Antineutrophil Antibody Research Articles

Treatment response and complications of older patients with ANCA(antineutrophil cytoplasmatic antibody)-associated vasculitis

ANCA (antineutrophil cytoplasmatic antibody)-associated vasculitis (AAV) mainly affects elderley people but adjusted therapy concepts for this patient group are lacking. The aim of this study was therefore to analyze differences in course and outcome of patients with AAV with respect to age. 62patients were analyzed for treatment response, of whom 53 (85%) experienced adverse events (AE and SAE) that could be evaluated. Older (> 65 yrs.) versus younger (< 65 yrs.) patients were compared. Treatment response was assessed at 6months, complications were assessed over 18months. Treatment response was not seen to differ by age groups. In multiple logistic regression, pulmonary involvement (OR = 6,9; CI = 1,7-27,8, p < 0,01) and ΔGFR [ml/min] (OR = 0,93; CI = 0,89-0,97, p < 0,01) were predictors of SAE. 14patients had more than 1SAE. Again, pulmonary involvement (28,2% vs. 78,6%, p < 0,01) was a risk factor and older patients (78,6% vs. 43,6%, p = 0,025) were more frequently affected. Patients with multiple SAEs received glucocorticoids of more than 5 mg/d for longer periods of time (171± 65days vs. 120± 70days, p = 0,03). No differences were found between older and younger patients with regard to treatment response. Multiple SAEs occurred more frequently in elderly patients. There was a correlation between pulmonary manifestation and duration of glucocorticoid therapy with a complicated course. The most frequent SAEs were infections requiring hospitalisation. Therapy for elderly patients should be individualized with the goal of a fast reduction of glucocorticoids. Special monitoring is indicated for elderly patients, especially those with pulmonary involvement.

Neutrophil Depletion Attenuates Acute Liver Stress after Exhaustive Exercise in Mice.

Exhaustive exercise induces acute liver stress; however, the precise mechanisms remain unclear. We investigated the effects of neutrophil depletion in male C57BL/6J mice. Male C57BL/6J mice were divided into four groups: sedentary with control antibody ( n = 20), sedentary with antineutrophil antibody ( n = 20), exhaustive exercise with control antibody ( n = 20), and exhaustive exercise with antineutrophil antibody ( n = 20). Antineutrophil antibodies (1A8) or control antibodies were administered intraperitoneally before running on a treadmill. Immediately and at 24 h after running to exhaustion on a treadmill at a 7% gradient and a speed of 24 m·min -1 , blood neutrophil counts were measured by flow cytometry. Plasma activities of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were also measured. Hematoxylin-eosin staining was performed to calculate the liver stress score, and hepatic tumor necrosis factor-α was measured using enzyme-linked immunosorbent assay. Exercise increased blood neutrophil and neutrophil infiltration into the liver. Plasma AST and ALT activities were significantly higher immediately after exhaustive exercise than after sedentary control (AST, sedentary with control antibody: 52.2 ± 0.4, exhaustive exercise with control antibody: 210.0 ± 19.8; ALT, sedentary with control antibody: 29.8 ± 2.2, exhaustive exercise with control antibody: 87.2 ± 15.8). However, AST and ALT activities were significantly decreased with the 1A8 antibody (AST, 102.2 ± 12.9; ALT, 39.2 ± 4.0). In addition, the liver stress score increased after exercise but was significantly reduced by prior 1A8 antibody administration. The 1A8 antibody treatment also decreased hepatic tumor necrosis factor-α levels after exhaustive exercise. These results suggested that neutrophils play a critical role in increasing liver stress by regulating inflammation.

Diagnosis and management of isolated neutropenia: A survey of pediatric hematologist oncologists.

Isolated neutropenia is a common referral to pediatric hematology oncology (PHO) physicians. There are no established consensus guidelines in the diagnosis and management of patients with isolated, asymptomatic, and incidentally discovered neutropenia. A survey was distributed to PHO physicians on the American Society of Pediatric Hematology Oncology member discussion page to determine the common diagnostic and management decisions regarding patients with isolated neutropenia and to explore beliefs regarding the term "benign ethnic neutropenia." One hundred twenty-six PHO attending physicians completed the survey. The most common tests reportedly ordered for this patient population included complete blood cell count (CBC) (98%), peripheral smear (75%), antineutrophil antibody testing (29%), and immunoglobulins (24%). Providers were more likely to order an antineutrophil antibody in toddlers (p=.0085), and antinuclear antibody (ANA) panels in adolescents (p<.001). Half of providers do not request additional CBCs prior to their initial consultation, and most suggest referring patients with mild neutropenia after confirming a declining absolute neutrophil count (ANC) (51%). The three most important factors influencing ongoing follow-up included: history of recurrent/severe infections (98%), family history of blood disorders (98%), and more severe/progressively worsening neutropenia (97%). Seventy percent of respondents have diagnosed patients with "benign ethnic neutropenia," and 75% support replacement of the term to "typical neutrophil count with Fy(a-/b-) status," if confirmed with red cell phenotyping. We identified practice patterns of PHO physicians for the diagnosis and management of patients referred for asymptomatic and isolated neutropenia. These data provide the framework to conduct cost-effectiveness studies.

