Timing of human preimplantation embryonic development is confounded by embryo origin.

STUDY QUESTION: To what extent do patient- and treatment-related factors explain the variation in morphokinetic parameters proposed as embryo viability markers? SUMMARY ANSWER: Up to 31% of the observed variation in timing of embryo development can be explained by embryo origin, but no single factor elicits a systematic influence. WHAT IS KNOWN ALREADY: Several studies report that culture conditions, patient characteristics and treatment influence timing of embryo development, which have promoted the perception that each clinic must develop individual models. Most of the studies have, however, treated embryos from one patient as independent observations, and only very few studies that evaluate the influence from patient- and treatment-related factors on timing of development or time-lapse parameters as predictors of viability have controlled for confounding, which implies a high risk of overestimating the statistical significance of potential correlations. STUDY DESIGN, SIZE, DURATION: Infertile patients were prospectively recruited to a cohort study at a hospital fertility clinic from February 2011 to May 2013. Patients aged <38 years without endometriosis were eligible if ≥8 oocytes were retrieved. Patients were included only once. All embryos were monitored for 6 days in a time-lapse incubator. PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 1507 embryos from 243 patients were included. The influence of fertilization method, BMI, maternal age, FSH dose and number of previous cycles on timing of t2-t5, duration of the 2- and 3-cell stage, and development of a blastocoel (tEB) and full blastocoel (tFB) was tested in multivariate, multilevel linear regression analysis. Predictive parameters for live birth were tested in a logistic regression analysis for 223 single transferred blastocysts, where time-lapse parameters were investigated along with patient and embryo characteristics. MAIN RESULTS AND THE ROLE OF CHANCE: Moderate intra-class correlation coefficients (0.16-0.31) were observed for all parameters except duration of the 3-cell stage, which demonstrates that embryos from one patient elicit clustering at a patient level. No single patient- and treatment-related factor was found to systematically influence the timing from cleavage to blastocyst stage, which indicates that no individual patient-related factor can be identified that separately explains the clustering throughout the entire developmental stages. The blastocyst parameters were more affected by patient-related factors than cleavage stage parameters, as tEB occurred significantly later with older age (0.29 h/year (95% confidence interval: CI 0.03; 0.56)), while both tEB and tFB occurred significantly later with increasing dose of FSH (tEB: 0.12 h/100 IU FSH (95% CI 0.01;0.24); tFB 0.14 h/100 IU FSH (95% CI 0.03;0.27)) and with more previous attempts (tEB: 1.2 h/attempt (95% CI 0.01;2.5); tFB 1.4 h/attempt (0.10;2.7)). Fertilization method affected timing of the first division, with ICSI embryos cleaving significantly faster than IVF embryos (-3.6% (95% CI -6.4; -0.77)), whereas no difference was found in the subsequent divisions. The univariable regression analysis identified female age, cumulative FSH dose, degree of blastocyst expansion, score of the inner cell mass and timing of full blastocyst formation as predictors of live birth. The timing of full blastocyst formation (tFB) did not remain significant when adjusting for age, number of previous cycles and cumulative FSH dose, which were the parameters shown to influence tFB in the mixed regression model. LIMITATIONS, REASONS FOR CAUTION: Only good prognosis patients were enrolled, so these results may not be generalized to all infertile women. Not all patient-related factors were investigated. WIDER IMPLICATIONS OF THE FINDINGS: Our findings underline the importance of treating embryos as dependent observations and suggest a high risk of patient-based confounding in retrospective studies. The impact of confounders and the embryo origin needs to be addressed in order to apply appropriate statistical models in observational studies. Furthermore, this observation emphasizes the need for RCTs for evaluating use of time-lapse parameters for embryo selection. STUDY FUNDING/COMPETING INTERESTS: Funding for the cohort study was provided by the Lippert Foundation, the Toyota Foundation, the Aase og Einar Danielsen foundation and NordicInfu Care research grant. Research at the Fertility Clinic, Aarhus University Hospital is supported by an unrestricted grant from MSD and Ferring. K.K. is funded by a grant from the Danish Council for Independent Research Medical Sciences. The authors declare no competing interest.

