Quantifying the Likelihood of Structural Models through a Dynamically Enhanced Powder X-Ray Diffraction Protocol.

Structurally characterizing new materials is tremendously challenging, especially when single crystal structures are hardly available which is often the case for covalent organic frameworks. Yet, knowledge of the atomic structure is key to establish structure-function relations and enable functional material design. Herein, a new protocol is proposed to unambiguously predict the structure of poorly crystalline materials through a likelihood ordering based on the X-ray diffraction (XRD) pattern. Key of the procedure is the broad set of structures generated from a limited number of building blocks and topologies, which is submitted to operando structural characterization. The dynamic averaging in the latter accounts for the operando conditions and inherent temporal character of experimental measurements, yielding unparalleled agreement with experimental powder XRD patterns. The proposed concept can hence unquestionably identify the structure of experimentally synthesized materials, a crucial step to design next generation functional materials.

The short IQCODE as a predictor for delirium in hospitalized geriatric patients.

BACKGROUND: Delirium is a serious complication, which occurs frequently in older patients with pre-existing cognitive impairment. There is a need for a simple tool to assess chronic cognitive impairment and the associated risk of delirium during hospitalization. AIMS: To assess the usefulness of the short IQCODE questionnaire in predicting delirium during hospitalization in older patients in a geriatric ward. METHODS: A prognostic study in the Geriatric Department at Aarhus University Hospital, Aarhus Denmark. Consecutive patients were enrolled during March to December, 2017. After consent of the patient, the staff interviewed the relatives by phone using the short IQCODE questionnaire. Delirium was assessed morning and evening until discharge by the Confusion Assessment Method. The ability of short IQCODE to predict delirium was examined. RESULTS: Three hundred and fifty-three patients were eligible, and 306 completed the IQCODE. Delirium occurred among 19% of the patients during hospitalization. The IQCODE score was associated with the risk of delirium with a receiver operating characteristic (ROC) area of 0.72. A cut-point of 3.3 could separate the patients in a larger group with a risk of approximately 26% to develop delirium and a smaller group having a risk of approximately 6%. CONCLUSION: The IQCODE is a useful tool to predict delirium among older inpatients, but it may not stand alone. It can be a useful supplement to other clinical information and observations in detecting patients needing dementia-friendly treatment and care.

Low risk of venous thromboembolism following early pregnancy loss in pregnancies conceived by IVF.

STUDY QUESTION: What is the risk of venous thromboembolism (VTE) in the 12 weeks following early pregnancy loss in pregnancies conceived by IVF? SUMMARY ANSWER: The VTE risk is low in the 12 weeks following early pregnancy loss in pregnancies conceived by IVF. WHAT IS KNOWN ALREADY: There is an excess VTE risk during first trimester in complete IVF pregnancies leading to delivery. It is unknown whether this excess VTE risk also is present in IVF pregnancies terminated by early pregnancy loss (implantation failure, missed abortion, or spontaneous or induced abortion before 10 completed gestational weeks). STUDY DESIGN, SIZE, DURATION: A nationwide registry-based cohort study including all Danish IVF pregnancies registered in the Danish IVF Register between 1995 and 2005. Women who underwent frozen embryo replacement or oocyte donation were not included. PARTICIPANTS/MATERIALS, SETTING, METHODS: We included all 24 931 IVF treatments leading to pregnancy among 19 260 women. We identified 16 701 complete IVF pregnancies, 7567 IVF pregnancies with early pregnancy loss, and 663 IVF pregnancies terminated by late abortion (≥gestational weeks 10). We followed women for 12 weeks after termination of pregnancy and calculated the absolute risk of VTE during follow-up with 95% CI. As a relative risk estimate, we calculated the risk ratio for VTE following IVF pregnancies with early loss compared to the VTE risk following complete IVF pregnancies. MAIN RESULTS AND THE ROLE OF CHANCE: During the 12 weeks of follow-up, only one case of VTE occurred in the group of early pregnancy loss, none in the late abortion group, and 13 VTE cases occurred in complete IVF pregnancies. The VTE risk per 10 000 pregnancies was 1.3 [0.03-7.4] for IVF pregnancies with early loss and 7.8 [4.1-13.3] for complete pregnancies; the corresponding risk ratio was 0.17 [0.02-1.3]. Thus, we found a low absolute VTE risk in the 12 weeks following early pregnancy loss in IVF pregnancies. The relative VTE risk was low in comparison to the VTE risk in early gestation reported for complete IVF pregnancies and for Danish background pregnancies. LIMITATIONS, REASONS FOR CAUTION: Despite the use of complete nationwide data, only a few VTE events were available for the statistical analyses thereby limiting the precision of our estimates. We included both inpatient and outpatient VTE hospital diagnoses, but we cannot rule out the occurrence of VTE cases not diagnosed at hospital and hence not registered in the National Patient Registry. No information was available on the use of prophylactic low molecular weight heparin: access to such might have helped to explain our findings, but would not have changed our conclusions. WIDER IMPLICATIONS OF THE FINDINGS: The results of the present study do not indicate a need for prophylactic anticoagulant therapy in women suffering early IVF pregnancy loss without other risk factors. STUDY FUNDING/COMPETING INTEREST(S): No external funding was obtained for this study. There are no conflicts of interest to declare.

