Case report
Frontotemporal lobar degeneration with MAPT mutation in an Italian-Polish family. A case report

Introduction

Frontotemporal lobar degeneration (FTLD) represents a large heterogeneous group of familial and sporadic diseases, in which the neurodegenerative process involves mainly the frontal and temporal lobes (Fig. 1) [3]. The group of these diseases constitutes about 20% of presenile dementia cases [18]. Hereditary forms of FTLD are associated with mutations of multiple genes, including MAPT (microtubule-associated protein tau gene), PGRN (progranulin gene), VCP (valosin-containing protein gene), CHMP2B (charged multivesicular body protein 2B gene), TDP-43 (TAR DNA-binding protein 43) and other mutations [4,17]. MAPT mutations are identified in up to 20% of familial FTLD [9,20]. The MAPT gene codes the microtubule-associated protein tau, which is involved in microtubule assembly and stabilization, neuronal polarity, and axonal transport in the brain [7,20]. The tau protein is mainly synthesized in the peripheral and central nervous system (central nervous system [CNS], neurons and neuroglia), but it is also expressed in other tissues (heart, skeletal muscle, lung, kidney and others) [1,8,16]. The MAPT gene locus on chromosome 17 q21 has two major haplotypes, H1 and H2. MAPT consists of 16 exons and encodes 6 human brain isoforms of tau protein with 3 or 4 repeat domains (3R and 4R) [2,5,15]. Tau protein inclusions in different tauopathies have specific morphology and distribution. In the nervous system of FTLD patients carrying the MAPT mutation, inclusions of both 3R tau and 4R isoforms of tau protein are found. The 3R isoform of tau protein inclusions are observed in FTLD with Pick bodies, while the 4R isoform of tau protein occurs, inter alia, in such diseases as corticobasal degeneration, progressive supranuclear palsy and argyrophilic grain disease [6]. More than 40 mutations have been identified in the MAPT gene in individuals from more than 100 families with FTLD [6,12,27]. Depending on the mutation in the MAPT gene, we can observe different neuropathologic and clinical symptoms. There are also differences between family members and families suffering from FTLD with the same MAPT mutations. The main hallmarks of this disease are neurofibrillary pathology, neuronal inclusions, tau-positive neuroglial inclusions, both in astrocytic and oligodendrocytic cells and also the coiled bodies [5,11,13,26]. In this study, we investigated immunohistochemically, using antibodies against 3R- and 4R-tau isoforms, genetic and ultrastructural features of two members of Italian-Polish familial FTLD with dementia and movement disorders.

Material and methods

The reported cases

The pedigree consists of 25 Italian-Polish family members in four generations. The first tested proband, a 58-year-old man (Fig. 2, II.2), died one year after the onset of the first dementia symptoms. At the age of 49, he had a cardiac pacemaker implanted. The patient complained of muscle weakness. At the end of his life, bulbar syndrome occurred. The diagnosis of Alzheimer’s disease was suspected. The proband’s sister (Fig. 2, II.1) died at the age of 68. She presented with behavioral abnormalities from the age of 59. Primarily, the loss of awareness, memory, interest and empathy was noted. Gradually social withdrawal and logopenic aphasia progressed. At the age of 61 she was afflicted by brain hemorrhagic stroke. At the end of her life, pyramidal and Parkinsonian syndromes were reported. Magnetic resonance imaging (MRI) showed cerebral atrophy in frontal and temporal lobes (Fig. 3).

Methods

Brain examinations were performed. Samples were taken from the brain structures of the affected male proband (II.2) and his sister (II.1). They were fixed in 10% buffered formalin and paraffin embedded. The specimens were stained with hematoxylin-eosin, Bielshowsky, Yamamoto, and Gallyas methods. Immunohistochemical studies were performed with antibodies to glial fibrillary acidic protein (GFAP, DAKO 1 : 70), anti-tau (DAKO 1 : 100), anti-Tau, 3R-repeated isoform RD3 (Millipore, 1 : 1000), anti-tau-4R repeated isoform RD4 (Millipore, 1 : 1000), ubiquitin (DAKO, 1 : 35), -amyloid (DAKO, 1 : 70) and -synuclein (Leica, 1 : 20). For electron microscope evaluation small fragments of brains were taken from formalin or paraffin blocks. After deparaffinizing and/or washing in water, the material was fixed in 2.5% glutaraldehyde and postfixed in 2% OsO4, then routinely processed to Spurr resin. Ultrathin sections were stained with uranyl acetate and lead citrate and examined in an electron microscope (Opton DPS 109). Blood samples for DNA analysis were protected. Genomic DNA was isolated from peripheral blood leukocytes using standard methods. The male proband was screened for mutations in: PSEN1, APP, PGRN, MAPT, and C9ORF72. The presence of the mutation also was confirmed in the proband’s sister. The absence of the mutation IVS10+3G> A MAPT was confirmed in the control group of 150 elderly, neurologically healthy subjects from the Polish population. Haplotype associated with the mutation was identified by the presence or absence of 238 bp deletion in intron 9 and by genotyping five SNPs (rs1467967, rs242557, rs3785883, rs2471738 and rs7521) [22]. The mutation-associated haplotype was determined based on the comparison of diplotypes identified in the male proband and his sister.

