Differences in sex and genetic status affect the disruption of NMDAR-related amino acid homeostasis in Parkinson's disease

Differences in sex and genetic status affect the disruption of NMDAR-related amino acid homeostasis in Parkinson’s disease | medRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-P4HH5NV'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search Differences in sex and genetic status affect the disruption of NMDAR-related amino acid homeostasis in Parkinson’s disease Isar Yahyavi , View ORCID Profile Federica Carrillo , View ORCID Profile Tommaso Nuzzo , Anna Di Maio , View ORCID Profile Sara Pietracupa , View ORCID Profile Nicola Modugno , View ORCID Profile Francesco Errico , View ORCID Profile Teresa Esposito , View ORCID Profile Alessandro Usiello doi: https://doi.org/10.1101/2025.08.08.25333287 Isar Yahyavi 1 CEINGE Biotecnologie Avanzate , Franco Salvatore, Naples, Italy ; 2 Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Università degli Studi della Campania “Luigi Vanvitelli” , 81100, Caserta, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site Federica Carrillo 3 Institute of Genetics and Biophysics, Italian National Research Council CNR , 80131, Naples, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Federica Carrillo Tommaso Nuzzo 1 CEINGE Biotecnologie Avanzate , Franco Salvatore, Naples, Italy ; 2 Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Università degli Studi della Campania “Luigi Vanvitelli” , 81100, Caserta, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Tommaso Nuzzo Anna Di Maio 1 CEINGE Biotecnologie Avanzate , Franco Salvatore, Naples, Italy ; 2 Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Università degli Studi della Campania “Luigi Vanvitelli” , 81100, Caserta, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site Sara Pietracupa 4 IRCCS INM Neuromed , 86077, Pozzilli, Italy ; 5 Department of Human Neuroscience, Sapienza University of Rome , Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Sara Pietracupa Nicola Modugno 4 IRCCS INM Neuromed , 86077, Pozzilli, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Nicola Modugno Francesco Errico 1 CEINGE Biotecnologie Avanzate , Franco Salvatore, Naples, Italy ; 6 Department of Agricultural Sciences, University of Naples “Federico II” , 80055, Portici, Italy Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Francesco Errico Teresa Esposito 3 Institute of Genetics and Biophysics, Italian National Research Council CNR , 80131, Naples, Italy ; 4 IRCCS INM Neuromed , 86077, Pozzilli, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Teresa Esposito For correspondence: teresa.esposito{at}igb.cnr.it alessandro.usiello{at}unicampania.it Alessandro Usiello 1 CEINGE Biotecnologie Avanzate , Franco Salvatore, Naples, Italy ; 2 Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, Università degli Studi della Campania “Luigi Vanvitelli” , 81100, Caserta, Italy ; Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Alessandro Usiello For correspondence: teresa.esposito{at}igb.cnr.it alessandro.usiello{at}unicampania.it Abstract Full Text Info/History Metrics Supplementary material Data/Code Preview PDF Abstract Sex and genetic differences influence the epidemiology of Parkinson’s disease (PD). However, their impact on the homeostasis of N-methyl-D-aspartate receptor (NMDAR)-related amino acids and their precursors remains unexplored. In this study, we measured serum levels of these neuroactive molecules using high-performance liquid chromatography (HPLC) in a genetically and clinically well-characterized cohort of PD patients (n = 245) and healthy controls (n = 203). PD cases were stratified by sex and subtype: idiopathic (n = 121) and genetic (n = 124), the latter including carriers of pathogenic variants in LRRK2 , TMEM175 , PARK2 , PINK1 , PARK7 , and GBA1 . We observed marked sex- and genotype-specific alterations in the serum profiles of D- and L-amino acids. Men with idiopathic conditions exhibited significant reductions in NMDAR-related amino acids and their precursors, including L-glutamate, L-aspartate, glycine, D-serine, L-serine, L-glutamine, and L-asparagine, compared to controls. Conversely, male patients with genetic PD showed a selective decrease in L-aspartate. Female PD patients, regardless of genetic status, did not show significant amino acid reductions. Intriguingly, D-serine levels positively correlated with motor worsening in patients as indexed by MDS-UPDRS III scores in TMEM175 mutation carriers of both sexes. Lastly, targeted genetic screening revealed sex- and subtype-specific associations with GRIN2A polymorphisms, suggesting a genetic contribution to the changes in NMDAR subunit composition reported in PD. Our findings reveal a previously unrecognized impact of sex and genotype differences on NMDAR-related amino acid balance in PD. These results highlight the importance of considering these factors in the discovery of biomarkers and in the development of personalized therapeutic strategies for managing PD. Introduction Parkinson’s disease (PD) is the second most common neurodegenerative disorder ( Bloem et al., 2021 ). Its pathophysiology is characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta combined with the accumulation of intraneuronal proteinaceous aggregates of α-synuclein (α-syn), forming the Lewy bodies (LBs) ( Vázquez-Vélez and Zoghbi, 2021 ). PD clinical manifestations include both motor symptoms, including tremors at rest, bradykinesia, rigidity, and postural instability, and non-motor symptoms, such as constipation, depression, anosmia, and sleep disorders, which often precede motor onset. In later stages, cognitive impairment and dysautonomia may also emerge ( Obeso et al., 2017 ; Vázquez-Vélez and Zoghbi, 2021 ). Although PD affects both sexes, it exhibits notable sex-specific differences in incidence, clinical manifestations and treatment responses ( Cattaneo and Pagonabarraga, 2025 ; Pringsheim et al., 2014 ). Increasing evidence suggests that sex hormones, particularly estrogens, may exert neuroprotective effects in the nigrostriatal pathway degeneration ( Bourque et al., 2024 ; Cerri et al., 2019 ; Smith and Dahodwala, 2014 ). Genetic background also significantly influences disease onset and PD progression ( Day and Mullin, 2021 ; Guadagnolo et al., 2021 ). Recent studies have focused on identifying monogenic disease-causing genes ( Lim et al., 2024 ; Vollstedt et al., 2023 ). While L15% of patients report a family history of PD, 5–10% carry monogenic mutations with Mendelian inheritance. However, the majority of PD cases are idiopathic ( Bloem et al., 2021 ), likely resulting from complex gene-environment interactions ( Kalinderi et al., 2016 ; Tambasco et al., 2016 ). Beyond dopaminergic neuron degeneration, glutamatergic dysfunction also contributes to PD pathogenesis ( Almohmadi et al., 2025 ; Wang et al., 2020 ). Altered glutamatergic neurotransmission within the basal ganglia, especially involving N-methyl-D-aspartate ionotropic receptor (NMDAR), has been widely reported ( Campanelli et al., 2022 ; Dunah et al., 2000; Gasparini et al., 2013 ; Pagonabarraga et al., 2021 ; Zhang et al., 2019 ). Alterations in NMDAR expression and activity in the caudate-putamen are implicated in maladaptive synaptic plasticity, L-DOPA induces-dyskinesia and cognitive deficits in both PD patients and animal models ( Calabresi et al., 2008 ; Campanelli et al., 2022 ; Gardoni and Di Luca, 2015 ). As endogenous co-agonists, glycine (Gly) and D-serine (D-Ser) have gained attention for their role in modulating NMDAR activity and potential therapeutic value in PD ( Gelfin et al., 2012 ; Heresco-Levy et al., 2013 ). Interestingly, in vivo studies in rodent and monkey models of PD showed that enhanced stimulation at the glycine-binding site of NMDAR facilitates striatal dopaminergic reinnervation and improves motor and non-motor dysfunctions ( Frouni et al., 2022 , 2021 ; Frouni and Huot, 2022 ; Ho et al., 2011 ; Schmitz et al., 2013 ; Schneider et al., 2000 ; Zhang et al., 2021). Consistent with these findings, elevated levels of D-Ser and its precursor, L-Ser, have been reported in the post-mortem putamen of MPTP-intoxicated monkeys ( Nuzzo et al., 2019 ; Serra et al., 2023 ) and caudate-putamen of PD patients (Gervasoni et al., 2025a; Di Maio et al., 2023 ). Additionally, increased levels of both serine enantiomers, and reduced L-glutamate (L-Glu) levels were found in the cerebrospinal fluid (CSF) of drug-naïve PD patients compared to individuals with other neurodegenerative diseases ( Di Maio et al., 2023 ). More recently, serum HPLC and metabolomic analyses in PD patients and age-matched healthy controls further confirmed significant dysregulation of serine enantiomers and glutamate levels in individuals with PD (Gervasoni et al., 2025b; Imarisio et al., 2024 ). Although these exploratory studies suggest a disruption of NMDAR-related amino acids homeostasis in this neurodegenerative disease (NDD), it remains unclear whether these metabolic alterations are influenced by patient sex or PD subtype. To address this gap, we investigated the influence of sex and genetic differences in modulating serum levels of NMDAR-related amino acids and their precursors, including L-Glu, L-aspartate (L-Asp), D-Ser, L-Ser, Gly, L-glutamine (L-Gln) and L-asparagine (L-Asn) in a large and well-characterized cohort of PD patients and healthy controls (HC). Patients were stratified by sex and PD subtype: idiopathic PD (without known pathogenic mutations and without rare variants in at-risk PD genes) and genetic PD (harboring mutations in LRRK2 , TMEM175 , PARK2 , PINK1 , PARK7 , or GBA1 ). We