Cerebrospinal fluid and serum protein markers in autism: A co-twin study.

No robust biomarkers have yet been identified for autism spectrum disorder (ASD) or autistic traits. Familial factors likely influence biomarkers such as protein concentrations. Comparing twins with ASD or high autistic traits to the less affected co-twin allows estimating the impact of familial confounding. We measured 203 proteins in cerebrospinal fluid (n = 86) and serum (n = 127) in twins (mean age 14.2 years, 44.9% females) enriched for ASD and other neurodevelopmental conditions. Autistic traits were assessed by using the parent-report version of the Social Responsiveness Scale-2. In cerebrospinal fluid, autistic traits correlated negatively with three proteins and positively with one. In serum, autistic traits correlated positively with 15 and negatively with one. Also in serum, six were positively-and one negatively-associated with ASD. A pathway analysis of these proteins revealed immune system enrichment. In within twin pair analyses, autistic traits were associated with serum B-cell activating factor (BAFF) only, whereas Cystatin B (CSTB) remained significantly associated with ASD. These associations did not remain significant when only considering monozygotic twins. For the remainder, the within-pair analysis indicated familial confounding, including shared environment and genes, influencing both autism and protein levels. Our findings indicate proteins involved in immunity as putative biomarkers of autistic traits and ASD with partial genetic confounding. Although some results are in line with previous studies in general, further studies are needed for replication.

Neonatal CSF vasopressin concentration predicts later medical record diagnoses of autism spectrum disorder.

Autism spectrum disorder (ASD) is a brain disorder characterized by social impairments. ASD is currently diagnosed on the basis of behavioral criteria because no robust biomarkers have been identified. However, we recently found that cerebrospinal fluid (CSF) concentration of the "social" neuropeptide arginine vasopressin (AVP) is significantly lower in pediatric ASD cases vs. controls. As an initial step in establishing the direction of causation for this association, we capitalized upon a rare biomaterials collection of newborn CSF samples to conduct a quasi-prospective test of whether this association held before the developmental period when ASD first manifests. CSF samples had been collected in the course of medical care of 0- to 3-mo-old febrile infants (n = 913) and subsequently archived at -70 °C. We identified a subset of CSF samples from individuals later diagnosed with ASD, matched them 1:2 with appropriate controls (n = 33 total), and quantified their AVP and oxytocin (OXT) concentrations. Neonatal CSF AVP concentrations were significantly lower among ASD cases than controls and individually predicted case status, with highest precision when cases with comorbid attention-deficit/hyperactivity disorder were removed from the analysis. The associations were specific to AVP, as ASD cases and controls did not differ in neonatal CSF concentrations of the structurally related neuropeptide, OXT. These preliminary findings suggest that a neurochemical marker of ASD may be present very early in life, and if replicated in a larger, prospective study, this approach could transform how ASD is detected, both in behaviorally symptomatic children, and in infants at risk for developing it.

Cerebrospinal fluid vasopressin and symptom severity in children with autism.

Autism is a brain disorder characterized by social impairments. Progress in understanding autism has been hindered by difficulty in obtaining brain-relevant tissues (eg, cerebrospinal fluid [CSF]) by which to identify markers of disease and targets for treatment. Here, we overcome this barrier by providing evidence that mean CSF concentration of the "social" neuropeptide arginine vasopressin (AVP) is lower in children with autism versus controls. CSF AVP concentration also significantly differentiates individual cases from controls and is associated with greater social symptom severity in children with autism. These findings indicate that AVP may be a promising CSF marker of autism's social deficits. Ann Neurol 2018;84:611-615.

CSF concentrations of 5-methyltetrahydrofolate in a cohort of young children with autism.

OBJECTIVE: To examine the association between cerebral folate deficiency and autism, this study examined CSF 5-methyltetrahydrofolate (5-MTHF) concentrations in a group of young children with autism, investigated the natural variation in CSF 5-MTHF over time, and assessed the relationship between CSF 5-MTHF and symptoms. METHODS: CSF was collected from 67 children with a diagnosis of DSM-IV-TR autistic disorder (age, mean ± SD 43 ± 11 months), with a second CSF sample obtained 1-3 years later on 31 of these subjects (time to follow-up, 30 ± 8 months). RESULTS: At time 1, 7% (5/67) of participants had 5-MTHF <40 nmol/L. At follow-up, 23% (7/31) of participants had 5-MTHF <40 nmol/L (only one of whom had been low at time 1). A moderate correlation with a very wide confidence interval (CI) was observed between time 1 and time 2 CSF 5-MTHF measurements (Pearson r[p] = 0.38 [0.04]; 95% CI 0.02-0.64). Neither the CSF 5-MTHF levels nor changes over time correlated with the clinical features of autism. CONCLUSIONS: CSF 5-MTHF levels vary significantly over time in an unpredictable fashion and do not show a significant relationship to typical clinical features of autism. Reduced CSF 5-MTHF levels are a nonspecific finding in autism. Our data do not support the use of lumbar puncture for assessment of CSF 5-MTHF in autism.

