Gabriele Stumm

Differential Transcription and Translation of Immediate Early Genes in the Gerbil Hippocampus after Transient Global Ischemia

Excitotoxic activation of glutamate receptors is thought to be a key event for the molecular pathogenesis of postischemic delayed neuronal death of CA-1 neurons in the gerbil hippocampus. Glutamate receptor stimulation also causes induction of transcription factors that belong to the class of immediate early genes. We examined the expression of six different immediate early genes in the gerbil hippocampus after transient global ischemia. Comparative analysis of c-fos and Krox-24 expression was carried out in the same animals at the transcriptional and translational level by in situ hybridization and immunocytochemistry. Postischemic synthesis of four additional immediate early gene (IEG)–encoded proteins (FOS-B, c-JUN, JUN-B, and JUN-D) was investigated by immunocytochemistry at recirculation intervals between 1 and 48 h. After 5 min of ischemia, transcription of c-fos and Krox-24 mRNA was induced in all hippocampal subpopulations with peak expression at 1 h after recirculation. In vulnerable CA-1 neurons, increased transcription of c-fos and Krox-24 was not followed by translation into protein. Induction of immediate early gene-encoded proteins was restricted to neuronal populations less vulnerable to brief ischemia and identified neurons that are targets of glutamate receptor-mediated neurotoxicity but that are destined to survive. Our data indicate an asynchronous synthesis and persistence of individual IEG-encoded proteins in these neurons. The staggered induction implies that combinatorial changes of transcription factors allow a differential postischemic regulation of target gene expression both spatially and over time.

A missense mutation in the WD40 domain of murine Lyst is linked to severe progressive Purkinje cell degeneration

Disturbance of intracellular trafficking plays a major role in several neurodegenerative disorders including Alzheimer or Parkinson’s disease. The Chediak–Higashi syndrome (CHS), a life-threatening autosomal recessive disease with frequent mutations in the LYST gene, and its animal model, the beige mouse, are both characterized by lysosomal defects with accumulation of giant lysosomes. Clinically they manifest as hypopigmentation, abnormal bleeding and increased susceptibility to infection with various degrees of involvement of the nervous system. In the course of a recessive N-ethyl-N-nitrosurea (ENU) mutagenesis screen, we identified the first murine missense mutation in the lysosomal trafficking regulator gene (LystIng3618) located at a highly conserved position in the WD40 protein domain. Nearly all described human Lyst alleles lead to protein truncation and fatal childhood CHS. Only four different missense mutations have been reported in patients with adolescent or adult forms of CHS involving the nervous system. Interestingly, the LystIng3618 model presents with a predominant neurodegenerative phenotype with progressive degeneration and loss of Purkinje cells and lacks severe impairment of the immune system. Therefore, the LystIng3618 allele could represent a new model for adult CHS with neurological impairment. It could also provide an important tool to elucidate the role of neuronal lysosomal trafficking in the pathophysiology of neurodegeneration.

Expression of the c-erbB-2-encoded oncoprotein and progesterone receptor in human meningiomas

The present study investigated the expression of c-erbB-2 in 59 meningiomas, including different histological subtypes and anaplastic variants, by immunocytochemistry and molecular biological techniques. Immunohistochemistry using the monoclonal antibody FWP-51 directed against c-erbB-2-encoded oncoprotein gp185 demonstrated variable degrees of immunoreactivity in all meningiomas. The intensity of immunostaining correlated with the degree of expression as assessed by Western analysis in 28 meningiomas using polyclonal antiserum 21N. There was no correlation between the degree of expression and histological variants. Immunoreactivity of all menigiomas was distinctly less intense, however, than that of the human breast cancer cell line SK-BR-3, and slightly lower than that of brain metastases of breast and ovarian carcinomas that served as positive controls for both methods. By Southern analysis all meningiomas showed a single copy of the c-erbB-2 gene. Non-neoplastic arachnoid cap cells also exhibited c-erbB-2 expression and the degree of immunoreactivity was comparable with the majority of meningiomas. These data argue against an overexpression of c-erbB-2 in meningiomas, but rather indicate a cell-type-specific constitutive expression of the c-erbB-2 gene product in meningiomas and their putative progenitor cells. Since a subgroup of meningiomas is known to express progesterone receptors (PR), gp185 immunoreactivity was compared to the hormone receptor status using monoclonal antibody KD68. Fifty-six percent meningiomas showed PR immunoreactivity, but there was no statistically significant correlation with the degree of gp185 expression.

Quantitative DNA analysis of an intracerebrally transplanted brain tumour model after experimental chemotherapy with BCNU and CCNU

In the present study we investigated the susceptibility of high passages of the rat glial transplantation tumour G-XIII to chemotherapy using nitrosourea compounds. We observed a significant increase in lifespan (ILS) of animals treated with BCNU (37%, p < 0.01) and CCNU (27%, p < 0.01). There were no difference in the efficiency between these two substances. Using a semi-quantitative score system no histopathological changes were observed which were associated with the response to therapy. The only predicative parameter in the present study was the quantitative DNA distribution pattern. There was a close correlation between treatment and the occurrence of unimodal DNA distribution patterns indicating clonal regrowth of recurrent tumours. Moreover, we also observed a correlation of the DNA distribution pattern of recurrent tumours with the result of experimental chemotherapy. Survival times of animals suffering from tumours containing unimodal DNA histogram was significantly longer than survival times of rats with multimodal DNA distribution, i.e. bimodal or broad DNA histograms. A unimodal, near-diploid stem line was only present in treated animals suggesting that these clones are more resistant against therapy using nitrosourea compounds. Our data indicate DNA cytophotometry as comprehensive tool for the monitoring of therapy response and the design of experimental chemotherapy using rat glial tumours.