Development and External Validation of a Prediction Model for Colorectal Cancer Among Patients Awaiting Surveillance Colonoscopy Following Polypectomy.

Background and Aims: Demand for surveillance colonoscopy can sometimes exceed capacity, such as during and following the coronavirus disease 2019 pandemic, yet no tools exist to prioritize the patients most likely to be diagnosed with colorectal cancer (CRC) among those awaiting surveillance colonoscopy. We developed a multivariable prediction model for CRC at surveillance comparing performance to a model that assigned patients as low or high risk based solely on polyp characteristics (guideline-based model). Methods: Logistic regression was used for model development among patients receiving surveillance colonoscopy in 2014-2019. Candidate predictors included index colonoscopy indication, findings, and endoscopist adenoma detection rate, and patient and clinical characteristics at surveillance. Patients were randomly divided into model development (n = 36,994) and internal validation cohorts (n = 15,854). External validation was performed on 30,015 patients receiving surveillance colonoscopy in 2020-2022, and the multivariable model was then updated and retested. Results: One hundred fourteen, 43, and 71 CRCs were detected at surveillance in the 3 cohorts, respectively. Polyp size ≥10 mm, adenoma detection rate <32.5% or missing, patient age, and ever smoked tobacco were significant CRC predictors; this multivariable model outperformed the guideline-based model (internal validation cohort area under the receiver-operating characteristic curve: 0.73, 95% confidence interval (CI): 0.66-0.81 vs 0.52, 95% CI: 0.45-0.60). Performance declined at external validation but recovered with model updating (operating characteristic curve: 0.72 95% CI: 0.66-0.77). Conclusion: When surveillance colonoscopy demand exceeds capacity, a prediction model featuring common clinical predictors may help prioritize patients at highest risk for CRC among those awaiting surveillance. Also, regular model updates can address model performance drift.

Glucose-6-phosphate dehydrogenase deficiency accelerates arterial aging in diabetes.

AIMS: High glucose levels and Glucose-6-Phosphate Dehydrogenase deficiency (G6PDd) have both tissue inflammatory effects. Here we determined whether G6PDd accelerates arterial aging (information linked stiffening) in diabetes. METHODS: Plasma glucose, interleukin 6 (IL6), and arterial stiffness (indexed as carotid-femoral Pulse Wave Velocity, PWV) and red blood cell G6PD activity were assessed in a large (4448) Sardinian population. RESULTS: Although high plasma glucose in diabetics, did not differ by G6DP status (178.2 ± 55.1 vs 169.0 ± 50.1 mg/dl) in G6DPd versus non-G6PDd subjects, respectively, IL6, and PWV (adjusted for age and glucose) were significantly increased in G6PDd vs non-G6PDd subjects (PWV, 8.0 ± 0.4 vs 7.2 ± 0.2 m/sec) and (IL6, 6.9 ± 5.0 vs 4.2 ± 3.0 pg/ml). In non-diabetics, neither fasting plasma glucose, nor IL6, nor PWV were impacted by G6PDd. CONCLUSION: G6PDd in diabetics is associated with increased inflammatory markers and accelerated arterial aging.

EDA ligand triggers plasma membrane trafficking of its receptor EDAR via PKA activation and SNAP23-containing complexes.

BACKGROUND: Ectodysplasin-A (EDA), a skin-specific TNF ligand, interacts with its membrane receptor EDAR to trigger EDA signaling in skin appendage formation. Gene mutations in EDA signaling cause Anhidrotic/Hypohidrotic Ectodermal Dysplasia (A/HED), which affects the formation of skin appendages including hair, teeth, and several exocrine glands. RESULTS: We report that EDA triggers the translocation of its receptor EDAR from a cytosolic compartment into the plasma membrane. We use protein affinity purification to show that upon EDA stimulation EDAR associates with SNAP23-STX6-VAMP1/2/3 vesicle trafficking complexes. We find that EDA-dependent PKA activation is critical for the association. Notably, either of two HED-linked EDAR mutations, T346M and R420W, prevents EDA-induced EDAR translocation; and both EDA-induced PKA activation and SNAP23 are required for Meibomian gland (MG) growth in a skin appendage model. CONCLUSIONS: Overall, in a novel regulatory mechanism, EDA increases plasma membrane translocation of its own receptor EDAR, augmenting EDA-EDAR signaling in skin appendage formation. Our findings also provide PKA and SNAP23 as potential targets for the intervention of HED.