Diagnosis and Management of Isolated Neutropenia: A Survey of Pediatric Hematologists Oncologists

Background: Isolated neutropenia is a common referral to pediatric hematology oncology. Guidelines exist regarding the management of severe congenital neutropenia as well as the differential diagnosis for isolated neutropenia. However, no expert guidelines exist to inform practice patterns on the diagnosis and management of patients with asymptomatic neutropenia or individuals with typical neutrophil counts associated with the Duffy null phenotype (Fy[a-/b-]). Understanding the practice patterns of pediatric hematologists oncologists may inform future efforts to reduce unnecessary medical testing and follow-up in these patient populations. Methods: We distributed a twenty-one-question electronic survey to pediatric hematologists oncologists through the American Society of Pediatric Hematology Oncology's member discussion board from March 2022 until April 2022. The purpose of the survey was to identify practice patterns regarding patients referred for isolated neutropenia. Eligible participants included attending physicians and those who regularly see referrals for neutropenia. All trainees, advanced care providers, and individuals who do not regularly see patients with neutropenia were excluded. The survey explored: 1) common diagnostic studies physicians use to evaluate patients referred with asymptomatic and isolated neutropenia, 2) frequency of patient follow-up, 3) use of limited red cell phenotyping in pediatric patients with isolated neutropenia, 4) common terminology used to describe isolated neutropenia, and 5) whether physicians support removal of the term "benign ethnic neutropenia" from practice. We also presented two clinical case vignettes of asymptomatic and isolated neutropenia in a one-year-old male and a twelve-year old female. We compared management responses to the two vignettes to general neutropenia practice patterns. Results: One hundred and twenty-six pediatric hematology oncology attending physicians who regularly see neutropenia referrals completed the survey and were included in the analysis. Selected survey responses are shown in the Table. The most common tests ordered for pediatric patients with isolated neutropenia include complete blood cell count (CBC) (98%), peripheral smear (75%), anti-neutrophil antibody testing (29%), and immunoglobulins (24%). In the clinical vignettes, providers were more likely to order an anti-neutrophil antibody in toddlers (p=0.0085), and antinuclear antibody (ANA) panels in adolescents (p<0.001). Half of providers ask for additional CBCs prior to their initial consultation. Most respondents suggest referring patients with mild neutropenia after demonstrating a decline in absolute neutrophil count (ANC) (51%). For patients with mild neutropenia, most providers order an additional CBC to ensure resolution of their neutropenia or do not follow after their initial evaluation (87%). For patients with moderate neutropenia, most providers continue to follow patients even after their initial evaluation (46%). The three most important factors that influence continued follow-up of patients with isolated neutropenia include: history of recurrent/severe infections (98%), family history of blood disorders (98%), and more severe/progressively worsening neutropenia (97%). Seventy percent of respondents have diagnosed patients with the term "benign ethnic neutropenia," and 75% support replacement of the term to "typical neutrophil count with Fy(a-/b-) status," if confirmed with limited red cell phenotyping. Conclusion: We identify the common practice patterns of pediatric hematologists oncologists caring for patients referred for asymptomatic and isolated neutropenia. Data from this study is providing the framework for a cost-effectiveness study aimed to reduce the healthcare and family burden of referrals for isolated neutropenia to pediatric hematologists oncologists. Furthermore, this survey demonstrates support for replacement of the term "benign ethnic neutropenia" with "typical neutrophil count with Fy(a-/b-) status" in applicable patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Assessment of the Frequency of Variants of Undetermined Significance (VUS) in Adult Patients with Chronic Idiopathic Neutropenia Studied for Myeloid Gene Mutations By Next Generation Sequencing

Background: Chronic Idiopathic Neutropenia (CIN) is a neutrophil disorder characterized by the persistent and unexplained reduction in the number of peripheral blood (PB) neutrophils. The diagnosis of CIN is based on exclusion criteria, namely the absence of clinical/laboratory evidence of any underlying disease or use of drugs that might be associated with neutropenia, absence of history of exposure to irradiation or use of chemical compounds, negative antineutrophil antibodies, and normal bone marrow (BM) morphology and karyotype. We have previously performed next generation sequencing (NGS) analysis of genes recurrently mutated in myeloid malignancies in a cohort of CIN patients and we found that 11.35% of the patients display clonal hematopoiesis in term of presence of pathogenic/likely pathogenic variants with VAF higher than 2%. Variants of undetermined significance (VUS) were ruled out from the analysis and their potential significance was not assessed. Aims: In the current study we have assessed the presence, frequency and potential clinical significance of VUS detected in our cohort of CIN patients previously studied for clonal hematopoiesis. Methods: Genomic DNA was extracted from CIN patients' BM or PB samples, sequencing libraries were prepared and subjected to targeted NGS on an Ion S5 Prime Sequencer (Thermo Fisher Scientific) using a panel of 38 genes recurrently mutated in myeloid malignancies (Tsaknakis G et al. Blood. 2021;138:1249-1257). Results: NGS analysis of myeloid genes has been performed in 139 CIN patients (106 female subjects and 33 male subjects, aged 21 to 89 years with a median age of 59 years). Re-analysis of patients' NGS data and assessment of VUS revealed that 35 out of 139 patients (25.18%) carried one or more VUS (a total of 39 VUS were detected, all with an allelic frequency of 50% and all missense variants). Genetic variants were evaluated according to the ACMG guidelines and classified as VUS. The majority of VUSs were observed in ATM (n=7) and SETBP1 (n=6) genes followed by TET2, NOTCH1 (4 occurrences respectively) and PLCG2 (n=3). Two different patients shared the same variant in TET2 gene whereas two siblings shared the same variant in SETBP1 gene. Pathogenicity score calculation based on the combined evidence from multiple other in-silico predictors identified 3 variants in 3 different genes as having the highest probability to be pathogenic. Namely these were PLCG2K456E,CXCR4R235C and SF3B1T56I. PLCG2 variants have been implicated in familial cold autoinflammatory syndrome 3 (PLCG2 associated antibody deficiency and immune dysregulation, PLAID) and in autoinflammation, antibody deficiency, and immune dysregulation syndrome (APLAID) whereas CXCR4 variants are known to be associated with Warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome. SF3B1 mutations have not yet been shown to be involved in hereditary disease. Conclusions: Our study of VUS assessment in a cohort of 139 well characterized adult CIN patients identified 39 VUS, all heterozygous, in a total of 35 patients. Further steps include paired analysis with constitutional reference tissue to determine the germline nature of the mutations. Moreover, these variants should be functionally tested and integrated with functional results and clinical phenotypes to more accurately interpret their potential contribution to disease onset. This study of VUS analysis in CIN patients is anticipated to contribute to the better understanding of the pathogenetic causes of this rare disease.

