Aqueductal CSF stroke volume is associated with the burden of perivascular space enlargement in chronic adult hydrocephalus.

The inflow of CSF into perivascular spaces (PVS) in the brain is crucial for clearing waste molecules. Inefficiency in PVS flow leads to neurodegeneration. Failure of PVS flushing is associated with CSF flow impairment in the intracranial hydrodynamic condition of CSF hypo-pulsatility. However, enlarged PVS (ePVS), a finding indicative of PVS flow dysfunction, is also present in patients with derangement of CSF dynamics characterized by CSF hyper-pulsatility, which increases CSF flow. Intriguingly, two opposite intracranial hydrodynamic conditions would lead to the same result of impairing the PVS flushing. To investigate this issue, we assessed the subsistence of a dysfunctional interplay between CSF and PVS flows and, if the case, the mechanisms preventing a hyper-pulsatile brain from providing an effective PVS flushing. We analyzed the association between phase contrast MRI aqueductal CSF stroke volume (aqSV), a proxy of CSF pulsatility, and the burden of ePVS in chronic adult hydrocephalus, a disease involving a broad spectrum of intracranial hydrodynamics disturbances. In the 147 (85 males, 62 females) patients, the age at diagnosis ranged between 28 and 88 years (median 73 years). Ninety-seven patients had tri-ventriculomegaly and 50 tetra-ventriculomegaly. According to the extent of ePVS, 113 patients had a high ePVS burden, while 34 had a low ePVS burden. aqSV, which ranged between 0 and 562 µL (median 86 µL), was increased with respect to healthy subjects. Patients presenting with less ePVS burden had higher aqSV (p < 0.002, corrected for the multiple comparisons) than those with higher ePVS burden. The present study confirmed the association between CSF dynamics and PVS flow disturbances and demonstrated this association in intracranial hyper-pulsatility. Further studies should investigate the association between PVS flow failure and CSF hypo- and hyper-pulsatility as responsible/co-responsible for glymphatic failure in other neurodegenerative diseases, particularly in diseases in which CSF disturbances can be corrected, as in chronic adult hydrocephalus.

Case report: A multiple sclerosis patient with imaging features of glymphatic failure benefitted from CSF flow shunting.

The derangement of CSF circulation impacts the functions of the glymphatic-lymphatic system (G-Ls), which regulates solute trafficking and immune surveillance in the CNS. The G-Ls failure leads to the dysregulation of clearance of waste molecules in the brain and to an altered CNS immune response. The imaging features of dilated perivascular spaces imply the impairment of the G-Ls. We report on the case of a patient with primary progressive multiple sclerosis and dilatation of perivascular spaces, who transiently improved after CSF shunt diversions. The underlying mechanisms remain to be determined and at this stage, it is not possible to link CSF diversion to an effect on MS pathology. However, this observation provides the rationale to incentivize research in the largely unknown area of CSF dynamic disturbances on G-Ls failure and ultimately in neurodegeneration.

Aqueductal CSF stroke volume measurements may drive management of shunted idiopathic normal pressure hydrocephalus patients.

CSF shunting with adjustable valve is the treatment of idiopathic normal pressure hydrocephalus. The opening pressure valve setting is left to the neurosurgeon's experience. Aqueductal CSF stroke volume by phase-contrast magnetic resonance measures the CSF passing through the Sylvian aqueduct and it changes with intracranial hydrodynamics. We sought to identify a window of stroke volume differences associated with the best clinical outcome and lowest rate of complications. The records of 69 patients were reviewed. At every clinical check, stroke volume, opening pressure valve, clinical outcome, and CSF overdrainage were analyzed. The correlation between stroke volume differences and negative outcome was also analyzed. The median follow-up was 2.3 years (range 0.3-10.4 years). The odds of negative outcome between two consecutive checks significantly increased by 16% (95%CI 4-28%, p = 0.006). Taking the lowest risk group as reference, the odds ratio of negative outcome was 1.16 (95%CI 0.51-2.63, p = 0.726) for SV differences less than - 37.6 µL, while it was 1.96 (95%CI 0.97-3.98, p = 0.062) for stroke volume changes above + 23.1 µL. Maintaining stroke volume values within a definite range might help maximize clinical benefit and avoid the risk of CSF overdrainage.

