Neutrophil to lymphocyte ratio is a transdiagnostic biomarker of depression and structural and functional brain alterations in older adults

Abstract

The neutrophil to lymphocyte ratio (N:L) is an emergent transdiagnostic biomarker shown to predict peripheral inflammation as well as neuropsychiatric impairment. The afferent signaling of inflammation to the central nervous system has been implicated in the pathophysiology of sickness behavior and depression. Here, the N:L was compared to structural and functional limbic alterations found concomitant with depression within a geriatric cohort. Venous blood was collected for a complete blood count, and magnetic resonance imaging as well as phenotypic data were collected from the 66 community-dwelling older adults (aged 65–86 years). The N:L was regressed on gray matter volume and resting-state functional connectivity (rsFC) of the subgenual anterior cingulate (sgACC). Thresholded parameter estimates were extracted from structural and functional brain scans and bivariate associations tested with scores on the geriatric depression scale. Greater N:L predicted lower volume of hypothalamus and rsFC of sgACC with ventromedial prefrontal cortex. Both parameters were correlated (p < 0.05) with greater symptomology in those reporting moderate to severe levels of depression. These findings support the N:L as a transdiagnostic biomarker of limbic alteration underpinning mood disturbance in non-treated older adults.

1. Introduction

The chronicity of low-grade inflammation has of recent garnered significant attention in the etiology of depression, particularly amongst older adults (Krishnadas and Cavanagh, 2012). Concomitant with immunosenescent processes and age-related multi-morbidity, persons aged over 65 years are susceptible to chronically elevated levels of peripheral inflammation (Orimo et al., 2006). Elevated levels of pro-inflammatory cytokines may subjugate the brain altering dopaminergic and glutamatergic neurotransmission within the neural circuits underpinning mood disturbance (Miller and Raison, 2016). As the scope of pro-inflammatory biomarker involvement in the manifestation of depression broadens it becomes important to investigate upstream changes in adaptive and innate immune function from which these age-related inflammatory-immune mechanisms progress.

The relative state of peripheral neutrophilia to lymphocytopenia has emerged as a potential transdiagnostic biomarker of neuropsychiatric disturbance. The largest pool of innate immune cells, i.e., neutrophils, are driven by cortisol levels and contribute to nonspecific inflammation via phagocytosis, chemotaxis, release of reactive oxygen species, degranulation, and the production and liberation of cytokines which affect BBB function (Aubé et al., 2014). Although neutrophil recruitment is key for host defense, the overexuberance of these cells brought on by trauma or physiologic stress appears to be amplified in older adults and lead to a paradoxical increase in systemic damage to bodily tissues(Liew and Kubes, 2019). In juxtaposition, through the anti-inflammatory effects of regulatory T-cells on the activated state of microglia and the tryptophan-metabolizing enzyme indoleamine-2,3-dioxygenase enzyme peripheral has been implicated in mitigation of depressive symptoms (Laumet et al., 2018). Moreover, the age-related changes in mitochondrial stress and telomerase activity within peripheral lymphocytes resulting in leukocytosis and lymphocytopenia are further implicated in age-related mood disturbance (Lindqvist et al., 2015). Hence, driven by disease and immunosenescent processes in addition to elevated presence of endogenous neuroendocrine response to acute physiologic stress, the neutrophil to lymphocyte ratio (NLR) has emerged as a highly reproducible index of the severity of stress and systemic inflammation amongst critically ill patients following trauma, major surgery or sepsis (Zahorec, 2001; Rainer et al., 1999). This relatively low-cost marker of neutrophil expansion is also used to indicate life-threatening inflammation and organ failure in older adults (Demir and YÜCEL M., 2020; Song et al., 2020). Yet, amongst older adults, the prognostic capacity for the NLR in neuropsychiatric and neurodegenerative disease remains unclear.

