Phenotypic and functional characterisation of CCR7+ and CCR7- CD4+ memory T cells homing to the joints in juvenile idiopathic arthritis.

The aim of the study was to characterise CCR7+ and CCR7- memory T cells infiltrating the inflamed joints of patients with juvenile idiopathic arthritis (JIA) and to investigate the functional and anatomical heterogeneity of these cell subsets in relation to the expression of the inflammatory chemokine receptors CXCR3 and CCR5. Memory T cells freshly isolated from the peripheral blood and synovial fluid (SF) of 25 patients with JIA were tested for the expression of CCR7, CCR5, CXCR3 and interferon-gamma by flow cytometry. The chemotactic activity of CD4 SF memory T cells from eight patients with JIA to inflammatory (CXCL11 and CCL3) and homeostatic (CCL19, CCL21) chemokines was also evaluated. Paired serum and SF samples from 28 patients with JIA were tested for CCL21 concentrations. CCR7, CXCR3, CCR5 and CCL21 expression in synovial tissue from six patients with JIA was investigated by immunohistochemistry. Enrichment of CD4+, CCR7- memory T cells was demonstrated in SF in comparison with paired blood from patients with JIA. SF CD4+CCR7- memory T cells were enriched for CCR5+ and interferon-gamma+ cells, whereas CD4+CCR7+ memory T cells showed higher coexpression of CXCR3. Expression of CCL21 was detected in both SF and synovial membranes. SF CD4+ memory T cells displayed significant migration to both inflammatory and homeostatic chemokines. CCR7+ T cells were detected in the synovial tissue in either diffuse perivascular lymphocytic infiltrates or organised lymphoid aggregates. In synovial tissue, a large fraction of CCR7+ cells co-localised with CXCR3, especially inside lymphoid aggregates, whereas CCR5+ cells were enriched in the sublining of the superficial subintima. In conclusion, CCR7 may have a role in the synovial recruitment of memory T cells in JIA, irrespective of the pattern of lymphoid organisation. Moreover, discrete patterns of chemokine receptor expression are detected in the synovial tissue.

Histochemical study of Dom mouse: A model for Waardenburg-Hirschsprung's phenotype.

BACKGROUND/PURPOSE: The spontaneous mouse mutant Dominant megacolon (Dom) represents the model of the Waardenburg-Hirschsprung's disease, a syndromic pathology, characterized by the association of pigmentation defects (PD), deafness, and Hirschsprung's disease (HD). The defect in Dom mouse is caused by a spontaneous mutation of the gene encoding the Sry-related transcription factor Sox10. This mutation affects several aspects of neural crest development leading to combined enteric innervation and pigmentation defects, both in mouse and human. The purpose of this report is to define, by enzymo-histochemical techniques routinely used for the diagnosis of human Hirschsprung's disease (AChE, LDH, NADPH-diaphorase), the innervative patterns of the affected gut. METHODS: Fifty-four siblings of Heterozygous Dom/+ mice underwent autopsy and were genotyped by direct sequencing of polymerase chain reaction (PCR) products for Sox10 mutations. The enteric nervous system of all the mice was studied by histochemical techniques indicated above. RESULTS: Genotyping showed that 43 mice were Dom/+ and 11 were Wild type +/+. Wild-type +/+ mice were used as control. The correspondence between genotype and at least 1 phenotypic aspect (PD or dysganglionosis) was present in 93% of cases (41 of 43). Among the Dom/+ mice, dysganglionosis was present in 79% of cases and PD in 90% of cases. Moreover, among Dom/+ mice, excluding those whose mantle was not evaluated as dead just after birth, PD and dysganglionosis (complete phenotype) were present in 68% of cases. CONCLUSIONS: The histochemical methods that we used proved to be useful for identification of different aganglionic (AG), hypoganglionic (HG), and normoganglionic segments of Dom/+ mouse gut studied in longitudinal sections. Unlike humans, control mice (Wild type +/+) presented a rich component of AChE nerve fibers, whereas Dom/+ mice with dysganglionosis presented a decrease in AChE-positive nerve fibers. These data confirm the variable phenotypic penetrance in heterozygous mice. Because dysganglionosis in this animal model (Dom/+) was evident in 79% of cases (AG or HG), we concluded that Dom mice could represent important models for further experimental studies.

Neuroblastic tumors associated with opsoclonus-myoclonus syndrome: histological, immunohistochemical and molecular features of 15 Italian cases.

