The dynamic changes of zinc transporter 8 autoantibodies in Czech children from the onset of Type 1 diabetes mellitus.

AIMS: The prevalence of autoantibodies to zinc transporter 8 (ZnT8) in Czech children at the onset of Type 1 diabetes mellitus and dynamic changes in ZnT8 autoantibody levels during disease progression were studied. The value of ZnT8 autoantibody measurements in diagnosis of Type 1 diabetes was assessed. METHODS: Serum samples from 227 children with newly diagnosed Type 1 diabetes and from 101 control children without diabetes were analysed in a retrospective cross-sectional study. One hundred and seventy-one samples from 116 of the patients with diabetes were analysed in a follow-up study at (median) intervals of 1, 3, 5 and 10 years after onset of Type 1 diabetes. ZnT8 autoantibodies were measured using a bridging enzyme-linked immunosorbent assay, while antibodies to glutamic acid decarboxylase, insulinoma antigen 2 and insulin were measured by radioimmunoassays. RESULTS: ZnT8 autoantibodies were detected in 163/227 (72%) of children at Type 1 diabetes onset and in 1/101 (1%) of the control subjects. Sixteen out of 227 (7%) patients with Type 1 diabetes were antibody negative based on three antibodies (glutamic acid decarboxylase, insulinoma antigen 2 and insulin). This false-negative rate was reduced to 10/227 (4.4%) (P < 0.05) after inclusion of ZnT8 autoantibody measurements. Of the children, 142/227 (63%) were positive for at least three antibodies and the most common combination was insulinoma antigen 2, glutamic acid decarboxylase and ZnT8. ZnT8 autoantibody levels decreased over time after Type 1 diabetes onset and the presence and level of ZnT8 autoantibodies correlated with IA-2 autoantibodies. CONCLUSIONS: A ZnT8 autoantibody enzyme-linked immunosorbent assay showed 72% disease sensitivity and 99% specificity at Type 1 diabetes onset. Measurements of ZnT8 autoantibodies are important for Type 1 diabetes diagnosis and should be included in the panel of autoantibodies tested at the onset of Type 1 diabetes.

Healthy first-degree relatives of patients with type 1 diabetes exhibit significant differences in basal gene expression pattern of immunocompetent cells compared to controls: expression pattern as predeterminant of autoimmune diabetes.

Expression features of genetic landscape which predispose an individual to the type 1 diabetes are poorly understood. We addressed this question by comparing gene expression profile of freshly isolated peripheral blood mononuclear cells isolated from either patients with type 1 diabetes (T1D), or their first-degree relatives or healthy controls. Our aim was to establish whether a distinct type of 'prodiabetogenic' gene expression pattern in the group of relatives of patients with T1D could be identified. Whole-genome expression profile of nine patients with T1D, their ten first-degree relatives and ten healthy controls was analysed using the human high-density expression microarray chip. Functional aspects of candidate genes were assessed using the MetaCore software. The highest number of differentially expressed genes (547) was found between the autoantibody-negative healthy relatives and the healthy controls. Some of them represent genes critically involved in the regulation of innate immune responses such as TLR signalling and CCR3 signalling in eosinophiles, humoral immune reactions such as BCR pathway, costimulation and cytokine responses mediated by CD137, CD40 and CD28 signalling and IL-1 proinflammatory pathway. Our data demonstrate that expression profile of healthy relatives of patients with T1D is clearly distinct from the pattern found in the healthy controls. That especially concerns differential activation status of genes and signalling pathways involved in proinflammatory processes and those of innate immunity and humoral reactivity. Thus, we posit that the study of the healthy relative's gene expression pattern is instrumental for the identification of novel markers associated with the development of diabetes.

Enhanced STAT3 phosphorylation and PD-L1 expression in myeloid dendritic cells indicate impaired IL-27Ralpha signaling in type 1 diabetes.

Interleukin 27 (IL-27), a member of the IL-12 family, is important for T cell differentiation; however, little is known about its effect on dendritic cells (DCs). IL-27 can activate multiple signaling cascades, including the JAK/STAT pathway, and depending on the setting it can both promote and antagonize inflammatory responses. An anti-inflammatory function of IL-27 has been reported in several autoimmune diseases; however, in type 1 diabetes (T1D), an autoimmune disease where autoreactive cytotoxic T cells attack insulin-producing beta cells, IL-27 has been shown to have a dual role and contradictory effects. Here, we show impaired IL-27 signaling in a large cohort of T1D patients (n = 51) compared to age- and gender-matched healthy donors. Increased expression of the IL-27 receptor subunit IL-27Ralpha mRNA in purified myeloid DCs (mDCs), detected by gene expression microarrays was mirrored by enhanced signal transduction in T1D mDCs in response to IL-27 stimulation. Higher STAT phosphorylation in T1D patients was also accompanied by elevated expression of the inhibitory molecules PD-L1, PD-L2 and PD-1, which may suggest not only immunomodulatory mechanisms of IL-27 in T1D but also a compensatory effort of T1D dendritic cells against the ongoing inflammation.