further assessed the relationship between amino acid levels and demographic or clinical variables, including age, disease duration, age at onset, Levodopa Equivalent Daily Dose (LEDD), and motor symptom severity (Movement Disorder Society-Unified Parkinson’s Disease Rating Scale III, MDS-UPDRS III). Our findings reveal a previously unrecognized contribution of sex and genotype to the regulation of blood NMDAR-related amino acid homeostasis in patients with PD. Results Clinical and demographic characteristics of the study cohort We analyzed demographic and clinical data of 245 PD patients and 203 HC ( Table 1 ) . PD patients were significantly older and had a higher proportion of males compared to HC ( Table 1 ) . Within the PD cohort, 121 patients were classified as idiopathic PD (iPD; 65 males, 56 females) and 124 as genetic PD (gPD; 64 males, 60 females), based on the presence of pathogenic mutations ( Table 2 ; Suppl. Table 1 ). View this table: View inline View popup Download powerpoint Table 1. Demographic and clinical features of Parkinson’s disease patients and healthy controls subjects. View this table: View inline View popup Download powerpoint Table 2. Demographic and clinical features of idiopathic PD, genetic PD and HC groups. Both iPD and gPD groups differed from HC in terms of age (median: iPD and gPD 68 years; HC 61 years) and sex distribution. However, iPD and gPD patients were comparable in sex prevalence, age, age at onset, LEDD, and motor symptom scores (MDS-UPDRS III) ( Table 2 ) . Notably, gPD patients had longer disease duration (median [IQR], 6 [3–10] years), compared to iPD group (5 [3– 8] years) ( Table 2 ) . Sex-stratified analysis revealed that iPD and gPD males were clinically indistinguishable across all variables ( Suppl. Table 2 ). In contrast, gPD females showed earlier age at onset (61.5 [52.5-65] years) and longer duration of motor symptoms (7 [3.5-12] years) than iPD females, while other measures remained comparable ( Suppl. Table 2 ). Among gPD patients, no significant differences were observed across carriers of GBA1 (13 males; 17 females), LRRK2 (6 males; 11 females), PARK2/PINK1/PARK7 (22 males; 18 females), or TMEM175 (21 males; 12 females) in age, age at onset, disease duration, LEDD, and MDS-UPDRS III ( Suppl. Table 3 ). PD patients show lower serum L-aspartate and L-glutamate levels compared to healthy controls To explore the influence of PD on blood NMDAR-related amino acids and their precursors, we first conducted an overall data analysis without stratifying patients by sex or genotype. We measured serum levels of D-Ser, L-Ser, Gly, L-Glu, L-Gln, L-Asp, and L-Asn in 245 PD patients and 203 HC by HPLC analysis ( Fig. 1a ; Suppl. Table 4 ). Also, we evaluated the D-Ser/total Ser and L-Gln/L-Glu ratios as indicators of conversion of D-Ser and L-Glu from their precursors, L-Ser and L-Gln, respectively. Non-parametric Mann-Whitney analysis revealed significantly elevated serum D-Ser levels and D-Ser/total Ser ratio in PD cases compared with HC ( Fig. 1b ; Suppl. Table 4 ). Conversely, L-Glu, L-Asp, and L-Asn levels were significantly lower in PD patients compared to controls ( Fig. 1b ; Suppl. Table 4 ). Additionally, we observed a mild increase of L-Gln/L-Glu ratio in the PD group. The remaining amino acids level showed no significant differences between groups ( Fig. 1b ; Suppl. Table 4 ). Download figure Open in new tab Figure 1. D- and L-amino acids levels in the serum of Parkinson’s disease patients and healthy subjects. (a) Representative HPLC chromatogram illustrating the separation and detection of D-serine (D-Ser), L-serine (L-Ser), glycine (Gly), L-glutamate (L-Glu), L-glutamine (L-Gln), L-aspartate (L-Asp), and L-asparagine (L-Asn) peaks from a blood serum sample. The magnification of D-Ser peak is shown. (b) Violin plots representing D-Ser, L-Ser, D-Ser/total Ser ratio, Gly, L-Glu, L-Gln, L-Gln/L-Glu ratio, L-Asp, and L-Asn levels in the serum of Parkinson’s disease patients (PD, N=245) compared to healthy control subjects (HC, N=203). The amino acid content is expressed as µM. In each sample, free amino acids were detected in a single run. Dots represent the single subjects’ values, while lines illustrate the median with interquartile range. * p <0.05, ** p <0.01, *** p <0.001 (Mann–Whitney U test). Only significant differences confirmed by age-, sex-, and LEDD-adjusted ANCOVA are displayed. To account for potential confounding factors, we used ANCOVA analysis on natural log-transformed data, adjusting for age, sex, and LEDD. This analysis confirmed that variations in D-Ser/total Ser and L-Gln/L-Glu ratios, as well as L-Glu and L-Asp levels, remained significantly associated with the clinical condition ( Fig. 1b ; Suppl. Table 4 ). Sex stratification reveals male-specific alterations in NMDAR-related amino acid levels in PD We next evaluated whether sex modulates the alteration of NMDAR-related amino acids and their precursors in PD versus sex-matched HC ( Fig. 2 ) . These analyses revealed that the dysregulation observed in the overall PD cohort ( Fig. 1b ) was largely driven by male patients ( Fig. 2 ). Specifically, men with PD showed significantly reduced L-Asp and L-Glu levels compared to male controls ( Fig. 2a ; Suppl. Table 5 ). In addition, lower serum L-Ser, Gly, L-Gln, and L-Asn levels were detected in male PD patients ( Fig. 2a ; Suppl. Table 5 ). These findings were further confirmed by ANCOVA, adjusting for age and LEDD ( Fig. 2a ; Suppl. Table 5) . Yet, a significant increase in the D-Ser/total Ser ratio was found in male patients relative to controls ( Fig. 2a ; Suppl. Table 5) . Download figure Open in new tab Figure 2. Serum D- and L-amino acids content in male and female Parkinson’s disease patients and healthy subjects. Analysis of D-serine (D-Ser), L-serine (L-Ser), D-Ser/total Ser ratio, glycine (Gly), L-glutamate (L-Glu), L-glutamine (L-Gln), L-Gln/L-Glu ratio, L-aspartate (L-Asp), and L-asparagine (L-Asn) levels in the serum of male (a) and female (b) Parkinson’s disease patients (male, N=129; female, N=116) compared to healthy control (HC) subjects (male, N=80; female N=123). The amino acid content is expressed as µM. In each sample, free amino acids were detected in a single run. Dots represent the single subjects’ values, while lines illustrate the median with interquartile range. * p <0.05, ** p <0.01, *** p <0.001 (Mann–Whitney U test). Only significant differences confirmed by age- and LEDD-adjusted ANCOVA are displayed. Interestingly, in female PD patients, we found increased serum levels of D-Ser and Gly, as well as elevated L-Gln/L-Glu ratio, compared to HC ( Fig. 2b ; Suppl. Table 5 ). However, following ANCOVA adjustment for age and LEDD, only the L-Gln/L-Glu ratio remained statistically significant ( Fig. 2b ; Suppl. Table 5 ). Collectively, these findings reveal a sex-specific dysregulation of D and L-amino acids in PD, with male patients exhibiting more extensive and pronounced amino acids reductions compared to females. Genotype stratification reveals predominant NMDAR-related amino acid dysregulations in idiopathic PD subtype To investigate whether genotype differences impact the serum levels of NMDAR-related amino acids and their precursors, we stratified the PD cohort into two disease subtypes: iPD comprising patients without known genetic etiology, and gPD including patients carrying at least one mutation in the most frequently mutated PD genes such as LRRK2 , TMEM175 , PARK2 , PINK1 , PARK7 , and GBA1 ( Table 2 ; Suppl. Table 1 ). Amino acid levels in both PD subgroups were then compared to those of HC. Notably, data analyses revealed that only iPD patients exhibited significantly lower concentrations of L-Glu, L-Asp, L-Ser, and L-Asn, as well as elevated L-Gln/L-Glu ratio, compared to HC ( Fig. 3 ; Suppl. Table 6 ). These findings remained significant after ANCOVA adjustment for age, sex and LEDD ( Suppl. Table 6) . In contrast, gPD patients showed a selective increase in D-Ser levels compared to controls ( Fig. 3 ; Suppl. Table 6) . No significant changes were detected in all the other amino acids tested ( Fig. 3 ; Suppl. Table 6) . Download figure Open in new tab Figure 3. Serum D- and L-amino acids profile in idiopathic PD, genetic PD and healthy controls. Analysis of D-serine (D-Ser), L-serine (L-Ser), D-Ser/total Ser ratio, glycine (Gly), L-glutamate (L-Glu), L-glutamine (L-Gln), L-Gln/L-Glu ratio, L-aspartate (L-Asp), and L-asparagine (L-Asn) levels in the serum of idiopathic PD (iPD, N=121), genetic PD (gPD, N=124) and healthy controls (HC, N=203). The amino acid content is expressed as µM. In each sample, free amino acids were detected in a single run. Dots represent the single subjects’ values, while lines illustrate the median with inter quartile. * p <0.05, ** p <0.01, *** p <0.001 (post hoc Dunn’s test). Significant post hoc results are shown when Kruskal–Wallis test was confirmed by age-, sex-, and LEDD-adjusted ANCOVA. Finally, we directly compared amino acid levels between the iPD and gPD subgroups. Mann-Whitney analysis revealed that iPD patients had significantly lower L-Glu, L-Ser, D-Ser, L-Asn, and Gly levels than gPD patients ( Suppl. Fig. 1 ). These differences were confirmed by ANCOVA correction for age, sex, LEDD and disease duration. No significant differences were found for the other tested amino acids ( Suppl. Fig. 1 ). Sex and genotype-stratified analysis reveals pronounced amino acids dysregulation in men with idiopathic PD To explore how sex and genotype influence blood D and L-amino acid levels in Parkinson’s disease, we compared serum concentrations of these biomolecules among individuals with iPD, gPD, and HC