Summary
Background
Hidradenitis suppurativa (HS) is a chronic inflammatory disease, characterized by painful, purulent and destructive skin alterations in intertriginous areas.
Objectives
We ...investigated the expression and role in HS of granulocyte colony‐stimulating factor (G‐CSF), the regulator of neutrophil biology, as clinical signs of a neutrophilic granulocyte‐driven inflammation are distinctive in the disease.
Methods
Skin and blood samples obtained from different cohorts of patients with HS and control individuals were assessed by RNA sequencing, quantitative polymerase chain reaction on reverse transcribed mRNA, and/or enzyme‐linked immunosorbent assay. Mechanistic studies using keratinocytes, dermal fibroblasts, immune cell populations and skin biopsies were performed.
Results
G‐CSF was abundant in HS skin, particularly in inflamed nodules and abscesses. Its levels even exceeded those found in other inflammatory skin diseases. Interleukin (IL)‐1 and IL‐17, respectively, induced G‐CSF production by fibroblasts and keratinocytes. These effects were enhanced by tumour necrosis factor (TNF)‐α and IL‐36. Accordingly, fibroblasts separated from HS lesions expressed G‐CSF, and IL‐1 receptor antagonist reduced G‐CSF levels in explanted HS skin. G‐CSF blood levels positively correlated with severity of HS. Elevated lesional G‐CSF receptor levels were linked to upregulation of molecules that contribute to prolonged activation of neutrophils by components of bacteria and damaged host cells formyl peptide receptor 1 (FPR1), FPR2 and free fatty acid receptor 2 (FFAR2), neutrophil survival TNF receptor superfamily member 10C (TNFRSF10C/TRAIL‐R3) and TNF receptor superfamily member 6B, kinases (tyrosine‐protein kinase HCK and hexokinase 3), and skin destruction MMP25 (matrix metalloproteinase 25) and ADAM8 (disintegrin and metalloproteinase domain‐containing protein 8). G‐CSF elevated the expression of FPR1, FFAR2, and TNFRSF10C/TRAIL‐R3 in neutrophils and synergized with bacterial components to induce skin‐destructive enzymes.
Conclusions
The G‐CSF pathway engages both tissue and immune cells, is strongly activated in HS lesions, and offers the opportunity to target the neutrophil‐driven inflammation.
What is already known about this topic?
Hidradenitis suppurativa (HS) is a chronic debilitating skin disorder with a very high, unmet medical need.
The diseased skin in patients with HS shows distinct features of a neutrophil‐driven inflammation (e.g. abscess formation, purulent discharge).
Granulocyte colony‐stimulating factor (G‐CSF) is the major regulator of neutrophil development, survival and function.
What does this study add?
HS lesions show highly increased levels of G‐CSF and its receptor.
Major G‐CSF inducers are interleukin (IL)‐1β and IL‐17.
In neutrophils, G‐CSF upregulates receptors for components of bacteria and damaged host cells, decoy receptors for apoptosis inducers and proteases.
The production of skin‐destructive enzymes induced by bacterial components is strengthened by G‐CSF in neutrophils.
G‐CSF inducers and molecules upregulated by G‐CSF in neutrophils in vitro are abundant in HS lesions.
What is the translational message?
G‐CSF is the central element of a pathogenetic pathway in HS.
The G‐CSF pathway may contribute to the persistence of abscesses, purulent secretion and progressive skin structure destruction.
Targeting G‐CSF or its pathway elements may represent an approach for the treatment of HS and other conditions with neutrophil‐driven inflammation and skin destruction.
Linked Comment: E.J. Giamarellos‐Bourboulis. Br J Dermatol 2021; 185:15–16.
The adoptive transfer of natural regulatory T cells (nTreg) is a new option to reshape undesired immune reactivity in autoimmunity and transplantation toward “tolerance.” The first clinical trials ...using adoptive transfer of polyclonal nTreg demonstrated safety and hints of efficacy. However, the low frequencies of antigen‐specific cells among the pool of polyclonal nTreg and their broad antigen nonspecific suppression are limitations of this approach regarding efficacy and safety. Recently, the isolation and expansion of (allo)antigen‐specific nTreg have successfully been achieved by using Treg‐specific activation markers but the yield is relatively low. Here, we describe a novel good manufacturing practice (GMP)‐compatible expansion protocol of alloantigen‐specific nTreg based on the stimulation of nTreg by allogeneic activated B cells. Their functionality and specificity are superior compared to polyclonal nTreg both in vitro and in vivo. Employing an allogeneic B cell bank, designed to cover the majority of HLA types, allows fast GMP‐compliant manufacturing for donor‐specific nTreg for clinical application in organ and stem cell transplantation. TCR repertoire analyses by next generation sequencing revealed impressive expansion by several log‐steps of even very low‐abundance alloantigen‐specific nTreg clones. This novel method offers a simple approach for expanding antigen‐specific nTreg and is characterized by high replicability and easy transferability to full GMP standards.