Administration of Obinutuzumab, and Not Rituximab, during Induction and Maintenance for Follicular Lymphoma Increases the Likelihood of Developing Delayed Neutropenia

The results of the PRIMA trial published a decade ago, substantiated the benefit of maintenance therapy with anti-CD20 monoclonal antibodies (MoAb), following chemo-immunotherapeutic induction, for prolongation of the progression-free survival (PFS) of patients with high-tumor- burden follicular lymphoma (FL). Rituximab-associated delayed neutropenia, defined as an event occurring after recovery of cytopenias related to the chemotherapy backbone, has been reported in lymphoma patients, with the prevalence varying between 8-27%. Several mechanisms have been implicated in its pathogenesis, mainly related to B-cell recovery, such as a disrupted stromal-derived factor-1 gradient and the generation of anti-neutrophil antibodies. The available data associated with anti-CD20 MoAb-mediated delayed neutropenia during the prolonged (2-year) maintenance treatment in FL are limited, and the consequences of its occurrence remain unclear. The aims of this study were to assess the prevalence of neutropenia and risk factors for its development in FL patients during maintenance therapy as well as to evaluate its severity and impact on decisions regarding modification of the original treatment protocol. In this retrospective study, clinical data were retrieved from electronic medical records of FL patients treated at the Rambam Health Care Campus between 2006 (when maintenance therapy was first applied) and 2021. Continuous variables were statistically processed using the Mann Whitney U test and categorical variables - using Fisher exact tests. Statistical significance was defined as p <0.05. During the study period, a total of 305 patients treated for high-tumor-burden FL were identified in the institutional database; 164 of these patients (53.8%) also received maintenance therapy with anti-CD20 MoAb (rituximab or obinutuzumab). Data on 157/164 patients were included in the final analysis after the exclusion of 7 patients due to their concomitant autoimmune disease. During maintenance therapy, 37/157 (23.6%) patients developed delayed neutropenia, with 4/37 (11%) of them experiencing at least one recurrent event. This relatively high percentage of delayed events is comparable only to that observed in a study which evaluated heavily pretreated lymphoma patients (Cattaneo C et al. Leuk Lymphoma, 2006), exposed to highly myelotoxic regimens or fludarabine and did not include a maintenance phase. Median time from the start of maintenance to the first neutropenic episode in the current study was 5 (range 1.5-12) months and the majority of these events were of high grade (III and IV in 24% and 38% of patients, respectively). Despite the fact that 15/37 (40%) patients who developed this complication were hospitalized with neutropenic fever, the occurrence of neutropenia led to early discontinuation of maintenance treatment solely in 14% of the cohort and had no effect on PFS, measured at a median follow-up of 91.5 months. Univariate and multivariate analyses identified the use of the obinutuzumab/bendamustine (O/B) combination for induction therapy, or maintenance with obinutuzumab (O) as the only risk factors of delayed neutropenia development in patients on maintenance therapy (odds ratios in case of O/B induction followed by O maintenance: 4.433 [95%CI=1.387-14.17, p=0.012]; in case of O maintenance: 3.048 [95%CI=1.207-7.697, p=0.018]). All other laboratory and clinical factors examined did not impact late neutropenia occurrence (Table 1). The current study demonstrates that the administration of obinutuzumab both as part of induction with bendamustine and as maintenance therapy increases the likelihood of developing anti-CD20 MoAb-induced delayed neutropenia, suggesting that pharmacokinetic differences between type I and type II MoAb are partly responsible for this effect. Moreover, the findings regarding the prevalence of neutropenia and time to its first episode in this study cohort do not support the involvement of processes related to B-cell recovery in the pathogenesis of this complication during maintenance treatment, and warrant investigation of alternative mechanisms. The risk factors for delayed neutropenia in FL patients on maintenance therapy, identified in this study, should be considered in the treatment paradigm for older frail patients who are more prone to complications of neutropenic fever. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

IS GFI-1 A TRIGGER TO DEVELOPMENT OF AUTOIMMUNE NEUTROPENIA?