Influence of maternal hyperglycaemia on cord blood mononuclear cells in response to diabetes-associated autoantigens.

UNLABELLED: Perfect maternal diabetes compensation is crucial for the outcome of the baby. However, little is known how hyperglycaemia influences the specific immune response. Furthermore, babies of type 1 diabetes (T1D) mothers have less risk of development T1D than babies with a T1D father. This study aimed to analyze the effect of maternal hyperglycaemia on newborns with focus on the response to diabetes-associated autoantigens. POPULATIONS: (1) Newborns of T1D mothers split into groups according to maternal diabetes compensation during the 3rd trimester: perfect (n = 15) or acceptable (n = 25) compensation. (2) newborns with T1D father (n = 12) (3) newborns with a mother treated for either gestational or type 2 diabetes (n = 10) (4) control newborns (n = 25). Spontaneous as well as diabetes-associated autoantigen-stimulated production of 23 cytokines and chemokines were tested using protein microarray. In addition, the influence of glucose on cytokine and chemokine responsiveness was analyzed in vitro. The study groups differed in their spontaneous as well as stimulated cytokine and chemokine spectra. A prominent Th1 response (high IFN-gamma) from autoantigen stimulation was observed especially in babies of T1D fathers (P = 0.001) and also in mothers with perfect diabetes compensation during the 3rd trimester (P = 0.016) in comparison with control newborns. By contrast, cord blood mononuclear cells cultivated in vitro in high glucose concentration decreased the diabetogenic stimulated Th1 cytokine response. Maternal 'sweet' as well as 'autoimmune environment' may both lead to lower occurrence of T1D within their offspring. Further studies will reveal the exact immunological mechanism of this observation.

CD 127- and FoxP3+ expression on CD25+CD4+ T regulatory cells upon specific diabetogeneic stimulation in high-risk relatives of type 1 diabetes mellitus patients.

Abnormalities in CD4+CD25+ regulatory T cells (Treg) may contribute to type 1 diabetes (T1D) development. First-degree relatives of T1D patients are at increased risk especially when they carry certain HLA II haplotypes. Using two novel markers of CD4+CD25+ Treg (CD127- and FoxP3+ respectively), we evaluated number and function of Treg after specific stimulation with diabetogeneic autoantigens in 11 high-risk (according to HLA-linked risk) relatives of T1D patients and 14 age-matched healthy controls using a cytokine secretion assay based on interferon-gamma (IFN-gamma) production. High-risk relatives of T1D patients had significantly lower pre- and post-stimulatory number of CD127- Treg than that of healthy controls (P < 0.05). Labelling Treg with FoxP3+ demonstrated similar trend but did not reach statistical significance. Although the stimulation with diabetogenic autoantigens did not lead to a significant change in number of Treg in both groups, the defective function of Treg was performed by significantly higher activation of diabetogeneic T cells in high-risk relatives of T1D patients compared to healthy controls (P < or = 0.02). Individuals at increased HLA-associated genetic risk for T1D showed defects in Treg.

Determinants of preclinical atherosclerosis are different in type 1 and type 2 diabetic women.

Diabetes mellitus type 2 ranks among the strongest predictors of cardiovascular diseases (CVD) while the association of type 1 diabetes with CVD is more complex. We studied differences between type 1 and 2 diabetic women regarding association of cardiovascular risk factors with preclinical atherosclerosis expressed as intima-media thickness of common carotid (IMT CCA) and femoral arteries (IMT CFA) measured by high resolution ultrasound. Women with type 1 (n=203) and type 2 diabetes (n=123) were examined with regard to the presence of cardiovascular risk factors. In type 1 diabetic women strong association between IMT CCA and body mass index, waist circumference, and total body fat was found in contrast to type 2 diabetic women. In type 2 diabetic women strong association between IMT CCA and fasting glucose, glycated hemoglobin, and atherogenic index of plasma (log TG/HDL cholesterol) was observed in contrast to type 1 diabetic women. In type 1 diabetic women, IMT CFA was associated with body fat in contrast to type 2 diabetic women. Preclinical atherosclerosis in type 1 diabetic women was strongly associated with factors reflecting body fat and its distribution, while in type 2 diabetic women preclinical atherosclerosis was associated with markers reflecting glucose and lipid metabolic disorders.