within each sex group. Statistical analysis revealed that iPD males exhibited significant reductions in L-Asp, L-Glu, L-Ser, D-Ser, Gly, L-Gln, and L-Asn levels compared to HC ( Fig. 4a ; Suppl. Table 7 ). In contrast, gPD males showed only a significant decrease in L-Asp levels compared to HC ( Fig. 4a ; Suppl. Table 7) . Additionally, an increased D-Ser/total Ser ratio was observed in both iPD and gPD males compared to controls ( Fig. 4a ; Suppl. Table 7 ). All these variations remained significant after ANCOVA adjustment for age and LEDD (Suppl. Table 7) . Download figure Open in new tab Figure 4. Serum D- and L-amino acids content in male and female subgroups of idiopathic PD, genetic PD and healthy controls. Analysis of D-serine (D-Ser), L-serine (L-Ser), D-Ser/total Ser ratio, glycine (Gly), L-glutamate (L-Glu), L-glutamine (L-Gln), L-Gln/L-Glu ratio, L-aspartate (L-Asp), and L-asparagine (L-Asn) levels in the serum of male (a) and female (b) idiopathic PD (iPD; male N=65, female N=56), genetic PD (gPD; male N= 64, female N= 60) and healthy controls (HC, male N=80, female N=123). The amino acid content is expressed as µM. In each sample, free amino acids were detected in a single run. Dots represent the single subjects’ values, while lines illustrate the median with interquartile range. * p <0.05, ** p <0.01, *** p <0.001 (post hoc Dunn’s test). Significant post hoc results are shown when Kruskal–Wallis test was confirmed by age-, and LEDD-adjusted ANCOVA. In women, Kruskal–Wallis analysis revealed an increased L-Gln/L-Glu ratio in iPD and elevated D-ser levels in gPD compared to HC. However, after adjustment for age and LEDD using ANCOVA, only the alteration in the L-Gln/L-Glu ratio remained statistically significant ( Fig. 4b ; Suppl. Table 8 ). Finally, we compared iPD and gPD patients within each sex. In males, iPD patients had significantly lower levels of D-Ser, L-Ser, Gly, L-Glu, L-Gln, L-Asp, and L-Asn compared to gPD patients ( Fig. 5a ; Suppl. Table 9) , with findings confirmed by ANCOVA adjusting for age, LEDD, and disease duration (Suppl. Table 9). In contrast, no significant differences were observed between iPD and gPD females, except for an increase in D-Ser levels in gPD patients, which did not remain significant after ANCOVA correction. Download figure Open in new tab Figure 5. Comparison of serum D- and L-amino acid levels in male and female Parkinson’s disease patients compared between idiopathic and genetic forms. Analysis of D-serine (D-Ser), L-serine (L-Ser), D-ser/total Ser ratio, glycine (Gly), L-glutamate (L-Glu), L-glutamine (L-Gln), L-Gln/L-Glu ratio, L-aspartate (L-Asp), and L-asparagine (L-Asn) in the serum of male (a) and female (b) idiopathic PD (iPD; male N=65, female N=56) and genetic PD (gPD; male N= 64, female N=60). The amino acid content is expressed as µM. In each sample, free amino acids were detected in a single run. Dots represent the single subjects’ values, while lines illustrate the median with interquartile range. * p <0.05, ** p <0.01, *** p <0.001 (Mann–Whitney U test). Only differences confirmed by age-, LEDD- and disease duration-adjusted ANCOVA are displayed. Overall, these results indicate a sex- and genotype-specific pattern of circulating NMDAR-related amino acid dysregulation in PD compared to controls. Intriguingly, a pronounced disruption of glutamatergic amino acid homeostasis is observed exclusively in men with iPD, while women, regardless of subtype, exhibited only mild or inconsistent alterations. Analysis of specific pathogenic mutations reveals limited impact on NMDAR-related amino acid levels in genetic PD patients We next investigated whether distinct pathogenic mutations in LRRK2 , GBA1 , TMEM175 , or PARK2/PINK1/PARK7 genes impact on circulating levels of NMDAR-related amino acids within male and female PD patient sub-cohorts. Statistical analysis showed no significant amino acid alterations in either men or women carrying these mutations compared to sex-matched HC (Suppl. Fig. 2a, b) . However, a clear trend toward increased D-Ser levels was observed in female mutation carriers, particularly in those with LRRK2 mutation, although this alteration did not reach statistical significance (Suppl. Fig. 2b) . All other amino acid levels were comparable across all mutation groups of both sexes (Suppl. Fig. 2a, b). Finally, we directly compared amino acid levels among PD patients carrying different pathogenic mutations within male and female sub-cohorts. Statistical analysis revealed no significant differences across LRRK2 , GBA1 , TMEM175 , and PARK2/PINK1/PARK7 mutation carriers, in either sex. However, a trend toward increased D-Ser levels were observed specifically in female patients with LRRK2 mutations, compared to those with other genetic variants (Suppl. Fig. 3) . Altogether, these HPLC findings suggest that distinct pathogenic mutations do not significantly impact the circulating levels of NMDAR-related amino acids and their precursors, except for a trend of female-specific upregulation of D-Ser associated with LRRK2 . Correlation analysis between amino acid levels and age or age at disease onset reveals sex- and subtype-specific patterns We explored the relationships between NMDAR-related amino acids and demographic or clinical variables across iPD, gPD and HC groups within male and female cohorts ( Table 2 ) . Spearman’s correlation analysis in male revealed no significant associations between circulating amino acid levels and age in both HC and all PD subjects regardless of disease subtype ( Table 3 ; Suppl. Fig. 4a) . In contrast, serum D-Ser levels and the D-Ser/total Ser ratio positively correlated with age in female HC ( Table 3 ; Suppl. Fig. 4b) . Similarly, iPD women showed a positive correlation between D-Ser levels and age. In contrast, no such associations were found in female gPD patients ( Table 3 ; Suppl. Fig. 4b) . View this table: View inline View popup Download powerpoint Table 3. Correlations analysis between serum levels of D- and L-amino acids with age of idiopathic PD, genetic PD and healthy individuals. Then, we examined correlations between amino acid levels and clinical features, including age at onset, disease duration, and LEDD ( Table 4 ) . Interestingly, in male patients, we found no significant correlations, either in iPD or gPD subgroups ( Table 4 ; Suppl. Fig. 5a) . In women, only D-Ser levels positively correlated with age at onset in iPD group ( Table 4 ; Suppl. Fig. 5b) , confirming previous observation ( Imarisio et al., 2024 ). Contrary to our previous findings showing a negative correlation between D-Ser levels and both disease duration and LEDD ( Imarisio et al., 2024 ), the current analysis in a larger, genetically characterized cohort did not confirm significant associations with either of these variables across PD subgroups ( Table 4 ) . Similarly, no significant correlations were found between other amino acids and clinical features across PD subgroups ( Table 4 ) . All reported associations with disease duration and LEDD were not confirmed by partial correlation analyses ( Table 4 ) . View this table: View inline View popup Download powerpoint Table 4. Correlation analysis of serum D- and L-amino acids level with age of disease onset, LEDD and disease duration of PD groups. Overall, our results suggest that the relationship between NMDAR-related amino acid levels and demographic or clinical features is modulated by both sex and PD subtype. D-serine levels correlate with motor symptoms severity in male and female TMEM175 mutation carriers We examined whether serum levels of NMDAR-related amino acids were correlated with the severity of motor symptoms in iPD and gPD patients, stratified by sex ( Fig. 6a-d ; Table 5 ) . Download figure Open in new tab Figure 6. Correlation of serum D-serine and L-serine concentrations with MDS-UPDRS III of idiopathic PD and genetic PD patients stratified for sex. Scatterplots showing the association of D-serine (D-Ser) or L-serine (L-Ser) content with MDS-UPDRS III of male ( a, c, e) and female ( b, d, f ) idiopathic PD ( a, b ), genetic PD ( c, d ) and genetic PD stratified for GBA1 , LRRK2 , PARK2/PINK1/PARK7 and TMEM175 pathogenic variants ( e, f ). Best fit lines and 95% confidence intervals are shown. * p < 0.05,** p < 0.01, Spearman’s correlation test confirmed by partial correlation analysis. View this table: View inline View popup Download powerpoint Table 5. Correlation analysis of serum D- and L-amino acids levels with MDS-UPDRS III of patients with idiopathic and genetic PD. Spearman’s correlation analysis showed no significant correlations between amino acid levels and MDS-UPDRS III in iPD patients of both sex ( Table 5 ) . Conversely, a positive correlation between D-Ser levels and motor symptom severity was observed in both male and female gPD patients ( Fig. 6a-d ; Table 5 ) . Additionally, only in male gPD patients, a negative correlation emerged between the L-Gln/L-Glu ratio and motor symptom scores ( Table 5 ) . The reported associations were confirmed by partial correlation analyses adjusted for age, disease duration, and LEDD ( Table 5 ) . Finally, to determine whether these correlations were specific to a particular genetic condition, we stratified gPD patients according to their specific pathogenic mutations ( Fig. 6e, f ) . Interestingly, data analysis revealed a direct correlation between D-Ser levels and the severity of motor symptoms specific to patients harboring the TMEM175 mutation, irrespective of their sex ( Fig. 6e, f ) . Our findings suggest that the correlations between blood NMDAR-related amino acid levels and motor symptoms severity is influenced by both sex