Transplantation of umbilical cord blood mononuclear cells increases levels of nerve growth factor in the cerebrospinal fluid of patients with autism.

We aimed to evaluate the levels of growth factors in the cerebrospinal fluid (CSF) of patients with autism after transplantation of umbilical cord blood mononuclear cells (CBMNCs). Fourteen subjects diagnosed with autism received transplantation of CBMNCs first through intravenous infusion, and three times subsequently through intrathecal injections. A 2-mL sample of CSF was taken before each intrathecal injection. CSF levels of nerve growth factor (NGF), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) were determined by enzyme-linked immunosorbent assay. All data are reported as means ± SD and were analyzed using the SPSS 10.0 software. One-way analysis of variance with post-hoc F-and Q-tests were performed for comparisons. NGF levels in the CSF were significantly increased after transplantation (213.54 ± 56.38 after the third versus 28.32 ± 12.22 ng/L after the first transplantation; P < 0.05), while VEGF and bFGF levels did not change significantly. Therefore, transplantation of CBMNCs could increase NGF levels in the CSF of patients with autism.

Effects of fluoxetine treatment on striatal dopamine transporter binding and cerebrospinal fluid insulin-like growth factor-1 in children with autism.

A positive effect of fluoxetine has been shown in some children with autism. The present study was undertaken to correlate striatal dopamine transporter (DAT) binding and cerebrospinal fluid insulin-like growth factor-1 (CSF-IGF-1) with clinical response in autistic children (n=13, age 5-16 years) after a 6-month fluoxetine treatment. Good clinical responders (n=6) had a decrease (p=0.031) in DAT binding as assessed using single-photon emission computed tomography with [123I]-nor-ß-CIT, whereas poor responders had a trend to an increase. An increase in CSF-IGF-1 (p=0.003) was detected after the treatment period, but no correlation between the clinical response and CSF-IGF-1 was found. In conclusion, fluoxetine decreases DAT binding indicating alleviation of the hyperdopaminergic state and increases CSF-IGF-1 concentration, which may also have a neuroprotective effect against dopamine-induced neurotoxicity in autistic children.

Profiling for novel proteomics biomarkers in neurodevelopmental disorders.

Protein biomarker discovery from biological fluids, such as serum, has been widely applied to disorders such as cancer and has more recently also been utilized in neuro-psychiatric disorders with relatively clear biological causes, such as Alzheimer's disease and schizophrenia. The application of the associated technologies for the identification of protein biomarker signatures in neurodevelopmental disorders, such as autism spectrum disorder and attention deficit hyperactivity disorder, is comparatively less well established. The aim of this article is to provide an overview of the various protocols available for such analysis, discuss reports in which these techniques have been previously applied in biomarker discovery/validation in neurodevelopmental disorders, and consider the future development of this area of research.

Phenotypic expression of autoimmune autistic disorder (AAD): a major subset of autism.

BACKGROUND: Autism causes incapacitating neurologic problems in children that last a lifetime. The author of this article previously hypothesized that autism may be caused by autoimmunity to the brain, possibly triggered by a viral infection. This article is a summary of laboratory findings to date plus new data in support of an autoimmune pathogenesis for autism. METHODS: Autoimmune markers were analyzed in the sera of autistic and normal children, but the cerebrospinal fluid (CSF) of some autistic children was also analyzed. Laboratory procedures included enzyme-linked immunosorbent assay and protein immunoblotting assay. RESULTS: Autoimmunity was demonstrated by the presence of brain autoantibodies, abnormal viral serology, brain and viral antibodies in CSF, a positive correlation between brain autoantibodies and viral serology, elevated levels of proinflammatory cytokines and acute-phase reactants, and a positive response to immunotherapy. Many autistic children harbored brain myelin basic protein autoantibodies and elevated levels of antibodies to measles virus and measles-mumps-rubella (MMR) vaccine. Measles might be etiologically linked to autism because measles and MMR antibodies (a viral marker) correlated positively to brain autoantibodies (an autoimmune marker)--salient features that characterize autoimmune pathology in autism. Autistic children also showed elevated levels of acute-phase reactants--a marker of systemic inflammation. CONCLUSIONS: The scientific evidence is quite credible for our autoimmune hypothesis, leading to the identification of autoimmune autistic disorder (AAD) as a major subset of autism. AAD can be identified by immune tests to determine immune problems before administering immunotherapy. The author has advanced a speculative neuroautoimmune (NAI) model for autism, in which virus-induced autoimmunity is a key player. The latter should be targeted by immunotherapy to help children with autism.

