Association of CHRDL1 Mutations and Variants with X-linked Megalocornea,Neuh?user Syndrome and Central Corneal Thickness

We describe novel CHRDL1 mutations in ten families with X-linked megalocornea (MGC1). Our mutation-positive cohort enabled us to establish ultrasonography as a reliable clinical diagnostic tool to distinguish between MGC1 and primary congenital glaucoma (PCG). Megalocornea is also a feature of Neuhäuser or megalocornea-mental retardation (MMR) syndrome, a rare condition of unknown etiology. In a male patient diagnosed with MMR, we performed targeted and whole exome sequencing (WES) and identified a novel missense mutation in CHRDL1 that accounts for his MGC1 phenotype but not his non-ocular features. This finding suggests that MMR syndrome, in some cases, may be di- or multigenic. MGC1 patients have reduced central corneal thickness (CCT); however no X-linked loci have been associated with CCT, possibly because the majority of genome-wide association studies (GWAS) overlook the X-chromosome. We therefore explored whether variants on the X-chromosome are associated with CCT. We found rs149956316, in intron 6 of CHRDL1, to be the most significantly associated single nucleotide polymorphism (SNP) (p = 6.81×10⁻⁶) on the X-chromosome. However, this association was not replicated in a smaller subset of whole genome sequenced samples. This study highlights the importance of including X-chromosome SNP data in GWAS to identify potential loci associated with quantitative traits or disease risk.     

Reproducibility of In Vivo Corneal Confocal Microscopy Using an Automated Analysis Program for Detection of Diabetic Sensorimotor Polyneuropathy

Objective

In vivo Corneal Confocal Microscopy (IVCCM) is a validated, non-invasive test for diabetic sensorimotor polyneuropathy (DSP) detection, but its utility is limited by the image analysis time and expertise required. We aimed to determine the inter- and intra-observer reproducibility of a novel automated analysis program compared to manual analysis.

Methods

In a cross-sectional diagnostic study, 20 non-diabetes controls (mean age 41.4±17.3y, HbA1c 5.5±0.4%) and 26 participants with type 1 diabetes (42.8±16.9y, 8.0±1.9%) underwent two separate IVCCM examinations by one observer and a third by an independent observer. Along with nerve density and branch density, corneal nerve fibre length (CNFL) was obtained by manual analysis (CNFL_MANUAL), a protocol in which images were manually selected for automated analysis (CNFL_{SEMI-AUTOMATED}), and one in which selection and analysis were performed electronically (CNFL_{FULLY-AUTOMATED}). Reproducibility of each protocol was determined using intraclass correlation coefficients (ICC) and, as a secondary objective, the method of Bland and Altman was used to explore agreement between protocols.

Results

Mean CNFL_Manual was 16.7±4.0, 13.9±4.2 mm/mm² for non-diabetes controls and diabetes participants, while CNFL_{Semi-Automated} was 10.2±3.3, 8.6±3.0 mm/mm² and CNFL_{Fully-Automated} was 12.5±2.8, 10.9 ± 2.9 mm/mm². Inter-observer ICC and 95% confidence intervals (95%CI) were 0.73(0.56, 0.84), 0.75(0.59, 0.85), and 0.78(0.63, 0.87), respectively (p = NS for all comparisons). Intra-observer ICC and 95%CI were 0.72(0.55, 0.83), 0.74(0.57, 0.85), and 0.84(0.73, 0.91), respectively (p<0.05 for CNFL_{Fully-Automated} compared to others). The other IVCCM parameters had substantially lower ICC compared to those for CNFL. CNFL_{Semi-Automated} and CNFL_{Fully-Automated} underestimated CNFL_Manual by mean and 95%CI of 35.1(-4.5, 67.5)% and 21.0(-21.6, 46.1)%, respectively.

Conclusions

Despite an apparent measurement (underestimation) bias in comparison to the manual strategy of image analysis, fully-automated analysis preserves CNFL reproducibility. Future work must determine the diagnostic thresholds specific to the fully-automated measure of CNFL.     

