Design of an adaptive randomized clinical trial of intravenous citrulline for sickle cell pain crisis in the emergency department.

Background: Vaso-occlusive pain crisis (VOC) is the most frequent cause for Emergency Department (ED) visits and hospital admissions for patients with sickle cell disease (SCD). Nitric oxide plays a critical role in the pathogenesis of vaso-occlusion. The amino acid, citrulline, is the main endothelial nitric oxide booster that offers the potential to ameliorate vaso-occlusion and decrease the risk of hospitalization. Objective: In this two-part study, the goal of the first part is to determine the pharmacokinetic profile of intravenous (IV) l-citrulline and optimal dose for the second part of the study, which is to determine the efficacy and tolerability of the intervention in patients with SCD. Design: A phase I/IIA open-label dose-finding study with subsequent double-blind, placebo-controlled, randomized Study of l-citrulline in children and adolescents with SCD presenting to the ED in VOC. Methods: Part 1: Subjects experiencing VOC are enrolled in an open-label, ascending dose of IV l-citrulline to identify the optimum dose with endpoints of pharmacokinetic parameters, pain scores, reduction of opioid use, quality of life, proportion admitted to the hospital for treatment of pain, readmission rates, and assessment of adverse events. Part 2 of the trial is a double-blind, placebo-controlled adaptive "pick-the-winner" design to evaluate the efficacy and tolerability of IV l-citrulline in patients with SCD while receiving standard of care therapy for VOC. Summary: This ED based sickle cell adaptive trial will determine the optimal dose for IV citrulline and whether the intervention improves outcome as a potential novel therapy for VOC in SCD.

Depression, anxiety and PTSD symptoms before and during the COVID-19 pandemic in the UK.

BACKGROUND: The impact of the coronavirus disease 2019 (COVID-19) pandemic on mental health is still being unravelled. It is important to identify which individuals are at greatest risk of worsening symptoms. This study aimed to examine changes in depression, anxiety and post-traumatic stress disorder (PTSD) symptoms using prospective and retrospective symptom change assessments, and to find and examine the effect of key risk factors. METHOD: Online questionnaires were administered to 34 465 individuals (aged 16 years or above) in April/May 2020 in the UK, recruited from existing cohorts or via social media. Around one-third (n = 12 718) of included participants had prior diagnoses of depression or anxiety and had completed pre-pandemic mental health assessments (between September 2018 and February 2020), allowing prospective investigation of symptom change. RESULTS: Prospective symptom analyses showed small decreases in depression (PHQ-9: -0.43 points) and anxiety [generalised anxiety disorder scale - 7 items (GAD)-7: -0.33 points] and increases in PTSD (PCL-6: 0.22 points). Conversely, retrospective symptom analyses demonstrated significant large increases (PHQ-9: 2.40; GAD-7 = 1.97), with 55% reported worsening mental health since the beginning of the pandemic on a global change rating. Across both prospective and retrospective measures of symptom change, worsening depression, anxiety and PTSD symptoms were associated with prior mental health diagnoses, female gender, young age and unemployed/student status. CONCLUSIONS: We highlight the effect of prior mental health diagnoses on worsening mental health during the pandemic and confirm previously reported sociodemographic risk factors. Discrepancies between prospective and retrospective measures of changes in mental health may be related to recall bias-related underestimation of prior symptom severity.

Gaslighting in alleged assault ascertained as Munchausen syndrome transcending to malingering in highly counterintuitive self inflicted acid burns.

A factitious disorder leading to the self-infliction of highly counter-intuitive burns was diagnosed in a middle-aged female. The injuries were otherwise alleged to have been sustained by assault inflicted upon her by an unknown person. The case was diagnosed by medico-legal interpretation of injuries, in spite of a highly deceptive and concocted history by the patient and her husband. The entity was unique in being associated with magnificent primary, secondary and tertiary gains. The exploitation of the morbid sequel to malinger by the patient, and the involvement of the husband for the prolongation of the illness of his wife for financial gains as gaslighting was highly unusual. The self-infliction of injuries over hands is seen in factitious disorder. However, a combination of a guarded self-immersion of the hands and feet in a corrosive by an illiterate female, followed by malingering to earn livelihood is unprecedented in factitious disorders. The delayed presentation which required amputation of all the limbs to save the life of the patient is a glaring highlight of this case.

Correction: Investigation of common, low-frequency and rare genome-wide variation in anorexia nervosa.

The fortieth author's name was listed incorrectly. The correct presentation is A Keski-Rahkonen.

