Bridging the notch: quantification of the end diastolic notch to better predict fetal growth restriction.

PURPOSE: We aimed to evaluate several quantitative methods to describe the diastolic notch (DN) and compare their performance in the prediction of fetal growth restriction. MATERIALS AND METHODS: Patients who underwent a placental scan at 16-26 weeks of gestation and delivered between Jan 2016 and Dec 2020 were included. The uterine artery pulsatility index was measured for all of the patients. In patients with a DN, it was quantified using the notch index and notch depth index. Odds ratios for small for gestational age neonates (defined as birth weight <10th and <5th percentile) were calculated. Predictive values of uterine artery pulsatility, notch, and notch depth index for fetal growth restriction were calculated. RESULTS: Overall, 514 patients were included, with 69 (13.4%) of them delivering a small for gestational age neonate (birth weight<10th percentile). Of these, 20 (20.9%) had a mean uterine artery pulsatility index >95th percentile, 13 (18.8%) had a unilateral notch, and 11 (15.9%) had a bilateral notch. 16 patients (23.2%) had both a high uterine artery pulsatility index (>95th percentile) and a diastolic notch. Comparison of the performance between uterine artery pulsatility, notch, and notch depth index using receiver operating characteristic curves to predict fetal growth restriction <10th percentile found area under the curve values of 0.659, 0.679, and 0.704, respectively, with overlapping confidence intervals. CONCLUSION: Quantifying the diastolic notch at 16-26 weeks of gestation did not provide any added benefit in terms of prediction of neonatal birth weight below the 10th or 5th percentile for gestational age, compared with uterine artery pulsatility index.

Cytogenetic outcomes following a failed cell-free DNA screen: a population-based retrospective cohort study of 35,146 singleton pregnancies.

BACKGROUND: Cell-free fetal DNA screening is routinely offered to pregnant individuals to screen for aneuploidies. Although cell-free DNA screening is consistently more accurate than multiple-marker screening, it sometimes fails to yield a result. These test failures and their clinical implications are poorly described in the literature. Some studies suggest that a failed cell-free DNA screening result is associated with increased likelihood of cytogenetic abnormalities. OBJECTIVE: This study aimed to assess the association between a failed cell-free DNA test and common aneuploidies. The objectives were to determine: (1) the proportion of test failures on first and subsequent attempts, and (2) whether a failed cell-free DNA screen on first attempt is associated with increased likelihood of common aneuploidies (trisomies 21, 18, and 13, and sex chromosome aneuploidies). STUDY DESIGN: This was a population-based retrospective cohort study using data from Ontario's prescribed maternal and child registry, Better Outcomes Registry and Network Ontario. The study included all singleton pregnancies in Ontario with an estimated date of delivery from September 1, 2016 to March 31, 2019 that had a cell-free DNA screening record in the registry. Specific outcomes (trisomies 21, 18, and 13, and sex chromosome aneuploidies) of pregnancies with a failed cell-free DNA screen on first attempt were compared with those of pregnancies with low-risk cell-free DNA-screening results using modified Poisson regression adjusted for funding status (publicly funded vs self-paid), gestational age at screening, method of conception, and maternal age for autosomal aneuploidies. RESULTS: Our cohort included 35,146 pregnancies that had cell-free DNA screening during the study period. The overall cell-free DNA screening failure rate was 4.8% on first attempt and 2.2% after multiple attempts. An abnormal cytogenetic result for trisomies 21, 18, and 13, or sex chromosome aneuploidies was identified in 19.4% of pregnancies with a failed cell-free DNA screening for which cytogenetic testing was performed. Pregnancies with a failed cell-free DNA screen on first attempt had a relative risk of 130.3 (95% confidence interval, 64.7-262.6) for trisomy 21, trisomy 18, or trisomy 13, and a risk difference of 5.4% (95% confidence interval, 2.6-8.3), compared with pregnancies with a low-risk result. The risk of sex chromosome aneuploidies was not significantly greater in pregnancies with a failed result compared with pregnancies with a low-risk result (relative risk, 2.7; 95% confidence interval, 0.9-7.9; relative difference, 1.2%; 95% confidence interval, -0.9 to 3.2). CONCLUSION: Cell-free DNA screening test failures are relatively common. Although repeated testing improves the likelihood of an informative result, pregnancies with a failed cell-free DNA screen upon first attempt remain at increased risk for common autosomal aneuploidies, but not sex chromosome aneuploidies.

