Specific Granule Deficiency Due To Novel Homozygote SMARCD2 Variant.

Background: Specific granule deficiency (SGD) is a rare immunodeficiency associated with CCAT/enhancer-binding protein epsilon (CEBPE) gene variants. It can cause severe recurrent infections and is lethal without successful stem cell transplantation. Few cases with SGD of both type 1 and type 2 have been described in the literature. In this study, we present the first report of a case with a novel homozygous c.511 C > T (p.Gln171Ter) mutation in the SMARCD2 gene of SGD type 2, which was successfully treated with bone marrow transplantation. Case: A male infant presented to our neonatal intensive care unit on the second day of life with an icteric appearance and mild hypotonia. He was evaluated for immunodeficiency as the cause of delayed cord separation and refractory neutropenia. At 6 weeks of age, SGD type 2 with a new variant was diagnosed and successfully treated by bone marrow transplantation. Conclusion: SGD is an immunodeficiency disease that is quite rare. However, we believe that SGD diagnosis and associated new variants can be detected more frequently with the widespread use of all whole-exome sequencing techniques.

Defective neutrophil development and specific granule deficiency caused by a homozygous splice-site mutation in SMARCD2.

BACKGROUND: SMARCD2 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily D, member 2) has recently been shown to have a critical role in granulopoiesis in humans, mice, and zebrafish. Our patient presented with delayed cord separation, failure to thrive, and sepsis. Retrospective whole-exome sequencing confirmed a homozygous splice-site mutation in SMARCD2. OBJECTIVE: We sought to provide the second description of human SMARCD2 deficiency and the first functional analysis of human primary SMARCD2-deficient cells. METHODS: Heparinized venous blood and bone marrow were collected from the patient after obtaining informed consent. Patient leukocytes and CD34+ cells were then isolated, phenotyped, and assessed functionally. RESULTS: Circulating neutrophils appeared phenotypically immature, lacking multilobed nuclei, and neutrophil granules lacked lactoferrin but showed normal levels of myeloperoxidase. Neutrophil oxidative burst was preserved in response to phorbol 12-myristate 13-acetate. Patient bone marrow-derived neutrophils and white blood cells showed a severely impaired chemotactic response. Furthermore, white blood cells showed defective in vitro killing of Staphylococcus aureus, consistent with a specific granule deficiency. Finally, patient bone marrow-derived CD34+ cells showed markedly impaired in vitro expansion and differentiation toward the neutrophil lineage. Before her molecular diagnosis, our patient underwent hematopoietic stem cell transplantation and is well 8 years later. CONCLUSIONS: This report highlights an important role for SMARCD2 in human myelopoiesis and the curative effect of hematopoietic stem cell transplantation for the hematopoietic features of SMARCD2 deficiency.

Coronavirus disease 2019 (COVID-19) related cytopenia: A protocol for systematic review and meta-analysis.

BACKGROUND: In December 2019, the novel coronavirus pneumonia was detected in Wuhan and named COVID-19. It is an international outbreak of the respiratory illness caused by severe acute respiratory syndrome coronavirus 2. Recent papers pointed out the cytopenia in COVID-19 patients including lymphopenia, neutrophilia, thrombocytopenia and lower level of hemoglobin had prognostic significance. This systemic review and meta-analysis summaries the latest evidence from available data and determine the hematological abnormality caused by severe acute respiratory syndrome coronavirus 2 and potential efficacy on the outcomes in patients with COVID-19. METHODS: This protocol for a systematic reviews and meta-analysis will be performed according to the preferred reporting items for systematic reviews and meta-analysis protocols 2015 guidelines. The database of Cochrane Library, PUBMED, EMBASE, Medline, Web of Science, Google Scholar, CNKI, WanFang, as well as gray literatures from the inception to present will be comprehensively and systematically searched without limitations of regions or language. The main study outcomes will be the mortality of COVID-19 patients. The meta-analysis was performed by RevMan V.5.3 program and Stata V.12.0 software after 2 reviewers independently selected literature, data extraction, bias risk evaluation and study quality assessment. Any disagreement will be resolved by consensus to the third researcher. RESULTS: This systematic review and meta-analysis may help provide clarify on the effect of cytopenia in patients with COVID-19. The result will be published at a peer-reviewed journal. CONCLUSIONS: This proposed study will evaluate the existing evidence on the effectiveness of cytopenia in COVID-19 patients. ETHIC AND DISSEMINATION: The content of this article does not involve moral approval or ethical review because no individual data will be collected. PROSPERO REGISTRATION: CRD42020187524.

