Malaria transmission and naturally acquired immunity to PfEMP-1

Why there are so few gametocytes (the transmission stage of malaria) in the blood of humans infected with Plasmodium spp. is intriguing. This may be due either to reproductive restraint by the parasite or to unidentified gametocyte-specific immune-mediated clearance mechanisms. We propose another mechanism, a cross-stage immunity to Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP-1). This molecule is expressed on the surface of the erythrocyte infected with either trophozoite or early gametocyte parasites. Immunoglobulin G antibodies to PfEMP-1, expressed on both life cycle stages, were measured in residents from an area where malaria is endemic, Papua New Guinea. Anti-PfEMP-1 prevalence increased with age, mirroring the decline in both the prevalence and the density of asexual and transmission stages in erythrocytes. These data led us to propose that immunity to PfEMP-1 may influence malaria transmission by regulation of the production of gametocytes. This regulation may be achieved in two ways: (i) by controlling asexual proliferation and density and (ii) by affecting gametocyte maturation.     

Phenotype, cytotoxic, and helper functions of T cells from varicella zoster virus stimulated cultures of human lymphocytes

Blood mononuclear cells (MNC) were stimulated with VZV in the form of live cell-associated virus, glutaraldehyde-fixed VZV-infected fibroblasts or an extracted VZV antigen. After 7 days of culture with live virus, IL2 receptors (IL2R) were found on CD4 and CD8 cells while cultures stimulated with fixed or extracted antigen had IL2R only on CD4 cells. Clones derived by limiting dilution from these cultures were tested for specificity by cytotoxicity to autologous VZV-superinfected B lymphoblasts, and by proliferation in the presence of antigen and antigen presenting cells. The CD8+ clones were not VZV specific in tests of cytotoxicity or proliferation. 50% of the CD4+ clones with specificity for VZV also proliferated in cultures stimulated with individual VZV glycoproteins gp I, II or III. Included amongst these were clones cytotoxic for lymphoblast targets prepared with live VZV. Most of the VZV-specific T cell clones which failed to lyse targets prepared with live VZV lysed targets prepared with heat killed VZV. Target cells were susceptible to lysis after VZV antigens were no longer demonstrable on the cell surface by immunofluorescence and monoclonal antibodies to VZV glycoproteins failed to block cytotoxicity. 5 of 8 VZV-specific CD4+ clones provided antigen-specific help to B cells for IgG antibody production. The data are consistent with the view that CD4+ T cells respond to processed VZV antigen fragments, and that these fragments include epitopes present on gp I, gp II and gp III. Additionally, some the cloned progeny of VZV specific T cells were bifunctional (expressing cytotoxicity and help for B cells) in tissue culture.     

Gain of 1q and loss of 22 are the most common changes detected by comparative genomic hybridisation in paediatric ependymoma

Ependymomas are the third most common brain tumour in the paediatric population. Although cytogenetic and molecular analyses have pinpointed deletions of chromosomes 6q, 17, and 22 in a subset of tumours, definitive patterns of genetic aberrations have not been determined. In the present study, we analysed 40 ependymomas from paediatric patients for genomic loss or gain using comparative genomic hybridisation (CGH). Eighteen of the tumours (45%) had no detectable regions of imbalance. In the remaining cases, the most common copy number aberrations were loss of 22 (25% of tumours) and gain of 1q (20%). Three regions of high copy number amplification were noted at 1q24-31 (three cases), 8q21-23 (two cases), and 9p (one case). Although there was no association with the loss or gain of any chromosome arm or with benign versus anaplastic histologic characteristics, the incidence of gain of 7q and 9p and loss of 17 and 22 was significantly higher in recurrent versus primary tumours. This study has identified a number of chromosomal regions that may contain candidate genes involved in the development of different subgroups of ependymoma.     

Use of tissue recombination to predict phenotypes of transgenic mouse models of prostate carcinoma

