Modulation by decitabine of gene expression and growth of osteosarcoma U2OS cells in vitro and in xenografts: Identification of apoptotic genes as targets for demethylation

<p>Abstract</p> <p>Background</p> <p>Methylation-mediated silencing of genes is one epigenetic mechanism implicated in cancer. Studies regarding the role of modulation of gene expression utilizing inhibitors of DNA methylation, such as decitabine, in osteosarcoma (OS) have been limited. A biological understanding of the overall effects of decitabine in OS is important because this particular agent is currently undergoing clinical trials. The objective of this study was to measure the response of the OS cell line, U2OS, to decitabine treatment both <it>in vitro </it>and <it>in vivo</it>.</p> <p>Results</p> <p>Microarray expression profiling was used to distinguish decitabine-dependent changes in gene expression in U2OS cells, and to identify responsive loci with demethylated CpG promoter regions. U2OS xenografts were established under the sub-renal capsule of immune-deficient mice to study the effect of decitabine <it>in vivo </it>on tumor growth and differentiation. Reduced nuclear methylation levels could be detected in xenografts derived from treated mice by immunohistochemistry utilizing a 5-methylcytidine antibody. Decitabine treatment reduced tumor xenograft size significantly (p < 0.05). Histological analysis of treated U2OS xenograft sections revealed a lower mitotic activity (p < 0.0001), increased bone matrix production (p < 0.0001), and a higher number of apoptotic cells (p = 0.0329). Microarray expression profiling of U2OS cultured cells showed that decitabine treatment caused a significant induction (p < 0.0025) in the expression of 88 genes. Thirteen had a ≥2-fold change, 11 of which had CpG-island-associated promoters. Interestingly, 6 of these 11 were pro-apoptotic genes and decitabine resulted in a significant induction of cell death in U2OS cells <it>in vitro </it>(p < 0.05). The 6 pro-apoptotic genes (<it>GADD45A</it>, <it>HSPA9B</it>, <it>PAWR</it>, <it>PDCD5</it>, <it>NFKBIA</it>, and <it>TNFAIP3</it>) were also induced to ≥2-fold <it>in vivo</it>. Quantitative methylation pyrosequencing confirmed that the tested pro-apoptotic genes had CpG-island DNA demethylationas a result of U2OS decitabine treatment both <it>in vitro </it>and in xenografts</p> <p>Conclusion</p> <p>These data provide new insights regarding the use of epigenetic modifiers in OS, and have important implications for therapeutic trials involving demethylation drugs. Collectively, these data have provided biological evidence that one mode of action of decitabine may be the induction of apoptosis utilizing promoter-CpG demethylation of specific effectors in cell death pathways in OS.</p

Loss of immune escape mutations during persistent HCV infection in pregnancy enhances replication of vertically transmitted viruses

Globally, about 1% of pregnant women are persistently infected with the hepatitis C virus (HCV)1. Vertical transmission occurs in 3–5% of cases2 and accounts for most new childhood HCV infections1,3. HCV-specific CD8+ cytotoxic T-lymphocytes (CTLs) play a vital role in the clearance of acute infections4–6, but in the 60–80% of infections that persist these cells become functionally exhausted or select mutant viruses that escape T-cell recognition7–9. Increased HCV replication during pregnancy10,11 suggests that maternofetal immune tolerance mechanisms12 may further impair HCV-specific CTLs, limiting their selection pressure on persistent viruses. To assess this possibility, we characterized the circulating viral quasispecies during and after consecutive pregnancies. This revealed a loss of some escape mutations in class I epitopes in pregnancy associated with emergence of more fit viruses13. CTL selection pressure was reimposed after childbirth, when escape mutations in these epitopes again predominated in the quasispecies and viral load dropped sharply14. Importantly, viruses transmitted perinatally were those with enhanced fitness due to reversion of escape mutations. Our findings indicate that immunoregulatory changes of pregnancy reduce CTL selection pressure on HCV class I epitopes, thereby facilitating vertical transmission of viruses with optimized replicative fitness

