Retroviral gene transfer to primitive normal and leukemic hematopoietic cells using clinically applicable procedures.

Clinical uses of gene transfer to bone marrow transplants require the establishment of a reproducible method for infecting large numbers of very primitive hematopoietic cells at high efficiency using cell-free retrovirus-containing media. In this study we report the results of experiments with preparations of a high-titer (2-5 x 10(7)/ml) helper-free recombinant neo(r) retrovirus that indicate this goal can now be achieved based on measurements of gene transfer efficiencies to cells referred to as long-term culture initiating cells (LTC-IC) because they give rise to clonogenic cells after greater than or equal to 5 wk in long-term culture (LTC). Intermittent, repeated exposure of normal human marrow mononuclear cells to virus-containing supernatant over a 3-d period of cell maintenance on an IL-3/granulocyte colony-stimulating factor (G-CSF) producing stromal layer resulted in gene transfer efficiencies to LTC-IC of 41%; a level previously obtainable only using co-cultivation infection techniques. Marrow cells enriched greater than or equal to 500-fold for LTC-IC (1-2% pure) by flow cytometry showed gene transfer efficiencies of 27% when infected in a similar fashion over a shorter period (24 h), but in the presence of added soluble IL-3 and G-CSF without stromal feeders, and this increased to 61% when Steel factor was also present during the infection period. By using a less highly enriched population of LTC-IC obtained by a bulk immunoselection technique applicable to large-scale clinical marrow harvests, gene transfer efficiencies to LTC-IC of 40% were achieved and this was increased to 60% by short-term preselection in G418. Southern analysis of DNA from the nonadherent cells produced by these LTC over a 6-wk period provided evidence of clonal evolution of LTC-IC in vitro. Leukemic chronic myelogenous leukemia LTC-IC were also infected at high efficiency using the same supernatant infection strategy with growth factor supplementation. These data demonstrate the feasibility of using cell-free virus preparations for infecting clinical marrow samples suitable for transplantation, as well as for further analysis of human marrow stem cell dynamics in vitro.

Zidovudine use in pregnancy: a report on 104 cases and the occurrence of birth defects.

As more women of childbearing age are being identified as HIV infected, vertical transmission to the fetus and/or neonate is an increasingly significant therapeutic problem. Currently the use of zidovudine is one of the few specific measures available, and as a potentially teratogenic and fetotoxic agent, any decision for its use requires evaluation of the potential for fetal damage. In a series of 104 cases of intentional or inadvertent use of zidovudine at differing gestations in pregnancy, there were eight spontaneous first trimester abortions, eight therapeutic terminations, and eight cases of fetal abnormality occurring among a total of 88 cases where the pregnancy progressed. Analysis and correlation of antenatal data and drug therapy with individual cases failed to show any specific abnormality that could reasonably be attributed to zidovudine therapy. While not proving safety, these data add to previous smaller series with similar findings, thus lending tenuous support to the use of this agent. Continuing studies are required, particularly to clarify the possibility of long-term developmental defects.

Screening for gestational diabetes in a multi-ethnic population.

A multi-ethnic population was screened for gestational diabetes mellitus (GDM) using a venous plasma glucose estimation, 1 h following a standard 50 g glucose load. A significant difference in the ethnic distribution of screen-positivity was found. Amongst the screen-positive group the odds ratio (OR) for special care baby facility (SCBU) admission and birthweight > 3999 g were both increased (OR = 1.87 and 1.99). Only limited diagnostic testing by a glucose tolerance test (GTT) could be achieved for the screen-positive population. For patients with confirmed GDM (two or more abnormal values on a GTT) the OR for SCBU admission was further increased to 5.1, while the OR for increased birthweight was only 1.34. Clinical attention should be directed towards outcome assessment in order to properly evaluate the nature of and place for screening for GDM in multi-ethnic populations.

Gestational diabetes screening of a multiethnic, high-risk population using glycated proteins.

In populations with a high incidence of gestational diabetes (GDM), any form of oral glucose testing for screening or diagnosis excessively strains the health care system. We investigated the value of glycated proteins as potential screening tests in 430 pregnant women, i.e. protein corrected fructosamine (cFRUC) and hemoglobin A1c (HbA(1c)) both alone and in combination for a GDM diagnosis confirmed by the 'gold standard' 100-g oral glucose tolerance test (OGTT). Two cut-off values were used for each test, the upper to rule in and the lower to rule out GDM. At the lower cut-off values for cFRUC of 210 micromol/l and HbA(1c) of 5%, the sensitivities achieved were 92.2 and 92.1% while the negative predictive values were 88.9 and 86.9%, respectively. The upper cut-off values did not achieve acceptable positive predictive values to be useful for ruling in GDM. Screening of our multiethnic, high-risk pregnant population with a combination of cFRUC and HbA(1c) on a single fasting sample would have avoided the cumbersome OGTT (by ruling out GDM) in 37.9% women with only a 3.9% misclassification rate. This potentially simpler approach, though not universally applicable, would be clinically useful and more acceptable to patients in selected high-risk populations.

