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Secreted phospholipases A(2) (sPLA(2)s) are molecules released in plasma and biological fluids of patients with systemic inflammatory, autoimmune and allergic diseases. These molecules exert proinflammatory effects by either enzymatic-mechanisms or through binding to surface molecules expressed on inflammatory cells. sPLA(2)s are released at low levels in the normal airways and tend to increase during respiratory allergies (e.g., rhinitis and bronchial asthma) as the result of local secretion. Several sPLA(2) isoforms are expressed in the human lung and some of them (e.g., group IIA and group X) are released in the airways of patients with rhinitis or asthma. Mast cells play a major role in the pathogenesis of respiratory allergies and other chronic inflammatory lung diseases. Recent evidence indicates that mast cells purified from human lung express most of the sPLA(2) isoforms so far described. IgE-mediated activation of these cells induce the release of sPLA(2)s suggesting that mast cells are a main source of extracellular sPLA(2)s during allergic reactions. Once released, sPLA(2)s may contribute to the generation of eicosanoids (e.g., PGD(2) and LTC(4)) and to the release of preformed mediators (e.g., histamine) by an autocrine loop involving the interaction of sPLA(2)s with surface molecules such as heparan sulphate proteoglycans or the M-type receptor. Thus, mast cell-derived sPLA(2)s may play an important role in the initiation and amplification of the inflammatory reactions in patients with allergic rhinitis and bronchial asthma.

Angiogenesis and lymphangiogenesis are distinct and complex processes requiring a finely tuned balance between stimulatory and inhibitory signals. Immune and inflammatory cells can contribute to these processes by multiple mechanisms: directly by producing a broad array of angiogenic growth factors, and indirectly by secreting several cytokines, chemokines and other mediators able to coordinate the cell-cell interactions. Immune cells can stimulate or inhibit angiogenesis/lymphangiogenesis, depending on their activation status and subset specificity. We summarize recent findings reporting the expression and activity of angiogenic and lymphangiogenic factors and their receptors and coreceptors in immune cells. It is evident that modulation of angiogenesis and lymphangiogenesis by the innate and adaptive immune cells (mast cells, macrophages, dendritic cells, basophils, eosinophils, and some subsets of T cells) is a highly complex process not yet completely understood.

Background: Morbidity and mortality of primary and secondary antibody deficiencies (AD) are frequently associated with diagnostic delays. These could be avoided by a combination of factors including a widespread and effective development in screening tests. Methods: Calculated globulin (CG), derived from the difference between serum total protein and albumin levels, reflects immunoglobulin serum levels and has shown to have a predictive value in the early diagnosis of antibody deficiencies. This study investigated the possibility to use low levels of CG to detect antibody deficiency in an Italian University Hospital. Results: First, we conducted an analysis of anonymized adult samples collected at our biochemistry laboratory with a range of calculated globulin levels from 15 to 22 g/l. A CG cut-off of 19 g/l detected subjects with IgG lower than 600 mg/dl with a sensitivity of 70% and a specificity of 75%. To further verify the clinical usefulness of CG, we retrospectively evaluated the relationship between CG values and serum IgG levels in 38 patients diagnosed with CVID at our Institution. Using a CG cut-off of 19 g/l, we detected antibody deficiency in 97.3% (37/38) of the subjects present in our cohort. Conclusion: Finally, we chose a CG value of 19 g/l as the cut-off for a prospective AD screening program. The results of this study show that a screening CG test can be used as a tool to reduce diagnostic delays, improve long-term prognosis and reduce the healthcare costs of antibody deficiency.

Adiponectin (Acrp30) is an adipokine widely studied for its beneficial metabolic properties. It circulates as low molecular weight (LMW), medium molecular weight (MMW), and high molecular weight (HMW) oligomers. The latter exerts the most potent biological effects. Acrp30 attracted renewed interest with the finding that it was associated with the development and progression of immune disorders. The mechanisms underlying this association and the role of Acrp30 in the pathophysiology of immune-mediated conditions remain unknown. Common variable immunodeficiency (CVID) is a primary immunodeficiency characterized by chronic activation of the immune system, impaired antibody production, and imbalanced cytokine production. In the attempt to shed light on the expression of Acrp30 in CVID, we: (a) investigated total Acrp30 and its oligomerization state in CVID patients undergoing maintenance Ig replacement therapy; (b) assessed the effects of Ig replacement therapy on Acrp30 expression in treatment-naïve CVID patients, namely, patients not treated before diagnosis, before and after the first Ig administration; and (c) evaluated the correlation between Acrp30 levels and clinical phenotypes of the disease. As controls, we analyzed healthy subjects and patients affected by a non-immunodeficiency chronic inflammatory demyelinating polyneuropathy (CIDP), before and after Ig infusion. We found that total Acrp30 and HMW oligomers were decreased in CVID but not in CIDP patients versus controls. Moreover, Acrp30 levels were correlated with IgA levels and were associated with two CVID phenotypes, namely, autoimmune cytopenia and enteropathy. Receiver operating characteristic curve analysis indicated that Acrp30 modulation is specific for CVID patients. Acrp30 and HMW levels quickly and dramatically increased after Ig infusion only in eight treatment-naïve CVID patients but not in five CIDP patients. This finding indicates that Ig administration per se is not able to induce an increase of Acrp30, but the specific cellular and/or molecular background proper of CVID seems to be essential. In conclusion, our data indicate that Acrp30 is specifically related to CVID activity. Further studies are required to understand the biological role of Acrp30 and its possible use as disease biomarker in CVID.

