Population-based association studies are used to identify common susceptibility variants for complex genetic traits. These studies are susceptible to confounding from unknown population substructure. Here we apply a model-based clustering approach to our case-control study of stroke among young women to examine if self-reported ethnicity can serve as a proxy for genetic ancestry.Structure program was used to iteratively evaluate for K = 1 to 5 potential genetic-based subpopulations. Evaluating the population as a whole, the Structure output plateaued at K = 2 clusters. 98% of self-reported Caucasians had an estimated probability ≥50% of belonging to Cluster 1, while 94% of self-reported African-Americans had an estimated probability ≥50% of belonging to Cluster 2. Stratifying the participants by self-reported ethnicity and repeating the analyses revealed the presence of two clusters among Caucasians, suggesting that potential substructure may exist.A population-based case-control study of stroke among women aged 15-49 identified 361 cases of first ischemic stroke and 401 age-comparable control subjects. Thirty single nucleotide polymorphisms (SNPs) throughout the genome unrelated to stroke risk and with established ancestry-based allele frequency differences were genotyped in all participants. The Among our combined sample of African-American and Caucasian participants there is no large unknown subpopulation and self-reported ethnicity can serve as a proxy for genetic ancestry. Ethnicity-specific analyses indicate that population substructure may exist among the Caucasian participants indicating that further studies are warranted. Population-based case-control studies are used to identify common susceptibility variants for complex genetic traits; however, population stratification may confound their results ,2. PopulThe Stroke Prevention in Young Women (SPYW) Study is a population-based case-control study initiated to examine risk factors for first ischemic stroke in women aged 15-49. All participants were identified from the same population including all of Maryland (except the far Western panhandle), Washington DC, and the southern portions of both Pennsylvania and Delaware. The methods for discharge surveillance, chart abstraction, and case adjudication have been described previously  includin2 > 10) in allele frequencies between individuals from Utah with European ancestry (CEU) and individuals from Nigeria (YRI) [Twenty ancestry informative markers (i.e. SNPs) were chosen from a HapMap panel previously shown to differ (χia (YRI) . Ten addGenotyping was conducted using DNA isolated from whole blood using the QIAamp DNA Blood Maxi Kit . SNP genotyping was performed by either TaqMan  or iPLEX  methodologies. For each SNP, genotyping for all cases and controls was performed on the same platform.2 test). Major allele frequency differences between self-reported Caucasians and African-Americans were calculated (χ2 test). Analyses were performed using SAS®, Version 9.1   exhibited genotype call rates less than 80% and one SNP (rs2695) did not exhibit a difference in allele frequencies between our Caucasian and African-American populations. Hence, 26 SNPs distributed throughout the genome were included in the analyses ) was generated. Similar self-reported ethnicity-specific analyses were also performed.Model-based clustering for inferring population structure was performed using software . An admiStructure was implemented to assess information about the distribution of Q, the estimated membership coefficients for each individual in each cluster. When this function is activated, the output file includes the left- and right-hand ends of the probability intervals for each q(i). The ANCESTDIST (Boolean) function of Demographic and risk factor characteristics by case-control status are described in Table 2 comparison values.Table Structure output (log Pr (X | K) (denoted in Table Table 1) log Pr (X | K) plateaus at K = 2.2) Dirichlet parameter for amount of admixture (α) converges to a value < 0.2 once the Markov chain converges.3) Most individuals are strongly assigned to one of the two populations.Figure Structure ANCESTDIST option provided the 90% probability intervals for each individual. Of the 760 individuals, 130 (17%) have overlapping probability intervals. Hence, 83% of the study population demonstrated individual ancestry proportion estimates that had non-overlapping 90% probability intervals.The Ethnicity specific exploratory analyses (demonstrated in Table Our results indicate that among the combined sample of African-American and Caucasian participants, self-reported ethnicity can serve as a proxy for genetic ancestry or relatedness. Furthermore, no large unknown subpopulation was identified. The ethnicity-specific analyses demonstrate no clear substructure in self-reported African American participants. This differs from the accepted idea that greater genetic diversity, as measured by linkage disequilibrium, is seen in populations of African origin. The lack of substructure in our African-American participants may be related to limitations of our panel. Interestingly, the ethnicity-specific analyses do demonstrate that some population substructure may exist among self-reported Caucasian participants. Evaluation of substructure in Americans of European decent has shown a course separation of European populations along a northeast to southwest axis . In thisIn summary, among the combined population, a small number of individuals were genetically more consistent with the other ancestry. Specifically, with a 50% ancestry threshold, 22 self-reported African-Americans were more consistent with Caucasian ancestry, while 10 self-reported Caucasians were more consistent with African-American ancestry. This information may be incorporated into future association analyses in various ways. Individuals not satisfying an ethnicity-based ancestry threshold could simply be removed from the study. Alternatively, as mentioned above, more null markers could be genotyped to improve the ancestry classification. Lastly, a variable incorporating percentage of ancestry could be introduced into the association analyses.Among our combined sample of African-American and Caucasian participants there is no large unknown subpopulation and self-reported ethnicity can serve as a proxy for genetic ancestry or relatedness. Ethnicity-specific analyses indicate that population substructure may exist among the Caucasian participants indicating that further studies are warranted.The authors declare that they have no competing interests.All authors certify that they participated in the conceptual design of this work, the analysis of the data, and the writing of the manuscript to take public responsibility for it. All authors reviewed the final version of the manuscript and approve it for publication. JBM, JWC, BDM and SJK: participated in the writing of the initial draft. JWC, MAW, BJS and SJK: participated in data collection. JBM, TDH and OCS: participated in the genotyping. JBM, JWC, JRO, LRM, BDM, and JDS: participated in the data analysis. All authors provided critiques of the final manuscript.


Background: Alzheimer's disease (AD) is the major cause of dementia in population aged over 65 years, accounting up to 70% dementia cases. However, validated peripheral biomarkers for AD diagnosis are not available up to present. In this study, we adopted a new strategy of combination of computational prediction and experimental validation to identify blood protein biomarkers for AD.Methods: First, we collected tissue-based gene expression data of AD patients and healthy controls from GEO database. Second, we analyzed these data and identified differentially expressed genes for AD. Third, we applied a blood-secretory protein prediction program on these genes and predicted AD-related proteins in blood. Finally, we collected blood samples of AD patients and healthy controls to validate the potential AD biomarkers by using ELISA experiments and Western blot analyses.Results: A total of 2754 genes were identified to express differentially in brain tissues of AD, among which 296 genes were predicted to encode AD-related blood-secretory proteins. After careful analysis and literature survey on these predicted blood-secretory proteins, ten proteins were considered as potential AD biomarkers, five of which were experimentally verified with significant change in blood samples of AD vs. controls by ELISA, including GSN, BDNF, TIMP1, VLDLR, and APLP2. ROC analyses showed that VLDLR and TIMP1 had excellent performance in distinguishing AD patients from controls . Further validation of VLDLR and TIMP1 by Western blot analyses has confirmed the results obtained in ELISA experiments.Conclusion: VLDLR and TIMP1 had better discriminative abilities between ADs and controls, and might serve as potential blood biomarkers for AD. To our knowledge, this is the first time to identify blood protein biomarkers for AD through combination of computational prediction and experimental validation. In addition, VLDLR was first reported here as potential blood protein biomarker for AD. Thus, our findings might provide important information for AD diagnosis and therapies. Alzheimer's disease (AD) is the major cause of dementia in population aged over 65 years, accounting up to 70% dementia cases . This di42 peptide and increased concentration of tau proteins in CSF of AD patients compared to controls might work as diagnostic biomarkers for AD  patients who later progressed to AD from those unchanged or converted to other dementia . PraticoIn this study, we conducted a combination of computational prediction and experimental validation to identify potential blood protein biomarkers for AD. We firstly analyzed previously published gene expression data of brain tissues from AD patients to identify differentially expressed genes for AD. Furthermore, we applied a blood-secretory protein prediction program on these genes to predict AD-related proteins in blood. Finally, several potential blood protein biomarkers for AD were selected and verified by enzyme-linked immunosorbent assay (ELISA) experiments and Western blot analyses on blood samples from AD patients and healthy controls. This work provides a more specific and effective way to investigate blood protein biomarkers for AD.The schematic diagram of the workflow in this study was given as Figure Brain tissue-based gene expression data of AD patients were collected from GEO database . Two sert-test would be used to detect DEPs. However, our results showed that the values of many examined probes did not fit normal distribution, Wilcoxon rank sum test  was computed