Understanding neutropenia secondary to intrinsic or iatrogenic immune dysregulation.

As a key member of the innate and adaptive immune response, neutrophils provide insights into the hematopoietic and inflammatory manifestations of inborn errors of immunity (IEI) and the consequences of immunotherapy. The facile recognition of IEI presenting with neutropenia provides an avenue for hematologists to facilitate early diagnosis and expedite biologically rationale care. Moreover, enhancing the understanding of the molecular mechanisms driving neutropenia in IEI-decreased bone marrow reserves, diminished egress from the bone marrow, and decreased survival-offers an opportunity to further dissect the pathophysiology driving neutropenia secondary to iatrogenic immune dysregulation, eg, immune checkpoint inhibitors and chimeric antigen receptor T-cell therapy.

Congenital Hypopigmentary Disorders with Multiorgan Impairment: A Case Report and an Overview on Gray Hair Syndromes.

The term congenital hypopigmentary disorders refers to a wide group of heterogeneous hereditary diseases, clinically characterized by inborn pigmentary defects of the iris, hair, and/or skin. They include Gray Hair Syndromes (GHSs), a rare group of autosomal recessive genodermatosis hallmarked by inborn silvery gray hair. GHSs encompass Griscelli, Chediak⁻Higashi, Elejalde, and Cross syndromes, which are all characterized by a broad spectrum of severe multisystem disorders, including neurological, ocular, skeletal, and immune system impairment. In this manuscript, we describe in detail the clinical, trichoscopic, and genetic features of a rare case of Griscelli syndrome; moreover, we provide an overview of all the GHSs known to date. Our report highlights how an accurate clinical examination with noninvasive methods, like trichoscopy, may play a crucial rule in diagnosis of rare and potentially lethal genetic syndromes such as Griscelli syndrome, in which timely diagnosis and therapy may modify the clinical course, quality of life, and likelihood of survival.

Morphological and functional analysis of beige (Chèdiak-Higashi syndrome) mouse mast cells with giant granules.

Chèdiak-Higashi syndrome is a rare autosomal recessive disease that causes hypopigmentation, recurrent infections, mild coagulation defects and neurological problems. Beige mice carry a mutation in the lysosome trafficking regulator (LYST) gene and display some of the key characteristics of human Chèdiak-Higashi syndrome, in particular, a high susceptibility to infection due to aberrant natural killer (NK) cell and polymorphonuclear leucocyte function. Morphological analysis of beige mice reveals the presence of enlarged lysosomes in a variety of cell types, including leucocytes, hepatocytes, fibroblasts and renal tubule cells. To examine the process of granule maturation and degranulation in beige mice mast cells, morphological studies have been conducted using a combination of electrophysiological techniques; however, few functional studies have been conducted with mast cells, such as mediator release. The aim of the present study was to determine the morphological and functional characteristics of skin and peritoneal mast cells and bone marrow-derived mast cells of homozygous (bg/bg) and heterozygous (bg/+) beige mice and wild-type (+/+) mice. The histamine concentration was lower in the peritoneal and bone marrow-derived mast cells of bg/bg mice compared with those of bg/+ and +/+ mice, but the histamine release response was potentiated. In vivo studies of passive cutaneous anaphylaxis showed no differences between bg/bg mice and either bg/+ or +/+ mice. Although bg/bg mast cells with enlarged granules display specific exocytotic processes in vitro, the consequences of mast cell activation in beige mice were similar to those of wild-type mice in vivo.

An actin cytoskeletal barrier inhibits lytic granule release from natural killer cells in patients with Chediak-Higashi syndrome.