Asthma is a heterogeneous disease characterized by chronic airway inflammation that results in a wide spectrum of clinical manifestations. Patients with severe asthma represent a substantial share of consumption of healthcare resources and hospitalization. Moreover, these patients are at risk of increased morbidity and mortality. Recently, several phenotypes and endotypes of asthma have been identified. The identification of specific subtypes of asthma is fundamental for optimizing the clinical benefit of novel treatments. Although in most patients the disease can be controlled by some combination of pharmacologic agents, in some 5-10% of patients the disease remains uncontrolled. Several monoclonal antibodies (mAbs) targeting pathogenetic molecules (e.g., IgE, IL-5, IL- 5Rα, IL-4, IL-13, TSLP) are currently available or under development for the treatment of different forms of severe type 2 asthma. The identification of diagnostic and predictive biomarkers (e.g., IgE, blood eosinophil count, FeNO, periostin, etc.) has revolutioned the field of targeted therapy in severe asthma. Monoclonal antibodies targeting Th2-driven inflammation are generally safe in adult patients with moderate-to-severe asthma. The long-term safety of these biologics is a relevant issue that should be addressed. Unfortunately, little is known about non-type 2 asthma. Further studies are needed to identify biomarkers to guide targeted therapies of different forms of non-type 2 asthma.

Immunoglobulin therapy is the administration of human polyvalent IgG and represents the most effective treatment to prevent recurrent infections in antibody deficiency patients. Primary antibody deficiency represents the main indication of immunoglobulin replacement therapy and includes a wide range of disorders characterized by impaired antibody production in response to pathogens and recurrent infections. However, not all primary antibody deficiency patients require immunoglobulin replacement. Indeed, immunoglobulin preparations are expensive and, once prescribed, usually result in lifelong therapy. Moreover, many patients significantly benefit from a long-term antibiotic prophylaxis and a prompt begin of antibiotic therapy in case of infectious events. Even more controversial is the decision to initiate immunoglobulin replacement therapy in secondary antibody deficiency, a heterogeneous and expanding group including B-cell lymphoproliferative syndromes, protein losing states and therapeutic agents. This review seeks to define the indication to immunoglobulin replacement in primary and secondary antibody deficiency disorders, distinguishing those in which the beginning of immunoglobulin therapy is always indicated at the same time as the diagnosis has been made, from those lacking of defined indication to replacement therapy. In addition, we propose a clinical approach, mainly based on the evaluation of infectious history, vaccine response and bronchiectasis finding, to support the decision to initiate immunoglobulin therapy in an individual patient.

Glucocorticoids and immunosuppressive drugs represent the first-line treatment of eosinophilic granulomatosis with polyangiitis (EGPA, former Churg-Strauss syndrome), even though the combined therapy is not successful in achieving the disease remission in some patients with neurological or cardiac involvement. We describe a case of an EGPA male patient with impaired left ventricular function not responsive to glucocorticoid and immunosuppressive therapy. We observed that high-dose (2g/kg/4weeks) intravenous immunoglobulin (IVIG) therapy significantly improved cardiac function, which was deteriorated after reducing IVIG dose at 0.5g/kg/4weeks, and was restored increasing again IVIG dose to 2g/kg/4weeks. The finding highlights the relevance of IVIG as treatment of choice in EGPA patients with cardiac involvement not responsive to the standard glucocorticoid and immunosuppressive therapy. Moreover, at a follow-up of 24months, the continuance of high dose IVIG therapy was required to maintain a sustained remission of the heart failure.

In this study, we compared intravenous immunoglobulins (IVIG) and subcutaneous immunoglobulins (SCIG) in terms of serum IgG concentration and incidence of infections in patients with hypogammaglobulinemia secondary to chemo-immunotherapy regimens including the anti-CD20 monoclonal antibody rituximab. Fourteen patients with a B-cell lymphoproliferative disease treated for at least six months with a rituximab-including chemo-immunotherapy regimen were recruited. Mean serum levels of IgG were higher during replacement therapy than at the end of rituximab treatment (p < 0.001). Moreover, serum IgG level was higher during replacement therapy with SCIG than with IVIG (p < 0.001). No differences in the incidence of infections were observed. Although the non-randomized design and the small number of patients do not allow definitive conclusions to be drawn, study results suggest that higher mean serum IgG levels are reached when using the subcutaneous route after a switch from the intravenous regimen, and that IVIG and SCIG offer comparable protection against infections.

Protein Fv, an endogenous protein produced in the liver, is released in biological fluids during viral hepatitis. Acute and chronic viral hepatitis can be associated with cardiovascular derangements. Protein Fv induced the release of histamine, tryptase and the de novo synthesis of prostaglandin D2 and cysteinyl leukotriene C4 from mast cells isolated from human heart tissue (HHMC). Protein Fv absorbed with protein A-Sepharose coated with polyclonal IgG did not induce histamine secretion. The maximal percent histamine secretion induced by protein Fv correlated (rs = 0.60; p < 0.05) with that induced by anti-IgE, whereas there was no correlation between the release caused by proteins Fv and C5a. Preincubation of HHMC with protein Fv or anti-IgE caused complete cross-desensitization to subsequent challenge with heterologous stimulus. HHMC from which IgE had been dissociated no longer released histamine in response to anti-IgE and protein Fv. A human monoclonal IgE blocked both anti-IgE- and protein Fv-induced release. Three human monoclonal IgM VH3+ inhibited protein-Fv-induced secretion of histamine from HHMC, whereas monoclonal IgM VH6+ did not inhibit the release induced by protein Fv. Protein Fv acts as an endogenous immunoglobulin superantigen by interacting with the VH3 domain of IgE to induce the release of mediators from HHMC.

