The need for a supply of high quality organic vegetable seeds

Production of high quality organic vegetable seeds encounters several challenges. Research is performed to support seed companies in producing vigorous and healthy organic vegetable seeds. Examples are provided with respect to research on seed vigour, determining critical control points to avoid disease transmission to the seeds, seed treatments with natu-ral compounds, new seed sorting techniques and enhancement of the natural plant defence. It is noticed that the restrictions in EU regulation 2092/91 on organic production are at present blocking the use of promising natural and sustainable components for seed treatments

Splicing of the platelet‐derived‐growth‐factor A‐chain mRNA in human malignant mesothelioma cell lines and regulation of its expression

Platelet‐derived‐growth‐factor (PDGF) A‐chain transcripts differing in the presence or absence of an alternative exon‐derived sequence have been described. In some publications, the presence of PDGF A‐chain transcripts with this exon‐6‐derived sequence was suggested to be tumour specific. However, in this paper it was shown by reverse‐transcription polymerase‐chain‐reaction (PCR) analysis that both normal mesothelial cells and malignant mesothelioma cell lines predominantly express the PDGF A‐chain transcript without the exon‐6‐derived sequence. This sequence encodes a cell‐retention signal, which means that the PDGF A‐chain protein is most likely to be secreted by both cell types. In cultured normal mesothelial cells, the secreted PDGF A‐chain protein might be involved in autocrine growth stimulation via PDGF α receptors. However, human malignant mesothelioma cell lines only possess PDGF β receptors. If this also holds true in vivo, the PDGF A‐chain protein produced and secreted by malignant mesothelial cells might have a paracrine function. In a previous paper, we described elevated expression of the PDGF A‐chain transcript in human malignant mesothelioma cell lines, compared to normal mesothelial cells. In this paper, the possible reason for this elevation was studied. First, alterations at the genomic level were considered, but cytogenetic and Southern‐blot analysis revealed neither consistent chromosomal aberrations, amplification nor structural rearrangement of the PDGF A‐chain gene in the malignant cells. Possible differences in transcription rate of the PDGF A‐chain gene, and stability of the transcript between normal and malignant cells, were therefore studied. The presence of a protein‐synthesis inhibitor, cycloheximide, in the culture medium did not significantly influence the PDGF A‐chain mRNA level in normal mesothelial and malignant mesothelioma cell lines. Furthermore, nuclear run‐off analysis showed that nuclear PDGF A‐chain mRNA levels varied in both cell types to the same extent as the levels observed in Northern blots. Taken together, this suggests that increased transcription is the most probable mechanism for the elevated mRNA level of the PDGF A‐chain gene in human malignant mesothelioma cell lines.</p

Splicing of the platelet‐derived‐growth‐factor A‐chain mRNA in human malignant mesothelioma cell lines and regulation of its expression

Platelet‐derived‐growth‐factor (PDGF) A‐chain transcripts differing in the presence or absence of an alternative exon‐derived sequence have been described. In some publications, the presence of PDGF A‐chain transcripts with this exon‐6‐derived sequence was suggested to be tumour specific. However, in this paper it was shown by reverse‐transcription polymerase‐chain‐reaction (PCR) analysis that both normal mesothelial cells and malignant mesothelioma cell lines predominantly express the PDGF A‐chain transcript without the exon‐6‐derived sequence. This sequence encodes a cell‐retention signal, which means that the PDGF A‐chain protein is most likely to be secreted by both cell types. In cultured normal mesothelial cells, the secreted PDGF A‐chain protein might be involved in autocrine growth stimulation via PDGF α receptors. However, human malignant mesothelioma cell lines only possess PDGF β receptors. If this also holds true in vivo, the PDGF A‐chain protein produced and secreted by malignant mesothelial cells might have a paracrine function. In a previous paper, we described elevated expression of the PDGF A‐chain transcript in human malignant mesothelioma cell lines, compared to normal mesothelial cells. In this paper, the possible reason for this elevation was studied. First, alterations at the genomic level were considered, but cytogenetic and Southern‐blot analysis revealed neither consistent chromosomal aberrations, amplification nor structural rearrangement of the PDGF A‐chain gene in the malignant cells. Possible differences in transcription rate of the PDGF A‐chain gene, and stability of the transcript between normal and malignant cells, were therefore studied. The presence of a protein‐synthesis inhibitor, cycloheximide, in the culture medium did not significantly influence the PDGF A‐chain mRNA level in normal mesothelial and malignant mesothelioma cell lines. Furthermore, nuclear run‐off analysis showed that nuclear PDGF A‐chain mRNA levels varied in both cell types to the same extent as the levels observed in Northern blots. Taken together, this suggests that increased transcription is the most probable mechanism for the elevated mRNA level of the PDGF A‐chain gene in human malignant mesothelioma cell lines.</p