Global vitamin D levels in relation to age, gender, skin pigmentation and latitude: an ecologic meta-regression analysis.

UNLABELLED: We performed a meta-analysis of cross-sectional studies on serum 25(OH)D status globally. Serum 25(OH)D levels on average were 54 nmol/l, were higher in women than men, and higher in Caucasians than in non-Caucasians. There was no trend in serum 25(OH)D level with latitude. Vitamin D deficiency was widespread. INTRODUCTION: We studied vitamin D status (expressed as serum 25-hydroxy-vitamin D [25(OH)D]) in native subjects worldwide. METHODS: Meta-analysis and meta-regression of studies reporting on 25(OH)D in healthy subjects retrieved from Pubmed, Embase and Web of Science using the terms "serum", "25-hydroxy-vitamin D", "cholecalciferol", and "human". A total of 394 studies were included. RESULTS: The mean 25(OH)D level was 54 nmol/l (95% CI: 52-57 nmol/l). Women had borderline significantly higher 25(OH)D levels than men, and Caucasians had higher levels than non-Caucasians. 25(OH)D levels were higher in subjects aged >15 years than in younger subjects. Unadjusted there was no significant decrease in 25(OH)D with latitude (slope of curve -0.03 +/- 0.12 nmol/l per degree latitude north or south of equator, p = 0.8). There was a significant decline with latitude for Caucasians (-0.69 +/- 0.30 nmol/l per degree, p = 0.02), but not for non-Caucasians (0.03 +/- 0.39 nmol/l per degree, p = 0.14). After adjustment for age, gender, and ethnicity, no overall correlation was present between 25(OH)D and latitude (-0.29 +/- 0.24 nmol/l per degree, p = 0.23). CONCLUSION: There was no overall influence of latitude on 25(OH)D. However, in separate analyses 25(OH)D decreased with latitude in Caucasians but not in non-Caucasians. A widespread global vitamin D insufficiency was present compared with proposed threshold levels.

Longitudinal data analysis: an application to construction of a natural history profile of Duchenne muscular dystrophy.

A 30-month prospective study of 27 Scandinavian boys with confirmed diagnosis of Duchenne muscular dystrophy was carried out to construct profiles of the natural history of the disease. Assessments which included measures of voluntary muscle strength and function were done at 3 monthly intervals except for the first and second which were separated by 1 month. Recently developed statistical methods for analysis of longitudinal data with repeated observations on the same individual were used avoiding the problem of induced serial correlations. This allowed for the construction of both reference and prediction profiles for the variables %MRC, motor ability, walking time for 10 m and the sum of myometry of seven muscle groups.

Treatment of breast-cancer-related lymphedema with or without manual lymphatic drainage--a randomized study.

A prospective randomized study was carried out to investigate whether the addition of manual lymphatic drainage (MLD) to the standard therapy could improve treatment outcome in women with lymphedema of the ipsilateral arm after breast cancer treatment. Forty-two patients were randomly assigned to receive standard therapy or standard therapy plus MLD 8 times in 2 weeks and training in self-massage. The standard therapy consisted of use of a compression garment, exercises and information about lymphedema and skin care. The efficacy of treatment was evaluated by reduction in lymphedema volume during treatment and by improvement in symptoms potentially related to lymphedema. The patients were followed-up for a total of 12 months. The study showed that both groups obtained a significant reduction in edema and that MLD did not contribute significantly to reduce edema volume.

Changes in left ventricular function and mass during serial investigations after valve replacement for aortic stenosis.