Seroprevalence and mother-to-infant transmission of hepatitis C in asymptomatic Egyptian women.

OBJECTIVES: In this study, we sought to determine (1) the prevalence of hepatitis C virus (HCV) RNA, or its antibodies, in a healthy parturient Egyptian population and (2) the risk of mother-to-infant transmission in this population. METHOD: The serum of 499 pregnant Egyptian women was tested for anti-HCV with ELISA-3 and for HCV RNA by polymerase chain reaction (PCR). Neonatal cord blood and infant blood were similarly tested for anti-HCV and HCV RNA. RESULTS: Recombinant immunoblot assay (RIBA) detected anti-HCV in 65/499 (13%) women; of these, 20/65 (31%) were PCR-positive. The total number of babies born was 499. Of the original group, 97 mothers and infants (HCV-negative) were lost to follow up and were excluded. Sixty-five anti-HCV-positive infants were born vaginally to the 65 anti-HCV-positive mothers, of which twenty (31%) corresponding mothers and babies were also positive for HCV RNA. Of these twenty babies, three died of hepatocellular disease by six months of age; sixteen developed chronic liver disease; the remaining nine remained asymptomatic but were serologically and PCR-positive. The mother-to-infant transmission rate was significantly increased (5%; P < 0.0001). Of the seropositive children, 45/65 (69%; P < 0.0001) seroreverted by eighteen months of age. CONCLUSION: There is a high prevalence of anti-HCV in healthy pregnant Egyptian women and vertical transmission is a major risk for chronic HCV carriers.

PIP: Hepatitis C virus (HCV) seropositivity among Egyptian women has been estimated at 13-22%. This study investigated the magnitude of risk of vertical HCV transmission in 499 consecutive, apparently healthy Egyptian women attending antenatal clinics at United Arab Emirates university teaching hospitals in 1994-96. 65 maternal serum samples (13%) were positive for HCV by confirmatory recombinant immunoblot assay and the infants of all these women were anti-HCV positive. HCV RNA was detected by polymerase chain reaction in 20 (31%) of these maternal-infant pairs. Of the 20 infants who were HCV RNA-positive at birth, 4 died of hepatocellular disease before 18 months of age and the remaining 16 developed chronic liver disease. 45 (69%) of the 65 infants seropositive at birth seroconverted over the course of the 18-month follow-up period (mean age of seroconversion, 15 months). These findings indicate that high levels of HCV viremia, like enhanced levels of HIV viremia, facilitate maternal-infant virus transmission. The presence of HCV RNA in the cord blood of the 3 infants who died in the first 3 months of life suggests transplacental pathogenesis in a small proportion of cases. Acquisition at delivery remains the predominant mode of HCV transmission, however, and can be reduced by appropriate screening and counseling of pregnant seropositive chronic carriers of disease.

Dynamic and steady state response of heart rate to orthostatic stress in normotensive and hypertensive pregnant women.

We determined the dynamic and steady state responses of heart rate (HR) to orthostatic stress (standing up) in normotensive and hypertensive pregnant women. Using a continuous recording with servo-photosphygmography, HR response to change in posture from left lateral recumbent position to standing was analysed. The subjects were divided into five groups comprising: Groups I, II and III: normotensive pregnant women in each of the three trimesters of pregnancy (total n = 77); Group IV: women with gestational proteinuric hypertension (GPH) in the third trimester (n = 16); Group V: age-matched non-pregnant normotensive controls (n = 15). The HR reacted with a typical overshoot response to this orthostatic change with HR rising to a peak and then settling to a new but higher steady state. Change in steady state HR from lying to standing (delta HR), rate of rise of HR in response to standing (i.e. the acceleration slope (HRon)), and rate of fall of HR after reaching the peak (i.e. deceleration slope (HRoff)) were evaluated from standing heart rate time curves. HRon in response to standing showed a downward trend with gestation (ANOVA, P < 0.05) in normotensive gravida. The deceleration slope (HRoff) showed a distinct gestational age-related decrease from first to third trimester in normotensive women (ANOVA, P < 0.01). The most striking observation was that the slope of HRoff for the GPH group was significantly steeper than that of normotensive women of comparable gestational age (unpaired t-test P < 0.01) and approximated to that of the non-pregnant group. The difference in HR response between normotensive women and those with GPH in the third trimester suggests it may have potential as a new marker for pre-eclampsia.

Grandmultiparity--not to be feared? An analysis of grandmultiparous women receiving modern antenatal care.

