Authors/ Authors:
Res. See. Dr. Caglar Charles Daniel JAICKS, Dr. Instructor Member Perihan ÇAM RAY, Prof. Dr. Özlem GÖRÜROĞLU, Prof. Dr. Gonca Gul CELIK, Prof. Dr. Ayşegül Yolga TAHİROĞLU, Dr. Instructor Member Zeliha HAYTOĞLU
SUMMARY
The aim of this study, which was carried out as a pilot study; The aim of this study is to measure serum melatonin levels and to investigate the relationship between sleep habits and clinical symptoms between groups of patients diagnosed with autism spectrum disorder and healthy controls. Cases aged 2-8 years diagnosed with autism spectrum disorder according to DSM-V and healthy children of similar age and gender in the control group were evaluated. Vitamin D level, ferritin, serum iron and iron binding capacity of all patients were checked, as well as endogenous melatonin levels, and a pediatric sleep habits questionnaire was filled. Autism Behavior Checklist (ABC – Autism Behavior Checklist) and Modified Early Childhood Autism Screening Scale (M-CHAT – The Modified Checklist for Autism in Toddlers) were applied to the autism spectrum disorder group. The relationship between the blood values obtained and the completed questionnaire results was evaluated. Melatonin levels were compared between the patients with autism spectrum disorder and the control group. A total of 71 patients were included. 81.7% (n=58) of the participants were male. The mean age of the autism spectrum disorder group was 44.4±20.4 months, while the mean age of the control group was 51.2±20 months (p=0.104). When the data obtained from the sleep questionnaire were compared between the two groups, the “daytime sleepiness” subscale score was higher in the control group, while the “bed-wetting” subscale score was higher in the autism spectrum disorder group (p=0.036 and p=0.008, respectively). There was no significant correlation between the survey results obtained from the patients and the melatonin levels obtained. Melatonin levels were 823.2±237.9 U/L in the autism spectrum disorder group and 677.4±254.7 U/L in the control group. This difference between the two groups was found to be statistically significant (p=0.027). When the patients were compared by dividing by gender, only male patients were found to differ between the two groups (p=0.020). It was observed that there was no significant difference between girls in terms of melatonin levels (p=0.608). In the light of the data we obtained in our study, it was observed that there were sleep problems in autism spectrum disorder and therefore, melatonin levels measured during the day were higher than in healthy children. This suggests that there may be desensitization to melatonin receptors in the ASD group, and that melatonin levels differ according to gender, suggesting that melatonin treatment modalities applied in ASD patients may be arranged according to gender.
Keywords: Melatonin, Autism Spectrum Disorder, Sleep Disorders
ABSTRACT
The aim of this study is to evaluate the relationship between serum melatonin levels, sleep habits and clinical features of children with autism spectrum disorder compared to healthy controls. Our research is planned as a controlled prospective project. In this study, participants were included among patients who applied to Cukurova University Child and Adolescent Psychiatry Department; children between ages 2 and 8, diagnosed with Autism Spectrum Disorder (ASD) according to DSM-V criteria. healthy children with similar ages and sex were included in the study to be the healthy control group; to be matched with ASD group. ASO, vitamin D level, ferritin, serum iron, iron binding capacity and melatonin levels of all patients were analyzed. Besides, child sleep habits questionnaire was applied to all participants. Autism Behavior Checklist (ABC) and The Modified Checklist for Autism in Toddlers (M-CHAT) was applied to ASD group. The data obtained from all pariticipants including the lab results, questtionnaires, checklists were evaluated. Serum melatonin levels of both groups were compared. A total of 71 participants were included in our research. Among the participants, 81.7% (n=58) was male. The average age, defined as months, in ASD group was 44.4±20.4 months, whereas the healthy controls average age was 51.2±20 months (p=0.104). After comparing the results of both groups obtained from Child Sleep Habits Questionnaire, “daytime sleepiness” subscale was higher in the control group, whereas “bedwetting” subscale in ASD group was higher (respectively p=0.036 and p=0.008). There was no correlation between the checklist scores and melatonin levels of the patients. The average melatonin levels in ASD group was 823.2±237.9 U/L, whereas 677.4±254.7 U/L in the healthy control group. This finding is statistically significant between the two groups (p=0.027). When the ASD group was categorized according to sex, only male patients showed statistically significance results (p=0.020). There was no statistical significance among female participants (p=0.608). The data obtained in this research reveals that in children with ASD there are sleep disorders and due to this fact the daytime serum melatonin levels is higher in ASD group compared to healthy controls. This could further be explained with melatonin receptor desensitization in ASD group. Besides, the serum melatonin levels showed difference among males and females and this could be a reason to modulate the exogenous melatonin treatment according to sex traits.