Development and Validation of a Diabetic Retinopathy Risk Stratification Algorithm.

OBJECTIVE: Although diabetic retinopathy is a leading cause of blindness worldwide, diabetes-related blindness can be prevented through effective screening, detection, and treatment of disease. The study goal was to develop risk stratification algorithms for the onset of retinal complications of diabetes, including proliferative diabetic retinopathy, referable retinopathy, and macular edema. RESEARCH DESIGN AND METHODS: Retrospective cohort analysis of patients from the Kaiser Permanente Northern California Diabetes Registry who had no evidence of diabetic retinopathy at a baseline diabetic retinopathy screening during 2008-2020 was performed. Machine learning and logistic regression prediction models for onset of proliferative diabetic retinopathy, diabetic macular edema, and referable retinopathy detected through routine screening were trained and internally validated. Model performance was assessed using area under the curve (AUC) metrics. RESULTS: The study cohort (N = 276,794) was 51.9% male and 42.1% White. Mean (±SD) age at baseline was 60.0 (±13.1) years. A machine learning XGBoost algorithm was effective in identifying patients who developed proliferative diabetic retinopathy (AUC 0.86; 95% CI, 0.86-0.87), diabetic macular edema (AUC 0.76; 95% CI, 0.75-0.77), and referable retinopathy (AUC 0.78; 95% CI, 0.78-0.79). Similar results were found using a simpler nine-covariate logistic regression model: proliferative diabetic retinopathy (AUC 0.82; 95% CI, 0.80-0.83), diabetic macular edema (AUC 0.73; 95% CI, 0.72-0.74), and referable retinopathy (AUC 0.75; 95% CI, 0.75-0.76). CONCLUSIONS: Relatively simple logistic regression models using nine readily available clinical variables can be used to rank order patients for onset of diabetic eye disease and thereby more efficiently prioritize and target screening for at risk patients.

Trends and Risk Factors for Venous Thromboembolism Among Hospitalized Medical Patients.

Importance: While hospital-associated venous thromboembolism (HA-VTE) is a known complication of hospitalization, contemporary incidence and outcomes data are scarce and methodologically contested. Objective: To define and validate an automated electronic health record (EHR)-based algorithm for retrospective detection of HA-VTE and examine contemporary HA-VTE incidence, previously reported risk factors, and outcomes. Design, Setting, and Participants: This cohort study was conducted using hospital admissions between January 1, 2013, and June 30, 2021, with follow-up until December 31, 2021. All medical (non-intensive care unit) admissions at an integrated health care delivery system with 21 hospitals in Northern California during the study period were included. Data were analyzed from January to June 2022. Exposures: Previously reported risk factors associated with HA-VTE and administration of pharmacological prophylaxis were evaluated as factors associated with HA-VTE. Main Outcomes and Measures: Yearly incidence rates and timing of HA-VTE events overall and by subtype (deep vein thrombosis, pulmonary embolism, both, or unknown), as well as readmissions and mortality rates. Results: Among 1 112 014 hospitalizations involving 529 492 patients (268 797 [50.8%] women; 75 238 Asian [14.2%], 52 697 Black [10.0%], 79 398 Hispanic [15.0%], and 307 439 non-Hispanic White [58.1%]; median [IQR] age, 67.0 [54.0-79.0] years), there were 13 843 HA-VTE events (1.2% of admissions) occurring in 10 410 patients (2.0%). HA-VTE events increased from 307 of 29 095 hospitalizations (1.1%) in the first quarter of 2013 to 551 of 33 729 hospitalizations (1.6%) in the first quarter of 2021. Among all HA-VTE events, 10 746 events (77.6%) were first noted after discharge. In multivariable analyses, several factors were associated with increased odds of HA-VTE, including active cancer (adjusted odds ratio [aOR], 1.96; 95% CI, 1.85-2.08), prior VTE (aOR, 1.71; 95% CI, 1.63-1.79), and reduced mobility (aOR, 1.63; 95% CI, 1.50-1.77). Factors associated with decreased likelihood of HA-VTE included Asian race (vs non-Hispanic White: aOR, 0.65; 95% CI, 0.61-0.69), current admission for suspected stroke (aOR, 0.73; 95% CI, 0.65-0.81), and Hispanic ethnicity (vs non-Hispanic White: aOR, 0.81; 95% CI, 0.77-0.86). HA-VTE events were associated with increased risk of readmission (hazard ratio [HR], 3.33; 95% CI, 3.25-3.41) and mortality (HR, 1.63; 95% CI, 1.57-1.70). Conclusions and Relevance: This study found that HA-VTE events occurred in 1.2% of medical admissions, increased over time, and were associated with increased adverse outcomes. These findings suggest that approaches designed to mitigate occurrence and outcomes associated with HA-VTE may remain needed.