An allogeneic B cell bank allows fast and GMP‐compatible expansion of even low‐abundance allospecific nTreg clones, which are more potent suppressors of specific alloresponses than their polyclonal counterparts, both in vitro and in vivo.
Clonotype analysis is essential for complete characterization of antigen‐specific T cells. Moreover, knowledge on clonal identity allows tracking of antigen‐specific T cells in whole blood and tissue ...infiltrates and can provide information on antigenic specificity. Here, we developed a next generation sequencing (NGS)‐based platform for the highly quantitative clonotype characterization of T cells and determined requirements for the unbiased characterization of the input material (DNA, RNA, ex vivo derived or cell culture expanded T cells). Thereafter we performed T cell receptor (TCR) repertoire analysis of various specimens in clinical settings including cytomegalovirus (CMV), polyomavirus BK (BKV) reactivation and acute cellular allograft rejection. Our results revealed dynamic nature of virus‐specific T cell clonotypes; CMV reactivation was linked to appearance of new highly abundant antigen‐specific clonalities. Moreover, analysis of clonotype overlap between BKV‐, alloantigen‐specific T cell–, kidney allograft‐ and urine‐derived lymphocytes provided hints for the differential diagnosis of allograft dysfunction and enabled appropriate therapy adjustment. We believe that the established approach will provide insights into the regulation of virus‐specific/anti‐tumor immunity and has high diagnostic potential in the clinical routine.
This study describes the development and characterization of T cell receptor repertoire analysis based on Next Generation Sequencing and its application for complex differential diagnosis of posttransplant kidney dysfunction.
Although the immunomodulatory potency of mesenchymal stromal cells (MSC) is well established, the mechanisms behind are still not clear. The crosstalk between myeloid dendritic cells (mDC) and ...natural killer (NK) cells and especially NK cell-derived interferon-gamma (IFN-γ) play a pivotal role in the development of type 1 helper (Th1) cell immune responses. While many studies explored the isolated impact of MSC on either in vitro generated DC, NK, or T cells, there are only few data available on the complex interplay between these cells. Here, we investigated the impact of MSC on the functionality of human mDC and the consequences for NK cell and Th1 priming in vitro and in vivo. In critical limb ischemia patients, who have been treated with allogeneic placenta-derived mesenchymal-like stromal cells (PLX-PAD), no in vivo priming of Th1 responses toward the major histocompatibility complex (MHC) mismatches could be detected. Further in vitro studies revealed that mDC reprogramming could play a central role for these effects. Following crosstalk with MSC, activated mDC acquired a tolerogenic phenotype characterized by reduced migration toward CCR7 ligand and impaired ability to stimulate NK cell-derived IFN-γ production. These effects, which were strongly related to an altered interleukin (IL)-12/IL-10 production by mDC, were accompanied by an effective prevention of Th1 priming in vivo. Our findings provide novel evidence for the regulation of Th1 priming by MSC via modulation of mDC and NK cell crosstalk and show that off-the-shelf produced MHC-mismatched PLX-PAD can be used in patients without any sign of immunogenicity.
Despite remarkable progress in organ transplantation through the development of a wealth of immunosuppressive drugs highly effective at controlling acute rejection, two major problems still remain, ...the loss of transplants due to chronic rejection and the growing number of sensitized recipients due to previous transplants, transfusions or pregnancies. Induction of immune tolerance appears to be the only way to curb this complex situation. Here we describe that a therapy, already successfully used to restore immune tolerance to self‐antigens in overt autoimmunity, is effective at promoting transplant tolerance. We demonstrate that a short low‐dose course with CD3 antibodies started after transplantation, at the time of effector T cell priming to alloantigens, induces permanent acceptance of fully mismatched islet allografts. Mechanistic studies revealed that antigen‐specific regulatory and effector T cells are differentially affected by the treatment. CD3 antibody treatment preferentially induces apoptosis of activated alloreactive T cells which is mandatory for tolerance induction. In contrast, regulatory T cells are relatively spared from CD3 antibody‐induced depletion and can transfer antigen‐specific tolerance thus arguing for their prominent role in sustaining long‐term graft survival.