Keywords: Melatonin, Autism Spectrum Disorder, Sleep Disorders
Introduction: Due to cognitive functions such as sleep disorder, working memory and attention, it negatively affects the daily lives of the cases. It has been shown that the serum melatonin level and the level of melatonin metabolites in the urine are low in ASD cases. [i] In physiological conditions, the concentration of melatonin is regulated according to the circadian rhythm, low during the day and high at night.[ii] Peak secretion is usually around 2 in the morning and is also called the dark hormone. Polymorphism in Acetyl-Serotonin O-Methyl Transferase may be related to low melatonin in ASD. [iii]Melatonin receptor agonists can treat insomnia and behavioral symptoms by reducing nitrosative/oxidative stress and inflammation in ASD. Melatonin levels also differ by gender. It is 2-3 times more common in girls. For this reason, ASD may be less common in girls (as it prevents oxidative damage to DNA). In a study conducted with 14 ASD cases, serum melatonin 24-hour circadian rhythm concentration was checked, lower levels were found in ASD at night compared to healthy controls, no circadian variation was found in the 10-14 age group, and an inverted rhythm was found in the 4-14 age group. [iv]
Materials and Methods: Our study was designed as a controlled and prospective study. Our study was started with the approval of Çukurova University Faculty of Medicine Non-Invasive Clinical Research Ethics Committee. Our work; Cases aged 2-8 years who applied to our outpatient clinic between August 2021 and January 2022 and were diagnosed with Autism Spectrum Disorder according to DSM-V, and a healthy control group matched for sex and age were included in the study. Medical diseases known to affect cognitive and brain function, cancer, bipolar disorder, psychotic disorders, epilepsy, any diagnosis of neurological diseases, electroconvulsive therapy in the last 3 months, intraocular surgery or laser therapy, ocular diseases such as retinitis pigmentosa, Traveling between continents in the last 1 month, sleep apnea and sleep disorders due to a medical condition, autoimmune diseases, being deaf or dumb, psychotropic, neurotropic, SSRI, anti-psychotic, SNRI, mood stabilizer, stimulant and benzodiazepine group medication Exclusion criteria are those who use drugs and other drugs known to change sleep patterns.
Psychometric Measurements
DSM– V diagnostic criteria, Autism Behavior Checklist (ABC-Autism Behavior Checklist), Modified Checklist for Autism in Toddlers / M-CHAT, Child Sleep Habits Questionnaire.