Identification of a novel TBX5 c.755 + 1 G > A variant and related pathogenesis in a family with Holt-Oram syndrome.

The proband with congenital heart disease and abnormal thumb was clinically diagnosed as Holt-Oram syndrome (HOS). A novel variant, T-box transcription factor 5 (TBX5) c.755 + 1 G > A, was identified in the proband via whole exome sequencing and validated using Sanger sequencing. Pedigree analysis and clinical examinations revealed three/seven individuals over three generations within the family, with features suggestive of HOS. Deep amplicon sequencing confirmed that the allele frequencies of the novel variant in the proband (III-1), her brother (III-2), and her mother (II-2) were 50%, 48.3%, and 38.1%, respectively, indicating that III-1 and III-2 harbored heterozygous variants, while II-2 harbored mosaic heterozygous variants. The minigene splicing assay showed that the novel variant affected the normal splicing of exon 7, resulting in the production of abnormal TBX5 transcripts. Reverse transcription-quantitative polymerase chain reaction and western blot analyses revealed that the novel variant upregulated TBX5 expression at the transcriptional and translational levels. Nuclear localization assay demonstrated impaired nuclear localization of the mutant TBX5. Cell viability assay revealed the inhibition of cell activity by the mutant TBX5. Our findings indicate that the novel variant was potentially induced HOS, probably by causing aberrant splicing, reducing the enrichment of nuclear TBX5 protein, and inhibiting cellular proliferation.

Modified multiple marker aneuploidy screening as a primary screening test for preeclampsia.

BACKGROUND: Abnormal levels of maternal biochemical markers used in multiple marker aneuploidy screening have been associated with adverse pregnancy outcomes. This study aims to assess if a combination of maternal characteristics and biochemical markers in the first and second trimesters can be used to screen for preeclampsia (PE). The secondary aim was to assess this combination in identifying pregnancies at risk for gestational hypertension and preterm birth. METHODS: This case-control study used information on maternal characteristics and residual blood samples from pregnant women who have undergone multiple marker aneuploidy screening. The median multiple of the median (MoM) of first and second trimester biochemical markers in cases (women with PE, gestational hypertension and preterm birth) and controls were compared. Biochemical markers included pregnancy-associated plasma protein A (PAPP-A), placental growth factor (PlGF), human chorionic gonadotropin (hCG), alpha feto-protein (AFP), unconjugated estriol (uE3) and Inhibin A. Logistic regression analysis was used to estimate screening performance using different marker combinations. Screening performance was defined as detection rate (DR) and false positive rate (FPR). Preterm and early-onset preeclampsia PE were defined as women with PE who delivered at < 37 and < 34 weeks of gestation, respectively. RESULTS: There were 147 pregnancies with PE (81 term, 49 preterm and 17 early-onset), 295 with gestational hypertension, and 166 preterm birth. Compared to controls, PE cases had significantly lower median MoM of PAPP-A (0.77 vs 1.10, p < 0.0001), PlGF (0.76 vs 1.01, p < 0.0001) and free-ß hCG (0.81 vs. 0.98, p < 0.001) in the first trimester along with PAPP-A (0.82 vs 0.99, p < 0.01) and PlGF (0.75 vs 1.02, p < 0.0001) in the second trimester. The lowest first trimester PAPP-A, PlGF and free ß-hCG were seen in those with preterm and early-onset PE. At a 20% FPR, 67% of preterm and 76% of early-onset PE cases can be predicted using a combination of maternal characteristics with PAPP-A and PlGF in the first trimester. The corresponding DR was 58% for gestational hypertension and 36% for preterm birth cases. CONCLUSIONS: Maternal characteristics with first trimester PAPP-A and PlGF measured for aneuploidy screening provided reasonable accuracy in identifying women at risk of developing early onset PE, allowing triage of high-risk women for further investigation and risk-reducing therapy. This combination was less accurate in predicting women who have gestational hypertension or preterm birth.

Performance of a universal prenatal screening program incorporating cell-free fetal DNA analysis in Ontario, Canada.

