Disrupted hypothalamic functional connectivity related to cognitive impairment after diffuse axonal injury.

ABSTRACT: This study aims to investigate whether there is imaging evidence of disrupted hypothalamic functional connectivity (FC) in patients with diffuse axonal injury (DAI) and relationships with cognitive impairment.Resting-state functional magnetic resonance imaging (fMRI) data were acquired from acute patients with diagnosed DAI (n = 30) and healthy controls (HC) (n = 30). We first assessed hypothalamic FC with seed-based analysis. Furthermore, the lateral and medial hypothalamic seed was selected to show distinct functional connectivity in DAI. In addition, partial correlation was used to measure the clinical associations with the altered hypothalamic FC in DAI patients.Compared with HC, DAI group showed significantly increased hypothalamic FC with superior temporal gyrus, and the regions around the operculum. Furthermore, there was a significant negative correlation between the connectivity coefficient of hypothalamus to right and left superior temporal gyrus and the disability rating scale scores in DAI group. When the seed regions were divided into lateral and medial hypothalamus, except for increased connectivity of medial hypothalamus (P < .01 with correction), we more observed that decreased left lateral hypothalamic connectivity was positively correlated with mini-mental state examination (MMSE) scores.Our results suggest that there are alterations of hypothalamic FC in DAI and offer further understanding of clinical symptoms including related cognitive impairment.

Introduction

Diffuse axonal injury (DAI) is thought to be caused by shearing forces, but far from a proven disease, impulse and momentum fluid waves and other factors are currently being explored.[ Pathologically, DAI encompasses a spectrum of effects from primary mechanical breaking of the axonal cytoskeleton, to transport interruption, swelling and proteolysis, through secondary physiological changes.[ Axonal degeneration arising from DAI is conventionally recognized as a progression from disruption in axonal transport leading to axonal swelling followed by secondary disconnection and, finally, Wallerian degeneration.[ Morphologically intact axons with disrupted physiology may contribute to the pathological milieu leading to clinical dysfunction across a wide range of injury severities.[ Loss of consciousness used to be associated with DAI, but it is not a prerequisite for DAI.[ The full range of DAI effects are unknown, especially regarding milder injuries.[ Conventional MRI can be helpful in revealing obvious DAI lesions, diffusion-weighted imaging can be used for detecting lesions with vasogenic and cytotoxic edema,[ and susceptibility weighted imaging can be used to detect characteristic microhemorrhages with high sensitivity.[ These studies suggested that the subcortical region, the corpus callosum, the brain stem and the cerebellum were common vulnerable areas in patients with DAI, but the condition is not restricted to the white matter (WM) structures.[

Recently, advanced neuroimaging techniques have been used in DAI patients; for example, voxel-based morphometry analysis for volumetric MR imaging detected marked volume reduction in several WM areas in a patient with DAI,[ and diffusion tensor imaging showed that the abnormal metrics of diffusion tensor imaging may correlate with the timing of head injury, severity biomarkers and long-term prognosis.[ A study of executive control tasks using functional MRI confirmed that augmented functional recruitment may be a neural marker of capacity or efficiency limits in chronic-stage, moderate-to-severe DAI patients.[ These discoveries might better explain the clinical symptoms, injury mechanism, and even the prognosis in patients with DAI. However, these findings cannot fully account for the clinical symptoms, for example, deficits in working memory, emotion disorders, anorexia, obesity, central fever, reproduction and sexual dysfunction, and other cognitional impairments in DAI patients, which are all closely related to hypothalamic dysfunction and neuropeptide dysregulation.[