Laboratory data: ASO, vitamin D level, ferritin, serum iron and iron binding capacity, as well as endogenous melatonin levels, which are routinely requested from patients diagnosed with ASD, were measured. Blood samples were collected in ACD-A tubes between 8.30-10.30 in the morning (background light intensity <2 lx, participants were allowed to sleep between 23:00 and 06:00 in the evening). In addition, the function of the pineal gland can be roughly evaluated in studies where melatonin metabolites are evaluated non-invasively. For this reason, it was decided to measure blood melatonin levels in our study. For the measurement of serum melatonin level, 5 ml blood sample was taken from the brachial vein of the forearm, and centrifugation was applied. The obtained serum samples were stored as 2 ml at -80°C. All patient and control blood samples were taken into Becton Dickinson vacuum-gel tubes. Serum was obtained by centrifugation at 2000 rpm for 20 minutes. Serum MELATONIN (SUNRED brand BT LAB Cat.NO EA0013Ge) level was measured by enzyme-linked Immunosorbent assay (ELISA) in accordance with the kit protocols. Serum samples were diluted according to the kit procedure. Serial dilutions were made according to the standards kit procedure. 50 microliters of standard or 40 microliters of patient and control serum were added to each determined well. 10 microliters of Biotin Antigen was added to the samples. They were incubated at 37oC at the specified time intervals in accordance with the procedure. 50 mL of stop solution was added. It was immediately read at 450 nm. The resulting concentration values were multiplied by the dilution coefficient. Serum melatonin level concentration was found. Serum melatonin levels were measured as pg/mL. In addition, chemiluminescence method with DXI 800 device for B12 and ferritin, photometric method with AU5800 device for iron and iron binding capacity, immunoturbidimetric method with IMMAGE 800 device for anti-streptolysin O and HPLC method with Thermo Dionex device for vitamin D were used. Melatonin levels of patients diagnosed with ASD and healthy controls were compared.
Statistics: Statistical analysis of the data was performed using the SPSS 25.0 (SPSS Inc., Chicago, Illinois) program. Normality test of numerical variables was determined by Kolmogorov-Smirnov test. Numerical variables were summarized as mean (±standard deviation). Categorical variables were given as numbers and percentages. When comparing numerical variables between the two groups, Mann Whitney U test was used because the variables did not show normal distribution. Chi-square and Fisher’s exact tests were used when comparing categorical variables. The correlation between the measurement variables was compared with the Pearson correlation test and the correlation coefficients were calculated. Statistical significance level was taken as 0.05 for each test.
Results: 38 patients with autism spectrum disorder and 33 healthy children evaluated in the control group were included in the study. The mean age of the patients with autism spectrum disorder was 44.4±20.4 months, while the mean age of the children in the control group was 51.2±20 months (p=0.104). While the male sex ratio was 86.8% in the autism spectrum disorder group, this ratio was 75.8% in the control group (p=0.228) (Table 1). When the laboratory data obtained from the patients were compared, it was observed that vitamin B12 levels were significantly higher in the group with autism spectrum disorder (p=0.019). There was no significant difference between the two groups in terms of hemogram values (p>0.05). Likewise, no significant difference was observed between the two groups in terms of iron, iron binding capacity, ferritin and vitamin D values (p>0.05). Anti-streptolysin O levels were found to be significantly lower in the autism spectrum disorder group (p=0.012), (Table 2). When the data obtained from the child sleep habits questionnaire were compared between the two groups, it was observed that there was a difference between the two groups only in terms of “daytime sleepiness” and “bed-wetting” subscales (p=0.036 and p=0.008, respectively). While the daytime sleepiness score was lower in patients with autism spectrum disorder than in the other group; Bedwetting score was higher. It was observed that there was no difference between the two groups in terms of other data and subscales (p>0.05), (Table 3). When the patients with autism spectrum disorder are clinically divided into mild, moderate and severe, and examined in terms of laboratory findings; Vitamin D levels of patients diagnosed with severe autism spectrum disorder were found to be significantly lower than other groups (p=0.005). Likewise, the iron binding capacity was found to be significantly lower in patients with severe ASD (p=0.043). It was determined that there was no significant correlation between melatonin and the scores obtained from the M-Chat and ABC scales (p>0.05), (Table 4). Likewise, no significant correlation was observed between melatonin and other laboratory parameters (p>0.05), (Table 5). It was observed that there was no significant difference between the three groups in terms of melatonin and other laboratory parameters (p>0.05). Melatonin levels were 823.2±237.9 U/L in the autism spectrum disorder group and 677.4±254.7 U/L in the control group. This difference between the two groups was found to be statistically significant (p=0.027). When the patients were compared by dividing by gender, only male patients were found to differ between the two groups (p=0.020). It was observed that there was no significant difference between girls in terms of melatonin levels (p=0.608), (Table 6). There was no significant correlation between melatonin and scores obtained from M-Chat and ABC scales (p>0.05). Likewise, no significant correlation was observed between melatonin and other laboratory parameters (p>0.05), (Table 7).
