Transfusion for sickle cell disease in pregnancy: a single-centre survey.

INTRODUCTION: Sickle cell disease in pregnancy carries a high risk of maternal and fetal adverse outcomes. The use of prophylactic transfusions to reduce the risk of sickle complications is controversial. Current UK standards do not recommend the routine use of transfusion for sickle pregnancy. We examined transfusion episodes during sickle pregnancies in a single centre over an 11-year period. METHODS: We conducted a retrospective observational study of all pregnancies in patients with sickle cell disease who attended the joint obstetric/haematology clinic over an 11-year period. All pregnancies were managed according to a local protocol, which did not recommend routine transfusion. RESULTS: A total of 38 pregnancies (HbSS 22, HbSC 13, Hb S/beta thalassaemia 3) were included, with a mean age at booking of 29 years. A total of 61% of pregnancies required on-demand or emergency transfusion during the course of pregnancy or post-partum. Women requiring a transfusion during pregnancy had a higher mean number of hospital admissions in the previous year (1·11 vs 0·15, P = 0·057), a significantly lower mean steady-state haemoglobin (85·0 vs 99·6 g L-1 , P = 0·003) and a significantly lower mean haemoglobin at the pregnancy booking visit. (86·1 vs 99·5 g L-1 , P = 0·02). CONCLUSION: In sickle pregnancies assigned to standard management in a single centre, a high proportion of women required on-demand transfusion. Possible pre-pregnancy factors predictive of a need for transfusion include lower baseline haemoglobin and number of hospital admissions in the previous 12 months.

A critique of current practice of transvaginal pudendal nerve blocks: a prospective audit of understanding and clinical practice.

Pudendal nerve blocks are a pre-requisite to forceps delivery without regional anaesthesia. Their efficacy is dependent on introducing local anaesthetic in close proximity to the pudendal nerve and allowing sufficient time for its onset of action. An audit of 57 obstetricians evaluated their clinical technique against standards using both a questionnaire and adapted model pelvis. The majority of participants were unable to describe correctly the point of infiltration and were unaware of the lag time required to effect adequate analgesia. We identify a deficiency in training and describe a method by which training can be facilitated and assessed.

Identification of early predictive imaging biomarkers and their relationship to serological angiogenic markers in patients with ovarian cancer with residual disease following cytotoxic therapy.

BACKGROUND: Patients with recurrent ovarian cancer often achieve partial response following chemotherapy, resulting in persistent small volume disease. After completion of treatment, the dilemma of when to initiate subsequent chemotherapy arises. Identification of biomarkers that could be used to predict when subsequent treatment is needed would be of significant benefit. DESIGN: Twenty-three patients with advanced ovarian cancer and residual asymptomatic disease following chemotherapy underwent dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) at study entry, 4, 8, 12, 18 and 26 weeks or disease progression. A subgroup of patients provided plasma samples within which a panel of angiogenic biomarkers was quantified. RESULTS: By 4 weeks, significant differences in whole tumour volume, enhancing fraction and Ca125 were observed between patients whose disease progressed by 26 weeks and those who remained stable. Significant correlations between plasma soluble vascular endothelial growth factor receptor-1 (sVEGFR-1) and sVEGFR-2 concentrations, and blood volume and tumour endothelial permeability surface area product measured by DCE-MRI were observed. CONCLUSIONS: Imaging markers have a potential role in early prediction of disease progression in patients with residual ovarian cancer and may supplement current measures of progression. The correlation of DCE-MRI and serological biomarkers suggests that tumour angiogenesis affects these markers through common biological means and warrants further investigation.

Weekly paclitaxel in the treatment of recurrent ovarian carcinoma.

