A comprehensive description of kidney disease progression after acute kidney injury from a prospective, parallel-group cohort study.

Acute kidney injury (AKI) is associated with adverse long-term outcomes, but many studies are retrospective, focused on specific patient groups or lack adequate comparators. The ARID (AKI Risk in Derby) Study was a five-year prospective parallel-group cohort study to examine this. Hospitalized cohorts with and without exposure to AKI were matched 1:1 for age, baseline kidney function, and diabetes. Estimated glomerular filtration rate (eGFR) and the urinary albumin:creatinine ratio (uACR) were measured at three-months, one-, three- and five-years. Outcomes included kidney disease progression, heart failure episodes and mortality. In 866 matched individuals, kidney disease progression at five years was found to be significantly increased in 30% of the exposed group versus 7% of those non-exposed (adjusted odds ratio 2.49 [95% confidence interval 1.43 to 4.36]). In the AKI group, this was largely characterized by incomplete recovery of kidney function by three months. Further episodes of AKI during follow-up were significantly more common in the exposed group (odds ratio 2.71 [1.94 to 3.77]) and had an additive effect on risk of kidney disease progression. Mortality and heart failure episodes were more frequent in the exposed group, but the association with AKI was no longer significant when models were adjusted for three-month eGFR and uACR. In a general hospitalized population, kidney disease progression after five years was common and strongly associated with AKI. Thus, the time course of changes and the attenuation of associations with adverse outcomes after adjustment for three-month eGFR and uACR suggest non-recovery of kidney function is an important assessment in post-AKI care and a potential future target for intervention. STUDY REGISTRATION: ISRCTN25405995.

Nuclear-Import Receptors Reverse Aberrant Phase Transitions of RNA-Binding Proteins with Prion-like Domains.

RNA-binding proteins (RBPs) with prion-like domains (PrLDs) phase transition to functional liquids, which can mature into aberrant hydrogels composed of pathological fibrils that underpin fatal neurodegenerative disorders. Several nuclear RBPs with PrLDs, including TDP-43, FUS, hnRNPA1, and hnRNPA2, mislocalize to cytoplasmic inclusions in neurodegenerative disorders, and mutations in their PrLDs can accelerate fibrillization and cause disease. Here, we establish that nuclear-import receptors (NIRs) specifically chaperone and potently disaggregate wild-type and disease-linked RBPs bearing a NLS. Karyopherin-ß2 (also called Transportin-1) engages PY-NLSs to inhibit and reverse FUS, TAF15, EWSR1, hnRNPA1, and hnRNPA2 fibrillization, whereas Importin-α plus Karyopherin-ß1 prevent and reverse TDP-43 fibrillization. Remarkably, Karyopherin-ß2 dissolves phase-separated liquids and aberrant fibrillar hydrogels formed by FUS and hnRNPA1. In vivo, Karyopherin-ß2 prevents RBPs with PY-NLSs accumulating in stress granules, restores nuclear RBP localization and function, and rescues degeneration caused by disease-linked FUS and hnRNPA2. Thus, NIRs therapeutically restore RBP homeostasis and mitigate neurodegeneration.

Nuclear localized C9orf72-associated arginine-containing dipeptides exhibit age-dependent toxicity in C. elegans.

A hexanucleotide repeat expansion mutation in the C9orf72 gene represents a prevalent genetic cause of several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. Non-canonical translation of this repeat gives rise to several distinct dipeptide protein species that could play pathological roles in disease. Here, we show in the model system Caenorhabditis elegans that expression of the arginine-containing dipeptides, but not alanine-containing dipeptides, produces toxic phenotypes in multiple cellular contexts, including motor neurons. Expression of either (PR)50 or (GR)50 during development caused a highly penetrant developmental arrest, while post-developmental expression caused age-onset paralysis. Both (PR)50- and (GR)50-green fluorescent protein tagged dipeptides were present in the nucleus and nuclear localization was necessary and sufficient for their toxicity. Using an inducible expression system, we discovered that age-onset phenotypes caused by (PR)50 required both continual (PR)50 expression and an aged cellular environment. The toxicity of (PR)50 was modified by genetic mutations that uncouple physiological aging from chronological aging. However, these same mutations failed to modify the toxicity of (GR)50, suggesting that (PR)50 and (GR)50 exert their toxicity through partially distinct mechanism(s). Changing the rate of physiological aging also mitigates toxicity in other C. elegans models of ALS, suggesting that the (PR)50 dipeptide might engage similar toxicity mechanisms as other ALS disease-causing proteins.