BACKGROUND AND AIM OF THE STUDY: The potential for left ventricular hypertrophy regression and associated functional improvements may well be the underlying mechanism of results in general after valve replacement for aortic stenosis. The study aim was to investigate preoperative predictors and the time course of such ventricular changes. METHODS: Forty-six patients (mean age 61 years; range: 24-82 years) with aortic stenosis were prospectively followed with serial investigations (Doppler echocardiography, radionuclide ventriculography) at eight days (n = 43), three months (n = 42) and 18 months (n = 39) after valve replacement with a mechanical valve (19-29 mm). The postoperative course of left ventricular ejection fraction (EF), fast filling fraction, mass index and end-diastolic volume index (EDVi) was analyzed in an independent increments statistical model for repeated measurements. RESULTS: EF rose marginally, from 59+/-15% preoperatively to 64+/-16% (p <0.05) at 18 months, independently related to preoperative EF (p = 0.0001) and fast filling fraction (p = 0.0001). Changes in fast filling fraction were similarly predicted by the preoperative starting point (p = 0.003) and by preoperative left ventricular systolic radius:wall thickness ratio (p = 0.0002) with an inverse relation (the larger the chamber and the poorer its contractility, the lower the postoperative fast filling fraction). Mass index was independently related to the time point of postoperative measurement, indicating continuing regression of hypertrophy, from 200+/-66 g/m2 preoperatively to 148+/-49 g/m2 at 18 months (p <0.0001), when only 18% of the patients had normal mass index. Mass index was also independently related to preoperative left ventricular end-systolic dimension index (p = 0.0008) with a constant influence, and systolic wall stress (p = 0.0009) which was modified by time: the influence of wall stress was significant at eight days, weak at three months, and pronounced at 18 months. Left ventricular EDVi after surgery was inversely related to preoperative EF modified by time. Associated coronary artery disease, size of the prosthetic valve, and peak gradient across the valve (mean 15 mmHg; range: 7-26 mmHg at 18 months) did not influence any of the four target variables. CONCLUSION: We conclude that left ventricular response to valve replacement for aortic stenosis is predictable. A significant reduction in hypertrophy occurs during the first 18 postoperative months, but to a normal ventricular mass in only a minority of patients. Insufficient regression of hypertrophy was related to indices of irreversible myocardial disease, which also prevented functional ventricular improvement despite successful valve replacement and a hemodynamically well functioning valve.

A randomized comparative study of two methods for controlling Tendo Achilles contracture in Duchenne muscular dystrophy.

A 30-month prospective randomized study of 27 Scandinavian boys with confirmed diagnosis of Duchenne muscular dystrophy was done to compare the effect of passive stretching combined with the use of night splints (group A) or passive stretching (group B) on the evolution of Tendo Achilles contractures. Assessments were based on the methodology of Scott et al. (Muscle Nerve 1982;5:291-301)Analysis of the pattern and mechanism of dropout was done to eliminate bias between the two groups. Logistic regression showed that Tendo Achilles contracture was the most important variable (P=0.0020) for dropout. Methods of statistical analysis for longitudinal data avoiding induced serial correlations were used in the analysis. The expected annual change in Tendo Achilles contracture was found to be 23% less in group A than in group B after equalization for total muscle strength (%MRC).

No seasonal variation of insulin sensitivity and glucose effectiveness in men.

Insulin resistance is of pathogenetic importance for the development of non-insulin-dependent diabetes mellitus (NIDDM). However, not much is known about the variation in insulin sensitivity in the individual over longer periods. Consequently, we measured insulin sensitivity (Si) and glucose effectiveness (Sg) in healthy young men (N = 10) 5 times over a period of 15 months using a frequently sampled intravenous glucose tolerance test (FSIVGTT) with minimal-model analysis (study of seasonality). The maximal aerobic capacity (V(O2)max), fat-free mass, body mass index (BMI), and 24-hour ambulatory blood pressure (BP) were also assessed. Furthermore, we performed a study designed to evaluate the day-to-day variation in Si and Sg (study of day-to-day variation). Here, we studied Si and Sg in healthy young men (n = 8) within 2 weeks. In the study of seasonality, the coefficient of variation (CV) for Si was 24.0%, whereas the CV for Sg was 26.0%. Anticipating a seasonal variation in Si following a sine curve with a cycle length of 1 year and an unknown phase and amplitude, we tested this hypothesis with a multiple linear regression model that allows for different levels of Si between individuals, and failed to detect any impact due to this. Si (mean +/- SD, 1.17 +/- 0.28 x 10(-4) x min(-1) x pmol/L(-1), P = .38), Sg (0.023 +/- 0.006 min(-1), P= .71), fasting insulin (21.2 +/- 7.3 pmol/L, P= .98), V(O2)max (3.8 +/- 0.6 L/min, P= .13), and fat-free mass (64.9 +/- 2.5 kg, P = .92) were constant over time. In the study of day-to-day variation, we found a CV for Si of 17.3% and a CV for Sg of 23.3%. In conclusion, we found that the variations in Si and Sg were slightly higher than those found in studies performed to establish the day-to-day variation. However, no significant seasonal variation in Si and Sg was evident in this group of healthy young lean caucasian men. Consequently, indices of Si and Sg obtained at different times of the year appear comparable.