Increased venous thrombosis incidence in pregnancies after in vitro fertilization.

STUDY QUESTION: Is venous thrombosis risk increased in pregnancies after in vitro fertilization? SUMMARY ANSWER: The venous thrombosis incidence was significantly increased in pregnancies after in vitro fertilization; especially in the first trimester and in the first 6 weeks post-partum. WHAT IS KNOWN ALREADY: In vitro fertilization without pregnancy is not associated with increased venous thrombosis incidence. STUDY DESIGN, SIZE, DURATION: This national register-based cohort study covered the period from 1995 to 2005. PARTICIPANTS/MATERIALS, SETTING, METHODS: All Danish pregnancies conceived by in vitro fertilization (n = 18 787) were included. Venous thrombosis incidence rates in pregnancies after in vitro fertilization were compared with venous thrombosis incidence rates in reference pregnancies, by calculating incidence rate ratios. MAIN RESULTS AND THE ROLE OF CHANCE: In total, 48 cases were identified. In pregnancies after in vitro fertilization, the overall venous thrombosis incidence rate was 28.6 per 10 000 pregnancy-years (95% confidence interval (CI) 20.6-39.6) in comparison to 10.7 per 10 000 woman-years in reference pregnancies. Post-partum, the venous thrombosis incidence rate was 27.9 per 10 000 woman-years (95% CI 15.8-49.1) after in vitro fertilization in comparison to 17.5 per 10 000 woman-years in reference pregnancies. The overall venous thrombosis incidence rate ratio during in vitro fertilization (IVF) pregnancies compared with reference pregnancies was 3.0 (95% CI 2.1-4.3). The venous thrombosis incidence rate ratios during pregnancy were 2.8 (95% CI 1.9-4.1) in singleton IVF pregnancies and 4.4 (95% CI 2.4-8.3) in multiple IVF pregnancies, compared with reference pregnancies. The venous thrombosis incidence rate ratio post-partum was 1.2 (95% CI 0.6-2.8) for singleton IVF pregnancies and 3.9 (95% CI 1.7-8.8) for multiple IVF pregnancies compared with reference pregnancies. The post-partum venous thrombosis risk was higher in multiple IVF pregnancies compared with singleton IVF pregnancies. Maternal age, smoking and parity did not significantly affect the venous thrombosis risk. Ovarian hyperstimulation syndrome and polycystic ovarian syndrome did increase the risk of venous thrombosis during pregnancy. Caesarean section also increased the post-partum venous thromboembolism risk, but the increase was not significant. LIMITATIONS, REASONS FOR CAUTION: Other known confounders in our reference population could have contributed to the results. Access to such data may have helped to explain the observations, but would not have changed the conclusion that IVF pregnancies have an increased risk of venous thrombosis compared with other pregnancies. WIDER IMPLICATIONS OF THE FINDINGS: Our study adds new insights by demonstrating an excess venous thrombosis incidence post-partum after in vitro fertilization. The high venous thrombosis incidence in first trimester after in vitro fertilization supports previous studies. Our findings are generalizable to other Western Countries. STUDY FUNDING/COMPETING INTERESTS: Expenses for the acquirement of data were covered by a grant from The Secretary of Doctors further education, Central Denmark Region. None of the authors have any competing interests to declare. TRIAL REGISTRATIONS NUMBER: Not applicable.

Exploring the phase stability in interpenetrated diamondoid covalent organic frameworks.

