Molecular characterization in an Italian patient with plasminogen deficiency and ligneous conjunctivitis

Plasminogen plays an important role in fibrinolysis and wound healing. Plasminogen deficiency is a rare disease associated with ligneous conjunctivitis. Since today 23 different mutations on PLG gene have been reported. We report a case of 3 year old Italian boy presenting ligneous conjunctivitis and low plasma levels of plasminogen (26%). Clinical history is unremarkable, and both parents are clinically healthy. Previously researches reported an extraordinary sequence homology between PLG genes, apo(A) genes and PRGs genes responsible of co-amplification problems of DNA fragments during PCR with a possible solutions based on amplification of PLG exons followed by subcloning and sequencing. We tried to carry out a faster strategy using Long PCR to analyse the gene portion between 5’UTR and intron 5. The PCR product subsequently were extracted from the gel and used to perform a series of nested PCR on amplification of every single exons for further sequencing analysis. Exons 11, 12, 16 and 17 were amplified using the forward primer of the upstream exon and the reverse primer of the downstream exon; the product was gel extracted and sequenced too. All remaining exons without any co-amplification problem, were amplified and sequenced. This strategy revealed a missense homozygous mutation in exon 2, an A to G substitution leading to an amino acid exchange (Lys to Glu) at codon 19 (K19E). The same mutation in heterozygous state was present in both parents. This mutation has been previously reported in German, Italian and Scottish patients. Since the K19E mutation was also reported in homozygous state in a healthy Scottish blood donor with low levels of PLG and no clinical manifestations, its clinical significance and allele distribution in Europe populations could be the matter of further investigations.