and genetic factors in patients with PD. NMDAR subunit gene variants exhibit sex- and subtype-specific association in Parkinson’s disease Beyond the occurrence of circulating NMDAR-related amino acids, we examined whether sex and genetic status of PD patients influence the genetic architecture of genes encoding NMDAR subunits, including GRIN1 , GRIN2A , and GRIN2B . We first applied Principal Component Analysis (PCA) to assess and account for population structure within the study cohort, a key step in genetic association studies to control for population stratification, which could otherwise lead to spurious associations. PCA disclosed that the cohort was highly genetically homogeneous ( Suppl. Fig. 6a ), with PC1 and PC2 contributing to 25% of the total variance (Suppl. Fig. 6b) . All subsequent association analyses we adjusted for age, sex and 10 principal components. We initially applied Sequence Kernel Association Test – Optimal (SKAT-O) test to evaluate the joint effects of multiple rare exonic, non-synonymous variants (Minor Allele Frequency, MAF<0.01, MAF<0.005 and MAF<0.001) in GRIN1 , GRIN2A and GRIN2B genes on PD susceptibility. This was done in the same cohort used for HPLC analysis stratified by sex and PD subtype. No significant associations were observed between rare variants in these genes and PD risk. Next, we investigated whether common variants (MAF>0.01) in GRIN1 , GRIN2A and GRIN2B genes were associated with PD susceptibility. A total of 70 variants were identified within the genomic regions from gene start to end (based on GRCh38 Whole Exome Sequencing, WES, data), including exons, 5’ and 3’ untranslated regions (UTRs) and intronic regions near splicing junctions. Association analyses were performed on male and female PD patients, stratified for PD subtype, and HC used for HPLC experiments. In addition, other case-control cohorts were considered; i) Mediterranean Neurological Institute (MNI) cohort (stratified for sex and PD subtype): PD (n=193 MNI-iPD, n=227 MNI-gPD) versus MNI-HC (n=282), stratified by sex; ii) MNI overall cohort (no stratification): MNI-PD (n=804) versus MNI-HC (n=282); iii) Parkinson disease Genetic Consortium (PDGC) cohort (4586 PD patients) versus United Kingdom (UK) biobank (43989 controls from general population) (no stratification). The first three analyses used an additive genetic model with PLINK2 software, while the fourth used Fisher’s exact test since data is available only as aggregated genotypes. We also calculated the odds ratio (OR) value for each association test to assess the direction and strength of the association between the genetic variants and PD condition. An odds ratio (OR) less than 1 suggests that the associated variant has a protective effect, indicating that it is more frequent in controls than in PD patients. Conversely, an OR greater than 1 indicates a risk effect, with the associated allele more frequent in PD patients than in HC. In our study cohort, significant associations were identified for two intronic variants: rs11866570 (p value (p) = 0.007, OR = 0.02) in GRIN2A and rs11055581 (p = 0.01, OR = 215) in GRIN2B in male iPD patients compared to sex-matched HC ( Table 6 ). No variants were associated with iPD in women ( Table 6 ). View this table: View inline View popup Download powerpoint Table 6. Common variants in GRIN2A and GRIN2B genes encoding NMDAR subunits associated with PD in case-control study Interestingly, in male gPD patients, the following variants were associated with PD: rs62621078 ( GRIN2A intronic variant, p = 0.005, OR = 123) and rs7301328 ( GRIN2B synonymous variant, p = 0.01, OR = 0.46), whereas in female gPD patients we observed rs6497540 ( GRIN2A intronic variant, p = 0.004, OR = 0.38) and rs1806201 ( GRIN2B synonymous variant, p = 0.004, OR = 0.35) ( Table 6 ). No variants in GRIN1 were associated with PD in our cohorts. Although none of these associations remained significant after correction for multiple testing (threshold set to 7.1×10 -4 considering 70 variants), this analysis suggests at a nominal significance level, that genetic variations in GRIN2A and GRIN2B may contribute to PD susceptibility or protection, in a sex- and PD subtype-specific manner. To validate these findings, we analyzed the MNI cohort, stratified by sex and PD subtype. The rs11866570 variant in GRIN2A was significantly associated with PD in male iPD patients (p = 0.005, OR = 0.24) but not female, supporting its protective effect (as indicated by OR < 1). We then analyzed both the full MNI cohort and the PDGC/UK Biobank cohort, without stratification to assess the broader relevance of the variants not replicated in the stratified analysis. The intronic rs11055581 variant in GRIN2B showed association with PD in both MNI cohort (p = 0.003; OR = 148) and PDGC cohort (p < 0.00001; OR = 1.12) (Suppl. Table 10) . This supports its potential role as PD susceptibility factor. The two synonymous GRIN2B variants, rs7301328 and rs1806201, were associated with PD only in the PDGC cohort (p < 0.00001; OR = 0.91, p = 0.005; OR = 0.94, respectively) (Suppl. Table 10) . Given the large sample size in the PDGC/UK cohort, these variants may act as mild protective factors for PD. Our findings suggest that genetic variability in GRIN2A and GRIN2B , but not GRIN1 , may contribute to PD susceptibility or protection in a sex- and subtype-specific manner. In particular, the GRIN2A intronic variant rs11866570 emerged as a potential protective factor for male idiopathic PD, and was replicated in an independent cohort. The novel identified variants in GRIN2A and GRIN2B are associated with increased gene expression in the basal ganglia of the human brain To further investigate the functional effects of the GRIN2A and GRIN2B variants associated with PD, we surveyed the Genotype-Tissue Expression (GTEx) portal ( https://www.gtexportal.org/home/ ), which links genetic variation with tissue-specific gene expression. We specifically examined the impact of the identified single-nucleotide-polymorphisms (SNPs) on gene expression in various human brain regions, including amygdala, anterior cingulate cortex (BA24), caudate basal ganglia, cerebellum, cortex, frontal cortex (BA9), hippocampus, hypothalamus, nucleus accumbens, putamen, and substantia nigra. It should be noted that GTEx data are not stratified by sex or PD subtype. The protective rs11866570-C allele in GRIN2A , associated with PD only in male iPD patients, was significantly associated with increased GRIN2A expression in cerebellum (p=2.8e -5 ) and putamen (p=3.5e- 3 ) and decreased expression in caudate (p=5.5e- 4 ) ( Fig 7 a ; Suppl. Table 11 ). Since this allele is associated with reduced PD risk, we can hypothesize that male iPD patients may exhibit lower GRIN2A expression in the cerebellum and putamen, and higher expression in the caudate, compared to other PD subtypes. Similarly, increased GRIN2B expression was observed in association with the two protective alleles rs7301328-C and rs1806201-A, which were identified in the large PDCG/UK case-control cohort. In particular, the rs7301328-C allele was associated with increased GRIN2B expression in the substantia nigra (p=6.06e- 3 ) ( Fig. 7 b ; Suppl . Table 12 ), while, the rs1806201-A allele was associated with increased GRIN2B expression in multiple regions, including amygdala (p=7.55e- 4 ), putamen (p=1.3e- 4 ), caudate (p=3.88e- 4 ), cortex (p=1.74e- 4 ) and hypothalamus (p=6.43e- 4 ) ( Fig. 7 c; Suppl. Table 13 ). Given their putative protective effect, these results suggest that PD patients may exhibit relatively lower GRIN2B expression in these brain regions. Download figure Open in new tab Figure 7. Genotype-Tissue Expression (Gtex) analysis of GRIN2A and GRIN2B transcripts in human brain regions associated with the presence of the allele variants rs11866570-C, rs7301328-C and rs1806201-A. Violin plots show significantly increased expression of GRIN2A transcript associated with the presence of the protective allele C in Cerebellum and Putamen (a) . On the contrary, reduced expression was observed in Caudate (a) . In b is shown the significant increased expression of GRIN2B transcript in the Substantia nigra in the presence of the protective allele rs7301328-C. In c is reported the increased expression of GRIN2B transcript in Amygdala, Caudate, Putamen, Cortex and Hypothalamus associated with the protective allele rs1806201-A. Discussion Previous studies have reported significant amino acid metabolism alterations in the serum or plasma of PD patients ( Akdas et al., 2024 ; Figura et al., 2018 ; Gervasoni et al., 2025b; Imarisio et al., 2024 ; JiménezLJiménez et al., 2020). However, recent meta-analyses have highlighted inconsistencies across metabolomic findings (JiménezLJiménez et al., 2020; Luo et al., 2024 ), likely reflecting differences in methodological platform and cohort stratification. In particular, the contribution of sex and genetic background in modulating amino acid metabolism in PD remains poorly understood. In the present study, we addressed this gap by performing a comprehensive HPLC characterization of serum levels of D- and L-amino acids involved in NMDAR signaling, a pathway increasingly implicated in PD pathophysiology and L-DOPA effects ( Campanelli et al., 2022 ; Di Maio et al., 2023 ; Nuzzo et al., 2019 ; Serra et al., 2023 ). Our findings indicate a significant dysregulation in the circulating levels of NMDAR-related amino acids and their precursors in PD patients, which is influenced by both sex and genotype. Notably, male patients showed marked reductions in several amino acids, including L-Ser, L-Glu, L-Gln, L-Asp and L-Asn, accompanied by a significant