PURPOSE: To study the effect of weekly paclitaxel in the treatment of recurrent ovarian and peritoneal carcinoma. METHODS: A retrospective analysis of patients treated at Christie Cancer Centre between May 2003 and May 2005 was carried out. RESULTS: Fortynine patients with recurrent ovarian and peritoneal carcinoma were treated. The mean duration of treatment was 11 weeks, with 27 (54%) patients receiving 12 or more treatments. The most frequent non-haematological toxicities reported were mild nausea, constipation, lethargy and neuropathy. Moderate anaemia was noted in 50% of patients. Radiological assessment by CT scanning showed that complete or partial responses were achieved in 28% of patients. CA125 response was demonstrated in 63% of patients. Median time to recurrence was 149 days and median survival was 359 days. CONCLUSION: This study provides evidence for the role of weekly paclitaxel in the treatment of recurrent ovarian and peritoneal carcinoma even in a drug-resistant setting following multiple lines of prior therapy.

Facilitation of Vaginal Delivery in an Infant with Complete Heart Block Secondary to Maternal Anti-Ro Antibodies.

Congenital heart block (CHB) is a rare disorder that may be associated with a high morbidity and even mortality, with a risk of death both in utero and during infancy. Women with serum titres of anti-Ro and/or anti-La antibodies carry a risk of CHB of 1-5% in their offspring, with a recurrence risk of approximately 20%. We present a case of a 36-year-old female with a pregnancy complicated by congenital heart block. Autoimmune profiling at booking showed she was positive for lupus anticoagulant and anti-Ro antibodies. A fetal echocardiogram at 21 + 3 showed complete heart block. She was monitored throughout the remainder of her pregnancy with serial growth scans, cardiovascular profiling, and BPP scoring. She had a normal vaginal delivery at term to a female infant.

The interval from surgery to chemotherapy in the treatment of advanced epithelial ovarian carcinoma.

BACKGROUND: To study the effect of the interval between surgery and the start of chemotherapy in the treatment of patients with advanced ovarian cancer. METHODS: We stratified patients according to the start of platinum-based chemotherapy in group 1 (within 4 weeks from surgery), group 2 (between 4 and 8 weeks) and group 3 (between 8 and 12 weeks). RESULTS: Three hundred and ninty-four stage III ovarian cancer patients were analysed. In the multivariate analysis there were no differences in survival according to the interval between surgery and chemotherapy among the three groups. The independent prognostic variables were type of procedure (p = 0.014), performance status (p = 0.040) and post-chemotherapy CA-125 (p < 0.0001). CONCLUSIONS: The interval between surgery and chemotherapy does not affect outcome.

DNA-dependent protein kinase is not required for accumulation of p53 or cell cycle arrest after DNA damage.

In response to DNA damage, cells transduce a signal that leads to accumulation and activation of p53 protein, transcriptional induction of several genes, including p21, gadd45, and gadd153, and cell cycle arrest. One hypothesis is that the signal is mediated by DNA-dependent protein kinase (DNA-PK), which consists of a catalytic subunit (DNA-PKcs) and a regulatory subunit (Ku). DNA-PK has several characteristics that support this hypothesis: Ku binds to DNA damaged by nicks or double-strand breaks, DNA-PKcs is activated when Ku binds to DNA, DNA-PK will phosphorylate p53 and other cell cycle regulatory proteins in vitro, and DNA-PKcs shares homology with ATM, which is mutated in ataxia telangiectasia and involved in signaling the p53 response to ionizing radiation. The hypothesis was tested by analyzing early passage fibroblasts from severe combined immunodeficient mice, which are deficient in DNA-PK. After exposure to ionizing radiation, UV radiation, or methyl methane-sulfonate, severe combined immunodeficient and wild-type cells were indistinguishable in their response. The accumulation of p53, induction of p21, gadd45, and gadd153, and arrest of the cell cycle in G1 and G2 occurred normally. Therefore, DNA-PK is not required for the p53 response or cell cycle arrest after DNA damage.

Chromosomal instability is correlated with telomere erosion and inactivation of G2 checkpoint function in human fibroblasts expressing human papillomavirus type 16 E6 oncoprotein.