Soft porous crystals, which are responsive to external stimuli such as temperature, pressure, or gas adsorption, are being extensively investigated for various technological applications. However, while substantial research has been devoted to stimuli-responsive metal-organic frameworks, structural flexibility in 3D covalent organic frameworks (COFs) remains ill-understood, and is almost exclusively found in COFs exhibiting the diamondoid (dia) topology. Herein, we systemically investigate how the structural decoration of these 3D dia COFs-their specific building blocks and degree of interpenetration-as well as external triggers such as temperature and guest adsorption may promote or suppress their phase transformations, as captured by a collection of 2D free energy landscapes. Together, these provide a comprehensive understanding of the necessary conditions to design flexible diamondoid COFs. This study reveals how their flexibility originates from the balance between steric hindrance and dispersive interactions of the structural decoration, thereby providing insight into how new flexible 3D COFs can be designed.

No evidence that assisted reproduction increases the risk of thrombosis: a Danish national cohort study.

BACKGROUND: Case reports have reported venous and arterial thromboses in women undergoing assisted reproduction. No large systematic studies on the risk of thrombosis have been published. The objective of our study was to investigate whether the risk of thrombosis is increased in women undergoing assisted reproduction. METHODS: A national register-based cohort study was conducted on all women undergoing IVF or ICSI treatment in Denmark from 1994 to 2005. Data were obtained from the National Patient Registry and the IVF Registry. Women with prior malignant or cardiovascular disease were excluded. Thrombosis occurring within the first 6 and 12 months after assisted reproduction was considered potentially related to the treatment. Thromboses during pregnancy as well as the pregnancy-related diagnoses were excluded from the statistical analysis. The incidence rates of venous and arterial thromboses were compared with previously published estimates of the risk of thrombosis among young Danish women. RESULTS: We analyzed 30 884 Danish women undergoing 75 141 treatments from 1994 to 2005. The mean age of the women at first treatment was 32.3 years. The delivery rate per cycle was 22%. The incidence rate ratio, with 95% confidence interval (CI), of venous thrombosis within 6 months was 0.95 (CI: 0.38-1.95). The incidence rate ratio of arterial thrombosis within 6 months was 0.36 (CI: 0.04-1.30). CONCLUSIONS: Our study showed no evidence that assisted reproduction increases the risk of thrombosis.

Evidence for an early degradative event to the insulin molecule following binding to hepatocyte receptors.

We have used photoreactive insulin analogues to investigate as related processes, early structural modification of the receptor-bound insulin molecule and internalisation of the insulin-receptor complex. In isolated rat hepatocytes an initial modification of bound insulin leads to the generation of a molecular species unchanged in molecular weight but with reduced receptor and antibody binding affinities and altered electrophoretic mobility. Using photoreactive insulin analogues and density gradient cell fractionation the insulin receptor complex has been shown to undergo internalisation from the plasma membrane to a low density vesicular fraction, the endosome. No labelled material was found in lysosomal fractions after up to 10 min incubation at 37 degrees C. The degree of labelling of the endosome fraction depended on the position of the photoreactive group within the insulin molecule. The data suggest that before or during endocytosis, a small peptide is proteolytically cleaved from the C terminus of the insulin B chain.

Demonstration that the insulin receptor undergoes an early structural modification following insulin binding.

Processing of the insulin receptor by hepatocytes was studied using a 125I-labelled photoreactive insulin derivative which could be covalently attached to the receptor and facilitate the analysis of receptor structure in isolated subcellular fractions by SDS-polyacrylamide gel electrophoresis. Following binding at the cell surface, the label was rapidly internalised and located in a low-density subcellular fraction ('endosomes'). The intact receptor (350 000 molecular weight) and binding (alpha) subunit (135 000), produced by in vitro disulphide reduction of the samples, were found in the plasma membrane fraction but not in endosomes. In endosomes, the label was concentrated in a band at 140 000 (non-reduced) which on reduction generated species of 100 000 and 68 000 predominantly. The insulin receptor therefore undergoes an early structural change during endocytosis. This modification does not involve complete disulphide reduction and may be due to a proteolytic event.

Patients' attitudes toward detection of heterozygous familial hypercholesterolemia.