Accurate noninvasive quantitation of blood flow, cross-sectional lumen vessel area and wall shear stress by three-dimensional paraboloid modeling of magnetic resonance imaging velocity data.

OBJECTIVES: We present a new method in which a priori knowledge of the blood velocity fields within the boundary layer at the vessel wall, combined with acquisition of high resolution magnetic resonance imaging (MRI) blood velocity data, allow exact modeling at the subpixel level. BACKGROUND: Methods are lacking for accurate, noninvasive estimation of blood flow, dynamic cross-sectional lumen vessel area and wall shear stress. METHODS: Using standard acquisition of MRI blood flow velocity data, we fitted all data points (n = 69) within the boundary layer of the velocity profile to a three-dimensional paraboloid, which enabled calculation of absolute volume blood flow, circumferential vessel wall position, lumen vessel area and wall shear stress. The method was tested in a 8.00 +/ 0.01-mm diameter glass tube model and applied in vivo to the common carotid artery of seven volunteers. RESULTS: In vitro the lumen area was assessed with a mean error of 0.6%. The 95% confidence interval included the specified tube dimensions. Common carotid mean blood flow was 7.42 ml/s, and mean (standard error) diastolic/systolic vessel area was 33.25 (0.72 [2.2%])/43.46 (0.65 [1.5%]) mm2. Mean/peak wall shear stress was 0.95 (0.04 [4.2%])/2.56 (0.08 [3.1%]) N/m2. CONCLUSIONS: We describe a new noninvasive method for highly accurate estimation of blood flow, cross-sectional lumen vessel area and wall shear stress. In vitro results and statistical analysis demonstrate the feasibility of the method, and the first in vivo results are comparable to published data.

Automatic accurate non-invasive quantitation of blood flow, cross-sectional vessel area, and wall shear stress by modelling of magnetic resonance velocity data.

OBJECTIVES: To apply a new, automatic and non-invasive method for quantification of blood flow, dynamic cross-sectional vessel area, and wall shear stress (WSS) by in vivo magnetic resonance velocity mapping of normal subjects. DESIGN: Prospective, open study. MATERIALS: Six young volunteers. METHODS: A three-dimensional paraboloid model enabling automatic determination of blood flow, vessel distensibility and WSS was applied to blood velocity determinations in the common carotid artery. Blood flow was also determined by a manual edge detection method. RESULTS: Using the new method, the common carotid mean blood flow was 7.28 (5.61-9.63) (mean (range)) ml/s. By the manual-method blood flow was 7.21 (5.55-9.60) ml/s. Mean luminal vessel area was 26% larger in peak systole than in diastole. Mean/peak WSS was 0.82/2.28 N/m2. Manually and automatically determined flows correlated (r2 = 0.998, p < 0.0001). WSS and peak centre velocity were associated (r2 = 0.805, p < 0.0001). CONCLUSIONS: Blood flow, luminal vessel area dilatation, and WSS can be determined by the automatic three-dimensional paraboloid method. The hypothesis of association between peak centre velocity and WSS was not contradicted by the results of the present study.

Increasing age-specific incidence of hip fractures in a Danish municipality.