Mast cells produce a wide spectrum of mediators and they have been implicated in several physiopathological conditions (e.g. allergic reactions and certain tumors). Pharmacologic agents that modulate the release of mediators from mast cells has helped to elucidate the biochemical mechanisms by which immunological and non-immunological stimuli activate these cells. Furthermore, the study of surface receptors and signaling pathways associated with mast cell activation revealed novel pharmacologic targets. Thus, the development of pharmacologic agents based on this new wave of knowledge holds promise for the treatment of several mast cell-mediated disorders.

To investigate the mechanisms of anaphylactoid reactions to radiocontrast media, in vitro mediator release induced by three iodinated contrast agents was examined using peripheral blood basophils and mast cells purified from human lung parenchyma, heart, and skin tissues. Three iodinated contrast agents, sodium and meglumine salts of ioxaglic acid, sodium and meglumine salts of ioxithalamic acid, and ioversol, were incubated with basophils purified from peripheral blood and human mast cells isolated and purified from different anatomical sites. Release of preformed (histamine and tryptase) and de novo synthesized mediators (prostaglandin D2 and leukotriene C4) into the supernatans was determined at various contrast medium concentrations after incubation for 60 min. Ioxaglate (0.2–0.3 M), ioxithalamate (0.3–0.5 M), and to a lesser extent ioversol (0.3–0.5 M) induced histamine release from basophils in a concentration-dependent manner. All three induced the release of preformed mediators (histamine and tryptase) from human lung, but not from skin mast cells. They also induced histamine and tryptase release from human heart mast cells. However, they did not induce the de novo synthesis of leukotriene C4 or prostaglandin D2 from human basophils or any type of mast cell examined. Cross-linking of IgE by anti-IgE induced the release of leukotriene C4 or prostaglandin D2 from human basophils or mast cells. Mannitol, an osmotic stimulus, induced the release of histamine from human basophils, but to a lesser extent from mast cells. These results show that different contrast media can differ in their ability to release mediators from enriched preparations of human basophils and mast cells. The three contrast agents examined act on basophils and mast cells as incomplete secretagogues, causing the release of preformed mediators, but not the de novo synthesis of chemical mediators. It may be useful to measure plasma tryptase levels to detect adverse reactions caused by iodinated radiographic contrast materials.

Oxatomide (OXA), a histamine H1 receptor antagonist, is effective in the treatment of patients with allergic rhinitis, some allergic skin disorders, and bronchial asthma. We have characterized the effect of OXA on the immunologic release of preformed (histamine and tryptase) and de novo synthesized mediators (leukotriene C4:LTC4 and prostaglandin D2:PGD2) from human basophils and mast cells purified (from 10 to 82%) from human lung parenchyma (HLMC) and skin tissue (HSMC). Preincubation (15 min, 37 degrees C) of basophils with OXA (10(-7)-10(-5) M) before Der p I antigen or anti-IgE challenge concentration-dependently (10-40%) inhibited the immunologic release of histamine and LTC4. OXA (10(-7)-10(-5) M) also inhibited (10-40%) histamine, tryptase and LTC4 release from HLMC activated by anti-IgE. In addition, OXA caused a concentration-dependent inhibition of histamine, tryptase and PGD2 release from HSMC immunologically challenged with a monoclonal antibody against the alpha chain of the high affinity receptor for IgE (anti-Fc epsilon RI) or anti-IgE. These results demonstrate that OXA exerts anti-inflammatory activities by inhibiting the release of preformed and de novo synthesized mediators from human basophils and mast cells.

Common variable immunodeficiency is the most common form of symptomatic primary antibody failure in adults and children. Replacement immunoglobulin is the standard treatment of these patients. By using a differential proteomic approach based on 2D-DIGE, we examined serum samples from normal donors and from matched, naive, and immunoglobulin-treated patients. The results highlighted regulated expression of serum proteins in naive patients. Among the identified proteins, clusterin/ApoJ serum levels were lower in naive patients, compared to normal subjects. This finding was validated in a wider collection of samples from newly enrolled patients. The establishment of a cellular system, based on a human hepatocyte cell line HuH7, allowed to ascertain a potential role in the regulation ofCLUgene expression by immunoglobulins.

Human heart mast cells (HHMC), by elaborating vasoactive mediators, cytokines and chemokines, are the main primary effector cells of anaphylaxis. Mast cells have been identified perivascularly, close to myocytes and in the arterial intima in human heart tissue. Mast cells isolated from human heart tissue (HHMC) of patients undergoing cardiac transplantation express high-affinity receptors for IgE (FcepsilonRI) and C5a receptors. Activation of HHMC in vitro with anti-IgE or anti-FcepsilonRI induced the release of preformed mediators (histamine, tryptase, chymase, and renin) and the de novo synthesis of LTC(4) (approximately =18 ng/l0(6) cells) and PGD(2) (approximately =18 ng/l0(6) cells). Complement is activated and anaphylatoxin forms during anaphylaxis. C5a causes rapid release of histamine and tryptase from HHMC. These cells are activated in vitro by therapeutic (general anesthetics, protamine, etc.) and diagnostic agents (radiocontrast media, etc.) which can cause anaphylactoid reactions. Low concentrations of histamine and cysteinyl leukotrienes given to subjects undergoing diagnostic catheterization caused significant systemic and coronary hemodynamic effects. These results indicate that HHMC probably have a role in anaphylactic reactions.