Brain morphometry volume in autistic spectrum disorder: a magnetic resonance imaging study of adults.

BACKGROUND: Several prior reports have found that some young children with autism spectrum disorder [ASD; including autism and Asperger's syndrome and pervasive developmental disorder - not otherwise specified (PDD-NOS)] have a significant increase in head size and brain weight. However, the findings from older children and adults with ASD are inconsistent. This may reflect the relatively small sample sizes that were studied, clinical heterogeneity, or age-related brain differences. METHOD: Hence, we measured head size (intracranial volume), and the bulk volume of ventricular and peripheral cerebrospinal fluid (CSF), lobar brain, and cerebellum in 114 people with ASD and 60 controls aged between 18 and 58 years. The ASD sample included 80 people with Asperger's syndrome, 28 with autism and six with PDD-NOS. RESULTS: There was no significant between-group difference in head and/or lobar brain matter volume. However, compared with controls, each ASD subgroup had a significantly smaller cerebellar volume, and a significantly larger volume of peripheral CSF. CONCLUSIONS: Within ASD adults, the bulk volume of cerebellum is reduced irrespective of diagnostic subcategory. Also the significant increase in peripheral CSF may reflect differences in cortical maturation and/or ageing.

Elevation of tumor necrosis factor-alpha in cerebrospinal fluid of autistic children.

Recent reports implicating elevated cytokines in the central nervous system in a small number of patients studied with autism have reported clinical regression. These studies have not focused on tumor necrosis factor-alpha as a possible marker for inflammatory damage. A series of 10 children with autism had clinical evaluation of their serum and spinal fluid for inflammatory changes and possible metabolic disease as part of their neurological evaluation. Elevation of cerebrospinal fluid levels of tumor necrosis factor-alpha was significantly higher (mean = 104.10 pg/mL) than concurrent serum levels (mean = 2.78 pg/mL) in all of the patients studied. The ratio of the cerebrospinal fluid levels to serum levels averaged 53.7:1. This ratio is significantly higher than the elevations reported for other pathological states for which cerebrospinal fluid and serum tumor necrosis factor-alpha levels have been simultaneously measured. This observation may offer a unique insight into central nervous system inflammatory mechanisms that may contribute to the onset of autism and may serve as a potential clinical marker. More controlled study of this potentially important observation may prove valuable.

Cerebrospinal fluid insulin-like growth factors IGF-1 and IGF-2 in infantile autism.

There has been little exploration of major biologic regulators of cerebral development in autism. We measured insulin-like growth factors (IGF) -1 and -2 from cerebrospinal fluid (CSF) by radio immunoassay in 25 children with autism (median age 5y 5mo; range 1y 11mo-15y 10mo; 20 males, 5 females), and in 16 age-matched comparison children without disability (median age 7y 4mo; range 1y 1mo-15y 2mo; eight males, eight females). IGF-1 and -2 concentrations were further correlated with age of patients and head size. CSF IGF-1 concentration was significantly lower in patients with autism than in the comparison group. The CSF concentrations of children with autism under 5 years of age were significantly lower than their age-matched comparisons. The head circumferences correlated with CSF IGF-1 in children with autism but no such correlation was found in the comparison group. There was no difference between the two groups in CSF IGF-2 concentrations. No patients with autism had macrocephaly. We conclude that low concentrations of CSF IGF-1 at an early age might be linked with the pathogenesis in autism because IGF-1 is important for the survival of Purkinje cells of the cerebellum. The head growth might be explained by the actions of IGF-1 and -2 reflected in CSF concentrations.

Administration of secretin for autism alters dopamine metabolism in the central nervous system.

