Isolation of human eosinophil phospholipase D.

Phospholipase D preferentially contained in human eosinophil polymorphonuclear leukocytes as compared to other leukocytes was isolated by sequential asion and cation exchange chromatography and gel filtration. The purified eosinophil enzyme specifically liberated choline from I-alpha-phosphatidyl choline with a pH optimum of 4.5-6.0 and exhibited a pI of 5.8-6.2 on polyacrylamide-gel isoelectric focusing, which are properties shared by phospholipase D from plant sources; however, its apparent mol wt of 60,000 is approximately one-half that of the plant enzymes. Eosinophil and cabbage phospholipase D inactivated a partially purified rat platelet-activating factor (PAF) in a time- and dose-dependent reaction. The cleavage of this PAF activity was attributed to the inherent phospholipase D activity of the eosinophil enzyme since the two activities chromatographed together at each purification step, and there was apparent reciprocal inhibition of choline-generating activity by PAF and of PAF-inactivating activity by phosphatidyl choline. Thus, possible regulatory functions of the eosinophil in immediate hypersensitivity reactions include inactivation of a PAF by phospholipase D as well as degradation of slow-reacting substance of anaphylaxis by arylsulfatase B.

Selective deficiency of immunoglobulin A2.

A case of familial selective IgA2 deficiency is described. The mother had no detectable IgA2, but a low level of IgA1. She had anti-alpha 2 antibodies of the IgG class. One of her daughters also lacked IgA2 with a normal level of IgA1. The analysis of the immunoglobulin haplotypes of the family suggested the deletion of the alpha 2-gene. In addition, the analysis of B lymphocytes of mother and daughter showed the absence of IgA2-bearing cells. Upon stimulation with pokeweed mitogen, the B cells differentiated into IgA1-containing plasma cells, but IgA2-containing cells were not found. The results suggest a defect in the generation of intraclonal B cell isotype diversity. The molecular basis of this phenomenon is unknown.

Implantation of cultured thymic fragments in patients with acquired immunodeficiency syndrome.

Cultured thymic fragments were implanted in one patient with acquired immunodeficiency syndrome (AIDS)-related complex (ARC) and in eight AIDS patients with opportunistic infections (OIs, four patients), Kaposi's sarcoma (KS, two patients), or both (two patients). Thereafter, objective clinical improvement was noted in one patient with OI, and a stable symptom-free condition was observed in the ARC patient and in two other patients with OIs. However, the ARC patient and two of the three patients with OIs developed infections three to six months after implantation. A fourth case of OI and the patients with KS showed progression of the disease. Peripheral blood investigations for counts of total leukocytes, lymphocytes, and T-lymphocyte subsets as well as for lymphocyte stimulation with mitogens showed no changes interpretable as an improvement of the cellular immune deficiency status. We conclude that cultured thymic fragments have no distinct in vivo effect on the course of AIDS, except for a temporary clinical improvement or a period of stable condition in some patients with OIs.

The relevance of IgA deposits in vessel walls of clinically normal skin. A prospective study.

Granular deposits of IgA are known to occur in the walls of superficial vessels in apparently healthy skin of patients with primary IgA nephropathy, Henoch-Schönlein purpura, or alcoholic liver disease, but the specificity of the finding is still a matter of debate. We investigated the disease specificity of these deposits in the skin of patients with kidney and liver diseases. The sensitivity of the finding for the diagnosis of primary IgA was 75%, the specificity was 88%. The sensitivity for the diagnosis of alcoholic liver disease was 71%, but the specificity was only 60%. The specificity for the diagnosis of Henoch-Schönlein purpura, primary IgA nephropathy, or alcoholic disease (in a group of 1,030 patients with various diseases) was 94%. Immunoelectron-microscopic investigation showed the IgA deposits localized within the endothelial cell and in the subendothelial rim.

The thymus in "bare lymphocyte" syndrome: significance of expression of major histocompatibility complex antigens on thymic epithelial cells in intrathymic T-cell maturation.