across samples for each probe. As a whole, probes with q-value < 0.05 and FC > 1.2 were considered up-regulated, and those with q-value < 0.05 and FC < 0.833 were down-regulated. Finally, we chose the differentially expressed probes with consistent change trend in these two datasets to map to their corresponding genes, which were considered to be differentially expressed genes for AD.We first identified differentially expressed probes (DEPs), and then mapped these probes to their genes. The following procedure was used to identify DEPs for each dataset. Kolmogorov–Smirnov test  was usedsum test  was applHochberg  method wAll differentially expressed genes were analyzed for prediction whether their protein products could be secreted into blood through a program developed by Juan Cui et al . The basIn this work, ELISA experiments were carried out on blood samples from AD patients and healthy individuals to validate the predicted blood protein biomarkers for AD. The research protocol of this study was approved by the Human Research Ethics Committee of Shenzhen University and had been performed in accordance with the ethical standards. A total of 123 subjects were enrolled in experiment from Shenzhen People's Hospital and the Eighth Affiliated Hospital of Sun Yat-sen University, including 54 AD patients and 69 healthy subjects. Informed consents were obtained from all participants in accordance with the Declaration of Helsinki prior to their inclusion in this study. All the patients were diagnosed by neuropsychiatrists in the hospital according to the criteria of Diagnostic and Statistical Manual of Mental Disorders-Fourth Edition (DSM-IV). The average age of the patients and controls were 74.3 (ranged from 52 to 93) and 73.9 (ranged from 53 to 94), respectively. The ratio of male to female was about 2:3. In each ELISA experiment, blood samples were selected from AD patients and age- and gender-matched healthy controls. Blood samples (5 ml) were collected using glass tubes. Serums were separated by centrifugation at 3000 g for 10 min, and then subdivided into aliquots and stored at −80°C for further use.For ELISA experiments, commercial ELISA kits for proteins gelsolin (GSN), brain-derived neurotrophic factor (BDNF), metalloproteinase inhibitor 1 (TIMP1), pigment epithelium-derived factor (SERPINF1) and amyloid-like protein 2 (APLP2) were bought from Uscn Life Science Inc. . The catalog numbers of these ELISA kits were SEA372Hu, SEA011Hu, SEA552Hu, SEB972Hu, and SEG122Hu, respectively. Additionally, ELISA kits of inositol 1,4,5-trisphosphate receptor-interacting protein (ITPRIP), transmembrane emp24 domain-containing protein 10 (TMED10), very low-density lipoprotein receptor (VLDLR), mitogen-activated protein kinase 8 (MAPK8) and mitogen-activated protein kinase 1 (MAPK1) were bought from Sbj Biological technology Co., Ltd.  with catalog numbers of SBJ-H2157, SBJ-H2158, SBJ-H1100, SBJ-H2160, and SBJ-H2161, respectively. The concentrations of these proteins were measured under the manufacturer's instructions. The total protein concentrations of samples were determined using bicinchoninic acid (BCA) protein assay kit with product No. 23227 .T-test was applied to make differential analysis on normalized protein concentrations of AD samples vs. controls, and then FDR  curve analysis was carried out to evaluate the power of these proteins in distinguishing AD samples from healthy controls, which was generated by using package pROC on R  were separated by SDS-PAGE on 12% polyacrylamide gels. After electrophoresis, the proteins were transferred onto 0.2 μm polyvinylidene fluoride (PVDF) membranes , and the membranes were blocked with 5% nonfat-dried milk in Tris-buffered saline  for 1 h and then washed with TBS containing 0.4% (v/v) tween 20 (TBST), followed by incubation with primary antibodies  against VLDLR and TIMP1 overnight at 4°C and horseradish peroxidase (HRP)-conjugated secondary antibody  for 2 h at room temperature. The membranes were washed three times each for 10 min in TBST and developed with enhanced chemiluminescence (ECL) kit . Immunoreactive signals were detected using a Kodak Image Station 4000M imaging system . Quantitative analysis was performed on the protein bands by ImageJ analysis software . Equal amount of proteins were separated by SDS-PAGE and stained with Coomassie blue, which was used as the loading control.t-test to examine any significant differences between ADs and controls by GraphPad Prism 7 software  and presented as the means ± the standard errors of the means (SEM). Differences were considered significant with p-value < 0.05.The data of Western blot were analyzed using the two-tailed Student's Two brain tissue-based gene expression datasets of AD patients were downloaded from GEO database. There were 5481 DEPs (2511 up-regulated and 2970 down-regulated) identified in GSE48350 and 12115 DEPs (4675 up-regulated and 7440 down-regulated) in GSE5281. Further comparing analysis was made on these two groups of DEPs, and 1545 probes (corresponding to 1186 genes) and 1981 probes (corresponding to 1568 genes) were found consistently up- and down-regulated in these two datasets, respectively . In addiIt is well known that blood-brain barrier (BBB) controls substances exchange strictly between brain and blood. However, some evidence indicates that breakdown of BBB may account for AD occurrence or aggravation and could enhance the movement of proteins between brain and blood in either direction , 30. TheTo gain a comprehensive understanding of these predicted AD-related blood-secretory proteins, we carried out GO enrichment analysis using DAVID . A varieTo further choose precise and important candidate biomarkers for AD, we manually checked the relationship between these proteins and AD through database and literature studies. First, we collected a total of 1493 AD-related genes from three databases, 1291 from GAD , 169 froIn order to explore the relationship between these 35 proteins and AD pathology, we made a protein-protein interaction analysis through the online sever LENS . A netwoBased on the gene expression levels of these 35 proteins in AD samples and their functional annotations, 10 proteins were chosen for experimental verification. They are GSN, BDNF, TIMP1, SERPINF1, ITPRIP, TMED10, VLDLR, MAPK8, APLP2, and MAPK1.ELISA experiments were performed to examine the protein levels in blood samples from AD patients and healthy controls. Figure ROC curve analyses were used to evaluate the performance of the five significantly changed proteins in distinguishing AD samples from controls Figure . We founBased on the ELISA analyses, VLDLR and TIMP1 were chosen for further validation of their abilities in identifying the samples of AD patients by Western blot analyses. The serum samples of 5 AD patients and 5 age- and gender-matched healthy controls were used to detect the expression levels of these two proteins. After densitometry analysis on Western blots, VLDLR and TIMP1 were found down- and up-regulated in AD patients respectively as shown in Figure AD is the major cause of dementia. However, there are no valid biomarkers for AD diagnosis in blood so far. In this study, we searched for potential protein biomarkers of AD in blood through computational prediction combined with experimental verification. Based on this strategy, we predicted 296 AD-related blood-secretory proteins, which were predominant enriched in protein phosphorylation, microtubule-based process, mitochondria and MAP kinase activity. As widely known, AD is characterized by neurodegenerative plaques and neurofibrillary tangles in brain . Tau proAfter careful analyses on these 296 proteins, 10 proteins were chosen for experimental validation by ELISA. Five proteins  were verified to be differentially expressed in AD patients vs. controls, suggesting that they might serve as potential biomarkers for AD in blood. Among them, GSN, BDNF, and TIMP1 have been reported to be potential blood protein biomarkers for AD in previous studies , 48, 49,GSN was reported to be implicated in AD due to its level changed with AD progression . GSN couBDNF could support the survival of existing neurons and encourage the growth and differentiation of new neurons and synapses , 53. PreTIMP1 is a tissue inhibitor of MMP9 and plays an important role in the development of AD for its function of inflammatory mediation . MMP9 waVLDLR is an apolipoprotein E receptor involved in synaptic plasticity, learning, and memory . It was APLP2, an APP like protein, could bind to synaptic signaling molecules exhibiting synaptogenic activity . FurtherAs a whole, this novel biomarker discovery strategy, namely computational prediction combined with experimental verification, provides some potential blood biomarkers for AD. To our knowledge, this is the first report to use such a strategy for AD blood biomarker discovery. Meanwhile, VLDLR is the first time reported here as potential protein biomarker for AD in blood. In addition, this strategy for biomarker discovery could also be used for discovering biomarkers of other nervous system diseases such as Parkinson's disease. Worth noting, this method provides an effective way to find pathology-associated biomarkers in blood, but there are still some shortages in this strategy that could affect our results. For example, there might be some false positive blood-secretory proteins coming from the computational prediction, so the sensitivity of the blood-secretory protein predictor need to be improved in the future. Additionally, gene expression changes in ADs vs. controls could not accurately reflect their proteins' expression changes, so the predicted proteins need to be validated on large scale blood samples further.A total of 2754 genes were identified differentially expressed in brain tissues of AD, among which 296 genes were predicted to encode blood-secretory proteins. GO enrichment analysis on the predicted blood-secretory proteins suggested that they were associated with AD and might act as candidate protein biomarkers of AD in blood. Furthermore, ten proteins were chosen for validation by ELISA and five proteins  were validated changed significantly in serum samples of AD vs. controls. ROC curves analyses on these five proteins showed that VLDLR and TIMP1 were with more power in distinguishing AD samples from controls. Western blot analyses on VLDLR and TIMP1 were further revealed that they might serve as potential blood biomarkers for AD. Obviously, further studies are required to confirm these findings.FY, QL, and JN conceived and designed this study. YZ collected data from database and literature. KZ and SX designed and performed the experimental