Afferent signaling of an inflammatory peripheral state is a central tenet of the geriatric manifestation of depression (Alexopoulos and Morimoto, 2011). Meta-analyses on the correlates of depression implicate chronically and experimentally elevated levels of the acute-phase reactant C-reactive protein (CRP), Interleukins (IL)-6, IL-1, and tumour necrosis factor alpha (TNF-α) in the severity of mood disturbance (Valkanova et al., 2013; Haapakoski et al., 2015; Howren et al., 2009). Levels of CRP are most germane to geriatric populations as they are found elevated amongst depresses older adults living with(Lu et al., 2013; Viscogliosi et al., 2013) or without (van den Biggelaar et al., 2007) chronic disease. Neutrophilia in combination with lymphocytopenia are closely intertwined with systemic inflammation in older adult cohorts. Most recently, literature has emerged correlating NLR with depression severity in middle- to older-aged adults diagnosed with major depressive disorder (MDD) (Sunbul et al., 2016; Kayhan et al., 2017; Arabska et al., 2018). Furthermore, NLR is higher in persons experiencing manic bipolar disorder episodes comparison with depressive bipolar and MDD suggesting the increased likelihood for severe neuropsychiatric disease with increasing NLR (Mazza et al., 2019). Although compelling, it remains unclear whether a neutrophilic environment predicts greater severity of depressive symptomology amongst individuals not currently treated for MDD and what are the neural regions implicated in this process.

In recent years, attention has turned to elucidating the role of peripheral inflammatory-immune processes in the structural integrity and functional connectivity of limbic regions implicated in depression (Alexopoulos and Morimoto, 2011). In a meta-analysis of functional Magnetic Resonance Imaging (fMRI) and positron emission tomography studies, peripheral markers of inflammation correlated with greater co-activation throughout limbic regions of interest (ROI) including the ventromedial prefrontal cortex (vmPFC), anterior cingulate cortex (ACC), temporal cortices, amygdala, hippocampus, hypothalamus, striatum, insula as well as other subcortical regions extending into the midbrain and brainstem (Kraynak et al., 2018). Notably, the subgenual region of the ACC (sgACC) was amongst the ROIs surviving the more stringent statistical correction. In the context of depression and sickness behavior, the sgACC is primarily implicated in behavioral withdrawal, resource conservation, and the promotion of safety behaviors (Marsland et al., 2017). This neurobehavioral adaptation of what may be collectively described as sickness behaviors further involves interactions between neuroendocrine and inflammatory-cytokine signaling response (Critchley et al., 2003). Not only do acute inflammatory changes predict enhanced sgACC activity during limbic processing, but increased connectivity of the sgACC with the amygdala, vmPFC, and nucleus accumbens (Harrison et al., 2009a). In addition to these experimentally induced changes in limbic activity neuroimaging the functional connectivity of these regions at rest has also garnered interest. Unlike task-related blood oxygen level dependent (BOLD) activity, resting state functional connectivity (rsFC) reflects the intrinsic energy demands of neuronal assemblies of brain regions involved in a common behavior (Lewis et al., 2009). Meta-analytic findings also implicate increased limbic connectivity with the sgACC in the presentation of depressive rumination (Hamilton et al., 2015). Although brain structure does not always implicate function deficits volumetric reductions of the sgACC are reported in depressed individuals (Kaltenboeck and Harmer, 2018; Schmaal et al., 2017; Campbell et al., 2004). Furthermore, in a large sample of non-demented older adults, serum levels of TNF-α and IL-1β were associated with smaller volume in the sgACC and inferior parietal lobule after controlling for age-related factors including APOE ε4 status, cardiovascular risk and geriatric depression (Corlier et al., 2018). Although inflammatory signaling in the periphery may contribute to the structural and functional alterations implicated in mood disturbance it remains unclear whether the upstream markers of immune dysfunction, such as the NLR, can be related to the neurobehavioral processes involved in the manifestation of geriatric depression.

The NLR has emerged as a potential transdiagnostic marker for psychiatric disease (Brinn and Stone, 2020). The sgACC is a region of interest implicated in the inflammatory signaling of mood disturbance, however, it is unclear whether the neural integrity and connectivity of this and other limbic regions relates to the NLR in older adults with subclinical depressive symptoms is unclear. Hence, the current study aimed to determine whether the neural correlates of the NLR, indexed by whole-brain volume and rsFC of the sgACC, predict the severity of geriatric depression in a cohort of older adults not diagnosed or treated for MDD.