Facets of Personality and Risk of Cognitive Impairment: Longitudinal Findings in a Rural Community from Sardinia.

BACKGROUND: Few studies have examined the associations between personality facets and dementia risk and rarely included individuals from rural settings or with low education. OBJECTIVE: To examine the association between personality and the risk of cognitive impairment. METHODS: Participants (N = 1,668; age 50 to 94 at baseline; 56.4% women; 86.5% less than high school diploma) were from a rural region of Sardinia (Italy) who completed the Revised NEO Personality Inventory (NEO-PI-R) during the first wave (2001-2004) and the Mini-Mental State Examination (MMSE) at waves two to five (2005-2021). Cox regression was used to test personality and covariates as predictors of cognitive impairment based on MMSE education-adjusted cutoffs. RESULTS: During the up to 18-year follow-up (M = 10.38; SD = 4.76), 187 individuals (11.2%) scored as cognitively impaired. Participants with higher neuroticism (particularly the depression facet [HR = 1.22, 95% CI = 1.06-1.40]), and lower agreeableness (particularly the modesty facet [HR = 0.83, 95% CI = 0.71-0.97]) and lower conscientiousness (particularly the dutifulness facet [HR = 0.78, 95% CI = 0.67-0.92]) were at higher risk of cognitive impairment. Lower warmth ([HR = 0.75, 95% CI = 0.65-0.87], facet of extraversion) and ideas ([HR = 0.76, 95% CI = 0.65-0.89], facet of openness) were also associated with increased risk of impairment. These associations were virtually unchanged in models that accounted for other risk factors, including smoking, depression, obesity, hypertension, diabetes, and apolipoprotein E (APOE) ɛ4 carrier status. Across the five domains, sex and the APOE variant did not moderate the associations. CONCLUSION: In a sample with demographic characteristics underrepresented in dementia research, this study identifies personality domains and facets most relevant to the risk of cognitive impairment.

Genome-wide meta-analysis of iron status biomarkers and the effect of iron on all-cause mortality in HUNT.

Iron is essential for many biological processes, but iron levels must be tightly regulated to avoid harmful effects of both iron deficiency and overload. Here, we perform genome-wide association studies on four iron-related biomarkers (serum iron, serum ferritin, transferrin saturation, total iron-binding capacity) in the Trøndelag Health Study (HUNT), the Michigan Genomics Initiative (MGI), and the SardiNIA study, followed by their meta-analysis with publicly available summary statistics, analyzing up to 257,953 individuals. We identify 123 genetic loci associated with iron traits. Among 19 novel protein-altering variants, we observe a rare missense variant (rs367731784) in HUNT, which suggests a role for DNAJC13 in transferrin recycling. We further validate recently published results using genetic risk scores for each biomarker in HUNT (6% variance in serum iron explained) and present linear and non-linear Mendelian randomization analyses of the traits on all-cause mortality. We find evidence of a harmful effect of increased serum iron and transferrin saturation in linear analyses that estimate population-averaged effects. However, there was weak evidence of a protective effect of increasing serum iron at the very low end of its distribution. Our findings contribute to our understanding of the genes affecting iron status and its consequences on human health.

PLAC1 affects cell to cell communication by interacting with the desmosome complex.