Technical report: Inter- and intra-rater reliability of regional gastrointestinal transit times measured using the 3D-Transit electromagnet tracking system.

BACKGROUND: The 3D-Transit electromagnet tracking system is an emerging tool for the ambulatory assessment of gastrointestinal (GI) transit times and motility patterns, based on the anatomical localization of ingestible electromagnetic capsules. Currently, 3D-Transit recordings are manually analyzed to extract GI transit times. As this is a subjective method, there is some inherent variability in the measurements, which may be experience-dependent. We therefore assessed inter- and intra-rater reliability of GI transit times from 3D-Transit recordings. METHODS: Thirty-six 3D-Transit recordings (17 female; median age: 34 years [range: 21-80]) were analyzed twice by 3 raters with varying experience. Each rater manually identified the timestamps when a capsule progressed from antrum to duodenum, and from ileum to right colon. These timestamps, along with the ingestion and expulsion times, were used to determine whole gut (WGTT), gastric emptying (GET), small intestinal (SITT) and colonic (CTT) transit times. Reliability was determined using interclass correlation coefficients (ICCs). KEY RESULTS: For capsule progression timestamps, the most and mid-experienced raters had fair to good inter- and excellent intra-rater reliability (ICCmin-max  = 0.61-1.00), whereas the inexperienced rater had poor to fair inter- and poor intra-rater reliability (ICCmin-max  = 0.28-0.55). GET and SITT reliability between the most and mid-experienced raters was fair (ICCmin-max  = 0.61-0.73), while reliability between these raters and the inexperienced rater was poor to fair (ICCmin-max  = 0.28-0.55). CTT reliability was excellent between and within all raters (ICCmin-max  = 0.92-0.99). CONCLUSIONS & INFERENCES: Inexperienced raters provide the least reliable measurements from 3D-Transit recordings, which confirms requirement for adequate training. Automation may improve the reliability of measurements.

Investigation of common, low-frequency and rare genome-wide variation in anorexia nervosa.

Anorexia nervosa (AN) is a complex neuropsychiatric disorder presenting with dangerously low body weight, and a deep and persistent fear of gaining weight. To date, only one genome-wide significant locus associated with AN has been identified. We performed an exome-chip based genome-wide association studies (GWAS) in 2158 cases from nine populations of European origin and 15 485 ancestrally matched controls. Unlike previous studies, this GWAS also probed association in low-frequency and rare variants. Sixteen independent variants were taken forward for in silico and de novo replication (11 common and 5 rare). No findings reached genome-wide significance. Two notable common variants were identified: rs10791286, an intronic variant in OPCML (P=9.89 × 10-6), and rs7700147, an intergenic variant (P=2.93 × 10-5). No low-frequency variant associations were identified at genome-wide significance, although the study was well-powered to detect low-frequency variants with large effect sizes, suggesting that there may be no AN loci in this genomic search space with large effect sizes.

Structural effects of intra-articular TGF-ß1 in moderate to advanced knee osteoarthritis: MRI-based assessment in a randomized controlled trial.

BACKGROUND: To determine effects of allogeneic human chondrocytes expressing TGF-ß1 (TG-C) on structural progression of MRI features of knee osteoarthritis over a 1 year period. METHODS: This phase II randomized controlled trial of TG-C included patients with moderate to advanced osteoarthritis. Patients were randomized to receive an intraarticular 3:1 mixture of non-transduced allogeneic human chondrocytes and TG-C or placebo. 3 T MRI was acquired for all patients at baseline and follow-up (3, 6 and 12 months). MRIs were assessed using the WORMS system including cartilage damage, bone marrow lesions (BMLs), meniscal damage/extrusion, Hoffa-, effusion-synovitis, and osteophytes. Analyses were performed on a whole knee level, compartmental level, and subregional level. Binary logistic regression with Generalized Estimating Equation was used to compare risks of progression, adjusting for baseline age and gender. Mann - Whitney - Wilcoxon tests were used to assess differences for continuous variables. RESULTS: Fifty-seven Patients were included in the TG-C group and 29 in the placebo group. At 12 months, knees in the TG-C group showed less progression of cartilage damage compared to placebo on a whole knee level (34.6% vs. 47.9%; adjusted RR 0.7, 95%CI [0.5-1.1], p = 0.077). Less progression of Hoffa-synovitis and effusion-synovitis was observed in the TG-C group compared to placebo (9.6% vs. 21.1%, adjusted RR 0.5, 95%CI [0.2,1.2], p = 0.115). No statistically significant differences were seen for BMLs, meniscal damage and osteophytes. CONCLUSIONS: Intraarticular treatment with TG-C showed fewer patients in the treated group with progression in structural OA features and other MRI-defined inflammatory markers such as Hoffa-synovitis and effusion-synovitis. However, no differences were observed in regard to progression of BMLs and meniscal damage, or hypertrophic osteophyte formation. TRIAL REGISTRATION: NCT01221441 .Registered 13th October, 2010.

