Martina Tagliaferri's thesis analyzes iron homeostasis and oxidative stress in fibroblasts from a patient with neuroferritinopathy caused by a mutation in the L-ferritin gene. The mutation alters the electrostatic potential of the ferritin C-terminus. The patient's fibroblasts show increased iron-dependent oxidative stress, reduced viability when treated with iron, compromised mitochondrial function, and ferritin iron aggregates, similarly to neuroferritinopathy in nervous tissue. Preliminary data suggest an iron-dependent cell death pathway involving oxidative stress may be responsible for death of the patient's fibroblasts.

1. UNIVERSITA' VITA-SALUTE SAN RAFFAELE
Facoltà di Medicina e Chirurgia
Corso di Laurea Magistrale in Biotecnologie mediche, molecolari e cellulari
Tesi di Laurea di: Martina Tagliaferri
Relatore: Sonia Levi
Correlatore: Paolo Santambrogio
IRON HOMEOSTASIS AND OXIDATIVE STATUS ALTERATION IN
NEUROFERRITINOPATHY AFFECTED PATIENT’S FIBROBLASTS, AN EXAMPLE OF
NEURODEGENERATION
Neuroferritinopathy (NP) is a dominantly inherited late onset monogenic disorder caused by
mutations in the L-ferritin gene. It belongs to a group of monogenic extrapyramidal disorders called
Neurodegeneration with Brain Iron Accumulation (NBIA) diseases characterized by focal
accumulation of iron in the basal ganglia, detected by MRI. Main symptoms include movement
disorders, spasticity and cognitive impairment and the age of onset varies among diseases. Up to
date, 7 mutations in the L-ferritin gene sequence have been proven to cause NP, all of them are
located in the fourth and last exon of the gene and affect the C-term of the protein in term of both
length and sequence. In humans L-chains coassemble with H-chains to form cytosolic ferritin, an
heteropolimeric spheroidal protein composed by 24 subunits. These subunits shape an internal
cavity in which excess iron is stored as ferrihydrite. In wt chains Cterm region presents an α-helix
structure called E- helix which is fundamental to form the hydrophobic channel of ferritin, located
at 4-fold symmetry axis of the molecule and involved in proton export during iron mineralization.
To assess the relation between ferritin sequence variation and function alteration we took into
consideration the c.469_484dup16 mutation characterizing the only NP Italian patient’s fibroblasts.
Computational analysis revealed that the C-term structure of the mutated L-chain doesn’t seem
compromised. However, according to this analysis, the electrostatic potential changes dramatically
in this area compared to wild type ferritin. Iron metabolism and oxidative status in patient’s
fibroblast showed increased iron-dependent oxidative stress, reduced viability, worsen by iron
treatment, and compromised
mitochondrial function, compared to the controls. Moreover patient’s fibroblasts showed ferritin
iron aggregates, a phenotype typical of the nervous tissue of Neuroferritinophaty’s patients. This
iron-dependent toxic effect seemed to induce cell death, although not through apoptosis or necrosis
pathway. An very recently described iron-dependent pathway of cell death that involves oxidative
stress is now under investigation in our laboratory and preliminary data suggested that it could be
responsible for patient’s fibroblasts death. These data show as patient’s fibroblasts display alteration
in iron metabolism, protein aggregation and increased oxidative stress as seen in
neuroferritinopathy patients’ nervous system, despite fibroblast, in patients don’t seem to be
affected. For the first time a mitochondrial involvement has been described, further characterization
are needed to fully elucidate the pathologic mechanism.