BACKGROUND: Chediak-Higashi syndrome (CHS) is a rare disorder caused by biallelic mutations in the lysosomal trafficking regulator gene (LYST), resulting in formation of giant lysosomes or lysosome-related organelles in several cell types. The disease is characterized by immunodeficiency and a fatal hemophagocytic lymphohistiocytosis caused by impaired function of cytotoxic lymphocytes, including natural killer (NK) cells. OBJECTIVE: We sought to determine the underlying biochemical cause of the impaired cytotoxicity of NK cells in patients with CHS. METHODS: We generated a human cell model of CHS using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) technology. We used a combination of classical techniques to evaluate lysosomal function and cell activity in the model system and super-resolution microscopy to visualize F-actin and lytic granules in normal and LYST-deficient NK cells. RESULTS: Loss of LYST function in a human NK cell line, NK92mi, resulted in inhibition of NK cell cytotoxicity and reproduced other aspects of the CHS cellular phenotype, including the presence of significantly enlarged lytic granules with defective exocytosis and impaired integrity of endolysosomal compartments. The large granules had an acidic pH and normal activity of lysosomal enzymes and were positive for the proteins essential for lytic granule exocytosis. Visualization of the actin meshwork openings at the immunologic synapse revealed that the cortical actin acts as a barrier for secretion of such large granules at the cell-cell contact site. Decreasing the cortical actin density at the immunologic synapse or decreasing the lytic granule size restored the ability of LYST-deficient NK cells to degranulate and kill target cells. CONCLUSION: The cortical actin and granule size play significant roles in NK cell cytotoxic function. We present evidence that the periodicity of subsynaptic actin is an important factor limiting the release of large lytic granules from NK cells from patients with CHS and could be a novel target for pharmaceutical intervention.

NK cell effector functions in a Chédiak-Higashi patient undergoing cord blood transplantation: Effects of in vitro treatment with IL-2.

NK cell cytotoxicity in Chédiak-Higashi syndrome (CHS) is strongly impaired as lytic granules are not released upon NK-target cell contact, contributing to several defects typical of this severe immunodeficiency. Correction of NK cell defects in CHS should improve the outcome of hematopoietic stem-cell transplantation, proposed as therapy. We investigated NK cell functions in a CHS patient before and after cord-blood transplantation, and the ability of in vitro IL-2 treatment to restore them. Before the transplant, the strong defect in NK cell-mediated natural and antibody-dependent cytotoxicity, as well as in IFN-γ production, could be restored up to normal levels by in vitro IL-2 treatment. This cytokine also caused the appearance of smaller lysosomal granules and their orientation towards the NK-target cell contact area, thus suggesting that IL-2 had a more general capacity to restore NK cell effector functions. Moreover after the transplant, although the successful engraftment, NK cell cytotoxicity resulted still partially impaired at one year, almost normal at ten years and, anyhow, fully recovered by in vitro IL-2 treatment. Taken together, our results indicate that IL-2 had a wide capacity to restore NK cell effector functions, being able to reverse the altered cytotoxic activity, lytic granule pattern, and cytokine production observed in the CHS patient.

Neutrophil functional disorder in childhood.

Neutrophil functional disorders thought to be uncommon, yet important as a cause of morbidity and mortality in infants and children. During the first years of life, when the immune system is still not completely mature, when the viral infections are frequent and antibiotic overuse can damage and alter the immune response, the inadequate nutrition followed with iron deficient anemia and malnutrition can lead the child`s organism in state of immunodeficiency. Sometimes is difficult to distinguish at the beginning weather the cause of patient suffering from frequent infections is existing of primary immunodeficiency disorder or the cause of the immunodeficiency state is just from exogenous factors. Fortunately, primary immune deficiencies are rare diseases and only 6-7% of all of them, due to the neutrophilic functional disorders. Unfortunately, many exogenous and environmental factors have influence to the immune system, and the percentage of secondary caused neutrophilic functional disorders is much higher and should be considered when children are investigated for immunodeficiency. So, when to suspect neutrophil functional disorder? The hallmarks for diseases related to the neutrophilic functional disorders are discussed in this article.

Skin fibroblasts from individuals with Chediak-Higashi Syndrome (CHS) exhibit hyposensitive immunogenic response.

