Alessandra Fierabracci

Long term follow up of children with myocarditis treated by immunosuppression and of children with dilated cardiomyopathy

Objective: To describe the treatment and long term outcome after immunosuppressive treatment of children with myocarditis. Methods and results: 114 patients with newly diagnosed dilated cardiomyopathy were divided into three groups, according to the histological pattern: group A, acute myocarditis; group B, borderline myocarditis; and group C, non-inflammatory cardiomyopathy. Groups A and B were treated with cyclosporine and prednisone in addition to conventional treatment. Survivors of the whole cohort were analysed for 13 year transplant-free survival and assessed for left ventricular function. Event-free survival at 13 years was 97 (3)% for group A, 70 (8)% for group B, and 32 (7)% for group C (p < 0.0001). It was 96 (4)% at one year and 83 (5)% at 13 years for the cumulated myocarditis group (A and B). Cardiac function recovered completely in 79% of survivors in group A, 64% in group B, and 36% in group C. The rate of complete recovery in the cumulated group (A and B) was 70%. Conclusions: The high long term survival rate of this cohort of children with myocarditis is probably due to the effect of short term immunosuppression. This result differs from previously published series of conventionally treated children, whose survival probability at one year was about 60%.

Retroviral superantigens and type 1 diabetes mellitus

Matters Arising diagnosed for less than three weeks), and four healthy Retroviral Superantigens and Type 1 adult controls. All subjects were of Caucasoid extrac-Diabetes Mellitus tion. RT-PCRs, with the Conrad et al. (1997) protocol and reagents, generated a band of the expected size (489 bp) for IDDMK 1,2 22 with U3-R-poly(A) primer from In 1994, Conrad et al. reported initial results that rekin-all samples, but in both the presence and absence of dled the debate about the involvement of an infectious RT. The U3-R-poly(A) primer also generated a 489 bp agent in type 1 diabetes. They analyzed the T cell (anti-band from genomic DNA samples. The RT-PCR product gen) receptor repertoire of T lymphocytes that had in-hybridized with the U3-R probe, as reported by Conrad vaded the pancreatic islets of two recently deceased, et al. (1997). However, DNase treatment of the RNA acutely diabetic patients. Antigen receptors carrying samples prevented generation of the PCR product and the V␤7 variable region were shown to be strikingly en-its detection by Southern blot. We could not achieve riched in the T cells, evoking the involvement of a super-specificity for RNA, by extending 5Ј the U3-R-poly(A) antigen (SAG). Recently, Conrad et al. (1997) (see also primer from four to 11Ts and/or by subtracting eight of Benoist and Mathis, 1997) described the isolation of a the 3Ј nucleotides, whereas the Pittsburgh and London hitherto unknown endogenous HERV-K10-like retroviral laboratories (see below) detected viral RNAs with modi-genome, IDDMK 1,2 22, related to mouse mammary tumor fied primers, although these were also not disease spe-virus, whose env gene encoded a SAG. The SAG stimu-cific. It should be noted, however, that the various modi-lated V␤7-bearing T cells, but not those that expressed fied primers were not identical. other V␤ genes. Transcripts derived from, or at least Conrad et al. (1997) added RT and PCR reagents to related to, the IDDMK 1,2 22 genome were detectable the same reaction tube. When we added the PCR re-by RT-PCR in the plasma of 10/10 newly diagnosed type 1 agents after the RT reactions, the yield of the U3-R-diabetes patients, but not from 10 age-matched, nondia-poly(A) product significantly increased, but again it was betic control individuals. In order to detect IDDMK 1,2 22 not specific for RNA. The increased yield was most likely RNA, Conrad et al. (1997) performed RT-PCR on total due to separation of Pwo, a DNA polymerase with 3Ј–5Ј …

Identification of an adult stem/progenitor cell-like population in the human thyroid