Histological correlation between different centers using the skin explant model to predict graft-versus-host disease following bone marrow transplantation.

Acute graft-versus-host disease (GVHD) remains the major complication of allogeneic stem cell transplantation (SCT) with an incidence of 40-60% and a mortality of up to 50%. Several assays have been developed to try to predict the development of GVHD including the mixed lymphocyte culture reaction, cytotoxic and helper T lymphocyte precursor frequency assays. In the Northern region of England we have used an in vitro skin explant model for predicting GVHD in MHC compatible bone marrow transplant recipients since 1988. The aims of the present study was to test the reproducibility of the model in two other bone marrow transplant centers in Europe. The assay consists of incubating patient skin explants with effector cells from mixed donor versus recipient lymphocyte cultures and the subsequent detection of graft-versus-host reactions by histopathological grading (0-IV) of the skin explants. 503 slides from 134 patients were evaluated. All were graded for negative GVHR grade 0-I or positive grade II-IV. Results from control and test slides significantly correlated between centers to the p value of 0.0001 by Fisher's exact probability test. These results show that the skin explant assay is reproducible between centers and supports the continued use of the assay to predict GVHD in allogeneic stem cell transplant recipients.

Anti-keratin antibodies in patients with juvenile idiopathic arthritis.

OBJECTIVE: We discuss the presence of anti-keratin antibodies (AKA) of the IgG class in patients with defined juvenile idiopathic arthritis (JIA). METHODS: An indirect immunofluorescence test with rat oesophagus substrate was used for the detection and quantification of AKA antibodies in patients'sera. RESULTS: Overall 30/60 patients with JIA had sera positiveforAKA (50%, p=0.0005) ranging from 1:20 to 1:160 dilutions. Using the classification criteria for childhood idiopathic arthritis, AKA occurred in 2/7 patients with systemic disease (28.6%), in 13/30 patients with RF negative polyarthritis (43.3%, p=0.008) and in 12/18 RF positive polyarthritis (66.7%, p=0.002). AKA were also found in a small cohort of patients with oligoarthritis (1/3) and psoriatic arthritis (2/2). AKA positivity occurred in 3/26 healthy controls at a 1:20 dilution. The presence ofAKA was correlated as well as with the severity of the disease. Our study revealed that AKA was present overall in 16/29 patients (55.2%) with severe JIA and in 11/26 patients (42.3%) with non-severe disease. We also observed that AKA remained positive regardless of disease activity. AKA were detectable in 44.4% patients with active JIA and in 45.9% patients in the complete or near remission. CONCLUSION: Our data suggest that AKA are present in patients with JIA. However no correlation with severity or disease activity was observed.

Pregnancy in a woman suffering from type 1 diabetes associated with Addison's disease and Hashimoto's thyroiditis (fully developed Autoimmune Polyglandular Syndrome Type 2).

In this article the pregnancy of a woman suffering from the complete triad typical of Autoimmune Polyglandular Syndrome Type 2 (Addison's disease + type 1 diabetes + Hashimoto's thyroiditis) is reported. By using insulin pump therapy with insulin lispro, it was possible to balance diabetes control with changes of steroid replacement therapy. Pregnancy was uneventful until week 27, when signs of preeclampsia occurred. The boy was born without difficulty at gestational age 37 weeks by planned cesarean section but signs of diabetic fetopathy (macrosomia, hypoglycaemia and hypocalcaemia) were expressed. He required a short course of hydrocortisone therapy. He made a good and rapid recovery. The mother made a good post-operative recovery too, but 4 months after the delivery microalbuminuria as well as mild hyperuricemia are still present. Interdisciplinary approach and very careful observation of the mother as well as of the child enabled successful outcome of this highly risky pregnancy.

In vitro autoreactivity against skin in rheumatoid arthritis: are peripheral blood mononuclear cells of patients with rheumatoid arthritis able to lyze autologous keratinocytes?