Increasing numbers of studies have indicated that damage to the hypothalamic cell bodies and hypothalamic microstructure are related to hypothalamic dysfunction and neuropeptide dysregulation in mild traumatic brain injury (mTBI) patients.[ The mTBI patients also showed disrupted hypothalamic resting-state networks based on blood oxygenation level-dependent (BOLD) task-free functional MRI (called resting-state functional MRI, rs-fMRI).[ For example, headache frequency and headache intensity related with hypothalamic functional disconnections in mTBI with post-traumatic headache.[ As a special type of traumatic brain injury, DAI patients may have extensive axonal injury that also involves the hypothalamic functional connectivity (FC) pathways and causes associated cognitive and behavioral symptoms. Increasing numbers of DAI studies have found disrupted regional, long-distance, and even network connectivity in patients with DAI.[

On basis of previous studies, the present study hypothesized that there are FC changes distributed in the hypothalamus for DAI that are associated with the clinical assessment scales. Furthermore, this study aimed to explore the FC alterations in two important subregions, lateral hypothalamus (LH) and medial hypothalamus (MH), because of their functional differences. As a motivation-cognition interface, LH is necessary for reward learning acquisition, memory recall and behavior expression[ while MH is involved in resting and eating disorders.[

Materials and methods

Participants

The DAI patients were recruited from the emergency department of our hospital between January 2018 and May 2020. Thirty right-handed patients with DAI (from 20–60 years) were admitted to this study ultimately by strict inclusion criteria. Inclusion criteria included

a traumatic brain injury with rapid acceleration or deceleration,

immediate loss of consciousness, disorientation, or amnesia,

MRI demonstrating traumatic micro-bleeds or non-hemorrhagic lesion.

In fact, there is no standard for hemorrhage, thus, to minimize bleeding interference with the BOLD signal, we set larger hemorrhage as 10 mL with subjectively. Exclusion criteria included other forms of traumatic brain injury, including herniations, subdural hematomas, or intracerebral hematomas. Thirty age, sex, and education-matched healthy controls (HC) were recruited through local advertisements. All patients in this study provided written informed consent before examination. The study was approved by the local ethics committee of the Taizhou People's Hospital.

Clinical assessment

The main clinical assessments included:

the Glasgow Coma Scale for assessment of conscious impairment;

the Disability Rating Scale (DRS) for measurement of disability;

the Mini-Mental State Examination (MMSE) for rapid assessment of a person's cognitive state and

the Motor Assessment Scale to assess everyday motor function.

MRI acquisition

A 3.0 T MRI scanner (Skyra, Siemens, Enlargen, Germany) with an 8-channel receiver array head coil was used. The resting-state functional MRI and high-resolution T1-weighted data were collected for all participants. The resting-state functional MR data were acquired by gradient echo-planer imaging sequences with the following parameters: repetition time/echo time = 2000/25 ms; slices = 36; thickness = 4 mm; gap = 0 mm; field of view  = 240 mm × 240 mm; acquisition matrix = 64 × 64; and flip angle (FA) = 90°. The fMRI sequence scan took 8 minutes and 8 second. The anatomical T1-weighted MR images were obtained with a T1-weighted 3D spoiled gradient-echo sequence as follows: repetition time/echo time = 1900/2.48 ms; thickness = 1 mm; slices = 170; FA = 8°; field of view = 256 mm × 256 mm; gap = 0 mm; and acquisition matrix = 256 × 256. The structural sequence scan took 5 minutes and 26 second.

MRI preprocessing

Resting-state fMRI data were analyzed using Data Processing & Analysis of Brain Imaging (DPABI, http://www.rfmri.org/dpabi) and Statistical Parametric Mapping (SPM12, http://www.fil.ion.ucl.ac.uk/spm). The first 10 volumes were discarded and the remaining 230 functional images were realigned and adjusted for slice timing. We excluded the images who had a head motion greater than 3.0 mm or a rotation in the x, y, or z directions higher than 3.0°. Frame displacement (FD) values were further computed for each subject to reflect the temporal derivative of the movement parameters.[ Data were spatially normalized to the Montreal Neurological Institute template (resampling voxel size = 3 × 3 × 3 mm3). After normalization, 24 head motion parameters, mean FD, and mean time series of global, WM, cerebrospinal fluid (CSF) signals were included in the regression analysis. Finally, the data was then filtered with a band-pass filter preserving signals between (0.01–0.1 Hz).