Early-onset malignant phyllodes breast tumor in a patient with germline pathogenic variants in NF1 and BRCA1 genes.

We present a 24-year-old female patient affected by neurofibromatosis type 1 (NF1) who developed a malignant phyllodes tumor of the breast. The molecular studies showed that the patient carried a heterozygous inactivating deleterious variant in BRCA1 inherited from the father associated with a germline de novo pathogenic alteration in NF1; the tumor presented a biallelic inactivation of both genes. Therefore, tumor analyses helped to establish that the germline NF1 and BRCA1 variants were in cis on the paternal chromosome. This last information is important to provide adequate genetic counselling regarding the risk of recurrence in the offspring, as well as opportunity for early intervention. In conclusion, we present the first case of a malignant phyllodes tumor of the breast in patient carrying pathogenic variants in NF1 and BRCA1. Further studies will be necessary to understand if the phyllodes histotype represents a very rare component of NF1-associated breast cancer.

Accuracy and safety of 1-day external lumbar drainage of CSF for shunt selection in patients with idiopathic normal pressure hydrocephalus.

OBJECTIVEThree to five days of external lumbar drainage (ELD) of CSF is a test for ventriculoperitoneal shunt (VPS) selection in idiopathic normal pressure hydrocephalus (iNPH). The accuracy and complication rates of a shorter (1-day) ELD procedure were analyzed.METHODSData of patients with iNPH who underwent 1-day ELD to be selected to undergo VPS placement with a programmable valve in the period from 2005 to 2015 were reviewed. Patients experiencing VPS complications, valve malfunctioning, or with less than 1 year of follow-up were excluded. The ability of 1-day ELD to predict VPS outcome at 1- and 12-month follow-up was assessed by calculating sensitivity, specificity, and positive and negative predictive values.RESULTSOf 93 patients who underwent 1-day ELD, 3 did not complete the procedure. Of the remaining 90 patients, 2 experienced transient nerve root irritation. Twenty-four patients had negative test outcomes and 66 had positive test outcomes. Nine negative-outcome patients had intraprocedural headache, which showed 37.5% sensitivity (95% confidence interval [CI] 19.5%-59.2%) and 100% specificity (95% CI 93.1%-100%) as predictors of negative 1-day ELD outcome. Sixty-eight patients (6 with negative and 62 with positive outcomes) underwent VPS insertion, which was successful in 0 and 58 patients, respectively, at 1-month follow-up. Test sensitivity and specificity in predicting surgical outcome at 1-month follow-up were 100% (95% CI 92.3%-100%) and 60% (95% CI 27.4%-86.3%), respectively, with 94.1% accuracy (95% CI 85.6-98.4%). Among the 1-day ELD-positive patients, 2 showed no clinical benefit at 12 months follow-up. Test sensitivity and specificity in predicting surgical outcome at 12-month follow-up was 100% (95% CI 92.5%-100%) and 75.0% (95% CI 35.6%-95.5%), respectively, with 97.1% (95% CI 89.8%-99.6%) accuracy.CONCLUSIONSOne-day ELD is a reliable tool in iNPH management, with low complication risk and short trial duration. The test is very consistent in predicting who will have a positive outcome with VPS placement, given the high chance of successful outcome at 1- and 12-month follow-up; negative-outcome patients have a high risk of unsuccessful surgery. Intraprocedural headache is prognostic of 1-day ELD negative outcome.

Glaucoma in patients with shunt-treated normal pressure hydrocephalus.