INTRODUCTION: X-linked PLAC1 is highly expressed in placenta during embryogenesis, and when ablated in mice, causes aberrant placental cell layer organization. It is also highly expressed in many types of cancer cell-lines. Although it has been shown that it promotes AKT phosphorylation in cancer cells, the exact mechanism by which it influences placental layer differentiation is unclear. METHODS: To investigate the mechanism of action of PLAC1 we did cell fractionation and immunoprecipitation of the protein and Mass Spectrometry analysis to identify its interaction partners. The associated proteins were directly tested for interactions by co-transfection with PLAC1 and immunoprecipitation. Mutations in the ZP-N domain of PLAC1 were introduced to assess its involvement in the interactions. RESULTS: We provide evidence that Desmoglein-2 (DSG2), a component of the membrane-associated desmosomal complex, directly interacts with PLAC1. Mutations of cysteines in ZP-N domain disrupt the interaction between PLAC1 and DSG-2. DISCUSSION: Because desmosomes are responsible for establishing lateral cell-cell junctions, we suggest that direct interaction with the lateral junction protein complex may be implicated in the PLAC1 effects on cell-cell interactions, and thereby on the layer structure of the placenta.

Arterial stiffness and multiple organ damage: a longitudinal study in population.

AIMS: Previous cross-sectional observation identified arterial aging, indexed as pulse-wave velocity (PWV), as a key determinant of the simultaneous multiple organ damage (heart, carotid artery, and kidney). The aim of the present cohort study is to investigate trajectories of repeated measures of PWV and traditional CV risk factors in subjects who eventually presented clinical evidence of multiple organ damage in the SardiNIA study. METHODS AND RESULTS: Organ damage was measured in the heart (left ventricular hypertrophy, LVH), the common carotid artery (intima-media thickness > 0.9 mm and/or plaque), and the kidney (eGFR < 60 ml/min/1.73 m2) of 2130 men and women of a broad age range participating the SardiNIA study. SHATS was defined as the simultaneous occurrence of all the three-organ damages. Trajectory in traditional CV risk factors and PWV was analyzed retrospectively (four observations over 9 years) according to the number of organ damage (from 0 to 3). Compared to subjects with no organ damage, after controlling for traditional CV risk factors, each 1 m/s increase in baseline PWV was accompanied by a 93% higher odds of developing SHATS; and each 1 cm/s (0.01 m/s) annual increase in PWV by a 31% greater odds of developing SHATS. CONCLUSIONS: Arterial stiffness, a proxy of arterial aging that can be measured clinically as PWV, is an integrated predictive marker of multiple age-associated organ damage recognized as clinical diseases.

Homeostatic Control of Sebaceous Glands by Innate Lymphoid Cells Regulates Commensal Bacteria Equilibrium.

Immune cells and epithelium form sophisticated barrier systems in symbiotic relationships with microbiota. Evidence suggests that immune cells can sense microbes through intact barriers, but regulation of microbial commensalism remain largely unexplored. Here, we uncovered spatial compartmentalization of skin-resident innate lymphoid cells (ILCs) and modulation of sebaceous glands by a subset of RORγt+ ILCs residing within hair follicles in close proximity to sebaceous glands. Their persistence in skin required IL-7 and thymic stromal lymphopoietin, and localization was dependent on the chemokine receptor CCR6. ILC subsets expressed TNF receptor ligands, which limited sebocyte growth by repressing Notch signaling pathway. Consequently, loss of ILCs resulted in sebaceous hyperplasia with increased production of antimicrobial lipids and restricted commensalism of Gram-positive bacterial communities. Thus, epithelia-derived signals maintain skin-resident ILCs that regulate microbial commensalism through sebaceous gland-mediated tuning of the barrier surface, highlighting an immune-epithelia circuitry that facilitates host-microbe symbiosis.

Exome Chip Meta-analysis Fine Maps Causal Variants and Elucidates the Genetic Architecture of Rare Coding Variants in Smoking and Alcohol Use.