To describe a child carrying a GFI1 -variant with autoimmune neutropenia. Single subject case report. This is a case report of a 1 year-and-10-months-old female presenting severe chronic neutropenia (0,03 × 10 9 /L) with mild infections. The frequency of infections reduced as she got older. Physical examinations outside of infections were always normal and she had expected development. She did not have monocytosis or other hematologic disorders, and lymphocytes evaluation by flow cytometry analysis resulted normal. Viral diseases were negative, immunoglobulins dosage was normal for the age group, and screening for systemic autoimmune disease (sAID) was negative. Bone marrow aspirate analysis with karyotype resulted normal, without maturation arrest or dysplasia. Inborn errors of metabolism (IRM) and immunity (IRI) were ruled out. Antineutrophil (anti-HNA) autoantibody was detected on two occasions: at ages of 2-years-and-9-months-old and 5-years-and-3-months-old, presenting the same intensity and the same pan-reactivity to the FCγRIIIb glycoprotein. She has no siblings, but her father presented recurrent otitis until 6-year-old and hemogram with a persistent mild neutropenia documented since childhood. Ethnic benign neutropenia was ruled out. Both expressed Fyb on erythrocytes and were heterozygotes for single nucleotide polymorphism rs2814778 in the ACKR1 . She did not present remission of the disease until the age of 5 years and 7 months. A whole exome sequencing, showed a heterozygotic non-pathogenic GFI-1 variant in exon 6 (c.929G>A:p.Arg310Gln). The variant, confirmed by Sanger sequencing, was also present in her father. He has no antineutrophil antibodies detected to date. Most cases of pAIN are diagnosed between 3 to 8 months. Spontaneous remission in about 90% could be expected in 7 to 24 months of diagnosis or up to age of 4-5 years. In this reported case, the patient had severe neutropenia demonstrated since 1 year-and-10-month-old. Even 45 months after the beginning she remains neutropenic, carrying anti-HNA autoantibody with the same pan-reactivity features. The persistence of anti-HNA with no specificity may be associated to persistent autoimmune neutropenia (AIN) as well as a characteristic of secondary AIN, but screening for both sAID and IEI were negative. Due to the persistence of neutropenia, a whole exome sequencing was performed, presenting GFI-1 variant inherited from her father, also neutropenic. The patient does not present features of GFI-1 -variants, clinical course and the bone marrow were not compatible with SCN. In addition to regulating myelopoiesis, GFI-1 also plays a role in the maturation of B and T lymphocytes, and in process of antigen-mediated lymphocyte activation. Although mature B-cells do not express GFI-1 , this loss may be correlated with the increase of some IgG fractions, and some works have correlated GFI-1 changes with autoimmune conditions and could be the cause of AIN reported. For patients with persistent pAIN, perfoming tests to detect a genetic involvement may be useful to understand the cause of the disease. In this case, the presence of GFI-1 variant could be the trigger of AIN reported.

An update on the incidence, risk factors and mechanisms of rituximab-associated neutropenia.

With the increasing application of rituximab (RTB) in hematological diseases and autoimmune diseases (AIDs), we have gradually increased our awareness of the adverse reaction of rituximab-associated neutropenia (RAN), but little is known about its true incidence rate, susceptibility risk factors and exact pathogenesis. At present, research groups have conducted a large number of studies on different populations. The team found that age (> 60), advanced disease, systemic lupus erythematosus (SLE) and combined cyclophosphamide therapy were independent risk factors for RAN. However, its exact mechanism is not completely clear. Several hypotheses have been put forward to solve this question, including the production of anti-neutrophil antibodies after RTB, the generation disorder and neutrophil maturation stagnation caused by abnormal B-cell reconstruction, and the amplification of T-large granular lymphocyte population that may induce neutrophil apoptosis. However, there are still many unsolved problems in all aspects of RAN. This article is an update of the incidence rate, risk factors and mechanisms of RAN.

Clinical Challenge in Urology: A Nonhealing Wound on the Glans Penis

A 63-year-old male presented to his surgeon 6 months following circumcision for phimosis with complaints of a nonhealing wound on the glans penis. He reported persistent bleeding, pain, and progressive difficulty urinating in the months following the procedure. He was noted to have a grossly dehisced circumcision, with erythematous, beefy granulation tissue involving the entire glans (Fig. 1). Biopsy taken from the glans demonstrated pseudoepitheliomatous hyperplasia, intraepidermal microabscess formation, and focal subepidermal vesiculation overlying brisk inflammation including neutrophils, plasma cells, eosinophils, and lymphocytes (Fig. 2). Infectious workup revealed MRSA on tissue culture (treated without clinical improvement), negative HSV PCR, and negative special stains and tissue cultures for mycobacterial and fungal species. Further laboratory evaluation included rapid plasma reagin (RPR), antineutrophilic cytoplasmic autoantibody (ANCA), Hepatitis A/B/C, HIV, Lyme, antineutrophilic antibody (ANA), and tuberculosis interferon-γ release assay (IGRA)—all were unremarkable/negative. There was no clinical improvement with topical lidocaine, antibiotic ointment, testosterone, halobetasol cream, and courses of oral antibiotics and fluconazole.

Case 302: Supratentorial Lymphocytic Inflammation with Parenchymal Perivascular Enhancement Responsive to Steroids.

