Cryptic splicing events in the iron transporter ABCB7 and other key target genes in SF3B1-mutant myelodysplastic syndromes.

The splicing factor SF3B1 is the most frequently mutated gene in myelodysplastic syndromes (MDS), and is strongly associated with the presence of ring sideroblasts (RS). We have performed a systematic analysis of cryptic splicing abnormalities from RNA sequencing data on hematopoietic stem cells (HSCs) of SF3B1-mutant MDS cases with RS. Aberrant splicing events in many downstream target genes were identified and cryptic 3' splice site usage was a frequent event in SF3B1-mutant MDS. The iron transporter ABCB7 is a well-recognized candidate gene showing marked downregulation in MDS with RS. Our analysis unveiled aberrant ABCB7 splicing, due to usage of an alternative 3' splice site in MDS patient samples, giving rise to a premature termination codon in the ABCB7 mRNA. Treatment of cultured SF3B1-mutant MDS erythroblasts and a CRISPR/Cas9-generated SF3B1-mutant cell line with the nonsense-mediated decay (NMD) inhibitor cycloheximide showed that the aberrantly spliced ABCB7 transcript is targeted by NMD. We describe cryptic splicing events in the HSCs of SF3B1-mutant MDS, and our data support a model in which NMD-induced downregulation of the iron exporter ABCB7 mRNA transcript resulting from aberrant splicing caused by mutant SF3B1 underlies the increased mitochondrial iron accumulation found in MDS patients with RS.

An international data set for CMML validates prognostic scoring systems and demonstrates a need for novel prognostication strategies.

Since its reclassification as a distinct disease entity, clinical research efforts have attempted to establish baseline characteristics and prognostic scoring systems for chronic myelomonocytic leukemia (CMML). Although existing data for baseline characteristics and CMML prognostication have been robustly developed and externally validated, these results have been limited by the small size of single-institution cohorts. We developed an international CMML data set that included 1832 cases across eight centers to establish the frequency of key clinical characteristics. Of note, we found that the majority of CMML patients were classified as World Health Organization CMML-1 and that a 7.5% bone marrow blast cut-point may discriminate prognosis with higher resolution in comparison with the existing 10%. We additionally interrogated existing CMML prognostic models and found that they are all valid and have comparable performance but are vulnerable to upstaging. Using random forest survival analysis for variable discovery, we demonstrated that the prognostic power of clinical variables alone is limited. Last, we confirmed the independent prognostic relevance of ASXL1 gene mutations and identified the novel adverse prognostic impact imparted by CBL mutations. Our data suggest that combinations of clinical and molecular information may be required to improve the accuracy of current CMML prognostication.

Endocrine disruptor & nutritional effects of heavy metals in ovarian hyperstimulation.

PURPOSE: There is increasing concern that environmental chemicals have a direct effect on fertility. Heavy metals such as mercury have been shown to affect various organ systems in humans including nervous system and skin, however they could also act as endocrine disrupting chemicals adversely affecting fertility. Metals such as zinc and selenium are essential micronutrients with diverse functions that may be important for reproductive outcomes. We measured mercury, zinc and selenium levels in the hair, a reliable reflection of long term environmental exposure and dietary status, to correlate with the outcome of ovarian hyperstimulation for in vitro fertilisation (IVF) treatment. METHODS: We analysed the hair of 30 subfertile women for mercury, zinc and selenium using inductively coupled mass spectrometry. Each woman underwent one cycle of IVF treatment. Correlation between the levels of these trace metals and treatment outcomes was investigated. RESULTS: Thirty women were recruited with mean (±SD) age of 32.7(4.4) years and BMI of 25.4(5.0)kg/m(2). Hair mercury concentration showed a negative correlation with oocyte yield (p < 0.05,ßcoefficient 0.38) and follicle number (p = 0.03,ß coefficient0.19) after ovarian stimulation. Zinc and selenium levels in hair correlated positively with oocyte yield after ovarian stimulation (p < 0.05,ß coefficient0.15) and (p = 0.03,ß coefficient0.21) respectively. Selenium levels in hair correlated significantly with follicle number following stimulation (p = 0.04, ßcoefficient0.22). There was no correlation between mercury, zinc and selenium in hair and their corresponding serum levels. CONCLUSION: These data suggest that mercury had a deleterious effect whilst there was a positive effect for zinc and selenium in the ovarian response to gonadotrophin therapy for IVF. Hair analysis offers a novel method of investigating the impact of long-term exposure to endocrine disruptors and nutritional status on reproductive outcomes.