BACKGROUND: Smoking and alcohol use have been associated with common genetic variants in multiple loci. Rare variants within these loci hold promise in the identification of biological mechanisms in substance use. Exome arrays and genotype imputation can now efficiently genotype rare nonsynonymous and loss of function variants. Such variants are expected to have deleterious functional consequences and to contribute to disease risk. METHODS: We analyzed ∼250,000 rare variants from 16 independent studies genotyped with exome arrays and augmented this dataset with imputed data from the UK Biobank. Associations were tested for five phenotypes: cigarettes per day, pack-years, smoking initiation, age of smoking initiation, and alcoholic drinks per week. We conducted stratified heritability analyses, single-variant tests, and gene-based burden tests of nonsynonymous/loss-of-function coding variants. We performed a novel fine-mapping analysis to winnow the number of putative causal variants within associated loci. RESULTS: Meta-analytic sample sizes ranged from 152,348 to 433,216, depending on the phenotype. Rare coding variation explained 1.1% to 2.2% of phenotypic variance, reflecting 11% to 18% of the total single nucleotide polymorphism heritability of these phenotypes. We identified 171 genome-wide associated loci across all phenotypes. Fine mapping identified putative causal variants with double base-pair resolution at 24 of these loci, and between three and 10 variants for 65 loci. Twenty loci contained rare coding variants in the 95% credible intervals. CONCLUSIONS: Rare coding variation significantly contributes to the heritability of smoking and alcohol use. Fine-mapping genome-wide association study loci identifies specific variants contributing to the biological etiology of substance use behavior.

Adipocytokine correlations with thyroid function and autoimmunity in euthyroid sardinians.

OBJECTIVE: Cytokines release by adipocytes could interact with TSH secretion. We evaluated the relationship between adipocytokines and TSH. We further tested for association of cytokines and thyroid autoimmunity. METHODS: We conducted a cross-sectional study in a community-based sample including 5385 individuals (2964 female) with TSH within the reference range. Subjects who reported taking thyroid medications or drugs that alter thyroid function were excluded. TSH, FT4, adiponectin, leptin, antibody against thyroperoxidase and against thyroglobulin were measured. Linear and logistic regression models were used to test for association. RESULTS: Females had higher adiponectin and leptin level and increased frequency of thyroid antibodies. In multiple regression analysis TSH was directly associated with leptin (ß = 0.003, p = 0.001) and the presence of circulating antibody against thyroperoxidase (ß = 0.315, p < 0.001), but negatively associated with age (ß = -0.012, p < 0.001) and FT4 (ß = -0.359, p < 0.001). Adiponectin, the presence of antibody against thyroglobulin and smoking habit were not associated with TSH levels (p = 0.223, p = 0.174 and p = 0.788, respectively). Logistic regression analysis revealed that higher adiponectin levels were associated with thyroid autoimmunity. CONCLUSIONS: Leptin is positively associated with TSH levels in euthyroid individuals, suggesting an effect of the adipokine on TSH secretion. Our results support the hypothesis that the leptin and pituitary-thyroid axis might interact in the context of energy homeostasis. The effect of adiponectin on thyroid autoimmunity will require more studies.

Eda-activated RelB recruits an SWI/SNF (BAF) chromatin-remodeling complex and initiates gene transcription in skin appendage formation.

Ectodysplasin A (Eda) signaling activates NF-κB during skin appendage formation, but how Eda controls specific gene transcription remains unclear. Here, we find that Eda triggers the formation of an NF-κB-associated SWI/SNF (BAF) complex in which p50/RelB recruits a linker protein, Tfg, that interacts with BAF45d in the BAF complex. We further reveal that Tfg is initially induced by Eda-mediated RelB activation and then bridges RelB and BAF for subsequent gene regulation. The BAF component BAF250a is particularly up-regulated in skin appendages, and epidermal knockout of BAF250a impairs skin appendage development, resulting in phenotypes similar to those of Eda-deficient mouse models. Transcription profiling identifies several target genes regulated by Eda, RelB, and BAF. Notably, RelB and the BAF complex are indispensable for transcription of Eda target genes, and both BAF complex and Eda signaling are required to open chromatin of Eda targets. Our studies thus suggest that Eda initiates a signaling cascade and recruits a BAF complex to specific gene loci to facilitate transcription during organogenesis.

Foxc1 Ablated Mice Are Anhidrotic and Recapitulate Features of Human Miliaria Sweat Retention Disorder.

