A transgenic mouse model of hereditary motor and sensory neuropathy

Congenital Hypomyelination (CH) IS the most severe demyelinating form of Hereditary Motor and Sensory Neuropathies and manifests at birth in human. Some subtypes of CH are due to dominant mutations in the gene coding for PO glycoprotein, which functions as a homophilic adhesion protein, responsible for compaction of opposing myelin lamellae. By homologous recombination in ES cells, we have generated a mouse containing a nonsense mutation in the intracellular portion of PO (Q215X) that, in the heterozygous state, is associated with CH neuropathy in humans. This mutation is predicted to encode a truncated PO protein, lacking part of the cytoplasmic domain. Expression analysis demonstrated that Q215X heterozygous mice produce both wildtype PO and a smaller, truncated PO glycoprotein; furthermore, the levels of mRNA and protein produced by the mutated allele are less abundant, relative to the wildtype. We demonstrated then that this reduction in mRNA and protein levels could be paIiially explained by inefficient transcription of the mutated allele, due to the presence of a LoxP site with intron 5. Behavioral analysis of the Q215X1+ mice revealed reduced motor performance at 11 days after birth. Morphological analysis performed on sciatic nerves of mice between PI and P 14 revealed defects in the process of axonal sorting by Schwann cells, with the presence of bundles of mixed large and small calibre axons surrounded, but not ensheathed, by single Schwann cells. These morphogical defects are rescued after the second week of life: sciatic nerves from adult mice, in fact, show only mild hypomyelination, which is much less severe than the morphology reported in patients. From these preliminary data, we conclude that the Q215X mutation results in a truncated PO protein; since the phenotype of Q215X1+ mice and PO +1- mice differs, Q215X probably produces a gain of function. Finally, we studied in vivo the intracellular location of the truncated protein, in order to clarify aspects of the pathogenetic mechanism of the Q215X PO mutation: we found that the mutated protein is not properly trafficked within Schwann cells, being partially retained in the ER compartment. The phenotype we have observed in the mutant mice presents similarities to other hypomyelinating mice that carry mutations in different genes, that are all involved in the laminin pathway: this suggests that mutant PO may interfere with laminin signaling, required for the correct timing of axonal sorting by Schwann cells. Thus, from these data we conclude that the Q215X mouse is a partial model of Congenital Hypomyelination, less severely affected, if compared to human patients. This difference in disease severity could be partially explained by the inefficient transcription of the Q215X allele. Thus, we believe that the mechanism of this hypomyelination is likely to be related to the mechanism of the more severe neuropathy in human. This mouse will be useful to reveal the pathogenesis of the mutation

The MULTI-ACT model: the path forward for participatory and anticipatory governance in health research and care

The COVID-19 pandemic has unmasked even more clearly the need for research and care to form a unique and interdependent ecosystem, a concept which has emerged in recent years. In fact, to address urgent and unexpected missions such as "fighting all together the COVID-19 pandemic", the importance of multi-stakeholder collaboration, mission-oriented governance and flexibility has been demonstrated with great efficacy. This calls for a policy integration strategy and implementation of responsible research and innovation principles in health, promoting an effective cooperation between science and society towards a shared mission. This article describes the MULTI-ACT framework and discusses how its innovative approach, encompassing governance criteria, patient engagement and multidisciplinary impact assessment, represents a holistic management model for structuring responsible research and innovation participatory governance in brain conditions research

Prospective validation of the CLIP score: a new prognostic system for patient with cirrhosis and hepatocellular carcinoma

Prognosis of patients with cirrhosis and hepatocellular carcinoma (HCC) depends on both residual liver function and tumor extension. The CLIP score includes Child-Pugh stage, tumor morphology and extension, serum alfa-fetoprotein (AFP) levels, and portal vein thrombosis. We externally validated the CLIP score and compared its discriminatory ability and predictive power with that of the Okuda staging system in 196 patients with cirrhosis and HCC prospectively enrolled in a randomized trial. No significant associations were found between the CLIP score and the age, sex, and pattern of viral infection. There was a strong correlation between the CLIP score and the Okuda stage, As of June 1999, 150 patients (76.5%) had died. Median survival time was 11 months, overall, and it was 36, 22, 9, 7, and 3 months for CLIP categories 0, 1, 2, 3, and 4 to 6, respectively. In multivariate analysis, the CLIP score had additional explanatory power above that of the Okuda stage. This was true for both patients treated with locoregional therapy or not. A quantitative estimation of 2-year survival predictive power showed that the CLIP score explained 37% of survival variability, compared with 21% explained by Okuda stage. In conclusion, the CLIP score, compared with the Okuda staging system, gives more accurate prognostic information, is statistically more efficient, and has a greater survival predictive power. It could be useful in treatment planning by improving baseline prognostic evaluation of patients with RCC, and could be used in prospective therapeutic trials as a stratification variable, reducing the variability of results owing to patient selection