Viewing ultrasound scan images prior to termination of pregnancy: choice for women or conflict for ultrasonographers?

Establishing pregnancy location is key to minimising the risks of abortion, and establishing gestational age optimises women's choice of procedure and timing. There is limited information in UK practice about the views of women having an ultrasound before an abortion with regards to whether they would wish to see the scan image of their fetus. We therefore surveyed the views of women having pre-abortion ultrasound scans and the views of ultrasonographers, using self-completed anonymous questionnaires. Responses were available for analysis from 191 women aged 16 to over 45. A total of 25 (13%) women had seen their scan. Of those who did not see their scan, 40 (24%) said they would have liked to see it, to aid their decision-making, as a right, or out of curiosity. Nine out of 17 ultrasonographers completed their questionnaires. Four ultrasonographers thought that women having scans before an abortion should see their scan images if they wished and another four thought they should not because of possible psychological effects. Their response also highlighted possible effects of such scans on those performing them. Given the choice, a larger number of women would like to see their scan images than actually ask. Women request to see their pre-abortion ultrasound images for their own individual reasons, which should be respected. Ultrasonographers may have conflict or ethical dilemmas when performing pre-abortion scans and they should be supported in identifying women for whom looking at their scan images would be of benefit.

Deregulated gene expression pathways in myelodysplastic syndrome hematopoietic stem cells.

To gain insight into the molecular pathogenesis of the myelodysplastic syndromes (MDS), we performed global gene expression profiling and pathway analysis on the hematopoietic stem cells (HSC) of 183 MDS patients as compared with the HSC of 17 healthy controls. The most significantly deregulated pathways in MDS include interferon signaling, thrombopoietin signaling and the Wnt pathways. Among the most significantly deregulated gene pathways in early MDS are immunodeficiency, apoptosis and chemokine signaling, whereas advanced MDS is characterized by deregulation of DNA damage response and checkpoint pathways. We have identified distinct gene expression profiles and deregulated gene pathways in patients with del(5q), trisomy 8 or -7/del(7q). Patients with trisomy 8 are characterized by deregulation of pathways involved in the immune response, patients with -7/del(7q) by pathways involved in cell survival, whereas patients with del(5q) show deregulation of integrin signaling and cell cycle regulation pathways. This is the first study to determine deregulated gene pathways and ontology groups in the HSC of a large group of MDS patients. The deregulated pathways identified are likely to be critical to the MDS HSC phenotype and give new insights into the molecular pathogenesis of this disorder, thereby providing new targets for therapeutic intervention.

Insulin resistance and free androgen index correlate with the outcome of controlled ovarian hyperstimulation in non-PCOS women undergoing IVF.

BACKGROUND: The dual effects of insulin and androgen on the ovary act to promote early folliculogenesis. In the context of polycystic ovarian syndrome (PCOS), the presence of hyperinsulinaemia, resulting from increased insulin resistance (IR), and hyperandrogenaemia lead to the appearance of multiple antral follicles and frequently a multi-follicular response to gonadotrophin stimulation for assisted reproductive treatments (ARTs). The effect of IR and androgen status in women without PCOS on the follicular outcome of controlled ovarian hyperstimulation (COH) is not known. METHODS: We assessed the IR [using the homeostasis model assessment (HOMA)] and androgen status of 49 women without PCOS undergoing an ART cycle. This was then related to the treatment cycle outcome. RESULTS: We found a significant positive correlation between HOMA and BMI, and free androgen index (FAI) and testosterone. The FAI significantly positively correlated with total follicle count after COH. The total follicle count was significantly higher in those with a HOMA >2.5, and HOMA positively correlated with total follicle count in this group of IR women (HOMA > 2.5). CONCLUSIONS: Our results suggest a positive correlation of HOMA-IR levels above a threshold level of 2.5 and a continuous positive correlation of free androgen (FAI) to total ovarian follicle count following COH in the non-PCOS patient.