An in vitro skin explant model was originally developed to predict the occurrence and severity of acute graft-versus-host disease in allogeneic hematopoietic stem cell transplants. In previous studies we reported that peripheral blood mononuclear cells of patients with rheumatoid arthritis were able to induce graft-versus-host-like histopathological changes when co-cultured in vitro with autologous skin explants. The aim of the present study was to verify if observed skin damage was really of autoimmune origin. Using a 51chromium release cytotoxic assay we found that peripheral blood mononuclear cells of patients lyzed autologous keratinocytes (n=5 patients with rheumatoid arthritis) but not autologous lymphoblasts (n=4 with rheumatoid arthritis, n=8 patients with juvenile idiopathic arthritis). No specific lysis of keratinocytes or lymphoblasts was observed in healthy controls (n=15). We hypothesize that autologous peripheral blood mononuclear cells might recognize similar autoantigen(s) expressed on epidermal cells, which gives rise to an autoimmune response in the synovium.

[Levels of peripheral circulating nucleated erythrocytes in pregnant women for noninvasive prenatal diagnosis]. / Koncentrace fetálních jaderných erytrocytu cirkulujících v periferii gravidních zen pro úcely neinvazivní prenatální diagnostiky.

OBJECTIVE: Enrichment of nucleated red blood cells (NRBCs) from maternal blood for non-invasive prenatal diagnosis. DESIGN: Pilot study. SETTING: 2nd Clinic of Paediatrics, University Hospital Motol, Prague, Czech Republic. METHODS: Mononuclear cells were isolated from 13-28 ml of peripheral maternal blood between 13 and 37 weeks of gestation. Leukocytes from maternal peripheral blood were depleted from mononuclear cells by treatment with anti-CD14 and anti-CD45 microbeads and high-gradient magnetic cell separation (MACS) on VarioMACS. NRBCs were sorted from CD14-/CD45- fraction by positive selection using anti-CD71 microbeads on MiniMACS. All sorting steps were analysed by three-colour cytometric analysis with FACScan flow cytometer. RESULTS: In 68 out of 78 pregnant woman (87%) NRBCs were found in range 2 x 10(5) - 1.02 x 10(6). NRBC were enriched with an average enrichment rate of 138-fold ranging from 4-526 fold. In our cohort of pregnant woman the number of isolated NRBCs was individual. We identified NRBCs from the 13th week of gestation. CONCLUSION: The aim of the study is to establish and standardise the method of enrichment of NRBCs from maternal blood samples and verify the applicability of this alternative source for non-invasive prenatal diagnosis.

[Fetal nucleated erythrocytes in the peripheral circulation of pregnant women as a noninvasive method of fetal sex determination]. / Fetální jaderné erytrocyty cirkulující v periferii gravidních zen jako neinvazivní zdroj detekce pohlaví fetu.

OBJECTIVE: Determination of the sex of the foetus from enriched nuclear red blood cells (NRBC) circulating in maternal blood during pregnancy. METHODS: NRBC were enriched from 13-28 ml peripheral blood of 32 pregnant women using double MACS procedure. NRBCs were enriched by magnetic activated cell sorting using anti-CD71 (transferrin receptor) monoclonal antibodies. Unwanted leukocytes were depleted using monoclonal antibodies against CD14 and CD45. The sex of the foetus was analysed by using dual-colour FISH with X and Y specific probes. The experimental results obtained from the noninvasive procedure were compared to karyotype obtained from amniocentesis or chorionic villus sampling. RESULTS: In 15 out of 17 male foetuses we could identify one X and one Y signal. In another 15 pregnant women carrying female foetuses two X signals were observed. CONCLUSION: NRBC circulating in blood of pregnant women can be used as an alternative source for determination of the sex of the foetus with a risk of false negative results (2/17, 12%). The problem of false negative results can be solved by using more sophisticated methods of enrichment and preparedness of the slides for FISH analysis.

[A method for modeling skin explants--an in vitro predictive test for graft vs host disease in allogenic hematopoietic cell transplantation]. / Metoda modelu kozního explantátu--in vitro graft versus host disease prediktivní test pro alogenní transplantace hematopoetických bunek.