increase in D-Ser/total Ser ratio, compared to sex-matched controls. In contrast, female PD patients displayed no significant reduction in blood amino acid levels. Altogether these results are supported by complementary metabolomic investigations. An untargeted NMR-based analysis conducted on the same patient cohort confirmed a significant sex-related effect on amino acid metabolism, with male PD patients exhibiting more pronounced alterations than female patients compared to sex-matched controls. In line with our observations, independent metabolomic studies in other PD cohorts reported sex-specific differences in both amino acid and lipid metabolism ( Hu et al., 2024 ; Jellen et al., 2025 ; Luo et al., 2024 ; Meoni et al., 2022 ). Further stratification by genotype of PD patients revealed that male subjects with idiopathic condition exhibited the most pronounced amino acidic dysregulation, including a notable reduction of D-Ser, L-Ser, Gly, L-Glu, L-Gln, L-Asp and L-Asn, along with increased D-Ser/total Ser ratio, compared to sex-matched healthy controls. Conversely, male patients carrying pathogenic mutations showed more restricted alterations, limited to decreased L-Asp levels and elevated D-Ser/total Ser. Among females, only an increase in L-Gln/L-Glu ratio was observed in the idiopathic group, while no differences were found in female genetic PD patients. Noteworthy, our findings highlight robust abnormalities of NMDAR-related amino acids selectively in male PD patients, particularly those with idiopathic conditions. In contrast, the relative homeostasis stability of amino acid metabolism seen in female PD patients may indicate a protective effect of estrogen or sex chromosome-related genes, which may ultimately influence neurotransmission, oxidative stress, and inflammation ( Goyette et al., 2023 ; Hu et al., 2024 ; Meoni et al., 2022 ; Schaffner et al., 2025 ; Zlotnik et al., 2011 ). Additionally, to investigate how specific pathogenic mutations affect blood amino acid profiles, we further divided gPD patients based on mutations in the most frequently mutated PD genes: LRRK2 , TMEM175 , GBA1 , and PARK2/PINK1/PARK7 ( Blauwendraat et al., 2020 ; Carrillo et al., 2025 ; Gialluisi et al., 2021 ). HPLC analysis evidenced that none of these mutations were associated with significant changes in serum amino acid levels, regardless of sex. Although LRRK2, TMEM175, GBA1, and PARK2/PINK1/PARK7 are expressed in the central nervous system and peripheral organs ( https://www.proteinatlas.org/ ) ( Huh et al., 2023 ; Matsuda et al., 2013 ; Palomba et al., 2023 ; Safreena et al., 2024 ; Serdaroglu et al., 2005 ; Shimura et al., 2000 ; Singh et al., 2019 ; Tang et al., 2023 ; Taymans et al., 2023 ; Tsafaras and Baekelandt, 2022 ), and are known to impact mitochondrial function, lysosomal activity, autophagy, and oxidative stress regulation ( Biosa et al., 2017 ; Huh et al., 2023 ; Narendra and Youle, 2024 ; Safreena et al., 2024 ; Taymans et al., 2023 ; Wang et al., 2025 ; Wu et al., 2023 ), our findings suggest that these genes individually do not lead to detectable changes in blood levels of these molecules, as measured by HPLC. In contrast, significant systemic amino acid metabolism alterations may become apparent only when multiple risk factors – genetic, environmental, or sex-related – converge, as observed in idiopathic male patients. Although other investigations are warranted, our integrated design may help clarify discrepancies in amino acid levels across previous studies using blood samples (JiménezLJiménez et al., 2020; Luo et al., 2024 ), which lacked stratification by sex and genotype. Besides the central role of dopaminergic neurons degeneration ( Álvarez-Luquín et al., 2025 ; Zhou et al., 2023 ), PD is increasingly recognized as a heterogeneous, multiorgan disorder ( Wüllner et al., 2023 ; Zesiewicz, 2019 ). In this context, we hypothesize that the widespread deficiency of NMDAR-related amino acids and their precursors observed in idiopathic male patients may provide a rationale for patient stratification and serve as a non-invasive, serum-based signature of multisystemic dysfunction. In addition to organ-based alterations, other factors – such as dietary intake and gut microbiota composition – are known to influence circulating amino acid profiles and may contribute to the metabolic abnormalities seen in PD ( Wang et al., 2022 ; Zhang et al., 2022 ), particularly in the light of gastrointestinal dysfunctions and nutritional imbalance frequently reported in this disorder ( Fasano et al., 2015 ; Wang et al., 2022 ). Previous studies have shown that blood levels of several D- and L-amino acids analyzed in this work are linked with organ functions and metabolic parameters, including liver enzymes and renal clearance ( Hesaka et al., 2019 ; Kimura et al., 2023 ; Sasabe et al., 2016 ; Sasabe and Suzuki, 2018b ; Suzuki et al., 2022 ). These observations therefore raise the possibility that changes in serum levels of NMDAR-related amino acids and their precursors may also serve as surrogate indicators of the broader metabolic organ dysfunctions associated with PD. The observed decrease in L-Glu, L-Gln, D-Ser, L-Ser, Gly, L-Asn and L-Asp levels in idiopathic male patients suggests the disruption of various metabolic pathways, including protein, hormone and phospholipid biosynthesis, mitochondrial energy production and antioxidant defense ( Butterfield et al., 2022 ; Figura et al., 2018 ). The reduction in L-Glu – consistently reported in independent PD cohorts ( Akdas et al., 2024 ; Gervasoni et al., 2025b; Imarisio et al., 2024 ; JiménezLJiménez et al., 2020; Luo et al., 2024 ; Meoni et al., 2022 ) – is particularly relevant given its central role in redox homeostasis, a process known to be impaired in PD ( Anandhan et al., 2017 ). Along with cysteine and Gly, L-Glu is a precursor of glutathione, the major intracellular antioxidant ( Akdas et al., 2024 ; Luo et al., 2024 ; Smeyne and Smeyne, 2013 ). Thus, reduced serum levels of L-Glu and Gly may reflect impaired glutathione biosynthesis and antioxidant capacity. Supporting this interpretation, recent untargeted NMR-based metabolomic analyses on the same PD cohort have revealed significant alterations in glutathione metabolism in male PD patients compared to sex-matched healthy controls. Moreover, L-Glu, along with L-Gln and L-Asp, plays a central role in mitochondrial metabolism, serving as key substrates for anaplerotic reactions, energy production, and nitrogen balance. Interestingly, these metabolic pathways have consistently been reported as altered in PD, as demonstrated by independent metabolomic studies ( Bose and Beal, 2016 ; Gao et al., 2022 ; Gątarek et al., 2022 ; Gervasoni et al., 2025; Imarisio et al., 2024 ; Meoni et al., 2022 ; Tassone et al., 2023 ). Noteworthy, the combined reduction in L-Ser, D-Ser and Gly observed in idiopathic male patients underscores perturbation of serine-glycine metabolism in PD, consistent with disruption in this pathway detected in untargeted NMR-based metabolomics in the same subgroup. Physiologically, L-Ser is synthesized de novo, primarily from a glycolytic intermediate, and serves multiple essential roles as it is incorporated into membrane phospholipids, contributing to the structure of plasma/organelle membranes and myelin ( Esaki et al., 2015 ; Yoshida et al., 2004 ); provides one-carbon units for nucleic acid synthesis, methylation, and reductive metabolism; acts as a precursor for D-Ser and Gly, which act as co-agonists at NMDARs, modulating excitatory neurotransmission ( Pollegioni et al., 2021 ). Thus, perturbation in serine enantiomer metabolism may impact a broad range of central and peripheral processes implicated in PD. Given the prominent dysregulation in metabolically interconnected amino acids such as D-Ser, L-Ser and Gly observed in males with iPD, our previous study evaluating gene variants in SRR, DAO, DAOA, SHMT1, SHMT2, PHGDH, AMT, GCSH, and GLDC , which encode key enzymes of D-Ser, L-Ser and Gly metabolism, reported a significant association with PD, suggesting that they act as genetic modifiers for PD in specific cohorts stratified by sex and genetic status. In particular, we disclosed that two PHGDH alleles, enriched in idiopathic PD patients, were associated with reduced expression of PHGDH protein, suggesting a role of these variants in the serine dysmetabolism observed in these patients. The extent to which altered serum amino acid levels reflect basal ganglia changes remain uncertain. D-Ser case is particularly noteworthy, given its ability to cross the blood brain barrier (BBB), unlike the other amino acids detected ( Kasai et al., 2011 ; Pernot et al., 2012 ). In contrast to the decreased levels of both serine enantiomers observed in the serum of idiopathic PD male patients, earlier HPLC studies reported elevated D-Ser and L-Ser levels in the post-mortem striatal samples of PD patients and MPTP-lesioned monkeys, as well as in the cerebrospinal fluid of de novo PD patients ( Di Maio et al., 2023 ; Nuzzo et al., 2019 ; Serra et al., 2023 ). Moreover, our recent Ultra Performance Liquid Chromatography-Mass Spectrometry (UPLC-MS) analysis of post-mortem cortex, caudate, and putamen samples from both MPTP-treated monkeys and PD patients revealed that, among the 44 amino acids examined, only the serine enantiomers were consistently upregulated in the striatum of both species (Gervasoni et al., 2025a). The contrast with reduced serum levels observed in our cohort suggests that serum decreases in serine enantiomers are more likely indicative of systemic metabolic dysfunction rather than direct basal ganglia variations. However, it