The aim of this study was to investigate the histological, immunohistochemical and molecular features of a series of children with neuroblastic tumors (NTs) and opsoclonus-myoclonus syndrome (OMS). Of 1187 children (age 0-15 years) with previously untreated NTs registered between 1979 and 1995, 15 (1.3%) had OMS at presentation. The majority of patients showed favorable biological characteristics, such as lack of amplification of the neuroblastoma-associated avian myelocytomatosis homolog MYCN oncogene and aneuploid nuclear DNA content. Tumor histology was reviewed according to the International Neuroblastoma Pathology Classification. Histology of the 15 cases of NTs with OMS was ganglioneuroblastoma, intermixed, in 10 patients; ganglioneuroma, maturing, in 1; and neuroblastoma in 4. Of 15 tumors, 12 (10 ganglioneuroblastomas, 2 neuroblastomas) showed abundant interstitial or perivascular lymphoid infiltrates, the latter often organized in secondary lymphoid follicles. The three remaining cases had only minimal infiltrates. A review of 91 cases of age- and stage-matched neuroblastic tumors not associated with OMS tested as controls showed that the degree of lymphoid infiltration was significantly lower than that detected in OMS-related tumors. Furthermore, lymphoid follicles were always present in the latter tumors, whereas they were detected only in a few ganglioneuroma, intermixed tumors from the control group. In conclusion, ganglioneuroblastoma, intermixed subtype, lack of MYCN amplification, aneuploid DNA content and presence of lymphoid infiltrates may contribute to favorable prognosis in NTs associated with OMS.

Antitumor activity of a new orally active organotin compound: a preliminary study in murine tumor models.

The toxicity and antitumor activity of the novel organotin compound triethyltin(IV)lupinylsulfide hydrochloride (IST-FS 29), administered by the oral route, have been evaluated against three transplantable murine tumor models: P388 lymphocytic leukemia, B16F10 melanoma and 3LL Lewis lung carcinoma. Mild and reversible signs of acute toxicity such as behavioral symptoms, weight loss and histological alterations were mainly reported at the highest single dose of 28 mg/kg. Conversely, lower concentrations of compound ranging from 7 to 21 mg/kg did not result in major toxic effects, even after repeated dosing. The antitumor activity studies showed that fractionation dosing, rather than single bolus administration, over 1 week, might prove more active and better tolerated by allowing the achievement of the highest therapeutic total dose of IST-FS 29 (42 mg/kg). Indeed, repeated administrations of IST-FS 29 resulted in marked significant improvement of antitumor activity against B16F10 (50% of tumor volume inhibition, p = 0.0003) and, to a greater extent, 3LL (90% of tumor volume inhibition, p = 0.0001) tumors. These results indicate that IST-FS 29 might be a suitable candidate as an orally administrable anticancer drug and support its further development in human tumor xenografts.

Overexpression of vascular endothelial growth factor and its endothelial cell receptor KDR in type 1 leprosy reaction.

The sites of expression of vascular endothelial growth factor (VEGF) and of KDR, its endothelial cell receptor, were investigated in leprosy reaction Type 1, or reversal reaction (RR), by immunohistochemistry and in situ hybridization. In comparison with nonreactional leprosy, overexpression of both VEGF and KDR was seen in granuloma cells, especially epithelioid and foreign body-type giant cells, the epithelium and the vascular endothelium of RR specimens. In granuloma cells, hybridization for VEGF was stronger than immunostaining, a finding that may reflect the rapid turnover of VEGF in an immunologically dynamic situation such as RR. In the epidermis, double immunohistochemistry revealed VEGF overexpression in CDla-positive dendritic cells. The VEGF may not only be relevant for hyperpermeability and mononuclear cell differentiation (the key morphologic features in the acute, clinically evident phase of RR), but it could also be implicated in RR onset, when dendritic cells are activated in response to antigen stimulation.

Glomerular cell replication and cell loss through apoptosis in experimental diabetes mellitus.

AIM: To evaluate changes in the glomerular cell balance between replication and apoptosis in experimental diabetes mellitus (DM) in relation to morphometric data. METHODS: Adult Sprague-Dowley rats with streptozotocin-induced DM and controls of the same age and strain were sacrificed 4 and 8 weeks and 6 months after disease onset. Cell replication was demonstrated with MIB-5, and apoptosis with the terminal uridine nick end labeling technique. Glomerular size and glomerular cell population were estimated morphologically. RESULTS: Diabetic and control rats showed irrelevant MIB-5 positivity at all time points. Glomerular apoptosis was minimal in rats with 4 and 8 weeks of DM and in controls. Rats with 6 months of DM showed significantly higher glomerular apoptosis values than controls (2.49 +/- 0.25 vs. 0.65 +/- 0.16%; p < 0.001). The mean cell count per glomerular profile was significantly lower in these diabetic rats (64.02 +/- 1.93 vs. 78.27 +/- 0.99; p < 0.001), a change that correlated with that in apoptosis. The glomerular cell density was further decreased in diabetic rats because of the diabetic increase in mean glomerular volume (1.598 vs. 0.927 10(6) microm). CONCLUSIONS: Apoptosis is associated with loss of glomerular cells in rats with long-term, streptozotocin-induced DM and - to a considerably lower degree - in controls of the same age and strain. These changes could be relevant to glomerulosclerosis associated with long-term, streptozotocin-induced DM.