OBJECTIVE: To document the reproductive performance of grandmultiparous women receiving modern antenatal care. METHODS: A cross-sectional study of 2784 multiparous (882 grandmultiparous) mothers delivered in a base hospital obstetric unit staffed by western-trained midwives and consultant obstetrical staff. RESULTS: There were minimal differences in major antenatal, peripartum and neonatal outcome events, with the exception of a high rate of gestational and pre-existing diabetes. CONCLUSIONS: This data supports the opinion that grandmultiparity per se is not necessarily a major risk factor for either mother or fetus. Similarly, the mature grandmultigravida in this population was not at significantly increased risk of the alleged associations of increased parity and advancing maternal age, with the exception of diabetes. Together with the combined prevalence of maternal anemia this requires further investigation and probable intervention, particularly in the light of recent speculation concerning the fetal and infant origins of adult disease.

Birthweight/gestational age charts revisited.

Intra-uterine growth charts are widely used, but the data from which they are generated are imprecise and the (mis)use made of them is in danger of acceptance through repetition. To an obstetrician they have little use and the description 'intra-uterine growth chart' is misleading. This traditional analysis is reported to emphasize that a new approach to judging the normality or otherwise of intra-uterine development is needed, particularly to aid the clinician involved in management of the high risk pregnancy and preterm labour and delivery.

Pregnancy outcome data in a United Arab Emirates population: what can they tell us?

A cross-sectional study of pregnancy outcome in a national Arabic population is presented to highlight a unique reproductive environment compared with many other obstetric populations. A high pro-natal intensity is evident with significant numbers of grandmultipara (36.7%) and mature gravida (22.2%). These groups did not show any significant increase in adverse antenatal or intrapartum events apart from an increasing rate of gestational diabetes with maternal age. There was an average induction/augmentation rate of 10.2% with a low rate of instrumental delivery (2.3%) and a mean caesarean section rate of 6.9%. A major antenatal complication noted was the prevalance of 17.3% for a haemoglobin < 10 g/dl together with overall increased rates of pre-existing and gestational diabetes. In addition the incidence of preterm labour increased over each of the 3 years studied and is of concern.

Birth weight distribution in the United Arab Emirates.

Birth weight is a point measure of size, not a measure of growth and past presentation of birth weight data has confounded its use by maintaining an emphasis on size at birth rather than health at birth. This paper presents current data for a United Arab Emirates (UAE) national population for both international comparison and contrast with other Gulf populations. Unique local conditions include a high rate of diabetes in pregnancy, extreme grandmultiparity and significant numbers of mature gravida (age > 35 yrs), and emphasizes a need for a re-appraisal and redevelopment of this form of analysis as a part of the overall obstetric care of the pregnant population.

Isolation of a specific enhancer for echovirus 6 from uninfected permissive host cells.

A specific enhancer for the m+ variant of echovirus 6 was isolated from uninfected, permissive host cells. The enhancer transferred the susceptibility to virus infection from permissive cells to less susceptible cells. Enhancer activity in cultured, human cells (WISH) was released by these cells into extracellular fluids at a linear rate. Maximal enhancer activity was recovered from cell monolayers that were extracted with buffered salt solutions (pH 6.6) for 5 h at 37 C. Crude enhancer preparations contained substances that reduced virus titers in permissive host cells and suppressed some of the enhancer activity. Viral inhibitory activity was removed from cellular extracts by either differential centrifugation or ion exchange chromatography. The enhancer, in contrast to inhibitory substances, remained in supernatant fractions after centrifugation at 110,000 x g for 4 h and adsorbed to diethylaminoethyl-Sephadex columns. Enhancer preparations that were devoid of inhibitory activity increased the virus titers in less susceptible simian cell cultures (approximately 100-fold) to the titers attained by fully permissive human cell cultures. A direct relationship between protein concentration and enhancer activity in cellular extracts was demonstrated. Partial purification of the enhancer by differential centrifugation and ammonium sulfate precipitation increased the specific activity of enhancer (units/mg of protein) by 13-fold.

Psychological well-being as an outcome variable in the treatment of infertility by Clomiphine.

A sample of twenty-seven patients who had undergone extensive investigation and subsequent treatment for infertility were asked to complete a questionnaire constructed from items demonstrated to be valid components of the subjective states of happiness and well-being. With two exceptions, whose current subjective state seemed likely to have been adversely affected by serious current life-events, the respondents seemed equally happy whether or not their treatment had been successful. Adjustments to the infertility state are briefly discussed.

Toxic shock syndrome presenting to a paediatric service.

A case of toxic shock syndrome occurring in a 13-year-old and presenting to a paediatric service is described. Some implications are discussed including an approach to menstrual protection and the question of future contraceptive needs.

Vaginal delivery in the grand multipara following previous lower segment cesarian section.