Outcomes of Induction vs Prelabor Cesarean Delivery at \u3c33 Weeks for Hypertensive Disorders of Pregnancy

BACKGROUND: Hypertensive disorders of pregnancy are the leading cause of indicated preterm birth; however, the optimal delivery approach for pregnancies complicated by preterm hypertensive disorders of pregnancy remains uncertain. OBJECTIVE: This study aimed to compare maternal and neonatal morbidity in patients with hypertensive disorders of pregnancy who either went induction of labor or prelabor cesarean delivery at \u3c33 \u3eweeks\u27 gestation. In addition, we aimed to quantify the length of induction of labor and rate of vaginal delivery in those who underwent induction of labor. STUDY DESIGN: This is a secondary analysis of an observational study which included 115,502 patients in 25 hospitals in the United States from 2008 to 2011. Patients were included in the secondary analysis if they were delivered for pregnancy associated hypertension (gestational hypertension or preeclampsia) between 23 RESULTS: A total of 471 patients met inclusion criteria, of whom 271 (58%) underwent induction of labor and 200 (42%) underwent prelabor cesarean delivery. Composite maternal morbidity was 10.2% in the induction group and 21.1% in the cesarean delivery group (unadjusted odds ratio, 0.42 [0.25-0.72]; adjusted odds ratio, 0.44 [0.26-0.76]). Neonatal morbidity in the induction group vs the cesarean delivery was 51.9% and 63.8 %, respectively (unadjusted odds ratio, 0.61 [0.42-0.89]; adjusted odds ratio, 0.71 [0.48-1.06]). The frequency of vaginal delivery in the induction group was 53% (95% confidence interval, 46.8-58.7) and the median duration of labor was 13.9 hours (interquartile range, 8.7-22.2). The frequency of vaginal birth was higher in patients at or beyond 29 weeks (39.9% at 24 CONCLUSION: Among patients delivered for hypertensive disorders of pregnancy \u3c3

Predictive Performance of Newborn Small for Gestational Age by a US Intrauterine versus Birth Weight-derived Standard for Short-Term Neonatal Morbidity and Mortality

Objective: The use of birth weight standards to define small for gestational age (SGA) may fail to identify neonates affected by poor fetal growth since they include births associated with sub-optimal fetal growth. Our objective was to compare intrauterine versus birth weight-derived standards to define newborn SGA to predict neonatal morbidity and mortality. Study Design: This was a secondary analysis of a multi-center observational study of 118,422 births. Liveborn singleton, non-anomalous newborns born at 23–41 weeks were included. Those with missing gestational age estimation or without a 1st or 2nd trimester ultrasound to confirm dating, birth weight, or neonatal outcome data were excluded. Birth weight percentile was computed using an intrauterine standard (Hadlock, Radiology 1991) and a birth weight-derived standard (Olsen, Pediatrics 2010). We compared the test characteristics of SGA (birth weight \u3c10th percentile) by each standard to predict a composite neonatal morbidity and mortality outcome (death prior to discharge, NICU admission \u3e48 hours, respiratory distress syndrome, sepsis, necrotizing enterocolitis, grade III or IV intraventricular hemorrhage, or seizures). Severe composite morbidity was analyzed as a secondary outcome and was defined as death, NICU admission \u3e7 days, NEC, grade 3–4 IVH, or seizures. The areas under the curve (AUC) using receiver-operating characteristic methodology and proportions of the primary outcome by SGA status were compared by gestational age category at birth (\u3c34, 340–366, ≥37 weeks). Results: Of 115,502 mother-newborn dyads in the parent study, 78,203 (67.7%) were included, with the majority of exclusions occurring because of missing or inadequate dating information, multiple gestations, or delivery outside the gestational age range. The primary composite outcome occurred in 9.5% (95% CI 9.3–9.7) and the severe composite outcome occurred in 5.3% (5.1–5.4). SGA was diagnosed by intrauterine and birth weight-derived standards in 14.8% and 7.4%, respectively (p\u3c0.001). Neonates considered SGA only by the intrauterine standard experienced the primary outcome more than twice as often as those considered non-SGA by both standards (18.4% vs 7.9%, p\u3c0.001). For prediction of the primary outcome, SGA by the intrauterine standard had higher sensitivity (29% vs 15%, p\u3c0.001) but lower specificity (87% vs 93%, p\u3c0.001) than by the birth weight standard. Both standards had weak performance overall, though the intrauterine standard had a higher AUC (0.58 vs 0.53, p\u3c0.001). When sub-analyzed by gestational age at birth, the difference in AUCs was only present among preterm deliveries 34 to 36 competed weeks. Neither standard demonstrated any discrimination for morbidity prediction among term births (AUC = 0.50 for both). When prediction of severe morbidity was compared, the intrauterine still had better overall prediction than the birth weight standard (AUC 0.65 vs 0.57, p\u3c0.001), though this also varied by gestational age at birth. Conclusion: Among non-anomalous neonates, neither intrauterine nor birth weight-derived standards for SGA accurately predicted neonatal morbidity and mortality, with no discriminatory ability at term. SGA intrauterine standards performed better than birth weight standards

