Chapter 13 : Molecular and Clinical Evaluation of Atypical Hemolytic Uremic Syndrome

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Familial hemolytic uremic syndrome (HUS) was first described in 1956; both autosomal dominant and recessive forms of inheritance have been reported. The chapter discusses three pieces of evidence which suggested that complement genes apart from the gene for factor H were involved in the pathogenesis of atypical HUS. Structurally, membrane cofactor protein (MCP) consists of four alternatively spliced isoforms that coexist on most cells. The extracellular domain is composed of four control protein modules (CCPs). Western blot, flow cytometry, cell surface labeling, and pulse-chase analysis all showed that the mutant protein was retained intracellularly. Noris et al. described a family in which two siblings were affected by HUS. Mutation screening in both revealed a heterozygous 2-bp deletion in MCP exon 7, which encodes CCP4. The functional effect of this mutation is similar to the deletion mutation and results in half of the normal level of cell surface expression of MCP. The two techniques most frequently used for mutation scanning are denaturing high-performance liquid chromatography (DHPLC) and single-strand conformation polymorphism (SSCP). DHPLC is based on heteroduplex analysis in which mutant and wild-type sequences present in a PCR product are heated and allowed to reanneal slowly. This results in the formation of two heteroduplexes and two homoduplexes. MLPA (multiplex ligationdependent probe amplification) is a relatively new method for detecting deletions and duplications in a simple two-stage procedure and gives dosage information for up to 45 exons in one test.

Citation: Strain L, Goodship J, Goodship* T. 2006. Molecular and Clinical Evaluation of Atypical Hemolytic Uremic Syndrome, p 118-123. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch13

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Denaturing Gradient Gel Electrophoresis
Factor H
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Factor H mutations associated with HUS are identified above the corresponding CCP. (A) Missense mutations with normal factor H levels. (B) Mutations associated with factor H deficiency (Fr, frameshift; X, premature stop codon; Δ, deletion). The amino acid numbering includes the 18-amino-acid signal peptide.

Citation: Strain L, Goodship J, Goodship* T. 2006. Molecular and Clinical Evaluation of Atypical Hemolytic Uremic Syndrome, p 118-123. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch13
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Methods for mutation detection

Citation: Strain L, Goodship J, Goodship* T. 2006. Molecular and Clinical Evaluation of Atypical Hemolytic Uremic Syndrome, p 118-123. In Detrick B, Hamilton R, Folds J (ed), Manual of Molecular and Clinical Laboratory Immunology, 7th Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815905.ch13

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