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A autoimmune Pulmonary Alveolar Proteinosis (PAP) patient with persistent disease underwent 3 Whole Lung Lavages (WLLs), 10 plasmapheresis sessions and further 3 WLL, from October 2004 to May 2007.

Background/Objectives: Increasing evidence suggests the importance of neuroinflammation in the pathogenesis of Alzheimer’s disease (AD), which is a complex neurodegenerative disorder. Complement activation occurs in the brain of patients with AD and seems to contribute to an important local inflammatory state. Increased expression of the fourth serum complement component 4 (C4) has been observed in AD patients in many studies. This protein has two isoforms, encoded by two genes: C4A and C4B localized to the HLA class III region. These genes exhibit copy number variations (CNVs) and this different gene copy number can influence C4 protein levels. We focalized our attention on these two genes, determining the distribution of CNVs in AD patients, compared with healthy controls, in order to analyse their possible involvement in AD pathogenesis. Methods: We investigated 191 AD patients and 300 healthy controls. The C4A and C4B copy numbers were assessed by quantitative PCR (qPCR). Results: The results obtained showed a statistically significant increase in the number of copies for both C4A and C4B in AD patients, compared with healthy controls (p Conclusion: The presence of high C4A and C4B copy numbers in AD patients could explain the increased C4 protein expression observed in AD patients, thus highlighting a possible role for C4A and C4B CNVs in the risk of developing AD.

Rationale: PAP is a ultra-rare disease, where surfactant accumulation within alveolar spaces could lead to respiratory failure and death. WLL is the current standard treatment for PAP, nevertheless disease persistence after treatment is reported in 30% of cases. Aim: This work is aimed to identify a common gene co-expression network able to predict the outcome of the WLL: total resolution/persistent improvement vs transient resolution/progressive deterioration. Methods: We applied a weighted genes co-expression network analysis to transcriptional profiles of total RNA, collected from peripheral blood mononuclear cells (PBMC) of 16 PAP patients who underwent WLL. After a 24-month follow up, they were dichotomized in positive vs negative outcome and a microarray analysis, with around 20000 genes screened (SurePrint G3 Human Gene, Agilent) was performed. Finally Gene Ontology (GO), Reactome, KEGG, and Disease Ontology enrichment analysis was performed for differentially expressed genes (DEGs). Results: Four biologically meaningful modules of genes highly connected were identified. In particular, one of these (comprehending 111 genes with a p value Conclusion: For the first time we provide a modular profile of PAP which could explain the regulatory mechanism underlying the clinical outcome.

Background: The laboratory diagnosis of alpha-1 antitrypsin deficiency (AATD) consists of alpha-1 antitrypsin (AAT) plasma levels quantification and AAT genotype and phenotype determination. Phenotyping helps to confirm the presence of the most common S and Z alleles and to detect rare variants. The classical method used to determine phenotype is IsoElectric Focusing (IEF), based on the separation of proteins according to their charge. In our lab, AATD diagnosis is lead on dried blood spot (DBS), drops of blood that have been dried onto a special filter paper, since 2006, in order to allow the centralization of AATD diagnosis in Italy. Objectives: We adapted a semi-automatic system for the determination of AAT phenotypes on DBS and we compared this method to the standard IEF performed with home made polyacrylamide gels 1 . Methods: The new method uses the Hydragel 18 A1AT Isofocusing® kit on the Sebia Hydrasys® System. The procedure consists of samples runs on ready-to-use agarose gels and immunofixation with an AAT specific antiserum labeled with peroxidase. Results: The Hydragel 18 A1AT Isofocusing® kit has been conceived to work with serum samples. To make the kit work with DBS samples we introduced some changes in the protocol, in particular regarding dilutions. We ran 100 DBS samples both with the old and the new method and we obtained superimposable results. Moreover, the new method made easier the identification of rare variants and clearer the Z-AAT pattern than old method. Conclusions: The new semi-automatic IEF method applied to DBS resulted to be easier, faster and more reproducible than the old manual procedure. 1.Ferrarotti I, et al. Transl Res. 150:267-74, 2007.