Immune Monitoring of Trans-endothelial Transport by Kidney-Resident Macrophages.

Small immune complexes cause type III hypersensitivity reactions that frequently result in tissue injury. The responsible mechanisms, however, remain unclear and differ depending on target organs. Here, we identify a kidney-specific anatomical and functional unit, formed by resident macrophages and peritubular capillary endothelial cells, which monitors the transport of proteins and particles ranging from 20 to 700 kDa or 10 to 200 nm into the kidney interstitium. Kidney-resident macrophages detect and scavenge circulating immune complexes "pumped" into the interstitium via trans-endothelial transport and trigger a FcγRIV-dependent inflammatory response and the recruitment of monocytes and neutrophils. In addition, FcγRIV and TLR pathways synergistically "super-activate" kidney macrophages when immune complexes contain a nucleic acid. These data identify a physiological function of tissue-resident kidney macrophages and a basic mechanism by which they initiate the inflammatory response to small immune complexes in the kidney.

An explanation of the pathogenesis of several autoimmune diseases in immuno-compromised individuals.

Some pathogen infections and immune system deficiencies have been linked to a few autoimmune diseases. However, the pathogenesis of most autoimmune diseases is unknown. An explanatory hypothesis for the pathogenesis of infection-initiated autoimmune diseases is provided. Virulent pathogen infections create extensive pathogen antigens that frequently require antibodies. These antibodies create extensive antigen-antibody immune complexes, which some immuno-compromised individuals will not adequately eliminate. This will cause inflammatory type III hypersensitivity symptoms, including protease releases that destroy epithelium, mesothelium and endothelium basement membranes, express new immunogenic antigens from previously sequestered basement membrane constituents, and ultimately induce new autoantibodies. This can continue after the infection ends, if the first wave of protease attacks on basement membranes induces new autoantibodies that cause new uncleared antigen-antibody immune complexes and type III hypersensitivity reactions. The secreted proteases and other enzymes will have preferred substrates and these proteases or other enzymes by themselves, or by their processed protein substrates, can express immunogenic antigens that induce new autoantibodies and initiate various autoimmune diseases. In summary, several autoimmune diseases can be initiated in immuno-compromised individuals during extensive pathogen infections, if these individuals have two immune problems: (a) slow or weak initial immune responses that result in a reliance on antibodies and (b) an inability to eliminate the resulting antigen-antibody immune complexes by phagocytosis. These two immune problems and the resulting immune system type III hypersensitivity reaction can explain the causation of several autoimmune diseases, including the most common and the rarest autoimmune diseases, both their differences and their similarities.

Type 3 hypersensitivity in COVID-19 vasculitis.

Coronavirus Disease 2019 (COVID-19) is an ongoing public health emergency and new knowledge about its immunopathogenic mechanisms is deemed necessary in the attempt to reduce the death burden, globally. For the first time in worldwide literature, we provide scientific evidence that in COVID-19 vasculitis a life-threatening escalation from type 2 T-helper immune response (humoral immunity) to type 3 hypersensitivity (immune complex disease) takes place. The subsequent deposition of immune complexes inside the vascular walls is supposed to induce a severe inflammatory state and a cytokine release syndrome, whose interleukin-6 is the key myokine, from the smooth muscle cells of blood vessels.

Complement C3-Targeted Therapy: Replacing Long-Held Assertions with Evidence-Based Discovery.

Complement dysregulation underlies several inflammatory disorders, and terminal complement inhibition has thus far afforded significant clinical gains. Nonetheless, emerging pathologies, fueled by complement imbalance and therapy-skewing genetic variance, underscore the need for more comprehensive, disease-tailored interventions. Modulation at the level of C3, a multifaceted orchestrator of the complement cascade, opens up prospects for broader therapeutic efficacy by targeting multiple pathogenic pathways modulated by C3-triggered proinflammatory crosstalk. Notably, C3 intervention is emerging as a viable therapeutic strategy for renal disorders with predominantly complement-driven etiology, such as C3 glomerulopathy (C3G). Using C3G as a paradigm, we argue that concerns about the feasibility of long-term C3 intervention need to be placed into perspective and weighed against actual therapeutic outcomes in prospective clinical trials.

New milestones ahead in complement-targeted therapy.