BACKGROUND: Chediak-Higashi Syndrome (CHS) is a rare autosomal recessive disease characterized by immunodeficiency, oculocutaneous albinism, neurological dysfunction, and early death. Individuals with CHS present with increased susceptibility to infections of the skin, upper-respiratory tract, gastrointestinal tract, and oral tissues. Classical CHS is caused by mutations in the gene encoding lysosomal trafficking regulator (LYST). Although defects in cytotoxic T cell lytic secretory granule secretion and neutrophil phagocytosis are suggested to contribute to the immunodeficiency in CHS, the underlying molecular mechanisms are unknown. We hypothesized that skin fibroblasts from CHS subjects exhibit impaired immune response due to defective trafficking of inflammatory factors. METHODS AND RESULTS: Primary skin fibroblasts from CHS subjects or healthy controls were assessed for genes encoding inflammatory response factors using PCR array. At baseline, we found CD14, IL1R1 and TLR-1 were down-regulated significantly (≥2 fold change) and the genes encoding TLR-3, IL-1ß and IL-6 were up-regulated in CHS cells compared to control cells. When challenged with E. coli lipopolysaccharide (LPS), CHS cells were less responsive than control cells, with only 8 genes significantly up-regulated (3-68 fold change) compared to baseline values, whereas 28 genes in control cells were significantly up-regulated at a much higher magnitude (3-4,629 fold change). In addition, 50% of the genes significantly up-regulated in LPS-treated control cells were significantly lower in LPS-treated CHS cells. IL-6, a fibroblast-derived proinflammatory cytokine essential for fighting infections was significantly lower in culture media of CHS cells with or without LPS. Furthermore, Western blot and immunofluorescent staining revealed that TLR-2 and TLR-4 were diminished on cell membranes of CHS cells and dissociated from Rab11a. CONCLUSIONS: For the first time, results from our study indicate defective trafficking of TLR-2 and TLR-4 contributes to the hyposensitive response of CHS skin fibroblasts to immunogenic challenge, providing a potential therapeutic target for clinical intervention in CHS.

[Screening for cytotoxic defects with flow cytometric detection of CD107α on natural killer cells and cytotoxic lymphocyte cells].

OBJECTIVE: To establish a novel flow cytometry-based assay for measuring the expression of lysosomal-associated membrane protein 1 (LAMP-1, CD107α) on the cell surface of natural killer (NK) cells and cytotoxic T lymphocyte (CTL) and evaluate the screening value of this assay for cytotoxic defects-related diseases such as familial hemophagocytic lymphopro-liferative (FHL) syndrome. METHOD: Three suspected Chediak-Higashi Syndrome (CHS) patients, three suspected FHL patients and 10 healthy children were enrolled in the study from October 2010 to June 2011. Their PBMCs were separated and activated overnight with IL-2. After the granule release of NK cells activated by phytohemagglutinin (PHA) and CD8+T cells by anti-CD3, the CD107α expression were analyzed by flow cytometry. The peripheral blood DNA and RNA of the patients were extracted to analyze the pathogenic genes via DNA-PCR/RT-PCR and direct sequencing. RESULT: The CD107α expression on CTL in the ten healthy children significantly increased after activation by anti-CD3 [(0.18 ± 0.07)% vs. (4.47 ± 2.36)%, P < 0.05] and NK cells after activation by PHA [(0.27 ± 0.07)% vs. (5.80 ± 2.83)%, P < 0.05]. The frequency of CD107α-expression NK cells in three suspected CHS after activation was significantly elevated when compared with the healthy control [0.5%, 0.6% vs. (5.80 ± 2.83)%] except patient 2. After the anti-CD3 activation, the frequency of CD107α expression on CTL cells also showed no significant difference [0.3%, 0.9%, 0.2% vs. (4.47 ± 2.36)%] in three patients. All of their mean fluorescence intensity (MFI) showed the same trend. Patient 1 and 3 were identified to have LYST mutations (Patient 1: c.5411-5414 del TTTC, L1741fsX1758 and c.7975 C > T, R2596X; Patient 3: c.4863G > A, R1563H and c.5392-5393delAA, E1739fsX1756). There was no mutation identified in the LYST gene for patient 2. CD107α expression of NK cells and CTL in the suspected FHL patients and in mirror of these findings, no underlying gene variation of PRF, MUNC13-4 and STX11 were identified. CONCLUSION: We developed a method to quantitatively assess cytotoxicity of the NK cells and CTL by measuring the expression of CD107α on the cell membrane, which appeared to be an effective and rapid screening test for cytotoxic defects-related diseases such as FHL and other HLH secondary to primary immunodeficiency.