Angiogenesis is a multistep complex phenomenon critical for several inflammatory and neoplastic disorders. Basophils, normally confined to peripheral blood, can infiltrate the sites of chronic inflammation. In an attempt to obtain insights into the mechanism(s) underlying human basophil chemotaxis and its role in inflammation, we have characterized the expression and function of vascular endothelial growth factors (VEGFs) and their receptors in these cells. Basophils express mRNA for three isoforms of VEGF-A (121, 165, and 189) and two isoforms of VEGF-B (167 and 186). Peripheral blood and basophils in nasal polyps contain VEGF-A localized in secretory granules. The concentration of VEGF-A in basophils was 144.4 ± 10.8 pg/106 cells. Immunologic activation of basophils induced the release of VEGF-A. VEGF-A (10–500 ng/ml) induced basophil chemotaxis. Supernatants of activated basophils induced an angiogenic response in the chick embryo chorioallantoic membrane that was inhibited by an anti-VEGF-A Ab. The tyrosine kinase VEGFR-2 (VEGFR-2/KDR) mRNA was expressed in basophils. These cells also expressed mRNA for the soluble form of VEGFR-1 and neuropilin (NRP)1 and NRP2. Flow cytometric analysis indicated that basophils express epitopes recognized by mAbs against the extracellular domains of VEGFR-2, NRP1, and NRP2. Our data suggest that basophils could play a role in angiogenesis and inflammation through the expression of several forms of VEGF and their receptors.

Human basophils were first identified by Paul Ehrlich (1878) thanks to the metachromatic staining properties of their cytoplasmic granules [1, 2]. In all mammalian species analysed so far, basophils and mast cells are the only cells that synthesize histamine and express plasma membrane receptors that bind with high affinity the Fce portion of IgE (FceRI) [3]. Human basophils derive from precursors that originate in the bone marrow and in foetal liver and that circulate in peripheral blood. Under normal conditions, basophils are never found in human healthy tissues. Basophils differentiate and mature in the bone marrow and circulate in the blood with a prevalence of ≅ 0.5% of total leukocytes [4]. Under certain circumstances, basophils can be recruited into the inflamed tissue during specific IgEdependent reactions and in association with a variety of pathologic conditions [5, 6, 7, 8]. Increasing evidence suggests that basophils and their mediators are involved in delayed-type hypersensitivity reactions in the human skin and in the lung [5,6, 8, 9].

Human heart mast cells (HHMC) have been identified in heart tissue, perivascularly, and in the intima of coronary arteries. In vitro activation of isolated HHMC induces the release of vasoactive and proinflammatory mediators (histamine, tryptase, and cysteinyl leukotriene C4[LTC4]). We investigated the effects of several bacterial proteins on HHMC activation in vitro. HHMC released histamine, tryptase, and LTC4in response toStaphylococcus aureusCowan 1 and the immunoglobulin (Ig)-binding protein A, but not toS. aureusWood 46, which does not synthesize protein A. The effect of protein A was inhibited by preincubation with monoclonal IgM VH3+. Some strains ofPeptostreptococcus magnusexpress an Ig light chain-binding surface protein called protein L. Such bacteria and soluble protein L stimulated the release of preformed and newly synthesized mediators from HHMC. Preincubation of HHMC with either protein A or protein L resulted in complete cross-desensitization to a subsequent challenge with the heterologous stimulus or anti-IgE. Monoclonal IgE (κ chains) blocked protein L-induced release, whereas IgE (λ chains) had no effect. Streptococcal protein G, formyl-containing tripeptide, and pepstatin A did not activate HHMC. Bacterial products protein A and protein L and intact bacteria (S. aureusandP. magnus) activate HHMC by acting as Ig superantigens.

Publisher Summary Within human heart tissue, mast cells lie between myocytes and in close contact with blood vessels. Mast cells are also localized in the coronary adventitia and in the shoulder region of coronary atheroma. The density of cardiac mast cells is higher in patients with dilated and ischaemic cardiomyopathy than in subjects without cardiovascular diseases. Intriguingly, in some of these conditions there is in situ evidence of mast cell activation. Observations by different groups of investigators, suggest that HHMCs play complex and significant roles in various pathophysiological conditions that involve the cardiovascular system. Direct activation of HHMCs by circulating antigens, autoantibodies and therapeutic and diagnostic substances injected intravascularly can explain some of the anaphylactic/anaphylactoid reactions caused by these agents. HHMCs possess FcɛRI, C5a receptors and IgE on their surface, which could explain the direct involvement of cardiac mast cells in systemic and cardiac anaphylaxis. Of particular interest is the observation that the eosinophil cationic proteins ECP and MBP can activate HHMCs to release preformed and de novo synthesized mediators. This novel observation suggests that, in allergic patients with eosinophilia, eosinophilic infiltration and the deposition of eosinophil granule proteins might, through direct activation of HHMCs, contribute to the cardiac derangements observed in some of the patients.