work. FY and LS processed the data and carried out the statistical analysis. YG and AL recruited and diagnosed the patients and provided the blood samples. FY and LS wrote the manuscript. All authors have read and proved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Dizziness is common among older people and is associated with a cascade of debilitating symptoms, such as reduced quality of life, depression, and falls. The multifactorial aetiology of dizziness is a major barrier to establishing a clear diagnosis and offering effective therapeutic interventions. Only a few multidisciplinary interventions of dizziness have been conducted to date, all of a pilot nature and none tailoring the intervention to the specific causes of dizziness. Here, we aimed to test the hypothesis that a multidisciplinary dizziness assessment followed by a tailored multifaceted intervention would reduce dizziness handicap and self-reported dizziness as well as enhance balance and gait in people aged 50 years and over with dizziness symptoms.n = 151) or to a tailored, multifaceted intervention (n = 154) comprising one or more of the following: a physiotherapist-led vestibular rehabilitation programme (35% [n = 54]), an 8-week internet-based cognitive-behavioural therapy (CBT) (19% [n = 29]), a 6-month Otago home-based exercise programme (24% [n = 37]), and/or medical management (40% [n = 62]). We were unable to identify a cause of dizziness in 71 participants . Primary outcome measures comprised dizziness burden measured with the Dizziness Handicap Inventory (DHI) score, frequency of dizziness episodes recorded with monthly calendars over the 6-month follow-up, choice-stepping reaction time (CSRT), and gait variability. Data from 274 participants  were included in the intention-to-treat analysis. At trial completion, the DHI scores in the intervention group  were significantly reduced compared with the control group (pre and post mean [SD]: 23.0 [15.8] and 21.8 [16.4]), when controlling for baseline scores . There were no significant between-group differences in dizziness episodes , CSRT performance , and step-time variability during gait . No serious intervention-related adverse events occurred. Study limitations included the low initial dizziness severity of the participants and the only fair uptake of the falls clinic  and the CBT interventions.We conducted a 6-month, single-blind, parallel-group randomized controlled trial in community-living people aged 50 years and over who reported dizziness in the past year. We excluded individuals currently receiving treatment for their dizziness, those with degenerative neurological conditions including cognitive impairment, those unable to walk 20 meters, and those identified at baseline assessment with conditions that required urgent treatment. Our team of geriatrician, vestibular neuroscientist, psychologist, exercise physiologist, study coordinator, and baseline assessor held case conferences fortnightly to discuss and recommend appropriate therapy (or therapies) for each participant, based on their multidisciplinary baseline assessments. A total of 305 men and women aged 50 to 92 years  were randomly assigned to either usual care  of dizziness can improve dizziness in middle-aged and older community-dwellers.We assessed 305 community-dwellers aged 50 years and above who reported dizziness in the past year on a range of questionnaires and tests of sensory and physical function, mental health, quality of life, and cardiovascular function, at baseline and 6 months.n = 151) or to a 6-month tailored, multifaceted intervention (n = 154) comprising one or more of the following: vestibular rehabilitation (35% [n = 54]), cognitive-behavioural therapy (CBT) for anxiety and depression (19% [n = 29]), a home-based exercise programme to train balance and lower-limb strength (24% [n = 37]), and/or medical management (40% [n = 62]).We randomly assigned the participants to usual care .We suggest that community-based dizziness clinics use existing healthcare services to implement tailored and multifaceted dizziness interventions based on multidisciplinary assessments. Dizziness is common in older people: prevalence rates in the community range between 10% and 30%  and incrThere is good evidence from Cochrane reviews, systematic reviews, and large randomized controlled trials for effective therapies addressing aspects of each disorder: vestibular rehabilitation for peripheral vestibular disorders  , syncopeOlder people often have symptomatology suggestive of more than one subtype of dizziness –21. Yet In summary, despite significant evidence of the multifactorial nature of dizziness with advancing age and the availability of a range of evidence-based therapies addressing multiple aspects of dizziness, no study to date has investigated the effectiveness of a multidisciplinary intervention to improve dizziness in the general older population. Therefore, within a randomized controlled trial design, we designed a comprehensive, multidisciplinary battery of vestibular, cardiovascular, neuromuscular, balance, and psychological assessments to improve the likelihood of obtaining a diagnosis for the symptom of dizziness in middle-aged and older people. The primary aim was to compare the effect of a tailored multifaceted intervention versus usual care on frequency of dizziness episodes, dizziness handicap, balance, and gait in people aged 50 years and over with self-reported dizziness. Secondary aims were to determine the effects of the programme on physiological falls risk, cardiovascular, and psychological measures associated with dizziness.https://bmcgeriatr.biomedcentral.com/articles/10.1186/s12877-017-0450-3. The study protocol is registered in the Australia New Zealand Clinical Trials Registry ACTRN12612000379819. A data safety monitoring committee involving a NeuRA senior scientist not involved in the study and the resident biostatistician oversaw the study.We conducted a 6-month single-blind, parallel-group randomized controlled trial in a single research centre  in Sydney, Australia. The trial was approved by the Human Research Ethics Committee of the University of New South Wales (HC 12152). All eligible participants provided written informed consent prior to participating in the study. Details of the protocol and study design have been reported previously  and can We recruited community-living middle-aged and older adults through advertisements; flyers in community facility, hospital, and university noticeboards; articles in newspapers and newsletters for older people; the NeuRA website, newsletter, and mailing list; and by mailbox drops within the local community. After an initial phone screening, eligible individuals were invited to participate if they (i) were aged 50 years and over, (ii) had experienced at least one significant episode of dizziness in the past 12 months, (iii) lived independently in the community, and (iv) were able to understand the English language. Individuals were excluded if they (i) had a degenerative neurological condition, (ii) were currently receiving treatment for their dizziness, (iii) had a cognitive impairment  , and/or Participants were randomly assigned to the intervention or control groups following baseline assessment and case conference, using a computer-generated random number schedule with permuted blocks . Allocation was concealed by using central randomization performed by NeuRA personnel not otherwise involved in the study. Outcome assessors, including those monitoring the dizziness episodes, were masked to study group allocation. Due to the nature of the intervention, it was not possible to blind the staff administering interventions or the participants. Participants were instructed not to inform the assessors of their intervention status.All eligible participants attended NeuRA for a 3-hour baseline assessment undertaken by trained research staff. The assessments included diagnostic tests for descriptive purposes  and for allocating intervention participants to treatment arms as well as baseline measures for the primary and secondary outcomes. Participants returned to NeuRA for reassessment of outcomes at the end of the trial (6 months post randomization).Multidisciplinary case conferences were held fortnightly throughout recruitment. At these meetings, consensus recommendations and baseline assessment results were used to prioritize and guide appropriate therapies for each participant . Panel mFor participants with vestibular disorders, vestibular rehabilitation was administered at the study site, during a single or several sessions depending on the pathology and its progress, by a physiotherapist (DM) specialized in vestibular rehabilitation. For patients presenting with BPPV, the physiotherapist performed canalith repositioning manoeuvres to treat the different variances of BPPV by removing vestibular debris from the semicircular canals . Given tIn both instances, during an initial 45-minute-long consultation, the physiotherapist reviewed the patients history, assessed them, and then prescribed 2 to 5 exercises (5- to 10-minute sets) for the patients to practice, a minimum of twice and up to 4 times a day, until symptom resolution. Subsequent appointments were scheduled 2 weeks apart to review the participants’ symptomatology and performance of the exercises and modify and/or increase the exercises and their intensity accordingly. An example of exercise progression would involve the transition from practicing static gaze stabilization exercises (during sitting or standing) to dynamic ones . All cases were resolved within 2 to 4 weeks, which is after 1 to 3 visits to the vestibular physiotherapist.https://www.acc.co.nz/assets/injury-prevention/acc1162-otago-exercise-manual.pdf). The timing of the last home visit was brought forward to 16 weeks to align within the 6-month intervention.The home-based Otago exercise programme was delivered by an exercise physiologist for participants with lower-limb muscle weakness and poor balance . This evParticipants with anxiety, depression, and/or fear of falling were provided with an online or booklet-based CBT programme with regular telephone support from a registered psychologist ,39. The Medical management intervention was divided into complex and simple cases. For complex cases, such as those with multiple comorbidities, frailty, cardiovascular problems , high physiological risk of falls and/or history of multiple falls, multiple medication use and likely drug interactions, etc.