Funding Medical Research Projects: Taking into Account Referees' Severity and Consistency through Many-Faceted Rasch Modeling of Projects' Scores

The funding policy of research projects often relies on scores assigned by a panel of experts (referees). The non-linear nature of raw scores and the severity and inconsistency of individual raters may generate unfair numeric project rankings. Rasch measurement (many-facets version, MFRM) provides a valid alternative to scoring. MFRM was applied to the scores achieved by 75 research projects on multiple sclerosis sent in response to a previous annual call by FISM-Italian Foundation for Multiple Sclerosis. This allowed to simulate, a posteriori, the impact of MFRM on the funding scenario. The applications were each scored by 2 to 4 independent referees (total = 131) on a 10-item, 0-3 rating scale called FISM-ProQual-P. The rotation plan assured "connection" of all pairs of projects through at least 1 shared referee.The questionnaire fulfilled satisfactorily the stringent criteria of Rasch measurement for psychometric quality (unidimensionality, reliability and data-model fit). Arbitrarily, 2 acceptability thresholds were set at a raw score of 21/30 and at the equivalent Rasch measure of 61.5/100, respectively. When the cut-off was switched from score to measure 8 out of 18 acceptable projects had to be rejected, while 15 rejected projects became eligible for funding. Some referees, of various severity, were grossly inconsistent (z-std fit indexes less than -1.9 or greater than 1.9). The FISM-ProQual-P questionnaire seems a valid and reliable scale. MFRM may help the decision-making process for allocating funds to MS research projects but also in other fields. In repeated assessment exercises it can help the selection of reliable referees. Their severity can be steadily calibrated, thus obviating the need to connect them with other referees assessing the same projects

Disruption of \u3ci\u3eMtmr2\u3c/i\u3e CMT4B1-like neuropathy with myelin outfolding and impaired spermatogenesis

Mutations in MTMR2, the myotubularin-related 2 gene, cause autosomal recessive Charcot-Marie-Tooth (CMT) type 4B1, a demyelinating neuropathy with myelin outfolding and azoospermia. MTMR2 encodes a ubiquitously expressed phosphatase whose preferred substrate is phosphatidylinositol (3,5)-biphosphate, a regulator of membrane homeostasis and vesicle transport. We generated Mtmr2-null mice, which develop progressive neuropathy characterized by myelin outfolding and recurrent loops, predominantly at paranodal myelin, and depletion of spermatids and spermatocytes from the seminiferous epithelium, which leads to azoospermia. Disruption of Mtmr2 in Schwann cells reproduces the myelin abnormalities. We also identified a novel physical interaction in Schwann cells, between Mtmr2 and discs large 1 (Dlg1)/synapse-associated protein 97, a scaffolding molecule that is enriched at the node/paranode region. Dlg1 homologues have been located in several types of cellular junctions and play roles in cell polarity and membrane addition. We propose that Schwann cell-autonomous loss of Mtmr2-Dlg1 interaction dysregulates membrane homeostasis in the paranodal region, thereby producing outfolding and recurrent loops of myelin

Disruption of Mtmr2 produces CMT4B1-like neuropathy with myelin outfolding and impaired spermatogenesis

Mutations in MTMR2, the myotubularin-related 2 gene, cause autosomal recessive Charcot-Marie-Tooth (CMT) type 4B1, a demyelinating neuropathy with myelin outfolding and azoospermia. MTMR2 encodes a ubiquitously expressed phosphatase whose preferred substrate is phosphatidylinositol (3,5)-biphosphate, a regulator of membrane homeostasis and vesicle transport. We generated Mtmr2-null mice, which develop progressive neuropathy characterized by myelin outfolding and recurrent loops, predominantly at paranodal myelin, and depletion of spermatids and spermatocytes from the seminiferous epithelium, which leads to azoospermia. Disruption of Mtmr2 in Schwann cells reproduces the myelin abnormalities. We also identified a novel physical interaction in Schwann cells, between Mtmr2 and discs large 1 (Dlg1)/synapse-associated protein 97, a scaffolding molecule that is enriched at the node/paranode region. Dlg1 homologues have been located in several types of cellular junctions and play roles in cell polarity and membrane addition. We propose that Schwann cell–autonomous loss of Mtmr2–Dlg1 interaction dysregulates membrane homeostasis in the paranodal region, thereby producing outfolding and recurrent loops of myelin