In the current study, the potential blocking ability of K+ channels encoded by the human ether-a-go-go related gene (HERG) by the piperazine H1 receptor antagonist cetirizine has been examined and compared with that of other second-generation antihistamines (astemizole, terfenadine, and loratadine). Cetirizine was completely devoid of any inhibitory action on HERG K+ channels heterologously expressed in Xenopus laevis oocytes in concentrations up to 30 microM. On the other hand, terfenadine and astemizole effectively blocked HERG K+ channels with nanomolar affinities (the estimated IC50 values were 330 and 480 nM, respectively), whereas loratadine was approximately 300-fold less potent (IC50 approximately 100 microM). In addition, in contrast to terfenadine, cetirizine did not show use-dependent blockade. In SH-SY5Y cells, a human neuroblastoma clone that constitutively expresses K+ currents carried by HERG channels (IHERG), as well as in human embryonic kidney 293 cells stably transfected with HERG cDNA, extracellular perfusion with 3 microM cetirizine did not exert any inhibitory action on IHERG. Astemizole (3 microM), on the other hand, was highly effective. Terfenadine (3 microM) caused a marked (approximately 80%) inhibition of IHERG in SH-SY5Y cells, whereas loratadine, at the same concentration, caused a 40% blockade. Furthermore, the application of cetirizine (3 microM) on the intracellular side of the membrane of HERG-transfected human embryonic kidney 293 cells did not affect IHERG, whereas the same intracellular concentration of astemizole caused a complete block. The results of the current study suggest that second-generation antihistamines display marked differences in their ability to block HERG K+ channels. Cetirizine in particular, which possesses more polar and smaller substituent groups attached to the tertiary amine compared with other antihistamines, lacks HERG-blocking properties, possibly explaining the absence of torsade de pointes ventricular arrhythmias associated with its therapeutical use.

Background Loratadine, a novel histamine H1-receptor antagonist, is effective in the treatment of patients with seasonal and perennial rhinitis and some allergic skin disorders. Histamine and other chemical mediators are synthesized and immunologically released by human peripheral blood basophils and tissue mast cells (Fc epsilon RI+ cells). Objective To evaluate the effects of loratadine and its main metabolite, desethoxylcarbonyl-loratadine (des-loratadine), on the immunological release of preformed (histamine and tryptase) and de novo synthesized mediators (leukotriene C4: LTC4 and prostaglandin D2:PGD2) from human Fc epsilon RI+ cells. Methods Human Fc epsilon RI+ cells purified from peripheral blood and from skin (HSMC) and lung tissue (HLMC) were preincubated with loratadine and des-loratadine before immunological challenge with Der p 1 antigen or anti-Fc epsilon RI. The release of preformed mediators (histamine and tryptase) and de novo synthesized eicosanoids was evaluated in the supernatants of human Fc epsilon RI+ cells. Results Preincubation (15 min, 37 degrees C) of purified (36-74%) basophils with loratadine (3 x 10(-6)-10(-4)M) and des-loratadine before Der p 1 antigen or anti-Fc epsilon RI challenge concentration-dependently (5-40%) inhibited the release of histamine and LTC4. Loratadine (3 x 10(-6)-10(-4)M) and des-loratadine also inhibited (10-40%) histamine, LTC4, and PGD2 release from purified HLMC (16-68%) activated by anti-Fc epsilon RI. Loratadine (3 x 10(-6)-10(-4)M) and des-loratadine caused concentration-dependent inhibition (10-40%) of histamine, tryptase, LTC4, and PGD2 release from purified HSMC (24-72%) immunologically challenged with anti-Fc epsilon RI. Conclusion These results indicate that loratadine and its main metabolite have anti-inflammatory activity by inhibiting the release of preformed and de novo synthesized mediators from human Fc epsilon RI+ cells.

Eosinophilia in humans is associated with eosinophil infiltration and cardiac localization of eosinophil granule proteins. Eosinophil cationic proteins are responsible for cardiac disease in some patients with eosinophilia. We have investigated the in vitro effect of four eosinophil granule proteins: eosinophil cationic protein (ECP), major basic protein (MBP), eosinophil-derived neurotoxin (EDN) and eosinophil peroxidase (EPO), on mast cells isolated from human cardiac tissue (HHMC). ECP and, to a lesser extent MBP (0.3-3 microM), but not EDN and EPO, stimulated the release of histamine and tryptase from HHMC. This release reaction induced by ECP and MBP was Ca(2+)- and temperature-dependent and was abolished by preincubation with anti-ECP and anti-MBP, respectively. The activation of HHMC by ECP and MBP was abolished by preincubation with 2-deoxy-D-glucose and antimycin A. These data demonstrate that some eosinophil cationic proteins, ECP and MBP, are selective activators of HHMC, thus contributing to the cardiac lesions in patients with eosinophilia.

Hypokalemic myopathy may occur in several infections. We report a case of severe and transient myopathy secondary to hypokalemia induced by chronic intenstinal infection withGiardia lamblia in a patient with common variable hypogammaglobulinemia. Hypokalemic myopathy is documented by serum enzymes, electromyography (reduction in the number of voluntarily activated motor unit action potentials and an increase in polyphasic motor unit action potentials, and pathological changes (hematoxylin-eosin, ATPase staining). The case reported involves hypokalemic myopathy induced by giardiasis in a patient with primary immunodeficiency; the histopathological changes observed in a skin/muscle biopsy from this patient are described for the first time.