—we organised an appointment to an outpatient Falls Clinic staffed by a geriatrician and a physiotherapist. Individualised interventions following comprehensive geriatric assessment comprised medical and medication management by a geriatrician and strength and balance training. For simpler issues, we sent a letter to participants’ general practitioners outlining factors identified at the baseline assessment that could contribute to dizziness or imbalance. These included problematic medication use, an abnormal electrocardiogram finding, the presence of low blood pressure that could be addressed with fluid intake or medication management, etc., undiagnosed reduced lower-limb sensation that warranted follow-up, and the use of multifocal glasses that could increase fall risk during ambulatory outdoor activities.During the 6-month trial period, participants assigned to the control group received usual care, i.e., care as usually received by patients in daily practice in this pragmatic clinical trial . We did The primary outcome measures captured 4 crucial aspects of the trial: dizziness episodes, dizziness handicap, balance, and walking stability. The total number of dizziness episodes during the 6-month follow-up were monitored with monthly dizziness diaries  and follSecondary outcome measures were used to elucidate how the interventions might have assisted in ameliorating dizziness symptoms. These measures included assessments of the following: orthostatic hypotension (positive tilt table test) , composiAdverse events  were monitored with monthly calendars and telephone calls as required. Uptake and adherence to all interventions was documented with therapist records and/or participant diaries. Deviations from the original protocol  were the inclusion of participants aged 50 to 64 years to increase generalizability to a wider demographic, as well as removal of the vertigo symptom scale as a secondary outcome measure because it was clear early in the study that this scale was not useful in documenting types of dizziness and was poorly completed by participants. Cost-effectiveness of the intervention will be addressed in a subsequent paper.A power analysis determined that 300 participants (150 per group) were required to provide 80% power to detect a statistically significant 20% between-group difference in the primary outcome measures. For these calculations, we assumed an alpha of 0.05 and a dropout rate of 15%. We assumed the following control group means (SDs) based on values from previous studies of older people: CSRT: 1,322 (331) milliseconds; step-time variability: 0.02 (0.01) seconds; and DHI score: 37 (2) ,45,52,53p < 0.05 were considered statistically significant. Holm-Bonferroni adjustments for multiple comparisons were also performed for the primary outcome measures. Analyses were conducted using the SPSS Version 24.0 software package .All analyses were completed with an intention-to-treat approach , and analysis of the primary outcomes was conducted masked to group allocation . Due to n = 151) or intervention (n = 154) groups. The included participants  were aged 50 to 92 years (mean [SD]: 67.8 [8.3]). All had intact cognitive function according to their performance on the GPCOG tool of cognitive impairment. Participants in the 2 groups were well-matched with regard to baseline characteristics . A total of 126 participants (41%) were identified with vestibular disorders , 54 (18%) with clinically significant symptoms of anxiety and/or depression, 79 (26%) with lower-limb weakness and poor balance, and 18 (6%) with multiple comorbidities. In addition, 107 participants (35%) had medical or medication issues that could precipitate dizziness warranting treatment or advice from a general practitioner. In 71 participants (23%), no cause of dizziness could be identified from the assessments, questionnaires, and interviews.Baseline assessments combined with dizziness history identified at least one potential cause of dizziness for most participants  uptake into the vestibular rehabilitation for those assigned to that therapy (n = 57)—5 participants could not be contacted by the vestibular physiotherapist, 2 declined participation because they lived too far from the assessment site, 2 others declined participation because they were not feeling dizzy anymore, 2 had medical issues, and 1 was not interested anymore. All of those who took up the vestibular rehabilitation programme did attend all their sessions with the vestibular physiotherapist. Uptake for the Otago home exercise programme in those assigned to that intervention (n = 39) was 85% (n = 33)—4 participants discontinued the programme for medical reasons, 1 was erroneously not enrolled in the programme, and 1 moved to a care facility. Regarding adherence to the Otago home exercise programme, 25 of 33 assigned participants (76%) completed 75% or more of the recommended number of exercise sessions (32 of 33 completed 50% or more of the recommended number of sessions). Uptake for the CBT programme was 31% among intervention participants whose anxiety and depression scores met the criteria for inclusion in this therapy (n = 31)—6 declined , 1 could not be contacted, and 15 expressed interest and enrolled in the course but never completed the modules. All the participants who took up the CBT completed the programme. Finally, among participants requiring medical management, only 2 of 9 intervention participants eligible for a falls clinic appointment took up the recommendation. Reasons for failing to attend the falls clinic intervention included living out of area and declining to attend (1), failing to obtain a referral from their general practitioner (4), concurrent referral by general practitioner to a neurology clinic (1), or already attending a community-health programme (1).There was a 79% (p = 0.003). There were no statistically significant between-group differences in dizziness episode frequency during the follow-up period, when controlling for length of follow-up . There were also no statistically significant between-group differences in CSRT performance  and step-time variability during gait   .The intention-to-treat analysis revealed no significant improvements in composite physiological fall risk and dynamic balance measures, anxiety, depression, neuroticism, fear of falling, or orthostatic hypotension .p = 0.002), CBT , and medical management . At completion of the trial and controlling for baseline performance, compared with their control counterparts, participants in the vestibular rehabilitation group also significantly improved their CSRT performance , and those in the exercise intervention group significantly reduced their composite physiological fall risk . At the 6-month follow-up, the CBT recipients significantly improved GAD-7 scores of anxiety .There were indications for intervention-specific improvements at the 6-month reassessment  Tables. This study investigated the effects of a multifactorial tailored intervention on dizziness handicap, frequency of dizziness episodes, and physical function in a sample of middle-aged and older people suffering from dizziness. Overall, our intervention significantly improved dizziness handicap. Given that around 12% of the community aged over 50 years report dizziness : 25.4 [18.3]), a between-group mean difference of 3.7 points in DHI score is between the 10% score change suggested by Treavalen  as clini170–337) ,23,28. TThe intervention-specific analyses revealed that the individual interventions were effective in managing specific aspects of dizziness, as hypothesized: fall risk  for the Otago exercise programme ,63, psycThe fortnightly multidisciplinary meeting was crucial to the success of the intervention because it allowed us to examine the participants’ results from an interdisciplinary perspective, which has been a major limitation of previous research in the field. We recommend that smaller clinics adopt a multidisciplinary approach to the assessment of their patients even if they do not have the specialist on site. Such approach can be done by selecting key measures from each domain.Strengths of the study included the broad sample of middle-aged and older people reporting dizziness, the multidisciplinary input and objective criteria for assigning participants to interventions,  the prosn = 71) had significantly lower mean DHI scores (mean [SD]: 15.8 [10.1]) than the rest of the sample . Other possible reasons include but are not limited to the following: the need for the condition to be present at baseline testing , a previous dizziness episode triggered by severe circumstances  unlikely to reoccur or be reproducible in the laboratory, and a rare cause requiring specific testing .We also acknowledge certain limitations. First, the single-centre nature of the study together with the reliance on self-selected volunteers as participants may limit the generalizability of the study. Second, our intervention comprised one or more individualized evidence-based components. As such, we cannot determine the effect of specific interventions, and it may be that certain interventions were key and others less efficacious. Third, in 23% of the sample, no cause of dizziness could be identified from the assessments, questionnaires, and interviews. Maarsingh and colleagues reported unclear causes of dizziness in only 8% of 417 older people comprehensively assessed in a primary care setting; however, their participants suffered from persistent dizziness  and accon ≤ 41) and delivered CBT either on its own or integrated within a vestibular rehabilitation programme [Fourth, uptake of the interventions was only good  to fair . Evident factors leading to non-adoption of the interventions included the receipt of healthcare through other care providers , the requirement of a general practitioner referral letter for a Falls Clinic appointment, delays in organising the interventions, and dizziness symptoms being considered insufficiently disabling by participants to warrant action. Some participants referred to CBT declined this intervention, reporting that they expected a therapy for dizziness and not one for anxiety and depression. Comparative data regarding uptake and adherence to CBT within dizziness interventions are lacking. In fact, to our knowledge, only few randomized controlled trials to reduce dizziness have included CBT as an intervention. These trials included small sample sizes Click here for additional data file.S2 Text(PDF)Click here for additional data file.S3 Text(DOCX)Click here for additional data file.S4 Text(DOCX)Click here for additional data file.S5 Text(DOCX)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 TableCBT, cognitive-behavioural therapy.(DOCX)Click here for additional data file.S5 Table(DOCX)Click here for additional data file.S1 Data(XLSX)Click here for additional data file.