Part one of this case appeared 4 months previously and may contain larger images. A 21-year-old immunocompetent man who was a long-term resident of Qatar presented to the emergency department with recurrent episodes of unprovoked generalized tonic-clonic seizures lasting 2-3 minutes that spontaneously resolved and were associated with postictal confusion. The patient also had progressive mild diplopia, intermittent dizziness, and numbness in the left arm over the course of 3 months. The patient did not have any other systemic symptoms or chronic medical diseases. He did not have any history of intake of illicit drugs, supplements, or regular medications; he had not received any recent vaccinations; and he had not undergone any surgical procedures. He had no history of travel. At presentation, vital signs were normal. Neurologic examination showed mild left homonymous hemianopia, normal gait with no cerebellar signs, and preserved sensations, power, tone, and reflexes in all four limbs. An electroencephalogram showed no epileptiform discharges. Chest CT and extensive laboratory work-up, including viral, fungal, bacterial, and parasite work-up, thyroid function tests, and immunologic blood tests yielded normal results. Those included normal complete and differential blood counts and normal serum chemistry. Serum analysis was negative for antinuclear antibody, Sjögren syndrome antigens A and B, cytoplasmic antineutrophil cvtoplasmic antibody, and paraneoplastic profile. Serum evaluation was also negative for human immunodeficiency virus type 1 and type 2 RNA, and Brucella, Schistosoma, and toxoplasma antibodies. Venereal Disease Research Laboratory (VDRL) and rapid plasma regain (RPR) test results were negative. Cerebrospinal fluid (CSF) analysis revealed clear fluid and normal pressure and biochemistry, except for elevated protein concentration (0.48 g/L) (normal range, 0.15-0.45 g/L). There were 43 leukocytes/µL (99% lymphocytes) (normal range, 0-5 leukocytes/µL; lymphocytes range, 40%-80%), with no atypical or malignant cells. CSF Gram staining, acid-fast staining, cryptococcal antigen, varicella-zoster virus polymerase chain reaction (PCR), herpes simplex virus PCR, VDRL, and RPR test results were negative. CSF cultures did not show any evidence of growth of bacteria, fungi, or acid-fast bacillus. CSF flow cytometry did not show a monoclonal lymphoid population. No CSF oligoclonal bands were detected. Conventional brain MRI with intravenous administration of contrast material and perfusion study were performed and included different sequences (Figs 1-3).

Role of inflammation in the pulmonary hemorrhage induced by a snake venom metalloproteinase

Abstract Snakebite envenoming (a neglected tropical disease that affects millions of people worldwide every year) is characterized by a complex pathophysiology. In cases of viperid envenomings, local and systemic hemorrhage is one of the major systemic effects that manifest in patients. This work focuses on studying pulmonary hemorrhage caused by a type P-III snake venom metalloproteinase (SVMP) extracted from Crotalus simus venom, and the extent to which inflammation affects the outcome of this hemorrhage. In order to determine the role of inflammation in pulmonary hemorrhage, we created a new automated image analysis methodology using the CellProfiler platform to quantitatively assess hemorrhage under different conditions, in conjunction with a statistical analysis program to process and analyze said image analysis results. With this toolset, we subsequently analyzed hemorrhage under the effects of treatment with three anti-inflammatory agents (a COX inhibitor, a TNF-ɑ inhibitor, and anti-neutrophil antibodies) in combination with the toxin on a murine model. Our findings conclude that inhibiting inflammation reduced lung hemorrhage significantly, which shows promise as a potential novel avenue for the treatment of snake envenomings. Moreover, we propose new hypotheses for how the toxin may enter lung capillaries and subsequently cause basement membrane damage which eventually leads to lung hemorrhage. This study was supported by Vicerrectori?a de Investigación, Universidad de Costa Rica (Project 741-B4-524), and by a fund for support of graduate studies of the Sistema de Estudios de Posgrado (Universidad de Costa Rica)

Coeliac Disease Presenting as Chronic Neutropenia and Leukopenia in a 14‐Year‐Old

Coeliac disease (CD) is an immune-mediated multisystem disorder due to gluten hypersensitivity. Common findings of CD include chronic diarrhoea, weight loss, short stature, iron and folate deficiency and delayed puberty.1, 2 Leukopenia and neutropenia are rare presenting features of CD.3, 4 We report a 14-year-old girl with neutropenia probably caused by CD resolving on a gluten-free diet (GFD). A previously healthy 14-year-old girl presented to her general practitioner with lethargy. Investigations revealed a normal haemoglobin and platelets, leukopenia (WCC 3.2 × 109/L), neutropenia (1.33 × 109/L) and low ferritin (18 μg/L). She was commenced on iron supplements. Repeat full blood count (FBC) in 4 months showed persistent leukopenia and neutropenia prompting a referral to a paediatric clinic in regional New South Wales. Further evaluation in clinic found no history of a viral infection or medications and no family history of a haematological disorder. Physical examination was unremarkable with no lymphadenopathy or hepatosplenomegaly; and the weight and height were at 25th and 75th percentiles. Subsequent FBCs, 8 and 11 months later demonstrated persistent leukopenia (3.6 and 3.1 × 109/L) and neutropenia (1.2 and 1.2 × 109/L). Blood picture, vitamin B12, folate, EBV and CMV serology, anti-neutrophil antibodies, ferritin and copper levels were normal. A presumptive diagnosis of benign chronic neutropenia was made, and in view of previous low ferritin coeliac serology was performed. Serum tTG IgA (>100 U/mL), anti-deamidated Gliadin IgG (80 U), anti-endomysial IgA (positive) were high. A duodenal biopsy revealed prominent villous atrophy with blunting and multiple intraepithelial lymphocytes (more than 30 lymphocytes per 100 enterocytes) suggestive of Marsh 3b type of CD. She was commenced on a GFD. An FBC 6 months after commencement of GFD revealed normal leukocyte and neutrophil counts and the coeliac antibodies were negligible. The causes of chronic neutropenia in children include congenital, cyclic, viral, medications, autoimmune, B12, folate, copper and zinc deficiency, and benign chronic. Neutropenia and leukopenia are uncommon manifestations in CD with only a few cases reported in literature.3, 4 Neutropenia in CD could be caused by deficiency of B12, folate, zinc and copper due to malabsorption of nutrients or else it could be due to autoimmunity. In our patient, B12, folate and copper levels were normal and zinc levels were not done. CD is an immune-mediated disease. In the absence of a nutritional deficiency, we postulate that an autoimmune reaction targeting clones of neutrophils or its precursors is the most likely cause of isolated leukopenia and neutropenia in our patient.4 Our case highlights that coeliac serology could be an important second-line investigation of children with persistent neutropenia. It also reiterates the famous medical maxim: uncommon presentations of common disease are more common than common presentations of uncommon disease.