Unrelated cord blood transplantation can restore hematologic and immunologic functions in patients with Chediak-Higashi syndrome.

CHS is a rare hereditary fatal disease, if not treated. APs occur in 85% of patients and are usually the main cause of mortality, and HSCT from HLA-matched related and unrelated donors is the only effective treatment for CHS and prevents recurrences of APs. We reviewed the records of three patients with CHS who underwent UCBT at KHCC. Records were examined for clinical features at the time of UCBT, conditioning regimens, morbidities, and outcomes. Conditioning comprised BU, cyclophosphamide, horse ATG, and etoposide. All patients tolerated the conditioning well. Two patients are alive, one with mixed and the other with full donor chimerism; hematologic and immunologic defects of CHS have been corrected in both patients. They show no evidence of recurrences of APs and have normal growth and development. In patients with CHS who lack HLA-matched related and unrelated donors, UCBT is a suitable alternative source of stem cells to restore immunologic and hematologic functions and prevent AP relapses, even in mixed chimeric states. Long follow-up and close monitoring are essential to evaluate the long-term benefits of using UCBT in patients with CHS.

Familial hemophagocytic lymphohistiocytosis: a model for understanding the human machinery of cellular cytotoxicity.

Cytotoxic T lymphocytes, natural killer cells, and NKT cells are effector cells able to kill infected cells. In some inherited human disorders, a defect in selected proteins involved in the cellular cytotoxicity mechanism results in specific clinical syndromes, grouped under the name of familial hemophagocytic lymphohistiocytosis. Recent advances in genetic studies of these patients has allowed the identification of different genetic subsets. Additional genetic immune deficiencies may also induce a similar clinical picture. International cooperation and prospective trials resulted in refining the diagnostic and therapeutic approach to these rare diseases with improved outcome but also with improved knowledge of the mechanisms underlying granule-mediated cellular cytotoxicity in humans.

[Secretory lysosome disorders in the immune synapse and other tissues]. / Los trastornos de la secreción lisosomal en la sinapsis inmune y otros tejidos.

INTRODUCTION: Haemophagocytic syndrome (HS) is a common manifestation of several congenital disorders characterised by a disruption of lysosomal secretion, interrupting the cytolytic pathway and triggering a dysfunction in the immune synapse. In this situation, the recognition of certain extra-immunological manifestations may help in the diagnostic process. PATIENTS AND METHODS: We describe the clinical and biological features present in two brothers with familial haemophagocytic lymphohistiocytosis type 3 (FHL-3), two patients with Griscelli syndrome type 2 (GS-2) and one patient with Chédiak-Higashi syndrome (CHS). RESULTS: Mutational assays at UNC13D were carried out on two brothers after diagnosing an early onset HS in the first one, yielding a positive result in both cases with a consequent diagnosis of FHL-3. The diagnosis of GS-2 was supported by positive results of mutational Rab27A studies in one patient with HS and abnormal pigmentation, and in her cousin who was affected by a similar abnormal pigmentation. The diagnosis of CHS was established in one patient with HS, abnormal pigmentation and atypical granules on cytological examination of a bone marrow smear. Diagnosis was confirmed in this patient by the finding of a homozygous LYST mutation. CONCLUSIONS: We point out the importance of recognising the presence of typical extra-immunological manifestations of certain congenital disorders of lysosome secretion in patients diagnosed with HS. The association of albinism and immunodeficiency has played a critical role in the recent identification of the molecular mechanism involved in these disorders.