Less than 1% of adult patients with schizophrenia taking clozapine develop agranulocytosis, and most of these cases occur within the first weeks of treatment. The human leukocyte antigen (HLA) region has been associated with genetic susceptibility to clozapine-induced agranulocytosis (single amino acid changes in HLA-DQB1 (126Q) and HLA-B (158T)). The current study aimed to evaluate the cost-effectiveness, from a healthcare provider’s perspective, of an HLA genotype-guided approach in patients with treatment-resistant schizophrenia who were taking clozapine and to compare the results with the current absolute neutrophil count monitoring (ANCM) schemes used in the USA. A semi-Markovian model was developed to simulate the progress of a cohort of adult men and women who received clozapine as a third-line antipsychotic medication. We compared current practices using two genotype-guided strategies: (1) HLA genotyping followed by clozapine, with ANCM only for patients who tested positive for one or both alleles (genotype-guided blood sampling); (2) HLA genotyping followed by clozapine for low-risk patients and alternative antipsychotics for patients who tested positive (clozapine substitution scheme). Up to a decision threshold of $3.9 million per quality-adjusted life-year (90-fold the US gross domestic product per capita), the base-case results indicate that compared with current ANCM, genotype-guided blood sampling prior to clozapine initiation appeared cost-effective for targeted blood monitoring only in patients with HLA susceptibility alleles. Sensitivity analysis demonstrated that at a cost of genotype testing of up to USD700, HLA genotype-guided blood monitoring remained a cost-effective strategy compared with either current ANCM or clozapine substitution. Treatment-resistant schizophrenia (TRS) is a highly disabling psychiatric disease that affects approximately one-third of psychotic patients over the course of their illness . ClozapiIn the 1990s, the human leukocyte antigen (HLA) region was found to be associated with genetic susceptibility for CIA . After mTo investigate whether the current US intensive blood monitoring might be restricted to high-risk individuals given new shared REMS in 2015, we developed a decision-analytical model to compare the current US ANCM scheme with two pharmacogenetically based schedules. We hypothesized that HLA genotyping was performed prior to clozapine initiation and conditioned the monitoring schedules. Two alternative schemes to current ANCM were defined: (I) clozapine for all patients. Targeted ANCM only in patients testing positive for one or both susceptibility alleles (genotype-guided sampling (GGS)), and (II) clozapine for patients testing negative for both susceptibility alleles plus an antipsychotic substitute for patients testing positive for one or both susceptibility alleles. No ANCM for any patients (clozapine substitution scheme (CSS))  GGS could be cost-effective because the reduced costs may offset the marginal effectiveness by removing long-term ANCM in lower-risk patients who tested negative for the HLA alleles; and that: (II) CSS could be cost-effective because it completely removes the costly ANCM due to switching from clozapine to an antipsychotic substitute in patients with a higher risk (incidence of CIA 1.8% instead of 0.7%), producing less follow-up burden.For the decision-analytical model, we developed a semi-Markovian framework allowing for probabilistic approaches and performed the cost-effectiveness analysis based on the US monitoring schedule and both abovementioned pharmacogenetic alternatives ; in contrast, the authors of a CIA Consortium publication [The study population consisted of adult men and women with TRS who received clozapine as a third-line antipsychotic medication. To maintain consistency with the analytical decision model, we specifically considered agranulocytosis cases  per patient over a 3-year period, which was employed because scant long-term estimates from registry-based prescribing systems are available beyond this follow-up duration. In accordance with the recommendations of the US Panel on Cost-Effectiveness in Health and Medicine  and to p9 per liter with potential infection, and, in some cases, sepsis-related death. Patients could be switched to an alternative antipsychotic for reasons unrelated to CIA  or die for other reasons unrelated to CIA-induced sepsis. The model was developed using the R® programming software package . Further details on model structure can be found elsewhere [We developed additional functions to integrate genotype-based schedules into a validated semi-Markovian model structure  with timlsewhere .http://www.redbook.com/redbook/online) [The cost-effectiveness analysis was conducted from a third-party payer perspective using direct medical expenditures (see Supplementary Table /online) , and clo/online) . We convQuality-of-life variations across the model were applied to each time period spent in HS, weighted by TRS utilities , and derWe conducted one-way sensitivity analyses to assess the robustness of the model outcomes by varying key parameters driving the model throughout credible ranges: sensitivity and specificity of genetic testing, risk of CIA, rate of severe infection in patients with agranulocytosis, case-fatality of sepsis in patients with agranulocytosis, utility of severe infectious episodes, TRS treated by clozapine, TRS treated by clozapine substitute, costs of genetic testing, clozapine substitutes, ANCM, sepsis management, and discount rates on costs and QALYs.We performed probabilistic sensitivity analyses to explore joint parameter uncertainty and whether parameter variability is translated into outcome variability. Parameter values were drawn at random from the assigned distributions for each of the 10,000 simulated cohorts, and 95% confidence intervals were obtained according to the 0.025 and 0.975 percentiles of the simulated results ). In all, 348 screened patients were needed to prevent one case of agranulocytosis. This estimate approximates a previous estimation with higher agranulocytosis prevalence but with a lower, single HLA-DQB1 genotyping sensitivity .In a cohort of 100,000 TRS patients taking clozapine, the 3-year all-cause mortality was 1.70% for the current US monitoring schedule, 1.71% for the GGS, and 1.78% for the CSS. The monitoring number required to avoid one death was nearly 10,000 patients in the current US ANCM scheme when compared with that of the GGS. Considering a 3-year period, the mean survival time adjusted for quality of life was 669.9 quality-adjusted life-days (QALD) for current US ANCM and 669.8 QALD for the GGS (−0.1), compared to 660.2 QALD for the CSS (−9.7). The mean patient cost was $13,694 in the current US ANCM, $13,091 in the GGS (-$603), and $13 738 (+$44) in the CSS  to save one QALY compared with the GGS  or by foregoing ANCM in all patients and selecting an alternative antipsychotic drug instead for patients with HLA alleles of susceptibility (CSS strategy). The results indicated the GGS was cost-effective to a WTP threshold of $3.9 million per QALY, although the results were sensitive to changes in the costs of genetic testing, CIA prevalence, and infection-related death rates.Although translation processes and clinical implementation of pharmacogenomics, including proof of cost-effectiveness, have been emphasized , these rThe relationship between CIA risk and the proportion of patients in the group with variant HLA alleles has been recently studied , 27. GivCost optimization through pharmacogenetic-guided treatments will permit the use of more efficient monitoring interventions for the management of patients taking antipsychotics, such as electrocardiographic screening to reduce drug-induced arrhythmias  and sudden cardiac death . The ratAccording to our results, the pharmacogenetic testing was cost-effective up to a cost of $700 per patient. From a probabilistic approach, and given a WTP of $50 000 per QALY, there is a higher than 99% chance that HLA allele genotyping is cost-effective if HLA test costs remain under $700. This estimate is plausible because pharmacogenetic testing based on genotyping for the two aforementioned allelic variants of HLA-DQB1 and HLA-B (126Q and 158T) is a routine procedure in laboratories that test for HLA histocompatibility. Moreover, precise epitope determination could be established by designing adequate primers that can be used for simple PCR amplification at a very low cost.The association of HLA alleles with an increased risk of agranulocytosis suggests an immune-mediated mechanism. The combination of genetic factors with clinical predictors of CIA, such as older age, sex, and the concomitant administration of other myelotoxic medications  could help narrow the group of patients with HLA alleles of susceptibility for long-term ANCM. Although there is no convincing evidence of the direct toxicity of the parent compound or its stable metabolites (demethylclozapine and clozapine N-oxide), myeloid precursors are the affected target cells. Indeed, various HLA specific alleles have been demonstrated to be directly involved in drug toxicities, such as HLA-B*57:01 with the abacavir-induced hypersensitivity reaction , HLA-B*1One limitation of these results is the moderate performance of the genetic testing, as it remains a driving parameter for clinically effective testing in a small group with a higher CIA prevalence (1.7%). Because the test has relatively low sensitivity, it is hardly advisable to stop, without pilot studies or transition periods, ANCM in all TRS patients at the beginning of clozapine treatment. Furthermore, because of limited long-term data, our analysis could not extend beyond three years without strong assumptions. Finally, because we used a third-party payer perspective, we could neither incorporate unintended follow-up benefit and disutility  associated with ANCM nor intangible costs, such as productivity loss related to premature death.The strengths of this study are the key parameters derived from the largest pharmacogenetic CIA study conducted to date. Additionally, the decision-analytical framework was based on a semi-Markovian model using a wide range of sensitivity analyses, which facilitated the probabilistic calculations of a single strategy and permitted transition between HS at any time during the treatment course in contrast with discrete-event models.In conclusion, a pharmacogenetically guided strategy based on examinations of independent amino acid changes in both HLA-B (158T) and HLA-DQB1 (126Q) appeared to be cost-effective, allowing for the relaxation of long-term ANCM in low-risk patients taking clozapine despite the moderate sensitivity of these allele tests.Similar analyses using less stringent monitoring schedules  should also be performed to assess the cost-effectiveness of pharmacogenetic-guided treatment and to calculate whether it is as economically worthwhile as in the US. Further research on HLA genome-wide genotyping and whole-exome sequencing will likely increase test performance and assist physicians in predicting agranulocytosis more efficiently, allowing a broader clinical application and supporting a review of the current monitoring schedules.Supplementary tables 1–3Supplemental Figure Legend