is also worth noting that earlier clinical studies were based on smaller cohorts and lacked sex or subtype stratification, which may account for some of the discrepancies with the more rigorously stratified approach of the present study. Considering the higher incidence of PD in men and that the idiopathic form accounts for over 80% of all cases, our findings have potential clinical implications. Specifically, reduced serum levels of NMDAR-related amino acids in this subgroup may help identify a broad population of patients who could benefit from targeted amino acid interventions ( Araújo et al., 2022 ; Bartl et al., 2022 ; Costa et al., 2023 ). In this respect, oral D-Ser supplementation has shown therapeutic effects in a small cohort of PD patients, improving motor and non-motor symptoms when used as an adjunct to standard PD therapy ( Gelfin et al., 2012 ; Heresco-Levy et al., 2013 ). Although the underlying mechanisms remain to be fully elucidated, recent evidence demonstrated that D-Ser enhances glutamatergic signaling at NMDARs on nigrostriatal dopaminergic neurons, thereby facilitating dopamine release ( Ringlet et al., 2025 ). Our recent preclinical findings also highlight D-Ser therapeutic potential in other neurodegenerative disorders. Specifically, elevated levels of this NMDAR co-agonist were shown to improve neurological symptoms in mouse models of spinal muscular atrophy ( Hassan et al., 2025 ), and to attenuate motor deficits, enhance blood–brain barrier integrity, and reduce neuroinflammation in experimental autoimmune encephalomyelitis (Usiello et al., 2025). Together, these findings support the potential of NMDAR-targeted metabolic strategies — particularly those involving D-Ser — as promising adjunctive therapies in PD and other neurodegenerative disorders. Furthermore, we found that sex significantly influences age-related variations in serum D-Ser levels in healthy individuals. Specifically, healthy women showed an age-dependent increase in both D-Ser levels and D-Ser/total Ser ratio, a trend not observed in men. A similar pattern was observed in women with idiopathic PD. This is in line with previously reported positive correlation between D-Ser and age in an independent cohort of PD women ( Imarisio et al., 2024 ), suggesting that such variation may reflect a physiological pattern rather than a disease-specific alteration. In contrast, women carrying genetic mutations did not display age-related variations in D-Ser, although the underlying reason remains unclear. Notably, female idiopathic PD patients also exhibited a direct correlation between D-Ser levels and age at disease onset, potentially reflecting age-related alterations in the regulation of this endogenous NMDAR-co-agonist. Meta-analysis studies indicate that sex and genetic factors can influence symptom severity in PD ( Russillo et al., 2022 ; Schaffner et al., 2025 ). In our cohort, however, iPD and gPD patients showed comparable motor symptoms severity across sexes. Despite pronounced disruptions in amino acid levels in idiopathic male patients, no significant correlations emerged between amino acid profiles and motor symptoms. These findings suggest that serum amino acid variations capture facets of iPD pathophysiology that are independent of motor symptom severity. In contrast, in the gPD subgroup, D-Ser levels were significantly correlated with MDS-UPDRS III scores, regardless of sex. Notably, TMEM175 mutation carriers displayed a selective correlation between circulating D-Ser levels and motor impairment. However, given the observational nature of this cross-sectional study, no causal inferences can be drawn. Future longitudinal investigations incorporating additional clinical and systemic parameters will be essential to unravel whether additional factors underlie the sex- and subtype-specific amino acid alterations. Additionally, to assess whether metabolic imbalance of NMDAR-related amino acids reflects underlying variations in the main NMDAR subunit genes, we screened polymorphisms in GRIN1 , GRIN2A and GRIN2B within our PD cohort. Consistent with prior evidence of altered NMDAR expression and function in PD (A W Dunah et al., 2000; Gardoni et al., 2012 , 2010 ; Hallett and Standaert, 2004 ; Nuzzo et al., 2019 ; Penney et al., 1996 ; Picconi et al., 2008 ) in the present work we identified allelic variants in GRIN2A and GRIN2B , suggesting a potential genetic contribution to the glutamatergic dysregulations observed in the disorder ( Campanelli et al., 2022 ). To strengthen these findings, we extended the analysis to two large independent case-control cohorts including our internal dataset (MNI-cohort) and the Parkinson Disease Genetic Consortium (PDGC)/UK Biobank cohort. In particular, we identified three variants in GRIN2B (rs11055581, rs7301328 and rs1806201) that were significantly associated with PD, regardless of gender and PD subtype, in the large PDGC/UK dataset. Interestingly, the rs11055581 variant showed association across both the MNI and the PDGC/UK datasets. These results strongly support a role for GRIN2B as a general PD susceptibility gene. Notably, the two synonymous variants rs7301328 and rs1806201 in GRIN2B , are associated with increased GRIN2B expression in key brain regions involved in PD pathophysiology, including the substantia nigra, basal ganglia, amygdala and cortex, as reported in GTEX database ( https://www.gtexportal.org/home/ ). In contrast, the three intronic variants (rs11866570, rs62621078 and rs6497540) identified in GRIN2A showed a subtype-specific distribution, being enriched in either idiopathic or genetic PD subgroups, but not associated with PD in large non-stratified case-control analyses. This suggests that GRIN2A may act as a context-dependent modifier rather than a primary susceptibility gene, in agreement with prior findings ( Nepal et al., 2019 ). Overall, our data indicate that genetic variation in NMDAR subunits, GRIN2A and GRIN2B , may contribute to PD risk differently: while GRIN2B emerges as a potential susceptibility gene with effects independent of sex and PD subtype, GRIN2A may act as a context-dependent modifier influenced by these factors. This divergence mirrors the biochemical asymmetry observed in our study, where serum levels of NMDAR-related amino acids were differentially modulated by sex and PD subtype. Although the direct interplay between these variants and peripheral amino acid profiles remains unclear, these findings support a converging model in which both genetic and metabolic perturbations in the NMDAR pathway influence PD pathophysiology. Future studies in other stratified cohorts will be crucial to validate these associations and elucidate the mechanistic link, if any, between GRIN2A / GRIN2B variants and systemic amino acid disruption in PD. A major strength of our study lies in its integrative design. To our knowledge, this is the most comprehensive investigation to date combining serum quantification of NMDAR-related amino acid, high-resolution genotyping - including a large panel of genetic markers ( Gialluisi et al., 2021 ; Carrillo et al., 2025 ) - and detailed clinical motor phenotyping in a large and well-characterized PD cohort. This multidimensional approach enabled us to delineate metabolic alterations across sex and PD subtypes with unprecedented resolution. However, some limitations must be acknowledged. First, the cross-sectional design of the study precludes causal inference regarding the relationship between amino acid variations and clinical features. Second, the absence of drug-naïve PD patients limits the extrapolation of our biochemical findings to untreated populations. Third, the exclusive inclusion of Caucasian individuals restricts the generalizability of our results to other ethnic backgrounds. Our findings indicate the following key points: 1. Sex and PD subtype significantly influence the balance of circulating NMDAR-related amino acids; 2. Men with idiopathic PD exhibit the most substantial reductions in NMDAR-related amino acids and their precursors, despite having similar LEDD, and motor symptoms severity when compared to subjects with gPD; 3. In women, amino acid levels remain largely unchanged, regardless of the PD subtype; 4. D-Ser levels show a positive correlation with motor impairment severity in both male and female carriers TMEM175 mutations; 5. Variants in the GRIN2B gene are associated with PD regardless of the context , indicating a general susceptibility, while variants in the GRIN2A gene may act as context-dependent modifiers influenced by sex or genetic background. In conclusion our results emphasize the need to consider sex and genetic background as critical variables in future clinical and translational research. This approach may be instrumental in identifying reliable biomarkers and therapeutic targets, ultimately enabling more precise and effective strategies for diagnosis, monitoring and treatment of PD. Material and Methods Study participants PD-cohort 245 independent and unrelated PD patients (129 males and 116 females; 86 familial and 159 sporadic cases) were enrolled in this study. These patients were part of the PD biobank of IRCCS Neuromed/IGB-CNR. The median age at diagnosis was 62.0 years (IQR 55.0-66.0). All the subjects were of European ancestry and were evaluated by qualified neurologists of the Parkinson Centre of the IRCCS INM Neuromed from June 2015 to December 2017, and from June 2021 to December 2023, with a thorough protocol comprising neurological examination and evaluation of non-motor domains. Information about family history, demographic characteristics, anamnesis, and pharmacological therapy was