OBJECTIVE: To examine the effect of attempting vaginal birth after cesarian section in the grand multipara with one previous cesarian section scar in the uterus. METHOD: Over 5-year period (1990-1994) mothers with 6 or more previous deliveries and with a previous section scar in the uterus were identified. The outcome in these patients who attempted vaginal birth was reviewed. RESULTS: Of the 85 patients with the combination of both grand multiparity and a previous cesarian section scar in the uterus, 45 attempted a trial of labor. Twenty-seven patients (60%) achieved successful uncomplicated vaginal delivery. There was a relatively high incidence of serious complications. CONCLUSION: Vaginal birth after cesarian section can be achieved in some grand multiparas with a previous scar in the uterus. There is an increased risk of serious complications. The labor should be very closely supervised and early intervention arranged if there is not smooth rapid progress.

An evaluation of fructosamine estimation in screening for gestational diabetes mellitus.

Although persuasive arguments against routine screening for gestational diabetes mellitus (GDM) have been made, it is widely but not universally performed as a part of antenatal care. There is no international agreement on methods or criteria used for screening (or for diagnosis), and administered glucose-load methods have significant practical difficulties in a busy antenatal clinic setting. However, recent evidence supports the concept of an increased level of importance being given to a diagnosis of GDM, with interest in the fetal and neonatal origins of adult disease being added to the short-term obstetric and fetal concern during pregnancy. A second generation fructosamine test, corrected for total protein, has been evaluated as a practical alternative to glucose screening for GDM in a busy, multi-ethnic antenatal clinic. This achieved a 79.4% sensitivity and a 77.3% specificity for a diagnosis of GDM confirmed by a glucose tolerance test using Carpenter's modified criteria. In view of the organizational simplicity of this sample/test requirement, a wider evaluation is suggested together with a re-evaluation of clinical outcome criteria rather than blood glucose levels alone.

Gestational diabetes and fetal macrosomia in a multi-ethnic population.

We set out to examine maternal and neonatal factors surrounding increased birthweight in a multi-ethnic population having an increased prevalence of diabetes mellitus. Additionally, to document the difference (if any) for such neonates in rates of obstetrical operative intervention at delivery where a specific diagnosis of maternal gestational diabetes mellitus had been made. This was an observational study of unselected mothers giving birth to a neonate of 4000 g or more. Data for this population concerning the results of antenatal screening and diagnostic testing for gestational diabetes mellitus were available as a subset of a larger independent and ongoing database. Odds ratios were used to compare group attributes subset on ethnic, diabetic screening and diabetic diagnostic status. Two ethnic groups showed an increased odds ratio for increased birthweight. A diagnosis of diabetes was associated with a twofold increase in caesarean section rate, and a significant increase in median birthweight when compared with screen positive/ diagnostic negative mothers. A total of 70.4% of mothers were overweight or obese while neonatal ponderal index showed a dependence on birthweight ( r2 = 0.17). We conclude that ethnic status is an important factor in assessing fetal size, as is maternal body mass index. A diagnosis of diabetes mellitus confers an increased risk of operative intervention at delivery. Our approach to the use of birthweight data requires re-assessment.

Fasting plasma glucose as a screening test for gestational diabetes in a multi-ethnic, high-risk population.

AIMS: Screening every pregnant woman for gestational diabetes mellitus (GDM), as widely recommended for high-incidence populations, strains the healthcare system excessively. This study investigated the value of fasting plasma glucose (FPG) as an alternative to the more cumbersome oral glucose tolerance test (OGTT). METHODS: One thousand six hundred and forty-four pregnant women in a multi-ethnic, high-risk population were evaluated by the FPG as a screening test among two principal subgroups, i.e. women (n = 1276) at risk for GDM on clinical grounds and those women (n = 368) with a positive post 50-g, 1-h plasma glucose challenge test (GCT). Two threshold values for FPG 'ruled in or ruled out' a GDM diagnosis, which was confirmed by the 3-h, 100-g OGTT, using Carpenter's modified criteria as the 'gold standard'. RESULTS: In the women with a positive clinical history, at an optimal cut-off value of FPG < 4.4 mmol/l to rule out GDM; a sensitivity of 94.7% was achieved, 21 (1.6%) women being false negatives. Using a FPG > or = 5.3 mmol/l to rule in GDM; the specificity was 94.0% with 53 (4.2%) women being classified as false positives. FPG would have eliminated need for the OGTT in 50.9% pregnant women (misclassification rate 5.8%). In the positive GCT group, using similar cut-offs for FPG, a sensitivity of 96.6% and specificity of 90.8% was achieved with a potential to avoid 51.6% OGTTs (misclassification rate 7.3%). The positive predictive value of the GCT was 31.8% compared to 80.2% for FPG at 5.3 mmol/l. CONCLUSIONS: While previously neglected as a screening test for GDM, in selected high-risk populations the FPG offers a potentially simple, practical algorithm to screen for GDM by being cost-effective and patient friendly. A wider application should be explored.