Mast cells are present in normal and even more abundant in diseased human heart tissue and their localization is of particular relevance to their function. Within heart tissue mast cells lie between myocytes and in close contact with blood vessels. They are also found in the coronary adventitia and in the shoulder regions of a coronary atheroma. The density of cardiac mast cells is markedly higher in some patients with myocarditis and dilated cardiomyopathy than in accident victims without cardiovascular diseases. More importantly, in some of these conditions there is in situ evidence of mast cell activation. We have described an original technique to isolate and purify HHMC for in vitro study. This procedure gives viable cells and after stimulation with immunological or non-immunological stimuli they release performed (histamine and tryptase) and newly generated mediators (PGD2 and LTC4). We have demonstrated that HHMC differ from those in other anatomical districts in that they are activated by specific immunological and non-immunological stimuli, and in their relation to the arachidonic acid metabolism, suggesting that the local microenvironment can influence their phenotypic and biochemical characteristics. Our own and other findings suggest that HHMC have complex and significant roles in different pathophysiological conditions involving the cardiovascular system. Direct activation of HHMC by therapeutic and diagnostic substances injected intravenously explains some of the anaphylactoid reactions caused by these agents. HHMC possess Fc epsilon RI and IgE bound to the surface and C5a receptors, which could explain how cardiac mast cells are involved in systemic and cardiac anaphylaxis. Cardiac mast cells and those in human coronary arteries also play a role in the early and late stages of atherogenesis and during ischemic myocardial injury. In conclusion, although studies of HHMC are in their infancy, their in vitro isolation may be useful in identifying additional mediators synthesized and released, stimuli relevant to human pathophysiology, and pharmacological agents selectively modulating the activation of these cells and their mediators. Drugs specifically acting on HHMC or on their mediators may eventually be useful in treating different cardiovascular diseases.

Mastocytosis is a disease characterized by an abnormal proliferation of tissue mast cells. The events primarily responsible for mast cell proliferation in mastocytosis are largely unknown, but a derangement of the network involving c-kit receptor and its natural ligand (stem cell factor, which promotes mast cell growth and differentiation in man) is likely to have a primary role in this disease. Mastocytosis comprises a wide spectrum of clinical conditions determined by the degree of mast cell proliferation, the organ systems involved, the age at onset and the association with hematologic diseases. Mastocytosis can occur in a pediatric or an adult form. In both groups of patients, the disease may be limited to the skin (cutaneous mastocytosis) or be systemic, involving predominantly the bone marrow and the gastrointestinal tract. The symptoms in patients with mastocytosis are generally related to the increased release of mast-cell-derived mediators, such as histamine, prostaglandin D2, peptide leukotrienes, platelet-activating factor, heparin and proteolytic enzymes. The measurement of these chemical mediators (histamine, tryptase and prostaglandin D2 and their metabolites) in body fluids is useful for the diagnosis and the laboratory evaluation of patients with systemic mastocytosis. As little is known about the pathogenesis of the different forms of mastocytosis, the treatment of the majority of these patients is largely symptomatic.

Background Angiogenesis and lymphangiogenesis are critical for several allergic, inflammatory, and neoplastic disorders. Mast cells infiltrate the sites of inflammation and tumors. Objective We sought to characterize the expression and functions of vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) in human mast cells. Methods VEGF expression was evaluated by means of RT-PCR and Western blotting in primary human lung mast cells and in the mast cell lines LAD-2 and HMC-1. Angiogenic activity of mast cell supernatants was determined by using the chick embryo chorioallantoic membrane assay. VEGFR expression was assessed by means of RT-PCR and flow cytometry. Modified Boyden chambers were used for chemotaxis assay. Results Human mast cells express VEGF-A, VEGF-B, VEGF-C, and VEGF-D at both the mRNA and protein level. Prostaglandin E 2 (PGE 2 ) enhanced the expression of VEGFA , VEGFB , and VEGFC , whereas an adenosine analog (5′-[N-ethylcarboxamido] adenosine [NECA]) increased VEGFA , VEGFC , and VEGFD expression. In addition, PGE 2 and NECA enhanced VEGF-A release, and supernatants of PGE 2 - and NECA-activated human lung mast cells induced angiogenic responses in the chorioallantoic membrane assay that were inhibited by an anti-VEGF-A antibody. Mast cells expressed mRNA for VEGFR1 and VEGFR2 . These receptors were present on the mast cell surface. VEGF-A 165 , VEGF-B 167 , VEGF-C, VEGF-D, and placental growth factor 1 induced mast cell chemotaxis. These chemotactic effects were mediated by the activation of both VEGFR-1 and VEGFR-2. Conclusion Our data indicate that human mast cells are both a source and a target of angiogenic and lymphangiogenic factors and therefore might play a role in inflammatory and neoplastic angiogenesis through the expression of several forms of VEGFs and their receptors.

Mast cells are present in the human heart [1], and coronary arteries of patients with ischemic heart disease contain more histamine and mast cells than do those of non cardiac patients [2]. Moreover, the in vitro immunologic activation of fragments of human right atrial appendages induces the release of histamine [1]. Human mast cells from lung, skin, gut, uterus, and synovia are immunologically and functionally heterogenous [3]. For instance, skin mast cells can be activated by a variety of secretagogues, e.g. substance P0 48/80, C5a, and morphine that do not activate other mast cell types [3]. The complex heterogeneity of human mast cells isolated from different organs suggests that certain functions of these cells are regulated by the local microenvironment. Therefore, it is important to develop techniques to retrieve mast cells from different anatomic sites. In this study we have identified some immunologic and non-immunologic stimuli that activate human heart mast cells (HHMC) and measured a number of preformed mediators released upon activation of these cells.