BACKGROUND: The emergence of cell-free fetal DNA (cfDNA) testing technology has disrupted the landscape of prenatal screening for trisomies 21 (T21) and 18 (T18). Publicly funded systems around the world are grappling with how to best integrate this more accurate but costly technology, as there is limited evidence about its incremental value in real-world conditions. The objectives of this study were to describe the population-based performance of Ontario's prenatal screening program, which incorporates publicly funded cfDNA screening for specific indications, and the effect of cfDNA testing on the screening and diagnostic choices made by pregnant people. METHODS: We conducted a retrospective, descriptive cohort study using routinely collected data from Better Outcomes & Registry Network (BORN) Ontario, which captures linked population data for prenatal and neonatal health encounters across Ontario. We included all singleton pregnancies with an estimated due date between Sept. 1, 2016, and Mar. 31, 2019, that underwent publicly funded prenatal screening in Ontario, and a comparison cohort from Apr. 1, 2012, and Mar. 31, 2013. We assessed performance of the screening program for the detection of T21 or T18 by calculating sensitivity, specificity, positive predictive value and negative predictive value against diagnostic cytogenetic results or birth outcomes. We assessed the impact of the program by calculating the proportion of T21 screen-positive pregnancies undergoing subsequent cfDNA screening and invasive prenatal diagnostic testing. RESULTS: The study cohort included 373 682 pregnancies. The prenatal screening program had an uptake of 69.9%, a screen-positive rate and sensitivity of 1.6% and 89.9% for T21, and 0.2% and 80.5% for T18, respectively. The test failure rate for cfDNA screening was 2.2%. Invasive prenatal diagnostic testing decreased from 4.4% in 2012-2013 to 2.4% over the study period; 65.2% of pregnant people who received a screen-positive result from cfDNA testing went on to have invasive prenatal diagnostic testing. INTERPRETATION: This publicly funded screening program, incorporating cfDNA analysis for common aneuploidies, showed robust performance, a substantial reduction in invasive prenatal diagnostic testing and that pregnant people exercise autonomy in their choices about prenatal screening and diagnosis.

Maternal Ethnicity and the Risk of Obstetrical Anal Sphincter Injury: A Retrospective Cohort Study.

OBJECTIVE: To explore the role of maternal ethnicity as a risk factor for obstetrical anal sphincter injury (OASI). METHODS: A retrospective cohort study of all women with singleton gestations who had a vaginal delivery at term, between January 2014 and October 2017, at a single center. OASI was defined as a third-degree perineal tear (anal sphincter complex) or a fourth-degree perineal tear (anorectal mucosa). The characteristics of women with and without OASIs were compared. Multiple logistic regression was performed to account for potential confounders, including ethnicity. RESULTS: During the study period, 11 012 women were eligible for inclusion, of whom 336 (3.1%) had an OASI; 313 (93.1%) had a third-degree tear, and 23 (6.9%) had a fourth-degree tear. Women with OASIs were characterized by younger maternal age (<35 years), Asian ethnicity, nulliparity, neonatal birth weight ≥3500 grams, midline and mediolateral episiotomy, second stage of labour lasting ≥60 minutes, and assisted vaginal delivery. After adjusting for potential confounders, Asian ethnicity remained independently associated with increased risk of OASI (adjusted odds ratio 2.07; 95% CI 1.6-2.7) whereas mediolateral episiotomy was independently associated with decreased risk of OASI (adjusted odds ratio 0.64; 95% CI 0.5-0.9). CONCLUSION: Asian ethnicity is independently associated with increased risk of OASI. Although midline episiotomy increases the risk of OASI, mediolateral episiotomy may protect against OASI, and should be considered in high-risk patients.

Risk of Severe Maternal Morbidity or Death in Relation to Prenatal Biochemical Screening: Population-Based Cohort Study.

