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Epstein-Barr virus (EBV) seronegativity is rare in people > 20 years old. However, some persons remain EBV-seronegative for nearly their whole lives. The aim of this study was to examine properties of the immune system of EBV-seronegative adults that could contribute to long-term EBV seronegativity. Therefore, differential blood cell counts and lymphocyte subpopulations were determined, and the production of interferon (INF)-alpha and -gamma and interleukin (IL)-6 and -2 in a whole blood assay was investigated. Whereas no differences in the distribution of lymphocyte subpopulations between EBV-seronegative and -positive adults were found, a significant higher percentage of monocytes in EBV-seronegative adults was observed. Significantly more IFN-alpha and IL-6 were detected in culture supernatants of EBV-seronegative persons after stimulation with Newcastle disease virus. In contrast, no differences in the induction of the lymphokines IFN-gamma and IL-2 were seen. These data suggest that faster and higher production of IFN-alpha and IL-6 amy protect EBV-seronegative adults against EBV infection.

Typing for human platelet antigens (HPA) is useful in a variety of clinical situations. We developed a method for genotyping for HPA-1, -2, -3 and -5 by means of the PCR amplification with sequence-specific primers (PCR-SSP) technique. Primer sets were designed to allow PCR amplification for all systems using the same assay conditions. Specificity and sensitivity of the method were assessed in a blind quality control study (n = 112). In 111 cases, results obtained by PCR-SSP were identical as compared with PCR-restriction fragment length polymorphism technique. One discrepancy was found to be due to a typing error in the data sheet. The results of the PCR-SSP technique were available within 3 h. We conclude that genotyping based on PCR-SSP enables rapid typing for HPA systems, which makes this technique feasible in most clinical settings where urgent HPA typing is required.

The pathogenesis of Parkinson's disease (PD) is largely unknown. Recently, several studies have presented evidence of an immunological dysfunction in patients suffering from PD. We studied the immune responsiveness of patients with idiopathic PD (n = 20) by investigation of the ability of peripheral blood mononuclear cells to produce cytokines after mitogenic stimulation in a whole blood assay. A group of age-related healthy blood donors served as control (n = 19). Additionally, white blood count, leukocyte differentiation and lymphocyte subtyping were performed. PD patients had a significantly higher neutrophil count, but analysis of T-cell subsets showed no difference between the two groups. In peripheral blood, secretion of interleukin-2 (IL-2) after mitogenic stimulation was significantly diminished in the patients' group (p < 0.01), whereas values of IFN-alpha 2, IL-6, IFN-gamma and sIL-2R were comparable in both groups. IL-2 production correlated negatively with the mean annual dose of levodopa treatment and correlated significantly (p < 0.002) with amantadine uptake. Analysis of sex, age, duration of illness and other drug intake revealed no correlation with cytokine release. Our findings support the view that there is a selective abnormality in the immune repertoire of peripheral blood lymphocytes in patients suffering from PD, the reasons for which need to be explored.

Platelet concentrates (PC) prepared from pooled buffy coat (BC-PC) contain a variable number of leukocytes from different donors. We questioned whether storage of BC-PC can lead to a lymphocyte activation in the sense of a mixed lymphocyte reaction. BC-PC were prepared from four ABO-identical buffy coats and we undertook leukocyte analyses and measurement of different cyto- kines on days 1,3 and 5 of PC storage (n = 72). Cytokine content was also deter- mined in freshly prepared plasma (n = 48) and PC prepared by thrombapheresis (SD-PC) (n = 12). As control, we studied lymphoproliferation of pooled periph- eral blood mononuclear cells from four individuals in 10 mixed lymphocyte cul- tures (MLCs) under optimal conditions. In the BC-PC, whole blood count and lymphocyte analysis showed a mean leukocyte contamination of 64f28 x 10' per unit with a proportion of lymphocytes of 66.7+13%. In the MLC, levels of in- terleukin-2 (IL-2) and interferon-y (IFN-y) were increased on day 3 and 5 of storage (p

We report the ultrastructural changes occurring in human platelets during eight days of storage. Extension of pseudopodia is frequently observed, but a concentration of organelles in the centre of the platelets is found only in a minor fraction (approximately 5%). Striking changes can be observed in both the granules and the open canalicular system. In fresh platelets, the latter often has the form of stacked membranes that have no lumen, but these membranes separate and spread with increasing storage time. However, the openings of this system on the outer surface of the platelet remain unchanged. Some of these features differ from the morphological description of platelets activated by thrombin or ADP, and suggest that the storage lesion is the result of a prolonged weak activation that leads to an incomplete release reaction within the first five days.

Inducer-specific gene loci and sex are known to play a role in the regulation of interferon-alpha (IFN-alpha) production in mice. Because little is known about the genetic control of the IFN-alpha system in man, we investigated the IFN-alpha production of 468 individuals by culturing peripheral blood with Newcastle disease virus (NDV) or Sendai virus (SDV). The IFN alpha release of different donors varied over a wide range. However, IFN-alpha production of 7 donors showed a donor-specific response over a period of 4 months, which led us to classify some donors as high and low responders. The amounts induced by NDV correlated positively to those induced by SDV. The donor's sex did not alter the IFN-alpha production significantly. The subjects were between 1 and 90 years in age. Highest IFN-alpha levels were obtained in children, followed by a gradual decline with age. Using a specific IFN-alpha-2 enzyme-linked immunosorbent assay (ELISA) and a bioassay, which detects all subtypes, our data showed that the net IFN-alpha production decreased with age. For further studies, we selected 17 low producers and 17 high producers. To analyze a possible influence of major histocompatibility complex (MHC) on IFN-alpha production, the HLA genotypes of 13 low producers and 12 high producers were determined. Here, no correlation between high or low production and HLA genotype was detectable.