OBJECTIVE: Changes in the pressure gradient between intraocular and intracranial compartments at the lamina cribrosa level are a possible explanation of normal tension glaucoma (NTG). Shunt-treated normal pressure hydrocephalus (NPH) is a model for testing whether the increase (time from disease onset to CSF shunt placement, i.e., "protection period") and decrease (time from shunt placement to observation, i.e., "exposure period") in intracranial pressure (ICP) are glaucoma protective or risk factors, respectively. The authors estimated the prevalence of NTG in patients with shunt-treated NPH and calculated the extent of optic nerve exposure to changes in the trans-lamina cribrosa gradient. METHODS: Data obtained in patients with NPH who had undergone ventriculoperitoneal (VP) shunt placement were analyzed. Patients with more than 6 months' follow-up, no pathologies associated with ICP changes or CSF dynamics disturbances, and no surgical or valve-related complications were scheduled for ophthalmic evaluation. RESULTS: Nine of 22 patients had NTG, which is about a 40-fold increase in rate compared with the rate in the general elderly population without hydrocephalus (p < 0.001). The median protection period was 12.0 months in patients with NTG and 18.0 months in those without NTG (p = 0.033). The median ICP decrease multiplied by duration of exposure in months was 76.0 mm Hg × months in the NTG group and 24.1 mm Hg × months in the no-NTG group (p = 0.048). The patients' median adjusted age (adjusted for "protection" and "exposure" times) was 85.1 years in the NTG group and 78.8 years in the no-NTG group (p = 0.001). CONCLUSIONS: A crucial risk factor for development of NTG in patients with shunt-treated NPH is the duration of optic nerve exposure to the lowering of ICP. Patients with NPH who are candidates for CSF shunting should be informed of the risk of incurring glaucoma. Longitudinal studies could provide estimates of tolerated times for a given ICP decrease.

Evaluation of a Next-Generation Sequencing Assay for BRCA1 and BRCA2 Mutation Detection.

The efficiency of a novel targeted next-generation sequencing (NGS) test, the Devyser BRCA kit, for a comprehensive analysis of all 48 coding exons of the high-risk breast/ovarian cancer susceptibility genes BRCA1 and BRCA2 has been assessed. The new assay intended to detect nucleotide substitutions, small deletions/insertions, and large deletions/duplications. To document the false-negative and false-positive rates of the NGS assay in the hands of end users, 48 samples with previously identified 444 small variants and seven gross rearrangements were analyzed, showing 100% concordance with gold standards. Furthermore, all other 43 variants (42 single-nucleotide variation or insertion/deletion variation and one copy number variation, whose significance is or may be of clinical value), which were called by the NGS assay in a prospectively analyzed 179-sample set, were confirmed by Sanger sequencing or multiplex ligation probe amplification, according to their nature. We conclude that the Devyser BRCA kit performed satisfactorily for use in a clinical laboratory.

A Founder Effect for the HGD G360R Mutation in Italy: Implications for a Regional Screening of Alkaptonuria.

We sought to establish rapid and specific genotyping methods for G360R mutation and for seven tightly linked markers in the homogentisate dioxygenase gene to address the question of whether G360R is a mutational hot spot or the result of a founder effect, as it has been repeatedly found in alkaptonuric patients from a geographic isolate in Italy.For G360R and single nucleotide polymorphism genotyping, high-resolution melting analysis was performed. Microsatellites were analysed by multiplex PCR and capillary electrophoresis. To investigate the natural history of the G360R mutation, we genotyped markers in 52 controls and in 8 unrelated patients from the UK and USA, who also segregated the G360R mutation, and calculated its age using DMLE+2.3 software.A distinct G360R-bearing haplotype was identified in all patients of Caucasian descent. Estimated mutation age was 545 generations (95% credible set, 402-854), suggesting that G360R arose in an ancestor who lived 8,000-10,000 years BC. Archaeological, historical and demographic data support that a G360R carrier has settled the remote valley where present-day population might have a heterozygote frequency of at least 6%.Given the late health-threatening complications of alkaptonuria and a cure within reach, inhabitants of this isolate would benefit from screening and genetic counselling.

Twelve novel HGD gene variants identified in 99 alkaptonuria patients: focus on 'black bone disease' in Italy.