Cell cycle checkpoints and tumor suppressor gene functions appear to be required for the maintenance of a stable genome in proliferating cells. In this study chromosomal destabilization was monitored in relation to telomere structure, lifespan control and G2 checkpoint function. Replicative senescence was inactivated in secondary cultures of human skin fibroblasts by expressing the human papillomavirus type 16 (HPV-16) E6 oncoprotein to inactivate p53. Chromosome aberrations were enumerated during in vitro aging of isogenic control (F5neo) and HPV-16E6-expressing (F5E6) fibroblasts. We found that structural and numerical aberrations in chromosomes were significantly increased in F5E6 cells during aging in vitro and fluorescence in situ hybridization (FISH) analysis using chromosome-specific probes demonstrated the occurrence of rearrangements involving chromosome 4 and 6 in genetically unstable F5E6 cells. Flow cytometry and karyotypic analyses revealed increased polyploidy and aneuploidy in F5E6 cells only at passages > 16, although these cells displayed defective mitotic spindle checkpoint function associated with inactivation of p53 at passages 5 and 16. G2 checkpoint function was confirmed to be gradually but progressively inactivated during in vitro aging of E6-expressing cells. Aging of F5neo fibroblasts was documented during in vitro passaging by induction of a senescence-associated marker, pH 6.0 lysosomal beta-galactosidase. F5E6 cells displayed extension of in vitro lifespan and did not induce beta-galactosidase at high passage. Erosion of telomeres during in vitro aging of telomerase-negative F5neo cells was demonstrated by Southern hybridization and by quantitative FISH analysis on an individual cell level. Telomeric signals diminished continuously as F5neo cells aged in vitro being reduced by 80% near the time of replicative senescence. Telomeric signals detected by FISH also decreased continuously during aging of telomerase-negative F5E6 cells, but telomeres appeared to be stabilized at passage 34 when telomerase was expressed. Chromosomal instability in E6-expressing cells was correlated (P < 0.05) with both loss of telomeric signals and inactivation of G2 checkpoint function. The results suggest that chromosomal stability depends upon a complex interaction among the systems of telomere length maintenance and cell cycle checkpoints.

Plasmacytoma-refractory BALB/cAnPt mice have naive T cell and highly specific B cell responses to antigen.

Recently, a reduction in the incidence of pristane-induced plasmacytomas in BALB/cAnPt (BALB/c) mice that were kept in viral specific pathogen-free (SPF) conditions has been reported. Environmentally, these SPF-BALB/c mice differed from conventionally-housed (CON) mice only in viral exposure and diet (i.e. sterilization of mouse chow), since microbial colonization of the intestinal tract was seen to be equivalent. This report assessed the ability of SPF- and CON-BALB/c mice to respond to immunologic challenge with soluble antigen, i.e. hen egg white lyzosyme (HEL), as a means of evaluating differences in T and B cell function and, indirectly, evaluating the possible effects these differences might have on plasmacytoma development. When cultured in vitro for 5 days with HEL, HEL-primed lymph node cells (LNC) from SPF-BALB/c mice proliferated to a significantly lesser extent than HEL-primed CON-BALB/c LNC. Moreover, HEL-induced production of IFN-gamma and IL-5 was significantly lower in SPF LNC. Serum IgG1 levels were 10-fold lower in SPF-BALB/c mice with, or without prior immunization with HEL and were not reconstituted by repeated injections of HEL in adjuvant. Serum IgM levels of SPF- and CON-BALB/c mice were equivalent. This reduction in immune responses could not be attributed to a lack of colonization of secondary lymphoid organs, since flow cytometric analysis of LNC revealed no difference in the number of recoverable cells and the proportion of lymphocyte subsets (CD4+, CD8+ and CD45+ cells) obtained from SPF- and CON-BALB/c mice. However, only CON LNC were induced to increase surface expression of CD44 after antigenic or mitogenic stimulation in vitro. Antibody responsiveness to HEL, as evidenced by serum anti-HEL binding or splenic hybridoma studies, demonstrated higher levels of IgG1 antibodies in CON BALB/c mice than in SPF mice. However, a greater proportion of the SPF IgG1 antibodies present were specifically directed against HEL, so that specific activity was greater in SPF-BALB/c mice. Therefore, while SPF BALB/c mice have a more restricted response to HEL than CON-BALB/c mice, those antibodies that are produced are more specifically directed against HEL with very little apparent bystander/polyclonal activation of multireactive cells. Resistance to plasmacytomas in SPF-BALB/c mice, therefore, may stem from a reduced number of circulating memory T and B cells, which are capable of reacting and/or crossreacting with a chronic inflammatory stimulus.

