Diurnal variations of resting-state fMRI data : a graph-based analysis

Circadian rhythms (lasting approximately 24 h) control and entrain various physiological processes, ranging from neural activity and hormone secretion to sleep cycles and eating habits. Several studies have shown that time of day (TOD) is associated with human cognition and brain functions. In this study, utilizing a chronotype-based paradigm, we applied a graph theory approach on resting-state functional MRI (rs-fMRI) data to compare whole-brain functional network topology between morning and evening sessions and between morning-type (MT) and evening-type (ET) participants. Sixty-two individuals (31 MT and 31 ET) underwent two fMRI sessions, approximately 1 hour (morning) and 10 h (evening) after their wake-up time, according to their declared habitual sleep-wake pattern on a regular working day. In the global analysis, the findings revealed the effect of TOD on functional connectivity (FC) patterns, including increased small-worldness, assortativity, and synchronization across the day. However, we identified no significant differences based on chronotype categories. The study of the modular structure of the brain at mesoscale showed that functional networks tended to be more integrated with one another in the evening session than in the morning session. Local/regional changes were affected by both factors (i.e., TOD and chronotype), mostly in areas associated with somatomotor, attention, frontoparietal, and default networks. Furthermore, connectivity and hub analyses revealed that the somatomotor, ventral attention, and visual networks covered the most highly connected areas in the morning and evening sessions: the latter two were more active in the morning sessions, and the first was identified as being more active in the evening. Finally, we performed a correlation analysis to determine whether global and nodal measures were associated with subjective assessments across participants. Collectively, these findings contribute to an increased understanding of diurnal fluctuations in resting brain activity and highlight the role of TOD in future studies on brain function and the design of fMRI experiments

Serotoninergic receptor ligands improve Tamoxifen effectiveness on breast cancer cells

Background: Serotonin (or 5-Hydroxytryptamine, 5-HT) signals in mammary gland becomes dysregulated in cancer, also contributing to proliferation, metastasis, and angiogenesis. Thus, the discovery of novel compounds targeting serotonin signaling may contribute to tailor new therapeutic strategies usable in combination with endocrine therapies. We have previously synthesized serotoninergic receptor ligands (SER) with high affinity and selectivity towards 5-HT2A and 5-HT2C receptors, the main mediators of mitogenic effect of serotonin in breast cancer (BC). Here, we investigated the effect of 10 SER on viability of MCF7, SKBR3 and MDA-MB231 BC cells and focused on their potential ability to affect Tamoxifen responsiveness in ER+ cells. Methods: Cell viability has been assessed by sulforhodamine B assay. Cell cycle has been analyzed by flow cytometry. Gene expression of 5-HT receptors and Connective Tissue Growth Factor (CTGF) has been checked by RT-PCR; mRNA levels of CTGF and ABC transporters have been further measured by qPCR. Protein levels of 5-HT2C receptors have been analyzed by Western blot. All data were statistically analyzed using GraphPad Prism 7. Results: We found that treatment with SER for 72 h reduced viability of BC cells. SER were more effective on MCF7 ER+ cells (IC50 range 10.2 μM - 99.2 μM) compared to SKBR3 (IC50 range 43.3 μM - 260 μM) and MDA-MB231 BC cells (IC50 range 91.3 μM - 306 μM). This was paralleled by accumulation of cells in G0/G1 phase of cell cycle. Next, we provided evidence that two ligands, SER79 and SER68, improved the effectiveness of Tamoxifen treatment in MCF7 cells and modulated the expression of CTGF, without affecting viability of MCF10A non-cancer breast epithelial cells. In a cell model of Tamoxifen resistance, SER68 also restored drug effect independently of CTGF. Conclusions: These results identified serotoninergic receptor ligands potentially usable in combination with Tamoxifen to improve its effectiveness on ER+ BC patients

Indicators for assessing the quality of refractive error care

Significance: Quality refractive error care is essential for reducing vision impairment. Quality indicators and standardized approaches for assessing the quality of refractive error care need to be established. Purpose: This study aimed to develop a set of indicators for assessing the quality of refractive error care and test their applicability in a real-world setting using unannounced standardized patients (USPs). Methods: Patient outcomes and three quality of refractive error care (Q.REC) indicators (1, optimally prescribed spectacles; 2, adequately prescribed spectacles; 3, vector dioptric distance) were developed using existing literature, refraction training standards, and consulting educators. Twenty-one USPs with various refractive errors were trained to visit optical stores across Vietnam to have a refraction, observe techniques, and order spectacles. Spectacles were assessed against each Q.REC indicator and tested for associations with vision and comfort. Results: Overall, 44.1% (184/417) of spectacles provided good vision and comfort. Of the spectacles that met Q.REC indicators 1 and 2, 62.5 and 54.9%, respectively, provided both good vision and comfort. Optimally prescribed spectacles (indicator 1) were significantly more likely to provide good vision and comfort independently compared with spectacles that did not meet any indicator (good vision: 94.6 vs. 85.0%, P =.01; comfortable: 66.1 vs. 36.3%, P <.01). Adequately prescribed spectacles (indicator 2) were more likely to provide good comfort compared with spectacles not meeting any indicator (57.7 vs. 36.3%, P <.01); however, vision outcomes were not significantly different (85.9 vs. 85.0%, P =.90). Good vision was associated with a lower mean vector dioptric distance (P <.01) but not with comfort (P =.52). Conclusions: The optimally prescribed spectacles indicator is a promising approach for assessing the quality of refractive error care without additional assessments of vision and comfort. Using USPs is a practical approach and could be used as a standardized method for evaluating the quality of refractive error care