Pathway based analysis of genotypes in relation to alcohol dependence.

We introduce a method for detecting variants in several genes of related function with small effect on a phenotype of interest. Our method uses logistic regression to test whether multiple alleles within a functional set have significantly higher than expected predictive value, even though none individually may have strong individual effects. We illustrate this method by testing seven gene sets (including 48 genes), from a study with 1350 single nucleotide polymorphisms in 130 addiction candidate genes studied in a sample of 575 alcohol dependence (AD) cases and 530 controls. We conclude that AD is related to variation in genes participating in Glutamate and γ-amino butyric acid signaling, as has been reported elsewhere, and in stress response pathways, but not with genes in several other systems implicated in other drugs of abuse.

A threonine to isoleucine missense mutation in the pericentriolar material 1 gene is strongly associated with schizophrenia.

Markers at the pericentriolar material 1 gene (PCM1) have shown genetic association with schizophrenia in both a University College London (UCL) and a USA-based case-control sample. In this paper we report a statistically significant replication of the PCM1 association in a large Scottish case-control sample from Aberdeen. Resequencing of the genomic DNA from research volunteers who had inherited haplotypes associated with schizophrenia showed a threonine to isoleucine missense mutation in exon 24 which was likely to change the structure and function of PCM1 (rs370429). This mutation was found only as a heterozygote in 98 schizophrenic research subjects and controls out of 2246 case and control research subjects. Among the 98 carriers of rs370429, 67 were affected with schizophrenia. The same alleles and haplotypes were associated with schizophrenia in both the London and Aberdeen samples. Another potential aetiological base pair change in PCM1 was rs445422, which altered a splice site signal. A further mutation, rs208747, was shown by electrophoretic mobility shift assays to create or destroy a promoter transcription factor site. Five further non-synonymous changes in exons were also found. Genotyping of the new variants discovered in the UCL case-control sample strengthened the evidence for allelic and haplotypic association (P=0.02-0.0002). Given the number and identity of the haplotypes associated with schizophrenia, further aetiological base pair changes must exist within and around the PCM1 gene. PCM1 protein has been shown to interact directly with the disrupted-in-schizophrenia 1 (DISC1) protein, Bardet-Biedl syndrome 4, and Huntingtin-associated protein 1, and is important in neuronal cell growth. In a separate study we found that clozapine but not haloperidol downregulated PCM1 expression in the mouse brain. We hypothesize that mutant PCM1 may be responsible for causing a subtype of schizophrenia through abnormal cell division and abnormal regeneration in dividing cells in the central nervous system. This is supported by our previous finding of orbitofrontal volumetric deficits in PCM1-associated schizophrenia patients as opposed to temporal pole deficits in non-PCM1-associated schizophrenia patients. Caution needs to be exercised in interpreting the actual biological effects of the mutations we have found without further cell biology. However, the DNA changes we have found deserve widespread genotyping in multiple case-control populations.

Alcohol dependence is associated with the ZNF699 gene, a human locus related to Drosophila hangover, in the Irish Affected Sib Pair Study of Alcohol Dependence (IASPSAD) sample.

Because tolerance is an important aspect of alcohol dependence (AD) in humans, recent evidence showing that the Drosophila gene hang is critically involved in the development of alcohol tolerance in the fly suggests that variation in related human loci might be important in the etiology of alcohol-related disorders. The orthology of hang in mammals is complex, but a number of human gene products (including ZNF699) with similar levels of amino-acid identity (18-26%) and similarity (30-41%), are consistently identified as the best matches with the translated hang sequence. We tested for association between the dichotomous clinical phenotype of alcohol dependence and seven single nucleotide polymorphisms (SNPs) in ZNF699 in our sample of 565 genetically independent cases and 496 siblings diagnosed with AD, and 609 controls. In analyses of genetically independent cases and controls, four of the seven single markers show strong evidence for association with AD (0.00003

Fine mapping of a susceptibility locus for bipolar and genetically related unipolar affective disorders, to a region containing the C21ORF29 and TRPM2 genes on chromosome 21q22.3.