Resolved lower limb muscle tone abnormalities in children with HIV encephalopathy receiving standard antiretroviral therapy

BackgroundThis short report arose from a follow-up study of children previously diagnosed with human immunodeficiency virus (HIV) encephalopathy and spastic diplegia and is among the first to describe that increased lower limb muscle tone in children with a confirmed HIV encephalopathy diagnosis may resolve over time in some cases.ResultsOf 19 children previously diagnosed with HIV encephalopathy and increased lower limb muscle tone, some were found to have resolved muscle tone abnormalities during a follow-up physical examination [resolved group, n=13, median age 9years 7months (interquartile range 7years 3months–10years 9months)] whereas others continued to show increased lower limb muscle tone at follow-up [unresolved group, n=6 median age 8years 6months (interquartile range 7years 9months–9years 7months)]. A review of clinical records showed no significant differences in age or follow-up time between the resolved and unresolved groups. However, the unresolved group appeared to have severe disease at an earlier age than the resolved group, based on the age at antiretroviral treatment initiation [median age at start of treatment 2years 3months (interquartile range 7months–5years 3months) vs. 8months (interquartile range 6–12months), p=0.08] and had more severe neurological signs at the initial assessment.ConclusionsIt is anticipated that this information may be of immediate value to those involved in the treatment of children with HIV encephalopathy and increased lower limb muscle tone whilst awaiting the outcome of future controlled clinical trials

Regulation of differential expression of platelet-derived growth factor α-and β-receptor mRNA in normal and malignant human mesothelial cell lines

In earlier studies we showed that the expression patterns of platelet-derived growth factor (PDGF) α- and β-receptors differ between normal and malignant mesothelial cell lines. Normal mesothelial cells predominantly express PDGF α-receptor mRNA and protein, whereas most malignant mesothelioma cell lines produce PDGF P-receptor mRNA and protein. In this paper we studied regulation of this differential PDGF receptor mRNA expression. Such an analysis is of importance in view of the suggested PDGF autocrine activity involving the PDGF β-receptor in mesothelioma cells. The results obtained in this study demonstrate that malignant mesothelioma cell lines are not only capable of PDGF β-receptor transcription but of α-receptor transcription as well, as evidenced from run off analysis and RT-PCR using α-receptor specific primers. However, the fact that PDGF α-receptor mRNA could not be detected by Northern blot analysis, even after cycloheximide treatment, suggests a difference in steady-state PDGF α-receptor mRNA expression levels between normal and malignant mesothelial cell lines, which is likely to be caused by a post-transcriptional mechanism. In normal mesothelial cells a half-life of more than 6 h was observed for PDGF α-receptor mRNA. In the majority of malignant mesothelioma cell lines clear PDGF β-receptor mRNA expression was seen. The half-life of the PDGF β-receptor transcript was at least 6 h in these cells. In contrast, hardly any PDGF β-receptor transcription was observed in run off assays in normal mesothelial cells, suggesting that differences in β-receptor transcriptional initiation most probably account for the inability to clearly detect PDGF β-receptor transcripts in these cells. Transforming growth factor β-1 (TGF-β1), which is being produced in active form by mesothelial cells was evaluated for its potential role in regulation of the differential PDGF receptor expression in these cells. Stimulation with TGF-β1 revealed decreased PDGF α-receptor mRNA expression in normal mesothelial cells. The effect on PDGF β-receptor mRNA in the malignant mesothelioma cell lines was variable. Although the TGF-β1 effect cannot entirely explain the differential PDGF receptor expression pattern, TGF-β1 may nevertheless play a role in downregulation of an (already) low PDGF α-receptor mRNA level in malignant mesothelioma cell lines.</p