The complement system is a powerful effector arm of innate immunity that typically confers protection from microbial intruders and accumulating debris. In many clinical situations, however, the defensive functions of complement can turn against host cells and induce or exacerbate immune, inflammatory, and degenerative conditions. Although the value of inhibiting complement in a therapeutic context has long been recognized, bringing complement-targeted drugs into clinical use has proved challenging. This important milestone was finally reached a decade ago, yet the clinical availability of complement inhibitors has remained limited. Still, the positive long-term experience with complement drugs and their proven effectiveness in various diseases has reinvigorated interest and confidence in this approach. Indeed, a broad variety of clinical candidates that act at almost any level of the complement activation cascade are currently in clinical development, with several of them being evaluated in phase 2 and phase 3 trials. With antibody-related drugs dominating the panel of clinical candidates, the emergence of novel small-molecule, peptide, protein, and oligonucleotide-based inhibitors offers new options for drug targeting and administration. Whereas all the currently approved and many of the proposed indications for complement-targeted inhibitors belong to the rare disease spectrum, these drugs are increasingly being evaluated for more prevalent conditions. Fortunately, the growing experience from preclinical and clinical use of therapeutic complement inhibitors has enabled a more evidence-based assessment of suitable targets and rewarding indications as well as related technical and safety considerations. This review highlights recent concepts and developments in complement-targeted drug discovery, provides an overview of current and emerging treatment options, and discusses the new milestones ahead on the way to the next generation of clinically available complement therapeutics.

A viral infection explanation for Kawasaki disease in general and for COVID-19 virus-related Kawasaki disease symptoms.

SARS-CoV-2, a new virus that appeared in Wuhan, China, in 2019 has approximately an 80% genomic match to the Severe Acute Respiratory Symptom (SARS) virus, which is known to come from a bat virus. Symptoms of Kawasaki disease in general and incomplete Kawasaki disease have been seen in a subset of pediatric patients having a current or previous infection of SARS-CoV-2. A viral infection, such as a SARS-CoV-2 virus infection, could result in extensive antigen-antibody immune complexes that cannot be quickly cleared in a subset of patients and thus create a type III hypersensitivity immune reaction and cause Kawasaki disease or Kawasaki disease symptoms (also known as multisystem inflammatory syndrome) in a subset of patients. Extensive binding of antibodies to viral antigens can create antigen-antibody immune complexes, which, if not eliminated in certain individuals having dysfunctional complement systems, can start inflammatory type III hypersensitivity symptoms, including protease releases that can disrupt epithelium, mesothelium, and endothelium basement membranes, and induce pervasive inflammation throughout the body. This could continue after SARS-CoV-2 infections end if the first wave of protease attacks on basement membranes created new secondary autoantibodies and new uncleared antigen-antibody immune complexes.

The many faces of Mac-1 in autoimmune disease.

Mac-1 (CD11b/CD18) is a ß2 integrin classically regarded as a pro-inflammatory molecule because of its ability to promote phagocyte cytotoxic functions and enhance the function of several effector molecules such as FcγR, uPAR, and CD14. Nevertheless, recent reports have revealed that Mac-1 also plays significant immunoregulatory roles, and genetic variants in ITGAM, the gene that encodes CD11b, confer risk for the autoimmune disease systemic lupus erythematosus (SLE). This has renewed interest in the physiological roles of this integrin and raised new questions on how its seemingly opposing biological functions may be regulated. Here, we provide an overview of the CD18 integrins and how their activation may be regulated as this may shed light on how the opposing roles of Mac-1 may be elicited. We then discuss studies that exemplify Mac-1's pro-inflammatory versus regulatory roles particularly in the context of IgG immune complex-mediated inflammation. This includes a detailed examination of molecular mechanisms that could explain the risk-conferring effect of rs1143679, a single nucleotide non-synonymous Mac-1 polymorphism associated with SLE.

Ubiquitin C-Terminal Hydrolase L1 is required for regulated protein degradation through the ubiquitin proteasome system in kidney.

