Table 2.

Genetic alterations in circadian rhythm of ASD individuals

References	Participants	Materials	Principal findings	Strengths and limitations
Nicholas et al. [27]	ASD probands and all their parents (N = 90, M = 65, F = 25 for the first stage of the study; N = 20, M = 14, F = 6 for the second stage of the study)	For screening candidate genes genotyping SNPs	Significant association (P < 0.05) for two single-nucleotide polymorphisms in per1 and two in npas2; in npas2 40 out of the 136 possible two-marker combinations were significant at the P < 0.05 level. The best result was between markers rs1811399 and rs2117714, P = 0.001. In per1 the significant result was for the markers rs2253820–rs885747. Epistatic clock genes may be involved in the etiology of autistic disorder. Problems in sleep, memory and timing are all characteristics of autistic disorder and aspects of sleep, memory and timing are each clock-gene-regulated in other species	Limitations: lack of a control group and the higher number of males compared to females
Hu et al. [5]	Three groups of autistic probands, selected after the exclusion of females, individuals with cognitive impairment, genetic or chromosomal abnormalities, born prematurely and comorbid psychiatric disorder and a control group (non-autistic controls)	DNA microarray analyses	In the most severely affected ASD group, 15 genes, which regulate circadian rhythm, have neurological and metabolic functions deregulated in ASD, were found. From other groups, 20 genes were pointed out, mostly located in non-coding regions and associated with androgen sensitivity	Limitations: the lack of the exact number of participants in each group and information about age and gender of the included people; epigenetic modifications related to inflammatory status Strengths: utility of subdividing individuals with ASD on the basis of cluster analyses of ADIR scores that incorporate all three core domains of ASD (as described in the accompanying manuscript)
Yang et al. [29]	ASD patients (N = 28, 14 of them with sleep problems and 14 without sleep problems. In the first group, M = 5 and F = 9, with an age range 3–28 years, while in the second group M = 12 and F = 2, with an age range 3–19 years) Healthy controls (N = 23)	Sequencing of the coding regions of 18 canonical clock genes and clock-controlled genes; direct sequence analyses verified detected mutations and additional control individuals were screened	Mutations in circadian-relevant genes affecting gene function are more frequent in patients with ASD than in controls. Circadian-relevant genes may be involved in the psychopathology of ASD	Limitations: small sample size Strength: presence of a control group (even if it is not known its internal composition)
Olde-Loohuis et al. [28]	Wistar rats	Rat mPFC collection, RNA isolation, RNA sequencing, gene ontology analysis, cDNA synthesis, qRT-PCR	Three different subsets of genes discovered: the first involved in the regulation of circadian rhythm, the second contributing to extracellular matrix, the third important to understand autism at a molecular level	Limitations: it is a study conducted on animals model, and it is difficult to make precise distinction between groups Strengths: it suggests a possible linking between circadian rhythm and molecular basis of autism

ASD autism spectrum disorder, SNPs single-nucleotide polymorphisms, ADIR autism diagnostic interview-revised, mPFC medial prefrontal cortex, qRT-PCR qReal time-polymerase chain reaction