Pediatric Benign Neutropenia: Assessing Practice Preferences in Canada.

Pediatric benign neutropenia is a self-limited condition with a benign clinical course. An approach to this condition is not well-defined in the literature. Our objective was to use a case-based survey to elucidate trends in the diagnosis and management of benign neutropenia among pediatric hematology/oncology practitioners in Canada. We received 46 completed surveys (response rate 66%). At initial presentation with fever and neutropenia, 67% of respondents recommended partial septic workup but 11% recommended no investigations. Nearly 70% recommended admission for empiric intravenous antibiotics, while 24% would discharge home without antibiotics. In a patient with fever and known neutropenia, respondents were more likely to pursue outpatient antibiotic therapy. For investigation of chronic neutropenia, most respondents (60%) do not use antineutrophil antibody testing. Common indications for bone marrow biopsy were severe infection, prolonged neutropenia, or before initiating granulocyte colony stimulating factor. Indications for granulocyte colony stimulating factor were based on severity and frequency of infection. Most respondents (84%) would not recommend antibiotic prophylaxis. Results demonstrate the considerable variability in management of benign neutropenia among pediatric hematology/oncology practitioners in Canada and highlight the need for prospective studies to establish diagnostic criteria for benign neutropenia and evaluate management of fever in this population.

Vascular thromboses with retroperitoneal fibrosis: a case report

BackgroundGranulomatosis with polyangiitis is a systemic inflammatory disease characterized by necrotizing vasculitis that affects small- and medium-sized blood vessels. Granulomatous inflammation affects the lungs, ears, nose, and throat, and commonly affects the kidneys, although the retroperitoneal tissue is rarely affected. Several studies have reported an increased risk of venous thromboembolism. Early diagnosis and treatment are of vital importance due to the rapid progression of the disease.Case presentationWe present the case of a 66-year-old Moroccan man followed for bilateral jugular thrombosis. Cavitary pulmonary nodules and retroperitoneal fibrosis with thrombosis involving several vascular territories were detected on thoracoabdominopelvic computerized tomography scan. Laboratory analyses revealed that the patient was positive for cytoplasmic antineutrophilic antibodies. The diagnosis of granulomatosis with polyangiitis was retained. Treatment with glucocorticoids and immunosuppressive agents resulted in significant clinical and radiological improvement over the following months.ConclusionsWe describe the diagnostic steps and the difficulty of managing this patient. Rare manifestations, such as retroperitoneal fibrosis, have been reported in the literature in association with granulomatosis with polyangiitis, and should not delay the diagnosis and treatment of granulomatosis with polyangiitis owing to its severity.

Case 302.

History A 21-year-old immunocompetent man who was a long-term resident of Qatar presented to the emergency department with recurrent episodes of unprovoked generalized tonic-clonic seizures lasting 2-3 minutes that spontaneously resolved and were associated with postictal confusion. The patient also had progressive mild diplopia, intermittent dizziness, and numbness in the left arm over the course of 3 months. The patient did not have any other systemic symptoms or chronic medical diseases. He did not have any history of intake of illicit drugs, supplements, or regular medications; he had not received any recent vaccinations; and he had not undergone any surgical procedures. He had no history of travel. At presentation, vital signs were normal. Neurologic examination showed mild left homonymous hemianopia, normal gait with no cerebellar signs, and preserved sensations, power, tone, and reflexes in all four limbs. An electroencephalogram showed no epileptiform discharges. Chest CT and extensive laboratory work-up, including viral, fungal, bacterial, and parasite work-up, thyroid function tests, and immunologic blood tests yielded normal results. Those included normal complete and differential blood counts and normal serum chemistry. Serum analysis was negative for antinuclear antibody, Sjögren syndrome antigens A and B, cytoplasmic antineutrophil cvtoplasmic antibody, and paraneoplastic profile. Serum evaluation was also negative for human immunodeficiency virus type 1 and type 2 RNA, and Brucella, Schistosoma, and toxoplasma antibodies. Venereal Disease Research Laboratory (VDRL) and rapid plasma regain (RPR) test results were negative. Cerebrospinal fluid (CSF) analysis revealed clear fluid and normal pressure and biochemistry, except for elevated protein concentration (0.48 g/L) (normal range, 0.15-0.45 g/L). There were 43 leukocytes/µL (99% lymphocytes) (normal range, 0-5 leukocytes/µL; lymphocytes range, 40%-80%), with no atypical or malignant cells. CSF Gram staining, acid-fast staining, cryptococcal antigen, varicella-zoster virus polymerase chain reaction (PCR), herpes simplex virus PCR, VDRL, and RPR test results were negative. CSF cultures did not show any evidence of growth of bacteria, fungi, or acid-fast bacillus. CSF flow cytometry did not show a monoclonal lymphoid population. No CSF oligoclonal bands were detected. Conventional brain MRI with intravenous administration of contrast material and a perfusion study were performed and included different sequences (Figs 1-3).