Alkaptonuria (AKU) is an autosomal recessive disorder caused by mutations in homogentisate-1,2-dioxygenase (HGD) gene leading to the deficiency of HGD enzyme activity. The DevelopAKUre project is underway to test nitisinone as a specific treatment to counteract this derangement of the phenylalanine-tyrosine catabolic pathway. We analysed DNA of 40 AKU patients enrolled for SONIA1, the first study in DevelopAKUre, and of 59 other AKU patients sent to our laboratory for molecular diagnostics. We identified 12 novel DNA variants: one was identified in patients from Brazil (c.557T>A), Slovakia (c.500C>T) and France (c.440T>C), three in patients from India (c.469+6T>C, c.650-85A>G, c.158G>A), and six in patients from Italy (c.742A>G, c.614G>A, c.1057A>C, c.752G>A, c.119A>C, c.926G>T). Thus, the total number of potential AKU-causing variants found in 380 patients reported in the HGD mutation database is now 129. Using mCSM and DUET, computational approaches based on the protein 3D structure, the novel missense variants are predicted to affect the activity of the enzyme by three mechanisms: decrease of stability of individual protomers, disruption of protomer-protomer interactions or modification of residues in the region of the active site. We also present an overview of AKU in Italy, where so far about 60 AKU cases are known and DNA analysis has been reported for 34 of them. In this rather small group, 26 different HGD variants affecting function were described, indicating rather high heterogeneity. Twelve of these variants seem to be specific for Italy.

Donor-Specific Anti-HLA Antibodies in Huntington's Disease Recipients of Human Fetal Striatal Grafts.

Fetal grafting in a human diseased brain was thought to be less immunogenic than other solid organ transplants, hence the minor impact on the efficacy of the transplant. How much prophylactic immune protection is required for neural allotransplantation is also debated. High-sensitive anti-HLA antibody screening in this field has never been reported. Sixteen patients with Huntington's disease underwent human fetal striatal transplantation in the frame of an open-label observational trial, which is being carried out at Florence University. All patients had both brain hemispheres grafted in two separate robotic-stereotactic procedures. The trial started in February 2006 with the first graft to the first patient (R1). R16 was given his second graft on March 2011. All patients received triple immunosuppressive treatment. Pre- and posttransplant sera were analyzed for the presence of anti-HLA antibodies using the multiplexed microsphere-based suspension array Luminex xMAP technology. Median follow-up was 38.5 months (range 13-85). Six patients developed anti-HLA antibodies, which turned out to be donor specific. Alloimmunization occurred in a time window of 0-49 months after the first neurosurgical procedure. The immunogenic determinants were non-self-epitopes from mismatched HLA antigens. These determinants were both public epitopes shared by two or more HLA molecules and private epitopes unique to individual HLA molecules. One patient had non-donor-specific anti-HLA antibodies in her pretransplant serum sample, possibly due to previous sensitization events. Although the clinical significance of donor-specific antibodies is far from being established, particularly in the setting of neuronal transplantation, these findings underline the need of careful pre- and posttransplant immunogenetic evaluation of patients with intracerebral grafts.

Human striatum remodelling after neurotransplantation in Huntington's disease.

BACKGROUND: Restoration of functions in Huntington's disease (HD) by neurotransplantation stems from the formation of a striatum-like structure capable of establishing host connections as a result of grafted striatal neuroblast maturation. For the first time, we demonstrated some developmental steps accomplished by progenitor cells in the brain of an HD patient and analysed the molecular asset of the human primordium. CASE REPORT: Surgery involved bilateral (two sessions) stereotactic, caudate-putaminal transplantation of whole ganglionic eminence fragments from single legally aborted fetuses. MRI showed that the tissue deposits of the left hemisphere grew and joined to constitute a single tissue mass that remodelled basal ganglia anatomy and remained stable in size over time. No evidence of graft growth was observed contralaterally. PET demonstrated increased striatal and stable cortical metabolism. Unified Huntington's Disease Rating Scale assessments demonstrated improvement of motor performances, which faded over the 36-month follow-up. Cognitive performance tended to decrease at a lower rate than before transplantation. CONCLUSION: The striatal primordium grew into the host brain and this process was associated with metabolic change and some clinical benefit. The study suggests the plasticity and reparative potential of un-manipulated primordium in an era where promising cell-based therapies are still in their infancy.