Thymic biopsies from two patients with combined immunodeficiency and defective expression of HLA class I and class II antigens on blood mononuclear cells ("bare lymphocyte" syndrome) were investigated. This made possible an evaluation of the significance of HLA antigen expression in a detailed (immuno)histologic study. Both thymuses showed a normal lobular architecture with distinct cortex-medulla areas, well-differentiated epithelium, including ultrastructurally defined subtypes, and Hassall's corpuscles. Normal numbers of lymphoid cells were present and normal T-cell phenotype was found. Using anti-HLA-A,B,C antisera, confluent staining of the medulla (stroma and lymphocytes) was observed. One of the thymuses was found to be negative for HLA class II antigen expression: the other revealed only HLA-DR positivity of nonlymphoid cells in the medulla. These cells were not of epithelial nature as judged from double staining with anti-keratin antibody. There was no expression of HLA-DC/DS. These observations differ from findings in the normal thymus, wherein epithelial cells in the cortex carry HLA class I and class II antigens, and epithelial cells in the medulla express HLA class I, and for a minor part class II antigens. The results indicate a normal sequential acquisition of T-cell differentiation antigens in the thymus of both cases. It is suggested that the expression of HLA class I and class II antigens on epithelial cells in the normal thymus cortex does not play a significant role in the sequential acquisition of differentiation antigens on T lymphocytes.

Chorea in systemic lupus erythematosus and "lupus-like" disease: association with antiphospholipid antibodies.

Twelve patients with chorea from a population of 500 patients with SLE and "lupus-like" disease were reviewed. Clinical histories, including time relationships of chorea to the systemic illness and other neurologic manifestations, are reported. Chorea appeared early in the course of disease in most patients, but the development of cerebral infarctions or TIAs occurred subsequently in seven of nine patients demonstrating antiphospholipid antibodies. The relationship of chorea to the presence of these antibodies in nine of 12 patients and the therapeutic outcome are briefly discussed.

Significance of anti-HBc in patients with hepatic disease. A comparative immunohistochemical study of blood and hepatic tissue.

Use of the immunofluorescent technic to examine liver sections disclosed different staining patterns for HBsAg and anti-HBc. In reverse, anti-HBc in serum can be detected by immunofluorescence using HBc-positive liver sections as substrate. In 57 of 89 sera tested, anti-HBc could be observed using immunofluorescence. Of these 57 sera, 31 contained HBsAg (54.5%), 15 contained anti-HBs (26.3%), and one contained both HBsAg and anti-HBs (1.7%). Ten patients (17.5%) had anti-HBc without HBsAg or anti-HBs; nine of these patients had disturbed hepatic functions. In eight cases biopsies of liver tissue showed lesions varying from steatosis to cirrhosis. In 32 anti-HBc-negative sera no HBsAg was found; transient anti-HBs was found in one case and persistent anti-HBs was also seen in one case. Biopsies of the livers of 27 anti-HBc-positive patients disclosed the whole spectrum of histologic lesions. There appear to be correlations between HBsAg in blood and in liver, and also between nuclear IgG, the HBsAg pattern in hepatic tissue, and active hepatic disease.

Alcoholic hepatic disease. Specificity of IgA deposits in liver.

The extent to which the immunofluorescent phenomenon of homogeneous deposition of IgA in the hepatic sinusoids (so-called continuous pattern) was specific for alcoholic hepatic disease was investigated. In 66 of 320 liver biopsy specimens a continuous IgA pattern was observed. Alcoholism was mentioned in the cases of 50 of the 66 patients (76%). The biopsy specimens in the remaining 254 cases continued scanty detectable IgA (discontinuous pattern) or none. In the latter group only eight patients (3%) had histories of alcoholism. A direct correlation between a continuous IgA pattern in the hepatic sinusoids and alcohol abuse is thus inferred (P less than 0.001). Additional findings of the concomitant occurrence of IgA in the perisinusoidal linings of the liver, the wall of superficial cutaneous capillaries, capillaries of the gut, and the glomerular mesangium in association with alcoholic hepatic disease further substantiates the concept of the existence of an IgA-associated disease.

Characteristics of IgA deposits in liver and skin of patients with liver disease.

The presence of IgA deposits in a continuous pattern along hepatic sinusoids is a specific entity for alcoholic liver disease. In superficial skin blood vessels of patients with liver disease, IgA deposits can occur. The authors characterized the deposits for IgA-subclass epitope expression and for macromolecular configuration (assessment of [hidden] J-chain determinants and of secretory component-binding capacity). A variety of monoclonal anti-IgA-subclass reagents were applied, which proved to be specific in control experiments on blastoid cells generated by pokeweed mitogen stimulation of blood mononuclear cells and frozen tissue sections of normal jejunum. IgA1 is the major component in IgA deposits in liver (n = 83) and skin (n = 31) of patients with liver disease. Macromolecular IgA is detectable in only one-fifth of the cases. The authors' data do not indicate that hepatic IgA deposits in liver disease are of gastrointestinal origin. Out of the circulating IgA pool, IgA1 appears to be most capable of being deposited in tissue.