SSBP1-Disease Update: Expanding the Genetic and Clinical Spectrum, Reporting Variable Penetrance and Confirming Recessive Inheritance

Purpose: To report novel genotypes and expand the phenotype spectrum of SSBP1-disease and explore potential disease mechanism. / Methods: Five families with previously unsolved optic atrophy and retinal dystrophy underwent whole genome sequencing as part of the National Institute for Health Research BioResource Rare-Diseases and the UK's 100,000 Genomes Project. In silico analysis and protein modelling was performed on the identified variants. Deep phenotyping including retinal imaging and International Society for Clinical Electrophysiology of Vision standard visual electrophysiology was performed. / Results: Seven individuals from five unrelated families with bilateral optic atrophy and/or retinal dystrophy with extraocular signs and symptoms in some are described. In total, 6 SSBP1 variants were identified including the previously unreported variants: c.151A>G, p.(Lys51Glu), c.335G>A p.(Gly112Glu), and c.380G>A, p.(Arg127Gln). One individual was found to carry biallelic variants (c.380G>A p.(Arg127Gln); c.394A>G p.(Ile132Val)) associated with likely autosomal recessive SSBP1-disease. In silico analysis predicted all variants to be pathogenic and Three-dimensional protein modelling suggested possible disease mechanisms via decreased single-stranded DNA binding affinity or impaired higher structure formation. / Conclusions: SSBP1 is essential for mitochondrial DNA replication and maintenance, with defects leading to a spectrum of disease that includes optic atrophy and/or retinal dystrophy, occurring with or without extraocular features. This study provides evidence of intrafamilial variability and confirms the existence of an autosomal recessive inheritance in SSBP1-disease consequent upon a previously unreported genotype

Adenoviral gene transfer of PLD1-D4 enhances insulin sensitivity in mice by disrupting phospholipase D1 interaction with PED/PEA-15.

Over-expression of phosphoprotein enriched in diabetes/phosphoprotein enriched in astrocytes (PED/PEA-15) causes insulin resistance by interacting with the D4 domain of phospholipase D1 (PLD1). Indeed, the disruption of this association restores insulin sensitivity in cultured cells over-expressing PED/PEA-15. Whether the displacement of PLD1 from PED/PEA-15 improves insulin sensitivity in vivo has not been explored yet. In this work we show that treatment with a recombinant adenoviral vector containing the human D4 cDNA (Ad-D4) restores normal glucose homeostasis in transgenic mice overexpressing PED/PEA-15 (Tg ped/pea-15) by improving both insulin sensitivity and secretion. In skeletal muscle of these mice, D4 over-expression inhibited PED/PEA-15-PLD1 interaction, decreased Protein Kinase C alpha activation and restored insulin induced Protein Kinase C zeta activation, leading to amelioration of insulin-dependent glucose uptake. Interestingly, Ad-D4 administration improved insulin sensitivity also in high-fat diet treated obese C57Bl/6 mice. We conclude that PED/PEA-15-PLD1 interaction may represent a novel target for interventions aiming at improving glucose tolerance

Platelet-rich plasma counteracts detrimental effect of high-glucose concentrations on mesenchymal stem cells from Bichat fat pad

Diabetic patients display increased risk of periodontitis and failure in bone augmentation procedures. Mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) represent a relevant advantage in tissue repair process and regenerative medicine. We isolated MSCs from Bichat's buccal fat pad (BFP) and measured the effects of glucose and PRP on cell number and osteogenic differentiation potential. Cells were cultured in the presence of 5.5-mM glucose (low glucose [LG]) or 25-mM glucose (high glucose [HG]). BFP–MSC number was significantly lower when cells were cultured in HG compared with those in LG. Following osteogenic differentiation procedures, calcium accumulation, alkaline phosphatase activity, and expression of osteogenic markers were significantly lower in HG compared with LG. Exposure of BFP–MSC to PRP significantly increased cell number and osteogenic differentiation potential, reaching comparable levels in LG and in HG. Thus, high-glucose concentrations impair BFP–MSC growth and osteogenic differentiation. However, these detrimental effects are largely counteracted by PRP

