge region of hair follicles induced hair follicle loss and tumorigenesis. Furthermore, RNA sequencing, proteomic and phosphopeptide analyses revealed that multiple cancer pathways are involved in tumor formation. Taken together, these findings indicate that constitutive activation of Nik in the bulge region induces tumorigenesis.

Empiric anti-Giardia therapy in non-diarrheal protein-losing enteropathy: A focus on children with monogenic humoral immunodeficiency.

Background and Aim: Chronic giardia infection can lead to non-erosive gastrointestinal disorders, including protein-losing enteropathy (PLE). This report describes non-diarrheal PLE in chronic giardiasis in children with defective humoral immunity. Methods: The retrospective report is related to 2 children known to have a monogenic inborn error of immunity. The first patient is a 13-year-old boy with X-linked agammaglobulinemia (Patient-1), and the second is a 5-year-old boy with NF-kB inducing kinase (NIK) deficiency infection. Frequency, growth status, and serum immunoglobulin-G (IgG) trough and albumin levels were monitored (Patient-2). Results: Patient-1 had more frequency of pneumonia but reported no symptoms of gastrointestinal disease, including alteration of bowel habits, change in stool consistency, nausea, vomiting, fatigue, bloating, and abdominal pain. No clinical oedema on examination. His weight remains static at 19-20 kg for about 1.5 years. Simultaneously, hypoproteinaemia was noted (Figure-1). A trial of increasing IVIG (0.7 - 1 g/kg) and the use of subcutaneous immunoglobulin (0.2 g/kg) did not reverse the biochemical and clinical situation. Hypoproteinaemia workup revealed normal liver and kidney functions, normal cardiac function, and no proteinuria. Interestingly, his upper endoscopy showed mild duodenitis and the presence of giardia lamblia at the luminal surface (Figure-2). Following this, a 2-week course of oral metronidazole (7.5 mg/kg/dose BID) resulted in a restoration of therapeutic serum IgG trough and albumin levels. Additionally, the child's nutritional status improved, and the frequency of respiratory infections dropped. In Patient-2, progressive hypoproteinaemia was noted over nine months. Similarly, no gastrointestinal complaints; however, the family reported foul-smelling semisolid stools with regular consistency. His IgG trough level was 2 g/l, and his albumin level was 20 g/l. Despite maximizing IVIG, he developed two episodes of pneumonia and once otitis media. An empiric oral metronidazole course resulted in the amelioration of symptoms and restoration of proteinemia. Meanwhile, faeces microscopy confirmed the presence of Giardia lamblia. Conclusions: Progressive hypoproteinaemia in children with humoral immunodeficiency is a clinical concern. Routine stool microscopy can identify giardia infection despite the ambiguity of gastrointestinal symptoms in such patients. However, a trial of empiric metronidazole therapy should be considered.

NF-κB-Inducing Kinase Provokes Insulin Resistance in Skeletal Muscle of Obese Mice.

Skeletal muscle is crucial for preserving glucose homeostasis. Insulin resistance and abnormalities in glucose metabolism result from a range of pathogenic factors attacking skeletal muscle in obese individuals. To relieve insulin resistance and restore glucose homeostasis, blocking the cell signaling pathways induced by those pathogenic factors seems an attractive strategy. It has been discovered that insulin sensitivity in obese people is inversely linked with the activity of NF-κB inducing kinase (NIK) in skeletal muscle. In order to evaluate NIK's pathological consequences, mechanism of action, and therapeutic values, an obese mouse model reproduced by feeding a high-fat diet was treated with a NIK inhibitor, B022. C2C12 myoblasts overexpressing NIK were utilized to assess insulin signaling and glucose uptake. B022 thus prevented high-fat diet-induced NIK activation and insulin desensitization in skeletal muscle. The insulin signaling in C2C12 myoblasts was compromised by the upregulation of NIK brought on by oxidative stress, lipid deposition, inflammation, or adenoviral vector. This inhibition of insulin action is mostly due to an inhibitory serine phosphorylation of IRS1 caused by ERK, JNK, and PKC that were activated by NIK. In summary, NIK integrates signals from several pathogenic factors to impair insulin signaling by igniting a number of IRS1-inhibiting kinases, and it also has significant therapeutic potential for treating insulin resistance.

Epidermal Growth Factor Receptor Mediates Neuronal Apoptosis After Subarachnoid Hemorrhage in Mice.

Early brain injury including neuronal apoptosis is a main contributor to neurological deterioration after subarachnoid hemorrhage (SAH). This study was aimed to investigate whether EGFR (epidermal growth factor receptor)/NFκB (nuclear factor-kappa B) inducing kinase (NIK)/NFκB (p65 and p50) pathway is involved in the neuronal apoptosis after SAH in mice. C57BL/6 adult male mice underwent endovascular perforation SAH modeling or sham-operation (n=286), and 86 mild SAH mice were excluded. In experiment 1, vehicle or an EGFR inhibitor (632.0 ng AG1478) was administered intraventricularly at 30 minutes postmodeling. At 24 or 72 hours, after neurological score was tested, brain water content, double immunolabeling with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and a neuronal marker antimicrotubule-associated protein-2 antibody, Western blotting using whole tissue lysate or nuclear protein extraction of the left cortex, and immunohistochemistry for cleaved caspase-3, phosphorylated (p-) EGFR, NIK, p-NFκB p65, and NFκB p105/50 were evaluated. In experiment 2, after sham or SAH modeling, AG1478+vehicle or AG1478+4.0 ng EGF was administered intraventricularly. The brain was used for TUNEL staining and immunohistochemistry after 24-hour observation. SAH group showed deteriorated neurological score (P<0.01, Mann-Whitney U test), more TUNEL- and cleaved caspase-3-positive neurons (P<0.01, ANOVA), and higher brain water content (P<0.01, Mann-Whitney U test), and these observations were improved in SAH-AG1478 group. Western blotting showed that expression levels of p-EGFR, p-p65, p50, and nuclear-NIK were increased after SAH (P<0.05, ANOVA), and decreased by AG1478 administration. Immunohistochemistry revealed these molecules localized in degenerating neurons. EGF administration resulted in neurological deterioration, increased TUNEL-positive neurons, and activation of EGFR, NIK, and NFκB. Activated EGFR, nuclear-NIK, and NFκB expressions were observed in cortical degenerating neurons after SAH, and were decreased by administration of AG1478, associated with suppression of TUNEL- and cleaved caspase-3-positive neurons. EGFR/NIK/NFκB pathway is suggested to be involved in neuronal apoptosis after SAH in mice.