Results

In both cases, the gross examination of the brains showed moderate symmetrical cortical atrophy in the frontal and temporal lobes. There was mild atrophy of the anterior part of the caudate nucleus, while other subcortical structures were normal. The lateral ventricles were intensively dilated. The substantia nigra of the mesencephalon and locus coeruleus of the pons were moderately depigmented. The cerebellum seemed to be normal. Routine histology of the light microscopy examination showed neocortical neuronal loss, mostly in the frontal and temporal lobes in layers II and III and also in the substantia nigra. This change was accompanied by moderate spongiosis and astrogliosis. Sections stained with either the Bielschowski, Yammamoto or Gallyas silver technique showed neurofibrillary tangles (NFTs) and neuropil threads (NTs) (Fig. 4A,B). Silver-stained fibrillary inclusions were found in Purkinje cells and neurons of the substantia nigra and also in the locus coeruleus (Fig. 4C-E). Numerous coiled bodies were observed in the white matter in frontal and temporal lobes (Fig. 4F,G). Ballooned neurons were detected in the pons and in the cerebral and cerebellar cortices (Fig. 4H). Tau-immunoreactive methods detected many more NFTs, coiled bodies and NTs than silver methods. Tau-immunoreactive deposits were abundant in the frontal and temporal cortex but also in the brainstem. They were visible in the form of granular deposits, tau-positive abundant neurites, neuropil threads, coiled bodies, Pick bodies, ballooned neurons and grain-like deposits (Fig. 5 and 6). Tau-3 repeated isoform positive Pick bodies in the granule neurons of the dentate gyrus of the hippocampus occurred only in the male proband (Fig. 5C), whereas intracytoplasmic deposits of tau-4 repeated isoform were observed in both cases (Fig. 5D and 6E). In the frontal cortex of the sister, tau-positive Pick bodies-like deposits were visible (Fig. 6F). A GFAP-positive tufted and monstrous astrocytes were observed mainly in the sister (Fig. 2, II.1) (Fig. 6H and 6J). Pathology of vascular walls was visible in both cases (Fig. 5J and 6J). Ultrastructural analysis of fibrillary inclusions showed tightly aggregated filaments of neurofibrillary tangles in the cytoplasm of nerve cells (Fig. 7). Astrocytes were often swollen with only a few short gliofilaments or with numerous gliofilaments filling the cytoplasm (Fig. 8). In their cytoplasm, lipofuscin deposits were located in the adjacent gliofilaments (Fig. 8). In capillaries, abundant collagen fibers were frequently found in the pericyte position and small clusters in different locations of the basement membrane (Fig. 9).
The analysis of the nucleotide sequence of the MAPT gene revealed an intron 10+3-splice site mutation (IVS10+3G>A, g.123806G>A) in both affected siblings. The analysis of diplotypes associated with mutation in the male proband and his sister showed that the IVS10+3G>A mutation was in the haplotype H1s background. The MAPT diplotype of the male proband was H1s/H1b, and that of his sister was H1s/H1j (Table I).

Discussion

We present a comprehensive analysis of members of the first Italian-Polish family with genetically confirmed FTLD with the IVS10+3G>A MAPT mutation. The mutation was previously reported as pathologic in another European population [19,26,28] and was absent in 138 patients with FTLD in the Polish population (data not published). The analysis of the MAPT gene in the two described cases showed that the IVS10+3G>A mutation was in the haplotype H1s background. There were differences in the MAPT diplotypes. The male proband’s diplotype was H1s/H1b, while his sister’s was H1s/H1j. It is possible that these variations affected the differences in the clinical course of the disease and in its neuropathological picture [28]. The male proband died one year after the onset of symptoms of dementia and 9 years after implantation of a cardiac pacemaker. The sister of the proband experienced brain hemorrhagic stroke 2 years after disease onset and died 9 years after the first symptoms of dementia. The mutation in the microtubule protein tau (MAPT) gene locus into haplotype H1 affected tau microtubule assembly and tau mRNA splicing. In addition, the increased inclusion of exon 10 in transcripts show an imbalance in the 3R and 4R tau isoforms [2,9,15].
The ratio of these two isoforms in normal conditions is generally equal (1 : 1), but it varies depending on the type of mutation [24]. Corticobasal degeneration (CBD) is characterized by increased 4R tau, whereas frontotemporal lobar degeneration with Pick bodies shares an overabundance of 3R tau (Fig. 1). FTLD with MAPT mutation is a biochemically heterogeneous disease associated with 3R and 4R tau deposits in various proportions, influencing the type of neuropathological changes. The main cellular function of tau protein is to assemble and stabilize microtubules and neural integrity. The aggregation of microtubules and creation of NFTs may be regarded as “toxic intensification” of their function in neurons [21,23]. These disorders may start aging processes leading finally to apoptosis. The clinical course of FTLD with MAPT mutation is also heterogeneous, because of the tau protein function in the cells outside the nervous system [1,8,16]. Ultrastructurally, in both cases, we observed neurofibrillary pathology and deposits of collagen in the vessel walls of the male proband’s brain [10]. Šerý et al. [24] suggest that vascular pathology influences pathological processes in Alzheimer’s disease [14]. Our report suggests that FTLD with MAPT mutation affects mainly the CNS, and neuropathological changes depend on the variant of the mutation. In the clinical course of this disease, damage of the cardiovascular system may also be observed.

Disclosure

Authors report no conflict of interest.

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