Haematological manifestations of COVID-19: From cytopenia to coagulopathy.

Emerging data from the management of patients with coronavirus disease 2019 (COVID-19) suggests multi-systemic involvement, including the hemopoietic system. The haematological manifestations of COVID-19 include blood count anomalies notably lymphopenia and neutrophilia which are of prognostic significance. Hyperferritinemia and elevated lactate dehydrogenase have also been associated with increased mortality. Furthermore, there is considerable evidence of a distinct coagulopathy associated with COVID-19 characterised by elevated D-dimers and an increased risk of thrombotic events. This comprehensive review summarises the latest evidence from published studies and discusses the implications of the various haematological manifestations of COVID-19 with a view to guiding clinical management and risk stratification in this rapidly evolving pandemic.

Neutrophil Defects and Diagnosis Disorders of Neutrophil Function: An Overview.

Primary disorders of neutrophil function result from impairment in neutrophil responses that are critical for host defense. This chapter summarizes inherited disorders of neutrophils that cause defects in neutrophil adhesion, migration, and oxidative killing. These include the leukocyte adhesion deficiencies, actin defects and other disorders of chemotaxis, hyperimmunoglobulin E syndrome, Chédiak-Higashi Syndrome, neutrophil specific granule deficiency, chronic granulomatous disease, and myeloperoxidase deficiency. Diagnostic tests and treatment approaches are also summarized for each neutrophil disorder.

CEBPE-Mutant Specific Granule Deficiency Correlates With Aberrant Granule Organization and Substantial Proteome Alterations in Neutrophils.

Specific granule deficiency (SGD) is a rare disorder characterized by abnormal neutrophils evidenced by reduced granules, absence of granule proteins, and atypical bilobed nuclei. Mutations in CCAAT/enhancer-binding protein-ε (CEBPE) are one molecular etiology of the disease. Although C/EBPε has been studied extensively, the impact of CEBPE mutations on neutrophil biology remains elusive. Here, we identified two SGD patients bearing a previously described heterozygous mutation (p.Val218Ala) in CEBPE. We took this rare opportunity to characterize SGD neutrophils in terms of granule distribution and protein content. Granules of patient neutrophils were clustered and polarized, suggesting that not only absence of specific granules but also defects affecting other granules contribute to the phenotype. Our analysis showed that remaining granules displayed mixed protein content and lacked several glycoepitopes. To further elucidate the impact of mutant CEBPE, we performed detailed proteomic analysis of SGD neutrophils. Beside an absence of several granule proteins in patient cells, we observed increased expression of members of the linker of nucleoskeleton and cytoskeleton complex (nesprin-2, vimentin, and lamin-B2), which control nuclear shape. This suggests that absence of these proteins in healthy individuals might be responsible for segmented shapes of neutrophilic nuclei. We further show that the heterozygous mutation p.Val218Ala in CEBPE causes SGD through prevention of nuclear localization of the protein product. In conclusion, we uncover that absence of nuclear C/EBPε impacts on spatiotemporal expression and subsequent distribution of several granule proteins and further on expression of proteins controlling nuclear shape.

Comparison of clinical utility between neutrophil count and neutrophil-lymphocyte ratio in patients with ovarian cancer: a single institutional experience and a literature review.