Three-year outcomes after acute kidney injury: results of a prospective parallel group cohort study.

OBJECTIVES: Using a prospective study design, we aimed to characterise the effect of acute kidney injury (AKI) on long-term changes in renal function in a general hospital population. PARTICIPANTS: Hospitalised patients with AKI (exposed) and hospitalised patients without AKI (non-exposed), recruited at 3 months after hospital admission. DESIGN: Prospective, matched parallel group cohort study, in which renal function and proteinuria were measured at 3 months, 1 year and 3 years. SETTING: Single UK centre. CLINICAL END POINTS: Clinical end points at 3 years were comparison of the following variables between exposed and non-exposed groups: renal function, prevalence of proteinuria and albuminuria and chronic kidney disease (CKD) progression/development at each time point. CKD progression was defined as a decrease in the estimated glomerular filtration rate (eGFR) of ≥25% associated with a decline in eGFR stage. RESULTS: 300 exposed and non-exposed patients were successfully matched 1:1 for age and baseline renal function; 70% of the exposed group had AKI stage 1. During follow-up, the AKI group had lower eGFR than non-exposed patients at each time point. At 3 years, the mean eGFR was 60.7±21 mL/min/1.73 m2 in the AKI group compared with 68.4±21 mL/min/1.73 m2 in the non-exposed group, p=0.003. CKD development or progression at 3 years occurred in 30 (24.6%) of the AKI group compared with 10 (7.5%) of the non-exposed group, p<0.001. Albuminuria was more common in the AKI group, and increased with AKI severity. Factors independently associated with CKD development/progression after AKI were non-recovery at 90 days, male gender, diabetes and recurrent AKI. CONCLUSIONS: AKI is associated with deterioration in renal function to 3 years, even in an unselected population with predominantly AKI stage 1. Non-recovery from AKI is an important factor determining long-term outcome.

Pur-alpha regulates cytoplasmic stress granule dynamics and ameliorates FUS toxicity.

Amyotrophic lateral sclerosis is characterized by progressive loss of motor neurons in the brain and spinal cord. Mutations in several genes, including FUS, TDP43, Matrin 3, hnRNPA2 and other RNA-binding proteins, have been linked to ALS pathology. Recently, Pur-alpha, a DNA/RNA-binding protein was found to bind to C9orf72 repeat expansions and could possibly play a role in the pathogenesis of ALS. When overexpressed, Pur-alpha mitigates toxicities associated with Fragile X tumor ataxia syndrome (FXTAS) and C9orf72 repeat expansion diseases in Drosophila and mammalian cell culture models. However, the function of Pur-alpha in regulating ALS pathogenesis has not been fully understood. We identified Pur-alpha as a novel component of cytoplasmic stress granules (SGs) in ALS patient cells carrying disease-causing mutations in FUS. When cells were challenged with stress, we observed that Pur-alpha co-localized with mutant FUS in ALS patient cells and became trapped in constitutive SGs. We also found that FUS physically interacted with Pur-alpha in mammalian neuronal cells. Interestingly, shRNA-mediated knock down of endogenous Pur-alpha significantly reduced formation of cytoplasmic stress granules in mammalian cells suggesting that Pur-alpha is essential for the formation of SGs. Furthermore, ectopic expression of Pur-alpha blocked cytoplasmic mislocalization of mutant FUS and strongly suppressed toxicity associated with mutant FUS expression in primary motor neurons. Our data emphasizes the importance of stress granules in ALS pathogenesis and identifies Pur-alpha as a novel regulator of SG dynamics.

Protein arginine methyltransferase 6 enhances polyglutamine-expanded androgen receptor function and toxicity in spinal and bulbar muscular atrophy.

Polyglutamine expansion in androgen receptor (AR) is responsible for spinobulbar muscular atrophy (SBMA) that leads to selective loss of lower motor neurons. Using SBMA as a model, we explored the relationship between protein structure/function and neurodegeneration in polyglutamine diseases. We show here that protein arginine methyltransferase 6 (PRMT6) is a specific co-activator of normal and mutant AR and that the interaction of PRMT6 with AR is significantly enhanced in the AR mutant. AR and PRMT6 interaction occurs through the PRMT6 steroid receptor interaction motif, LXXLL, and the AR activating function 2 surface. AR transactivation requires PRMT6 catalytic activity and involves methylation of arginine residues at Akt consensus site motifs, which is mutually exclusive with serine phosphorylation by Akt. The enhanced interaction of PRMT6 and mutant AR leads to neurodegeneration in cell and fly models of SBMA. These findings demonstrate a direct role of arginine methylation in polyglutamine disease pathogenesis.