2. Methods

2.1. Participants

The fMRI and physiological data were acquired from the Enhanced NKI Rockland Sample (Nooner et al., 2012a). Data collection for this study was approved by the Nathan Kline Institute and Montclair State University institutional review boards and all participants provided informed consent. Data collection involved a semi-structured diagnostic psychiatric interview, battery of psychiatric, cognitive, and behavioral assessments as well as a multimodal brain imaging session (Nooner et al., 2012a). The initial sample of individuals with structural and functional brain scans was n = 801. Individuals with a history of treatment or diagnosis for schizophrenia, Alzheimer’s disease, dementia, Parkinson’s Disease, Huntington’s Disease, Multiple Sclerosis, and loss of consciousness were excluded from the study. Seventy-eight older adults, (age of 65 years or older), provided a blood sample NLR assay. Of these, 66 individuals passed quality control for resting state fMRI. Of those individuals four did not have quantifiable T1-weighted scans.

2.2. Measures

2.2.1. Self-report

The Geriatric Depression Scale (GDS) is a 30-item “yes/no” self-report measure of depressive symptoms designed to distinguish depressed elders with neurovegetative symptoms from nondepressed elders with symptoms that overlap with depression yet are concomitant with normal aging or comorbid medical disorder (Yesavage et al., 1982). During administration, the individual is asked to report whether they have experienced several symptoms over the past week. The GDS has a maximum score of 30, with higher scores representing more severe symptoms. The GDS emphasizes cognitive symptoms of depression rather than somatic symptoms. The instrument shows good validity amongst healthy, medically ill, or mild to moderately cognitively impaired older adults(Jung et al., 1997).

2.2.2. Biomarker

The blood sample was obtained on day 1 and the R-fMRI scans on day 2 (~1–2 weeks apart) (Nooner et al., 2012a). A 5 ml venous blood sample was collected at study entry during the first visit and tested between 2 and 6 h for complete blood count including absolute number and percentage of neutrophils, lymphocytes, and monocytes. CBCs are performed in house on a Beckman Coulter LH780 using samples collected in K3EDTA. C-Reactive protein was sent out for reference at Bioreference Laboratories^™, using serum samples and would have to meet Bioreferences specimen requirements.

2.2.3. Neuroimaging

Acquisition.

A 3.0-T Siemens MAGNETOM Trio-Tim scanner was used to collect resting state scans with using the following imaging parameters: TR = 1400 ms, TE = 30 ms, slice thickness = 3.0 mm, flip angle = 65°, field of view = 224 mm, slices = 64, and voxel size = 2.0 × 2.0 × 2.0 mm. The acquisition time was 10-min for each participant using a multi-band imaging sequence. The subjects were instructed to lay still inside the scanner with their eyes open thinking of nothing and not to fall asleep. High-resolution anatomical images (MPRAGE) were acquired using the following scanning parameters: TR = 1900 ms, TE = 2.52 ms, slice thickness = 1.0 mm, flip angle = 9°, field of view = 256 mm, and voxel size = 1.0 × 1.0 × 1.0 mm. All fMRI data used in the analysis are part of the NKI Enhanced dataset made publicly available by the international neuroimaging data sharing initiative (Nooner et al., 2012b).

Structural Image Preprocessing.

Voxel Based Morphometry (VBM) (Ashburner, 2010) analysis was conducted using the following steps implemented in SPM12 software: 1) origin manually set to the anterior commissure; (2) images reoriented to fit a 2 mm Montreal Neurological Institute (MNI) template; (3) images segmented into gray matter, white matter, and cerebrospinal fluid-based probability maps using DARTEL templates; (4) deformations estimated; (5) Jacobian modulation performed to avoid volumetric changes during the normalization to MNI space; and (6) Jacobian-scaled warped gray matter tissue images were produced for statistical analysis.

fMRI pre-processing.