Discussion: The male sex ratio was 86.8% in the ASD patients included in our study, and the male sex ratio was 6.6 times higher. The differences in ASD according to gender are thought to be due to genetic characteristics. [v]In our study, red blood cell count, white blood cell count, hemoglobin, hematocrit, MCV and platelet values were found to be quite similar between children with ASD and healthy children (p>0.05). Vitamin B12 levels were found to be significantly higher in patients with ASD (respectively p=0.019). Anti-streptolysin O values were found to be significantly lower in ASD patients (p=0.012). Vitamin D level and iron binding capacity; were found to be significantly lower in the severe ASD group compared to the other groups (p=0.005 and p=0.043, respectively). In our study, when healthy children and children with autism spectrum disorder were compared in terms of sleep problems, it was observed that the total sleep duration was similar between the two groups (p=0.479). In terms of staying awake at night, it was found that patients with ASD had significantly longer awake times, but this difference was not statistically significant (p=0.052). There were no significant differences between the ASD and control groups in terms of difficulty in waking up in the morning, sleep fragmentation, sleep anxiety, sleep disturbance, parasomnia, morning awakening style, sleep duration, sleep transition, and need to sleep with others (p>0.05). It was determined that daytime sleepiness problem was observed less in ASD patients compared to the control group, but bedwetting problems were observed more frequently (p=0.036 and p=0.008, respectively). It has been determined that melatonin levels are high in ASD patients and melatonin levels decrease as the clinical situation worsens. In addition, in this study, although melatonin levels were lower than the control group in measurements made at night, melatonin levels were found to be higher than healthy children in measurements made during the daytime. [vi]In a meta-analysis study examining the relationship between ASD and melatonin, it was found that melatonin levels were lower in ASD patients than in healthy individuals in seven different studies. [vii] In addition, four different studies have shown that melatonin levels decrease as the clinical condition of ASD gets worse. [viii] However, in almost all of these studies, melatonin levels were measured at night. In the study of Ritvo et al., it was observed that the melatonin levels measured during the day were higher in patients with ASD. [ix] In our study, it was observed that serum melatonin levels were significantly higher in ASD patients than in healthy controls (p=0.027). In studies where melatonin levels were measured at night, lower melatonin levels were found in patients with ASD, while elevated melatonin levels were observed in patients with ASD in daytime melatonin levels. [x]This is an indication that the circadian rhythm of melatonin is impaired in patients with ASD. Although there are still missing data today, some genes that cause disruption of melatonin synthesis are thought to play a role in the etiology of ASD. In addition, the presence of some gene mutations that play a role in melatonin receptor desensitization is also blamed in the etiology of ASD. [xi]Melatonin receptor desensitization, which is thought to be observed in ASD patients, may also play a role in the ASD clinic. The increased daytime melatonin levels obtained in our study are in parallel with the literature. There is no current study in the literature examining gender-related melatonin values in patients with autism spectrum disorder. Studies in healthy people have shown that women’s melatonin levels are higher than men, and sometimes the difference can be up to 3 times higher. In our study, melatonin levels were found to be significantly higher in male patients between ASD patients and the control group (p=0.020). It was observed that melatonin levels were higher in girls with ASD, but this difference was not significant (p=0.608). High melatonin levels observed in women are thought to be a protective factor against ASD. It is a data that has been revealed in previous studies, especially that the circadian rhythm of melatonin works differently in ASD patients and that it is an inverted rhythm.
Limitations of the Study: One of the biggest limitations of our study is the evaluation of sleep habits in children using a questionnaire method that provides subjective data. In addition, at the time of blood analysis, the number of hours the participant slept in the previous night, what time he woke up in the morning or what time he slept at night was ignored. Since the number of girls among the applicants is low, it is thought that future studies should be conducted with multicenter, cohort and large study groups, with female gender-weighted participants, and with sleep monitoring in order to evaluate the sleep of the children studied more objectively, more precise results can be obtained.