Antisense proline-arginine RAN dipeptides linked to C9ORF72-ALS/FTD form toxic nuclear aggregates that initiate in vitro and in vivo neuronal death.

Expanded GGGGCC (G4C2) nucleotide repeats within the C9ORF72 gene are the most common genetic mutation associated with both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Sense and antisense transcripts of these expansions are translated to form five dipeptide repeat proteins (DRPs). We employed primary cortical and motor neuron cultures, live-cell imaging, and transgenic fly models and found that the arginine-rich dipeptides, in particular Proline-Arginine (PR), are potently neurotoxic. Factors that anticipated their neurotoxicity included aggregation in nucleoli, decreased number of processing bodies, and stress granule formation, implying global translational dysregulation as path accountable for toxicity. Nuclear PR aggregates were also found in human induced motor neurons and postmortem spinal cord tissues from C9ORF72 ALS and ALS/FTD patients. Intronic G4C2 transcripts, but not loss of C9ORF72 protein, are also toxic to motor and cortical neurons. Interestingly, G4C2 transcript-mediated neurotoxicity synergizes with that of PR aggregates, suggesting convergence of mechanisms.

The effects of acute kidney injury on long-term renal function and proteinuria in a general hospitalised population.

BACKGROUND: Acute kidney injury (AKI) is common in hospitalised patients and is associated with adverse long-term consequences. There is an urgent need to understand these sequelae in general hospitalised patients utilising a prospective cohort-based approach. We aimed to test the feasibility of study methodology prior to commencing a large-scale study and investigate the effects of AKI on chronic kidney disease (CKD) progression and proteinuria. METHODS: Pilot study testing novel methodology for remote patient recruitment within a prospective case-control design. 300 cases (hospitalised patients with AKI) and controls (hospitalised patients without AKI) were matched 1:1 for age and baseline estimated glomerular filtration rate (eGFR). 70% of cases had AKI stage 1, 16% AKI stage 2 and 14% AKI stage 3. Renal function and proteinuria were measured 3 and 12 months after hospital admission. RESULTS: The study met pre-defined recruitment, withdrawal and matching criteria. Renal function was worse in the AKI group at 3 (eGFR 61 ± 20 vs. 74 ± 23 ml/min/1.73 m(2), p < 0.001) and 12 months (eGFR 64 ± 23 vs. 75 ± 25 ml/min/1.73 m(2), p < 0.001). More cases than controls had CKD progression at 3 months (14 vs. 0.7%, p < 0.001). This difference persisted to 12 months, but there was no significant change between 3 and 12 months. Proteinuria and albuminuria were more prevalent in the AKI group and associated with CKD progression. CONCLUSIONS: We describe a method of remote patient recruitment which could be employed more widely for prospective observational studies. Even mild AKI is associated with long-term renal dysfunction. Further investigation using this methodology is now underway.

Protein arginine methyltransferase 1 and 8 interact with FUS to modify its sub-cellular distribution and toxicity in vitro and in vivo.

Amyotrophic lateral sclerosis (ALS) is a late onset and progressive motor neuron disease. Mutations in the gene coding for fused in sarcoma/translocated in liposarcoma (FUS) are responsible for some cases of both familial and sporadic forms of ALS. The mechanism through which mutations of FUS result in motor neuron degeneration and loss is not known. FUS belongs to the family of TET proteins, which are regulated at the post-translational level by arginine methylation. Here, we investigated the impact of arginine methylation in the pathogenesis of FUS-related ALS. We found that wild type FUS (FUS-WT) specifically interacts with protein arginine methyltransferases 1 and 8 (PRMT1 and PRMT8) and undergoes asymmetric dimethylation in cultured cells. ALS-causing FUS mutants retained the ability to interact with both PRMT1 and PRMT8 and undergo asymmetric dimethylation similar to FUS-WT. Importantly, PRMT1 and PRMT8 localized to mutant FUS-positive inclusion bodies. Pharmacologic inhibition of PRMT1 and PRMT8 activity reduced both the nuclear and cytoplasmic accumulation of FUS-WT and ALS-associated FUS mutants in motor neuron-derived cells and in cells obtained from an ALS patient carrying the R518G mutation. Genetic ablation of the fly homologue of human PRMT1 (DART1) exacerbated the neurodegeneration induced by overexpression of FUS-WT and R521H FUS mutant in a Drosophila model of FUS-related ALS. These results support a role for arginine methylation in the pathogenesis of FUS-related ALS.