also collected (the treatment of the PD groups consisted for the most part of a combination of levodopa and dopamine agonists) ( Gialluisi et al., 2021 ). Clinical criteria for diagnosis required the presence of at least two cardinal motor signs: asymmetric resting tremor, bradykinesia and rigidity, as well as a good response to levodopa and absence of other atypical features and causes of parkinsonism. Exclusion Criteria for enrolment were: i ) pre-existing psychiatric conditions; ii ) presence of neurodegenerative neurological diseases such as multiple sclerosis, lateral sclerosis amyotrophic, Alzheimer’s, neuromuscular pathologies, epilepsy; iii ) diagnosis of dementia; iv ) depression; v ) prolonged intake of anxiolytics, antidepressants, antipsychotics, hypnotic drugs, cognitive stimulants. The Movement Disorder Society revised version of the Unified Parkinson’s Disease Rating Scale Part III (33 items, maximum score 132; hereafter called UPDRS) ( Goetz et al., 2008 ) was used to assess clinical motor symptoms. These included language, facial expressions, tremor, rigidity, agility in movements, stability, gait and bradykinesia. Patients were analyzed during the ON period. This cohort includes 121 idiopathic and 124 genetic PD patients, carrying at least one pathogenic mutation in the most frequently mutated genes LRRK2 , GBA1 , TMEM175 and PARK2/PINK1/PARK7 (see next paragraph “Study Cohort and stratification factors” for the criteria used for patient selection). This cohort was used both for quantification of NMDAR-related amino acids by HPLC analysis and for genetic association analysis of GRIN1 , GRIN2A and GRIN2B genes. HC cohort 203 neurological controls (80 males and 123 females; median age 61.0 years; IQR 55.0-68.0) were selected for this study. Healthy controls matched for sex and age with PD patients and were negative for mutation/variant in PD genes (see next paragraph “Study Cohort and stratification factors” for the criteria used for patient selection). This HC cohort was used for both HPLC analysis and for genetic association analysis of NMDAR genes. The project was approved by the ethical committees of IRCCS Neuromed and written informed consent was signed by all the participants. All procedures involving human participants were approved by the Institutional Review Board of the IRCCS Neuromed Italy. The study protocols N°9/2015, N°19/2020, N°4/2023 have been registered in clinicaltrial.gov with the numbers NCT02403765 , NCT04620980 , NCT05721911 . Clinical investigations were conducted according to the principles expressed in the Declaration of Helsinki. Written informed consent was obtained from all participants. The research was carried out following the recommendations set out in the Global Code of Conduct for Research in Resource-Poor Settings. Study Cohort and stratification factors Stratification factors for PD include age of onset (early-vs late-onset PD), the presence or absence of family history (familial vs sporadic) and the presence of pathogenic mutations (monogenic vs idiopathic). Here, we classified patients based on their genetic background as “idiopathic-PD” and “genetic PD”. We choose not to use the classification “familial PD” or “sporadic PD” since this classification does not always reflect the presence/absence of mutation/variant. Indeed, in our cohort a consistent number of patients (41.1 %) mutated in LRRK2 were classified as “sporadic” at interview, and similar percentages (ranging from 50 to 65 %) emerged for patients carrying single pathogenic mutation in GBA1, PARK2, PINK1, PARK7 , and TMEM175 . This could depend on several factors, such as late age at the onset of PD, the presence of small nuclear families, and the reduced penetrance of PD mutations. Whole Exome Sequencing (WES) data were used for a comprehensive identification of the genetic background of the study cohort, including PD patients and HC. Considering the extreme genetic heterogeneity of PD, thirty-seven PD candidate genes were extracted from WES data. The selected genes include those reported in the literature as Mendelian PD genes ( PARK7 / DJ-1, DNAJC13, DNAJC6, EIF4G1, FBXO7, LRRK2, PARK2, PINK1, SNCA ) and those reported as PD genes/at risk factors ( AIMP2, ANKK1, ANKRD50, CHMP1A, GBA1, GIGYF2, GIPC1, GRK5, HMOX2, HSPA8, HTRA2, IMMT, KIF21B, KIF24, MAN2C1, PACSIN1, RHOT2, SLC25A39, SLC6A3, SLC6A3, SNCAIP, SPTBN1, TMEM175, TOMM22, TVP23A, UCHL1, VPS8, ZSCAN21 ) (AbouLSleiman et al., 2006; Djarmati et al., 2006 ; Gialluisi et al., 2021 ; Klein et al., 2005 ; Martin et al., 2011 ; Nuytemans et al., 2010 ; Palomba et al., 2023 ; Steinmetz et al., 2024 ; Sun et al., 2006 ; Verstraeten et al., 2015 ; Carrillo et al 2025 ). The cohort (both PD patients and HC) was analyzed for the presence of mutations (as reported in the literature or public mutation databases) or rare (Minor allele frequency (MAF) < 0.001) and highly deleterious exonic variants (including non-synonymous, splicing and indels; Combined Annotation Dependent Depletion (CADD) score ≥15) in the panel of the selected PD genes. We selected 121 PD patients that we named “idiopathic”, in which no mutation/variant was identified in the selected PD genes. The second group, that we named “genetic PD”, includes 124 PD-affected subjects carrying of at least 1 pathogenic mutation in one of the most frequently mutated PD genes, which includes 17 patients mutated in LRRK2 , 30 mutated in GBA1 , 40 with at least 1 mutation in PARK2/PINK1/PARK7 , 33 mutated in TMEM175, and four patients carrying two mutations in PD genes such as GBA1/TMEM175 , GBA1/PARK2 , GBA1/PINK1 and TMEM175/PARK2 , respectively. No pathogenic mutation was identified in SNCA , VPS35 , DNAJC13, DNAJC6, EIF4G1, FBXO7 and UCHL1 , in our cohort, by surveying the public mutations databases (LOVD - An Open Source DNA variation database system ( https://lovd.nl/ ) ( Suppl. Table 1 and Materials and Methods for the detailed list of mutations/variants). In particular, the seventeen LRRK2 mutated patients were carriers of the p.G2019S (14 patients), p.R1441C (2 patients) and p.A419V (1 patient) mutations. Thirty-three GBA1 mutated patients were carriers of the following pathogenic mutations: p.N409S (9 patients), p.E365K (7 patients), p.T408M (3 patients), p.L483P (2 patients), p.H294Q (2 patients) and p.R17G, p.D66E, p.V230E, p.P240S, p.H367Y, p.S403T, p.E427K, p.D448H, p.V499L, p.K505N (1 patient each). Forty-two PD patients were carriers of at least one pathogenic mutation in one of the recessive PD genes PARK2 , PINK1 , PARK7. 18 out of 42 PD patients were mutated in PARK2 including one homozygous patient (p.W453X/ p.W453X), 1 compound heterozygous (p.R275W/exon deletion) and 16 carrying a single heterozygous mutation such as p.R402C (4 patients), p.A82E (5 patients), and p.M192L, p.R234Q, p.T240M, p.V244L, p.R275W, p.E409X, p.T415N, p.W453X (1 patient each). Nine patients were carriers of a single heterozygous mutation in PINK1 including p.P196L (2 patients), p.W437X (2 patients), p.A168P, p.K186N, p.A291D, p.R326C, p.D525N (1 patient each). Sixteen patients were carriers of a single heterozygous variant in PARK7 of which 14 carrying the p.R98Q variant and 2 carrying the splicing variant c.252+2->A. Thirty-five PD patients were carriers of one pathogenic mutation in TMEM175 (as reported in our previous study ( Palomba et al., 2023 ) which includes p.V147Dfs*104 (4 patients), p.A429Qfs*120 (3 patients), p.A149Gfs*97 (3 patients), p.R35C (3 patients), p.L405V (4 patients), p.R414W (3 patients), p.R335H (3 patients), p.T105A (2 patients), p.I78T (2 patients), and p.R260C, p.A270T, p.I280M, p.P286L, p.A326V, p.S348L, p.A424V, p.R481W (1 patient each) (Suppl. Table 1). The healthy subjects (HC) that we included in the study were negative for mutation/variant in the selected panel of PD genes. Collection and storage of serum samples Blood sampling was performed after a 6-h fasting. Whole blood was collected by peripheral venipuncture into clot activator tubes and gently mixed. Sample was stored upright for 30 min at room temperature to allow blood to clot and centrifuged at 2000 ×g for 10 min at room temperature. Serum was aliquoted (0.5 ml) in polypropylene cryotubes and stored at −80 °C before usage. Unique anonymized codes have been assigned to the samples for processing and subsequent analysis, maintaining the confidentiality of personal data. HPLC analysis of amino acids content Serum samples (100 μl) were mixed in a 1:10 dilution with HPLC-grade methanol (900 μl) and centrifuged at 13,000 ×g for 10 min; supernatants were dried and then suspended in 0.2 M trichloroacetic acid (TCA). TCA supernatants were then neutralized with 0.2 M NaOH and subjected to precolumn derivatization with o-phthaldialdehyde /N-acetyl-L-cysteine in 50% methanol. Amino acids derivatives were resolved on a UHPLC Nexera X3 system (Shimadzu) by using a Shimpack GIST C18 3-μm reversed-phase column (Shimadzu, 4.0 × 150 mm) under isocratic conditions (0.1 M sodium acetate buffer, pH 6.2, 1% tetrahydrofuran, and 1 ml/min flow rate). A washing step in 0.1 M sodium acetate buffer, 3% tetrahydrofuran and 47% acetonitrile, was performed after every run. Identification and quantification of amino acids were based on retention times and peak areas, compared with those associated with external standards. The detected amino acids concentration was expressed as μM. Statistical analysis Clinical and demographic characteristics were described as median and the interquartile range (IQR) or absolute frequencies. Comparisons between PD patients and HC were evaluated using Mann Whitney U or Kruskal-Wallis test for continuous variables and Chi-Square test for