Morphometric study of histological changes in sublabial salivary glands due to aging process.

The sublabial salivary glands were studied by morphometric methods in 68 healthy volunteers to establish possible changes related to age in those tissue components that are affected in Sjögren's syndrome and connective tissue diseases (and which might stimulate Sjögren's syndrome). There was an increase in the amount of connective tissue and intralobular ducts with age and a corresponding decrease in acinar tissue. During the aging process changes in the intralobular ducts occurred: the outer and inner diameters of these ducts and the thickness of the epithelium decreased, but the ratio of the outer and inner diameters of the ducts remained constant. The amount of diffuse lymphoplasmacytic infiltrate and the vascularity of the tissue remains constant with age. In 15 of the subjects, however, discrete lymphocytic foci were seen and in six of these more than one focus/4 mm2 of salivary tissue was found, which has been described as suggestive of Sjögren's syndrome. The volume percentage of lymphocytic foci is constant during the aging process. The histological features commonly used to diagnose Sjögren's syndrome may occur in normal people, and false positive diagnoses will occur if these criteria are rigidly adhered to. Morphometry may provide more reliable criteria for distinguishing changes induced by inflammation and related to age which occur in salivary tissue.

Monotypic plasma cells in labial salivary glands of patients with Sjögren's syndrome: prognosticator for systemic lymphoproliferative disease.

AIMS: To determine the prevalence of plasma cell monotypia in labial salivary gland tissue of patients with and without Sjögren's syndrome, and to evaluate its relation to the development of systemic monoclonal lymphoproliferative disorders. METHODS: A quantitative immunohistological study was performed on labial salivary gland tissue of 45 patients with Sjögren's syndrome, 18 with rheumatoid arthritis without Sjögren's syndrome, and 80 healthy controls. In none of the patients with Sjögren's syndrome was there evidence of systemic monoclonal lymphoproliferative disease at the time of biopsy. RESULTS: Monotypic plasma cell populations, defined by a kappa:lambda ratio of > or = 3, were only observed in older patients (above 43 years) with Sjögren's syndrome. In almost all these patients monotypic plasma cell populations were present in multiple labial salivary gland tissues and the IgM/kappa monotypia was observed most frequently. The prevalence of monotypic plasma cell populations in the group with Sjögren's syndrome was 22% (10/45) and there was no significant predilection for primary Sjögren's syndrome. Of special clinical interest was the observation that progression to systemic monoclonal lymphoproliferative disease had occurred exclusively in this subgroup of patients with Sjögren's syndrome, with a prevalence of 30% (3/10). CONCLUSION: Quantitative immunohistological examination of labial salivary gland tissues provides pathologists with a simple method to select those patients with Sjögren's syndrome who have an increased relative risk at the time of biopsy to develop benign or malignant lymphoproliferative disorders.

Long-term course of tear gland function in patients with keratoconjunctivitis sicca and Sjögren's syndrome.

AIMS: To assess the course of tear gland function of patients with keratoconjunctivitis sicca (KCS) associated with primary (KCS-PSS) or secondary Sjögren's syndrome (KCS-SSS), and of patients with KCS not related to Sjögren's syndrome (KCS-NS). METHODS: In 106 patients with dry eye an ophthalmic diagnosis of KCS was made. Subsequent evaluations revealed a diagnosis of KCS-PSS in 31, KCS-SS in 19, and KCS-NS in 56 patients. Follow up assessments have been performed 10-12 years after initial diagnosis. RESULTS: At baseline and at follow up tear gland function tests were worse in patients with KCS-PSS compared with the other forms of KCS. At follow up in the KCS-SSS patient group the tear gland function variables returned to marginal normal limits. In contrast with expectation, a marked improvement of the tear gland function variables in the KCS-NS patient group was noted. CONCLUSIONS: In KCS-PSS patients tear gland function is characterised by a steady state situation. In KCS-SSS patients the normalisation of tear gland function variables most probably reflects a remission of the underlying disease. In view of the overall improvement in KCS-NS patients the term age related KCS should be avoided.