RNA-binding ability of FUS regulates neurodegeneration, cytoplasmic mislocalization and incorporation into stress granules associated with FUS carrying ALS-linked mutations.

Amyotrophic lateral sclerosis (ALS) is an uncommon neurodegenerative disease caused by degeneration of upper and lower motor neurons. Several genes, including SOD1, TDP-43, FUS, Ubiquilin 2, C9orf72 and Profilin 1, have been linked with the sporadic and familiar forms of ALS. FUS is a DNA/RNA-binding protein (RBP) that forms cytoplasmic inclusions in ALS and frontotemporal lobular degeneration (FTLD) patients' brains and spinal cords. However, it is unknown whether the RNA-binding ability of FUS is required for causing ALS pathogenesis. Here, we exploited a Drosophila model of ALS and neuronal cell lines to elucidate the role of the RNA-binding ability of FUS in regulating FUS-mediated toxicity, cytoplasmic mislocalization and incorporation into stress granules (SGs). To determine the role of the RNA-binding ability of FUS in ALS, we mutated FUS RNA-binding sites (F305L, F341L, F359L, F368L) and generated RNA-binding-incompetent FUS mutants with and without ALS-causing mutations (R518K or R521C). We found that mutating the aforementioned four phenylalanine (F) amino acids to leucines (L) (4F-L) eliminates FUS RNA binding. We observed that these RNA-binding mutations block neurodegenerative phenotypes seen in the fly brains, eyes and motor neurons compared with the expression of RNA-binding-competent FUS carrying ALS-causing mutations. Interestingly, RNA-binding-deficient FUS strongly localized to the nucleus of Drosophila motor neurons and mammalian neuronal cells, whereas FUS carrying ALS-linked mutations was distributed to the nucleus and cytoplasm. Importantly, we determined that incorporation of mutant FUS into the SG compartment is dependent on the RNA-binding ability of FUS. In summary, we demonstrate that the RNA-binding ability of FUS is essential for the neurodegenerative phenotype in vivo of mutant FUS (either through direct contact with RNA or through interactions with other RBPs).

Defining the cause of death in hospitalised patients with acute kidney injury.

BACKGROUND: The high mortality rates that follow the onset of acute kidney injury (AKI) are well recognised. However, the mode of death in patients with AKI remains relatively under-studied, particularly in general hospitalised populations who represent the majority of those affected. We sought to describe the primary cause of death in a large group of prospectively identified patients with AKI. METHODS: All patients sustaining AKI at our centre between 1(st) October 2010 and 31(st) October 2011 were identified by real-time, hospital-wide, electronic AKI reporting based on the Acute Kidney Injury Network (AKIN) diagnostic criteria. Using this system we are able to generate a prospective database of all AKI cases that includes demographic, outcome and hospital coding data. For those patients that died during hospital admission, cause of death was derived from the Medical Certificate of Cause of Death. RESULTS: During the study period there were 3,930 patients who sustained AKI; 62.0% had AKI stage 1, 20.6% had stage 2 and 17.4% stage 3. In-hospital mortality rate was 21.9% (859 patients). Cause of death could be identified in 93.4% of cases. There were three main disease categories accounting for three quarters of all mortality; sepsis (41.1%), cardiovascular disease (19.2%) and malignancy (12.9%). The major diagnosis leading to sepsis was pneumonia, whilst cardiovascular death was largely a result of heart failure and ischaemic heart disease. AKI was the primary cause of death in only 3% of cases. CONCLUSIONS: Mortality associated with AKI remains high, although cause of death is usually concurrent illness. Specific strategies to improve outcomes may therefore need to target not just the management of AKI but also the most relevant co-existing conditions.

Use of electronic results reporting to diagnose and monitor AKI in hospitalized patients.