Ubiquitin C-terminal hydrolase L1 (UCH-L1) is a major deubiquitinating enzyme of the nervous system and associated with the development of neurodegenerative diseases. We have previously shown that UCH-L1 is found in tubular and parietal cells of the kidney and is expressed de novo in injured podocytes. Since the role of UCH-L1 in the kidney is unknown we generated mice with a constitutive UCH-L1-deficiency to determine its role in renal health and disease. UCH-L1-deficient mice developed proteinuria, without gross changes in glomerular morphology. Tubular cells, endothelial cells, and podocytes showed signs of stress with an accumulation of oxidative-modified and polyubiquitinated proteins. Mechanistically, abnormal protein accumulation resulted from an altered proteasome abundance leading to decreased proteasomal activity, a finding exaggerated after induction of anti-podocyte nephritis. UCH-L1-deficient mice exhibited an exacerbated course of disease with increased tubulointerstitial and glomerular damage, acute renal failure, and death, the latter most likely a result of general neurologic impairment. Thus, UCH-L1 is required for regulated protein degradation in the kidney by controlling proteasome abundance. Altered proteasome abundance renders renal cells, particularly podocytes and endothelial cells, susceptible to injury.

The atypical chemokine receptor 2 limits renal inflammation and fibrosis in murine progressive immune complex glomerulonephritis.

The atypical chemokine receptor 2 (ACKR2), also named D6, regulates local levels of inflammatory chemokines by internalization and degradation. To explore potential anti-inflammatory functions of ACKR2 in glomerulonephritis, we induced autologous nephrotoxic nephritis in C57/BL6 wild-type and Ackr2-deficient mice. Renal ACKR2 expression increased and localized to interstitial lymphatic endothelium during nephritis. At two weeks Ackr2-/-mice developed increased albuminuria and urea levels compared to wild-type mice. Histological analysis revealed increased structural damage in the glomerular and tubulointerstitial compartments within Ackr2-/- kidneys. This correlated with excessive renal leukocyte infiltration of CD4+ T cells and mononuclear phagocytes with increased numbers in the tubulointerstitium but not glomeruli in knockout mice. Expression of inflammatory mediators and especially markers of fibrotic tissue remodeling were increased along with higher levels of ACKR2 inflammatory chemokine ligands like CCL2 in nephritic Ackr2-/- kidneys. In vitro, Ackr2 deficiency in TNF-stimulated tubulointerstitial tissue but not glomeruli increased chemokine levels. These results are in line with ACKR2 expression in interstitial lymphatic endothelial cells, which also assures efflux of activated leukocytes into regional lymph nodes. Consistently, nephritic Ackr2-/- mice showed reduced adaptive cellular immune responses indicated by decreased regional T-cell activation. However, this did not prevent aggravated injury in the kidneys of Ackr2-/- mice with nephrotoxic nephritis due to simultaneously increased tubulointerstitial chemokine levels, leukocyte infiltration and fibrosis. Thus, ACKR2 is important in limiting renal inflammation and fibrotic remodeling in progressive nephrotoxic nephritis. Hence, ACKR2 may be a potential target for therapeutic interventions in immune complex glomerulonephritis.

Immunogenicity and Immune Complex Disease in Preclinical Safety Studies.

This article summarizes a continuing education presentation on immunogenicity that was part of a continuing education course entitled, "Clinical Pathology of Biotherapeutics." Immunogenicity of a biotherapeutic can have diverse impacts including altered systemic exposure and pharmacologic responses and, in a fraction of the cases, safety concerns including cross-reactive neutralization of endogenous proteins or sequela related to immune complex disease (ICD). In most cases, immune complexes are readily cleared from circulation; however, based on physiochemical properties, insoluble complexes form, activate complement, and deposit in tissues. Using published information and personal experience, a set of repeat-dose monkey toxicity studies with manifestations suggestive of ICD was reviewed to summarize the spectrum of clinical and pathology findings. The most common live-phase observation linked to ICD was an acute postdosing reaction following multiple dose administrations characterized by generalized collapse and attributed to acute complement activation. Less common live-phase observations were related to syndromes such as a consumptive coagulopathy or a protein losing nephropathy. The most common histologic change attributed to ICD was multi-organ vascular/perivascular inflammation followed by glomerulonephritis. The presentation concluded with a description of the challenges in assessing the relevance of immunogenicity-related reaction in monkey to human clinical use.