BACKGROUND: Molecular biology has improved the diagnostic abilities of physicians and enabled them to identify apparently healthy persons with a high risk of genetic disease. OBJECTIVE: To examine the attitudes toward detection of disease and the present well-being in persons at risk of disease with a modifiable outcome, in this case heterozygous familial hypercholesterolemia (FH) and their hypercholesterolemic relatives. MATERIALS AND METHODS: A questionnaire was developed that collected information on the impact on well-being at present and at the time of diagnosis of hypercholesterolemia and on attitudes toward screening family members for heterozygous FH. It was mailed to 62 index patients with heterozygous FH, which was defined by using clinical criteria, and 108 hypercholesterolemic relatives. The response rate was 88%. Results were related to demographic data, experience of psychological or physiologic reactions in relation to awareness of hypercholesterolemia, cardiovascular symptoms, lipid-lowering drug treatment, and information on the DNA-based diagnosis. RESULTS: Of the respondents, anxiety was expressed by 44%, fear of coronary heart disease by 37%, and diminished well-being by 13%. These findings were most pronounced in persons who had experienced physiologic or psychological reactions at the time of diagnosis of hypercholesterolemia or who already had heart disease. Six percent regretted that they were aware of their disease diagnosis, and 84% were in favor of screening for affected individuals in families with a history of heterozygous FH. CONCLUSIONS: The results indicated that a substantial proportion of persons with heterozygous FH had some degree of anxiety. A small minority regretted that they were informed of the diagnosis of heterozygous FH, however, and a majority were in favor of family screening for heterozygous FH.

Incidence and late prognosis of cushing's syndrome: a population-based study.

The main purpose was to assess the incidence and late outcome of Cushing's syndrome, particularly in Cushing's disease. Information for all patients diagnosed with Cushing's syndrome during an 11-yr period in Denmark was retrieved. The incidence was 1.2-1.7/million.yr (Cushing's disease), 0.6/million.yr (adrenal adenoma) and 0.2/million.yr (adrenal carcinoma). Other types of Cushing's syndrome were rare. In 139 patients with nonmalignant disease, 11.1% had died during follow-up (median, 8.1 yr; range, 3.1-14.0), yielding a standard mortality ratio (SMR) of 3.68 [95% confidence interval (CI), 2.34-5.33]. The SMR was partly attributable to an increased mortality within the first year after diagnosis. Eight patients died before treatment could be undertaken. The prognosis in patients with malignant disease was very poor. Patients in whom more than 5 yr had elapsed since initial surgery were studied separately, including a questionnaire on their perceived quality of health. In 45 patients with Cushing's disease who had been cured through transsphenoidal neurosurgery, only 1 had died (SMR, 0.31; CI, 0.01-1.72) compared with 6 of 20 patients with persistent hypercortisolism after initial neurosurgery (SMR, 5.06; CI, 1.86-11.0). In patients with adrenal adenoma, SMR was 3.95 (CI, 0.81-11.5). The perceived quality of health was significantly impaired only in patients with Cushing's disease and appeared independent of disease control or presence of hypopituitarism. It is concluded that 1) Cushing's syndrome is rare and is associated with increased mortality, in patients with no concurrent malignancy also; 2) the excess mortality was mainly observed during the first year of disease; and 3) the impaired quality of health in long-term survivors of Cushing's disease is not fully explained.

Usefulness of ST deviation induced by prolonged hyperventilation as a predictor of cardiac death in angina pectoris.

One hundred ninety patients with chronic angina for an average of 2 years were followed. Forty-seven had at least 1 mm of ST deviation in response to provocation of coronary vasoconstriction by prolonged hyperventilation (group I); 143 had no ST deviation (group II). The angiographic response to this test was studied in 21 patients from group I, and revealed 25 to 100% diameter reduction; in group II 9 patients showed a 5 to 14% diameter reduction. In group I, 15 patients (32%) died (hazard rate = 0.17 deaths/patients X years) vs 18 (13%) in group II (hazard rate = 0.06) (p less than 0.01). Seven patients in group I (15%) and 3 in group 2 (2%) died while waiting for surgery (p less than 0.01). All patients who died had coronary stenoses of at least 70%. A Cox regression analysis, using 24 variables (invasive and noninvasive), showed a positive hyperventilation test (ST deviation at least 1 mm), low ejection fraction and systolic blood pressure of at least 160 mm Hg to be independent predictors of death (p less than 0.05). Considering only deaths in non-operated patients (patients waiting for surgery and patients not planned to undergo operation), a rate-pressure product/100 of 150 or less at exercise stress testing, left ventricular end-diastolic pressure of 15 mm Hg or more and duration of angina less than 1 year were also independent predictors of death. Thus, the hyperventilation test may be useful for identifying angina patients who are at high risk of cardiac death due to dynamic coronary obstructions.