Conclusion: Although it has already been stated in the literature that the relationship between autism spectrum disorder and melatonin levels is significant, that there may be an inverted oscillation rhythm, and that there is a significant relationship between sleep habits and clinical severity of autism, our study included patients who did not use any medication that could affect sleep and melatonin. selected, it differs from previous studies. We think that evaluation with larger and perhaps 24-hour serum samples to be made in this area, and evaluation with study groups with more female gender will contribute significantly to the literature.
Acknowledgment: Our project was supported by the Scientific Research Project Board of our university.
Attachments:
Table 1. Demographic data
|
OSB (n=38) |
Control (n=33) |
p |
|
|
Age (months), Mean±Sd |
44.4±20.4 |
51.2±20 |
0.104* |
|
Gender, n (%) |
0.228** |
||
|
Girl |
5 (13.2%) |
8 (24.2%) |
|
|
Male |
33 (86.8%) |
25 (75.8%) |
*: Mann-Whitney U test **: Chi-square test ASD: Autism spectrum disorder, Mean±Sd: Mean±standard deviation
Table 2. Comparison of laboratory data between groups
|
OSB (n=38) Mean±Sd |
Control (n=33) Mean±Sd |
p* |
|
|
RBC (1012/L) |
4.5±0.8 |
4.6±0.3 |
0.773 |
|
WBC (109/L) |
8.7±2.8 |
8.5±1.9 |
0.572 |
|
HGB (g/dL) |
12.1±2.2 |
12.1±0.8 |
0.415 |
|
HTC (%) |
35.3±6.5 |
35.6±2.3 |
0.471 |
|
MCV (fL) |
75.6±15.6 |
77.4±3.6 |
0.199 |
|
PLT (109/µL) |
336.5±109.2 |
345.6±109.5 |
0.904 |
|
Vitamin B12 (mg/L) |
366.1±180.2 |
257.2±284.9 |
0.019 |
|
Iron (g/dL) |
68±35.9 |
77.9±40.1 |
0.333 |
|
Iron binding capacity (g/dL) |
282.6±110.8 |
303.6±77.9 |
0.42 |
|
Ferritin (ng/mL) |
25±18.8 |
24.2±26.5 |
0.365 |
|
Vitamin D (ng/mL) |
20±13.7 |
16.8±9.5 |
0.278 |
|
Anti streptolysin-O (IU/mL) |
27.4±8.9 |
67.6±85.7 |
0.012 |
*: Mann-Whitney U test
ASD: Autism spectrum disorder, Mean±Sd: Mean±Standard deviation, RBC: Erythrocyte, WBC: White blood cell; HGB: hemoglobin, HCT: Hematocrit, MCV: Mean erythrocyte volume, PLT: Platelet,
Table 3. Comparison of the results of the child sleep habits questionnaire
|
OSB (n=38) Mean±Sd |
Control (n=33) Mean±Sd |
p* |
|
|
Total sleep time |
10.2±1.9 |
10.4±1.7 |
0.479 |
|
Night awake time |
45.5±54.9 |
18.3±24 |
0.052 |
|
Difficulty waking up in the morning |
5.7±2.2 |
5.8±1.9 |
0.703 |
|
Division of Sleep |
7.3±2.4 |
6.9±2 |
0.538 |
|
sleep anxiety |
5.7±2.1 |
6±1.8 |
0.456 |
|
Sleep disturbance |
3.8±1.5 |
4±1.4 |
0.303 |
|
parasomnia |
4.3±1.5 |
4.5±1.4 |
0.492 |
|
How to wake up in the morning |
2.5±0.8 |
2.9±1 |
0.079 |
|
sleep time |
4.7±1.8 |
4.5±1.7 |
0.732 |
|
transition to sleep |
3.4±1.3 |
3.2±1.1 |
0.531 |
|
the need to sleep with others |
4.2±1.3 |
3.6±1.3 |
0.072 |
|
daytime sleepiness |
3.2±1.2 |
3.8±1.3 |
0.036 |
|
bedwetting |
3.3±1.3 |
2.5±0.8 |
0.008 |
|
Total |