Linkage analyses of bipolar families have confirmed that there is a susceptibility locus near the telomere on chromosome 21q. To fine map this locus we carried out tests of allelic association using 30 genetic markers near the telomere at 21q22.3 in 600 bipolar research subjects and 450 ancestrally matched supernormal control subjects. We found significant allelic association with the microsatellite markers D21S171 (P=0.016) and two closely linked single-nucleotide polymorphisms, rs1556314 (P=0.008) and rs1785467 (P=0.025). A test of association with a three locus haplotype across the susceptibility region was significant with a permutation test of P=0.011. A two SNP haplotype was also significantly associated with bipolar disorder (P=0.01). Only two brain expressed genes, TRPM2 and C21ORF29 (TSPEAR), are present in the associated region. TRPM2 encodes a calcium channel receptor and TSPEAR encodes a peptide with repeats associated with epilepsy in the mouse. DNA from subjects who had inherited the associated marker alleles was sequenced. A base pair change (rs1556314) in exon 11 of TRPM2, which caused a change from an aspartic acid to a glutamic acid at peptide position 543 was found. This SNP showed the strongest association with bipolar disorder (P=0.008). Deletion of exon 11 of TRPM2 is known to cause dysregulation of cellular calcium homeostasis in response to oxidative stress. A second nonconservative change from arginine to cysteine at position 755 in TRPM2 (ss48297761) was also detected. A third nonconservative change from histidine to glutamic acid was found in exon 8 of TSPEAR. These changes need further investigation to establish any aetiological role in bipolar disorder.

Genome scan of Tourette syndrome in a single large pedigree shows some support for linkage to regions of chromosomes 5, 10 and 13.

OBJECTIVES: To localize genes influencing the susceptibility to Gilles de la Tourette syndrome (GTS) and associated chronic multiple tics (CMT). METHOD: A single, large, multiple affected pedigree containing 35 subjects diagnosed with GTS and a further 14 with CMT was genotyped for markers spanning the autosomes. Linkage analysis was carried out using classical lod score analysis and model-free lod score analysis. All markers were subjected to two-point analysis, and markers producing a two-point result significant at P<0.005 were subjected to three-point analysis using adjacent markers. RESULTS: The following markers produced at least one result significant at 0.005 using two-point analysis: D5S1981, D5S2050, D10S591, D10S189, D13S217, and D14S288. Three-point analysis with D5S2050 and D5S400 produced a lod of 2.9 with CMT. Three-point analysis of D10S591 and D10S189 produced lods of 1.9 with GTS and CMT. Three-point analysis of D13S217 and D13S171 produced a lod of 2.7 with GTS. No single haplotype appeared to account for the majority of cases within the pedigree. CONCLUSIONS: It seems likely that more than one susceptibility allele is present in the pedigree. Although none of the three positive regions is conclusively implicated, it seems probable that at least one contains a susceptibility locus. We recommend that association-based studies be carried out in these three regions to produce further evidence for a localization and to carry out fine-mapping.

Variation in the DCP1 gene, encoding the angiotensin converting enzyme ACE, is not associated with increased susceptibility to Alzheimer's disease.

OBJECTIVES: To attempt to replicate previous reports that polymorphic variation in the DCP1 gene causes increased susceptibility to the development of Alzheimer's disease, either on its own or in interaction with the effects of the gene for apolipoprotein E (APOE). METHOD: Subjects older than 65 years of age consisting of 81 dementia patients diagnosed as having possible or probable Alzheimer's disease and 68 controls were obtained from Camden, Islington and Harlow psychiatric services. Subjects were genotyped for APOE alleles e2, e3 and e4, and the common insertion/deletion polymorphisms for DCP1* I/D were genotyped. RESULTS: There was no statistically significant difference in the frequency of the DCP1* insertion/deletion alleles between the cases and controls (X2 =0.04, 1 degree of freedom, not significant). When subjects were subdivided according to whether they possessed at least one copy of the APOE e4 allele, there were still no differences in DCP1 allele frequencies between cases and controls. CONCLUSIONS: Further research is needed to elucidate any role that the DCP1 polymorphism may play in relation to Alzheimer's disease. Previous studies may be false positive, or inconsistency in replication may be due to heterogeneity.

Genetic association studies of schizophrenia using the 8p21-22 genes: prepronociceptin (PNOC), neuronal nicotinic cholinergic receptor alpha polypeptide 2 (CHRNA2) and arylamine N-acetyltransferase 1 (NAT1).