Non-invasive In-cell Determination of Free Cytosolic [NAD+]/[NADH] Ratios Using Hyperpolarized Glucose Show Large Variations in Metabolic Phenotypes

Accumulating evidence suggest that the pyridine nucleotide NAD has far wider biological functions than its classical role in energy metabolism. NAD is used by hundreds of enzymes that catalyze substrate oxidation and, as such, it plays a key role in various biological processes such as aging, cell death, and oxidative stress. It has been suggested that changes in the ratio of free cytosolic [NAD⁺]/[NADH] reflects metabolic alterations leading to, or correlating with, pathological states. We have designed an isotopically labeled metabolic bioprobe of free cytosolic [NAD⁺]/[NADH] by combining a magnetic enhancement technique (hyperpolarization) with cellular glycolytic activity. The bioprobe reports free cytosolic [NAD⁺]/[NADH] ratios based on dynamically measured in-cell [pyruvate]/[lactate] ratios. We demonstrate its utility in breast and prostate cancer cells. The free cytosolic [NAD⁺]/[NADH] ratio determined in prostate cancer cells was 4 times higher than in breast cancer cells. This higher ratio reflects a distinct metabolic phenotype of prostate cancer cells consistent with previously reported alterations in the energy metabolism of these cells. As a reporter on free cytosolic [NAD⁺]/[NADH] ratio, the bioprobe will enable better understanding of the origin of diverse pathological states of the cell as well as monitor cellular consequences of diseases and/or treatments.     

Hepatic Proliferation and Angiogenesis Markers Are Increased after Portal Deprivation in Rats: A Study of Molecular,Histological and Radiological Changes

MethodsWe conducted an experimental study comparing portacaval shunt (PCS), total portal vein ligation (PVL), and sham (S) operated rats. Each group were either sacrificed at 6 weeks (early) or 6 months (late). Arterial liver perfusion was studied in vivo using CT, and histopathological changes were noted. Liver mRNA levels were quantified by RT-QPCR for markers of inflammation (Il10, Tnfa), proliferation (Il6st, Mki67, Hgf, Hnf4a), angiogenesis: (Vegfa, Vegfr 1, 2 and 3; Pgf), oxidative stress (Nos2, and 3, Hif1a), and fibrosis (Tgfb). PCS and PVL were compared to the S group.ResultsPeriportal fibrosis and arterial proliferation was observed in late PCS and PVL groups. CT imaging demonstrated increased arterial liver perfusion in the PCS group. RT-QPCR showed increased inflammatory markers in PCS and PVL early groups. Tnfa and Il10 were increased in PCS and PVL late groups respectively. All proliferative markers increased in the PCS, and Hnf4a in the PVL early groups. Mki67 and Hnf4a were increased in the PCS late group. Nos3 was increased in the early and late PCS groups, and Hif1a was decreased in the PVL groups. Markers of angiogenesis were all increased in the early PCS group, and Vegfr3 and Pgf in the late PCS group. Only Vegfr3 was increased in the PVL groups. Tgf was increased in the PCS groups.ConclusionsPortal deprivation in rats induces a sustained increase in intrahepatic markers of inflammation, angiogenesis, proliferation, and fibrosis.     

Unique preservation of neural cells in Hutchinson- Gilford progeria syndrome is due to the expression of the neural-specific miR-9 microRNA

One puzzling observation in patients affected with Hutchinson-Gilford progeria syndrome (HGPS), who overall exhibit systemic and dramatic premature aging, is the absence of any conspicuous cognitive impairment. Recent studies based on induced pluripotent stem cells derived from HGPS patient cells have revealed a lack of expression in neural derivatives of lamin A, a major isoform of LMNA that is initially produced as a precursor called prelamin A. In HGPS, defective maturation of a mutated prelamin A induces the accumulation of toxic progerin in patient cells. Here, we show that a microRNA, miR-9, negatively controls lamin A and progerin expression in neural cells. This may bear major functional correlates, as alleviation of nuclear blebbing is observed in nonneural cells after miR-9 overexpression. Our results support the hypothesis, recently proposed from analyses in mice, that protection of neural cells from progerin accumulation in HGPS is due to the physiologically restricted expression of miR-9 to that cell lineage.     

Genetic and environmental variability of adult size in some stocks of the edible snail, Helix aspersa

This paper aims at providing a better knowledge of factors determining body size (especially the importance of heredity) in Helix aspersa Müller.
A preliminary investigation of snails from Maghreb allowed us to show the importance of heredity in the variability of body size and to produce an efficient design for a subsequent reliable estimation of heritabilities. All traits measured (diameter, height and weight) were highly correlated and weight appeared to be the most convenient measure of size.
The second experiment provided 4150 snails born from known individuals among 500 wild snails. Pedigrees were recorded. Weight and diameter revealed high heritabilities (>0.4), which is relevant for commercial selection since variability of both traits was important. The design also revealed a significant non-genetic maternal effect and also that offspring from pairs where only one animal laid were bigger than offspring from pairs where both animals laid. This surprising observation has to be confirmed and its mechanisms studied.