dichotomous variables. Comparison of serum amino acid levels between PD and HC groups was first performed using a Mann Whitney U or Kruskal-Wallis test followed by Dunn’s multiple comparisons post hoc, when required. Second, an ANCOVA model with “diagnosis” as between factors and age, sex and LEDD as covariate was used to control for the effect of these factors on serum amino acid levels. For sex stratified analysis, age- and LEDD-adjusted ANCOVA model was used. The correlation of serum amino acid concentration with age, age at PD onset, disease duration, LEDD and MDS-UPDRS III was evaluated with Spearman’s correlation test. Partial correlation analyses adjusted for the effect of potential confounders (age, disease duration, LEDD) were adopted to confirm the correlation between serum amino acid levels and PD clinical features (disease duration, LEDD and MDS-UPDRS III). Significance was set at p < 0.05 for all analyses. Data were analysed by using SPSS 26.0 software (IBM, Armonk, NY, USA). Study cohorts used for genetic analysis MNI-PD cohort The Mediterranean Neurological Institute (MNI)-PD cohort included 804 independent and unrelated PD patients (501 males; 300 familiar and 504 sporadic cases), for which Whole exome sequencing (WES) data are available. This cohort is part of the Parkinson’s disease Biobank of the IRCCS Neuromed and of the Institute of Genetics and Biophysics (CNR). All the subjects were of European ancestry and were evaluated by qualified neurologists of the Parkinson Centre of the IRCCS INM Neuromed from June 2015 to December 2017, and from June 2021 to December 2023, with a thorough protocol comprising neurological examination and evaluation of non-motor domains. Information about family history, demographic characteristics, anamnesis, and pharmacological therapy was also collected (the treatment of the PD groups consisted for the most part of a combination of levodopa and dopamine agonists) ( Gialluisi et al., 2021 ). This cohort was used to identify susceptibility or protective factors for PD. MNI-PD stratified cohort This cohort was selected from the entire MNI-PD cohort of 804 PD patients with the same criteria used to select the PD patients analyzed by HPLC and includes 193 idiopathic and 227 genetic PD patients carrying at least one pathogenic mutation in the most frequently mutated genes LRRK2 , GBA1 , TMEM175 and PARK2/PINK1/PARK7 (see the paragraph “Study Cohort and stratification factors” for the criteria used for patient selection). This cohort was used as validation cohort in genetic association analysis to confirm the association with PD in the cohort stratified by gender and genetic background. MNI-HC cohort 282 neurological healthy controls (HC) were recruited by the same group of neurologists, among the patients’ wives/husbands, after having ascertained the lack of neurological pathologies and the absence of affected family members. WES data were available for the entire cohort. The MNI-HC cohort was used for association analysis. Replication cohort 4586 PD patients (from PDGC cohort) and 43989 CNT (from UK biobank) whose data were downloaded from the PDGC Variant browser ( https://pdgenetics.shinyapps.io/VariantBrowser/ ). Associations analysis Principal Component Analysis (PCA) performed with Plink software was used to characterize the genetic diversity of the study sample (PD_MNI, CNT_MNI) ( Suppl . Fig. 6a ) ( Chang et al., 2015 ). The analysis was carried out by using common variants (Minor allele frequency MAF > 0.01), PC1 and PC2 were found to contribute to a variance of 25% among samples (Suppl. Fig. 6b ). To investigate the genetic contribution of rare variants MAF was set to: MAF < 0.01, MAF<0.005 and MAF< 0.001 on the phenotype, gene-based analyses were carried out using the unified Optimal Sequence Kernel Association Test (SKAT-O) of the R SKAT package ( http://cran.nexr.com/web/packages/SKAT/index.html ), age was used as covariate. To identify the genetic contribution given by common variants (MAF > 0.01) we adopted a logistic regression model trough plink2 software, by adjusting for age and the 10 principal components; we also adjusted for sex when we analyzed the entire cohort regardless of gender. p-values were adjusted for Bonferroni multiple testing correction. Expression studies in human brain regions The Genotype Tissue Expression (GTEx) portal ( https://www.gtexportal.org/home/ ) was accessed to obtain gene expression data of the identified SNPs. The analysis was performed on all available adult human brain regions (amygdala, anterior cingulate cortex BA24, caudate nucleus, putamen, substantia nigra, cerebellar hemispheres, cerebellum, cerebral cortex, frontal cortex BA9, hippocampus, hypothalamus, nucleus accumbens). Data Availability All data produced in the present work are contained in the manuscript Conflict of Interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be considered as a potential conflict of interest. Author contributions IY: Investigation, review & editing; FC: data analysis, review & editing; TN: data analysis, review & editing; ADM: Investigation, review & editing; NM: review & editing, SP: review & editing; FE: Writing – review & editing; TE; Funding acquisition, Project administration, Supervision, Writing – review & editing. AU: Conceptualization, Funding acquisition, Project administration, Supervision, Writing – review & editing. Acknowledgments This study was partially funded by Italian Ministry of University and Research (PRIN 2022 - COD. 2022XF7YYL_02 to AU and PRIN 2022 – COD. 2022W3RKLJ to TE). The work of A.U., T.N. and T.E. was supported by NEXTGENERATIONEU (NGEU) and funded by the Ministry of University and Research (MUR), National Recovery and Resilience Plan (NRRP), project MNESYS (PE0000006) – A Multiscale integrated approach to the study of the nervous system in health and disease (DN. 1553 11.10.2022). The work of T.E. was supported by Next Generation EU - PNRR M6C2 Investimento 2.1 valorizzazione e potenziamento della ricerca biomedica del SSN grant n. PNRR-MAD-2022-12375960 and grant n. PNRR-MCNT2-2023-12377375. TE was also supported by Ministry of Health, Ricerca Corrente. The authors are grateful to all the patients, their caregivers, the Clinical Parkinson’s Disease Center of IRCCS Pozzilli and the PD biobank of IRCCS Neuromed and IGB-CNR for the kind cooperation with this study. References Abou-Sleiman PM , Muqit MMK , McDonald NQ , Yang YX , Gandhi S , Healy DG , et al. A heterozygous effect for PINK1 mutations in Parkinson’s disease? 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Share Differences in sex and genetic status affect the disruption of NMDAR-related amino acid homeostasis in Parkinson’s disease Isar Yahyavi , Federica Carrillo , Tommaso Nuzzo , Anna Di Maio , Sara Pietracupa , Nicola Modugno , Francesco Errico , Teresa Esposito , Alessandro Usiello medRxiv 2025.08.08.25333287; doi: https://doi.org/10.1101/2025.08.08.25333287 Share This Article: Copy Citation Tools Differences in sex and genetic status affect the disruption of NMDAR-related amino acid homeostasis in Parkinson’s disease Isar Yahyavi , Federica Carrillo , Tommaso Nuzzo , Anna Di Maio , Sara Pietracupa , Nicola Modugno , Francesco Errico , Teresa Esposito , Alessandro Usiello medRxiv 2025.08.08.25333287; doi: https://doi.org/10.1101/2025.08.08.25333287 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Neurology Subject Areas All Articles Addiction Medicine (568) Allergy and Immunology (863) Anesthesia (300) Cardiovascular Medicine (4435) Dentistry and Oral Medicine (444) Dermatology (382) Emergency Medicine (608) Endocrinology (including Diabetes Mellitus and Metabolic Disease) (1509) Epidemiology (15229) Forensic Medicine (30) Gastroenterology (1124) Genetic and Genomic Medicine (6600) Geriatric Medicine (668) Health Economics (997) Health Informatics (4536) Health Policy (1368) Health Systems and Quality Improvement (1613) Hematology (541) HIV/AIDS (1264) Infectious Diseases (except HIV/AIDS) (15916) Intensive Care and Critical Care Medicine (1103) Medical Education (623) Medical Ethics (146) Nephrology (667) Neurology (6599) Nursing (346) Nutrition (998) Obstetrics and Gynecology (1144) Occupational and Environmental Health (957) Oncology (3332) Ophthalmology (974) Orthopedics (369) Otolaryngology (420) Pain Medicine (436) Palliative Medicine (130) Pathology (663) Pediatrics (1693) Pharmacology and Therapeutics (691) Primary Care Research (711) Psychiatry and Clinical Psychology (5447) Public and Global Health (9232) Radiology and Imaging (2198) Rehabilitation Medicine and Physical Therapy (1370) Respiratory Medicine (1196) Rheumatology (593) Sexual and Reproductive Health (712) Sports Medicine (530) Surgery (712) Toxicology (99) Transplantation (289) Urology (265) (function(){function c(){var b=a.contentDocument||a.contentWindow.document;if(b){var d=b.createElement('script');d.innerHTML="window.__CF$cv$params={r:'a00bbb251df4c13d',t:'MTc3OTYxOTczOA=='};var a=document.createElement('script');a.src='/cdn-cgi/challenge-platform/scripts/jsd/main.js';document.getElementsByTagName('head')[0].appendChild(a);";b.getElementsByTagName('head')[0].appendChild(d)}}if(document.body){var a=document.createElement('iframe');a.height=1;a.width=1;a.style.position='absolute';a.style.top=0;a.style.left=0;a.style.border='none';a.style.visibility='hidden';document.body.appendChild(a);if('loading'!==document.readyState)c();else if(window.addEventListener)document.addEventListener('DOMContentLoaded',c);else{var e=document.onreadystatechange||function(){};document.onreadystatechange=function(b){e(b);'loading'!==document.readyState&&(document.onreadystatechange=e,c())}}}})();