Transgenic mouse models of cancer represent a powerful approach for exploring disease processes and testing potential therapeutic interventions. Currently, it is difficult to predict if a specific genetic manipulation will result in a desirable phenotype. The present study tests the idea that tissue recombinants recapitulate the pathologic features of the neoplastic prostate seen in transgenic mice, and would thus be suitable predictive models for new mouse design. The large probasin-large T-antigen (LPB-Tag) transgenic lines 12T-7f and 12T-10 were used as a basis for this study. Tissue recombinants of bladder epithelium (BlE) and urogenital sinus mesenchyme (UGM) were implanted under the renal capsule of athymic mice. Recombinants composed of BlE from 12T-10 LPB-Tag and wild-type (wt) UGM faithfully recapitulated the histopathologic and temporal features of intact transgenic mice of this line. Tissue recombinants using BlE from 12T-7f mice and wt UGM developed epithelial proliferation with atypia that lacked the associated hypercellular stroma seen in the intact 12T-7f line. Recombinants using 12T-7f UGM demonstrated that the hypercellular stroma results from stromal cell expression of the SV40 large T antigen. Corresponding to the recombinant phenotypes, stromal Tag immunostaining was observed in prostate tissues from intact 12T-7f but not 12T-10 mice. Similar stromal expression of Tag was also noted in the hypercellular TRAMP prostatic stroma. Further analysis revealed a previously unreported pattern of SV40T expression in the LADY and TRAMP models including ductus deferens and seminal vesicle stroma as well as region and cell type-specific patterns in the epididymis. The present study demonstrates the utility of using tissue recombination to explore organ-specific phenotypes. Recombination strategies should enable quick and cost-effective screening for likely phenotypes in transgenic animals. This comparison of tissue recombination to existing models shows that this approach can elicit new information on well-characterized models.     

Expression and localization of mutant p16 proteins in melanocytic lesions from familial melanoma patients

Little is known about the correlation between the loss of p16 expression and tumor progression in familial melanoma; no systematic study has been conducted on p16 expression in melanocytic tumors from patients carrying germline CDKN2A mutations. We analyzed 98 early primary lesions from familial patients, previously tested for germline CDKN2A status, by quantitative immunohistochemistry using 3 p16 antibodies. We found that p16 expression was inversely correlated with tumor progression and was significantly lower in melanomas, including in situ lesions, than in nevi. Of other features analyzed, tumor thickness showed the most significant correlation with p16 levels. Lesions from mutation-negative patients displayed combined nuclear and cytoplasmic staining. However, some mutation-positive lesions (ie, G101W, 113insR, M53I, R24P, and 33ins24), including benign nevi, showed nuclear mislocalization, confirming previous studies suggesting that subcellular distribution indicates functional impairment of p16.     

Germline mutations of the CDKN2 gene in UK melanoma families

Germline mutations in CDKN2 on chromosome 9p21, which codes for the cyclin D kinase inhibitor p16, and more rarely, mutations in the gene coding for CDK4, the protein to which p16 binds, underlie susceptibility in some melanoma families. We have sequenced all exons of CDKN2 and analysed the CDK4 gene for mutations in 27 UK families showing evidence of predisposition to melanoma. Five different germline mutations in CDKN2 were found in six families. Three of the mutations (Met53Ile, Arg24Pro and 23ins24) have been reported previously. We have identified two novel CDKN2 mutations (88delG and Ala118Thr) which are likely to be associated with the development of melanoma, because of their co-segregation with the disease and their likely functional effect on the CDKN2 protein. In binding assays the protein expressed from the previously described mutation, Met53Ile, did not bind to CDK4/CDK6, confirming its role as a causal mutation in the development of melanoma. Ala118Thr appeared to be functional in this assay. Arg24Pro appeared to bind to CDK6, but not to CDK4. No mutations were detected in exon 2 of CDK4, suggesting that causal mutations in this gene are uncommon. The penetrance of these mutant CDKN2 genes is not yet established, nor is the risk of non-melanoma cancer to gene carriers.      

Persistence of varicella-zoster virus DNA in blood mononuclear cells of patients with varicella or zoster

Varicella-zoster virus (VZV) DNA was detectable by in-situ hybridization in blood mononuclear cells (MNCs) of patients with varicella or zoster for 2–56 days after the onset of a rash. VZV DNA was present in many MNCs from one acute varicella patient 2 days after the onset of the rash and was rarely found in MNCs during acute zoster, convalescent zoster, and convalescent varicella. The morphology of MNCs containing VZV was heterogenous, although most viral-DNA-containing MNCs were large monocytoid cells. Serial examination of blood MNCs from one adult with varicella revealed VZV DNA up until 8 weeks, but not 16 weeks, after the appearance of the rash; parallel studies in four zoster patients showed VZV DNA up until 3 weeks, but not later than 7 weeks after the appearance of the rash. These results indicate that MNCs become infected with VZV during the primary encounter with VZV (varicella) and during reactivation (zoster) and that infection continues for weeks after the onset of the skin rash. Furthermore, the detection of VZV DNA in blood MNCs of uncomplicated zoster patients coincides with the period during which these patients experience pain.