Clinical Phenotype and Mechanisms of Leukopenia/Neutropenia in Patients with Primary Sjögren's Syndrome.

Sjögren’s syndrome (SS) is a chronic, systemic autoimmune disease which afflicts mainly the exocrine salivary and lachrymal glands, leading to mouth and eye dryness. However, any organ can be affected during the disease course, resulting in a variety of clinical manifestations. Sjögren’s syndrome clinical manifestations can be classified into glandular (sicca manifestations or parotid swelling), extra-glandular, either nonspecific (arthralgias, arthritis, Raynaud’s phenomenon, fatigue) or peri-epithelial (primary biliary cirrhosis, interstitial nephritis, bronchiolitis), and extra-epithelial (palpable glomerulonephritis, peripheral neuropathy, purpura). In addition, SS patients display high risk for B cell lymphomas due to chronic antigenic stimulation. Although disease pathogenesis remains unclear, genetic, environmental, and immunologic factors are implicated. In the context of systemic autoimmune manifestations, SS patients may also present with hematologic abnormalities including anaemia, leukopenia (mainly neutropenia or lymphopenia), and thrombocytopenia. Although leukopenia has been reported as a laboratory finding in many case series or cohorts of SS patients and in very few studies it has been proposed as an independent risk factor for lymphoma, the clinical phenotype of SS patients with leukopenia/neutropenia and the implicated pathogenetic mechanisms have not been elucidated. In the current study, we intend to analyse the clinical phenotype of leukopenic/neutropenic SS patients and explore the possible pathogenetic mechanisms by detecting anti-neutrophil antibodies and investigate the role of apoptotic pathways, especially the contribution of TRAIL pathway and the cFLIP molecule.

Induction and maintenance of remission with mycophenolate mofetil in ANCA-associated vasculitis: a systematic review and meta-analysis.

Uncertainties exist about the use of mycophenolate mofetil (MMF) in anti-neutrophil cytoplasmatic antibody (ANCA)-associated vasculitis (AAV), particularly for remission maintenance. Systematic review and meta-analysis of phase II and III trials assessing the use of MMF in AAV, granulomatosis with polyangiitis and microscopic polyangiitis (MPA). A comprehensive search of several databases (Medline, EMBASE, Cochrane, Web of Science, Scopus) from inception to 5 May 2020 has been conducted. Trial data were extracted to estimate odds ratios (ORs) and estimates (ES) for MMF efficacy (remission-induction and maintenance). Severe adverse effects (SAEs) were collected. From 565 articles captured, 10 met the predefined criteria, 5 phase II and 5 III trials; 4 assessed remission-induction, 3 remission maintenance and 3 both. The pooled OR for remission-induction at 6 months was 1.06 (95% confidence interval 0.74, 1.52), with no significant difference by subgroup meta-analysis of trials stratified by different study-level features (i.e. kidney disease, MPA, myeloperoxidase-ANCA positivity, newly diagnosed disease) (P&gt;0.05). The overall ES for remission maintenance at the end of follow-up ranged between 51% and 91% (I2=74.8%). Subgroup meta-analysis identified kidney involvement as a possible source of heterogeneity, yielding a significantly higher rate of sustained remission in trials enrolling only patients with kidney involvement (92%, 76-100%) versus those enrolling patients with and without kidney involvement (56%, 45-66%). Results were similar in multiple sensitivity analyses. During follow-up, the frequency of SAEs in MMF-based treatment arms was 31.8%. In AAV, MMF use was significantly associated with higher sustained remission rates in trials enrolling only patients with kidney involvement. These findings might influence clinical practice.

Patho-Mechanisms for Hemorrhage/Sepsis-Induced Indirect Acute Respiratory Distress Syndrome: A Role for Lung TIE1 and Its Regulation by Neutrophils.

Severe hemorrhage (Hem) has been shown to be causal for the development of extra-pulmonary/indirect acute respiratory distress syndrome (iARDS) and is associated with severe endothelial cell (EC) injury. EC growth factors, Angiopoietin (Ang)-1 and -2, maintain vascular homeostasis via tightly regulated competitive interaction with the tyrosine kinase receptor, Tie2, expressed on ECs. Since it has been reported that the orphan receptor, Tie1, may be able to play a role in Ang:Tie2 signaling; we chose to examine Tie1's capacity to alter the lung Ang:Tie2 interaction in response to the sequential insults of shock/sepsis (cecal ligation and puncture [CLP]), culminating in iARDS. Male mice were subjected to Hem alone or sequential Hem followed 24 hours later by CLP that induces iARDS. Changes in lung and/or plasma levels of Tie1, Tie2, Ang-1, Ang-2, various systemic cytokine/chemokines and indices of lung injury/inflammation were then determined. The role of Tie1 was established by intravenous administration of Tie1 specific or control siRNA at 1 h post-Hem. Alternatively, the contribution of neutrophils was assessed by pre-treating mice with anti-neutrophil antibody depletion 48 h prior to Hem. Lung tissue levels of Tie1 expression elevated over the first 6 to 24 h post-Hem alone. Subsequently, we found that treatment of Hem/CLP mice with Tie1-specific siRNA not only decreased Tie1 expression in lung tissue compared to control siRNA, but, suppressed the rise in lung inflammatory cytokines, lung MPO and the rise in lung protein leak. Finally, much as we have previously shown that neutrophil interaction with resident pulmonary vascular ECs contribute significantly to Ang-2 release and EC dysfunction, central to the development of iARDS. Here, we report that depletion of neutrophils also decreased lung tissue Tie1 expression and increased Tie2 activation in Hem/CLP mice. Together, these data imply that shock-induced increased expression of Tie1 can contribute to EC activation by inhibiting Ang:Tie2 interaction, culminating in EC dysfunction and the development of iARDS.