Ten Weeks of Infection with a Tissue-Invasive Helminth Protects against Local Immune Complex-Mediated Inflammation, but Not Cutaneous Type I Hypersensitivity, in Previously Sensitized Mice.

In this study, we evaluated the effect chronic helminth infection has on allergic disease in mice previously sensitized to OVA. Ten weeks of infection with Litomosoides sigmodontis reduced immunological markers of type I hypersensitivity, including OVA-specific IgE, basophil activation, and mast cell degranulation. Despite these reductions, there was no protection against immediate clinical hypersensitivity following intradermal OVA challenge. However, late-phase ear swelling, due to type III hypersensitivity, was significantly reduced in chronically infected animals. Levels of total IgG2a, OVA-specific IgG2a, and OVA-specific IgG1 were reduced in the setting of infection. These reductions were most likely due to increased Ab catabolism as ELISPOT assays demonstrated that infected animals do not have suppressed Ab production. Ear histology 24 h after challenge showed infected animals have reduced cellular infiltration in the ear, with significant decreases in numbers of neutrophils and macrophages. Consistent with this, infected animals had less neutrophil-specific chemokines CXCL-1 and CXCL-2 in the ear following challenge. Additionally, in vitro stimulation with immune complexes resulted in significantly less CXCL-1 and CXCL-2 production by eosinophils from chronically infected mice. Expression of FcγRI was also significantly reduced on eosinophils from infected animals. These data indicate that chronic filarial infection suppresses eosinophilic responses to Ab-mediated activation and has the potential to be used as a therapeutic for pre-existing hypersensitivity diseases.

Immune Complex Membranoproliferative Glomerulonephritis Associated with Transjugular Intrahepatic Portosystemic Shunts in Alcoholic Cirrhosis: Two Case Reports.

OBJECTIVE: To report on 2 patients with alcoholic cirrhosis who were treated with transjugular intrahepatic portosystemic shunt (TIPS) placement. CLINICAL PRESENTATION AND INTERVENTION: The 2 patients had a history of alcoholic cirrhosis, and TIPS surgery was performed on them. In both cases, 4 months after TIPS placement, proteinuria was observed along with histological alterations characteristic of immune complex membranoproliferative glomerulonephritis (MPGN). CONCLUSION: The TIPS in one patient was successful without immediate complications, while the other patient was referred for a combined liver-kidney transplant. In both cases, immune complex MPGN might have developed after TIPS placement probably due to a reduced immune complex clearance.

ß7 Integrin controls mast cell recruitment, whereas αE integrin modulates the number and function of CD8+ T cells in immune complex-mediated tissue injury.

Immune complex (IC) deposition causes significant tissue injury associated with various autoimmune diseases such as vasculitis. In the cascade of inflammation, cell-to-cell and cell-to-matrix adhesion via adhesion molecules are essential. To assess the role of αE and ß7 integrin in IC-mediated tissue injury, peritoneal and cutaneous reverse-passive Arthus reaction was examined in mice lacking αE integrin (αE(-/-)) or ß7 integrin (ß7(-/-)). Both αE(-/-) and ß7(-/-) mice exhibited significantly attenuated neutrophil infiltration in the peritoneal and cutaneous Arthus reaction. ß7 integrin deficiency, not αE integrin deficiency, significantly reduced the number of mast cells in the peritoneal cavity, which was consistent with the result that mast cells expressed only α4ß7 integrin, not αEß7 integrin. αE(-/-) mice instead revealed the reduction of CD8(+) T cells in the peritoneal cavity, and nearly half of them in wild-type mice expressed αE integrin. These αE(+)CD8(+) T cells produced more proinflammatory cytokines than αE(-)CD8(+) T cells, and adoptive transfer of αE(+)CD8(+) T cell into αE(-/-) recipients restored cutaneous and peritoneal Arthus reaction. These results suggest that in the peritoneal and cutaneous reverse-passive Arthus reaction, α4ß7 integrin is involved in the migration of mast cells for initial IC recognition. αEß7 integrin, in contrast, contributes by recruiting αE(+)CD8(+) T cells, which produce more proinflammatory cytokines than αE(-)CD8(+) T cells and amplify IC-mediated inflammation.

B cell-specific loss of Lyn kinase leads to autoimmunity.