Urinary tract anomalies associated with congenital diaphragmatic defects.

Anomalies of the urinary tract occur in some 13-27% of infants with congenital posterolateral diaphragmatic defect (CDD) and are often severe (renal agenesis, dysplasia, hypoplasia, or hydronephrosis). To test the hypothesis that urinary and diaphragmatic anomalies share elements of pathogenesis, we reviewed 60 autopsy cases of CDD studied at our institution. Sixteen patients (27%) manifested anomalies of the urinary tract: 12 had markedly altered kidneys, 8 of which were unilateral and ipsilateral to the diaphragmatic defect. Among 27 patients free of gross urinary tract anomalies, kidney weights formed a skewed distribution, with most values above published norms for body weight; by analysis of covariance, kidney weight (as a function of body weight) was significantly greater in CDD than in a control population of infants free of chronic illnesses and congenital anomalies who died suddenly and unexpectedly. Differences in glomerular number and diameter could not be identified between the latter groups. In 71% of patients with isolated left CDD, the left kidney was heavier than the right, a reversal of the usual condition in infancy. These findings demonstrate that both marked and subtle changes of the urinary tract in CDD are generally ipsilateral to the diaphragmatic defect and suggest that the pathogenetic mechanisms responsible for urinary and diaphragmatic anomalies may overlap topographically. Aberrant morphogenesis within a developmental field or fields is one explanation for this.

Morbidity in Turner syndrome.

Turner syndrome afflicts approximately 50 per 100,000 females and is characterized by retarded growth, gonadal dysgenesis, and infertility. Much attention has been focused on growth and growth promoting therapies, while less is known about the natural course of the syndrome, especially in adulthood. We undertook this study to assess the incidence of diseases relevant in the study of Turner syndrome. The study period was from January 1, 1984 to December 31, 1993, and the study base was all women living in Denmark during the study period. We used data from the Danish Cytogenetic Central Register and the Danish National Registry of Patients to assess morbidity. This study supports several earlier studies reporting increased morbidity and confirms results of a recent study on cancer in Turner syndrome. Women with Turner syndrome seem to have an increased incidence of fractures, osteoporotic fractures in adulthood, and non-osteoporotic fractures in childhood. Furthermore, diabetes mellitus, both NIDDM and IDDM, was found with a markedly increased incidence in Turner syndrome, as well as ischemic heart disease, hypertension, and stroke. The risk of cancer, except cancer of the large bowel, does not seem to be elevated in Turner syndrome. Our data suggest that patients with Turner syndrome are extraordinarily prone to abnormalities constituting the metabolic syndrome (e.g., hypertension, dyslipidaemia, NIDDM, obesity, hyperinsulinemia and hyperuricemia). The present data may help to explain the decreased life span found in patients with Turner syndrome.

Time to pregnancy as a correlate of fecundity: differential persistence in trying to become pregnant as a source of bias.

BACKGROUND: Subfecundity is a frequent and often serious problem and it is important to identify its preventable determinants and to monitor fecundity over time. Since follow-up studies are difficult and expensive to conduct, time to pregnancy (TTP) in pregnant women is often used as a surrogate measure of fecundity. TTP data can be retrieved at low costs and they need no valid population registry as a source for sampling. While TTP may serve as a valid surrogate measure in many situations, its validity rests upon a number of assumptions. We have analysed one of these overlooked assumptions, the importance of persistence in trying to become pregnant. METHODS: By means of computer simulations we estimated bias caused by differences in persistence in pregnancy attempts. We investigated whether the assumptions made in the simulation were realistic by using empirical data from a European study. RESULTS: The mean waiting time to pregnancy and other estimates of subfecundity (or infertility) strongly depend upon the persistence of couples in pursuing a pregnancy. We show that even moderate changes in the planning behaviour considerably modify the waiting time distribution. Empirical data confirm that persistence in trying to become pregnant is age-related. CONCLUSIONS: Persistence in pregnancy attempts affects outcome measures of subfecundity in studies based upon TTP in pregnant women. It is likely that the length of time during which couples keep trying to become pregnant is influenced by a number of factors which would probably change over time or be different between populations to be compared.