Resting state scans were preprocessed using DPARSF-A in DPABI (http://rfmri.org/DPARSF) (Chao-Gan and Yu-Feng, 2010; Yan et al., 2016). The pipeline was implemented in MATLAB R2017a (MathWorks, Natick, MA, USA) using the following steps: 1) first five images were removed; 2) nuisance covariates (Friston 24 motion parameters, white matter, & cerebrospinal fluid) and linear trends were regressed out; 3) band-pass filtering at 0.01 to 0.1 Hz (Damoiseaux et al., 2006); 4) data were despiked through AFNI 3dDespike, realigned and normalized with DPARSF-A, and smoothed to 6 mm with AFNI 3dBlur; and 5) independent component analysis (ICA-FIX) was applied through FSL MELODIC to identify signal and noise components, which were extracted and transformed into 3 mm MNI space (i.e., subject space). Frramewise displacement (FD) was calculated (Power et al., 2012) and a total of 12 individuals were removed from the analysis due to excessive head motion (FD > 0.50).

Seed-to-whole-brain connectivity analysis.

Whole-brain rsFC was examined using a 6-mm spherical seed centered on the sgACC (MNI: x = 1, y = 22, z = −10). This region was defined from coordinates demonstrating maximal increase in response to administration of an endotoxin (Harrison et al., 2009a). Individual preprocessed data were used to extract fMRI time series data from the sgACC seed in the filtered data, and then Pearson’s correlation coefficients were calculated between the sgACC time series and the time series from all other voxels in the brain. The correlation coefficient at each voxel was transformed to a z-value using Fisher’s r-to-z transformation to enhance normality. The resultant sgACC functional connectivity map for each participant was entered into the group level analysis.

To identify significant associations between the inflammation index and functional connectivity of the sgACC, a voxel-wise multiple regression analysis was conducted using the group-level sgACC connectivity maps as the dependent variable, the inflammation index as the independent variable, gender, and age as covariates of no interest. Mean framewise displacement (FD), averaged across the entire time series, was added as a nuisance covariate to minimize the impact of head motion-related variance on group inference (Satterthwaite et al., 2013). The relationship between inflammation index and sgACC functional connectivity was tested using t-statistics and reported as a z-score after the t-value was transformed into the standard normal distribution. Clusters were considered statistically significant if they reached the extent threshold of p <0.001, uncorrected with a with an FWE-corrected cluster-forming threshold of p < 0.001.

Extraction of regions of interest (ROI).

The contrast parameter estimates were extracted from the seed-based connectivity or VBM regression analyses for voxels defining clusters within the ROIs that exhibited significant effects for high or low levels of inflammation at k ≥ 25 using the program MarsBaR (http://marsbar.sourceforge.net). Extracted parameter estimates were averaged within each ROI and imported into SPSS before being submitted to a partial correlation with GDS scores controlling for head motion and/or age.

3. Results

Sociodemographic and cardiometabolic data is provided in Table 1. The bivariate correlation between GDS scores and NLR was not significant (r = 0.158, p = 0.10). However, compared with individuals with an elevated NLR (>3) (mean = 5.53 ± 5.55) those with lower neutrophil ratios (mean = 2.86 ± 2.54) reported significantly lower levels of geriatric depression (t(64) = −2.65, p = 0.01).

Table 1.

Sample characteristics.

Characteristics	n = 66
Sociodemographic
Age	71.9 ± 5.6
Gender (Female %))	57.6
Ethnicity (%)
Caucassian	90.9
African-American	6.1
Hispanic	3.0
Primary language (%)
English	92.8
Socioeconomic Status	52.7 ± 8.1
Parents’ Socioeconomic Status	34.3 ± 14.3
Full Scale Sum T-score	217.7 ± 29.6
Medical History (%)
Cancer	25.8
Myocardial Infarction	3.0
Coronary Artery Disease	4.5
Hyperlipidemia	43.3
Hypertension	40.9
Irritable Bowel Syndrome	14.4
Crohn’s Disease	1.0
Ulcerative Colitis	2.1
Hepatitis	1.5
Type II Diabetes Mellitus	10.6
Arthritis	45.5
Psychiatric History (%)
Major Depression	4.5
Bipolar Disorder	1.0
Social Anxiety	4.5
Delusional Disorder	1.1
Specific Phobia	2.2
Posttraumatic Stress Disorder	2.2
Bereavement	1.1
Substance Abuse
Alcohol Abuse	4.3
Alcohol Dependence	2.2
Biomarkers
Body mass index	27.0 ± 5.1
Systolic blood pressure	128.6 ± 12.3
Diastolic blood pressure	75.8 ± 8.0
Neutrophil %	60.7
Lymphocyte %	27.9
Monocyte %	7.9
Hemoglobin A1c1	0.4 ± 1.4