Remarkably, these effects are maintained in tumors resistant to traditional microtubule-binding chemotherapies such as Taxol®.Multiple-drug resistance in human cancer is a major problem. To circumvent this issue, clinicians combine several drugs. However, this strategy could backfire resulting in more toxic or ineffective treatments. Carbon nanotubes (CNTs), and particularly multi-walled nanotubes (MWCNTs), display intrinsic properties against cancer interfering with microtubule dynamics and triggering anti-proliferative, anti-migratory and cytotoxic effects active-by-design nanocarriers, attempting to enhance the effect of broadly-used chemotherapies. We compare the cytotoxic and the anti-tumoral effect of 5-Fluorouracil (5-FU) -an antimetabolite treatment of various forms of cancer- with that of the drug physisorbed onto MWCNTs. Our results demonstrate how the total effect of the drug 5-FU is remarkably improved (50% more effective) when delivered intratumorally coupled to MWCNTs both in vitro and in vivo in solid tumoral models. Our results demonstrate how using MWCNTs as anti-cancer drug delivery platforms is a promising approach to boost the efficacy of traditional chemotherapies, while considerably reducing the chances of resistance in cancer cells.In the view of these properties, we investigate the use of MWCNTs in the development of  Unfortunately, most aggressive cancers metastasize to distant organs and develop resistance to chemotherapy. For this reason, clinicians often need to combine different drugs to simultaneously interrupt cell proliferation pathways at various points, boosting the potential cytotoxic effect of the treatment. Regrettably, anti-cancer drug combination does not always work. For instances, Taxolnterally . As an a mixture . Here, win vitro [in vivo [in vitro in cancer cells [in vivo [® [® -that binds a structural pocket in the β-tubulin polypeptide- could be significantly boosted when combined with MWCNTs, for these nanomaterials utilize an alternative microtubule stabilization mechanism, complementary to that Taxol® [Carbon nanotubes (CNTs) have shown excellent properties and applications in nanobiotechnology –5. In pain vitro  and in v[in vivo , and the[in vivo . The inc[in vivo , 11, ant[in vivo –14 and cer cells –17, and [in vivo , 19. In  vivo [® . Thus, tt Taxol® . Summariin vitro can degrade surface-oxidized MWCNTs in few days, reducing the length of the nanotubes in approximately 30%, and in vivo, in the tumoral tissue after triggering significant antitumoral effects [But despite these numerous advantages, the stigma of the structural similarity of CNTs with asbestos fibers has slowed down progress of these nanomaterials in medicine, and have been repeatedly excluded in the design of drug nanocarriers . Fortuna effects . Here wevia π-stacking has been broadly used for successful loading of different drugs onto graphene and MWCNTs preventing drug inactivation due to the binding procedure  during 140 h. Measurement of the collected samples using fluorescence spectroscopy suggested an initial phase of ‘burst’ release of the drug during the first 24 h, that corresponded to a 40–42% of the loaded 5-FU. This was followed by a ‘sustained release’ phase where the rest of the drug was slowly discharged in the course of several days ; and finally, to 100 μg/mL of 5-FU-MWCNTs. The effects of the treatments were compared 72 h after exposure to the three therapies.To evaluate the effect of the 5-FU loaded MWCNTs Both, the resuspended 5-FU and 5-FU-MWCNTs produced a potent anti-proliferative effect accompanied by an obvious increase in the cellular size compared to untreated controls, or to cells treated with p-MWCNTs. This phenotype suggested a possible blockage in the “S” part of the cell cycle Figure . Remarkain vitro and suggested a significant rise in the efficacy of the drug when applied decorating MWCNTs.Quantification of the cellular area in cultures treated with both, 5-FU and 5-FU-MWCNT, revealed a statistically significant 3-fold increase in cell size compared to untreated or p-MWCNTs controls . As prevTo perform an accurate qualitative and quantitative analysis of the cytotoxic effects of the treatment, we used flow cytometry. This statistically powerful technique allows the characterization of the cell cycle for each condition, and to quantify the percentage of apoptotic cells in approximately 10,000 cells per experiment. For the study we used human HeLa cells and murine B16F10 cells. This murine model of malignant melanoma cells is resistant to many traditional chemotherapeutic drugs  and, as in vitro, MWCNTs increase the effectiveness of the drug while significantly amplifying the cytotoxic effect of the two chemotherapies individually applied. Therefore, data indicate that the sequential inhibitory effect of the 5-FU at the “S” phase and the MWCNTs at the “M” phase of the cell cycle, as depicted in Figure Figure The anti-tumoral effect of 5-FU-MWCNTs was also tested on solid melanoma tumours produced by transplantation of B16-F10 cells. This system model is highly representative of most malignant tumours, being characterized by local acidosis, edema and abundant tumor-associated supporting stromal cells that include macrophages . As in pResults shown in Figure as-produced MWCNTs were washed with HCl to remove impurities, resuspended in distilled water by sonication and incubated with a solution of ca. 10 mg/mL 5-FU . The mixture was stirred in a vertical wheel at ca. 20 r.p.m. for 2 h at room temperature, centrifuged at 12000 g and washed with distilled water three times in repeated cycles of centrifugation /redispersion (High-purity (>98%) MWCNTs were obtained from Sigma Aldrich (Ref. 698849). The spersion .The concentration of the physisorbed 5-FU was estimated using fluorescence spectroscopy as described in the Supplementary Material . TGA was®). Microtubules were immunostained with B512 anti- α-tubulin antibody (Sigma-Aldrich®) and a secondary goat anti-mouse Alexa Fluor 488-conjugated IgG (Molecular probes®). Confocal laser scanning and phase contrast images were obtained with a Nikon A1R confocal microscope. Image processing was performed with the NIS-Elements Advanced Research software. All fluorescent images are pseudo-colored.HeLa cells were grown in cultured with Eagle’s Minimum Essential Medium (Biowhittaker™). Cells were fixed with 4% paraformaldehyde for imaging. DNA was stained with Hoechst dye  indicative of apoptosis. Three different replicas of the experimental analysis were performed on an average of 10,000 cells per condition using the FACS Diva software (Becton Dickinson). For illustrative purposes the original flow cytometry graphs have been substituted by pie charts displaying the calculated proportions of the different cell populations, including apoptotic cells in Figure t-test was used for statistical analysis and to evaluate significance that was stablished for a (*) p = 0.05 or a (**) p = 0.01. The confidence levels and total number of events included in the study (n) are indicated in the figure legends. Quantitative results are expressed as mean values with their corresponding standard error bars.A Student’s two-tailed ® (paclitaxel) or Epothylones. Indeed, the fact that MWCNTs can produce notable antitumoral effects in solid tumours generated by Taxol®-resistant cells suggests these nanofilaments can complement and significantly boost chemotherapy. Here we demonstrate how drug and nanomaterial therapies can complement each other, in vitro and in vivo in the treatment of cancer. In conclusion, these data invite to improve anti-cancer delivery systems considering the use of nanomaterials with intrinsic antitumoral properties as active excipients, to enhance the therapeutic effect of traditional chemotherapy, preventing drug resistance in cancer.The emergence of drug resistance depends on the genetic instability, heterogeneity and high mutational rate of tumour cells among others. Malignant cancer cells continuously develop new mechanisms of resistance to chemotherapy that include drug destruction, selection of mutations that inhibit drug binding to targets, drug efflux out of the cells, etc. The administration of chemotherapy loaded onto MWCNT used as nanocarriers can significantly improve many different aspects of the traditional chemotherapy. On one hand, the penetrating properties of these nanomaterials are interesting when considering, for instances, treatments applied topically, where CNTs can penetrate and spread into the affected lesion transporting the drug away from the application point. But more interestingly, MWCNTs have intrinsic antitumoral properties that can be exploited in cancer treatment interfering with microtubule dynamics, triggering effects similar to traditional drugs such as Taxol


Does the contribution of modifiable risk factors on area-based inequalities in untreated dental caries among Australian children differ by Indigenous status?In this cross-sectional study of Australian children, the association of modifiable risk factors with area-based inequalities in untreated dental caries among Indigenous and non-Indigenous Australian children differed substantially. Consumption of sugar-sweetened beverages was associated with dental caries for both groups, and irregular tooth brushing was also significantly associated with dental caries for Indigenous children.Targets to reduce consumption of sugar-sweetened beverages may reduce oral health inequalities for both groups; however, Indigenous children require additional focus on oral hygiene. This cross-sectional study estimates the association of modifiable risk factors with dental caries in Indigenous and non-Indigenous Australian children using data from the National Child Oral Health Study 2012-2014. Although the prevalence of untreated dental caries among Indigenous Australian children greatly exceeds the prevalence observed among non-Indigenous children, the associations of dental caries with risk factors is considered to be the same.To estimate the association of modifiable risk factors with area-based inequalities in untreated dental caries among Indigenous and non-Indigenous Australian children using decomposition analysis.Cross-sectional study using data from Australia’s National Child Oral Health Study 2012-2014, a nationally representative sample of both Indigenous and non-Indigenous children aged 5 to 14 years. Data analyses were completed in November 2018.Outcomes were the mean number of decayed tooth surfaces in the primary dentition for children aged 5 to 10 years and mean number of decayed tooth surfaces in the permanent dentition for children aged 8 to 14 years. The area-based measure was the school-based Index of Community Socio-Educational Advantage, with individual-level variables including sex, equivalized household income, tooth-brushing frequency, sugar-sweetened beverage (SSB) consumption, time from last dental visit, and residing in an area with water fluoridation.There were 720 Indigenous children aged 5 to 10 years, 14 769 non-Indigenous children aged 5 to 10 years, 738 Indigenous children aged 8 to 14 years, and 15 631 non-Indigenous children aged 8 to 14 years. For area-based inequalities in primary dentition among Indigenous children, two-thirds of the contribution was associated with SSB consumption , followed by irregular tooth brushing  and low household income . Among non-Indigenous children, almost half the contribution was associated with low household income , followed by SSB consumption  and residing in an area with nonfluoridated water . For area-based inequalities in permanent dentition among Indigenous children, 40.0%  of the contribution was associated with residing in an area with nonfluoridated water, followed by low household income  and consumption of SSBs . Among non-Indigenous children, the contribution associated with low household income, SSB consumption, and last dental visit more than a year ago were each 28.6%.The association of modifiable risk factors with area-based inequalities in untreated dental caries among Indigenous and non-Indigenous Australian children differed substantially. Targets to reduce SSB consumption may reduce oral health inequalities for both groups; however, Indigenous children require additional focus on oral hygiene. In Australia’s National Child Oral Health Study 2012-2014, 44% of Indigenous children had 1 or more