From 1970 to 1986 a total of 4578 hip fractures were registered in persons > or = 40 years in a Danish urban municipality. During that period, the annual number almost tripled. The statistical analysis revealed an increase in the age-specific incidence for both females and males (p < 0.0001). Furthermore, the number of fractures increased due to demographic aging. The overall incidence for individuals > or = 40 years increased 9-10% annually; from 2.1 to 5.4 per 1.000 females, and from 0.9 to 2.1 per 1.000 males. The female:male ratio was 2.8 without changes over time. If the observed increase in incidence is extrapolated to the next 17 years, a doubling of the annual number of fractures is predicted by the year 2002, whereas an unchanged incidence would result in a 20% increase in number.

Occurrence and incidence of the second hip fracture.

During a 16-year period, 256 second hip fractures were found in 3898 persons 40 years of age and older who had a previous hip fracture. Ninety-two percent of the second fractures were contralateral, and 68% of these were the same type as the first. Thus, 62% of the femoral neck and 72% of the trochanteric fractures were preceded by a contralateral fracture of the same type. The mean interval between fractures was 3.3 years, and there was no significant difference between genders or among fracture types. The risk of the first fracture was 1.6 per 1000 men per year and 3.6 per 1000 women, and for the second fracture 15 per 1000 men per year and 22 per 1000 women. This increase was highly significant for both genders, especially for men.

Age and sex as determinants of mortality after hip fracture: 3,895 patients followed for 2.5-18.5 years.

Between 1970 and 1985, a total of 3,895 persons > or = 40 years of age were treated for hip fracture in a Danish municipality. The female: male ratio was 2.7. The cumulative survival was estimated and compared with that of the age- and sex-matched general population. The analysis showed that (a) the mortality was increased compared with the expected mortality, especially during the first year (19% for women, 25% for men), but a statistically significant excess mortality could be demonstrated up to 10 years after a hip fracture; (b) the mortality increased with age, even after adjustment for the expected higher mortality of the elderly; and (c) the mortality was higher for men than for women, even after adjustment for the expected higher mortality of males. This difference was already present after the first 2 weeks among patients > or = 70 years of age.

Reliability of three computer methods in the analysis of ECG-gated radionuclide left ventriculography: interrecording, interobserver and intraobserver variability.

Three different computer methods for analysis of systolic and diastolic left ventricular function (ejection fraction, peak ejection rate, time to peak ejection rate, peak filling rate, time to peak filling rate, duration of fast filling phase, and fast filling fraction) as derived from ECG-gated radionuclide cardiography were compared in 30 patients with various diseases. The patients had two gamma camera recordings of the left ventricle performed immediately following one another during radionuclide (99mTc) equilibrium (3 x 10(6) counts, 16 frames/cycle, 64 x 64 pixels). Mean ECG R-R interval of the patients remained unchanged from first to second recording. The three computer methods were: (1) end-diastolic (ED) region of interest (ROI) analysis based on manually defined ED-ROI; (2) multi (M) ROI, manually defined ROI for each frame; and (3) semiautomatic (SA) ROI, ROI for each frame defined by an SA edge detection technique. With the 16 frame points as nodes, a 160-point time-activity curve was constructed for each of the three methods by use of a spline function. A tailored multiway analysis of variance showed that the M-ROI method had the highest interindividual range of values of the function parameters and the smallest interrecording, interobserver, and intraobserver variabilities. In theory this implies a better diagnostic sensitivity and specificity for the M-ROI method as compared with the other two methods.

Intralesional and metastatic heterogeneity in malignant melanomas demonstrated by stereologic estimates of nuclear volume.

Regional variability of nuclear 3-dimensional size can be estimated objectively using point-sampled intercepts obtained from different, defined zones within individual neoplasms. In the present study, stereologic estimates of the volume-weighted mean nuclear volume, nuclear vv, within peripheral and central zones of cutaneous, primary, and secondary malignant melanomas were obtained. For comparisons, estimates of nuclear vv were also obtained from primary and metastatic melanomas, based on stereologic measurements taking the whole tumor sectional areas into consideration. Estimates of nuclear vv were on average larger in the peripheral zones of primary melanomas, than nuclear vv in central zones (2p = 6.7 x 10(-4), whereas no zonal differences were demonstrated in metastatic lesions (2p = 0.21). A marked intraindividual variation was demonstrated between primary and corresponding secondary melanomas (2p = 1.7 x 10(-39). These results do not necessarily indicate clonal heterogeneity, but are in agreement with the hypothesis of polyclonal and monoclonal composition of primary and metastatic neoplasms, respectively. Estimates of nuclear vv based on measurements of whole sectional areas of primary melanomas showed large interindividual variation. This finding emphasizes that unbiased estimates of nuclear vv are robust to regional heterogeneity of nuclear volume and thus suitable for purposes of objective, quantitative malignancy grading of melanomas.