OBJECTIVE: We retrospectively investigated the prognostic significance and clinical utility of pretreatment neutrophilia and elevated neutrophil-lymphocyte ratio (NLR) in patients with epithelial ovarian cancer. METHODS: Clinical data were collected from 344 surgically staged ovarian cancer patients between April 2007 and March 2016 and retrospectively reviewed. Neutrophilia and elevated NLR were defined as a neutrophil count ≥ 8,000/µl and an NLR ≥ 4.0, respectively. Univariate or multivariate analysis was conducted to evaluate the association between pretreatment neutrophilia or elevated NLR and clinicopathological characteristics, optimal surgery rate, progression-free survival (PFS) and disease-specific survival (DSS). Finally, we compared the clinical utility between neutrophil count and NLR by receiver operating characteristic (ROC) analysis. RESULTS: Pretreatment neutrophilia and elevated NLR were observed in 24 (7.0%) and 142 (41.3%) patients, respectively. In univariate analysis, both neutrophilia and elevated NLR were found to be associated with short PFS and DSS (p < 0.005). Multivariate analysis showed that neutrophilia and elevated NLR were predictors for shorter survival. In ROC analysis, the NLR tended to have a greater area under the ROC curve (AUC) value than the neutrophil count in predicting recurrence (0.7011 vs 0.6516, p = 0.0546) and had a significantly greater AUC value in predicting DSS (0.7249 vs 0.6379, p = 0.0182). Finally, based on the neutrophil count and NLR, we divided the patients into 3 prognostic groups-high-risk group (elevated NLR with neutrophilia), intermediate-risk group (elevated NLR without neutrophilia), and low-risk group (normal NLR), which allows for individualized and accurate survival estimates. CONCLUSIONS: Pretreatment neutrophilia and elevated NLR are independent poor prognostic factors in epithelial ovarian cancer patients. The NLR was superior to neutrophil count in predicting the survival of epithelial ovarian cancer patients.

Proposed diagnostic criteria and classification of basophilic leukemias and related disorders.

Basophils form a distinct cell lineage within the hematopoietic cell family. In various myeloid neoplasms, including chronic myeloid leukemia, basophilia is frequently seen. Acute and chronic basophilic leukemias, albeit rare, have also been described. However, no generally accepted criteria and classification of basophilic leukemias have been presented to date. To address this unmet need, a series of Working Conferences and other meetings were organized between March 2015 and March 2016. The current article provides a summary of consensus statements from these meetings, together with proposed criteria to delineate acute basophilic leukemia (ABL) from chronic basophilic leukemia (CBL) and primary forms of the disease where no preceding myeloid malignancy is detected, from the more common 'secondary' variants. Moreover, the term hyperbasophilia (HB) is proposed for cases with a persistent peripheral basophil count ⩾1000 per µl of blood. This condition, HB, is highly indicative of the presence of an underlying myeloid neoplasm. Therefore, HB is an important checkpoint in the diagnostic algorithm and requires a detailed hematologic investigation. In these patients, an underlying myeloid malignancy is often found and is then labeled with the appendix -baso, whereas primary cases of ABL or CBL are very rare. The criteria and classification proposed in this article should facilitate the diagnosis and management of patients with unexplained basophilia and basophil neoplasms in routine practice, and in clinical studies.

Neutrophilic dermatosis of the dorsal hands: a restrictive designation for an acral entity.

In 2000, Galaria et al. proposed the designation neutrophilic dermatosis of the dorsal hands (NDDH). The authors describe a case of NDDH with predominant involvement of the palmar aspect of the hands in a patient suffering from lung cancer, a possible paraneoplastic manifestation. Therefore, the term NDDH is not accurate because palmar manifestations of this dermatosis are also possible.

Role of the Leucine Zipper Domain of CCAAT/ Enhancer Binding Protein-Epsilon (C/EBPε) in Neutrophil-Specific Granule Deficiency.

Neutrophil-specific granule deficiency (SGD) is a rare autosomal recessive primary immunodeficiency characterized by bilobed neutrophil nuclei and lack of neutrophil-specific granule proteins such as lactoferrin. A deficiency of a myeloid-specific transcription factor, CCAAT/enhancer binding protein-epsilon (C/EBPε), has been identified as a cause of SGD. C/EBPε binds to DNA though its basic leucine zipper (bZIP) domain, and regulates terminal differentiation of neutrophils and expression of specific granule genes. Homozygous frameshift mutations resulting in loss of the bZIP domain have been reported in two patients with SGD. A recent observation showed that a homozygous 2-aa deletion in the bZIP domain with normal DNA-binding and dimerization abilities causes SGD by impairing protein-protein interactions with other transcription factors, indicating that multiple molecular mechanisms can lead to SGD. Studies of patient-derived mutations and analysis of C/EBPε knockout mice have shown the importance of the bZIP domain for the essential functions of C/EBPε.