Sweat glands are critical for thermoregulation. The single tubular structure of sweat glands has a lower secretory portion and an upper reabsorptive duct leading to the secretory pore in the skin. Genes that determine sweat gland structure and function are largely unidentified. Here we report that a Fox family transcription factor, Foxc1, is obligate for appreciable sweat duct activity in mice. When Foxc1 was specifically ablated in skin, sweat glands appeared mature, but the mice were severely hypohidrotic. Morphologic analysis revealed that sweat ducts were blocked by hyperkeratotic or parakeratotic plugs. Consequently, lumens in ducts and secretory portions were dilated, and blisters and papules formed on the skin surface in the knockout mice. The phenotype was strikingly similar to the human sweat retention disorder miliaria. We further show that Foxc1 deficiency ectopically induces the expression of keratinocyte terminal differentiation markers in the duct luminal cells, which most likely contribute to keratotic plug formation. Among those differentiation markers, we show that Sprr2a transcription is directly repressed by overexpressed Foxc1 in keratinocytes. In summary, Foxc1 regulates sweat duct luminal cell differentiation, and mutant mice mimic miliaria and provide a possible animal model for its study.

Menopause modulates the association between thyrotropin levels and lipid parameters: The SardiNIA study.

OBJECTIVE: Thyroid hormone influences lipoprotein metabolism. The role of menopausal status in this association has not been extensively studied. The aim of the present study is to evaluate the association between lipid parameters and mild elevations of thyrotropin (TSH), and whether menopause influences this relationship. STUDY DESIGN: A cross-sectional study was conducted with a sample of 2,914 women (aged 14-102 years) from the SardiNIA study. MAIN OUTCOME MEASURES: The association of TSH with blood lipid levels was examined using regression analyses, according to menopausal status. RESULTS: Postmenopausal women had lower serum TSH concentrations and higher levels of total cholesterol, low-density lipoprotein cholesterol (LDLc), high-density lipoprotein cholesterol (HDLc), and triglycerides than did premenopausal women (p=0.001 or less for all). In premenopausal women, after adjusting for the confounders age, BMI, smoking, insulin and glycaemia, TSH showed a direct relation to the levels of total cholesterol (ß=0.046, p=0.010), LDLc (ß=0.044, p=0.016) and triglycerides (ß=0.085, p<0.001), but no association with HDLc level. In the postmenopausal group, TSH was directly associated only with triglyceride levels (ß=0.103, p=0.014). CONCLUSIONS: The association between mild elevation of TSH and lipid levels is influenced by menopausal status. Further research is needed to clarify this finding.

Induction of specific neuron types by overexpression of single transcription factors.

Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of the combined action of multiple added growth factors, which generally tend to result in mixed populations of neurons. Here, we report that overexpression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron lineages. Analysis of data on gene expression changes 2 d after induction of each of 185 TFs implicated candidate TFs for further ESC differentiation studies. Induction of 23 TFs (out of 49 TFs tested) for 6 d facilitated neural differentiation of ESCs as inferred from increased proportion of cells with neural progenitor marker PSA-NCAM. We identified early activation of the Notch signaling pathway as a common feature of most potent inducers of neural differentiation. The majority of neuron-like cells generated by induction of Ascl1, Smad7, Nr2f1, Dlx2, Dlx4, Nr2f2, Barhl2, and Lhx1 were GABA-positive and expressed other markers of GABAergic neurons. In the same way, we identified Lmx1a and Nr4a2 as inducers for neurons bearing dopaminergic markers and Isl1, Fezf2, and St18 for cholinergic motor neurons. A time-course experiment with induction of Ascl1 showed early upregulation of most neural-specific messenger RNA (mRNA) and microRNAs (miRNAs). Sets of Ascl1-induced mRNAs and miRNAs were enriched in Ascl1 targets. In further studies, enrichment of cells obtained with the induction of Ascl1, Smad7, and Nr2f1 using microbeads resulted in essentially pure population of neuron-like cells with expression profiles similar to neural tissues and expressed markers of GABAergic neurons. In summary, this study indicates that induction of transcription factors is a promising approach to generate cultures that show the transcription profiles characteristic of specific neural cell types.

Emergence of undifferentiated colonies from mouse embryonic stem cells undergoing differentiation by retinoic acid treatment.