Variations in the apparent pH set point for activation of platelet Na-H antiport.

To explore the role of the Na-H antiport in essential hypertension, we studied the kinetics of cytosolic pH and external sodium activation of this transport system in platelets from 65 normotensive and essential hypertensive subjects on and off antihypertensive medications. Subjects included both blacks and whites, as well as men and women. The fluorescent dye 2'7-bis(carboxyethyl)-5,6-carboxyfluorescein was used to monitor the cytosolic pH in these cells. Platelets from black (hypertensive and normotensive) men and hypertensive white men demonstrated a highly significant alkaline shift in the apparent cytosolic pH set point for activation of the Na-H antiport. For the hypertensive subgroups, the cytosolic pH set point values (mean +/- SEM) were: white men, 7.45 +/- 0.052; white women, 7.04 +/- 0.089; black men, 7.66 +/- 0.148; and black women, 7.20 +/- 0.082. For the normotensive subgroups, the cytosolic pH set point values were: white men, 7.13 +/- 0.034; white women, 7.05 +/- 0.036; black men, 7.50 +/- 0.110; and black women, 7.20 +/- 0.176 (p = 0.0016 for race and p = 0.0001 for gender, using a three-way analysis of variance by race, gender, and hypertension). There were no race-, gender-, or blood pressure-related differences among the various cohorts in the kinetics of sodium activation of the Na-H antiport, the cellular buffering power, and basal pH. These results suggest that at basal pH the Na-H antiport is quiescent in platelets from both black and white women and normotensive white men.(ABSTRACT TRUNCATED AT 250 WORDS)

Blood pressure inversely correlates with thrombin-evoked calcium rise in platelets.

Earlier investigations showed a positive correlation between basal cytosolic free calcium in human platelets and blood pressure; however, recent studies have failed to show this relation. We undertook the present work to examine which platelet cytosolic calcium parameters (namely, cytosolic calcium in resting or stimulated states in calcium-containing and calcium-free media) present the least variability and best correlation with blood pressure. We studied 17 healthy white men on three different occasions separated by 1- and 4-week intervals. Their manual and ambulatory automated 24-hour blood pressure measurements were correlated with cytosolic calcium in resting and stimulated (thrombin-treated) fura 2-loaded platelets. The following cytosolic calcium parameters were measured in 1 mmol/L calcium and calcium-free media: basal cytosolic calcium, peak thrombin-evoked cytosolic calcium, and post-transient cytosolic calcium 5 minutes after thrombin treatment. The highest and lowest coefficients of variation were respectively shown by the basal cytosolic calcium (22.8%) and peak thrombin-evoked cytosolic calcium (10.1%) in calcium medium. Basal cytosolic calcium did not correlate with any of the blood pressure parameters. Of the cytosolic calcium parameters, peak thrombin-evoked cytosolic calcium in calcium medium demonstrated consistent (negative) correlations with blood pressure, with better correlations shown with diastolic than systolic blood pressure of both automated and manual blood pressure readings. Peak thrombin-evoked cytosolic calcium in calcium medium showed similar correlations with nighttime and daytime automated blood pressure measurements. There were no correlations between peak thrombin-evoked cytosolic calcium in calcium-free medium and blood pressure.(ABSTRACT TRUNCATED AT 250 WORDS)

The ABL-MYC retrovirus generates antigen-specific plasmacytomas by in vitro infection of activated B lymphocytes from spleen and other murine lymphoid organs.