To identify gray matter areas that were associated with a high or low peripheral inflammation index, voxel-wise multiple regression analysis was tested in SPM12 controlling for the effects of age gender, and total gray matter volume. The results indicate that three clusters surviving an uncorrected primary threshold of p < 0.001 and a secondary cluster-level FDR-corrected threshold of p < 0.01 showed greater gray matter volume that corresponded with a lower index of peripheral inflammation (see Fig. 1). These clusters included the hypothalamus (k = 137, T = 4.68, FWE < 0.001, FDR < 0.001), the left mid-posterior insula (k = 82, T = 4.45, FWE = 0.048, FDR < 0.01), and the left parahippocampal gyrus (k = 108, T = 4.29, FWE = 0.014, FDR < 0.001). The trending correlation for parameter estimates extracted from the hypothalamus with the GDS scores emerged for the entire cohort (r = −0.181, p = 0.073, n = 66), however, upon restricting the sample to those with moderate to severe depressive symptomology, a significant negative correlation emerged (r = −0.452, p = 0.023, n = 20).

Fig. 1.

Three volumetric regions in the hypothalamus (MNI x = −6, y = −2, z = −12), left mid-posterior insula (x = −36, y = −8, z = −6) and the left parahippocampal gyrus (x = −24, y = − 8, z = −36) corresponding to lower neutrophil-to-lymphocyte ratios (uncorrected primary p < 0.001 and secondary cluster-level FDR p < 0.01).

Based on the hypothesis put forth regarding the sgACC as an anterior cortical structure involved in the interoceptive signaling of inflammation, this structure was used as the seed ROI for a whole brain functional connectivity analysis with the inflammatory index as a covariate of interest. The results showed one large cluster emerging from an area in the left ventromedial prefrontal cortex (x = −30, y = −57, z = −3), (Brodmann Area 10), wherein increased rsFC with the sgACC was associated with lower NLR (k = 132, T = 4.79, FWE < 0.001, FDR < 0.01) (see Fig. 2). Parameter estimates extracted from left BA10 did not correlate with GDS scores when controlling for age and head-motion for the entire cohort (r = −0.137, p > 0.05, n = 66). However, upon restricting the sample to individuals with moderate to severe depression, parameter estimates from BA10 negatively predicted GDS scores (r = −0.383, p < 0.05, n = 20).

Fig. 2.

Greater resting state functional connectivity of the subgenual cingulate with an area in the left ventromedial prefrontal cortex (BA10) (x = −30, y = −57, z = −3), that corresponds with lower ratio of neutrophils to lymphocytes (k = 132, T = 4.79, FWE < 0.001, FDR < 0.01).

4. Discussion

The aim of the current study was to assess the relationship of the NLR, an emergent transdiagnostic biomarker of neuropsychiatric dysfunction, to geriatric depression and its structural and functional neural underpinnings. In this sample a NLR ratio of more than 3:1 was associated with greater endorsement of depressive symptomology on a scale validated in geriatric populations. Previous studies examining systemic markers of critical illness commonly implicate inflammatory biomarkers and acute-phase reactants in mood disturbance amongst older adults (Kushner, 2001; Michaud et al., 2013; Puzianowska-Kuźnicka et al., 2016; Smith et al., 2018). The sgACC was selected as the a priori region of interest because its activity reflects acute changes in peripheral levels of inflammation (Harrison et al., 2009b). This region exhibited greater rsFC with the left vmPFC in conjunction with a lower NLR. This sgACC connectivity to the vmPFC was inversely associated with severity of geriatric depressive symptoms, albeit amongst those endorsing moderate to severe symptoms. In the full sample, greater NLR was associated with volumetric deficits in the hypothalamus, left mid-posterior insula, and parahippocampal cortex. Parameter estimates of hypothalamus volume were inversely associated with moderate to severe depression symptoms with a similar trend found across the entire spectrum of symptom severity.