The collagen content (measured as myocardial concentration of hydroxyproline) and dry weight (expressed as ventricle weight to body weight ratio) were determined in the left ventricle of male Sprague-Dawley rats (200--220 g b.wt.) exposed to a simulated altitude of 7,000 m for 18 h a day for 10 days in a hypobaric chamber. Hypoxia resulted in a significant increase (P less than 0.001) in the mass of the left ventricle with a concomitant significantly increased collagen concentration (P less than 0.001). The data indicate that hypoxia effects the synthesis of a significant amount of connective tissue in the left ventricle, which is the ventricle not exposed to pressure load. These results may be related to clinical, hemodynamic, and pathologic observations showing the left ventricular dysfunction in patients with chronic respiratory insufficiency. Since the amount of collagen in the left ventricle might interfere with contractile function, it is suggested that the hypoxia in these patients could affect the left ventricular myocardium via a direct action on the connective metabolism.

Fenoterol, a β2-adrenergic agonist recently introduced to treat asthmatic disorders, inhibits antigen-induced histamine release from human basophil leukocytes and lung mast cells in a dose-dependent fashion. The dose-response inhibition curve is paralleled by a fenoterol-induced increase in the cAMP levels of human leukocyte preparations. The relationship between the effect of fenoterol and cAMP level is supported by the finding that the β2-adrenergic agonist only inhibits the first stage of antigen-induced histamine release and not the release caused by the Ca2+ ionophore, A23187. Propranolol, a competitive antagonist of β2-adrenergic receptor, blocks the inhibition of release and the cAMP accumulation caused by fenoterol. Finally, theophylline, a cAMP phosphodiesterase inhibitor, synergistically potentiates the inhibitory effect of fenoterol on histamine release and the accumulation of cAMP. These data suggest that fenoterol may modulate the in vivo release of the mediators of immediate hypersensitivity reactions via the activation of β2-adrenergic receptor linked to adenylate cyclase on human basophils and mast cells.

The authors performed a number of experiments aiming at quantifying the infarct size in Sprague-Dawley male rats. The experimental model employed was isoproterenol (ISP) induced, infarctlike myocardial lesions. In quantifying the extent of the infarct area they compared the cross reliability of enzymatic and histological methods. ISP was administered in two subcutaneous injections (50 mg/100 Gm body weight) at a 24 hour interval. At the peak of the necrotic process (48 hrs after the first injection), the heart was subjected to enzymatic or histological studies. Thus, a group of infarcted animals (n = 25) and their controls (n = 15) underwent the assessment of creatine kinase (CK) content in the homogenate of the myocardium. Another group of animals (n = 15) was used for histological observations, including measurement of infarct size by planimetry from histological sections. A good relationship was observed between the percentage of necrotic tissue calculated on the basis of CK values (69.11 +/- 2.39%) and the infarction area assessed by planimetry (72.37 +/- 2.69%) (mean +/- standard error of the mean [SEM]). Thus, the present study confirms that ISP-induced myocardial lesions are a simple and reliable model for experimental infarct. Moreover, the assessment of the CK content in the heart is correlated with histological studies by planimetry, and is suggested as a direct enzymatic method to quantify the extent of myocardial necrosis.

Two different conditions responsible for cardiac hypertrophy in the rat were investigated: the first one is isoproterenol-induced myocardial infarct, the second is exposure to hypoxia (0.42 atmospheres/24 h) in hypobaric chamber. To demonstrate that in both experimental models stimulation of protein synthesis is an absolute requirement to induce cardiac hypertrophy, a variety of techniques were employed including: evaluation of dry heart weight value, concentration of radiothallium (201T1) in the heart and effects of inhibitors of protein synthesis (Puromycin). Experimental results have showed: (A) a significant increase (p0.001 as compared to control group) of dry heart weight values both in isoproterenol-treated and hypoxic rats; (B) a significant increase over control group (p0.001) in myocardial 201T1 concentration in isoproterenol-treated rats; (C) total inhibition of cardiac hypertrophy in Puromycin treated group subjected to hypoxia. Finally, on the basis of different mortality observed in infarcted (85.0%) or hypoxic (5.0%) rats treated with Puromycin (40 mg/Kg body weight i.p.) a different role of cardiac hypertrophy in 2 experimental conditions is postulated: in the case of infarct-like lesions the cardiac hypertrophy has compensatory significance; under hypoxic stimulus is only due to increased cardiac work.

Summary. The number of erythroid burst- (BFU-E) and colony-forming units (CFU-E), as well as of myeloid-macrophage colony-forming units (CFU-C), has been evaluated in tibial marrow and spleen of ex-hypoxic polycythaemic mice, at sequential time intervals after the end of hypoxia. In both marrow and spleen, the kinetics of the CFU-E pool is characterized by a sharp fall from above normal to lower than normal values. BFU-E and CFU-C however rise from below normal to higher than normal levels. These results have been correlated with both the erythropoietin (Ep) and the erythropoietic activity curves. It is apparent that Ep levels largely control both the differentiation and the amplification of the CFU-E pool and it is suggested that Ep may act as a ‘survival factor’ at the CFU-E level and/or increase the flow of cells from BFU-E to CFU-E. The difference in response between CFU-E and BFU-E favours a clearcut distinction between these populations, whereas the similarity between the BFU-E and CFU-C response suggests a close relationship between these two cell populations. It is also of interest that the murine spleen functions as a large reservoir of erythroid microenvironment for hypoxia-induced stress erythropoiesis.