*: Mann-Whitney U test
ASD: Autism spectrum disorder, Mean±Sd: Mean±standard deviation
Table 4. M-Chat and ABC evaluation results of patients with autism spectrum disorder
|
OSB (n=38) Mean±Sd |
|
|
Risky questions (M-Chat) |
5±2.9 |
|
Total (M-Chat) |
13.2±4.8 |
|
Sensory (ABC) |
11.2±5.9 |
|
Relationship Building (ABC) |
15.9±8.1 |
|
Body and object use (ABC) |
16.6±9.5 |
|
Language skills (ABC) |
14.1±5.8 |
|
Social and self-care (ABC) |
12.6±4.7 |
|
Total (ABC) |
70.4±25.3 |
ASD: Autism spectrum disorder, ABC: Autism Behavior Checklist M-Chat: Modified Infancy Autism Screening Scale Mean±Sd: Mean±Standard deviation
Table 5. Comparison of laboratory data of autism spectrum disorder patients according to their clinical status
|
Clinical ASD Level |
||||
|
Light (n=17) Mean±Sd |
Middle (n=17) Mean±Sd |
Heavy (n=4) Mean±Sd |
p* |
|
|
RBC (1012/L) |
4.3±1.1 |
4.6±0.3 |
4.8±0.3 |
0.334 |
|
WBC (109/L) |
8.6±3.1 |
8.9±2.8 |
8.2±0.9 |
0.907 |
|
HGB (g/dL) |
11.5±3.1 |
12.6±1.1 |
12.7±1 |
0.361 |
|
HTC (%) |
33.7±9 |
36.3±3.1 |
37.4±2.3 |
0.393 |
|
MCV (fL) |
71.2±22.4 |
79.2±3.8 |
78.6±7.1 |
0.308 |
|
PLT (109/µL) |
313.2±107 |
350.9±121.2 |
374.5±35.3 |
0.471 |
|
Vitamin B12 (mg/L) |
411.7±199.4 |
330.1±174.9 |
325±62 |
0.383 |
|
Iron (g/dL) |
68.2±29.9 |
73.6±37.2 |
43.5±52.3 |
0.328 |
|
Iron binding capacity (g/dL) |
296.2±94.7 |
299.3±92.9 |
153.5±181.2 |
0.043 |
|
Ferritin (ng/mL) |
23.7±13.5 |
28.7±24.2 |
14.9±7.8 |
0.397 |
|
Vitamin D (ng/mL) |
26±15.7 |
17.9±8.6 |
3.3±4.3 |
0.005 |
|
Anti-streptolysin O (IU/mL) |
29.5±12.7 |
25.9±3.8 |
25±0 |
0.442 |
|
Melatonin (U/L) |
805.7±244.1 |
872±237.8 |
690±200 |
0.367 |
*: Mann-Whitney U test
ASD: Autism spectrum disorder, Mean±Sd: Mean±Standard deviation, RBC: Erythrocyte, WBC: White blood cell; HGB: hemoglobin, HCT: Hematocrit, MCV: Mean erythrocyte volume, PLT: Platelet,
Table 6. Comparison of melatonin levels between groups
|
Melatonin Level (U/L) |
p* |
||
|
OSB Mean±Sd |
Control Mean±Sd |
||
|
Girl (n=13) |
838.6±221.8 |
766.2±254.7 |
0.608 |
|
Male (n=58) |
820.8±243.5 |
649.0±262.8 |
0.020 |
|
Total (n=71) |
823.2±237.9 |
677.4±254.7 |
0.027 |
*: Mann-Whitney U test
ASD: Autism spectrum disorder, Mean±Sd: Mean±standard deviation
Table 7. Correlation analysis of melatonin and other parameters
|
Parameters |
melatonin |
|
|
Total (M-Chat) |
r |
0.063 |
|
Total (ABC) |
r |
-0.066 |
|
Anti-streptolysin O (IU/L) |
r |
-0.141 |
|
Vitamin D (ng/mL) |
r |
-0.062 |
|
Ferritin (ng/mL) |
r |
0.010 |
|
Vitamin B12 (mg/L) |
r |
-0.004 |
|
Hemoglobin (g/dL) |
r |
0.196 |
|
Hematocrit (%) |
r |
0.201 |
r: Pearson correlation coefficient
ABC: Autism Behavior Checklist M-Chat: Modified Infancy Autism Screening Scale
resources
[i] Rossignol DA, Frye RE. Melatonin in autism spectrum disorders. Current clinical pharmacology. 2014;9(4):326-34.