OBJECTIVE: This study aimed to examine whether prenatal biochemical screening analytes are associated with an increased risk of severe maternal morbidity (SMM) or maternal mortality. STUDY DESIGN: This population-based cohort study includes all women in Ontario, Canada, who underwent prenatal screening from 2001 to 2011. Increasing fifth percentiles of the multiple of the median (MoM) for alphafetoprotein (AFP), total human chorionic gonadotropin, unconjugated estriol (uE3), dimeric inhibin-A (DIA), and pregnancy-associated plasma protein A were evaluated. An abnormally high concentration (>95th percentile MoM) for each analyte, individually and combined, was also evaluated. The main outcome assessed was the adjusted relative risk (aRR) of SMM or maternal mortality from 20 weeks' gestation up to 26 weeks thereafter. RESULTS: Among 748,972 pregnancies, 11,177 resulted in SMM or maternal mortality (1.5%). Except for uE3, the aRR of SMM or maternal mortality increased in association with increasing fifth percentiles of the MoM for all analytes. AFP (aRR: 2.10; 95% confidence interval [CI]: 1.97-2.25) and DIA (aRR: 2.33; 95% CI: 1.98-2.74) > 95th versus ≤ 5th percentile of the MoM were especially associated with SMM or death. CONCLUSION: Women with abnormally high concentrations of certain prenatal biochemical analytes may be at a higher risk of SMM or death in pregnancy or postpartum.

Racial differences in contribution of prepregnancy obesity and excessive gestational weight gain to large-for-gestational-age neonates.

OBJECTIVES: To examine the racial differences in the population attributable fraction (PAF) of prepregnancy obesity and excessive gestational weight gain to large-for-gestational-age (LGA) neonates. METHODS: We conducted a population-based retrospective cohort study among all women who had prenatal screening and had a singleton live birth in a hospital (1 April 2016-31 March 2017) using data from Ontario birth registry in Canada. We used multivariable log-binomial regression models to estimate the PAF and 95% confidence interval (CI) of LGA neonates due to prepregnancy obesity and excessive gestational weight gain. All models were stratified by race (White, Asian, and Black). RESULTS: Of the 74,402 eligible women, the prevalence of prepregnancy obesity, excessive gestational weight gain, and LGA neonate was 21.1%, 60.0%, and 11.3%, respectively, for Whites; 9.3%, 45.9%, and 5.4%, respectively, for Asians; and 28.6%, 52.4%, and 7.9%, respectively, for Blacks. The association of prepregnancy obesity was greater than that of excessive gestational weight gain on LGA for all racial groups. Excessive gestational weight gain contributed more than prepregnancy obesity in Whites (PAF 32.9%, 95% CI [30.3-35.5%] and 16.6%, 95% CI [15.3-17.9%], respectively, for excessive gestational weight gain and prepregnancy obesity) and in Asians (PAF 32.1%, 95% CI [27.2-36.7%] and 11.8%, 95% CI [9.5-14.1%], respectively, for excessive gestational weight gain and prepregnancy obesity). Prepregnancy obesity (PAF 22.8%, 95% CI [17.1-28.1%]) and excessive gestational weight gain (PAF 20.1%, 95% CI [4.7-33.0%]) contributed to LGA neonates almost the same in Blacks. CONCLUSIONS: Excessive gestational weight gain contributed more to LGA neonates than prepregnancy obesity in Whites and Asians, while there was no difference between excessive gestational weight gain and prepregnancy obesity in their contributions to the LGA neonates in Blacks. The differences are mostly driven by the differential prevalence of the two risk factors across racial groups.

A 2-year review of publicly funded cell-free DNA screening in Ontario: utilization and adherence to funding criteria.

OBJECTIVE: Ontario offers a publicly funded modified contingent model of prenatal screening for aneuploidy in which cell-free DNA (cfDNA) screening is covered for pregnancies at higher risk of fetal aneuploidy. The objective of this study was to review utilization of provincially funded cfDNA screening and adherence to the criteria laid out in Ontario prenatal screening guidelines. METHODS: This was a descriptive cohort study using data collected by Ontario's prescribed maternal and child registry. The study population included all pregnant individuals who received cfDNA screening from January 2016 to December 2017. RESULTS: The most common criteria for provincially funded cfDNA screening were advanced maternal age ≥40 years (37.7%), positive multiple marker screen (34.1%), modifying risk factors such as ultrasound soft markers (7.1%), and previous aneuploidy (5.5%). The audit demonstrated that 2.9% of funded cfDNA screens tests did not meet funding criteria, and that 11.4% of self-paid cfDNA screens could have been publicly funded. CONCLUSION: Reviewing and auditing the application of criteria for funded cfDNA screening using prescribed registry data allows an opportunity to identify areas where targeted education may improve adherence to standardized screening protocols, and provides a basis for reassessment of the funding model.