There is evidence that tissue-specific stem cells reside in certain adult tissues. Their specific properties remain elusive, because they are rare and heterogeneous in parent tissues; furthermore, technical difficulties have been encountered in the identification and characterization of their progeny. The aim of this study was to isolate stem/progenitor cells from the human thyroid. We devised a method based on the enzymatic digestion of fresh surgical thyroid specimens, followed by culture of cells in the presence of epidermal growth factor and basic fibroblast growth factor. We also used markers that identify and characterize these cells. Spheroids with self-replicative potential were obtained from all thyroid specimens. The isolated population contained a subset of CD34+ CD45- cells and it was able, in differentiation conditions, to generate follicles with thyroid hormonal production. In support of the plasticity concept, we obtained evidence that, when most freshly isolated spheroids were co-cultured with a neuroblastoma cell line, they produced progeny expressing the neuronal marker beta-tubulin III. Spheroids were also able to undergo adipogenic differentiation when cultured in adipogenic medium. We conclude that a predominant functional type of stem/progenitor cell exists within the thyroid, with an intrinsic ability to generate thyroidal cells and the potential to produce non-thyroidal cells.

Identifying thyroid stem/progenitor cells: advances and limitations

Continuing advances in stem cell science have prompted researchers to envisage the potential application of stem cells for the management of several debilitating disorders, thus raising the expectations of transplant clinicians. In particular, in order to find a source of adult stem cells alternative to embryonic stem cells (ESCs) for the exploration of novel strategies in regenerative medicine, researchers have attempted to identify and characterise adult stem/progenitor cells resident in compact organs, since these populations appear to be responsible for physiological tissue renewal and regeneration after injury. In particular, recent studies have also reported evidence for the existence of adult stem/progenitor cell populations in both mouse and human thyroids. Here, I provide a review of published findings about ESC lines capable of generating thyroid follicular cells, thyroid somatic stem cells and cancer stem cells within the thyroid. The three subjects are analysed by also considering the criticism recently raised against their existence and potential utility. I comment specifically on the significance of resident thyroid stem cells in the developmental biology of the gland and their putative role in the pathogenesis of thyroid disorders and on the protocols employed for their identification. I finally provide my opinion on whether from basic science results obtained to date it is possible to extrapolate any convincing basic for future treatment of thyroid disorders.

Identification and characterization of adult stem/progenitor cells in the human bladder (bladder spheroids): perspectives of application in pediatric surgery.

There is evidence that tissue-specific stem cells reside in certain adult tissues. Their specific properties remain elusive, because they are rare in parent tissues and heterogeneous; furthermore, technical difficulties have been encountered in their identification and the characterization of their progeny. The aim of this study was to isolate stem/progenitor cells from the human bladder. We have devised a method for isolating stem/progenitor cells from the human bladder. This is based on the enzymatic digestion of fresh surgical bladder specimens, followed by culture of cells in the presence of EGF and bFGF. We also used markers that identified and finally characterized these cells. Spheroids with self-replicative potential were obtained from all bladder specimens. The isolated population contained a subset of CD34+ CD45- cells. These spheroids represent a predominant functional type of stem/progenitor cells within the human bladder. This envisage their potential use for the treatment of animal models in pediatric surgery.

OSTEOPONTIN IS AN AUTOANTIGEN OF THE SOMATOSTATIN CELLS IN HUMAN ISLETS: IDENTIFICATION BY SCREENING RANDOM PEPTIDE LIBRARIES WITH SERA OF PATIENTS WITH INSULIN-DEPENDENT DIABETES MELLITUS

Random peptide libraries (RPLs) screening with IDDM sera has identified 5 disease-specific mimotopes displayed on phage (phagotopes). We characterised one phagotope (CH1p), by raising a rabbit antibody against the peptide insert on phage, which was employed in immunohistochemistry, Western blotting and cDNA libraries screening. The CH1p mimotope was detected in somatostatin cells of human islets and experimentally raised anti-osteopontin antibodies or human sera positive for the phagotope, detected a similar subpopulation of islet cells. The screening of cDNA library identified a clone corresponding to human osteopontin. In summary, RPLs proved to be successful in the identification of a novel islet-related autoantigen (osteopontin), whose significance in disease remains to be established.

