Analysis of lead, arsenic and calcium content in the hair of children with autism spectrum disorder

Autism Spectrum Disorder (ASD) is one of the most common neurodevelopmental impairments (in 2014 it was diagnosed in 1 out of 68 children) and is four times more common in boys than girls [1] (https://​www.​cdc.​gov/​ncbddd/​autism/​data.​html). This ratio is higher in those with a mild course of the disorder compared to individuals with an acute course of autism. Over 50% of children with autism have an intellectual disability, while every third child develops epilepsy and every second child suffers from speech disorders. Apart from intellectual deficits, ASD may also be accompanied by metabolic disorders, motor organ development abnormalities, and chronic somatic diseases [1] (https://​www.​cdc.​gov/​ncbddd/​autism/​data.​html).

In previous years, an increase in the number of diagnosed ASD cases has been observed, which allows to hypothesize that chemical contamination in the environment influences these sorts of developmental disorders [2‐4]. Studies indicate the neurotoxic effect of numerous substances, including heavy metals (on mitosis, cell differentiation, synapse formation, oxidative stress, endoplasmic reticulum stress, and essential metalloprotein disruption, apoptotic processes, level of neurotransmitters). Lead (Pb) and arsenic (As) are well-established neurotoxicants known to cross the blood–brain barrier and affect neurodevelopment [1, 5‐7]. To give an example, Pb impairs intellectual development, causes behavioral disorders and motor hyperactivity. It can also have adverse effects on the health of children, causing behavioral and neurological problems, and a reduction in IQ scores [1, 5, 6, 8‐11]. Similar effects can be observed in the case of As. Arsenic exposure significantly affects brain morphology, resulting in gliosis, neuronal degeneration, a decrease in cognitive abilities, attention, comprehension, language skills, and reduces intelligence quotient (IQ) scores [1, 5, 6, 11‐14]. The high levels of heavy metal antagonists (e.g. Ca) in the body act protectively to a certain degree. However, a Ca deficiency can further increase the toxic effect of Pb and As [7, 8, 15].

ASD is a neurodevelopmental disorder, this suggests a possible role of heavy metals in its underlying cause (induction). However, investigations on associations between ASD and neurotoxic heavy metals are inconclusive. Over the last decades, numerous studies have reported a relationship between ASD and Pb and As exposure [5, 11‐13, 16‐24]. However, there are also some reports that conclude that heavy metals exposure is not a risk factor for ASD [6, 14, 25‐29]. Therefore, an accurate evaluation of the relationship between ASD and heavy metals needs more specific research. This study is the first in Poland and may be an important contribution to the body of knowledge on the subject of metal concentration in the hair of children with ASD.

The aim of the study was to analyze Pb, As, and Ca concentrations in the hair of children with ASD and a control group.

Table 1 presents a comparison of mean levels of concentration, medians, scopes and standard deviations of selected trace elements in the hair of autism spectrum children and those without neurological disorders.

Mean Ca level in the hair of children with ASD (254.7 ± 91.96 mg kg¹) was lower than the mean level of this element in the control group (312.8 ± 86.89 mg kg^− 1) (statistically significant differences, p = 0.002). Mean As concentration in the hair of children with ASD was 0.216 ± 0.09 mg kg^− 1 and was statistically significantly higher (p < 0.001) than the mean concentration of this element in the hair of children without neurological disorders (0.061 ± 0.03 mg kg^− 1). According to observations, mean Pb concentration in the hair of children with ASD was 2-fold higher compared to mean Pb concentration in the control group (6.028 ± 0.69 mg kg^− 1 and 3.415 ± 1.207 mg kg^− 1, respectively) (statistically significant differences, p < 0.001) (Table 1, Fig. 1).

We found weak negative correlations between the levels of the analyzed metals. In the hair of children with ASD Ca was negatively correlated with Pb (r = − 0.18, p > 0.05) and positively with As (r = 0.11, p > 0.05). Statistically significant moderate positive correlations between Pb and As (r = 0.36, p < 0.05) in the hair of children with ASD were observed (Table 2).

There is a list of 201 industrial chemical agents that have a neurotoxic effect on humans (including Pb and As). They may cause autism and other disorders such as attention deficit, mental retardation, and cerebral palsy [1, 16, 17, 23, 24].