BACKGROUND AND OBJECTIVES: Many patients with AKI are cared for by non-nephrologists. This can result in variable standards of care that contribute to poor outcomes. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: To improve AKI recognition, a real-time, hospital-wide, electronic reporting system was designed based on current Acute Kidney Injury Network criteria. This system allowed prospective data collection on AKI incidence and outcomes such as mortality rate, length of hospital stay, and renal recovery. The setting was a 1139-bed teaching hospital with a tertiary referral nephrology unit. RESULTS: An electronic reporting system was successfully introduced into clinical practice (false positive rate, 1.7%; false negative rate, 0.2%). The results showed that there were 3202 AKI episodes in 2619 patients during the 9-month study period (5.4% of hospital admissions). The in-hospital mortality rate was 23.8% and increased with more severe AKI (16.1% for stage 1 AKI versus 36.1% for stage 3) (P<0.001). More severe AKI was associated with longer length of hospital stay for stage 1 (8 days; interquartile range, 13) versus 11 days for stage 3 (interquartile range, 16) (P<0.001) and reduced chance of renal recovery (80.0% in stage 1 AKI versus 58.8% in stage 3) (P<0.001). Utility of the Acute Kidney Injury Network criteria was reduced in those with pre-existing CKD. CONCLUSIONS: AKI is common in hospitalized patients and is associated with very poor outcomes. The successful implementation of electronic alert systems to aid early recognition of AKI across all acute specialties is one strategy that may help raise standards of care.

Novel anti-inflammatory compound SEN1176 alleviates behavioral deficits induced following bilateral intrahippocampal injection of aggregated amyloid-ß1₋42.

Behavioral effects of a novel anti-inflammatory SEN1176 were investigated. This pyrrolo[3,2-e][1,2,4]triazolo[1,5-a]pyrimidine suppresses amyloid-ß (Aß)1-42-induced macrophage production of nitric oxide, TNF-α, IL-1ß, and IL-6 in a dose-dependent fashion, an activity profile consistent with SEN1176 being a neuroinflammation inhibitor. Using male Sprague-Dawley rats, SEN1176 was examined relative to detrimental behavioral effects induced following bilateral intrahippocampal (IH) injections of aggregated Aß1-42. The rats were trained to respond under an alternating-lever cyclic-ratio (ALCR) schedule of food reinforcement, enabling measurement of parameters of operant performance that reflect aspects of learning and memory. Under the ALCR schedule, orally administered SEN1176 at 5, 20, or 30 mg/kg was effective in reducing the behavioral deficit caused by bilateral IH aggregated Aß1-42 injections in a dose-related manner over a 90-day treatment period. SEN1176 at 20 and 30 mg/kg significantly reduced lever switching errors and, at doses of 5, 10, and 30 mg/kg, significantly reduced incorrect lever perseverations, indicating a reduction of the behavioral deficit induced as a result of inflammation following IH Aß1-42 injections. When treatment with SEN1176 was instigated 30 days after IH Aß1-42 injections, it resulted in progressive protection, and withdrawal of SEN1176 treatment 60 days after IH Aß1-42 injections revealed partial retention of the protective effect. SEN1176 also significantly reduced numbers of activated astrocytes adjacent to the aggregated Aß1-42 injection sites. These results indicate the potential of SEN1176 for alleviating chronic neuroinflammatory processes related to brain Aß deposition that affect learning and memory in Alzheimer's disease.

Evolution of coronary artery calcification in patients with chronic kidney disease Stages 3 and 4, with and without diabetes.

BACKGROUND: The purpose of this study was to report the evolution of coronary artery calcification (CAC) in subjects with chronic kidney disease Stages 3 and 4 comparing those with and without diabetes. We previously reported prevalence in the same population. METHODS: CAC was measured using multi-slice computer tomography. We prospectively followed up 103 patients for 2 years, 49 with diabetes and 54 without diabetes. Demographic, routine biochemistry, calcification inhibitors and bone mineral density data were collected and analysed. Evolution of CAC was defined as those with a difference of ≥ 2.5 U between baseline and final square root CAC scores. RESULTS: There were more progressors in the group with diabetes, 24 compared to 12 in the group without diabetes (P= 0.004). When diabetes was present, CAC progressed equally in men and women. Risk factors for evolution of CAC included age, baseline CAC score and serum phosphate levels. Baseline CAC score, phosphate and body mass index were independent predictors for the increase of CAC score during the study period. Severity of CAC was greater in the diabetes group (median CAC score at baseline in the group with diabetes 154 increased to 258 2 years later, P < 0.001). CONCLUSIONS: Evolution of CAC is greater in older patients and those with diabetes, where the gender advantage of being female is lost. Serum phosphate level, despite being within the normal range and virtually no use of phosphate binders, was also a risk factor. Further studies are required to determine the levels of serum phosphate required to minimize cardiovascular risk.