We evaluated the clinical effects of intravenously administered secretin in 12 children with autism (age range: 4-6 years, median age: 9 years, boy:girl=8:4). In addition, we investigated the association between improvement in symptoms and changes in the cerebrospinal fluid (CSF) homovanillic acid (HVA),5-hydroxyindole-3-acetic acid (5-HIAA), and 6R-5,6,7,8-tetrahydro-L-biopterin (BH(4)) levels after administration. After administration of secretin, the Autism Diagnostic Interview-Revised (ADI-R) score improved in 7 of the 12 children. However, the score deteriorated in 2 of the 12 children (in the item of 'restricted and repetitive, stereotyped interests and behaviors'). The HVA and BH(4) levels in CSF were increased in all children with improvement in the ADI-R score. In contrast, no patient without the elevation of the BH(4) level showed improvement in the score. These findings suggest that secretin activated metabolic turnover of dopamine in the central nervous system via BH(4), improving symptoms.

Cerebrospinal fluid and serum markers of inflammation in autism.

Systemic immune abnormalities have no known relevance to brain dysfunction in autism. In order to find evidence for neuroinflammation, we compared levels of sensitive indicators of immune activation: quinolinic acid, neopterin, and biopterin, as well as multiple cytokines and cytokine receptors, in cerebrospinal fluid and serum from children with autism, to control subjects with other neurologic disorders. In cerebrospinal fluid from 12 children with autism, quinolinic acid (P = 0.037) and neopterin (P = 0.003) were decreased, and biopterin (P = 0.040) was elevated, compared with control subjects. In sera from 35 persons with autism, among cytokines, only tumor necrosis factor receptor II was elevated compared with controls (P < 0.02). Decreased quinolinic acid and neopterin in cerebrospinal fluid are paradoxical and suggest dysmaturation of metabolic pathways and absence of concurrent infection, respectively, in autism. Alternatively, they may be produced by microglia but remain localized and not expressed in cerebrospinal fluid.

Cerebral folate deficiency with developmental delay, autism, and response to folinic acid.

The authors describe a 6-year-old girl with developmental delay, psychomotor regression, seizures, mental retardation, and autistic features associated with low CSF levels of 5-methyltetrahydrofolate, the biologically active form of folates in CSF and blood. Folate and B12 levels were normal in peripheral tissues, suggesting cerebral folate deficiency. Treatment with folinic acid corrected CSF abnormalities and improved motor skills.

Neuropeptides and non-human primate social deficits associated with pathogenic rearing experience.

There is a persuasive evidence that autism is highly heritable and likely to be substantially determined by polygenic mechanisms. Nevertheless, some intriguing findings in children raised in conditions of extreme social deprivation suggest that an autistic-like syndrome may occur as a consequence of environmental conditions. A particularly close model of this human syndrome has been studied in rhesus monkeys for almost half a century. Monkeys reared in pathogenic rearing conditions manifest considerable deficits in social interaction and increased self-directed behaviors. We have been interested in the possibility that disruptions in normal social development in non-human primates might be expressed in neuropeptide systems which have emerged in rodent studies as important candidates for a unique social biology. In recent studies, we have described persistently reduced CSF OT levels in male rhesus monkeys with significant social deficits. We also found that OT levels were positively related to the expression of affiliative social behaviors. Alterations were also detected in both CRH and AVP receptor binding patterns in limbic structures likely to influence social and emotional development. Taken together, these data suggest that abnormal rearing influences the development of brain systems critical to normal social and emotional competence in rhesus monkeys and may contribute to the development of autistic-like symptomatology associated with pathogenic rearing histories.

[Efficacy of secretin for the treatment of autism].

We administered secretin intravenously to 14 patients with autism (9 to 14 years, 10 males; 4 females), and evaluated its clinical effect. We also measured cerebrospinal fluid (CSF) levels of homovanillic acid (HVA) and 5-hydroxy-indole-3-acetic acid (5HIAA) before and after 4 weeks treatment, and compared them with the grade of improvement of the clinical symptoms assessed by the scores of Autism Diagnostic Interview-Revised (ADI-R). After injection of secretin, the ADI-R score increased in 8 patients, but declined in 3. Improvement was observed in functions such as sociability (interpersonal relationships), communication and speech improved, whereas in the others. symptoms such as hyperkinesias and stereotyped behavior became worse. The CSF levels of HVA was significantly increased in all of the patients showing an improvement in the ADI-R score. SHIAA levels also tended to increase, although this increase was not significant. These findings suggest that secretin promotes the metabolism of serotonin and dopamine in the central nervous system, which may contribute to improvement in clinical symptoms of autism.

Neurotrophic factors in the pathogenesis of Rett syndrome.