ABL-MYC is a recombinant retrovirus that constitutively expresses the v-abl and c-myc oncogenes. When used to infect immunized mice this virus rapidly and efficiently induces plasmacytomas of which an unusually high percentage secrete antigen (Ag)-specific monoclonal antibodies. These findings suggested that ABL-MYC targets Ag-stimulated B cells for transformation and that infection of lymphoid cells in vitro might be a useful, alternative method for generating monoclonal, Ag-specific plasmacytomas (ASPCTs). Therefore, we used helper virus-free ABL-MYC to infect suspensions of cells from spleens and other lymphoid organs from mice that had been immunized with a variety of Ags and transplanted them into naive mice. The results show that ABL-MYC preferentially transforms splenocytes that are Ag-reactive. They also demonstrate that ASPCTs can be produced by in vitro infection of cell suspensions from the spleen, lymph nodes and Peyer's patches of mice that had been immunized intraperitoneally with sheep red blood cells, Escherichia coli core RNA polymerase or Epstein-Barr virus gp340 protein or immunized orally with live Giardia lamblia parasites. The ASPCTs usually consisted of one to three colnes, secreted antibodies that were quantitatively and qualitatively similar to those obtained from hybridomas, and could continue to secrete Ag-reactive antibody over eight transplant generations.

A method for transition smoothing of sleep--waking states.

A method was developed for sequential redistribution of short-duration sleep--waking states into the long-duration behavioral state(s) that surrounds these intervals. This methodological approach is useful in determination of general sleep--waking patterns such as state durations, interstate intervals, and sleep--waking cycles. The redistribution sequences had no effect on total amount of sleep or waking categories, but did decrease stage 1 with successive redistribution steps. Examination of the slope of the curve (number of occurrences or mean epoch duration divided by successive redistributions) can be used to determine the appropriate "smoothing" for electroencephalogram (EEG) state data via location of the breakpoint in the curve. The low-pass redistribution technique has been particularly useful for smoothing large volumes of EEG state data for subsequent analyses without significantly changing the various sleep-waking states.

Effects of morphine alone and in combination with naloxone or d-amphetamine on shock-maintained behavior in the squirrel monkey.

Key-pressing behavior in the squirrel monkey was maintained under an 8-min fixed-interval (FI) schedule of electric-shock delivery. The acute i.m. administration of morphine prior to a daily session decreased response rates at doses of 1.0--3.0 mg/kg but had little systematic effect on rate at doses of 0.03-0.3 mg/kg. When naloxone was administered concomitantly with morphine prior to a session, 0.01 mg/kg naloxone required a three-fold increase in the dose of morphine necessary to obtain decreased response rates, 0.1 mg/kg naloxone required a 30-fold increase in morphine, and 1.0 mg/kg required more than a 30-fold increase in morphine. Moreover, the administration of naloxone with morphine resulted in increased rates of responding at certain combinations of doses of the two drugs. The administration of d-amphetamine (0.03 or 0.1 mg/kg) alone increased mean response rates under the FI schedule; when combined with 0.03-0.3 mg/kg morphine the increases in responding were greater than obtained with d-amphetamine alone. The negative slope of the linear regression lines relating the effects of morphine to control rates of responding engendered under the FI schedule was decreased when morphine was combined with naloxone, but not with d-amphetamine. These results show that naloxone, but not d-amphetamine, can antagonize the response-rate decreasing effect of morphine when responding in the squirrel monkey is maintained by response-produced electric shock.

Serotonergic modulation of the discriminative-stimulus effects of cocaine in squirrel monkeys.