Fragmentation of trimethoprim and other compounds containing alkoxy-phenyl groups in electrospray ionisation tandem mass spectrometry.

This work describes electrospray ionisation tandem mass spectrometry studies of trimethoprim and a series of structurally similar compounds containing alkoxy-phenyl groups; using accurate mass measurement to confirm the proposed fragmentations. Radical cations were observed in the spectra obtained for some of the compounds, as well as uncommon fragmentations showing losses of CH4 and C2H6, whereas other compounds showed the formation of even electron ions. Possible structures for the fragment ions have been proposed and explanations for the different types of fragmentations based on the structures of the compounds. In addition an alternate structure for a fragment ion previously reported for tandem mass spectrometry of trimethoprim has been proposed, based on accurate mass measurement.

Longitudinal changes of insulin-like growth factors and their binding proteins throughout normal pregnancy.

BACKGROUND: Insulin-like growth factors (IGFs) are anabolic proteins that are essential regulators of cell division, differentiation and growth. We describe the longitudinal changes in IGF-I, IGF-II and the binding proteins IGFBP-1, -2 and -3 before and during normal pregnancy. METHOD: Serum samples were taken before conception and then at 12, 24 and 36 weeks of gestation in 41 healthy women with uncomplicated pregnancies. We measured IGF-I using an automated chemiluminescent method, IGF-II and IGFBP-2 using in-house radioimmunoassays (RIAs), and IGFBP-1 and IGFBP-3 using commercial enzyme-linked immunosorbent assay (ELISA) and RIA kits, respectively. Because of the potential haemodilution effects during pregnancy, albumin was also measured in all samples. RESULTS: There was a significant fall in IGF-I during the first (36%) and second trimesters (21%) followed by an increase of 25% at 36 weeks. During pregnancy, the mean IGF-II concentrations fell by 12% at 12 weeks, 8% at 24 and 8% at 36 weeks compared with pre-conception values. When IGF-II results were adjusted for the haemodilution of pregnancy, its concentrations increased. During pregnancy, there was a rapid increase in mean IGFBP-1 levels by 17-fold (12 weeks), 24-fold (24 weeks) and 25-fold (36 weeks). IGFBP-2 concentrations fell after conception but started to increase towards term. This increase was more significant when adjusted for haemodilution. In contrast, IGFBP-3 concentrations increased significantly throughout pregnancy. CONCLUSION: Our data on the physiological changes of IGFs and their binding proteins add further evidence of the vital roles of these hormones throughout normal pregnancy.

Cervical screening in women over the age of 50: results of a population-based multicentre study.

OBJECTIVE: It has been suggested that women over 50 with a satisfactory negative smear history are at low risk for dyskaryosis and might be suitable for withdrawal from the cervical screening programme. The objectives of this study are to document the pattern of dyskaryosis in the cervical smears of women over 50 and to relate the risk of dyskaryosis in these women to the prior smear history. DESIGN: Available computerised smear data were analysed. SETTING: Five regions in England and Scotland; Aberdeen, Dundee, Birmingham, Gateshead and Nottingham. POPULATION: All women aged 50 or over who had a satisfactory smear between 1988 and 1996. METHODS: Smear results were sorted into individual smear records. The first smear after the age of 50 was identified as well as all smears before and after the age of 50. MAIN OUTCOME MEASUREMENTS: The smear history before and after the age of 50 was determined for all women. The relative risk of dyskaryosis as well as the time to dyskaryosis was calculated for women whose raw data were available. RESULTS: The study included 170,436 women with at least one satisfactory smear after 50. No results of previous smears were available in 90,546 (54%) of women but 36,512 (21%) of women had a satisfactory negative smear history. Women with prior dyskaryosis or borderline nuclear abnormalities (BNA) had an increased risk of dyskaryosis after the age of 50 compared with women with a negative history (RR 4.39 and 3.08 respectively). It was notable that 1.8% of women with a negative history still demonstrated subsequent dyskaryosis. CONCLUSIONS: Women with either dyskaryosis or BNA before 50 are not suitable for withdrawal from cervical screening. Well-screened women with a negative smear history at the age 50 still have a residual risk of subsequently developing a new abnormality.