Seed-based connectivity and group analyses

According to previous studies,[ the seed region of interest (ROI) of the hypothalamus were at x = 2, y = −1, z = −12 (radius = 2 mm), Moreover, we also select the MH and LH, which were 2 paired spheres 2 mm in radius as seed regions, centered at x = ± 4, y = −2, z = −12 and at x = ± 6, y = −9, z = −10, respectively. The BOLD signals were extracted as averages from each of MH and LH seeds. For each subject, we used seed-based analyses to computed the Pearson correlation coefficient as function connectivity between the averaged BOLD time course of each seed ROI and the rest of the brain voxels. Then, a Fisher z-transform was applied to improve the normality of the correlation coefficients. A one-sample t-test was performed on each of the Z maps for DAI and HC, then a two-sample t-test was used for 2 groups with age, sex, education, and mean FD as covariates. We also performed a two-way ANOVA with age as a covariate on z maps to confirm the group (DAI vs. HC) main effect and to examine the seed (LH vs MH) main effect and group by seed interaction.

Statistical analysis

Demographic factors between the 2 groups and clinical characteristics were evaluated using SPSS 22.0 (SPSS Inc., Chicago, IL). A one-sample t-test was used to explore the spatial distribution of hypothalamic FC patterns. The individual z-values were then entered into a one-sample t-test in DPABI to identify brain regions that showed significant correlations with each seed region for each group to control the possibility of Type I errors. We utilized a permutation test (permuted 1000 times) with Threshold-Free Cluster Enhancement to threshold our results at P < .01 corrected (age, sex, and mean FD as covariates), which is a program of multiple comparison correction integrated into DPABI. Two-sample t-test was performed to identify regions with a significant difference between DAI and HC groups in hypothalamic connectivity; permutation test with Threshold-Free Cluster Enhancement at P < .01 corrected, age, sex, education, and mean FD were regarded as covariates. Partial correlations were calculated as the measure of association between the hypothalamic functional index and clinical symptom severity after correction for sex, age and mean FD.

Results

Demographic characteristics

As shown in Table 1, patients had a relatively short injury duration of averages 10.7 weeks, and displayed moderate to severe disability (DRS scores = 9.07 ± 4.40) and moderate cognitive dysfunction (MMSE scores = 20.63 ± 4.08), as is expected in the result of DAI.

Table 1

Demographic characteristics in DAI patients and HC.

Characteristics	DAI patients (n = 30)	HC (n = 30)	P value
Age (years)	46.30 ± 11.93	48.07 ± 10.55	.546
Sex (Female/Male)	9/15	10/16	.944
Education (years)	12.60 ± 2.91	12.80 ± 3.68	.816
FD values	0.17 ± 0.09	0.21 ± 0.12	.210
Disease duration (weeks)	10.70 ± 3.93	–	–
GCS scores	9.83 ± 2.37	–	–
DRS scores	9.07 ± 4.40	–	–
MMSE scores	20.63 ± 4.08	–	–
MAS scores	33.13 ± 4.87	–	–

Alteration of hypothalamic FC in DAI patients

DAI patients showed significantly increased hypothalamic FC with the superior temporal gyrus (STG), the temporal operculum and the insula (Fig. 1). Furthermore, DAI patients showed increased medial hypothalamic FC with the inferior frontal gyrus, the frontal operculum, the insula and cuneus (Fig. 2). When compared with HC, decreased left lateral hypothalamic FC with the basal ganglia, cingulate cortex, and other structures was found in the patients with DAI (Fig. 3). No significantly differences on the right lateral hypothalamic connectivities were found between the DAI and HC group. The detail of significantly different brain regions between DAI and HC groups are shown in Table 2.

Figure 1

The group differences of the hypothalamic FC pattern between the DAI and HC groups (PT correction with TFCE at P < .01).