The correspondence between neutrophilia and hypothalamus atrophy in relation to depressive symptomology is a novel finding amongst older adults. In addition to breakdown of the blood brain barrier the accumulation of neutrophils in the brain parenchyma can perpetuate local microglial activation yielding neurogenerative effects (Cernackova et al., 2020; Zhang et al., 2016). Hypothalamic inflammation is commonly observed in conjunction with atrophy in this region. Chronic interoceptive signaling of an inflammatory peripheral state has been linked to subsequent reductions in insula volume over time (Craig and Craig, 2009; Nusslock and Miller, 2016; Jiang et al., 2015). Specifically, induction of systemic inflammation following endotoxin administration is shown to induce abnormal glucose metabolism in both anterior and posterior insula (Lekander et al., 2016). Meta-analytic findings highlight a strong association between markers of systemic inflammation and resting BOLD activity in left posterior and right anterior insula (Kraynak et al., 2018). Although the causal effect for systemic inflammation on volumetric brain reduction has been mainly inferred amongst older adults (Taki et al., 2013), some mechanistic evidence has been levied. A longitudinal study genotyping sequences for IL-1β C-511 T and CRP polymorphisms measured 2 years apart found these pro-inflammatory biomarkers accurately predicted shrinkage within the entorhinal and ambient gyri of the parahippocampal cortex (Persson et al., 2014; Ladenvall et al., 2006; Reitz et al., 2007; Meisenzahl et al., 2001). Notably, the hypothalamus was the only correlate of greater NLR and geriatric depressive symptom severity. The hypothalamus has long been implicated in major depression by virtue of its role in the adaptive control of energy homeostasis (Cernackova et al., 2020; Pimentel et al., 2014; Valdearcos et al., 2015; Mravec et al., 2019). For example, obesity-mediated inflammation has not only been linked to atrophy of hypothalamic nuclei (Cazettes et al., 2011; Puig et al., 2015), but also M1-activation, HPA-axis reactivity, and total mood disturbance in older adults (Hryhorczuk et al., 2013; Soczynska et al., 2011; Schachter et al., 2018; Martinac et al., 2014). Thus, the pattern of volumetric deficits observed in relation to the NLR supports this measure as a transdiagnostic marker of the limbic atrophy concomitant with depressive disorder.

Coinciding with extant literature implicating inflammation with changes in mood and coactivation patterns with the sgACC (Kraynak et al., 2018; Harrison et al., 2009a; O’Connor et al., 2009), lower rsFC was observed between sgACC and vmPFC amongst individuals with greater NLR. Despite a growing number of studies implicating rsFC between the vmPFC and sgACC in inflammation-based mood disturbance the precise neurobiological mechanism remains unclear (Beckmann et al., 2009; Torta and Cauda, 2011; Yu et al., 2011). Neuroanatomical tracing studies show the murine homologue of the vmPFC is laden with cytokine receptors that mediate activity with an extensive network of subcortical regions including hypothalamus as well as parabrachial and solitary nuclei of the brainstem (Vertes, 2004; Gabbott et al., 2005). Furthermore, brainstem and hypothalamus expression of cytokines, chemokines, and pro-inflammatory transcription factors are observed following systemic inflammation induced by changes in diet(Dalvi et al., 2017) or stress (Sirivelu et al., 2012; Kanemitsu, 2000). Indeed, the highest density of IL-1 receptors are found in the preoptic, supraoptic, and paraventricular areas of the hypothalamus (Konsman et al., 2004). It should be noted that IL-1 and CRP share common gene variation (Eklund et al., 2003; Berger et al., 2002), proliferate in tandem (Pue et al., 1996), and are both linked to major depression (Howren et al., 2009). Intriguingly, thrombocytosis, in association with elevated production of chemotactic proteins for neutrophil proliferation is shown to coincide with elevated levels of TNF-α and the IL-1 family (Strieter et al., 1989; Clancy et al., 2017). Although altered interaction of activated microglia with neurons and axons are an established downstream mechanism of inflammatory signaling to the brain (Fujita and Yamashita, 2021), research has recently emerged implicating neutrophils in cerebrovascular function. In a study examining a mouse model for Alzheimer’s disease found that an the administration of a neutrophil-specific signaling marker that increases migration of neutrophils toward sites of inflammation by modulation of β2-integrin-dependent adhesion resulted in the stalling of blood flow in cerebral capillary segments (Hernandez et al., 2019). According to the neurovascular coupling hypothesis, increased cerebral blood flow may be coupled with higher degree of functional brain connectivity (Zhu et al., 2017). Nevertheless, if a neutrophilic environment is conducive to neuroinflammatory signaling and cerebrovascular dysfunction, then the volumetric reductions of the hypothalamus in conjunction with mitigated vmPFC connectivity with sgACC may reflect independent pathways for inflammatory-immune dysfunction to subjugate the limbic regions that signal sickness behavior (Jin et al., 2016; Dantzer, 2009; Hennessy et al., 2017).