Arsenic (As) is an element that is common in nature and is toxic. It is absorbed into the body primarily through the digestive tract and the respiratory system. In the human body, As compounds may inhibit the activity of over 200 enzymes. The effects of chronic As intoxication include neurotoxic effects in the central and the peripheral nervous system. The symptoms may manifest as sensory changes, muscle sensitivity, sensation of stinging and tingling (paresthesis), weakness, progressing muscle flaccidity, and polyneuropathy with sensory loss. A particularly unfavorable impact is exerted on the central nervous system, and in the case of pregnant women on the development of the fetal nervous system, since it crosses the blood-placenta barrier and accumulates in fetal epithelial tissue in the early stages of pregnancy. In children whose mothers were exposed to As compounds, the following symptoms are observed: difficulty with learning, memory, concentration, cognitive and behavioral disorders. Children exposed to this element may also present the symptoms of retarded development or inhibited physical and mental development [1, 5, 6, 13, 23, 29‐35].

Lead (Pb) is one of the most dangerous metals due to its common occurrence in the environment. It is highly toxic, has the ability to easily cross biological barriers, and accumulates in the internal organs [16, 17, 36]. Pb may affect the functions of both the central and peripheral nervous system as well as the senses. Numerous studies have confirmed that excessive exposure to Pb results in, among others, convulsions, changes in brain functions, encephalogram changes as well as acute encephalopathy and other brain disorders. In cases when Pb concentration in the plasma is over 30 mg·L^− 1, gradual changes in behavior and cognitive functions may occur manifesting as irritability, sensory disorders, apathy, impaired visual-motor coordination, and longer time of responding to stimuli [1, 5, 6, 8, 15, 22‐24, 29, 37‐40].

The nervous systems of fetuses and small children show higher sensitivity to Pb intoxication compared to adults. This results from a greater susceptibility of their brains to disorders (especially in the period of fast growth). Functional changes of the brain develop in children at over 10 mg Pb/10^− 1 L of blood causing most of all decreased IQ. Also, Pb in children leads to disorders of cognitive functions, difficulty learning (mostly reading, learning languages, and mathematics), lack of concentration, impairment of a short-time memory, recurring epileptic seizures, irritability, hyperactivity as well as impairment of speech, vision and even personality (e.g. aggression) [6, 15, 31, 32, 37‐42].

In our research, mean As and Pb content in the hair of children with ASD was statistically significantly higher compared to the mean content of these elements in the hair of children from the control group (Table 1, Fig. 1). Numerous studies have confirmed that heavy metals play a crucial role in the development of autism spectrum disorders [1, 5, 11‐13, 16‐22, 24]. Yasuda et al. [16, 17] demonstrated increased Pb concentrations in 4.8% of children with ASD and As in 2.6% (maximal Pb concentration amounted to 24.9 ppm, while As 1.7 ppm). Fido and Al-Saad [25] also observed a significantly higher Pb concentration in 4–7 year old children with autism compared to normal individuals (6.75 vs 3.20 mg·kg^− 1). Similar Pb and As content were found in the hair of ASD children from different parts of the world, Saudi Arabia [11, 12, 18, 43], Kuwait [25], Oman [19], India [20], Egypt [21, 22], Italy [26], and the USA [5, 29, 44].

A study conducted on a large group of Mexican children showed a positive correlation between Pb concentration in the blood and increased severity of overactivity symptoms. Pb content showed no correlation with attention disorder, which is characteristic of full blown ADHD (attention-deficit hyperactivity disorder) [45]. Meta-analysis of 33 studies published in the years 1972–2010 conducted on 10,232 children showed a positive dependency between Pb concentration in the blood and hair and overactivity and attention disorders [41]. According to observations, low Pb concentration in the blood (< 3.4 mg·L^− 1) may be a factor that additionally increases the risk of genetically conditioned ADHD. It was demonstrated that exposure to Pb causes cognitive deficits similar to those observed in ADHD children as well as impaired attention and cognitive function. Also, studies conducted on children indicated that Pb impairs the processes of inhibiting response and attention in school-age children. It has also been proven that in the case of blood Pb concentration > 2.0 mg·L^− 1 the risk of developing ADHD increases over four times [46].

