Essential fatty acids and phospholipase A₂ in autistic spectrum disorders

Abstract

A health questionnaire based on parental observations of clinical signs of fatty acid deficiency (FAD) showed that patients with autism and Asperger's syndrome (ASP) had significantly higher FAD scores (6.34±4.37 and 7.64±6.20, respectively) compared to controls (1.78±1.68). Patients with regressive autism had significantly higher percentages of 18:0,18:2n-6 and total saturates in their RBC membranes compared to controls, while 24:0, 22:5n-6, 24:1 and the 20:4n-6/20:5n-3 ratio were significantly higher in both regressive autism and ASP groups compared to controls. By comparison, the 18:1n-9 and 20:4n-6 values were significantly lower in patients with regressive autism compared to controls while 22:5n-3, total n-3 and total dimethyl acetals were significantly lower in both regressive autism and ASP groups compared to controls. Storage of RBC at −20°C for 6 weeks resulted in significant reductions in highly unsaturated fatty acid levels in polar lipids of patients with regressive autism, compared to patients with classical autism or ASP, or controls. Patients diagnosed with both autism and ASP showed significantly increased levels of EPA (∼200%) and DHA (∼40%), and significantly reduced levels of ARA (∼20%), 20:3n-6 and ARA/EPA ratio in their RBC polar lipids, when supplemented with EPA-rich fish oils, compared to controls and non-supplemented patients with autism. Patients with both regressive autism and classical autism/Asperger's syndrome had significantly higher concentrations of RBC type IV phospholipase A₂ compared to controls. However, patients with autism/ASP, who had taken EPA supplements, had significantly reduced PLA₂ concentrations compared to unsupplemented patients with classical autism or ASP.

Introduction

The prevalence of autistic spectrum disorders (ASD) has increased dramatically over the past 10 years and recent studies of children under 5 years found prevalences of 1 in 166 [1]. While the reasons for this increase are unclear, and likely to be multi-factorial, the potential burden on health, education and welfare budgets will be substantial. Current estimates are of lifetime costs around £3 m per patient with a total annual cost to the UK economy of £1 billion, possibly rising to £5 billion if numbers continue to escalate [2]. The steep increase in ASD diagnosis in Scottish schoolchildren is shown in Fig. 1.

Neuronal membranes are rich in lipids, especially phospholipids (PL), and can contain up to 80% lipid by weight [3]. Neural tissue contains high concentrations of highly unsaturated fatty acids (HUFA) in particular arachidonic acid (20:4n-6; ARA) and docosahexaenoic acid (22:6n-3; DHA). These fatty acids comprise between 15% and 30% of the dry weight of neural tissues [3]. The occurrence of a “phospholipid spectrum disorder”, whereby there is an increased loss of HUFAs from the sn 2 position of red blood cell (RBC) membrane PL, has been recorded in a number of studies in schizophrenic patients [4] and reductions in erythrocyte HUFA have also been observed in patients with attention-deficit hyper-activity disorder (ADHD) [5], [6]. Similar reductions in RBC membrane HUFA were also observed in a single case study of a patient with autism [7]. This loss of membrane HUFA may arise if the rate of removal exceeds the rate of repair and/or if the rate of phospholipid repair (synthesis) is reduced. Such changes can arise from overactive/over-expressed phospholipase A₂ (PLA₂) enzymes, which remove HUFA from the sn 2 position of membrane PL [8].

Parents were asked to complete a health questionnaire on their children which included a checklist of clinical indicators of fatty acid metabolism (excessive thirst, urination, dry skin, dry hair, dandruff, soft and brittle nails, follicular keratosis) which had been used previously to assess patients with ADHD and dyslexia [5], [9]. Each indicator was scored on a severity of 0–3 and a total fatty acid deficiency score (FAD) was obtained for each patient. These authors suggested that a FAD score >3 was indicative of EFA deficiency. In the present study the values were 6.34±4.37, 7.64±6.20 and 1.78±1.68 for the autism, Asperger's syndrome (ASP) and control groups, respectively (Fig. 2).

Major fatty acids of polar lipids from RBC of patients having regressive autism, classical autism/ASP and controls (not age matched) are shown in Table 1. In this study classical autism was described as autism apparent very early in development, probably being present from birth, whereas regressive autism described individuals who passed all developmental criteria up to 18–36 months and thereafter regressed into autism. Patients with regressive autism had significantly higher percentages of 18:0,18:2n-6 and total saturates in their RBC membranes compared to controls while 24:0, 22:5n-6, 24:1 and the ARA/EPA ratio were significantly higher in both regressive autism and ASP groups compared to controls. Conversely, the 18:1n-9 and ARA values were significantly lower in patients with regressive autism compared to controls while 22:5n-3, total n-3 and total dimethyl acetals were significantly lower in both regressive autism and ASP groups compared to controls. Changes in RBC fatty acid composition, especially reduced ARA and DHA, have been described in a number of other neurological disorders including schizophrenia [10], ADHD [6], depression [11] and bipolar disorder [12]. A recent study in children with ASD identified reduced levels of HUFA in plasma polar lipids [13]. Dimethyl acetals are derived from plasmalogens which are important mediators of brain function [14].

In comparison to samples from patients with classical autism/ASP, and controls, the HUFA composition of the regressive autism group changed dramatically following storage at −20°C. Percentages of all HUFA were reduced by 60–82% following 6 weeks storage at −20°C (Fig. 3). This dramatic loss of HUFA from RBC membranes in cold storage may indicate increased metabolism of phospholipids, due to elevated PLA₂ activity and/or concentration (phospholipid breakdown) or reduced fatty acyl CoA ligase/fatty acyl transferase activity and/or concentration (phospholipid synthesis) [8]. In addition, loss of HUFA may be due to increased lipid peroxidation [15].

The present study has identified significantly increased RBC type IV PLA₂ activity in both patients with regressive autism and classical autism/ASP, compared to controls (Fig. 4). A number of recent studies have identified elevated concentration of RBC PLA₂ in a range of neurological disorders including schizophrenia, depression, bipolar disorder and dyslexia [16].

The HUFA composition of RBC from patients with ASD were measured following supplementation with EPA-rich fish oils for at least 6 months (Table 2). Percentages of RBC 20:3n-6 were significantly reduced in supplemented patients compared to controls while both EPA and DHA were increased (∼200% and 40%, respectively) compared to controls and unsupplemented patients with autism. ARA percentages were significantly reduced (∼20%) following EPA supplementation. EPA supplementation has been shown to have significant benefits in a range of disorders including schizophrenia, depression, bipolar disorder, ADHD, dyslexia and dyspraxia [17]. Parents of children with autism who supplemented their children with EPA-rich fish oils reported improvements in general health and reduced infections, sleep patterns, cognitive and motor skills, concentration, eye contact and sociability, as well as reductions in irritability, aggression and hyperactivity. A few parents reported improved cognitive skills and concentration but increased hyperactivity and behavioural problems following supplementation. In some cases these problems could be overcome by giving an evening primrose oil (EPO) supplement prior to starting the EPA oils or increasing the EPO dosage while maintaining the EPA supplement. Interestingly, RBC PLA₂ concentrations in children with ASD who had been supplemented with EPA were significantly lower than unsupplemented children with classical autism/ASP. However, the efficacy of EPA supplementation could not be confirmed as the PLA₂ concentration had not been measured in the EPA group before supplementation.