| Objective: The serotonin system interact with the circadian rhythm regulation.In specific brain regions,the expression of some key genes of the serotonin system(such as Tryptophan hydroxylase-2,TPH2)and the concentration of serotonin fluctuated periodically for about 24 hours.The serotonin system may be involved in the regulation of circadian rhythm by affecting the synchronization function of the suprachiasmatic nucleus(SCN).To explore the genetic and brain function alterations in MDD patients with circadian rhythm disturbance,we investigate the genetic susceptibility and resting-state functional MRI(rs-f MRI)brain characteristics of MDD patients with early-wakening symptom from the perspective of circadian rhythm disturbance.Method: A total of 249 subjects who met the inclusion criteria from The Affiliated Brain Hospital of Nanjing Medical University were enrolled,including 185 patients with depression and 64 healthy controls.Patients were grouped according to the Hamilton depression scale 6th item scores from 24-item Hamilton Rating Scale for Depression(HAMD-24): 1 or 2 for waking early in the morning(WE group,n=123)and 0 for not waking early in the morning(NWE group,n=62).We also recruited 64 healthy controls(HC group).Venous blood and resting state functional magnetic data were collected,and the data processing and analysis methods were as following:1 The relationship between gene polymorphism and MDD patients with sleep related circadian rhythm disturbanceThe next-generation sequencing technology was performed to genotyping the TPH2 gene polymorphism.Based on the clinical group,logistic regression model was employed for genetic data analysis and adjusted for age,gender,education,marriage condition and family history using the PLINK software,which aim to invested a significant allele associated to MDD patients with sleep related circadian rhythm disturbance.All the results were corrected by Bonferroni multiple correction2 The effect of rs4290270 polymorphism of TPH2 gene on rs-f MRI characteristics in MDD patients2.1 The functional connectivity difference between patients who carry or not carry the risky alleleAfter processed with head motion correction,co-registration,segmentation,regression the head motion and spatial normalization etc.,the functional connectivity map between left and right SCN with other voxels in the whole brain was calculated in the Matlab software.All the FC map were divided into two groups based on the allele genotype which obtained from the previous result.To obtain the significantly FC values,the independent t test was performed and the Alphasim test for multiple corrections.2.2 The dynamic community features difference between patients who carry or not carry the risky alleleSliding time window algorithm was used to slide the preprocessed time series to obtain the overlapping sub-time series matrix.The LASSO model was used to sparse the matrix and then the dynamic functional connection matrix is constructed.Dynamic modularization algorithm was performed to divide the functional connectivity of each window into modules,which can be divided into the same module or different modules according to the tightness of the connection between nodes.According to the divided module matrix,the module conversion frequency of a certain brain region was defined as flexibility value,and the probability of assigning certain brain regions to the same module was the MA(module allegiance)value of these brain regions.The information of subject groups was same with which in the 2.1,the permutation test was performed in the Matlab software to compare the flexibility 16 nodes of the visual network and MA values of the visual network,and p < 0.05 was considered as significant.The Bonferroni method was performed for multiple comparison corrections.3 Mediating effect of the rs-f MRI characteristics in the relationship between the rs4290270 polymorphism and the clinical features of MDD patientsTo investigate the potential relationship among rs4290270 polymorphism,rs-f MRI characteristics and clinical features of MDD patients,we performed the mediation analysis in the R software by using mediate package.The genotype of rs4290270 was treated as the independent variable(X);the abnormal rs-f MRI characteristics such as FC 、flexibility and MA values which representing endophenotype was treated as intermediary variable(M)respectively;and the clinical features which obtained from HAMD-24 was treated as dependent variable(Y)respectively.Although there was no significant difference in some demographic information,they were still included in the covariate treatment considering the potential contribution to the pathogenesis of MDD.Results: 1 The relationship between gene polymorphism and MDD patients with sleep related circadian rhythm disturbanceAll selected SNPs distributions were in Hardy-Weinberg equilibrium,none SNPs had a call rate of > 10% and MAF < 0.05.The allele frequency for each group which were calculated in the logistic regression model was > 0.05.The correlations between MDD and HC,NWE and HC,WE and HC groups were then assessed.There is one significant result between the MDD and HC group(rs4290270;p = 0.016),however,it failed in the Bonferroni correction.The rs4290270 showed a significant association in the WE group(p = 0.0047;OR = 2.03);associating allele: T)but not in the NWE group(p = 0.1750;OR = 1.49).2 The effect of rs4290270 polymorphism of TPH2 gene on rs-f MRI characteristics in MDD patients 2.1 The functional connectivity difference between patients who carry or not carry the risky alleleA total of 158 subjects satisfied the quality control criteria,58 data for A allele group and 100 data for T allele group.Compared with the A allele group,increased functional connectivities of the right SCN with the right fusiform gyrus(34,-4,-44)and the right middle frontal gyrus(50,46,4)were observed in the T allele group.Additionally,there was a decrease in functional connectivities of the left SCN with the right lingual gyrus(12,-60,6)and the left calcarine sulcus in the T allele group(-26,-58,6).All results were corrected by Alpha Sim correction.2.2 The dynamic community features difference between patients who carry or not carry the risky alleleThe information of subjects groups were same with which in 2.1.Compared with the A allele group,the T allele group showed increased flexibility value of the right fusiform gyrus(30,-42,-15;p = 0.0029)which corrected by Bonferroni correction.In addition,the MA values in visual network also showed significant difference(p = 0.031)between A and T allele groups.3 Mediating effect of the rs-f MRI characteristics in the relationship between the rs4290270 polymorphism and the clinical features of MDD patients The mediation analysis showed that the coefficient of total effect was established according to the masking effect(c=-0.0738[-0.8947,0.6900](p = 0.8550).The coefficients a(p = 0.0008)and b(p = 0.0154)were both significant in the model.The result also showed a significant indirect effect of coefficient ab(ab=-0.2654,p = 0.0096),95% confidence interval [-0.5559,-0.05].The coefficient of direct effect was not significant(c ’=0.1916,p = 0.6427).To sum up,the flexibility value of the right fusiform gyrus mediate the relationship between the rs4290270 polymorphism and the cognitive factor score of HAMD-24.Conclusion: The TPH2 gene polymorphism plays a role in the sleep related circadian rhythm disturbance in MDD.In patients who carry the risk alleles,there was an abnormal FC values between SCN and the cortex around the calcarine sulcus,lingual gyrus,fusiform gyrus and middle frontal gyrus,which may reflect the abnormality of the accepting,processing and transmission of external information.Further study found that the right fusiform gyrus also showed abnormality in the frequency of modularization state transition and the abnormality dynamic community features of the visual network.It may reflect that the impaired function of the involved brain regions in processing complex information dynamic transformation and ultimately contribute to the cognitive impairment.However,there may exist more complex compensatory mechanisms for the cognitive function regulation in the higher cortex such as dorsolateral prefrontal cortex.These abnormal brain imaging features are underlying neuropathological basis for the sleep related circadian rhythm disturbance in MDD. |
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