Prenatal screening for trisomy 21: a comparative performance and cost analysis of different screening strategies.

BACKGROUND: Prenatal screening for chromosome aneuploidies have constantly been evolving, especially with the introduction of cell-free fetal DNA (cfDNA) screening in the most recent years. This study compares the performance, costs and timing of test results of three cfDNA screening implementation strategies: contingent, reflex and primary. METHODS: We modelled enhanced first trimester screening (eFTS) as the first-tier test in contingent or reflex strategies. cfDNA test was performed contingent on or reflex from eFTS results. A comparison was made between cfDNA screening using sequencing technology and Rolling Circle Amplification (RCA)/imaging solution. All model assumptions were based on results from previous publications or information from the Ontario prenatal screening population. RESULTS: At an eFTS risk cut-off of ≥1/1000, contingent and reflex cfDNA screening have the same detection rate (DR) (94%) for trisomy 21. Reflex cfDNA screening using RCA/Imaging solution provided the lowest false positive rate and cost. The number of women requiring genetic counselling and diagnostic testing was significantly reduced and women received their cfDNA screening result 9 days sooner compared with the contingent model. While primary cfDNA screening improved the trisomy 21 DR by 3-5%, it was more costly and more women required diagnostic testing. CONCLUSION: Reflex cfDNA screening is the most cost-effective prenatal screening strategy. It can improve the efficiency of prenatal aneuploidy screening by reducing the number of patient visits and providing more timely results.

Noninvasive Prenatal Testing for Trisomies 21, 18, and 13, Sex Chromosome Aneuploidies, and Microdeletions in Average-Risk Pregnancies: A Cost-Effectiveness Analysis.

OBJECTIVE: The cost effectiveness of noninvasive prenatal testing (NIPT) has been established for high-risk pregnancies but remains unclear for pregnancies at other risk levels. The aim was to assess the cost effectiveness of NIPT in average-risk pregnancies from the perspective of a provincial public payer in Canada. METHODS: A model was developed to compare traditional prenatal screening (TPS), NIPT as a second-tier test (performed only after a positive TPS result), and NIPT as a first-tier test (performed instead of TPS) for trisomies 21, 18, and 13; sex chromosome aneuploidies; and microdeletions in a hypothetical annual population cohort of average-risk pregnancies (142 000 to 148,000) in Ontario, Canada. A probabilistic analysis was conducted with 5000 repetitions. RESULTS: Compared with TPS, NIPT as a second-tier test detected more affected fetuses with trisomies 21, 18, and 13 (188 vs. 158), substantially reduced the number of diagnostic tests (i.e., chorionic villus sampling and amniocentesis) performed (660 vs. 3107), and reduced the cost of prenatal screening ($26.7 million vs. $27.6 million) annually. Compared with second-tier NIPT, first-tier NIPT detected an additional 80 cases of trisomies 21, 18, and 13 at an additional cost of $33 million. The incremental cost per additional affected fetus detected was $412 411. Extending first-tier NIPT to include testing for sex chromosome aneuploidies and 22q11.2 deletion would increase the total screening cost. CONCLUSIONS: NIPT as a second-tier test is cost-saving compared with TPS alone. Compared with second-tier NIPT, first-tier NIPT detects more cases of chromosomal anomalies but at a substantially higher cost.

Hormone Levels in Pregnancy and Subsequent Risk of Maternal Breast and Ovarian Cancer: A Systematic Review.