deciduous teeth with untreated dental caries, compared with 26% of non-Indigenous children.11 Among children, the literature suggests a number of risk factors for experience of dental caries, which conceptually can be grouped as related to sociodemographic characteristics, dental hygiene, diet, dental service use, and environment (water fluoridation). Sociodemographic factors include household income, which is a proxy for broader material influences that play in a role in oral health literacy, purchasing power, and general health education.12 Oral hygiene practices such as tooth-brushing frequency are associated with the physical removal of food and beverage residue in the oral cavity, lowering levels of Streptococcus mutans through reduction of plaque,13 and bolstering fluoride in saliva reservoirs if fluoridated toothpaste is used. Diet, specifically a cariogenic diet, is associated with dental caries, with recent evidence demonstrating that frequent consumption of sugar-sweetened beverages (SSBs) is one of the strongest associations.14 Frequent use of dental services provides preventive measures such as application of topical fluoride and general prophylaxis and enables any untreated dental caries to be restoratively managed.15 Consumption of fluoridated water is one of the most successful public health interventions regarding oral health and is especially beneficial in reducing oral health inequalities between Indigenous and non-Indigenous Australian children.16It is widely accepted that dental disease is socially patterned, with a plethora of research indicating that socially disadvantaged populations have substantially greater experience of dental disease than those who are more advantaged for both individual and area-based measures of inequality.While these factors associated with dental disease have been identified, to our knowledge, there has been no application of analysis that enables specific percentages of risk contribution to be calculated. Nor has there been any analysis of whether the risks, and contribution of these risks, might differ between Indigenous and non-Indigenous children. This is important for both targeting of effective oral health promotion initiatives and for policy implications in the allocation of scarce resources in the dental public health setting. This study, therefore, aims to analyze both area- and individual-level oral health inequalities among Australian children and, specifically, to assess the association of individual-level sociodemographic, oral hygiene, dietary, dental service use, and water fluoridation factors with area-based inequalities in untreated dental caries, stratified by Indigenous status. The hypothesis is that the percentage association of each factor will differ for Indigenous and non-Indigenous children.STROBE) reporting guideline. The study was reviewed and approved by the University of Adelaide Human Research Ethics Committee; research ethics committees within each jurisdiction; the 3 education sectors: public, independent, and Catholic schools; and the Indigenous Human Research Ethics Committee at the Menzies Institute. Parents provided signed, informed consent for their child to participate.This study is reported according to the Strengthening the Reporting of Observational Studies in Epidemiology  for caries assessment scores ranged from 0.67 to 0.99, indicating good to excellent reliability.Data were obtained from Australia’s National Child Oral Health Study 2012-2014 (NCOHS), a population-based cross-sectional survey of Australian children aged 5 to 14 years from both primary and secondary schools, for which data collection details have been previously described.7Child examination and questionnaire data were weighted separately for each state and territory by deriving survey weights, which were adjusted by school type and sociodemographic characteristics of participating children. Population estimates derived from the weighted sample closely reflected the true child population.The outcome variable was mean number of tooth surfaces with untreated decay for the primary dentition (ds) (age 5-10 years) and for the permanent dentition (DS) (age 8-14 years).17 School level was dichotomized into low  and high .The area-based measure was the school-based Index of Community Socio-Educational Advantage (ICSEA), a composite variable of school socioeconomic status that combines student characteristics (parent occupation and level of education) and school-area characteristics .18 Equivalized household income was grouped into 4 approximately equal quartiles, with quartile 1 being lowest and quartile 4 being highest. The dental hygiene variable included frequency of tooth brushing (<2 times per day and ≥2 times per day). The diet variable was number of glasses of SSB per day . Dental service use included time since last dental visit (≥18 months or <18 months). Environmental factors included exposure to water fluoridation (yes for ≥0.5 mg/L fluoride vs no for <0.5 mg/L fluoride). Fluoridation status of the local water supply was based on the child’s residential postcode. The Australian Research Centre for Population Oral Health maintains a postcode-level database of fluoride concentration in water supplies. The recommended range for fluoride in water supplies in Australia is 0.6 to 1.1 mg/L dependent on ambient temperature. If the fluoride level in the drinking water was at least 0.5 mg/L, the water was defined as fluoridated; otherwise, it was classified as nonfluoridated.Individual-level explanatory variables included sociodemographic, dental hygiene, diet, dental service use, and environmental factors. Sociodemographic factors included child sex and equivalized household income. Equivalized household income was derived by calculating an equivalence factor according to the modified Organisation for Economic Co-operation and Development equivalence scale, then dividing income by the factor. The equivalence factor was built up by allocating points to each person in a household  and then summing the equivalence points of all household members.Data were analyzed separately for Indigenous and non-Indigenous children for age groups 5 to 10 years and 8 to 14 years, which included descriptive multilevel multivariable regression, Blinder-Oaxaca, and Neumark decomposition analyses. Data analyses were completed in November 2018. The mean ds and DS were estimated according to each explanatory variable using a procedure in SUDAAN statistical software version 11.0.3 . Multilevel general linear regressions were used to build 2-level models in which individuals  were nested within schools . The random component (level 1 and 2 variances) and the fixed component (regression coefficients) of the models were estimated by using maximum likelihood to test the association between school level and untreated decayed surfaces after adjusting for individual-level explanatory variables. The ICC was estimated as the percentage of school-level variance in the total  variance to determine whether mean ds and DS vary notably across schools (ICC > 0):00 quantifies the variation in mean ds and DS across schools. The ICC value indicates the proportion of variance at the school level.19where ϭ2 is the average variance of individual’s (child’s) ds or DS within schools and τThe models were compared using an Akaike information criterion to assess goodness of fit and to determine which models were preferred. The lower values of Akaike information criterion indicate a better model fit. All models were developed and fitted using SAS statistical software version 9.4 (SAS Institute Inc).Z test with 2-tailed P < .05), were used to identify factors that explained most of the school-level inequalities in untreated dental caries using STATA version 14 statistical software (StataCorp LLC). Analyses used the surveys’ sampling probability weights, accounting for the sampling strategy and nonresponse, to provide population-representative data.Blinder-Oaxaca and Neumark decomposition analyses, straightforward statistical methods that use, in our case, socioeconomic area as a way to explain observed differences between groups  resided in homes with the lowest household income, compared with 28.7% of non-Indigenous children. Nearly half of Indigenous children (49.0%) brushed less than twice per day compared with 30.0% of non-Indigenous children. Approximately half of Indigenous children (49.4%) consumed 2 or more cups of SSB per day, compared with 26.0% of non-Indigenous children. Among Indigenous children, 27.1% had last visited a dentist more than 18 months previous, compared with 17.9% of non-Indigenous children. In all, 43.3% of Indigenous children resided in areas without water fluoridation, compared with 29.3% of non-Indigenous children. The mean DS for Indigenous children was nearly 3 times that of non-Indigenous children (0.8 vs 0.3). The highest levels of mean DS among Indigenous children were observed among those in the lowest school ICSEA level (1.0), those in the lowest household income quartile (0.8), those brushing less than twice daily (0.8), those consuming more than 2 SSBs per day (1.2), those who last received dental care more than 18 months previously (0.9), and those who did not reside in areas with fluoridated water (1.1).The decomposition models  demonstrThe study aims were to characterize area- and individual-level oral health inequalities among Indigenous and non-Indigenous Australian children and to assess the contributions associated with individual-level modifiable risk factors on area-based inequalities in untreated dental caries. Although modifiable risk factors that were most strongly associated with the gap in untreated dental decay explained by school-level socioeconomic status for both Indigenous and non-Indigenous children were largely the same , different risk factors were also observed. Examples of these factors were tooth-brushing frequency for Indigenous children and last dental visit for non-Indigenous children. The hypothesis that the percentage contribution associated with each risk factor would differ for Indigenous and non-Indigenous children proved true; there were substantial differences in the contributions associated with each of these risk factors. The findings have important policy translation implications, as they indicate that while targets to reduce consumption of SSBs may reduce oral health inequalities for all Australian children, an additional focus on oral hygiene is required for Indigenous children. This is important for both targeting of effective oral health promotion initiatives and for policy implications in the allocation of scarce resources in the public dental health setting.20 and suggests that, in the Australian context, there is something inherently unique in the social composition of Indigenous Australians that makes them more vulnerable to chronic disease health states  over and above indicators of social advantage.21It is important to comment on the overwhelming individual- and area-based oral health inequalities experienced by Indigenous children relative to their non-Indigenous peers. Irrespective of dentition, levels of untreated dental caries was 3 times higher among Indigenous children, which was not mitigated by area-based social advantage . This is contrary to evidence suggesting that positive oral health outcomes are associated with high socioeconomic status areas9 and Australia is not alone in its efforts to ameliorate Indigenous-related health inequities.24 Scholars and community leaders have insisted that Indigenous health needs to be considered as separate from the health of other racial and ethnic minority groups within a given country, owing to the sustained colonization, discrimination, and marginalization, along with policies that focus on assimilation and, in some cases, cultural annihilation experienced by these groups.25 