The Association of Race and Ethnicity with Severe Maternal Morbidity among Individuals Diagnosed with Hypertensive Disorders of Pregnancy

Objective: To examine whether there are racial disparities in severe maternal morbidity in patients with hypertensive disorder of pregnancy (HDP). Study Design: Secondary analysis of an observational study of 115,502 patients who had a live birth at ≥ 20 weeks in 25 hospitals in the US from 2008 to 2011. Only patients with HDP were included in this analysis. Race and ethnicity were categorized as non-Hispanic White, non-Hispanic Black and Hispanic and were abstracted from the medical charts. Patients of other races and ethnicities were excluded. Associations were estimated between race and ethnicity and the primary outcome of severe maternal morbidity, defined as any of the following: blood transfusion ≥4 units, unexpected surgical procedure, need for a ventilator ≥ 12 hours, intensive care unit (ICU) admission, or failure of ≥ 1 organ system, were estimated by unadjusted logistic and multivariable backward logistic regressions. Multivariable models were run classifying HDP into 3 levels: 1) gestational hypertension; 2) preeclampsia (mild, severe or superimposed); and 3) eclampsia or HELLP. Results: A total of 9,612 individuals with HDP met inclusion criteria. No maternal deaths occurred in this cohort. In univariable analysis, non-Hispanic White patients were more likely to present with gestational hypertension whereas non-Hispanic Black and Hispanic patients were more likely to present with preeclampsia. The frequency of the primary outcome, composite severe maternal morbidity, was higher in NHB patients compared with that in non-Hispanic White or Hispanic patients (11.8% vs. 4.5% in non-Hispanic White and 4.8% in Hispanic, p\u3c0.001). This difference was driven by a higher frequency of blood transfusions and ICU admissions among non-Hispanic Black individuals. Prior to adjusting the analysis for confounding factors, the odds ratio (OR) of primary composite outcomes in non-Hispanic black individuals was 2.85 (95% CI 2.38, 3.42) compared to non-Hispanic white. After adjusting for sociodemographic and clinical factors, hospital site, and the severity of HDP, the odds ratios of composite severe maternal morbidity did not differ between the groups (adjusted OR 1.26, 95% CI 0.95, 1.67 for non-Hispanic Black and adjusted OR 1.29, 95% CI 0.94, 1.77 for Hispanic, compared to non-Hispanic White patients). Sensitivity analysis was done to exclude one single site that was an outliner with the highest ICU admissions and demonstrated no difference in ICU admission by maternal race and ethnicity. Conclusions: Non-Hispanic Black patients with HDP had higher rates of the composite severe maternal morbidity compared with non-Hispanic White patients, driven mainly by a higher frequency of blood transfusions and ICU admissions. However, once severity and other confounding factors were taken into account, the differences did not persist