Hospital based screening of 65-73 year old men for abdominal aortic aneurysms in the county of Viborg, Denmark.

OBJECTIVE - To analyse the benefits of screening older men for abdominal aortic aneurysms. METHODS - A hospital based screening trial concerning 13 500 65-73 year old men using B-mode ultrasonographic scanning. To improve the response rate the invited men could change their appointment, and nonresponders were reinvited. RESULTS - Results from the first year of the trial are presented. Among 4404 invited, 3344 (76%) were scanned. The primary response rate was 64.8%, but a further 11.2% were scanned after revised appointments or reinvitation. The whole infrarenal aorta could be visualised in 97-6%, and the distal part in 99.7% of the scans. The time taken for each scan was 9-7 minutes and the costs per scan were $9.50. One hundred and fifty three subjects (4.6%) had aortic diameters of 25-29 min, and 141 (4.2%) had an abdominal aortic aneurysm, 19 (0.6%) above 49 mm in diameter. CONCLUSION - In Denmark the short term costs and benefits of screening older men for abdominal aortic aneurysms seem realistic. Long term costs and benefits need to be investigated.

Variation and information in white blood cell differential counts.

The magnitude and sources of variation in the white blood cell (WBC) count and differential count affect their information content and clinical value. This study describes components of variation in the WBC count and differential, estimates the magnitude of each component, and uses computer simulations to compare the information conveyed by the total WBC count and by the WBC differential count, expressed as the number of each type of cell and as the percentage of each cell type. Biologic variation is much greater than statistical sampling variation in differential WBC counts, even when a relatively small number of cells is classified. The commonly reported neutrophil percentage is inferior both to the neutrophil count and to the total WBC count in its ability to detect neutrophilia and to detect recovery from elevated levels. This conclusion holds for single as well as for sequential WBC differential determinations and regardless of the level of test result at which the clinician considers disease to be present. The total WBC count and the neutrophil count differ little in performance, so a neutrophilic patient's return to normal levels can safely be detected and monitored by relying on the less expensive total WBC count and foregoing repeated requests for differential counts.

The distribution of receptors for the pro-inflammatory cytokines interleukin (IL)-6 and IL-8 in the developing human fetus.

UNLABELLED: Interleukin-6 receptor alpha (IL-6R) and interleukin-8 receptor (IL-8RB) are widely expressed in adult human and murine tissues. Little is known about the expression of these receptors and the function of their ligands in the developing human fetus. OBJECTIVES: To determine the tissue distribution and cellular expression of IL-6R and IL-8RB in the developing human fetus. METHODS: Reverse transcription-PCR and immunohistochemical staining were performed on brain, spinal cord, eye, heart, lung, liver, spleen, adrenal, kidney, intestine, and placenta from fetuses of 8 and 16+/-2 weeks post-conception. RESULTS: IL-6R and IL-8RB mRNA were detected in all tissues tested at both time points. Immunoreactivity to anti-IL-6R antibody was present on neurons, and in neuropil of the brain, as well as in bone marrow, bronchi, hepatocytes, zona glomerulosa of the adrenal, glomerular cells in kidney, spleen, and placental trophoblasts. Cell-specific expression for IL-8RB in the central nervous system was localized to specific groups of neurons and astrocytes in the brain and spinal cord, including the neural retina. In somatic organs IL-8RB was detected in bone marrow, myocardiocytes, bronchiolar epithelial cells, hepatocytes, cells of the zona glomerulosa and the zona fasciculata of the adrenal, the collecting system of the kidney, enterocytes of the bowel and in placental cells. CONCLUSION: The widespread expression of these cytokine receptors suggests a nonhematopoietic role for their ligand in the developing fetus.

Tissue distribution of erythropoietin and erythropoietin receptor in the developing human fetus.

