Manuela Födinger

Methylenetetrahydrofolate reductase (MTHFR) plays a central role in the folate cycle and contributes to the metabolism of the amino acid homocysteine. It catalyzes the reduction of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate, thus generating the active form of folate required for remethylation of homocysteine to methionine. Deficiency of MTHFR may be associated with an increase in plasma homocysteine, which in turn is associated with an increased risk of vascular disease. This article summarizes the biochemistry, the function in the folate cycle, and the molecular genetics of this enzyme. Particular emphasis has been given to the role of two common polymorphisms (MTHFR 677C-->T, 1298A-->C) in cardiovascular disease, cerebrovascular disease, venous thrombosis, longevity, neural tube defects, pregnancy/preeclampsia, diabetes, cancer, psychiatry, renal failure and renal replacement therapy. Finally, the rare genetic defects underlying severe MTHFR deficiency are also considered.

Background: Patients receiving recombinant human erythropoietin (rHuEPO) may experience side effects arising from the vascular system. The underlying mechanisms, how-ever, are largely unknown. Methods: To elucidate downstream events following eryth-ropoietin receptor triggering, a differential display analysis of human vascular endothelial cell mRNA was performed. Results: We identified eight genes that were upregulated by rHuEPO as confirmed in two further independent cell culture experiments using a semiquantitative reverse tran-scriptase polymerase chain reaction (RT-PCR) protocol. The genes coded for proteins that may be assigned to four differ-ent groups: 1) proteins implicated in the regulation of vas-cular functions (thrombospondin-1, 20 kDa myosin regula-tory light chain; relative increase of rHuEPO induced mRNA levels: 155.2%, P=0.043; 137.6%, P=0.046, respectively); 2) gene products involved in gene transcription and/or transla-tion (c-myc purine-binding transcription factor PuF, trypto-phanyl-tRNA synthetase, S19 ribosomal protein; increase of mRNA levels: 126.4%, P=0.032; 150.9%, P=0.012; 134.9%, P=0.038); 3) subunits of mitochondrial enzymes related to energy transfer (NADH dehydrogenase subunit 6, cyto-chrome C oxidase subunit 1; increase of mRNA concentra-tions: 141.7%, P=0.007; 140.3%, P=0.01); and 4) regulators of signal transduction (protein tyrosine phosphatase G1, in-crease of transcript level: 160.3%, P=0.016). Conclusions: We report on novel molecular downstream events following rHuEPO receptor triggering of human vas-cular endothelial cells. We identified EPO-responsive imme-diate-early genes, coding for proteins involved in vascular functions, gene transcription, and/or translation, energy transfer, and signal transduction. Thus, our data provide new insights into the molecular changes induced by EPO in human vascular endothelial cells.

Recent genetic studies have led to the characterization of molecular determinants contributing to the pathogenesis of hyperhomocysteinemia. In this article we summarize the current insights into the molecular genetics of severe, moderate and mild hyperhomocysteinemia. We will consider deficiencies of the trans-sulfuration enzyme cystathionine β-synthase (gene symbol: CBS), and the disturbances of the remethylation enzymes 5,10-methylenetetrahydrofolate reductase (gene symbol: MTHFR), methionine synthase (gene symbol: MTR), and the recently identified methionine synthase reductase (gene symbol: MTRR). Furthermore, we will focus on clinically important genetic polymorphisms which are highly prevalent and thus of potential general interest.

The number of characterized monogenic and polygenic diseases is rising each year. In consequence, molecular diagnostics is faced with an ever increasing number of patient samples and with more and more heterogeneous genetic defects. The fusion of microelectronics and molecular biology has created a new technology (microelectronic miniaturization), which provides a rapid, efficient, and cost-effective tool in molecular diagnostics at a high-sample throughput. The biochip has recently been selected as one of the ten scientific highlights in the year 1998. The application of microelectronics ranges from the polymerase chain reaction (PCR), nucleotide sequence analysis via DNA-chips or capillary electrophoresis-chips to gene expression analysis. These microchips are suited for integration into fully automated systems, thus providing the basis for automation of molecular diagnostics. The present article summarizes important trends in molecular diagnostics and provides a glimpse on future technologies.

Background: Mutations in the carboxy termini of the beta subunit (hβENaC) and the gamma subunit (hγENaC) of the human epithelial sodium channel have been identified in patients with Liddle syndrome. Moreover polymorphisms have been described in these genes, the clinical relevance of which for progression to end-stage renal disease (ESRD) is unknown. We, therefore, have screened ESRD patients for putative variants of these genes. Methods: We investigated 256 chronic hemodialysis patients, including 123 patients with a history of hypertension as a cause of ESRD. Screening for mutations in the carboxy termini of hβENaC and hγENaC was accomplished by polymerase chain reaction amplification followed by single-strand conformation polymorphism analysis. Results: In 231 patients single-strand conformation polymorphism analysis of the polymerase chain reaction fragments of the hβENaC and hγENaC genes showed a similar migration pattern as compared with negative control subjects. In 25 patients a band shift was observed. However, sequence analysis in all these patients revealed wild-type sequence. Conclusions: The present study demonstrates the absence of genetic variants in the carboxy terminus of the hβENaC and hγENaC genes in Austrian patients with ESRD maintained on chronic hemodialysis treatment. Thus, mutations in these genes are unlikely to be associated with ESRD.

Recent molecular studies have resulted in the identification of genetic alterations underlying hereditary disorders of iron metabolism. One example is the discovery of the HFE gene that is mutated in patients suffering from hereditary hemochromatosis. This autosomal recessive disorder has an estimated carrier frequency that varies between 0.07 and 0.13, thus representing one of the most common genetically determined metabolic disorders. The identification of the hemochromatosis mutations has encouraged efforts to investigate other conditions with iron overload for a putative interaction with these genetic variants. Few data are already available suggesting, for example, that iron overload in patients with sporadic porphyria cutanea tarda is associated with mutations in the hereditary hemochromatosis gene. However, it is obvious that disorders of iron metabolism have a multifactorial pathogenesis, including environmental and genetic factors. Thus, many questions remain to be answered about whether a genetic predisposition exists for development of various iron-loading or iron-deficiency phenotypes. This review focuses on the most recent advances in the field of hereditary disorders of iron metabolism and discusses their potential implications for nephrologists.