Rett syndrome is characterized by disruption of a period of vigorous brain growth with synapse development. Neurotrophic factors are important regulators of neuronal growth, differentiation, and survival during early brain development. The aims of this study were to study the role of neurotrophic factors in Rett syndrome, specifically whether Rett syndrome has abnormal levels of specific neurotrophic factors in serum and cerebrospinal fluid and whether the changes differ from other neuropediatric patients, for example, those with infantile autism. Four neurotrophic factors were measured: nerve growth factor, brain-derived neurotrophic factor, glial cell line-derived neurotrophic factor, and insulin-like growth factor 1 from the frozen cerebrospinal fluid and from serum (except glial cell line-derived neurotrophic factor) by enzyme-linked immunosorbent assay and cerebrospinal fluid glutamate and aspartate by high-performance liquid chromatography (HPLC) method in patients with Rett syndrome. Insulin-like growth factor 1 was measured from the cerebrospinal fluid of patients with infantile autism. We found low concentrations of cerebrospinal fluid nerve growth factor in patients with Rett syndrome compared with control patients. The serum levels and other cerebrospinal fluid neurotrophic factor levels of the patients did not differ from the controls. Patients with Rett syndrome had high cerebrospinal fluid glutamate levels. Patients with infantile autism had low cerebrospinal fluid insulin-like growth factor 1 levels. Nerve growth factor acts especially on cholinergic neurons of the basal forebrain, whereas insulin-like growth factor 1 acts on cerebellar neurons. In Rett syndrome, the forebrain is more severely affected than the other cortical areas. In autism, many studies show hippocampal or cerebellar pathology. Our findings are in agreement with the different morphologic and neurochemical findings (brain growth, affected brain areas, neurotransmitter metabolism) in the two syndromes. Impairment in dendritic development in Rett syndrome could be the consequence of cholinergic deficiency and of neurotrophic factor/glutamate imbalance. Cholinergic gene expression might be influenced by the Rett syndrome gene directly or via the neurotrophic factor system.

Low levels of insulin-like growth factor-I in cerebrospinal fluid in children with autism.

Autism is a behaviourally defined syndrome characterized by disturbances of social interaction and communication and restrictions of behaviour patterns and imagination. The pathogenesis of autism is unknown but it is suspected that a number of genetic factors may be involved. Neurotrophic factors such as insulin-like growth factor-I (IGF-I) play a role in early brain development. The aim of this study was to determine whether IGF-I levels might be associated with the development of autism. IGF-I levels were measured in the CSF of 11 children with autism (4 females, 7 males; mean age 3.8 years, SD 1.1) using a sensitive radioimmunoassay method and compared with levels in 11 control participants (6 females, 5 males; mean age 3.8 years). Levels of IGF-I in the CSF were statistically significantly lower in the children with autism than in the control children (p=0.03). IGF-I may play a role in pathogenetic mechanisms of autism and the role of neurotrophic factors in autism and other neurodevelopmental diseases should be studied further.

Decreased cerebrospinal fluid levels of beta-phenylethylamine in patients with Rett syndrome.

To clarify the mechanism of brain impairment in Rett syndrome, we measured the cerebrospinal fluid levels of beta-phenylethylamine (PEA) in 17 patients with Rett syndrome. Findings were compared with those obtained in age-matched controls and diseased controls. The cerebrospinal fluid level of PEA was significantly lower in patients with Rett syndrome than in the controls (31% of control values). The alteration in the cerebrospinal fluid level of PEA may reflect dopamine system impairment in Rett syndrome.

Levels of cerebrospinal fluid nerve-growth factor differ in infantile autism and Rett syndrome.

Autism and Rett syndrome (RS) are both developmental disorders of unknown origin. Autism is a behaviourally defined syndrome. RS, which affects girls only, is characterized by a profound learning disability following early normal development, with a consistent cluster of clinical features. Differentiation of RS from infantile autism in the very early stages of the disorders is not always easy. Both syndromes still lack discriminative laboratory markers for accurate diagnosis and differentiation. We decided to compare the CSF nerve-growth factor (NGF) levels of children with infantile autism and children with RS using enzyme-linked immunosorbent assay (ELISA). Our findings of mainly normal CSF NGF in autism and low to negligible values in RS are in agreement with the different morphological and neurochemical findings (brain growth, affected brain areas, neurotransmitter metabolism) in the two syndromes. CSF NGF could be used as a biochemical marker for differentiation of patients with autism from those with RS.