Lupus anticoagulant: revival of an old phenomenon.

The term lupus anticoagulant (LAC) refers to antiphospholipid antibodies assessed by means of phospholipid-dependent coagulation tests. About two-thirds of LAC-positive patients described in the literature have systemic lupus erythematosus (SLE) or a lupus-like syndrome. The estimated prevalence in SLE is about 30%. LAC is a marker of a subset of patients characterized by a high prevalence of thromboembolic complications (present in 40% of the LAC-positive patients), fetal loss (related to placental infarction), thrombocytopenia, biologically false positive tests for syphilis, antinuclear antibodies and a positive Coombs' test. There is a strong correlation between the presence of LAC and antibodies to the phospholipid cardiolipin, which can be estimated with relatively simple solid phase assays. Studies demonstrating in some patients interactions between LAC and either humoral factors with important functions in the (patho-) physiology of thrombosis, endothelial cells or platelets strongly suggest that LAC represents autoantibodies with pathogenic significance.

The outcome of angiography in patients with Raynaud's phenomenon: an unexpected role for atherosclerosis and hypercholesterolemia.

OBJECTIVE: Upper extremity angiography can make an important contribution to the diagnosis in vasculopathy. The present study was designed to assess the diagnostic role of upper extremity angiography in patients with disturbed circulation of the hand, according to a standardised protocol. METHODS: The study was carried out in an outpatient setting in 103 patients suffering from bilateral Raynaud's phenomenon without any obvious underlying disease and who were unresponsive to nifedipine and aspirin. All patients had angiographies taken according to a standardized technique using vasodilating medication, and reviewed according to a standardised protocol that covered all the known characteristics of angiopathy such as diminished flow, stops, tortuosity, irregularity of the wall, tapering, collaterals and blushing. RESULTS: Standardised angiograms showed vasculopathy compatible with primary vasospasm in 42 patients [all women; mean age 35.1 years], atherosclerotic vascular disease in 44 patients [M/F 9/35; mean age 46.7 years], peripheral embolism in 8 patients [M/F 4/4; mean age 38.4 years], vasculitis in 3 patients [3 women; mean age 38 years] and Buerger's disease in 3 patients [3 men; mean age 47 years]. Inter-observer differences were present in 4 cases, but consensus could be reached through open discussion. An unexpected 47% of patients with atherosclerotic vascular disease had dyslipidemia, frequently of familial origin. CONCLUSIONS: The standardised angiography protocol proved to be helpful in the assessment of upper extremity angiography. Surprisingly, a high prevalence of angiographic abnormalities compatible with atherosclerotic vascular disease could already be diagnosed in relatively young patients with Raynaud's phenomenon, of whom 47% showed hypercholesterolemia.

Binding of antibodies to nonhistone nucleoproteins on keratinocytes in suspensions.

Antibody binding on the cell surface of epidermal cells, recently established on cultured neonatal foreskin cells, is supposed to play a role in the pathogenesis of in vivo antinuclear antibodies (ANA) of the skin. To study this phenomenon in suspensions of adult human keratinocytes, a cell system more closely related to the in vivo situation, we investigated the binding capacity of nine sera with various antibody profiles against nuclear components, as well as a murine monoclonal Sm-antibody. It was found that sera containing antibodies against nonhistone nucleoproteins bound to the cell surface of keratinocytes, whereas monospecific anti-dsDNA sera and the murine anti-Sm serum did not. This binding was found in both basal and suprabasal keratinocytes. The percentage of cells showing antibody binding was not significantly enhanced by preirradiation with ultraviolet light, as was found in a previously study. The cell surface binding is probably an antigen-antibody binding and not the result of cross-reactivity. Such cell surface binding may be important for the formation of in vivo ANA in the skin.

Quantitative immunohistologic and histomorphometric diagnostic criteria for Sjögren's syndrome.