Impaired glucose tolerance alters functional ability of peripheral blood-derived mononuclear cells in Asian Indian men.

AIM: To compare the adhesion, migration and endothelial differentiation potential of peripheral blood-derived mononuclear cells (PBMCs) obtained from drug-naive normal glucose tolerance (NGT) and impaired glucose tolerance (IGT) Asian Indian men. METHODS: Based on the 75-g oral glucose tolerance test, 30 NGT and 31 IGT subjects were recruited into the study. PBMCs were isolated from fasting blood using histopaque density gradient centrifugation. Isolated PBMCs were analysed for their ability to adhere to extracellular matrices, incorporation into tubular structures formed by matured endothelial cells and differentiation into endothelial cells upon 7-day culture in endothelial-specific growth medium. RESULTS: PBMCs obtained from IGT subjects exhibit poor adherence to fibronectin and reduced incorporation into tubular structures. Migration towards stromal cell-derived factor-1α (SDF-1α) in a trans-well filter assembly was also reduced for these cells. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) analysis revealed decreased expression of CXCR4 and ß2 integrin and increased expression of arginase II in IGT subjects. No differences were observed with regard to endothelial differentiation; however, cultured PBMCs of IGT subjects had decreased intracellular nitric oxide (NO) production. CONCLUSION: In pre-diabetic, Asian Indian men, PBMCs exhibit defective migration and homing potential.

γδT cells are prevalent in the proximal aorta and drive nascent atherosclerotic lesion progression and neutrophilia in hypercholesterolemic mice.

Unique innate immunity-linked γδT cells have been seen in early human artery lesions, but their role in lesion development has received little attention. Here we investigated whether γδT cells modulate atherogenesis in apolipoprotein E-deficient (ApoE KO) mice. We found that γδT cell numbers were markedly increased in the proximal aorta of ApoE-deficient vs. wild-type mice during early atherogenesis, particularly in the aortic root and arch, where they comprised most of the T cells and lesion progression is most rapid. γδT cells infiltrated intimal lesions in ApoE KO mice, but only the adventitia in wild-type mice, and were more prevalent than CD4+ T cells in early nascent lesions, as evaluated by en face confocal microscopy. These aortic γδT cells produced IL-17, but not IFN-γ, analyzed by ex vivo FACS. Furthermore, aortic arch lipid accumulation correlated strongly with abundance of IL-17-expressing splenic γδT cells in individual ApoE KO mice. To investigate the role of these γδT cells in early atherogenesis, we analyzed ApoE/γδT double knockout (DKO) compared to ApoE KO mice. We observed reduced early intimal lipid accumulation at sites of nascent lesion formation, both in chow-fed (by 40%) and Western diet-fed (by 44%) ApoE/γδT DKO mice. In addition, circulating neutrophils were drastically reduced in these DKO mice on Western diet, while expansion of inflammatory monocytes and splenic Th1 or Th17 lymphocytes was not affected. These data reveal, for the first time, a pathogenic role of γδT cells in early atherogenesis in ApoE KO mice, by mechanisms likely to involve their IL-17 production and induction of neutrophilia. Targeting γδT cells thus might offer therapeutic benefit in atherosclerosis or other inflammatory vascular diseases.

The role of leptin in the development of pulmonary neutrophilia in infection and acute lung injury.