While our findings reflect health system differences , they also reflect a maldistribution of the social determinants of oral health; that is, Indigenous and non-Indigenous inequalities in education, jobs, material security, and experiences of discrimination. Attitudinal and cultural barriers also play a role,26 for example, a lack of cultural awareness among non-Indigenous health professionals, Indigenous suspicion (and sometimes rejection) of Western health systems,27 and the absence  of treaty-based or formal recognition of Indigenous rights, including native title. Until only recently, substantial restrictions have been imposed on Indigenous Australian’s civil rights, residence, mobility, and employment.28Child oral health inequalities provide insight into social inequality in a given society,Untreated dental decay is an indicator of inability to access timely, appropriate, acceptable, and affordable dental care. The profound inequalities demonstrated indicate that Indigenous Australian children are not only disadvantaged with respect to dental disease, but also in their ability to access appropriate care for that disease. Untreated decayed teeth reflect not only unequal access to care, but also unequal social conditions , which lead to dental decay in the first instance.It is important to discuss the study limitations. The design was cross-sectional, meaning no causal inferences can be implied. The water fluoridation estimates were based on carer-provided residential histories, with only the current place of residence used to characterize exposure to water fluoridation. This could have led to some misclassification. The study also has some substantial strengths, including being a large, representative sample of Australian children (for both Indigenous and non-Indigenous populations) that implemented internationally accepted measures for both parental self-reported information and clinical assessments, a nationally used school-based social deprivation index, sophisticated weighting approaches, and complex multilevel and decomposition techniques.29 reducing oral health inequalities among and between Indigenous groups needs to be a public health priority at a global level.Our study provides new evidence on the magnitude of oral health inequalities experienced by Indigenous and non-Indigenous Australian children and the contributions of individual-level modifiable risk factors on area-based inequalities in untreated dental caries for these groups. Efforts by the dental profession—as well as policy makers and health professionals more generally—are required at both national and international levels to reduce barriers to access to and the availability of preventive and rehabilitative oral health services for Indigenous groups. As reported in the Geneva Declaration on the Health and Survival of Indigenous Peoples,


Introduction: Cancer antigen 125 (CA-125) is a tumor marker of ovarian cancer, which has shown to be increased in different cardiovascular diseases. Although the prognostic role of CA-125 in heart failure and coronary heart disease is well-established, there is little known about its role in acute myocardial infarction (AMI). In this study we aimed to evaluate the serum levels of CA-125 in patients with AMI and its prognostic role in evaluating the in-hospital outcome of AMI.Methods: We evaluated 120 male patients with AMI and 120 male normal subjects. CA-125 levels were measured upon the patient’s admission to hospital. The in-hospital major adverse cardiac events (MACE) and its predictors were also recorded for AMI patients.Results: CA-125 levels were significantly higher in AMI patients compared to normal subjects . We found significant positive correlations between CA-125 levels with creatine kinase-MB (CKMB)  and CTnI . The in-hospital MACE was observed in 19 cases (15.8%). Patients with MACE had significantly higher value of CA-125, CKMB and CTnI and lower LVEF compared to patients without MACE. CKMB  and CA-125 levels  were independent predictors of MACE.Conclusion: Serum CA-125 levels are significantly higher in male patients with AMI compared to normal subjects and have a significant role in predicting in-hospital MACE after AMI. In patients with higher CA-125 more aggressive treatment and close observation should be performed in order to reduce the possible adverse outcomes. In recent years, various studies have reported prognostic role of increased CA-125 in different cardiac diseases.9 In this study, we measured CA-125 levels in patients with acute MI and its prognostic role in evaluating the outcome of MI.Due to these findings, it is possible that CA-125 could be used as prognostic factor in myocardial infarction (MI). There are few previous studies reporting increased CA-125 levels in patients with MI.In this case control study, 120 eligible male patients admitted to Madani Heart Center, Tabriz, Iran between August 2014 and December 2014 with ST elevation acute MI (STEMI) who underwent primary percutaneous coronary intervention (PCI) were recruited. Control group were 120 male subjects who were evaluated for suspicion of heart disease with normal cardiac findings. Both groups were matched for baseline findings. As CA-125 is increased in genital disease in female patients including ovarian disease, to decrease the possible bias in Ca-125 levels, we only recruited male subjects. Exclusion criteria were history of congenital heart disease, congestive heart failure (CHF), vascular or valvular disease, cardiomyopathy, left ventricular systolic dysfunction with non-ischemic causes, atrial fibrillation, previous STEMI, regional-wall motion abnormalities in non–infarct-related regions, previous coronary artery bypass grafting (CABG), pleural or pericardial effusion, ascites, active infection requiring intravenous antibiotics, chronic renal or liver disease, malignancy, hematological disorders.d ≥ 0.30 as statistically significant in a two-tailed test with α=0.05 and power of 0.95. In order to reach better results, we included 120 patients in each group.Study sample was calculated n = 109 for each group using G*Power 3.1.3.9 software considering the effect size of 10 at the baseline. Left ventricular ejection fraction (LVEF) was measured by transthoracic echocardiography.Patients’ demographic, laboratory and clinical data were collected. All patients were classified by Killip classificationAll patients underwent PCI and the in-hospital outcome and major adverse cardiac events (MACE) including cardiac and non-cardiac death, MI, HF and need for revascularization were recorded.Blood samples were collected at the admission. Creatine kinase-MB (CKMB) was measured in three different periods and CTnI was measured twice and the peak CKMB and CTnI was included. CA-125 was measured with chemiluminescent enzyme immunoassay methods by using an Acculite CLIA, commercial kit . CA-125 normal level was 35 UI/mL.t test and Mann-Whitney U test were used to compare findings between groups. Correlations were analyzed using the Pearson correlation. Logistic regression analysis was used to define predictors of in-hospital MACE. A P value <0.05 was considered statistically significant.All data were analyzed using SPSS statistics version 17.0. Numerical variables were expressed as mean ± standard deviation (SD), whereas categorical variables were expressed as percentage. The chi-square, Fisher exact test, Student’s P < 0.001) and CTnI , while the correlations with other variables were not significant in patients with AMI.Most patients were in Killips class I  and few were in Killips class II and III . Patients had mean LVEF of 42.29±9.52%. We found significant correlations between CA-125 levels with CKMB  including Re-MI in 6 cases, revascularization in 11 cases, CABG in 2 cases, cardiac death in 2 and overall death in 4 cases. Few patients had 2-3 complications during the hospital stay. There were no cases of major bleeding or stroke. There were also no cases of new onset atrial fibrillation or HF after MI during the hospital stay. Demographic and laboratory findings among patients with and without MACE were evaluated and there was significantly higher levels of CA-125, CKMB and CTnI and lower LVEF in patients with MACE compared to those without MACE .P = 0.007] and CA-125 levels  were independent predictors of MACE. In fact, lower levels of CKMB and CA-125 had protective effect in occurrence of MACE.Significant findings were used in logistic regression analysis to find predictors of MACE  and obse12 Although there are few reports, the increased CA-125 levels is also documented in AMI.9The increased CA-125 levels have been reported in different cardiac pathologies including CHF, coronary heart disease (CHD), AF and valve disease such as aortic stenosis and mitral stenosis.13 also reported that CA-125 levels before and after aortic valve implantation could independently predict death and MACE.In this prospective study, we observed that AMI patients compared to normal subjects had significantly higher levels of CA-125 levels. The outcome of AMI was poor in older patients and in those with lower LVEF. Higher levels of CA-125, CKMB and CTnI and lower LVEF were found in patients with MACE than without MACE, of which CKMB and CA-125 were independent predictors of MACE. Husser et al7 These may also have a role in the development of atherosclerosis and its complications.14 Mechanical stress and inflammation could induce CA-125 synthesis from the mesothelial cells of the peritoneum, pleura, and pericardium.15 Considering these changes, CA-125 could act by inflammatory repose in CHD development and also AMI occurrence.There are some possible mechanisms in which CA-125 could be increased in cardiac disease. The role of hemodynamic abnormalities and inflammatory cytokines on the CA-125 levels have been recommended in malignant, non-malignant and chronic diseases like CHF.7 have suggested that ischemic injury of myocardium causing possible ventricular remodeling and heart enlargement would have a role in the secretion of CA125 in CHD patients. However, these findings need more investigations.Although myocardial production of CA-125 have not been documented, the possibility could not be excluded. Rong and colleagues16 In fact, CA-125 has shown considerable correlation to functional and clinical status of the patient and may demonstrate those at risk of adverse outcomes.17While the exact mechanism of CA-125 level increase in cardiac disease is not well understood, there are some properties that make CA-125 a promising prognostic tool: it is widely available and less expensive that other biomarkers. It is stable with long half life (more than a week) which is correlated with clinical status and prognosis.There were several limitations. This was a single center study that would limit to extend it to the general population. Also, the sample of patients and number of outcome, especially mortality rate were small. We did not recheck the CA-125 levels after the acute phase of the AMI to compare the possible changes in its levels and define its exact role in STEMI which could be another limitation of our study.As in this study we excluded women from the study to reduce the possible effects of gender and women related disease on CA-125 levels, it could be concluded that there is higher CA-125 levels in male patients with AMI and have significant role in predicting in-hospital MACE after AMI. In patients with higher CA-125 more aggressive treatment and close observation should be performed in order to reduce the possible adverse outcomes.The Ethics committee of Tabriz University of Medical Sciences approved the study and in-formed consent were obtained from all participants.None.This article was supported by Cardiovascular Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. Authors have no other funding support to disclose