OBJECTIVE: Erythropoietin receptors (Epo-R) have been demonstrated on several nonhematopoietic cell types in animal models and in cell culture. Our objective was to determine the tissue distribution and cellular specificity of erythropoietin (Epo) and its receptor in the developing human fetus. STUDY DESIGN: The expression of Epo and Epo-R mRNA was ascertained by RT-PCR for organs ranging in maturity from 5 to 24 weeks postconception. The cellular location of protein immunoreactivity was then determined using specific antiEpo and antiEpo-R antibodies. Antibody specificity was established by Western analysis. RESULTS: mRNA for Epo and Epo-R was found in all organs in the first two trimesters. Immunolocalization of Epo was limited to the liver parenchymal cells, kidney interstitial cells and proximal tubules, neural retina of the eye, and adrenal cortex. As development progressed, immunoreactivity in the kidney became more prominent. In contrast, immunoreactivity for Epo-R was widespread throughout the body, in cell types including endothelial cells, myocardiocytes, macrophages, retinal cells, cells of the adrenal cortex and medulla, as well as in small bowel, spleen, liver, kidney, and lung. CONCLUSIONS: The distribution of Epo and its receptor is more widespread in the developing human than was initially postulated. Epo-R is expressed on many cell types during early fetal development, leading us to speculate that Epo acts in concert with somatic growth and development factors during this period. Further investigation is required to understand the nonhematopoietic role of Epo during human development.

Thrombopoietin (Tpo) in the fetus and neonate: Tpo concentrations in preterm and term neonates, and organ distribution of Tpo and its receptor (c-mpl) during human fetal development.

Little is known about thrombopoietin (Tpo) production in human fetuses and neonates. As a step toward determining whether Tpo is relevant to platelet production in the fetus and neonate, we hypothesized that: (1) like other cytokines, Tpo is present in the cord blood in higher concentrations than in adult plasma; (2) Tpo and its receptor (c-mpl) are expressed in fetuses at, and following, 5-6 weeks post-conception (when platelet production begins); and (3) the sites of Tpo and c-mpl production in the fetus are similar to those of adults. We quantified Tpo, by ELISA, in the plasma of 50 adults, as well as in the umbilical cord plasma of 50 preterm and term infants. We also characterized, by RT-PCR, the organ distribution of Tpo and c-mpl during fetal development (at 8 and 16 weeks). Tpo concentrations were measurable (> or =41 pg/ml) in only two of the 50 adult samples (44 and 46 pg/ml), but in 24 of the 50 cord plasma samples (of the 24 samples, the median was 62 pg/ml; mean+/-SD, 80+/-39 pg/ml). Tpo levels did not correlate with either gestational age or platelet count at birth. Similarly to adults, in the fetal tissues, Tpo transcripts were found in all organs tested, but the most dense bands were from liver. C-mpl transcripts were also predominantly from liver. We conclude that: (1) Tpo is present in higher concentrations in cord plasma than in venous plasma of adults; (2) Tpo and c-mpl transcripts are detected in human fetuses as early as the onset of platelet appearance; and(3) Tpo and c-mpl have a similar organ distribution in fetuses and adults.

The first-appearance of neutrophils in the human fetal bone marrow cavity.

We sought to define the time of first-appearance of neutrophils within the developing human bone marrow cavity, and to compare this with the time of appearance of G-CSF and its receptor (G-CSF-R) at that site. We hypothesized that the onset of G-CSF production is an initiation signal for neutrophil production within the marrow cavity, and that therefore G-CSF mRNA and G-CSF protein in the marrow cavity would immediately precede the first-appearance of neutrophils. To test this, we determined the time of first-appearance of neutrophils in the clavicular marrow space using a monoclonal antibody against myeloperoxidase (MPOAb), and then validated these findings by flow cytometric analyses, for neutrophil cell-surface markers, of cells flushed from the marrow cavity. After thus defining the time of first-appearance of neutrophils, specific mRNA transcripts for G-CSF and G-CSF-R were sought from clavicles of varying gestational ages, using RT-PCR, and the presence of these proteins in the clavicles were sought using immunohistochemistry. We observed that; (1) MPO+ cells first appeared in the clavicular marrow cavity between the 10 to 11th weeks post-conception, (2) Flow cytometric analyses confirmed that these MPO+ marrow cells included CD11b+, CD15+ neutrophils, (3) Transcripts for G-CSF and G-CSF-R, and the specific G-CSF and G-CSF-R proteins, were present in the clavicles by 6 weeks post-conception, 4 to 5 weeks before the first-appearance of neutrophils. Thus, neutrophils first appear in the human clavicular marrow at 10-11 weeks post-conception, and G-CSF and G-CSF-R are present in the developing bone rudiment preceding the appearance of neutrophils. It is unclear whether neutrophils arise in the marrow cavity in response to the onset of production of G-CSF or to other initiation signals.