Sandra D’aLfonso

Recent research has indicated a connection between amyotrophic lateral sclerosis (ALS) and Huntington’s disease (HD), an inherited neurological condition caused by a trinucleotide CAG repeat expansion within exon 1 of the Huntingtin gene (HTT; MIM:613004).1 The same pathogenic CAG repeat expansions (40 or more CAG repeats) observed in patients with HD have been identified in patients with frontotemporal dementia (FTD)/ALS.1 Similarly, non-pathogenic intermediate-length CAG repeats in the ATXN2 gene are a well-established factor associated with increased ALS risk and faster disease progression.2 Given these observations, we investigated the impact of intermediate HTT alleles on survival in two cohorts of patients diagnosed with ALS.

Background: Recent studies identified > 100 non-HLA (human leukocyte antigen) multiple sclerosis (MS) susceptibility variants in Northern European populations, but their role in Southern Europeans is largely unexplored. Objective: We aimed to investigate the cumulative impact of those variants in two Mediterranean populations: Continental Italians and Sardinians. Methods: We calculated four weighted Genetic Risk Scores (wGRS), using up to 102 non-HLA MS risk variants and 5 HLA MS susceptibility markers in 1691 patients and 2194 controls from continental Italy; and 2861 patients and 3034 controls from Sardinia. We then assessed the differences between populations using Nagelkerke’s R² and the area under the Receiver Operating Characteristic (ROC) curves. Results: As expected, the genetic burden (mean wGRS value) was significantly higher in MS patients than in controls, in both populations. Of note, the burden was significantly higher in Sardinians. Conversely, the proportion of variability explained and the predictive power were significantly higher in continental Italians. Notably, within the Sardinian patients, we also observed a significantly higher burden of non-HLA variants in individuals who do not carry HLA risk alleles. Conclusions: The observed differences in MS genetic burden between the two Mediterranean populations highlight the need for more genetic studies in South Europeans, to further expand the knowledge of MS genetics.

In this chapter we will present the principal technical methods to genotype the HLA-DQA1* and -DQB1* alleles associated with celiac disease (CD), corresponding to the serological heterodimers HLA-DQ2 and -DQ8. We will present the methods specific for the genotyping of these heterodimers, which represents a common request from consultant doctors. Because these alleles are also common in healthy subjects, their presence is not diagnostic for CD. Conversely, their absence is more important because it excludes the disease, since CD patients negative for these heterodimers are very rare. Accordingly, HLA typing has been included as a useful test to exclude celiac disease in the ESPGHAN guidelines for diagnosis of celiac disease. The methods for HLA typing described in the present chapter are based on the following techniques: PCR-SSP (Polymerase Chain Reaction-Sequence Specific Primers): PCR with primers specific for HLA alleles encoding the CD risk heterodimers, whose presence is revealed through the electrophoresis of PCR products. Reverse PCR-SSOP (PCR-Sequence Specific Oligonucleotide Probes): PCR with primers specific for a single locus or a large group of alleles followed by hybridization with enzyme-conjugated probes specific for a single allele, immobilized on different supports (i.e., nitrocellulose strips), in which DNA-probes binding is revealed by the production of a colored precipitate derived from the enzymatic modification of a specific substrate. Real-Time PCR (RT-PCR): PCR with locus or allelic specific primers whose amplification is revealed by particular probes (i.e., Taqman probes) hybridizing the DNA template within the two PCR primers and emitting fluorescent while the PCR reaction occurs.

Frontotemporal dementia (FTD) may present with psychiatric symptoms, usually together with neurological ones and in cases with a family history of dementia. We describe the case of an FTD behavioural variant with a psychiatric presentation and a normal neurological examination, due to a C9Orf72 gene mutation. The patient was a 57 years-old Caucasian woman with a recent onset of bizarre behaviours and mystic delusions. She had a negative clinical history for previous psychiatric disorders and treatments and this was her first admission to a Psychiatry Ward. A careful assessment was performed including, beyond psychiatric evaluation, the following: blood sampling, neurological examination (including electroencephalogram, electroencephalogram with zygomatic electrodes, Positron Emission Tomography, Cerebrospinal Fluid Analysis), carotid artery Doppler ultrasound, brain Magnetic Resonance Imaging – angio Magnetic Resonance Imaging. Blood sampling for the genetic assessment of mutations associated to primary dementias was performed as well: the genes investigated were FUS, C9Orf72, PSEN-1, PSEN-2. Serological tests were negative, neurological examination was normal, instrumental examinations showed theta waves in the posterior temporal areas bilaterally and frontotemporal cortical atrophy bilaterally. The genetic assessment of mutations associated revealed she carried a GGGGCC hexanucleotide repeat expansion (at least 80 repeats) in C9Orf72 intron 1. Patients carrying the C9Orf72 mutation are likely to receive a psychiatric diagnosis (mainly mood disorder or schizophrenia) prior to correct diagnosis; this may be particularly problematic for those patients with no neurological signs to orientate diagnosis. Understanding the manner in which such FTD variant may present as a psychiatric syndrome, with a negative neurological examination, is essential to provide the best treatment for patients, as soon as possible, especially when the behavioural anomalies interfere with their care.