Do falls and falls-injuries in hospital indicate negligent care -- and how big is the risk? A retrospective analysis of the NHS Litigation Authority Database of clinical negligence claims, resulting from falls in hospitals in England 1995 to 2006.

BACKGROUND: Accidental falls are very common in older hospital patients -- accounting for 32% of reported adult patient safety incidents in UK National Health Service (NHS) hospitals and occurring with similar frequency in settings internationally. In countries where the population is ageing, and care is provided in inpatient settings, falls prevention is therefore a significant and growing risk-management issue. Falls may lead to a variety of harms and costs, are cited in formal complaints and can lead to claims of clinical negligence. The NHS Litigation Authority (NHSLA) negligence claims database provides a novel opportunity to systematically analyse such (falls-related) claims made against NHS organisations in England and to learn lessons for risk-management systems and claims recording. OBJECTIVES: To describe the circumstances and injuries most frequently cited in falls-related claims; to investigate any association between the financial impact (total cost), and the circumstances of or injuries resulting from falls in "closed" claims; to draw lessons for falls risk management and for future data capture on falls incidents and resulting claims analysis; to identify priorities for future research. METHODS: A keyword search was run on the NHSLA claims database for April 1995 to February 2006, to identify all claims apparently relating to falls. Claims were excluded from further analysis if, on scrutiny, they had not resulted from falls, or if they were still "open" (ie, unresolved). From the narrative descriptions of closed claims (ie, those for which the financial outcome was known), we developed categories of "principal" and "secondary" injury/harm and "principal" and "contributory" circumstance of falls. For each category, it was determined whether cases had resulted in payment and what total payments (damages and costs) were awarded. The proportions of contribution-specific injuries or circumstances to the number of cases and to the overall costs incurred were compared in order to identify circumstances that tend to be more costly. Means were compared and tested through analysis of variance (ANOVA). The association between categorical variables was tested using the chi-square test. RESULTS: Of 668 claims identified by word search, 646 met inclusion criteria. The results presented are for the 479 of these that were "closed" at the time of the census. Of these, 290 (60.5%) had resulted in payment of costs or damages, with the overall total payment being 6,200,737 pound (mean payment 12,945 pound). All claims were settled out of court, so no legal rulings on establishing liability or causation of injury are available. "Falls whilst walking;" "from beds or trolleys" ("with and without bedrails applied") or "transferring/from a chair" were the most frequent source of these claims (n = 308, 64.2%). Clear secondary contributory circumstances were identified in 190 (39.7%) of closed claims. The most common circumstances cited were "perioperative/procedural incidents" (60, 12.5%) and "requests for bedrails being ignored" (54, 11.3%). For primary injuries, "hip/femoral/pelvic fracture" accounted for 203 (42.4%) of closed claims with total payments of 3,228,781 pound (52.1% of all payments), with a mean payment 15,905 pound per closed case. A "secondary" contributory circumstance could be attributed in 133 (27.8%) of cases. Of these, "delay in diagnosis of injury," "recurrent falls during admission" and "fatalities relating to falls" were the commonest circumstances (n = 59, 12.2%). DISCUSSION: Although falls are the highest volume patient safety incident reported in hospital trusts in England, they result in a relatively small number of negligence claims and receive a relatively low total payment (0.019% in both cases). The mean payment in closed claims is also relatively small. This may reflect the high average age of the people who fall and difficulty in establishing causation, especially where individuals are already frail when they fall. The patterns of claims and the narrative descriptions provide wider lessons for improving risk-management strategies. However, the inherent limitations and biases in the data routinely recorded for legal purposes suggest that for more informative research or actuarial claims analysis, more comprehensive and systematic data to be recorded for each incident claim are needed.

Obstetrics and gynaecology trainees' knowledge of electrosurgical equipment.