The potential of multimer technologies in type 1 diabetes prediction strategies

Type 1 diabetes is an autoimmune disease which occurs in (human leukocyte antigen) genetically predisposed individuals as a consequence of the organ‐specific immune destruction of the insulin‐producing β cells in the islets of Langherans within the pancreas. Type 1 diabetes is the result of a breakdown in immune regulation that leads to expansion of autoreactive CD4+ and CD8+ T cells, autoantibody‐producing B lymphocytes and activation of the innate immune system.

Autoimmune Polyendocrinopathy-Candidiasis-Ectodermal Dystrophy: Report of Seven Additional Sicilian Patients and Overview of the Overall Series from Sicily

Background: Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a rare recessive inherited disease caused by the mutation of the AIRE gene on chromosome 21. To date, 8 Sicilian patients have been described and the R203X AIRE mutation was found to be the most common in this region. Aims: (1) To describe 7 additional Sicilian APECED patients and to review all 15 Sicilian APECED patients who have been investigated by our group in the last years, and (2) to report a novel AIRE gene mutation. Results: Among the 3 cardinal features of APECED, hypoparathyroidism has been already detected in all 15 patients, whereas Addisons disease and chronic mucocutaneous candidiasis have so far been found in 10/15 and 12/15 cases, respectively. In 2 consanguineous cases, AIRE gene analysis revealed a novel mutation, named IVS13+2T, in homozygosis. R203X was the most common mutation in this region (30% of alleles and 46.6% of patients), followed by R257X (20% of alleles and 40% of patients). Conclusions: Sicilian APECED patients are confirmed to have some peculiar characteristics from a clinical and genetic point of view. No correlations between genotype and phenotype were identified.

Type 1 diabetes in autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy syndrome (APECED): A “Rare” manifestation in a “Rare” disease

Type 1 autoimmune polyglandular syndrome (APS1) is a rare autosomal recessive disease, caused by mutations in the autoimmune regulator gene (AIRE); the encoded Aire protein plays an important role in the establishment of the immunological tolerance acting as a transcriptional regulator of the expression of organ-specific antigens within the thymus in perinatal age. While a high prevalence for this rare syndrome is reported in Finland and Scandinavia (Norway), autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy syndrome (APECED) cohorts of patients are also detected in continental Italy and Sardinia, among Iranian Jews, as well as in other countries. The syndrome is diagnosed when patients present at least two out of the three fundamental disorders including chronic mucocutaneous candidiasis, hypoparathyroidism, and Addison’s disease. Among the associated conditions insulin-dependent diabetes mellitus (Type 1 diabetes) has been rarely reported in different series of patients and occurring more frequently in Finnish APECED patients. In this review, we analyze the incidence of Type 1 diabetes as a clinical manifestation of APECED in different populations highlighting the peculiar genetic and immunological features of the disease when occurring in the context of this syndrome.

Detection of GAD65 Autoreactive T-Cells by HLA Class I Tetramers in Type 1 Diabetic Patients

Type 1 diabetes (T1D) is an autoimmune disease, in which pancreatic β cells are destroyed in genetically predisposed individuals. While the direct contribution of autoantibodies to the disease pathogenesis is controversial, it is generally recognised that the mechanism of β cell destruction is mediated by autoreactive T cells that had escaped the thymic selection. We aimed to design a method to detect circulating CD8+ T cells autoreactive against an epitope of the glutamic acid decarboxylase autoantigen, isoform 65 (GAD65) ex vivo in T1D patients by using HLA class I tetramers. Low frequencies of GAD65 peptide-specific CD8+ cytotoxic T lymphocytes were detected in peripheral blood lymphocytes (PBMC) of normal controls after GAD65 peptide-specific stimulation. Conversely, their frequencies were significantly higher than in controls in PBMC of T1D patients after GAD65 peptide stimulation. These preliminary data are encouraging in order to develop a reliable assay to be employed in large-scale screening studies.