Figure 2

The group differences of the left and right medial hypothalamic FC pattern between the DAI and HC groups (PT correction with TFCE at P < .01). MH = medial hypothalamus.

Figure 3

The group differences of the left lateral hypothalamic FC pattern between the DAI and HC groups (PT correction with TFCE at P < .01). LH = lateral hypothalamus.

Table 2

The significantly different brain regions in the hypothalamic ROI between the DAI and HC groups.

ROI	Brain region	BA	Cluster voxels	Peak MNI (X, Y, Z)	Peak T values	Effect size
Hypothalamus
	Temporal operculum/Insula	13,41	46	−36, −21, −6	4.293	1.450
	Superior temporal gyrus	22	22	54, −12, 0	5.898	1.650
	Superior temporal gyrus	22	21	−51, 6, 0	5.063	1.364
	Temporal operculum/Insula	13,41	26	−36, −33, 21	4.352	1.450
Left MH
	Temporal pole: superior gyrus	38	14	42, −6, −15	4.058	1.374
	Temporal operculum/Insula	13,41	83	−36, −24, −3	5.239	1.654
	Inferior frontal gyrus (orbital)	47	27	−42, 21, −6	4.239	1.297
	Superior temporal gyrus	22	23	54, −9, 0	5.273	1.455
Right MH
	Superior temporal gyrus	38	19	51, 6, −12	4.534	1.358
	Frontal operculum/Insula	13	353	−36, −30, −6	4.586	1.663
	Superior temporal gyrus/Rolandic operculum	22,13	144	54, −9, 0	6.137	1.531
	Cuneus	18	33	9, −84, 18	4.561	1.324
	Parietal operculum/insula	13	48	−30, −45, 15	4.372	1.819
Left LH
	Lentiform nucleus, Putamen	–	255	21, 9, 9	−4.956	1.204
	Precuneus/Anterior and middle cingulate/Middle frontal gyrus	7,9,10,24,31,32,33	1598	18, −6, 36	−5.500	1.680
	Lentiform nucleus, pallidum	–	10	−15, 6, −3	−4.558	1.196

In addition, we conducted a two-way group (DAI vs. HC) × seed (LH vs MH) ANOVA with age, sex, and education as covariates to examine the interaction effects. No significant results were found.

Correlation analysis

The hypothalamic FC to left and right STG showed significant negative correlations with DRS scores (r = −0.436, P = .026; r = −0.459, P = .018, respectively) (Fig. 4A–B). In the regions with altered sub-region (LM or MH) connectivity, the DRS scores were negatively associated with left MH connectivity to right STG (r = −0.552, P = .003) while the left LH connectivity to right putamen and left pallidum were positively correlated with MMSE scores (r = 0.417, P = .034; r = 0.420, P = .033, respectively) (Fig. 4C-E).

Figure 4

Partial correlations between the altered hypothalamic FC and clinical variables. The hypothalamic FC to left and right STG were negatively correlated with DRS scores (r = −0.436, P = .026; r = −0.459, P = .018, respectively) (Figure 4A–B). The left MH connectivity to right STG were negatively correlated with DRS scores (r = −0.552, P = .003) (Figure 4C). The left LH connectivity to right putamen and left pallidum were positively correlated with MMSE scores (r = 0.417, P = .034; r = 0.420, P = .033, respectively) (Figure 4D–E). LH = lateral hypothalamus, MH = medial hypothalamus, PAL = pallidum, PUT = putamen, STG = superior temporal gyrus.

Discussions

In this study, we have demonstrated that there is alteration in hypothalamic FC following traumatic axonal injury. This alteration is mainly reflected in the disability rating related to the increased hypothalamic connectivity with the STG and the regions around the operculum. Moreover, when divided into LH and MH, decreased LH connectivity was related to disrupted cognitive state in DAI patients.