Electrosurgical energy is potentially dangerous but is a very useful surgical tool. A survey was planned to assess the knowledge of electrosurgery among obstetrics and gynaecology trainees in the Yorkshire Deanery (training region). Questionnaires were given to trainees attending the Modular training programme and were collected on the same day. A total of 71 questionnaires were collected, which represents 62% of the 113 trainees in Yorkshire region: 39 participants were registrars; 27 senior house officers (SHO); four senior SHOs, and one Clinical Fellow. A total of 52% of trainees had only MRCOG Part 1 and 36% of trainees had MRCOG Part 2; 53% of trainees had attended a Basic Surgical Skills course and 10% of trainees were not aware that the 'blue pedal' is used for coagulation (all were SHO except one Specialist Registrar, SpR). A total of 71% of trainees answered incorrectly that coagulating current provides an initial high voltage peak that quickly dissipates, while cutting current provides a constant high energy waveform. Trainees who had attended a Basic Surgical Skills (BSS) course did not demonstrate better knowledge. Knowledge of electrosurgery therefore was found to be poor among obstetrics and gynaecology trainees. Attending a BSS course did not seem to improve it significantly. This study outlines a training need. Perhaps a refresher course in electrosurgery and more focussed training might be useful for trainees with a particular interest in operative gynaecology, to acquire the necessary information and skills.

Low IPSS score and bone marrow hypocellularity in MDS patients predict hematological responses to antithymocyte globulin.

Immunosuppressive therapy has been shown to induce sustained hematological responses in a subset of patients with myelodysplastic syndromes (MDS). In particular, antithymocyte globulin (ATG), a polyclonal immunoglobulin induces hematological responses in up to 60% of MDS patients. We report herein on the results of a retrospective multicenter study on the use of ATG in the treatment of 96 patients with MDS. Patients were evaluated for duration of response to ATG, as well as survival after administration of ATG. The median age of the cohort was 54.7 years (range: 19-75 years), with a median follow-up of 33.8 months (range: 0.8-133 months). A total of 40 patients (42%) achieved a hematological response, of which 30 patients (75%) had a durable hematological response lasting a median duration of 31.5 months (range: 6-92 months). On multivariate analysis, both low International Prognostic Scoring System (IPSS) and bone marrow (BM) hypocellularity were independent predictive factors for improved response to ATG (IPSS Int-2/high: odds ratio (OR) 0.08, P=0.018 and BM normo/hypercellularity: OR 0.49, P=0.012). In addition, IPSS was the sole predictor of overall survival, with Int-2/high risk patients having a significantly poorer survival outcome (OR 0.08, P<0.01). In conclusion, this study identifies BM hypocellularity and a low IPSS as important factors predicting response to ATG.

Ultrasound appearance of the uterus in women over 60 years of age on tibolone: is it a SERM?

OBJECTIVES: To evaluate the effects of long-term tibolone on the endometria of asymptomatic women over the age of 60 years. METHODS: An observational study of the ultrasound appearance of the endometria of women aged 60 years or older taking long-term tibolone. Those found to have a double-layer endometrial thickness greater than 4 mm were offered endometrial biopsy. RESULTS: Seventeen asymptomatic patients with a mean age of 61 years (range 60-73 years) and an average duration of tibolone use of 5 years (range 3-12 years) were recruited. Seven patients (41%) showed a thickened endometrium of more than 4 mm. Three of these had an area of translucency in the subendometrial space. Of the ten women with an endometrial thickness of less than 4 mm, four showed the presence of subendometrial fluid. Hysteroscopy was performed in five out of the seven women, as two did not accept further investigations. In all five women, the endometrial cavity was reported as atrophic and the histology showed an inactive basal type. In two of the five cases examined histologically, there were additional changes suggesting endometrial polyp, although the samples were from macroscopically non-polypoid endometrium. DISCUSSION: Endometrial thickness is increased in 41% of elderly women on long-term treatment with tibolone. However, at hysteroscopy, the endometrium was uniformly atrophic. Similar finding have been described in women using selective estrogen receptor modulators and a similar mechanism of action of these drugs on the uterus should be considered.

Testing the pluralist approach to sex: the influence of environment on synergistic interactions between mutation load and parasitism in Daphnia magna.

Both deleterious mutations and parasites have been acknowledged as potential selective forces responsible for the evolutionary maintenance of sexual reproduction. The pluralist approach to sex proposes that these two factors may have to interact synergistically in order to stabilize sex, and one of the simplest ways this could occur is if parasites are capable of causing synergistic epistasis between mutations in their hosts. However, the effects of both deleterious mutations and parasitism are known to be influenced by a range of environmental factors, so the nature of the interaction may depend upon the organisms' environment. Using chemically mutated Daphnia magna lines, we examined the effects of mutation and parasitism under a range of temperature and food regimes. We found that although parasites were capable of causing synergistic epistasis between mutations in their hosts, these effects were dependent upon an interaction between parasite genotype and temperature.