BACKGROUND: Acute graft versus host disease (GvHD) remains a severe complication of allogeneic haematopoietic stem cell transplantation (HSCT). Our study summaries results of skin explant assay (SEA) as a pretransplant GvHD predictive test in a cohort of paediatric (n = 33) and adult (a = 8) patients receiving grafts from their HLA identical siblings (n = 28), haploidentical relatives (n = 3) and unrelated donors (n = 10). Results GvHD prediction are correlated with the occurrence and severity of acute GvHD posttransplant and effect of GvHD prophylaxis on GvHD clinical outcome is evaluated. METHODS AND RESULTS: SEA utilises responding lymphocytes of the donor, which are sensitized firstly in vitro by mononuclears cells of patient in allogeneic mixed lymphocyte culture (MLC) and subsequently co-cultured with recipient's skin. Histopathological changes found in patients' skin explants are evaluated according to standard Lerner classification for acute GvHD. In general, GvHD predictive results in SEA correlated with GvHd clinical outcome in 28 out of 41 tested patients (68%, p = 0.015). In a cohort of HLA identical sibling transplants GvHD predictive results correlated with clinical manifestation of acute GvHD only in 15 out of 28 patients on individual GvHD prophylaxis. GvHD prophylaxis in the form of cyclosporine A (CsA) combined with short-term methotrexate (MTX) reduced the risk of acute GvHD in 10 out of 14 transplanted patients (71%) meanwhile CsA alone prophylaxis only in 1 out of 5 patients (20%). In a cohort of unrelated pairs on CsA/MTX prophylaxis combined with horse anti-lymphocyte globuline (ALG) correlated the GvHD prediction with GvHD clinical outcome (100%, p = 0.003). In all patients transplanted with the grafts from their haploidentical relatives the occurrence of severe GvHD was predicted. CONCLUSION: Skin explant assay helps identify pretransplant patients at higher risk of severe acute GvHD. GvHD predictive results enable the transplantation team to individualise GvHD prophylaxis and to optimise selection of the donor.

Reactivity to Helicobacter pylori antigens in patients suffering from thyroid gland autoimmunity.

The role of infection in autoimmunity is widely discussed. In this study we concentrated on relationship between HELICOBACTER PYLORI as a very important gastroduodenal pathogen and autoimmune thyroiditis (AT). Forty seven AT patients and 34 healthy controls were enrolled. They were split into: THP ( H.PYLORI positive patients, n=17), THN ( H.PYLORI negative patients, n=30), CP ( H.PYLORI positive controls, n=17) and CN groups ( H.PYLORI negative controls, n=17). By protein microarray we analysed production of 23 cytokines and chemokines prior and post stimulation with H.PYLORI lysate and its lipopolysaccharide (LPS). Reactivity to lysate as well as to bacterial LPS differed within groups. The lowest basal cytokine and chemokine production was observed in CN group but these subjects reacted significantly to specific stimulation by increasing IFN-gamma (in comparison with THP p=0.01 for LPS and p=0.004 for H.PYLORI lysate) and TGF-beta production (p=0.015 for LPS). In contrast, IL-10 and IL-5 were decreased in this group. In CP, THN and THP groups, we observed in general higher chemokine response. THP group increased proinflammatory IL-6 after specific stimulation as well (in comparison with CP p<0.0001 for LPS stimulation). We observed different "reactivity pattern" to H.PYLORI within groups with low basal cytokine and chemokine production in healthy H.PYLORI negative controls but with clear specific response in IFN-gamma and TGF-beta production in this group. Adequate immune reaction which is joined to appropriate immunoregulation leads to prevention of the chronic infection and on the other hand may prevent the development of "connected" diseases such as autoimmune.

Cord blood cytokine profile detection in neonates with T1D parents -- monitoring of cellular auto-reactivity using protein microarray.

Type 1 diabetes (T1D) is a great medical challenge and its incidence rises rapidly. T lymphocytes and their cytokine production are supposed to play a major role in T1D development. So far, there is no potent tool to recognize the early signs of cellular auto-reactivity which leads to beta-cell damage. The naïve immune system of the newborn (not yet influenced by external factors) can be used as an important model for T1D pathogenesis studies. Cord blood samples of 22 healthy neonates born at term to a diabetic parent (T1DR) and 15 newborns with no family history of any autoimmune disease (controls) were collected. Determination of 23 cytokines was performed before and after the stimulation with diabetogenic autoantigens using protein microarray. We observed lower basal production of all detected cytokines in the T1DR group - granulocyte/macrophage colony-stimulating factor (GM-CSF) (P = 0.025), growth regulated protein (GRO) (P = 0.002), GRO-alpha (P = 0.027), interleukin (IL)-1-alpha (P = 0.051), IL-3 (P = 0.008), IL-7 (P = 0.027), IL-8 (P = 0.042), monocyte chemoattractant proteins (MCP)-3 (P = 0.022), monokine-induced by IFN-gamma (MIG) (P = 0.034) and regulated upon activation normal T-cell express sequence (RANTES) (P = 0.004). Exclusively lower post-stimulative levels of G-CSF (P = 0.030) and GRO-alpha (P = 0.04) were observed in controls in comparison with the basal levels. A significant post-stimulative decrease in G-CSF (P = 0.030) and MCP-2 (P = 0.009) levels was observed in controls in comparison with T1DR neonates. We also observed the interesting impact of the risky genotype on the protein microarray results. Protein microarray seems to be a useful tool to characterize a risk pattern of the immune response for T1D also in newborns.