Subtle differences in CTL cytotoxicity determine susceptibility to hemophagocytic lymphohistiocytosis in mice and humans with Chediak-Higashi syndrome.

Perforin-mediated cytotoxicity is important for controlling viral infections, but also for limiting immune reactions. Failure of this cytotoxic pathway leads to hemophagocytic lymphohistiocytosis (HLH), a life-threatening disorder of uncontrolled T-cell and macrophage activation. We studied susceptibility to HLH in 2 mouse strains (souris and beige(J)) and a cohort of patients with partial defects in perforin secretion resulting from different mutations in the LYST gene. Although both strains lacked NK-cell cytotoxicity, only souris mice developed all clinical and histopathologic signs of HLH after infection with lymphocytic choriomeningitis virus. The 2 strains showed subtle differences in CTL cytotoxicity in vitro that had a large impact on virus control in vivo. Whereas beige(J) CTLs eliminated lymphocytic choriomeningitis virus infection, souris CTLs failed to control the virus, which was associated with the development of HLH. In LYST-mutant patients with Chediak-Higashi syndrome, CTL cytotoxicity was reduced in patients with early-onset HLH, whereas it was retained in patients who later or never developed HLH. Thus, the risk of HLH development is set by a threshold that is determined by subtle differences in CTL cytotoxicity. Differences in the cytotoxic capacity of CTLs may be predictive for the risk of Chediak-Higashi syndrome patients to develop HLH.

Improvement of deficient natural killer activity and delayed bactericidal activity by a thiol proteinase inhibitor, E-64-d, in leukocytes from Chediak-Higashi syndrome patients in vitro.

We previously reported that administration of a potent calpain inhibitor, E-64-d, which protects protein kinase C (PKC) from proteolysis, in a mouse model of Chediak-Higashi syndrome (CHS) (beige mice), decreases its susceptibility to Staphylococcus aureus infection. In the present study, we examined the in vitro effect of E-64-d on both deficient natural killer (NK) and delayed bactericidal activities of leukocytes from six CHS patients. Our results showed that pretreatment of peripheral blood mononuclear cells (PBMCs) obtained from CHS patients with E-64-d (1 microg/ml) significantly enhanced NK activity against K562 cells. The delayed bactericidal activity of polymorphonuclear cells (PMNs) against S. aureus also showed marked improvement. This was recovered to almost normal levels when PMNs were pretreated with E-64-d (1 microg/ml). On the other hand, the same concentration of E-64-d did not affect either the NK or bactericidal activity of normal controls. In addition, we confirmed that following E-64-d treatment, the abnormal down-regulation of PKC activity after concanavalin A (Con A) stimulation was eliminated in PBMCs obtained from CHS patients. To examine whether PKC is involved in the NK cell-mediated cytolysis and bactericidal activity of PMNs, two potent PKC inhibitors, chelerythrin and GO6976, were used. We found that chelerythrin inhibits NK activity of normal PBMCs in a dose-dependent manner, and GO6976 inhibits NK activity at doses that inhibit Ca(2+)-dependent PKC isozymes. These inhibitors also suppressed the bactericidal activity of PMNs against S. aureus. Taken together, our findings suggested that E-64-d improved the compromised NK and bactericidal activity of leukocytes from CHS patients by reversing the down-regulation of PKC activity.