Clinical and surgical data of affected members of a classic CFEOM 1 family

BACKGROUND: Congenital fibiosis of the extraocular muscles (CFEOM1) refers to a group of congenital eye movement disorders that are characterized by non-progressive restrictive ophthalmoplegia. We present clinical and surgical data on affected members of a classic CFEOM1 family. METHODS: Ten members of a fifteen-member, three-generation Italian family affected by classic CFEOM participated in this study. Each affected family member underwent ophthalmologic (corrected visual acuity, pupillary function, anterior segment and fundus examination), orthoptic (cover test, cover-uncover test, prism alternate cover test), and preoperative examinations. Eight of the ten affected members had surgery and underwent postoperative examinations. Surgical procedures are listed. RESULTS: All affected members were born with varying degrees of bilateral ptosis and ophthalmoplegia with both eyes fixed in a hypotropic position (classic CFEOM). The affected members clinical data prior to surgery, surgery procedures and postoperative outcomes are presented. On 14 operated eyes to correct ptosis there was an improvement in 12 eyes. In addition, the head position improved in all patients. CONCLUSIONS: Surgery is effective at improving ptosis in the majority of patients with classic CFEOM. However, the surgical approach should be individualized to each patient, as inherited CFEOM exhibits variable expressivity and the clinical features may differ markedly between affected individuals, even within the same family

Novel USH1G homozygous variant underlying USH2-like phenotype of Usher syndrome

PURPOSE: Usher syndrome (USH) is an autosomal recessive disorder characterized by congenital sensorineural hearing impairment and retinitis pigmentosa. Classification distinguishes three clinical types of which type I (USH1) is the most severe, with vestibular dysfunction as an added feature. To date, 15 genes and 3 loci have been identified with the USH1G gene being an uncommon cause of USH. We describe an atypical USH1G-related phenotype caused by a novel homozygous missense variation in a patient with profound hearing impairment and relatively mild retinitis pigmentosa, but no vestibular dysfunction. METHODS: A 26-year-old female patient with profound congenital sensorineural hearing loss, nyctalopia and retinitis pigmentosa was studied. Audiometric, vestibular and ophthalmologic examination was performed. A panel of 13 genes was tested by next-generation sequencing (NGS). RESULTS: While the hearing loss was confirmed to be profound, the vestibular function resulted normal. Although typical retinitis pigmentosa was present, the age at onset was unusually late for USH1 syndrome. A novel homozygous missense variation (c.1187T>A, p.Leu396Gln) in the USH1G gene has been identified as causing the disease in our patient. CONCLUSIONS: Genetic and phenotypic heterogeneity are very common in both isolated and syndromic retinal dystrophies and sensorineural hearing loss. Our findings widen the spectrum of USH allelic disorders and strength the concept that variants in genes that are classically known as underlying one specific clinical USH subtype might result in unexpected phenotypes

po 292 breast cancer stem cell reprogramming deciphering the impact of glucose and the contribute of tumour microenvironment

Introduction Diabetes-associated hyperglycemia is linked to poorer prognosis and survival in breast cancer (BC). Indeed, glucose can affect both tumour and tumour-surrounding cells. BC cells are embedded in an adipocyte-rich microenvironment, which, beside adipocytes, contains Stromal-Vascular Fraction Cells (SVFCs). In this scenario, epithelial and stromal compartments communicate through the release of soluble factors and establish an intricate crosstalk. Here, we analysed whether glucose could directly affect the phenotype of ER+ MCF7 BC cells and interfere with their interaction with adipose-derived SVFCs, thereby promoting tumour progression. Material and methods MCF7 cell stemness markers were measured by qReal-Time PCR. Adipose-derived (Ad-)SVFCs were obtained by mammary adipose tissue specimens of women undergoing plastic surgery. The trascriptome of MCF7 exposed to either low (LG-5.5 mM) or high glucose concentration (HG-25 mM) was obtained by RNA-Sequencing (Illumina HiSeq3000). Results and discussions HG exposure of MCF7 determined a significant increase of SOX2 mRNA levels as compared to LG, suggesting the induction of stemness programming. Co-culture with Ad-SVFCs in HG increased SOX2, NANOG and OCT4 mRNA levels in MCF7, as compared to isolated culture, indicating the involvement of SVF-produced soluble factors in BC stem cell reprogramming. Moreover, in presence of Ad-SVFCs and HG, MCF7 produced a higher number of mammospheres, which also displayed larger size. However, both in LG and in HG, conditioned media (CM) obtained from Ad-SVFCs produced no relevant effect on MCF7 stemness. Nevertheless, when Ad-SVFCs were pre-incubated with CM obtained from HG-treated MCF7, their CM very effectively increased OCT4, NANOG and SOX2 mRNA levels in MCF7. Thus, HG likely perturbs MCF7, which produce soluble factors leading Ad-SVFCs to release, in turn, reprogramming factors for BC cell stemness. In this regard, we have observed that HG modification of MCF7 transcriptome includes deregulation of 17 genes (pval=0.05) encoding for secreted proteins involved in cancer progression-related pathways, which may potentially play a role in tumour-stroma interactions. Conclusion Glucose affects BC stem cell reprogramming both directly and through Ad-SVFCs. Deciphering the mechanisms that govern this intricate crosstalk will pave the way to new targeted strategies to improve BC control in conditions of metabolic derangement