Short, low‐dose courses of CD3‐specific antibodies started after transplantation, at the time of effector T cell priming to alloantigens, induces immune tolerance to fully mismatched islet allografts by preferentially targeting activated alloreactive T cells while sparing regulatory T cells.
Abstract Infections are a leading cause of death in patients with acute CNS injury such as stroke. Recent experimental evidence indicated that stroke leads to suppression of innate and adaptive ...peripheral immune responses which predisposes to infection. However, less is known on phenotypic and functional immune alterations in correlation with the occurrence of infectious complications in patients with acute stroke. Experimental procedures: In the recently completed randomized, double blind, placebo-controlled Preventive Antibacterial Therapy in Stroke (PANTHERIS) trial on the efficacy of short-term antibacterial therapy to prevent the development of post-stroke infections, we assessed longitudinal changes in lymphocyte subpopulations and mitogen-induced lymphocytic interferon gamma (IFN)-γ production using flow cytometry in 80 patients with acute severe stroke at days 1, 3, 8, 90 and 180 after clinical onset. Plasma interleukin (IL)-6 and IL-10 concentration as well as urinary levels of norepinephrine and cortisol was assessed within the first 8 days after stroke. Patients of the placebo and verum (moxifloxacin) treatment groups who did or did not develop infections within 11 days after stroke were compared to identify immunological changes associated with the occurrence of post-stroke infections. Results: Rapid T-lymphopenia and long-lasting suppression of lymphocytic IFN-γ production were observed in all stroke patients. Patients of the placebo group who developed infections showed a trend toward greater decline of CD4+ Th cell counts and higher urinary levels of norepinephrine early after stroke than patients without infections. Onset of infections was accompanied with higher plasma IL-6 levels in the placebo group but not in the moxifloxacin group. In addition, an early rise in plasma IL-10 was detected in patients who developed infections despite preventive antibacterial treatment. Conclusion: A rapid loss and functional deactivation of T cells are common changes in stroke patients consistent with immunodepression after brain ischemia. A stronger decrease in cellular immune responses and an increased sympathetic activity after stroke are associated with a higher risk of infections. Increased plasma levels of the anti-inflammatory cytokine IL-10 early after stroke may identify patients who will not respond to preventive antibacterial therapy with moxifloxacin.
Interleukin (IL)-10 is an important immunoregulatory cytokine produced by many cell populations. Its main biological function seems to be the limitation and termination of inflammatory responses and ...the regulation of differentiation and proliferation of several immune cells such as T cells, B cells, natural killer cells, antigen-presenting cells, mast cells, and granulocytes. However, very recent data suggest IL-10 also mediates immunostimulatory properties that help to eliminate infectious and noninfectious particles with limited inflammation. Numerous investigations, including expression analyses in patients, in vitro and animal experiments suggest a major impact of IL-10 in inflammatory, malignant, and autoimmune diseases. So IL-10 overexpression was found in certain tumors as melanoma and several lymphomas and is considered to promote further tumor development. Systemic IL-10 release is a powerful tool of the central nervous system to prevent hyperinflammatory processes by activation of the neuro-endocrine axis following acute stress reactions. In contrast, a relative IL-10 deficiency has been observed and is regarded to be of pathophysiological relevance in certain inflammatory disorders characterized by a type 1 cytokine pattern such as psoriasis. Recombinant human IL-10 has been produced and is currently being tested in clinical trials. This includes rheumatoid arthritis, inflammatory bowel disease, psoriasis, organ transplantation, and chronic hepatitis C. The results are heterogeneous. They give new insight into the immunobiology of IL-10 and suggest that the IL-10/IL-10 receptor system may become a new therapeutic target.
Summary
Background
Acne inversa (AI)/hidradenitis suppurativa is a chronic inflammatory disease characterized by painful axillary, inguinal and perianal skin lesions with deep‐seated nodules, ...abscesses and fistulae.
Objectives
This study aimed to identify and characterize the key players in AI pathogenesis.
Methods
Epidemiological and anamnestic data for patients with AI were collected, and blood and skin samples were also taken. Healthy participants and patients with psoriasis served as controls. Assessment of samples and cultures of primary cells was performed by enzyme‐linked immunosorbent assay, quantitative polymerase chain reaction on reverse transcribed mRNA, and immunohistochemistry.