Short-Term Neonatal Outcomes of Pregnancies Complicated by Maternal Obesity

BACKGROUND: Maternal obesity complicates a high number of pregnancies. The degree to which neonatal outcomes are adversely affected is unclear. OBJECTIVE: This study aimed to evaluate neonatal outcomes of pregnancies complicated by maternal obesity. STUDY DESIGN: This study was a secondary analysis of a cohort of deliveries occurring on randomly selected days at 25 hospitals from 2008 to 2011. Data were collected by certified abstractors. This analysis included singleton deliveries between 24 and 42 weeks of gestation. Body mass index was calculated on the basis of maternal height and most recent weight before delivery. Normal and overweight (reference group; body mass index, 18.5-29.9 kg/m RESULTS: Overall, 52,162 patients and their neonates were included after propensity score matching. Of these, 21,704 (41.6%) were obese, 3787 (7.3%) were morbidly obese, and 590 (1.1%) were super morbidly obese. A total of 2103 neonates (4.0%) had the composite outcome. Neonates born to pregnant people with morbid obesity had a 33% increased risk of composite neonatal morbidity compared with those in the reference group (adjusted odds ratio, 1.33; 95% confidence interval, 1.17-1.52), but no significant association was observed for persons with obesity (adjusted odds ratio, 1.05; 95% confidence interval, 0.97-1.14) or with super morbid obesity (adjusted odds ratio, 1.18; 95% confidence interval, 0.86-1.64). CONCLUSION: Compared with the reference group, gravidas with morbid obesity were at higher risk of composite neonatal morbidity

Global, regional, and national burden of chronic kidney disease, 1990–2017 : a systematic analysis for the Global Burden of Disease Study 2017

Background Health system planning requires careful assessment of chronic kidney disease (CKD) epidemiology, but data for morbidity and mortality of this disease are scarce or non-existent in many countries. We estimated the global, regional, and national burden of CKD, as well as the burden of cardiovascular disease and gout attributable to impaired kidney function, for the Global Burden of Diseases, Injuries, and Risk Factors Study 2017. We use the term CKD to refer to the morbidity and mortality that can be directly attributed to all stages of CKD, and we use the term impaired kidney function to refer to the additional risk of CKD from cardiovascular disease and gout. Methods The main data sources we used were published literature, vital registration systems, end-stage kidney disease registries, and household surveys. Estimates of CKD burden were produced using a Cause of Death Ensemble model and a Bayesian meta-regression analytical tool, and included incidence, prevalence, years lived with disability, mortality, years of life lost, and disability-adjusted life-years (DALYs). A comparative risk assessment approach was used to estimate the proportion of cardiovascular diseases and gout burden attributable to impaired kidney function. Findings Globally, in 2017, 1·2 million (95% uncertainty interval [UI] 1·2 to 1·3) people died from CKD. The global all-age mortality rate from CKD increased 41·5% (95% UI 35·2 to 46·5) between 1990 and 2017, although there was no significant change in the age-standardised mortality rate (2·8%, −1·5 to 6·3). In 2017, 697·5 million (95% UI 649·2 to 752·0) cases of all-stage CKD were recorded, for a global prevalence of 9·1% (8·5 to 9·8). The global all-age prevalence of CKD increased 29·3% (95% UI 26·4 to 32·6) since 1990, whereas the age-standardised prevalence remained stable (1·2%, −1·1 to 3·5). CKD resulted in 35·8 million (95% UI 33·7 to 38·0) DALYs in 2017, with diabetic nephropathy accounting for almost a third of DALYs. Most of the burden of CKD was concentrated in the three lowest quintiles of Socio-demographic Index (SDI). In several regions, particularly Oceania, sub-Saharan Africa, and Latin America, the burden of CKD was much higher than expected for the level of development, whereas the disease burden in western, eastern, and central sub-Saharan Africa, east Asia, south Asia, central and eastern Europe, Australasia, and western Europe was lower than expected. 1·4 million (95% UI 1·2 to 1·6) cardiovascular disease-related deaths and 25·3 million (22·2 to 28·9) cardiovascular disease DALYs were attributable to impaired kidney function. Interpretation Kidney disease has a major effect on global health, both as a direct cause of global morbidity and mortality and as an important risk factor for cardiovascular disease. CKD is largely preventable and treatable and deserves greater attention in global health policy decision making, particularly in locations with low and middle SDI