Neutrophils of diploid and triploid rainbow trouts assessed by flow cytometry

Triploid rainbow trouts Oncorhynchus mykiss are reared in Brazil, but there is no data about haematological differences when compared to diploid ones. Due to the lack of information, the goal was to evaluate the neutrophils of adult diploid and triploid rainbow trouts O. mykiss by flow cytometry screening using a monoclonal anti-neutrophil antibody (TTL-5E9). In comparison, the population of neutrophils was proportionally larger in diploid trouts (9.94 ± 4.64% vs. 5.58 ± 1.09, p = 0.0481), but with cells showing smaller size and less complexity than the neutrophils of triploid trouts. As sorted neutrophils were not evaluated on phagocytic activity or apoptosis induction, it was not possible to establish neutrophil activity differences between ploidies.

Late Onset and Long Lasting Idiopathic and Autoimmune Neutropenia As Epiphenomena of Immune Dysregulation: Preliminary Data Study from the Italian Neutropenia Registry

▪INTRODUCTION: Idiopathic neutropenia (IN) of childhood is a benign, self-limiting disorder usually occurring in the first 3 years of life that differs from primary autoimmune neutropenia (AIN) because anti neutrophil antibodies are not detected on repeated indirect testing over time(1). Chronic idiopathic neutropenia (CIN) is a well characterized disorder of elderly with a peculiar immunological pattern(2). Pediatric AIN patients, with atypical features represented by longer disease duration (Long Lasting) or diagnosed later (Late Onset), were recently shown by our group to display a different immune-hematological profile vs typical primary AIN(3). Atypical pediatric IN subjects, i.e. diagnosed later or with longer disease duration, though occasionally reported, were never systematically investigated. In the present study we analyzed a cohort of young patients with atypical IN and AIN, diagnosed after the age of 3 years or with longer disease duration, aiming to identify an immunological signature that might predict their different outcome from classical AIN and IN of infancy.PURPOSE OF THE STUDY: to analyze a cohort of patients affected with AIN and IN rising > 3 years of age and with duration >12 months or rising <3 years of age but persisting over 36 months.MATERIALS AND METHODS: Clinical, immunological and genetic data (NGS panel of 160 immunodeficiency/disimmunity genes) of eligible patients were collected from the database of the Italian Neutropenia Registry.RESULTS: From 2005 to 2020, data from 46 patients (24F, 24 autoimmune and 22 Idiopathic Neutropenia) were retrieved. Median age at onset was 11.2 years (IQR13.2-16.7), median follow-up was 4.3 years (IQR 3.2-6.8). Neither autoimmunity nor additional cytopenias to neutropenia were present at onset. Cumulative incidence at 5 years of autoimmune manifestations (thyroiditis, arthritis, vitiligo, recurrent skin rash) was 11.8% (CI 95% 4.3-30). Throughout the follow up, infections occurred in 32/46 (70%) subjects and in only 16% were severe (meningitis, recurrent pneumonia, sepsis and pericarditis). Infections sites were upper respiratory (in 84% of subjects mouth and gums (47%), and skin (40%) ear (25%), lung (19%) and urinary tract (12%). Fever of unknown origin was detected in 40% of patients. Recurrent infections (more than 3/y) involved mouth (85%) and ear (88%). White Blood Cell, Absolute Neutrophil and Lymphocyte counts retrieved at the beginning of the study, in the middle and at the end of follow up are shown in Table 1. Leucocytes and Lymphocytes at the end of follow up were significantly lower than values seen at diagnosis (p < 0.001) whereas neutrophil count remained stable (p=ns). One third of the cohort had lower values than normal of CD19+ and CD3-CD56+CD16+ cells. B switched memory CD27+/IgD-/IgM- (in 94%) were lower, while marginal zone B lymphocytes CD27+/IgD+/IgM+ (62%) and Tγδ cells (70%) were increased than the normal population according to a pattern comparable to that described in chronic idiopathic neutropenia (CIN). Immunoglobulin serum levels (IgA, IgG and IgM) were below the normal value in 7.5% of the population, but specularly a small subset (7.5%) of IN showed an increase of IgM was seen, similar to what already descibed in adult CIN. The genetic study carried out in 32 patients showed in 5 (16%) pathogenic variants: of immunodeficit/dysregulation (2 TACI, 1 TINF2, 1 CARD11) and in 9 cases (28%) VUS in genes of the same groups (1CARD11, 4CASP10,1DDX41/SOCS1, 1TSR2/DCLRE1c, 1PIK3D, 1TERT/TACI).CONCLUSIONS: Atypical IN and AIN (of longer duration or occurring in advanced pediatric age) are different from those appearing in early infancy. They seem to display dysimmune/autoimmune features that is some case is proven to rely on a genetic dysimmune background. They might represent an anticipation of more complex autoimmune disorders which may fully manifest later in life. Interstingly these patients seem to share some features with CIN of adults. Definitive conclusions will be drawn by applying more comprehensive genetic nalysis (WES and WGS) on larger group of subjects.