Independent predictors of operative mortality and postoperative complications in surgically treated carcinomas of the oesophagus and cardia--is the aggressive surgical approach worthwhile?

During the 25 years 1960-84, 657 patients were operated on for carcinomas of the thoracic oesophagus (n = 347) or gastric cardia (n = 310). Resection was carried out in 514 (78%) and oesophagogastrostomy in 481 (73%). Overall operative mortality (defined as death within 30 days) was 19% (n = 122). Pulmonary complications developed in 167 patients (25%), cardiovascular complications in 100 (15%), and anastomotic leakage and mediastinitis in 36 (6%). After radical resection of a localised tumour (n = 144), or non-localised tumour (n = 224), pallative resection (n = 146), or exploration (n = 143), the operative mortality and five year cumulative survival were 10% and 26%, 15% and 8%, 27% and 0, and 24% and 0, respectively (p less than 0.01 and p less than 0.0001). Using logistic regression analysis several variables were found to be independent predictors of operative mortality, pulmonary complications, cardiovascular complications, and anastomotic leakage. The predictor variables reflected both general preoperative status of the patients, preexisting cardio-pulmonary diseases, stage of the cancer, and surgical procedures. Based on the final logistic regression models the patients were stratified into risk groups (12 for operative mortality, pulmonary complications, and cardiovascular complications, and eight for anastomotic leakage). Operative mortality varied from 0 to 80%, pulmonary complications from 3 to 100%, cardiovascular complications from 0 to 100%, and anastomotic leakage from 0 to 50% (p less than 0.0001 in each case). Given the high operative mortality and complication rates, and the low five year survival rate after palliative procedures or exploratory operations, a more selective surgical approach seems warranted. Patients likely to have a good response should be identified before operation.

Risk stratification and long-term results after surgical treatment of carcinomas of the thoracic esophagus and cardia. A 25-year retrospective study.

During 25 years (1960 to 1984), 657 patients (aged 22 to 91, mean 66 years) were operated on for carcinomas (squamous cell, n = 230; adenocarcinoma, n = 399; anaplastic, n = 28) of the thoracic esophagus (n = 347) or gastric cardia (n = 310). Esophagogastrectomy was accomplished in 514 patients, of whom 94% (n = 481) had an "inkwell" esophagogastrostomy performed. The hospital mortality rate (less than or equal to 30 days) was 19% and the 5-year cumulative survival rate was 9% +/- 1% (standard error). A Cox regression analysis enabled a detailed risk stratification of the patients. T, N, and M class and age were the strongest predictor variables. The general status of the patients, including pulmonary disease, also had a strong prognostic influence. Eight risk groups were identified having 1-year and 5-year survival rates of 71%/41% (n = 35), 69%/24% (n = 80), 47%/11% (n = 125), 30%/6% (n = 139), 12%/0% (n = 105), 6%/0% (n = 71), 2%/0% (n = 57), and 0%/0% (n = 45) (p less than 0.0001). Hospital mortality (from 6% to 42%) and complication rates increased significantly from the low-risk to high-risk groups. Comparisons with survival rates of background populations matched to each of the first four risk groups indicated that the benefit of surgical treatment may be underestimated if only patient survivals are judged. Use of modern endoscopic and noninvasive tests may yield a reliable TNM classification without surgical exploration. Given the extremely poor prognostic outlook together with high hospital mortality and complication rates of the four last risk groups, an aggressive surgical approach with resection whenever possible can no longer be regarded rational. Selection for surgical treatment should be based on a detailed risk estimation that takes into account both TNM classification and general patient status.