Preterm neonatal morbidity and mortality by gestational age: a contemporary cohort

Although preterm birth less than 37 weeks gestation is the leading cause of neonatal morbidity and mortality in the United States, the majority of data regarding preterm neonatal outcomes come from older studies, and many reports have been limited to only very preterm neonates. Delineation of neonatal outcomes by delivery gestational age is needed to further clarify the continuum of mortality and morbidity frequencies among preterm neonates

Serious maternal complications after early preterm delivery (24-33 weeks’ gestation)

To describe the prevalence of serious maternal complications following early preterm birth by gestational age (GA), delivery route and type of cesarean incision

Mapping geographical inequalities in access to drinking water and sanitation facilities in low-income and middle-income countries, 2000-17

Background: Universal access to safe drinking water and sanitation facilities is an essential human right, recognised in the Sustainable Development Goals as crucial for preventing disease and improving human wellbeing. Comprehensive, high-resolution estimates are important to inform progress towards achieving this goal. We aimed to produce high-resolution geospatial estimates of access to drinking water and sanitation facilities. Methods: We used a Bayesian geostatistical model and data from 600 sources across more than 88 low-income and middle-income countries (LMICs) to estimate access to drinking water and sanitation facilities on continuous continent-wide surfaces from 2000 to 2017, and aggregated results to policy-relevant administrative units. We estimated mutually exclusive and collectively exhaustive subcategories of facilities for drinking water (piped water on or off premises, other improved facilities, unimproved, and surface water) and sanitation facilities (septic or sewer sanitation, other improved, unimproved, and open defecation) with use of ordinal regression. We also estimated the number of diarrhoeal deaths in children younger than 5 years attributed to unsafe facilities and estimated deaths that were averted by increased access to safe facilities in 2017, and analysed geographical inequality in access within LMICs. Findings: Across LMICs, access to both piped water and improved water overall increased between 2000 and 2017, with progress varying spatially. For piped water, the safest water facility type, access increased from 40·0% (95% uncertainty interval [UI] 39·4–40·7) to 50·3% (50·0–50·5), but was lowest in sub-Saharan Africa, where access to piped water was mostly concentrated in urban centres. Access to both sewer or septic sanitation and improved sanitation overall also increased across all LMICs during the study period. For sewer or septic sanitation, access was 46·3% (95% UI 46·1–46·5) in 2017, compared with 28·7% (28·5–29·0) in 2000. Although some units improved access to the safest drinking water or sanitation facilities since 2000, a large absolute number of people continued to not have access in several units with high access to such facilities (>80%) in 2017. More than 253 000 people did not have access to sewer or septic sanitation facilities in the city of Harare, Zimbabwe, despite 88·6% (95% UI 87·2–89·7) access overall. Many units were able to transition from the least safe facilities in 2000 to safe facilities by 2017; for units in which populations primarily practised open defecation in 2000, 686 (95% UI 664–711) of the 1830 (1797–1863) units transitioned to the use of improved sanitation. Geographical disparities in access to improved water across units decreased in 76·1% (95% UI 71·6–80·7) of countries from 2000 to 2017, and in 53·9% (50·6–59·6) of countries for access to improved sanitation, but remained evident subnationally in most countries in 2017. Interpretation: Our estimates, combined with geospatial trends in diarrhoeal burden, identify where efforts to increase access to safe drinking water and sanitation facilities are most needed. By highlighting areas with successful approaches or in need of targeted interventions, our estimates can enable precision public health to effectively progress towards universal access to safe water and sanitation