Schizophrenia is a common, genetically heterogeneous disorder with a lifetime prevalence of approximately 1% in the general population. Linkage studies of affected families have now strongly implicated a susceptibility locus on chromosome 8p21-22. Tests of allelic association with markers on 8p21-22 should be able to localise any quantitative trait nucleotides (QTN's) or susceptibility mutations to within a few hundred kilobases. Three brain expressed candidate susceptibility genes, prepronociceptin (PNOC), neuronal cholinergic receptor, nicotinic, alpha polypeptide 2 (CHRNA2) and arylamine N-acetyltransferase 1 (NAT1) have been mapped to chromosome 8p21-22. A case-control, allelic association study was performed using a novel highly polymorphic dinucleotide repeat, D8S2611 near the PNOC gene, two previously characterised dinucleotide repeats, D8S131 and D8S131P at the CHRNA2 locus and an RFLP at the 3'UTR of the arylamine N-acetyltransferase 1 (NAT1) gene. No differences were found in allele frequencies between the patient and control groups. DNA variations or mutations at or near the three genes under study are unlikely to increase susceptibility to schizophrenia in our population sample.

Genomewide genetic linkage analysis confirms the presence of susceptibility loci for schizophrenia, on chromosomes 1q32.2, 5q33.2, and 8p21-22 and provides support for linkage to schizophrenia, on chromosomes 11q23.3-24 and 20q12.1-11.23.

We have performed genetic linkage analysis in 13 large multiply affected families, to test the hypothesis that there is extensive heterogeneity of linkage for genetic subtypes of schizophrenia. Our strategy consisted of selecting 13 kindreds containing multiple affected cases in three or more generations, an absence of bipolar affective disorder, and a single progenitor source of schizophrenia with unilineal transmission into the branch of the kindred sampled. DNA samples from these families were genotyped with 365 microsatellite markers spaced at approximately 10-cM intervals across the whole genome. We observed LOD scores >3.0 at five distinct loci, either in the sample as a whole or within single families, strongly suggesting etiological heterogeneity. Heterogeneity LOD scores >3.0 in the sample as a whole were found at 1q33.2 (LOD score 3.2; P=.0003), 5q33.2 (LOD score 3.6; P=.0001), 8p22.1-22 (LOD score 3.6; P=.0001), and 11q21 (LOD score 3.1; P=.0004). LOD scores >3.0 within single pedigrees were found at 4q13-31 (LOD score 3.2; P=.0003) and at 11q23.3-24 (LOD score 3.2; P=.0003). A LOD score of 2.9 was also found at 20q12.1-11.23 within in a single family. The fact that other studies have also detected LOD scores >3.0 at 1q33.2, 5q33.2, 8p21-22 and 11q21 suggests that these regions do indeed harbor schizophrenia-susceptibility loci. We believe that the weight of evidence for linkage to the chromosome 1q22, 5q33.2, and 8p21-22 loci is now sufficient to justify intensive investigation of these regions by methods based on linkage disequilibrium. Such studies will soon allow the identification of mutations having a direct effect on susceptibility to schizophrenia.

Localization of a Nod factor-binding protein in legume roots and factors influencing its distribution and expression.

The roots of the legume Dolichos biflorus contain a lectin/nucleotide phosphohydrolase (Db-LNP) that binds to the Nod factor signals produced by rhizobia that nodulate this plant. In this study we show that Db-LNP is differentially distributed along the surface of the root axis in a pattern that correlates with the zone of nodulation of the root. Db-LNP is present on the surface of young and emerging root hairs and redistributes to the tips of the root hairs in response to treatment of the roots with a rhizobial symbiont or with a carbohydrate ligand. This redistribution does not occur in response to a non-symbiotic rhizobial strain or a root pathogen. Db-LNP is also present in the root pericycle where its level decreases upon initiation of nodule formation. Maximum levels of Db-LNP are found in 2-d-old roots, and the expression of this root protein is increased when the plants are grown in the absence of NO(3)(-) and NH(4)(+). These results support the possibility that Db-LNP is involved in the initiation of the Rhizobium legume symbiosis.

Primary care perspective of prostate cancer screening after national guidance: a questionnaire survey.

National guidance (executive letter) EL(97)12 stated that population screening should not be provided by the NHS, or be offered to the public until there is effective screening technology for prostate cancer. The study set out to determine the views of general practitioners and, indirectly, their practice staff on prostate cancer screening in primary care upon receiving EL(97)12. This postal questionnaire survey reveals that 81% (95% CI 75% to 87%) of responding general practitioners in North Staffordshire agreed with EL(97)12 and one in ten said that the executive letter changed their views, suggesting that such national guidance has an effect.