OBJECTIVES: One of the hallmarks of severe pneumonia and associated acute lung injury is neutrophil recruitment to the lung. Leptin is thought to be up-regulated in the lung following injury and to exert diverse effects on leukocytes, influencing both chemotaxis and survival. We hypothesized that pulmonary leptin contributes directly to the development of pulmonary neutrophilia during pneumonia and acute lung injury. DESIGN: Controlled human and murine in vivo and ex vivo experimental studies. SETTING: Research laboratory of a university hospital. SUBJECTS: Healthy human volunteers and subjects hospitalized with bacterial and H1N1 pneumonia. C57Bl/6 and db/db mice were also used. INTERVENTIONS: Lung samples from patients and mice with either bacterial or H1N1 pneumonia and associated acute lung injury were immunostained for leptin. Human bronchoalveolar lavage samples obtained after lipopolysaccharide-induced lung injury were assayed for leptin. C57Bl/6 mice were examined after oropharyngeal aspiration of recombinant leptin alone or in combination with Escherichia coli- or Klebsiella pneumoniae-induced pneumonia. Leptin-resistant (db/db) mice were also examined using the E. coli model. Bronchoalveolar lavage neutrophilia and cytokine levels were measured. Leptin-induced chemotaxis was examined in human blood- and murine marrow-derived neutrophils in vitro. MEASUREMENTS AND MAIN RESULTS: Injured human and murine lung tissue showed leptin induction compared to normal lung, as did human bronchoalveolar lavage following lipopolysaccharide instillation. Bronchoalveolar lavage neutrophilia in uninjured and infected mice was increased and lung bacterial load decreased by airway leptin administration, whereas bronchoalveolar lavage neutrophilia in infected leptin-resistant mice was decreased. In sterile lung injury by lipopolysaccharide, leptin also appeared to decrease airspace neutrophil apoptosis. Both human and murine neutrophils migrated toward leptin in vitro, and this required intact signaling through the Janus Kinase 2/phosphatidylinositol-4,5-bisphosphate 3-kinase pathway. CONCLUSIONS: We demonstrate that pulmonary leptin is induced in injured human and murine lungs and that this cytokine is effective in driving alveolar airspace neutrophilia. This action appears to be caused by direct effects of leptin on neutrophils.

Crohn's disease presenting as a neutrophilic dermatosis in a 5-month-old boy.

A 5-month-old boy with a previous history of failure to thrive and poor feeding was admitted to the hospital with failure to thrive, oral ulcers, and a generalized vesiculopustular rash that demonstrated a subcorneal pustule and neutrophilic infiltrate on histology. Esophagogastroduodenoscopy and flexible sigmoidoscopy biopsies demonstrated chronic active colitis with granulomas, consistent with the diagnosis of Crohn's disease. Our case represents, to our knowledge, the youngest person reported with this condition in association with Crohn's disease.

Animal models of human granulocyte diseases.

In vivo animal models have proven very useful to the understanding of basic biologic pathways of the immune system, a prerequisite for the development of innovate therapies. This article addresses currently available models for defined human monogenetic defects of neutrophil granulocytes, including murine, zebrafish, and larger mammalian species. Strengths and weaknesses of each system are summarized, and clinical investigators may thus be inspired to develop further lines of research to improve diagnosis and therapy by use of the appropriate animal model system.

The molecular basis of leukocyte recruitment and its deficiencies.

The innate immune system responds to inflammation, infection and injury by recruiting neutrophils and other leukocytes. These cells are able to leave the intravascular compartment in a process called leukocyte recruitment. This process involves several distinct steps: selectin-mediated rolling, firm adhesion via integrins, postarrest modifications including adhesion strengthening and leukocyte crawling and finally transmigration into tissue. Genetic defects affecting the different steps of the cascade can result in severe impairment in leukocyte recruitment. So far, three leukocyte adhesion deficiencies (LAD I-III) have been described in humans. These LADs are rare autosomal recessive inherited disorders and, although clinically distinct, exhibit several common features including recurrent bacterial infections and leukocytosis. In LAD-I, mutations within the ß2-integrin gene result in a severe defect in ß2 integrin-mediated firm leukocyte adhesion. Defects in the posttranslational fucosylation of selectin ligands dramatically reduce leukocyte rolling and lead to LAD-II. Finally, LAD-III, also known as LAD-I variant, is caused by impaired integrin activation due to mutations within the kindlin-3 gene. This review provides an overview on the molecular basis of leukocyte adhesion and its deficiencies.