Summary Monitoring the storage lesion of platelet concentrates (PC) is commonly done by determining metabolic parameters, release of cytoplasmic markers and granules contents and watching the light microscopical appearance of platelets. Descriptions of ultrastructural changes are rare, and so we ascertained the changes in two important organelles, the α- and lysosomal granules and the open canalicular system (OCS), by morphometric analysis. Single donor PC's were prepared with a CS-3000 plus (Baxter) processor and stored under routine conditions. The volume fraction of the granules displayed a decrease to 43 % after 8 days of storage, and the OCS fraction increased to 143%. Moreover, changes in the morphology of granules and the development of OCS channels are described, and ballooning and even complete lysis of some platelets could be observed after only three days. Furthermore, the membrane flux in the formation of pseudopodia is discussed. Our results stress the need to establish better preparation and storage conditions for platelet concentrates.

BACKGROUND: Neuro-hormonal and hemostatic mechanisms are important in a wide range of psychological and cardiovascular diseases. The use of psychoactive drugs in mental illnesses is often involved with hematologic side effects including impaired platelet function. Subsequently, the risk for the development of cardiovascular diseases may be higher in these patients. Interestingly, platelets that play a key role in cardiovascular complications contain quite a number of neuronal receptors which are involved in psychotic disorders. It has been widely discussed whether psychoactive drugs used in the therapy of psychotic disorders have a direct effect on platelet function and whether the effects are transmitted through the corresponding receptors on the platelet surface. MATERIAL AND METHODS: In this study, we tested several psychoactive drugs regarding their impact on whole blood platelet aggregation. RESULTS: Antidopaminergics preferentially inhibited ADP-induced aggregation whereas anticholinergics mainly inhibited U46619-induced aggregation. Because platelets respond selectively to different psychoactive drugs we assume that corresponding receptors have a functional aspect on platelets and that receptor blockade affects platelet aggregation through different mechanisms. CONCLUSION: The knowledge about the effects of psychoactive drugs on platelet function may help to characterize neuronal receptors on platelets and may contribute to a better understanding of altered platelet function during therapy with psychoactive drugs.

The clinical and molecular findings in three patients with multiple sclerosis (MS) and additional type 1 diabetes mellitus are described. These patients all presented with a severe and progressive disease course of MS. Molecular testing for HLA class II genes demonstrated the presence of the haplotype DRB1 ∗ 0401, DQB1 ∗ 0302 in all patients. This haplotype is closely linked to type 1 diabetes mellitus and is increased among patients with the primary progressive subtype of MS. We conclude that the immunogenetic background in patients with diabetes mellitus may determine the severity and clinical course of MS in these individuals.

SUMMARY: Multipotent mesenchymal stromal cells (MSCs) are currently exploited in numerous clinical trials to investigate their potential in immune regulation, hematopoiesis, and tissue regeneration. The low frequency of MSCs necessitates cell expansion to achieve transplantable numbers. The challenge is to assure safe and high-quality cell production. GMP(Good Manufacturing Practice)-graded cell processing such as cell preparation, culture, and manipulation is mandatory for the progress of such advanced cell therapy. This review summarizes protocols to isolate MSCs from bone marrow and adipose tissue and to expand MSCs for clinical use focussing on culture media composition as well as culture devices and assays to ensure and control quality of the final product.

Background Homograft valves have been shown to be immunogenic, but it is unknown whether this affects valve function. Therefore, we prospectively studied the degree of histoincompatibility (defined as the number of human leukocyte antigen [HLA] mismatches between valve donor and recipient) and the response of the recipient (measured by antibodies against HLA) in relation to echocardiographic parameters of homograft valve function after the Ross procedure. Methods and Results Twenty-six patients (mean age 41±14 years; 20 males, 6 females) and the cryopreserved pulmonary homograft valves that were implanted during a Ross procedure were typed for HLA-A, HLA-B, and HLA-DR. After a mean follow-up of 15±6 months, 14 (54%) of the patients were anti–HLA class I antibody positive. In all but 1 patient, these antibodies were shown to be donor specific. During follow-up, there was a significant increase of the maximal (+6.2±7.1 mm Hg) and mean (+3.2±4.3 mm Hg) transhomograft pressure gradients but not of homograft regurgitation. Neither the number of HLA mismatches nor antibody status was found to have significant impact on homograft valve function. In a multivariate analysis, smaller homograft size ( P =0.001) and younger recipient age ( P =0.044) were shown to be significantly associated with increased transhomograft pressure gradients. Conclusions Implantation of a cryopreserved pulmonary homograft during the Ross procedure can induce a specific humoral response. We observed a significant increase of the transhomograft pressure gradients within 15±6 months after surgery. For this period, we were unable to demonstrate a relationship between this increase and the degree of histoincompatibility.

Background Interleukin-6 (IL-6) promoter polymorphisms at positions -597(G→A), -572(G→C) and -174(G→C) were shown to have a clinical impact on different major diseases. At present PCR-SSP protocols for IL-6 -597/-572/-174haplotyping are elaborate and require large amounts of genomic DNA. Findings We describe an improved typing technique requiring a decreased number of PCR-reactions and a reduced PCR-runtime due to optimized PCR-conditions. Conclusion This enables a fast and efficient determination of IL-6 -597/-572/-174haplotypes in clinical diagnosis and further evaluation of IL-6 promoter polymorphisms in larger patient cohorts.