The Lyn tyrosine kinase regulates inhibitory signaling in B and myeloid cells: loss of Lyn results in a lupus-like autoimmune disease with hyperactive B cells and myeloproliferation. We have characterized the relative contribution of Lyn-regulated signaling pathways in B cells specifically to the development of autoimmunity by crossing the novel lyn(flox/flox) animals with mice carrying the Cre recombinase under the control of the Cd79a promoter, resulting in deletion of Lyn in B cells. The specific deletion of Lyn in B cells is sufficient for the development of immune complex-mediated glomerulonephritis. The B cell-specific Lyn-deficient mice have no defects in early bone marrow B cell development but have reduced numbers of mature B cells with poor germinal centers, as well as increased numbers of plasma and B1a cells, similar to the lyn(-/-) animals. Within 8 mo of life, B cell-specific Lyn mutant mice develop high titers of IgG anti-Smith Ag ribonucleoprotein and anti-dsDNA autoantibodies, which deposit in their kidneys, resulting in glomerulonephritis. B cell-specific Lyn mutant mice also develop myeloproliferation, similar to the lyn(-/-) animals. The additional deletion of MyD88 in B cells, achieved by crossing lyn(flox/flox)Cd79a-cre mice with myd88(flox/flox) animals, reversed the autoimmune phenotype observed in B cell-specific Lyn-deficient mice by blocking production of class-switched pathogenic IgG autoantibodies. Our results demonstrate that B cell-intrinsic Lyn-dependent signaling pathways regulate B cell homeostasis and activation, which in concert with B cell-specific MyD88 signaling pathways can drive the development of autoimmune disease.

Differences in clinical findings, pathology, and outcomes between C3 glomerulonephritis and membranoproliferative glomerulonephritis.

BACKGROUND: To clarify the clinical manifestations of pediatric complement component C3 glomerulonephritis (C3GN), we retrospectively evaluated differences in the clinicopathological findings and prognosis between C3GN and immune-complex-mediated membranoproliferative glomerulonephritis (IC-MPGN). METHODS: Thirty-seven patients diagnosed with "idiopathic MPGN" were enrolled in this retrospective study. The patients were divided into two groups, with Group 1 consisting of 19 patients diagnosed with IC-MPGN and Group 2 consisting of 18 patients diagnosed with C3GN. The clinical findings and the prognosis were investigated for both groups. RESULTS: Thirteen patients in Group 2 were identified by mandatory annual school screening for urinary abnormalities. The incidence of macro-hematuria and the frequency of low serum C4 values were lower in Group 2 patients than in Group 1 patients. At the time of the second renal biopsy, urinary protein excretion, incidence of hematuria, frequency of low serum C3 values, and scores for mesangial proliferation, glomerular sclerosis, and interstitial fibrosis were higher in Group 2 patients than in Group 1 patients. At the most recent follow-up examination, the number of patients categorized as non-responding or with end-stage renal disease was higher in Group 2 patients than in Group 1 patients. CONCLUSIONS: Our results suggest that the treatment response and prognosis of patients with C3GN are worse than those of patients with IC-mediated MPGN. Therefore, in the clinical context regarding treatment options and prognosis, it may be useful to classify idiopathic MPGN as C3GN or IC-MPGN. In addition, long-term follow-up of C3GN is necessary.

Bronchial lesions of mouse model of asthma are preceded by immune complex vasculitis and induced bronchial associated lymphoid tissue (iBALT).

We systematically examined by immune histology the lungs of some widely used mouse models of asthma. These models include sensitization by multiple intraperitoneal injections of soluble ovalbumin (OVA) or of OVA with alum, followed by three intranasal or aerosol challenges 3 days apart. Within 24 h after a single challenge there is fibrinoid necrosis of arterial walls with deposition of immunoglobulin (Ig) and OVA and infiltration of eosinophilic polymorphonuclear cells that lasts for about 3 days followed by peribronchial B-cell infiltration and slight reversible goblet cell hypertrophy (GCHT). After two challenges, severe eosinophilic vasculitis is present at 6 h, increases by 72 h, and then declines; B-cell proliferation and significant GCHT and hyperplasia (GCHTH) and bronchial smooth muscle hypertrophy recur more prominently. After three challenges, there is significantly increased induced bronchus-associated lymphoid tissue (iBALT) formation, GCHTH, and smooth muscle hypertrophy. Elevated levels of Th2 cytokines, IL-4, IL-5, and IL-13, are present in bronchial lavage fluids. Sensitized mice have precipitating antibody and positive Arthus skin reactions but also develop significant levels of IgE antibody to OVA but only 1 week after challenge. We conclude that the asthma like lung lesions induced in these models is preceded by immune complex-mediated eosinophilic vasculitis and iBALT formation. There are elevations of Th2 cytokines that most likely produce bronchial lesions that resemble human asthma. However, it is unlikely that mast cell-activated atopic mechanisms are responsible as we found only a few presumed mast cells by toluidine blue and metachromatic staining limited to the most proximal part of the main stem bronchus, and none in the remaining main stem bronchus or in the lung periphery.