Numerous clinical control studies have proven increased levels of one or more toxic metals in urine, blood, hair, nails, teeth, and brain samples in autism spectrum children compared to children showing normal development [24, 47]. Al-Ayadhi [18] and Blaurock- Busch et al. [11, 12] observed significantly higher concentrations of heavy metals (Pb and As) in the hair of children with ASD compared to healthy children. Adams et al. [27, 47] also demonstrated that children with autism had higher Pb and As concentrations in urine (by 72 and 21%, respectively) and in the hair (by 30 and 20%, respectively) compared to healthy children.

There is increasing evidence that children with ASD have a considerable disability to process and eliminate toxins from the body [1, 5, 6, 14, 43]. Toxin accumulation leads to increased activity of free radicals in the body, which in turn affects the structure of the nervous system. It is worth noticing that detoxification and eliminating heavy metals involves their conjugation with glutathione, whose concentrations are considerably lower in ASD individuals [19].

Modern people’s exposure to the risk is complex. Mutual interactions may occur between particular elements. Toxic metals affect trace element absorption, while the interaction between essential elements and toxic metals affects threshold values and toxicity effects. Toxic metals may interact metabolically with nutritionally essential metals [11, 12, 18]. Resorption of some ions (Ca²⁺, Mg²⁺, Fe²⁺, Zn²⁺ or Cu²⁺) from the digestive tract is inhibited by Pb, which is indicative of absorption competitiveness of these elements in the intestines. Also, Pb impairs the metabolism of minerals and causes multisystemic toxic effects through the inactivation of many enzymes (it binds to sulfhydryl, carboxyl and amino groups of aminoacids and proteins), leads to impaired effects of vital cations (Ca, Zn and iron (Fe)), abnormal oxidation-reductive condition of cells, impaired structure of cellular membrane and receptor functions. Increased Pb content enhances Ca elimination, while decreased Ca content in the diet causes a higher accumulation of this metal, especially in bones [8, 15, 38, 39]. A Ca deficiency may additionally strengthen the toxic effects of Pb and affect cognitive and behavioral development in children. A significant converse dependency between Ca intake with diet and Pb concentration in the blood was observed as early as in the 1990s in 3000 American children in the NHANES II study [7].

According to our research, we found weak negative correlations between the levels of the analyzed metals. In the hair of children with ASD, Ca was negatively correlated with Pb and positively with As. Statistically significant moderate positive correlations between Pb and As in the hair of children with ASD were observed (Table 2). The antagonistic effects of toxic metals and bioelements have been confirmed by a number of studies [10‐12, 16, 18, 20, 27, 47‐50]. The mechanism through which Ca impairs As absorption and conversely has not been fully understood so far. Ca is able to participate in the inhibition and absorption of As through competitiveness connected with a common binding site with proteins in the intestines [16, 17, 33, 47]. Heavy metals, e.g. Pb and As, have a very similar toxic effect, which in turns leads to the increased severity of intoxication symptoms with concurrent exposure to more than one metal [1, 5, 6, 24, 33].

Ca is a mineral necessary for the normal functioning of the whole body. It regulates the nervous system by improving the transmission of nerve impulses (it is a transmitter of arousal states in nerve synapses). Low concentrations of Ca in the body cause the following symptoms: irritability and excitability, nervousness, oversensitivity and anxiety states. Chronic Ca deficiency in children may lead to mental disorders [7, 51].

In our study, mean Ca content in the hair of children in the ASD case group was lower than the mean content of this element in the control group (statistically significant differences) (Table 1, Fig. 1). Many authors have also observed significantly lower Ca content in children with autism compared to healthy children. This suggests that children with ASD are susceptible to Ca deficiency [16‐18]. The presented results have been reflected in studies by Japanese researchers who noticed a Ca deficiency in approximately 6% of children. A considerable deficiency of this element has been diagnosed mainly in children under 10 years old. The minimal Ca concentration in this group amounted to approximately 70 ppm [16, 17]. Apart from Ca deficiency, many children with autism also had a deficiency of other elements, including Zn, Mg, Mn or Cu [12, 16, 17, 19, 49]. It was also demonstrated that children with autism had significantly lower Ca content in the hair (by 20%) compared to normal children [27].

Despite a number of studies and analyses, autism is still not understood by scientists, because no specific factor causing this condition has been found. Each experiment provides new valuable data but fails to deliver an explanation for symptom occurrence.