Familial and acquired hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening condition characterized by uncontrolled hyperinflammation on the basis of various inherited or acquired immune deficiencies. Cardinal symptoms are prolonged fever, hepatosplenomegaly and cytopenias. Central nervous system (CNS) symptoms are common. Biochemical markers include elevated triglyceride and ferritin, high levels of the alpha chain of the soluble interleukin-2 receptor and low fibrinogen. Impaired function of natural killer (NK) cells and cytotoxic T-cells (CTL) is a characteristic of all forms of HLH. Genetic HLH occurs in familial forms (FHLH), in which HLH is the primary and only manifestation, and in association with the immune deficiencies Chédiak-Higashi syndrome (CHS), Griscelli syndrome (GS) and X-linked lymphoproliferative syndrome (XLP), in which secondary HLH occurs sporadically. Most patients with acquired HLH have no known underlying immune deficiency. Both acquired and genetic forms are triggered by infections, mostly viruses, or other stimuli. HLH also occurs as a complication of rheumatic diseases (macrophage activation syndrome) and of malignancies. The recent discovery of several genetic defects causing FHLH as well as the identification of the genes responsible for CHS, GS and XLP have underscored the role of granule (perforin/granzymes)-mediated cytotoxicity in both the killing of infected cells and the termination of the immune response. The immediate aim of therapy is suppression of the increased inflammatory response by immunosuppressive/immunomodulatory agents and cytotoxic drugs. Genetic cases can only be cured with stem cell transplantation. Awareness of the clinical symptoms and of diagnostic criteria for HLH is crucial to starting life-saving therapy in time.

Herpesvirus saimiri-transformed CD8+ T cells as a tool to study Chediak-Higashi syndrome cytolytic lymphocytes.

Cytolytic CD8+ T lymphocytes are the main cell type involved in the fatal lymphoproliferative-accelerated phase of the Chediak-Higashi syndrome (CHS). To generate a cellular tool to study the defects of this T cell subset in vitro, we have used Herpesvirus saimiri, a lymphotropic virus that transforms human T lymphocytes into extended growth and in addition, endows them with natural killer (NK) features. Transformed CHS CD8+ T cells were generated and characterized in comparison with healthy controls. The results showed that transformed CHS T cells maintained the defects described in primary CHS lymphocytes, such as giant secretory lysosomes and impaired NK and T cell receptor/CD3-induced, perforin-mediated cytolytic activity [which, however, could be restored after extended culture in the presence of interleukin-2 (IL-2)]. Upon activation with phorbol ester plus calcium ionophore or upon extended culture with IL-2, transformed CHS T cells showed normal, perforin-independent plasma membrane CD178/CD95L/FasL-mediated cytolytic activity but negligible secretion of microvesicle-bound CD95L. Transformed (and primary) CHS T cells were otherwise normal for cytolysis-independent activation functions, such as proliferation, surface expression of several activation markers including major histocompatibility complex class II, and cytokine or surface activation-marker induction. Therefore, the CHS protein [CHS1/LYST (for lysosomal traffic regulator)] can be dispensable for certain NK and T cell cytolytic activities of activated CHS CD8+ T lymphocytes, but it seems to be required for microvesicle secretion of CD95L. We conclude that transformed CHS T cells may be useful as a tool to study in vitro the relative role of CHS1/LYST in NK and T lymphocyte cytolysis and antigen presentation.

Characterization of spontaneous dermatitis in beige rats with IgE hyperproduction.

BACKGROUND: Atopic dermatitis (AD) is a common skin disease characterized by chronic recurrent eczematous lesions, but its exact etiology and mechanism are unclear. We found that beige rats (DAbg/bg), a mutant model of Chediak-Higashi syndrome, develop skin lesions characterized by pruritus, excoriation, erosion and alopecia. We describe the beige rat and examine its possible usefulness as an AD model. METHODS: Beige rats of 4, 8, 13, 16, 26 and 52 weeks were used. Histological analysis of the skin was performed. Plasma IgE and cytokines were measured. Th1 and Th2 cytokines and RANTES mRNA expression of skin and lymph nodes were evaluated. Passive cutaneous anaphylaxis (PCA) reactions were examined, and maximization tests were conducted. RESULTS: Skin lesions begin to develop with increases in serum IgE levels and the expression of IL-4 mRNA in the lymph node and skin. Histologically, skin lesions are characterized by acanthosis, ulceration and inflammatory cell infiltration in the dermis. Inflammatory cells consist of CD3+, CD4+, ED1+, ED2+ and I-A+ mononuclear cells, eosinophils, degranulated mast cells and neutrophils accompanying interleukin (IL)-4, interferon (IFN)-gamma and RANTES mRNA expressions of the skin. Inflammatory cells are reduced during chronification with decreased expressions of IL-4, IFN-gamma and RANTES mRNA. In addition, the rats show a high sensitivity to PCA reactions and maximization tests. CONCLUSIONS: Our results show that some of the skin lesions of beige rats are morphologically similar to human AD, being characterized by inflammatory cell composition in the acute phase, and increased IgE and RANTES levels. However, the inflammatory process and cytokine expression pattern are different from those in human AD.