Results
Of 35 mediators quantified in the blood of patients with AI, lipocalin‐2 (LCN2) appeared as one of the most significantly upregulated parameters compared with healthy participants 85·8 ± 12·2 (n = 18) vs. 41·8 ± 4·2 (n = 15); P < 0·001. Strongly elevated LCN2 expression was present in AI lesions, with granulocytes and keratinocytes being sources of this expression. In vitro, these cells upregulated LCN2 production in response to tumour necrosis factor (TNF)‐α, and a positive relationship between systemic TNF‐α and LCN2 levels (rs = 0·55, P = 0·011; n = 20) was evident for AI. LCN2 blood levels correlated with AI disease severity (rs = 0·65, P < 0·001; n = 29), but not with disease duration, age, sex, body mass index or smoking habit. Detailed analyses revealed a link with the number of skin regions containing nodules and fistulae, but not scars.
Conclusions
LCN2 might serve as a blood biomarker for the objective assessment of inflammatory activity in AI. We suggest a self‐amplification loop comprising TNF‐α, neutrophilic granulocytes and LCN2, which contributes to the recurrent skin neutrophil infiltration in AI, clinically evident as pus.
What's already known about this topic?
Acne inversa (AI) is a painful and debilitating chronic inflammatory disease with unknown pathogenesis and a profound medical need.
The efficacy of the recently approved anti‐tumour necrosis factor (TNF)‐α treatment suggests an important role for TNF‐α in the maintenance of AI lesions.
Physicians currently lack a commonly used dynamic scoring system and a blood biomarker for assessing the inflammatory activity of the skin alterations.
What does this study add?
Lipocalin‐2 (LCN2) is one of the most significantly upregulated blood parameters in patients with AI.
In AI lesions, granulocytes and, to a lower extent, keratinocytes produce LCN2.
While TNF‐α seemed to be the sole inducer of granulocyte LCN2, interleukin (IL)‐1β, IL‐17 and TNF‐α provoke keratinocyte LCN2 production.
In AI, LCN2 blood levels positively correlate with Sartorius score and with TNF‐α blood levels.
What is the translational message?
Quantifying LCN2 blood levels may be used for objective estimation of disease severity in patients with AI.
LCN2 blood levels should be tested as a predictor for therapy decisions and assessment of therapy response (especially for anti‐TNF‐α treatment).
Anti‐TNF‐α treatment may interrupt the suggested vicious circle comprising TNF‐α, neutrophilic granulocytes, and LCN2, that contributes to the high cutaneous number of neutrophils, clinically evident as pus in AI.
Linked Comment: Mössner. Br J Dermatol 2017; 177:1162–1164.
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Quality of life (QoL) plays a significant role in the treatment of dogs with idiopathic epilepsy (IE), yet is so far understudied. This study describes the outcome evaluation of an online ...questionnaire based on the carer's perception focusing on 62 QoL questions in 159 dogs with IE. Results showed that seizure frequency, but not seizure severity or presence of cluster seizures, was significantly associated with carer-perceived dog's QoL. Dogs receiving third-line antiepileptic drugs had a significantly lower perceived QoL than those that did not. Generalised linear mixed model analysis demonstrated that severity of the side effects sleeping more and ataxia were significantly associated with carer-perceived dog's QoL, with higher severities predicting lower QoL scores. The degree of carer acceptability of seizure frequency and severity was significantly associated with the dog's reported seizure frequency and severity. Moreover, there was a significant association between IE-related QoL changes of the dog and the carer, with reductions in perceived canine QoL scores associated with reductions in carer QoL, and vice versa. In conclusion, aspects of canine IE can affect both the carer and their dog's QoL. This has implications for the management and requires consideration when treatment options and outcomes are discussed.
The early identification of renal transplant recipients at enhanced risk of developing acute and subclinical rejection would allow individualized adjustment of immunosuppression before functional ...graft injury occurs and would exclude these patients from drug-weaning studies. Protein and reverse transcriptase-polymerase chain reaction-based analyses of candidate markers in urine open the opportunity to closely monitor kidney-transplanted patients non-invasively. The chemokine interferon-inducible protein 10 (IP-10; CXCL10) might be an interesting candidate to uncover ongoing immune processes within the graft. Urine samples from kidney-transplanted recipients were retrospectively analyzed for IP-10 mRNA and protein expression. IP-10 levels were correlated with the incidence of acute rejection episodes proven by histology and long-term graft function assessed by the glomerular filtration rate 6 months post transplantation. IP-10 expression in urine identified patients with ongoing acute rejection episodes several days before a biopsy was indicated by rising serum creatinine levels. Most importantly, elevated levels of urinary IP-10 protein within the first four postoperative weeks were predictive of graft function at 6 months even in the absence of acute rejection. These data reveal a correlation between elevated IP-10 expression in urine at early time points post-transplantation and intragraft immune activation that leads to acute rejection and compromised long-term graft function.
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