Regulation of differential expression of platelet-derived growth factor α-and β-receptor mRNA in normal and malignant human mesothelial cell lines

In earlier studies we showed that the expression patterns of platelet-derived growth factor (PDGF) α- and β-receptors differ between normal and malignant mesothelial cell lines. Normal mesothelial cells predominantly express PDGF α-receptor mRNA and protein, whereas most malignant mesothelioma cell lines produce PDGF P-receptor mRNA and protein. In this paper we studied regulation of this differential PDGF receptor mRNA expression. Such an analysis is of importance in view of the suggested PDGF autocrine activity involving the PDGF β-receptor in mesothelioma cells. The results obtained in this study demonstrate that malignant mesothelioma cell lines are not only capable of PDGF β-receptor transcription but of α-receptor transcription as well, as evidenced from run off analysis and RT-PCR using α-receptor specific primers. However, the fact that PDGF α-receptor mRNA could not be detected by Northern blot analysis, even after cycloheximide treatment, suggests a difference in steady-state PDGF α-receptor mRNA expression levels between normal and malignant mesothelial cell lines, which is likely to be caused by a post-transcriptional mechanism. In normal mesothelial cells a half-life of more than 6 h was observed for PDGF α-receptor mRNA. In the majority of malignant mesothelioma cell lines clear PDGF β-receptor mRNA expression was seen. The half-life of the PDGF β-receptor transcript was at least 6 h in these cells. In contrast, hardly any PDGF β-receptor transcription was observed in run off assays in normal mesothelial cells, suggesting that differences in β-receptor transcriptional initiation most probably account for the inability to clearly detect PDGF β-receptor transcripts in these cells. Transforming growth factor β-1 (TGF-β1), which is being produced in active form by mesothelial cells was evaluated for its potential role in regulation of the differential PDGF receptor expression in these cells. Stimulation with TGF-β1 revealed decreased PDGF α-receptor mRNA expression in normal mesothelial cells. The effect on PDGF β-receptor mRNA in the malignant mesothelioma cell lines was variable. Although the TGF-β1 effect cannot entirely explain the differential PDGF receptor expression pattern, TGF-β1 may nevertheless play a role in downregulation of an (already) low PDGF α-receptor mRNA level in malignant mesothelioma cell lines.</p

Expression of the wilms' tumor gene WT1 in human malignant mesothelioma cell lines and relationship to platelet‐derived growth factor A and insulin‐like growth factor 2 expression