PF-1355, a mechanism-based myeloperoxidase inhibitor, prevents immune complex vasculitis and anti-glomerular basement membrane glomerulonephritis.

Small vessel vasculitis is a life-threatening condition and patients typically present with renal and pulmonary injury. Disease pathogenesis is associated with neutrophil accumulation, activation, and oxidative damage, the latter being driven in large part by myeloperoxidase (MPO), which generates hypochlorous acid among other oxidants. MPO has been associated with vasculitis, disseminated vascular inflammation typically involving pulmonary and renal microvasculature and often resulting in critical consequences. MPO contributes to vascular injury by 1) catabolizing nitric oxide, impairing vasomotor function; 2) causing oxidative damage to lipoproteins and endothelial cells, leading to atherosclerosis; and 3) stimulating formation of neutrophil extracellular traps, resulting in vessel occlusion and thrombosis. Here we report a selective 2-thiouracil mechanism-based MPO inhibitor (PF-1355 [2-(6-(2,5-dimethoxyphenyl)-4-oxo-2-thioxo-3,4-dihydropyrimidin-1(2H)-yl)acetamide) and demonstrate that MPO is a critical mediator of vasculitis in mouse disease models. A pharmacokinetic/pharmacodynamic response model of PF-1355 exposure in relation with MPO activity was derived from mouse peritonitis. The contribution of MPO activity to vasculitis was then examined in an immune complex model of pulmonary disease. Oral administration of PF-1355 reduced plasma MPO activity, vascular edema, neutrophil recruitment, and elevated circulating cytokines. In a model of anti-glomerular basement membrane disease, formerly known as Goodpasture disease, albuminuria and chronic renal dysfunction were completely suppressed by PF-1355 treatment. This study shows that MPO activity is critical in driving immune complex vasculitis and provides confidence in testing the hypothesis that MPO inhibition will provide benefit in treating human vasculitic diseases.

Cholinergic receptors modulate immune complex-induced inflammation in vitro and in vivo.

Cholinergic neural output has been shown to modulate innate immune responses to infection, injury and ischemia through stimulation of α7 nicotinic acetylcholine receptors (α7nAChR) on mononuclear phagocytes. We tested the hypothesis that cholinergic neurotransmitters, similar to those released through activation of a neural reflex, regulate responses to products of the adaptive immune system, specifically immune complex (IC)-mediated activation of effector cells. In this study, we show that stimulation of α7nAChR on human polymorphonuclear neutrophils (PMNs) and blood mononuclear phagocytes in vitro attenuates C5aR- and FcγR-triggered generation of reactive oxygen species, expression of leukocyte markers involved in cell recruitment and adhesion, and release of TNF-α and other proinflammatory cytokines. We show that this pathway is operative in vivo. Ligation of cholinergic receptors blunts IC-triggered responses in the reverse peritoneal Arthus reaction in mice. The selective 7nAChR agonist GTS21 decreased PMN accumulation and release of cytokines and chemokines at sites of IC deposition. In addition, mice lacking α7nAChR had exaggerated responses to reverse peritoneal Arthus reaction characterized by increased infiltration of PMNs and elevated of levels of TNF-α and CXCL1 in peritoneal fluid compared with wild-type mice. Taken together, these findings suggest that cholinergic output has the potential to exert tonic inhibitory activity that dampens responses to ICs and C5a and thus may be a target to minimize tissue damage in autoimmune diseases.