In order to investigate the potential modulatory role of serotonin on the discriminative-stimulus effects of cocaine, two groups of squirrel monkeys were trained to discriminate 0.3 mg/kg or 1.0 mg/kg cocaine and saline under a two-lever drug-discrimination procedure. Substitution of a range of cocaine doses (0.03-1.7 mg/kg) occasioned orderly, dose-dependent increases in cocaine-lever responding. When administered alone, the non-selective serotonin direct agonist, quipazine, also occasioned increases in cocaine-lever responding which were more pronounced in subjects trained with the lower cocaine dose. When quipazine was administered in combination with cocaine, there was an increase in cocaine-lever responding, indicating an additive effect. The serotonin uptake inhibitor, fluoxetine, occasioned saline-lever responding when administered alone. However, in combination with cocaine, fluoxetine enhanced the discriminative effects of cocaine in subjects trained at the lower cocaine dose. The 5-HT2-selective antagonists, ketanserin and ritanserin, did not occasion cocaine-lever responding when administered alone. In combination with cocaine, ketanserin attenuated the discriminative effects of cocaine in most subjects, and ritanserin attenuated the discriminative effects of cocaine in subjects trained at the higher dose. These results indicate that the discriminative-stimulus effects of cocaine may be increased by direct- and indirect-acting serotonin agonists and attenuated by serotonin antagonists in squirrel monkeys.

Pharmacological interactions between serotonin and dopamine on behavior in the squirrel monkey.

The behavioral effects of GBR 12909, a selective dopamine uptake inhibitor, were determined in squirrel monkeys trained to respond under a fixed-interval (FI) schedule of stimulus termination and a second-order schedule of IV drug self-administration. Intermediate doses of GBR 12909 increased FI response rate markedly, and the highest dose decreased response rate below control values. The 5HT uptake inhibitors, alaproclate and fluoxetine, and the 5HT agonist, quipazine, attenuated the behavioral-stimulant effects of GBR 12909, whereas the 5HT2A/2C antagonist, ritanserin, enhanced the behavioral-stimulant effects of the lowest dose. GBR 12909 reliably maintained self-administration, and ritanserin increased response rate maintained by the highest dose. The dopamine agonist, quinpirole, increased FI response rate in only one of three subjects, and ritanserin enhanced the behavioral-stimulant effects of quinpirole in that subject. The dopamine agonist, apomorphine, only decreased FI response rate, and ritanserin did not alter its behavioral effects. The pharmacological profile of GBR 12909 administered alone and in combination with selective 5HT drugs in the present study was similar to that obtained previously with cocaine, further demonstrating that 5HT can reliably modulate the behavioral effects of psychomotor stimulants with prominent dopaminergic actions.

Behavioral evaluation of hemiparkinsonian MPTP monkeys following dopamine pharmacological manipulation and adrenal co-graft transplantation.

Bradykinesia and rigidity are the symptoms that most directly correlate with loss of striatal dopamine in Parkinson's disease. In the hemiparkinsonian (HP) monkey, this is represented by paucity of movement as measured by coli puterized movement analysis, diminished manual dexterity on clinical examination, and diminished performance on operant behavioral tasks. The present study used an MPTP-induced HP model in rhesus monkeys to evaluate the effectiveness of adrenal medullary and peripheral nerve co-grafts in diminishing parkinsonian symptoms. Unoperated controls (N = 4), surgical controls with caudate lesioning (N = 4), and caudate co-grafted (N = 4) HP monkeys demonstrated diminished movement in the home cage following MPTP. This behavior persisted in unoperated controls, but improved in both surgical control and co-grafted monkeys. Functional hand dexterity evaluations demonstrated similar impairment in all three groups but only surgical controls and co-grafted monkeys demonstrated improvement. In general, rotational behavior in response to apomorphine was consistent with recovery of function in surgical controls and co grafted monkeys, but marked between-subject variability precluded group statistical analyses. None of the monkeys could perform the operant task using the affected limb following MPTP. However, the performance of two co-grafted animals demonstrated partial recovery. L-dopa improved operant performance, demonstrating a dopaminergic component to the task. The results demonstrate recovery of behavioral function after surgical treatment, with adrenal co-grafted monkeys showing the greatest degree of improvement.