5. Limitations

Amongst the inherent limitations of this cross-sectional design is the inference of causality of NLR on volumetric atrophy and aberrant functional connectivity. Nor can we determine, with any degree of specificity, the chronicity of neutrophilia or inflamation in the current sample. However, supplementary analyses suggest individuals with NLR ratio greater than 3:1 were more likely to have detectable levels of CRP. Furthermore, literature does support neutrophilia is concomitant with immunosenescence and propensity for chronic inflammation amongst older adults (Uhl et al., 2016; Ward et al., 2011; Drew et al., 2018). As a function of hypothalamic atrophy and vmPFC dysconnectivity this study supports NLR as a transdiagnostic index of inflammatory-immune and mood disruption (Kushner, 2001; Michaud et al., 2013; Puzianowska-Kuźnicka et al., 2016). However, other mechanisms involving HPA-axis disruption have been implicated in the etiology of depression (Cernackova et al., 2020). Given the specificity of our volumetric findings and the role this structure plays in mediating glucocorticoid and inflammatory receptor signaling collection of basal cortisol measures would have helped to further support and elucidate the biobehavioral mechanism for hypothalamic inflammation in the geriatric presentation of depressive symptoms. Future studies should include a protocol for cortisol measurement, particularly given that cortisol levels sampled from awakening to pre-scan predicts aberrant rsFC of the medial frontal gyrus with limbic and non-limbic regions (Veer et al., 2012; Wu et al., 2015; Wang et al., 2018). Clinical implications for our findings are limited by the self-reported measure of depression. Although older adults are at greater risk for major depression they are less likely to endorse depressive symptomology (Gallo et al., 1994). Diagnostic interview may provide a more reliable index of depression, however, compared to other frequently used self-report indices the GDS demonstrates superior sensitivity in detecting depression in adults over the age of 65 (Lyness et al., 1997).

6. Clinical implications

Neural tracing studies show dense connection between sgACC and VMPFC lead a large field cortical network that actively responds to metabolic demands (Joyce and Barbas, 2018). Although the current study did not implicate hypothalamus connectivity with the sgACC in the presentation of the NLR or depression, activation of these subcortical nuclei may trigger an extended network that regulates internal homeostasis (Chiba et al., 2001; Freedman et al., 2000). Our neurobiological findings relating to the hypothalamus supports a mechanism whereby inflammation predicts mood disturbance amongst individuals exposed to chronic metabolic or inflammatory-immune dysfunction (Cernackova et al., 2020). It should be noted that selective serotonin-reuptake inhibitors show an inverse effect on inflammation, suggesting bidirectional relationships might be at play (Gałecki et al., 2018; Adzic et al., 2018). Moreover, preliminary yet compelling evidence suggests cumulative exposure to stress and major depression can induce reductions in leukocyte telomere length and activity (Kinser and Lyon, 2013). Concomitantly, waning NLR is observed in MDD following 3 months of selective serotonin-reuptake inhibitor treatment. Hence, future longitudinal studies should elucidate the bidirectional effects of im munosenescent processes and the neuropathophysiology processes underpinning the etiology of geriatric manifestations of depression, particularly in persons living with chronic disease.