Compared to HC, the DAI patients showed increased FC between the hypothalamus and STG, in which the connectivity strength was negatively correlated with the DRS scores. The STG is a site of multisensory integration that is implicated in the processing of visual analysis of body movement information.[ In previous studies, disrupted FC between the hypothalamus and the temporal gyrus was extensively explored in epilepsy, Alzheimer's disease and other neurological diseases.[ In DAI patients, loss of or decline in movement ability is 1 common sequela.[ Interestingly, a recent study showed decreased cerebral glucose metabolism in the right STG and cingulate gyrus in patients with DAI,[ supporting the idea that it is a compensatory contribution to altered function connectivity in a type of regional cerebral metabolism. The temporal operculum/insula, another region with increased FC with the hypothalamus, is involved in communication between internal and external information through emotional subjective awareness,[ especially auditory semiology.[ Recent functional neuroimaging suggested that the insula may also act as the interface between body awareness and movement in Parkinson's disease.[ “Functional plasticity” is hypothesized to change regional brain FC to delay functional decline. Zhou et al found that significantly decreased interhemispheric FC in callosotomy, and restoration with the increased intrahemispheric connectivity occurred 28 days after surgery, demonstrates the functional plasticity in animal model.[ Therefore, in this study, the increased hypothalamic connectivity within several regions around the operculum and STG might indicated a compensative mechanism with functional plasticity after traumatic axonal injury.[

In this study, the decreased LH connectivity was possibly due to direct injury in DAI, and structural alteration after injury may account for the decreased FC. Damage to the cingulum was found in DAI using diffusion tensor tractography.[ In a study of craniopharyngioma with hypothalamic lesions, the strongest association was found between the decline in visual episodic memory and WM alterations in the cingulum, with both loss of WM integrity and demyelination/edema.[ Moreover, decreased amplitudes of low-frequency fluctuations were reported in the cingulate gyrus and were related to working memory and processing speed.[ Furthermore, decreased connectivity in the middle frontal gyrus and superior frontal gyrus were shown in mild traumatic brain injury.[ Moreover, recent work suggests that the lateral hypothalamus may be regarded as a motivation-cognition interface and is associated with cognitive processes, such as associative learning and memory control feeding behavior.[ Our current results confirm that the dysfunction of the cingulum accounts for the cognitive impairment in patients with diffuse axonal injury, it is greatly significant for our understanding of the underlying pathology of DAI related cognitive impairment.

Several limitations in the current study need to be considered. First, since the study is cross-sectional with a relatively small sample size, it is not appropriate to make direct causal inferences regarding the relationships between the brain disconnections and clinical DAI characteristics. Increasing the sample size with our approach would increase our ability to make causal relationships between the disrupted hypothalamic FC and DAI-related cognitive impairment. Thus, further longitudinal MRI studies with large sample data should validate the findings in the future. Second, there were no specific clinical assessments for hypothalamic function, and clinical evaluations of healthy controls were inadequate; this deficiency structure. We selected multiple seeds based on previous studies, as image resolution and inter-subject variations in anatomy pose challenge should be considered. Finally, the hypothalamus is implicated in a variety of complicated physiological functions; therefore, the meaning of altered hypothalamic FC after diffuse axonal injury still requires to be explored further.

Conclusions

DAI patients showed that disability rating was associated with increased hypothalamic FC with the STG and the regions around the operculum, while decreased LH connectivity was related to the cognitive state of DAI patients. Our findings may provide further understanding of clinical symptoms including DAI-related cognitive dysfunction.

Author contributions

Conceptualization: Jian Zhang, Shaohua Ding.

Data curation: Jian Zhang.

Formal analysis: Jian Zhang, Yi Chang.

Investigation: Jian Zhang, Yi Chang.

Methodology: Jian Zhang, Yi Chang.

Software: Jian Zhang, Yi Chang.

Supervision: Shaohua Ding.

Validation: Shaohua Ding.

Writing – original draft: Jian Zhang.

Writing – review & editing: Shaohua Ding.