Retinoic acid (RA) is one of the most potent inducers of differentiation of mouse embryonic stem cells (ESCs). However, previous studies show that RA treatment of cells cultured in the presence of a leukemia inhibitory factor (LIF) also result in the upregulation of a gene called Zscan4, whose transient expression is a marker for undifferentiated ESCs. We explored the balance between these two seemingly antagonistic effects of RA. ESCs indeed differentiated in the presence of LIF after RA treatment, but colonies of undifferentiated ESCs eventually emerged from these differentiated cells - even in the presence of RA. These colonies, named secondary colonies, consist of three cell types: typical undifferentiated ESCs expressing pluripotency genes such as Pou5f1, Sox2, and Nanog; cells expressing Zscan4; and endodermal-like cells located at the periphery of the colony. The capacity to form secondary colonies was confirmed for all eight tested ESC lines. Cells from the secondary colonies - after transfer to the standard ESC medium - retained pluripotency, judged by their strong alkaline phosphatase (ALP) staining, typical colony morphology, gene expression profile, stable karyotype, capacity to differentiate into all three germ layers in embryoid body formation assays, and successful contribution to chimeras after injection into blastocysts. Based on flow cytometry analysis (FACS), the proportion of Zscan4-positive cells in secondary colonies was higher than in standard ESC colonies, which may explain the capacity of ESCs to resist the differentiating effects of RA and instead form secondary colonies of undifferentiated ESCs. This hypothesis is supported by cell-lineage tracing analysis, which showed that most cells in the secondary colonies were descendents of cells transiently expressing Zscan4.

Sex-Specific Parental Effects on Offspring Lipid Levels.

BACKGROUND: Plasma lipid levels are highly heritable traits, but known genetic loci can only explain a small portion of their heritability. METHODS AND RESULTS: In this study, we analyzed the role of parental levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and triglycerides (TGs) in explaining the values of the corresponding traits in adult offspring. We also evaluated the contribution of nongenetic factors that influence lipid traits (age, body mass index, smoking, medications, and menopause) alone and in combination with variability at the genetic loci known to associate with TC, LDL-C, HDL-C, and TG levels. We performed comparisons among different sex-specific regression models in 416 families from the Framingham Heart Study and 304 from the SardiNIA cohort. Models including parental lipid levels explain significantly more of the trait variation than models without these measures, explaining up to ≈39% of the total trait variation. Of this variation, the parent-of-origin effect explains as much as ≈15% and it does so in a sex-specific way. This observation is not owing to shared environment, given that spouse-pair correlations were negligible (<1.5% explained variation in all cases) and is distinct from previous genetic and acquired factors that are known to influence serum lipid levels. CONCLUSIONS: These findings support the concept that unknown genetic and epigenetic contributors are responsible for most of the heritable component of the plasma lipid phenotype, and that, at present, the clinical utility of knowing age-matched parental lipid levels in assessing risk of dyslipidemia supersedes individual locus effects. Our results support the clinical utility of knowing parental lipid levels in assessing future risk of dyslipidemia.

Genome-wide association study of susceptibility loci for breast cancer in Sardinian population.

BACKGROUND: Despite progress in identifying genes associated with breast cancer, many more risk loci exist. Genome-wide association analyses in genetically-homogeneous populations, such as that of Sardinia (Italy), could represent an additional approach to detect low penetrance alleles. METHODS: We performed a genome-wide association study comparing 1431 Sardinian patients with non-familial, BRCA1/2-mutation-negative breast cancer to 2171 healthy Sardinian blood donors. DNA was genotyped using GeneChip Human Mapping 500 K Arrays or Genome-Wide Human SNP Arrays 6.0. To increase genomic coverage, genotypes of additional SNPs were imputed using data from HapMap Phase II. After quality control filtering of genotype data, 1367 cases (9 men) and 1658 controls (1156 men) were analyzed on a total of 2,067,645 SNPs. RESULTS: Overall, 33 genomic regions (67 candidate SNPs) were associated with breast cancer risk at the p < 0(-6) level. Twenty of these regions contained defined genes, including one already associated with breast cancer risk: TOX3. With a lower threshold for preliminary significance to p < 10(-5), we identified 11 additional SNPs in FGFR2, a well-established breast cancer-associated gene. Ten candidate SNPs were selected, excluding those already associated with breast cancer, for technical validation as well as replication in 1668 samples from the same population. Only SNP rs345299, located in intron 1 of VAV3, remained suggestively associated (p-value, 1.16 x 10(-5)), but it did not associate with breast cancer risk in pooled data from two large, mixed-population cohorts. CONCLUSIONS: This study indicated the role of TOX3 and FGFR2 as breast cancer susceptibility genes in BRCA1/2-wild-type breast cancer patients from Sardinian population.