Sjögren's syndrome (SS) is a chronic auto-immune exocrinopathy, especially affecting the lacrimal and salivary glands. The aim of this study is to improve the diagnostic possibilities of the sublabial salivary gland (SSG) biopsy. The SSG biopsies of 19 patients with SS and 65 healthy control subjects were used in a quantitative immunohistologic and histomorphometric study. Statistical analysis of the immunohistochemical data resulted in a diagnostic criterion, which is based on the percentages of IgA- and IgG-containing plasma cells. Statistical analysis of 3 immunohistologic and 6 histomorphometric features resulted in a combined immunohistologic and histomorphometric criterion, which is based on 2 immunohistologic parameters (the percentages IgA- and IgG-containing plasma cells) and 3 histomorphometric parameters (the volume percentages of acini, intralobular ducts and diffuse lymphoplasmacytic infiltrate). The immunohistologic diagnostic criterion has a specificity of 95.4%, a sensitivity of 100% and an overall percentage of misclassification of 3.6%. The combined diagnostic criterion has a specificity of 98.5%, a sensitivity of 100% and an overall percentages of misclassification of 1.2%. Furthermore it reduces the number of false positive diagnoses with a factor 6 from 9% to 1.5%.

Quantitative immunohistologic study of lip biopsies. Evaluation of diagnostic and prognostic value in Sjögren's syndrome.

In a group of 45 patients with Sjögren's syndrome (SS) and 80 controls the high specificity (95%) and sensitivity (100%) of a recently proposed bivariate quantitative immunohistologic (QIH) criterion for SS, based on percentages of IgA and IgG-containing plasma cells in labial salivary gland (LSG) tissue, was confirmed. The best univariate QIH criterion for discrimination between LSG biopsies of SS patients and controls appeared to be based on the percentage of IgA containing plasma cells, and had a specificity of 99% and a sensitivity of 96%. A criterion based only on the percentages of IgM-containing plasma cells, proposed in another recent study, resulted in a high number (31%) of false negatives. Interobserver reproducibility of QIH diagnoses was excellent. Moreover it was demonstrated that accuracy, precision and the interobserver reproducibility of plasma cell counting depends on the choice of tissue fixation and immunohistologic staining procedure. The combination of formol sublimate fixation and peroxidase anti-peroxidase procedure appeared to be the best combination for QIH examination. Furthermore, in 2 SS patients systemic monoclonal IgM/kappa gammopathy was preceded by high predominance of IgM and kappa containing plasma cells in the plasma cellular infiltrate of the LSG tissue.

Anti-phospholipid antibodies and vascular pathology.

Over the past decade it has been recognized that there is an association between the presence of anti-phospholipid antibodies (lupus anticoagulant, anti-cardiolipin antibodies) and thromboembolic phenomena. This "anti-phospholipid syndrome", which was originally described in patients with lupus erythematosus also occurs in patients without an underlying systemic disease. An answer to the question whether anti-phospholipid antibodies cause thrombosis warrants further studies. Especially the recent findings on a role of plasma proteins in the binding of these antibodies to phospholipids will have to be taken into account. To obtain guidelines for optimal treatment of patients suffering from this syndrome, additional studies will have to be performed.

Polyarteritis nodosa in a patient with AIDS.

A 33-year-old male homosexual with the acquired immunodeficiency syndrome (AIDS) presented with pain, blue colouring and ulceration of fingers and toes. Angiography of the superior mesenteric artery and renal arteries showed vessel wall irregularities and aneurysms, characteristic of polyarteritis nodosa (PAN). We describe this patient because of the possible association between systemic vasculitis and AIDS.

Neuro-musculo-skeletal manifestations in primary Sjögren's syndrome.

Primary Sjögren's syndrome is a systemic autoimmune disorder whose main characteristics are dryness of the eyes and mouth, caused by lymphocytic infiltration of the exocrine glands. Patients may also show signs of extraglandular involvement of lung, liver, kidney and vessel walls, as well as of the central and peripheral nervous systems, muscles and joints. This article presents a review of the literature on extraglandular involvement of the peripheral and central nervous systems, muscles and joints. Several data support the hypothesis that vasculitis is the underlying mechanism. The need for an extended inventory of the extraglandular manifestations, preferentially linked to immunoserological and -histological investigations to gain more insight into the aetiology and pathogenesis is stressed. As far as the clinical picture is concerned, myalgia and arthralgia are often reported, but myositis and arthritis are rare. Data about the prevalence of peripheral and central nervous system involvement are conflicting: factors contributing to these differences are discussed. As insight into prognosis and therapy will strongly depend on the diagnostic criteria used, the need for international agreement on these is emphasized.