Immune regulatory T cells in siblings of children suffering from type 1 diabetes mellitus.

Patients with type 1 diabetes are suffering from defects in immune regulatory cells. Their siblings may be at increased risk of type 1 diabetes especially if they are carriers of certain human leucocyte antigen (HLA) alleles. In a prospective non-randomized study, we intended to evaluate 31 healthy siblings of paediatric patients with type 1 diabetes and explore immune regulatory populations of CD4+CD25+ T cells and natural killer (NK) T cells. Tested siblings of type 1 diabetes patients were stratified according to the HLA-associated risk of possible diabetes development. Immune regulatory function of CD4+CD25+ T cells was tested in vitro. Significant differences in CD4+CD25+ but not in NK T cells have been identified. Siblings of type 1 diabetes patients carrying high risk HLA alleles (DQA1*05, DQB1*0201, DQB1*0302) had significantly lower number of immune regulatory CD4+CD25+ T cells than the age-matched healthy controls or siblings carrying low-risk HLA alleles (DQB1*0301, DQB1*0603, DQB1*0602). Regulatory function of CD4+CD25+ T cells demonstrated a dose-escalation effect. In siblings of type 1 diabetes patients, the defect in immune regulatory CD4+CD25+ T cells exists in association with genetic HLA-linked risk for type 1 diabetes.

An in vitro skin explant assay as a predictive assay for graft-versus-host disease in a cohort of pediatric transplants.

Severe acute graft-versus-host disease (GvHD) remains a serious complication of allogeneic stem cell transplantation. An in vitro skin explant assay was used to predict the occurrence and severity of acute GvHD in a cohort of 30 pediatric patients undergoing human leucocyte antigen (HLA)-matched sibling transplants (20 patients) and matched or one antigen mismatched unrelated donor transplants (10 patients). In the cohort of HLA-matched sibling transplants, the result appeared to reflect the degree of GvHD prophylaxis. The skin explant assay correlated with GvHD outcome in 12 of 20 children, but this did not reach statistical significance (chi-square 0.95, d.f.=1, p=0.32). These results support previous observations. In this present cohort, patients were treated either with cyclosporin A (CsA) monotherapy (n=7) or with CsA plus additional methotrexate (MTX) (n=13). We have previously demonstrated that the skin explant assay was not as predictive in patients receiving CsA plus additional MTX compared to cohorts treated with CsA alone. In the group of patients treated with CsA alone, four of five patients (80%) predicted to develop GvHD developed acute GvHD of grade II or above; of two patients predicted to develop only grade 0-I GvHD, one patient developed no GvHD and the other grade II GvHD. In the CsA plus MTX group, nine patients were predicted to develop GvHD. Five of nine (55%) developed acute GvHD of grade II or above, while three of four with grade 0 or I skin explant assay results developed only grade 0-I GvHD. In a cohort of 10 patients who received unrelated donor transplants, the skin explant assay correlated with GvHD outcome in all 10 patients (Fisher's exact test p=0.008). Hence, the skin explant assay is a pretransplant in vitro GvHD predictive test that predicts the occurrence and severity of acute GvHD in pediatric unrelated donor transplants and to varying degrees, depending on the GvHD prophylaxis protocols, in HLA-matched sibling cohorts.

In vitro autoreactivity against skin and synovial cells in patients with juvenile idiopathic and rheumatoid arthritis.

In the present study we compared specific lysis of various autologous target cells in patients with juvenile idiopathic arthritis JIA; n = 8) or rheumatoid arthritis RA; n = 17) with those of healthy controls (n = 15). (51)Cr-release cytotoxic assay with autologous peripheral blood mononuclear cells as effector cells was used. When compared with controls, effector cells of patients with JIA or RA were found to lyse significantly autologous synovial cells (p < 0.0005) and epidermal keratinocytes (p < 0.0005), however, no difference was found for autologous dermal fibroblasts.