Low dose antithymocyte globulin for the treatment of older patients with aplastic anaemia.

We report 14 older patients with aplastic anaemia (AA) who were treated with 'low dose' antithymocyte globulin (ATG). The aims of the study were to assess the efficacy and safety of reduced dose ATG in patients over the age of 60 years. Median age was 71 years (range 62-74 years). At the study endpoint (response to treatment at 6 months) 12 patients were evaluable. All patients received lymphoglobuline (horse ATG; Genzyme) at a dose of 0.5vials/10kg/day for 5 days (5mg/kg/day, equivalent to one-third of the standard dose). There were no deaths attributed to ATG. Two patients died during follow-up, from sepsis and anaphylaxis following platelet transfusion, respectively. Only one of the 12 evaluable patients responded to treatment and remains transfusion independent at 14 months after ATG. These results suggest that this lower dose of ATG, though well tolerated, had low efficacy in the treatment of AA.

Is previous aberrant reproductive outcome predictive of subsequently reduced fecundity?

BACKGROUND: Effect of past reproductive performance on subsequent fecundity is uncertain. METHODS: A total of 2983 consecutive pregnant women self-completed questionnaires about time to pregnancy (TTP), pregnancy planning, previous pregnancies, contraceptive use, age, and individual/lifestyle variables. Outcome measures were: TTP, conception rates (CR) and, subfecundity odds ratio (OR; with 95% confidence intervals) before and after each outcome of last pregnancy. RESULTS: After miscarriage, TTP was longer than before miscarriage [2.1 (1.4-3.0), P < 0.001] and than TTP after livebirth [OR = 2.1 (1.6-2.6), P < 0.001]. Also subfecundity OR after miscarriage increased [1.7 (1.2-2.4), 1.8 (1.2-2.5), P = 0.001, 0.002 respectively]. This effect was more evident in older and obese women. Compared with livebirth, time to ectopic pregnancy (EP) was longer [OR = 13.8 (1.8-108.5), P = 0.001] but TTP after EP was not significantly different. Subfecundity OR relative to livebirth were 12.8 (3.6-45.0) (P<0.001) before, and 3.9 (1.4-11.0) (P=0.01) after, EP. The CR after EP increased 3-fold (1.1-8.3) over those prior to EP. Time to the terminated pregnancies even without contraceptive failures was shorter than that to livebirth [OR = 0.5 (0.3-0.7), P = 0.001] and than TTP after termination [0.35 (0.1-0.8), P = 0.001]. Also subfecundity OR increased after termination [7.2 (1.8-29.7), P = 0.02]. CONCLUSIONS: Miscarriers should be counselled about short-term reduction in subsequent fecundity, and earlier investigations should be considered in those who have other potential risk factors for reduced fertility. Further studies are required to clarify the relatively favourable effect on fecundity following EP and the relative reduction in fecundity after termination of pregnancy.

A national study examining the effect of making emergency hormonal contraception available without prescription.

BACKGROUND: In January 2001, emergency hormonal contraception was made available for women over the age of 16 years directly from a pharmacist without prescription. It is of interest whether this change in the UK has led to any improvements or deterioration in the service provided for the women who need it. METHODS: Self- completed, anonymous questionnaires were distributed to women requesting emergency hormonal contraception through a single group of pharmacies located throughout England, Wales and Scotland. RESULTS: A total 419 women returned completed questionnaires. A greater proportion of women were able to take emergency contraception within 24 h when they obtained their tablets directly from a pharmacy without a prescription (64% versus 46%, P = 0.029). Women who obtained their drugs directly from the pharmacist were just as well informed, just as likely to arrange regular follow-up and generally preferred this system, although they disliked having to pay. CONCLUSION: Making emergency hormonal contraception available without prescription has improved services to women who need them, but these improvements are quantitatively minimal, preventing only five additional pregnancies per 10,000 users.