OBJECTIVE: Some maternal hormone levels in pregnancy are associated with a higher risk of breast and ovarian cancer. This study systematically assessed the association between blood hormone levels measured in pregnancy and future risk of these cancers. METHODS: Two reviewers independently conducted a literature search of MEDLINE and EMBASE databases from January 1970 to August 2017. Studies were included that measured one or more serum hormone levels in pregnancy and later assessed for cancer. Cancer outcomes were considered by cancer type, each in relation to a specific maternal hormone. RESULTS: Eleven studies were included, comprising a total of 57 967 women. The interval between pregnancy and cancer onset varied from 4.1 to 20.5 years. Elevated serum chorionic gonadotropin (two of four studies) and alpha fetoprotein (two of three studies) were each associated with a lower risk of maternal breast cancer, whereas elevated estrone levels suggested a higher risk (one of three studies). Elevated testosterone (one of one study) and androstenedione (one of one study) were each associated with a significantly greater risk of sex-cord stromal ovarian tumours. Higher serum 17-hydroxyprogesterone was associated with an increased risk of sex-cord stromal (one of one study) and epithelial (one of one study) ovarian cancer. CONCLUSION: Observational studies suggest some degree of association between serum hormones measured in pregnancy and a woman's future risk of breast and ovarian cancer. More data are needed to determine sufficiently whether certain blood hormone levels measured in pregnancy are predictive of future cancer risk.

Enhanced First Trimester Aneuploidy Screening with Placental Growth Factor and Alpha Feto-Protein: Detection of Trisomies 18 and 13.

OBJECTIVES: To assess the performance of first trimester combined screening (FTS) when enhanced with placental growth factor and alpha feto-protein in the detection of trisomies 18 and 13. METHODS: A retrospective case-control study. Marker parameters were derived using frozen serum samples. Multivariate Gaussian modelling predicted the detection rate (DR) and false-positive rate (FPR) for trisomies 18 and 13 with FTS and enhanced first trimester screening (eFTS) using the risk of trisomy 21 alone and an additional risk cut-off for trisomy 18, or trisomies 18 or 13. RESULTS: There were 83 trisomy 18, 22 trisomy 13, and 588 controls. The median placental growth factor levels in trisomies 18 and 13 were 0.75 and 0.65 multiple of the median of controls, respectively (both P < 0.0001). There were no statistically significant differences in alpha feto-protein levels. Modelling predicts that using a trisomy 21 risk cut-off alone, at FPR of 3%, eFTS increases the DR for trisomies 18 and 13 by 0.6-0.8% compared with FTS. Additionally using a trisomy 18 risk cut-off, at an extra FPR of 0.2%, eFTS increased the DR by 0.6-0.9% over FTS; using a trisomy 18 or 13 risk cut-off did not further increase detection for FTS or eFTS. The increase in DR was greater at higher FPR. CONCLUSION: eFTS increases the detection of trisomies 18 and 13 to a small extent.

Comparing Maternal Serum Screening Markers Among IVF and Spontaneous Conceptions in Ontario Through Registry Data.

OBJECTIVE: The objectives of this study were as follows: (1) to investigate the accuracy of IVF identification on the prenatal screening record from prenatal screening laboratories; (2) to compare the screening markers in IVF and non-IVF pregnancies in the population of Ontario; and (3) to propose more appropriate IVF adjustment factors for the Ontario population. METHODS: Two years of IVF treatment, data from all fertility clinics in Ontario were merged with the corresponding prenatal screening data from all five prenatal screening labs. New adjustment factors for IVF were developed for each maternal serum screening marker and nuchal translucency measurement. Means and SDs and linear regression models were reported for all prenatal screening records, as well as for records that had IVF identified through the prenatal screening requisition and records that were identified through the Canadian Assisted Reproductive Technologies Register (CARTR) Plus database. RESULTS: Significant differences between IVF and non-IVF groups on the basis of the prenatal screening requisition information and CARTR Plus information were found among the ethnicity-adjusted mean multiple of the medians for alpha fetoprotein, first trimester pregnancy-associated plasma protein A, second trimester unconjugated estradiol, first trimester human chorionic gonadotropin, total human chorionic gonadotropin, and dimeric inhibin A. CONCLUSION: This study proposed alternate IVF adjustment factors that will produce more accurate screening results within the population of Ontario.

Human umbilical cord Wharton jelly cells promote extra-pancreatic insulin formation and repair of renal damage in STZ-induced diabetic mice.