Myocardial hypertrophy in Sprague-Dawley adult rats exposed to hypobaric hypoxia (0.40 atmosphere of air/18 h daily for 7 days) in a hypobaric chamber was investigated. Changes in the myocardial mass were evaluated on the basis of the dry heart weight and expressed as mg/100 g of total body weight (mean ± SEM). Data are presented indicating that: 1) chronic hypobaric hypoxia causes a significant degree of myocardial hypertrophy in rats; 2) hypertrophic process involves both ventricles (the right more than the left); 3) removal of the hypoxic stimulus leads to the disappearance of hypertrophy when evaluated as an increase in dry heart weight; 4) hypoxia affects the synthesis of a significant amount of connective tissue in the left ventricle, which is not exposed to pressure load. The role of neurohumoral factors (i. e., adrenergic stimulation and catecholamines) in the development of the ventricular hypertrophy is suggested.

Forskolin, a diterpene compound isolated from the roots of Coleus forskohlii, activates adenylate cyclase in membranes from a variety of mammalian tissues. We found that forskolin (10(-7) to 3 X 10(-5) M) caused a concentration-related inhibition of IgE-mediated release of histamine and peptide leukotriene C4 (LTC4) from human basophils and lung mast cells. There was a significant linear correlation between the per cent inhibition of histamine and LTC4 release from both cell types. However, in both systems forskolin exerted a significantly greater inhibitory effect on LTC4 release than on histamine release. The concentration-response inhibition curve was paralleled by a forskolin-induced rise in cAMP levels in human leukocyte and mast cell preparations. The relationship between the effect of forskolin and the cAMP concentration was supported by the finding that forskolin inhibited the "first stage" of antigen-induced histamine release, but not the release caused by the Ca2+ ionophore, A23187. Propranolol, a competitive beta-receptor antagonist, did not block the inhibition of mediator release or the cAMP accumulation caused by forskolin. These data suggest that forskolin modulates the release of mediators of immediate hypersensitivity reactions via the activation of adenylate cyclase in human basophils and mast cells.

The study is based on the experimental model of myocardial hypertrophy in rats. Male Sprague-Dawley adult rats were exposed to acute hypoxia in a hypobaric chamber (0.40 atmospheres of air for 24 hours). Myocardial hypertrophy was detected by wet and dry heart weight values when calculated on the basis of total body weight; i.e., ratio heart weight in mg to body weight in 100 g. The behaviour of myocardial hypertrophy was assessed after treatment of intact animals with Puromycin (10 mg/rat i.p.) or Actinomycin-D (100 micrograms/rat i.p.). The experimental results showed the following dry heart weight values (mean +/- SEM): room pressure 60.5 +/- 1.5; hypoxia 69.2 +/- 0.9; Actinomycin-D + hypoxia 63.5 +/- 0.6; Puromycin + hypoxia 62.4 +/- 1.5, (P less than 0.001). From the comparison of data reveals that inhibitors of protein synthesis completely inhibit the myocardial hypertrophy due to hypoxia. Thus it is documented that acute hypobaric hypoxia is able to induce in the rat an actual myocardial hypertrophy condition as expressed of the enhanced protein synthesis.

5'-N-ethylcarboxamideadenosine (NECA) greater than 2-chloroadenosine greater than adenosine greater than N6-(R-phenyl-isopropyl)-adenosine (R-PIA) inhibited in vitro anti-IgE-induced histamine and peptide leukotriene C4 (LTC4) release from human basophils in a concentration-dependent fashion. Micromolar concentrations of adenosine, NECA and R-PIA potentiated the anti-IgE-stimulated release of histamine and LTC4 from human lung parenchymal mast cells. Submillimolar concentrations of adenosine, NECA and R-PIA inhibited in a concentration dependent manner the release of histamine and prostaglandin D2 (PGD2) from skin mast cells challenged with anti-IgE. These results demonstrate marked heterogeneity of the modulatory effect exerted by adenosine on mediator release from human basophils and mast cells.

Basophil leukocytes and tissue mast cells are inflammatory cells that are found in virtually all human tissues. They appear to be involved in the pathogenesis of such allergic diseases as allergic rhinitis, bronchial asthma, anaphylaxis, atopic and contact dermatitis, chronic urticaria, and hypersensitivity pneumonitis. By releasing a variety of chemical mediators, they could also play a role in the pathophysiology of a wide range of inflammatory disorders of the joints, and of intestine, lung, coronary, and myocardial diseases. Although these two cell types are similar in several aspects, striking differences have also been observed. Moreover, human mast cells from different anatomical sites and within an individual tissue synthesize different mediators and have different release mechanisms. The recent advent of techniques that yield highly purified basophils and mast cells from diverse tissues will probably lead to major advancements in understanding the biochemical and pharmacological mechanisms that control the release process of these cells. The release of mediators from these cells is also controlled by a series of largely undefined biochemical steps that represent the basis of the concept of basophil and mast cell releasability. Alterations of basophil or mast cell releasability have already been detected in patients with allergic rhinitis, bronchial asthma, atopic dermatitis, and chronic urticaria. Taken together, these findings demonstrate that basophils, mast cells, and their chemical mediators play a pivotal role in several inflammatory disorders.

A single administration of testosterone propionate (TP) in ex-hypoxic polycythemic mice induces, at 24 hr after androgen, an amplification of the erythroid burst-forming unit (BFU-E or B) pool in marrow. This phenomenon is not associated with an amplification of the erythroid colony-forming unit (CFU-E or E) compartment and is followed by its depletion. In the other hand, the 36–49 hr rise of erythropoietin (Ep) levels in serum is followed by a 60-hr amplification of the E pool. It is suggested that the latter phenomenon is mediated by enhanced Ep production, whereas the early amplification of the B compartment may derive from a direct influence of TP at the stem cell level.