Establishment of a set of combined immunodeficient DA/Slc-Foxn1(rnu) Lyst(bg) congenic rat strains.

The congenitally athymic nude rat is used for studying cancer and transplantation owing to its hairlessness and T-cell defective function caused by the Foxn1(rnu) gene. However, NK cell activity of the nude rat is markedly increased. It is known that NK cells play a major role in rejection of xenografts and in cytotoxicity against tumor cells. Thus, the athymic nude rat with impaired NK cell activity should be a useful model for extensive studies. The DA-Lyst(bg)/Lyst(bg) rat, a model for human Chediak-Higashi syndrome (CHS) is characterized by diluted-coat color and impairment of NK cell activity. We planned to establish a combined immunodeficient double mutant rat introgressed with the Foxn1(rnu) and Lyst(bg) genes and a set of congenic strains having an identical genetic backgrounds simultaneously. Based on the phenotypic and genetic characteristics of the parental rat strains, the new strains were produced using continuous backcross and diagnosis with molecular genetic techniques. Each disease gene was diagnosed with PCR-RFLP or the long-nested PCR method. Furthermore, we used a marker-assisted congenic strategy based on scanning the genetic backgrounds of the parental rats with 461 rat microsatellite markers. We think that the newly established DA/Slc-Foxn1(rnu)/Foxn1(rnu) Lyst(bg)/Lyst(bg) double mutant will be useful as a severe disease model for human CHS, and the set of DA/Slc-Foxn1(rnu) Lyst(bg) congenic strains which have impaired NK cell activity and/or defective T cell function should be useful for studying in cancer research, xenotransplantation, immune function and other wide-ranging studies.

Lytic granules, secretory lysosomes and disease.

Lytic granules harbour many of the dangerous apoptosis-inducing molecules of the immune system, including perforin, granzymes and Fas ligand. Safe transport, storage and release of these lytic components is vital. As a secretory lysosome, the lytic granule is able to accomplish these roles, as well as conferring the lysosomal functions of cytotoxic T lymphocytes and natural killer cells. Secretory lysosomes are common to many other haemopoietic cells and also melanocytes. Many of the proteins used in lysosomal secretion are found in both melanocytes and hemopoietic cells, and are dysfunctional in genetic diseases with defects in these proteins. The genetically heterogeneous Hermansky-Pudlak syndrome represents an excellent model for revealing proteins involved in secretory lysosome functioning. However, studies of this disease reveal differences between the various different types of secretory lysosomes, including lytic granules.

Delivering the kiss of death.

Cytotoxic T lymphocytes and natural killer cells kill their targets by secreting specialized granules that contain potent cytotoxic molecules. Through the study of rare immunodeficiency diseases in which this granule pathway of killing is impaired, proteins such as Rab27a have been identified as components of the secretory machinery of these killer cells. Recent evidence suggests that the destruction of activated lymphocytes through granule-mediated killing may be an important mechanism of immunological homeostasis. Although the process by which this occurs is not yet known, it is possible that events taking place at the immunological synapse may render the killer cell susceptible to fratricidal attack by other killer cells.