BACKGROUND: We evaluated the therapeutic effect and fate of high doses of human umbilical cord Wharton jelly cells (hUCWJCs) after IP administration to streptozotocin (STZ)-induced diabetic mice. METHODS: Type 1 diabetes (T1D) was induced in Kunming mice via IP injection of STZ. hUCWJCs were labeled with 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI). Diabetic animals with sustained hyperglycemia for at least 2 weeks were administered 1 × 107 Dil-hUCWJCs via intraperitoneal injection. Insulin, glucagon and PDX-1 were detected by immunofluorescence with confocal microscopy. Serum mouse and human C-peptide was assayed in blood collected via intracardiac puncture. Specific ß-cell differentiation markers and human DNA were assessed using qPCR performed with 200 ng of target DNA. RESULTS: hUCWJCs migrated to the STZ-damaged organs and contributed to lower blood glucose levels in 30% of the treated mice. Confocal microscopy revealed the presence of resident insulin-positive cells in the liver and kidneys. hUCWJC-treated mice with restored hyperglycemia also showed increased serum mouse C-peptide levels. The qPCR results, particularly in the liver, revealed that after transplantation hUCWJCs upregulated genes of endocrine precursors but failed to express endocrine stage markers. Mice with restored hyperglycemia had reduced urinary volume and lacked glomerular hypertrophy, exhibiting a morphology resembling that of normal glomeruli. Moreover, we also verified that one of the possible mechanisms by which hUCWJCs exert immunosuppressive effects is through down-regulation of the cell surface receptor HLA-1. CONCLUSIONS: We confirmed the potential of IP administration of hUCWJCs and the capability of these cells to migrate to damaged tissues and promote insulin secretion from non-pancreatic local cells and to improve renal damage. These findings confer unique therapeutic properties to hUCWJCs, suggesting a promising future in the treatment of diabetes mellitus.

Enhanced First Trimester Screening for Trisomy 21 with Contingent Cell-Free Fetal DNA: A Comparative Performance and Cost Analysis.

OBJECTIVE: Prenatal screening for trisomy 21 is a standard of care. Emerging cell-free fetal DNA (cffDNA) technologies can improve screening performance, but they are expensive. This study was conducted to propose a contingent screening model that would incorporate cffDNA technology, would remain affordable, and could be applied equitably in a publically funded system. METHODS: Using performance and cost parameters from published literature, four prenatal screening strategies were compared. Scenario 1 modelled integrated prenatal screening (first trimester nuchal translucency and biochemical markers from both the first and second trimesters) with no cffDNA. Scenarios 2 and 3 modelled first trimester combined screening (FTS) and "enhanced FTS" (adding serum placental growth factor and alpha fetoprotein to FTS), respectively, with contingent cffDNA following a positive result. Scenario 4 modelled cffDNA as the primary screening test. RESULTS: Scenario 1 provides a known detection rate (DR) of 88%, with a false positive rate (FPR) of 3.3%. Scenarios 2 and 3 result in a DR of 94% and overall FPR of 0.59% and 0.33%, respectively, comparable to the DR of 96% and FPR of 0.1% with primary cffDNA (assuming the published test failure rate of 3%). The total cost, cost per woman screened, and cost per case of trisomy 21 detected were lower with scenario 3 (enhanced FTS with contingent cffDNA) compared with primary cffDNA or scenario 2 (FTS with contingent cffDNA). CONCLUSION: Enhanced FTS with contingent cffDNA following a positive result provides a similar performance to that of primary cffDNA at a substantially lower cost.

Research on the Vertical Transmission of Hepatitis C Gene from Father-to-child via Human Sperm.

BACKGROUND: The aim was to develop a better experimental model which could facilitate further studies assessing the vertical HCV gene transmission via human spermatozoa, and verify the possibility of father-to-child transmission of the HCV gene. METHODS: The recombinant plasmid pIRES2-EGFP-HCV C was constructed. Fluorescence in situ hybridization was performed to detect the integration of the HCV C gene in human sperm genome and in zygote's pronucleus. RESULTS: Successful construction of recombinant plasmid pIRES2-EGFP-HCV C was confirmed by restriction mapping, PCR, and sequencing. Positive HCV C DNA signals were observed in sperm heads, human sperm chromosomes and two-cell embryos in transfected samples. No positive signal was found in normal control and HCV infected groups. CONCLUSIONS: The recombinant plasmid pIRES2-EGFP-HCV C was successfully constructed. The HCV C gene was able to pass through the sperm membrane and integrate into the sperm genome. Human sperm carrying the HCV C gene was able to achieve normal fertilization. The replication of the sperm-mediated HCV C gene was synchronized with that of the host genome. Our results provide direct evidence for vertical transmission of the HCV C gene from father-to-child via human sperm.