[ii] Liebrich LS, Schredl M, Findeisen P, Groden C, Bumb JM, Nölte IS. Morphology and function: MR pineal volume and melatonin level in human saliva are correlated. J Magn Reson Imaging. 2014 Oct;40(4):966-71.
[iii] Doyen C, Mighiu D, Kaye K, Colineaux C, Beaumanoir C, Mouraeff Y, et. get. Melatonin in children with autistic spectrum disorders: recent and practical data. Eur Child Adolesc Psychiatry. 2011 May;20(5):231-9.
[iv] Kulman G, Lissoni P, Rovelli F, Roselli MG, Brivio F, Sequeri P. Evidence of pineal endocrine hypofunction in autistic children. Neuroendocrinology Letters. 2000;21(1):31-4.
[v] Melke J, Goubran Botros H, Chaste P, Betancur C, Nygren G, Anckarsäter H, et al. Abnormal melatonin synthesis in autism spectrum disorders. Mol Psychiatry. 2008 Jan;13(1):90-8.
[vi] Tordjman S, Anderson GM, Pichard N, Charbuy H, Touitou Y. Nocturnal excretion of 6-sulphatoxymelatonin in children and adolescents with autistic disorder. biological psychiatry. 2005;57(2):134-8.
[vii] Nir I, Meir D, Zilber N, Knobler H, Hadjez J, Lerner Y. Brief report: circadian melatonin, thyroid-stimulating hormone, prolactin, and cortisol levels in serum of young adults with autism. Journal of autism and developmental disorders. 1995;25(6):641-54.
[viii] Melke J, Goubran Botros H, Chaste P, Betancur C, Nygren G, Anckarsäter H, et al. Abnormal melatonin synthesis in autism spectrum disorders. molecular psychiatry. 2008;13(1):90-8.
[ix] Ritvo ER, Ritvo R, Yuwiler A, Brothers A, Freeman BJ, Plotkin S. Elevated daytime melatonin concentrations in autism: A pilot study. Eur Child Adolesc Psychiatry. 1993;2(2):75-8.
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[xi] Jonsson L, Ljunggren E, Bremer A, Pedersen C, Landén M, Thuresson K, et al. Mutation screening of melatonin-related genes in patients with autism spectrum disorders. BMC medical genomics. 2010;3(1):1-7.
resources
[1] Rossignol DA, Frye RE. Melatonin in autism spectrum disorders. Current clinical pharmacology. 2014;9(4):326-34.
[1] Liebrich LS, Schredl M, Findeisen P, Groden C, Bumb JM, Nölte IS. Morphology and function: MR pineal volume and melatonin level in human saliva are correlated. J Magn Reson Imaging. 2014 Oct;40(4):966-71.
[1] Doyen C, Mighiu D, Kaye K, Colineaux C, Beaumanoir C, Mouraeff Y, et. get. Melatonin in children with autistic spectrum disorders: recent and practical data. Eur Child Adolesc Psychiatry. 2011 May;20(5):231-9.
[1] Kulman G, Lissoni P, Rovelli F, Roselli MG, Brivio F, Sequeri P. Evidence of pineal endocrine hypofunction in autistic children. Neuroendocrinology Letters. 2000;21(1):31-4.
[1] Melke J, Goubran Botros H, Chaste P, Betancur C, Nygren G, Anckarsäter H, et al. Abnormal melatonin synthesis in autism spectrum disorders. Mol Psychiatry. 2008 Jan;13(1):90-8.
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