Mutations in the WT1 tumor suppressor gene are known to contribute to the development of Wilms' tumor (WT) and associated gonadal abnormalities. WT1 is expressed principally in the fetal kidney, developing gonads, and spleen and also in the mesothelium, which lines the coelomic cavities. These tissues develop from mesenchymal components that have subsequently become epithelialized, and it has therefore been proposed that WT1 may play a role in this transition of cell types. To test the possible involvement of this gene in malignant mesothelioma, we have first studied its expression in a panel of human normal and malignant mesothelial cell lines. WT1 mRNA expression levels varied greatly between the cell lines and no specific chromosomal aberration on 11p, which could be related to the variation in WT1 expression in these cell lines, was observed. Furthermore, no gross deletions, rearrangements, or functionally inactivating point mutations in the WT1 coding region were identified. All four WT1 splice variants were observed at similar levels in these cell lines. The WT1 gene encodes a zinc‐finger transcription factor and the four protein isoforms are each believed to act as transcriptional repressors of certain growth factor genes. Lack of WT1 expression is thus predicted to result in growth stimulation of tumor cells. Binding of one particular WT1 isoform construct to the insulin‐like growth factor 2 (IGF2) and platelet‐derived growth factor A (PDGFA) gene promoters has been demonstrated to result in repression of these genes in transient transfection studies. Analysis of IGF2 and PDGFA mRNA expression levels compared with WT1 mRNA expression levels failed to demonstrate an inverse correlation in the mesothelial cell lines, which endogenously express these genes. Finally, the putative role of WT1 in the transition of cell types was investigated. No obvious correlation between WT1 expression levels and cell morphology of the malignant mesothelial cell lines was evident from this study. Moreover, no change in WT1 expression was observed in normal mesothelial cells which were, by alteration of culture conditions, manipulated to switch from the mesenchymal to epithelial morphology.</p

Expression of the wilms' tumor gene WT1 in human malignant mesothelioma cell lines and relationship to platelet‐derived growth factor A and insulin‐like growth factor 2 expression

Mutations in the WT1 tumor suppressor gene are known to contribute to the development of Wilms' tumor (WT) and associated gonadal abnormalities. WT1 is expressed principally in the fetal kidney, developing gonads, and spleen and also in the mesothelium, which lines the coelomic cavities. These tissues develop from mesenchymal components that have subsequently become epithelialized, and it has therefore been proposed that WT1 may play a role in this transition of cell types. To test the possible involvement of this gene in malignant mesothelioma, we have first studied its expression in a panel of human normal and malignant mesothelial cell lines. WT1 mRNA expression levels varied greatly between the cell lines and no specific chromosomal aberration on 11p, which could be related to the variation in WT1 expression in these cell lines, was observed. Furthermore, no gross deletions, rearrangements, or functionally inactivating point mutations in the WT1 coding region were identified. All four WT1 splice variants were observed at similar levels in these cell lines. The WT1 gene encodes a zinc‐finger transcription factor and the four protein isoforms are each believed to act as transcriptional repressors of certain growth factor genes. Lack of WT1 expression is thus predicted to result in growth stimulation of tumor cells. Binding of one particular WT1 isoform construct to the insulin‐like growth factor 2 (IGF2) and platelet‐derived growth factor A (PDGFA) gene promoters has been demonstrated to result in repression of these genes in transient transfection studies. Analysis of IGF2 and PDGFA mRNA expression levels compared with WT1 mRNA expression levels failed to demonstrate an inverse correlation in the mesothelial cell lines, which endogenously express these genes. Finally, the putative role of WT1 in the transition of cell types was investigated. No obvious correlation between WT1 expression levels and cell morphology of the malignant mesothelial cell lines was evident from this study. Moreover, no change in WT1 expression was observed in normal mesothelial cells which were, by alteration of culture conditions, manipulated to switch from the mesenchymal to epithelial morphology.</p

Pitfalls in TCR gene clonality testing: teaching cases

Clonality testing in T-lymphoproliferations has technically become relatively easy to perform in routine laboratories using standardized multiplex polymerase chain reaction protocols for T-cell receptor (TCR) gene analysis as developed by the BIOMED-2 Concerted Action BMH4-CT98-3936. Expertise with clonality diagnostics and knowledge about the biology of TCR gene recombination are essential for correct interpretation of TCR clonality data. Several immunobiological and technical pitfalls that should be taken into account to avoid misinterpretation of data are addressed in this report. Furthermore, we discuss the need to integrate the molecular data with those from immunohistology, and preferably also flow cytometric immunophenotyping, for appropriate interpretation. Such an interactive, multidisciplinary diagnostic model guarantees integration of available data to reach the most reliable diagnosis