Effects of Hepatitis C Virus Infection on Human Sperm Chromosomes.

BACKGROUND: To determine the degree of chromosomal aberrations in the sperm of men with hepatitis C. METHODS: 36 subjects (20 in the healthy control group and 16 in the HCV infection group [genotype 1b]) were recruited. The cause of viral transmission was unknown in all patients. Sperm samples from the subjects were used for interspecies in vitro fertilization of zona-free golden hamster ova. The frequencies of spermatozoan aberrations were compared between the healthy control group and the HCV infection group. RESULTS: A total of 280 sperm chromosome complements were studied, including 129 complements from the 16 donors in the HCV infection group and 151 from the healthy control group. Of the 129 analyzable sperm metaphase spreads in the HCV infection group, 14 (10.85%) complements contained chromosomal aberrations, which was significantly higher than the number (9/151, 5.96%) in the healthy control group (p < 0.01). Moreover, in the HCV infection group, chromosomes frequently showed anomalies such as stickiness, clumping, and failure to stain, which prevented their analysis. CONCLUSIONS: HCV infection has mutagenic effects on the chromosomes in sperm and may lead to extensive heredi-tary effects owing to genetic alterations and/or chromosomal aberrations. In addition, there is the possibility of vertical transmission of HCV via the germ line.

First and second trimester maternal serum markers in pregnancies with a vanishing twin.

OBJECTIVE: The aim of this study was to assess the concentration of the first and second trimester maternal serum markers in pregnancies with a vanishing twin. METHODS: This is a retrospective case-control study of pregnancies screened for Down syndrome in one Ontario center. Singleton pregnancies with ultrasound evidence of a vanishing twin were identified, and each was matched with five normal singleton controls for ethnicity, maternal age, gestational age, and blood sampling date. The median MoM of the first and second trimester serum markers was compared between cases and controls. The differences were assessed using the Mann-Whitney U-test. RESULTS: The study included 174 pregnancies that had a vanishing twin. Compared with control pregnancies, pregnancy associated plasma protein A increased by 21% (p = 0.0026), alpha-fetoprotein (AFP) increased by 10% (p < 0.0001), and dimeric inhibin A (DIA) increased by 13% (p = 0.0470) in pregnancies with a vanishing twin. Unconjugated oestriol and total human chorionic gonadotrophin were not significantly changed in these pregnancies. CONCLUSIONS: Pregnancy associated plasma protein A is not an adequate marker for pregnancies with a vanishing twin. The impact of elevated AFP on risk estimation is offset by that of DIA to certain extent. Further studies are needed to establish an adequate adjustment method for AFP and DIA to improve the accuracy of screening results for these pregnancies.

First trimester screening for Down syndrome using nuchal translucency, maternal serum pregnancy-associated plasma protein A, free-ß human chorionic gonadotrophin, placental growth factor, and α-fetoprotein.

OBJECTIVE: The aim of this study was to assess the screening performance for Down syndrome using first trimester combined screening (FTS) and two additional markers, serum placental growth factor (PlGF) and α-fetoprotein (AFP). METHODS: This is a retrospective case-control study of 137 pregnancies affected by Down syndrome and 684 individually matched unaffected pregnancies. Stored serum samples were tested for all four markers, and results were expressed as multiples of the gestation-specific median (MoM). Multivariate Gaussian modeling was used to calculate risks for different combinations of markers and to predict the detection rate (DR) and false positive rate (FPR). The predicted performance of enhanced FTS (FTS plus PlGF and AFP) was compared with FTS; the performance without nuchal translucency (first trimester quad) was assessed. RESULTS: For affected pregnancies, the median PlGF level was 0.622 MoM and median AFP 0.764 MoM. Adding PlGF and AFP improved the screening performance. At 3% FPR, DR increased by 4.4% from 83.8% to 88.2% using enhanced FTS; at 95% DR, FPR decreased by 8.3%, from 19.3% to 11.0%. At 3% FPR, DR using first trimester quad test was 76.4%. CONCLUSIONS: The performance of FTS can be enhanced by adding PlGF and AFP. Even without nuchal translucency, the test would perform well.