Peripheral immune profile and neutrophil-to-lymphocyte ratio in progressive supranuclear palsy: Case–control study and meta-analysis

Eur J Neurol. 2024;31:e16451.   | 1 of 9 https://doi.org/10.1111/ene.16451 wileyonlinelibrary.com/journal/ene Received:29June2024 | Accepted:5August2024 DOI: 10.1111/ene.16451 ORIGINAL ARTICLE Peripheral immune profile and neutrophil- to- lymphocyte ratio in progressive supranuclear palsy: Case–control study and metaanalysis Laura Muñoz- Delgado1,2 | Antonio Luque- Ambrosiani1 | Belén Benítez Zamora1 | Daniel Macías- García1,2 | Silvia Jesús1,2 | Astrid Adarmes- Gómez1,2 | Elena Ojeda- Lepe1,2 | Fátima Carrillo1,2,3 | Pablo Mir1,2,3 This is an open access article under the terms of the CreativeCommonsAttribution-NonCommercial-NoDerivs License, which permits use and distribution in anymedium,providedtheoriginalworkisproperlycited,theuseisnon-commercialandnomodificationsoradaptationsaremade. ©2024TheAuthor(s).European Journal of NeurologypublishedbyJohnWiley&SonsLtdonbehalfofEuropeanAcademyofNeurology. 1Unidad de Trastornos del Movimiento, ServiciodeNeurología,Instituto de Biomedicina de Sevilla, Hospital UniversitarioVirgendelRocío/CSIC/ Universidad de Sevilla, Seville, Spain 2Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas(CIBERNED), Instituto de Salud Carlos III, Madrid, Spain 3Departamento de Medicina, Facultad de Medicina, Universidad de Sevilla, Seville, Spain Correspondence Pablo Mir, Unidad de Trastornos del Movimiento, Instituto de Biomedicina de Sevilla, Hospital Universitario Virgen del Rocío,Avda.ManuelSiurots/n,Seville 41013, Spain. Email: [email protected] Funding information Instituto de Salud Carlos III, Grant/ AwardNumber:PI16/01575,PI18/01898, PI19/01576andPI20/00613;Consejería deEconomía,Innovación,Cienciay Empleo,JuntadeAndalucía,Grant/Award Number:CVI-02526andCTS-7685; ConsejeríadeSaludyConsumo,Junta deAndalucía,Grant/AwardNumber: NEU-0005-2022;MinisteriodeCiencia eInnovación,Grant/AwardNumber: RTC2019-007150-1;ConsejeríadeSaludy Familia,JuntadeAndalucía,Grant/Award N u m b e r :  P I - 0 4 7 1 - 2 0 1 3 ,  P E - 0 2 1 0 - 2 0 1 8 ,  P I - 0 4 5 9 - 2 0 1 8  a n d  P E - 0 1 8 6 - 2 0 1 9 ;  ConsejeríadeTransformaciónEconómica, Industria, Conocimiento y Universidades, Grant/AwardNumber:PY20_00896 Abstract Background and purpose: Peripheral inflammation is probably involved in the pathogenesisofprogressivesupranuclearpalsy(PSP)anditmaybeacommonfeaturewith Parkinson'sdisease(PD).TheperipheralimmuneprofileinPSPremainsunclear,aswell as whether the inflammatory pathways differ from those in PD. The neutrophil-to- lymphocyteratio(NLR)hasbeenproventobeawell-establishedbiomarkerofsystemic inflammation. This study aimed to evaluate the peripheral immune profile in PSP compared with PD. Methods: Across-sectionalstudywasconductedincludingpatientswithPSPandPD andhealthycontrols(HCs).LeukocytesubpopulationsandtheNLRweremeasuredinperipheral blood. Multivariate linear regression and post hoc tests were applied. Electronic databasesweresearchedinNovember2023toperformmeta-analysestoclarifytheperipheral immune profile in PSP. Results: Ourcohortincluded121patientswithPSP,127patientswithPDand266HCs. TheNLRwashigherinPSPandPDcomparedwithHCs.PSPhadahigherneutrophil count compared with HCs. Whilst a lower lymphocyte count was found in PD compared withHCs,thelymphocytecountdidnotdifferbetweenPSPandHCs.Themeta-analyses supported this immune profile. Conclusions: PSPandPDshowanincreasedperipheralinflammationandahigherNLR compared with HCs. Different pathogenic inflammatory mechanisms are probably involved in PSP and PD, since in PSP this altered peripheral immune profile is mainly driven by neutrophils. Understanding the neutrophils' role in PSP may allow for the development of targeted therapies. KEYWORDS inflammation,neutrophil,neutrophil-to-lymphocyteratio,progressivesupranuclearpalsy 2 of 9 | MUÑOZ-DELGADO et al. INTRODUCTION Progressivesupranuclearpalsy(PSP)isarareadult-onsetneurodegenerative disease of unknown aetiology. Clinically, PSP is characterized by progressive gait instability, supranuclear gaze abnormalities, levodopa-unresponsive parkinsonism and frontal cognitive disturbances [1, 2]. Definitive diagnosis requires postmortem examination; however, current diagnostic criteria based on clinical manifestations enable sensitive and reliable clinical diagnosis of probable PSP and possible PSP [1, 2]. Höglinger and collaborators distinguished eight different phenotypes, although PSP with Richardson's syndrome (PSP-RS)andPSPwithpredominantparkinsonism(PSP-P)arethe most common, constituting 80%–90% of all PSP cases. Currently therearenoeffectivesymptomaticordisease-modifyingtreatments available, probably because the aetiopathogenesis of the disease has not yet been elucidated. Neuropathologically,PSPisconsidereda‘tauopathy’,characterizedbythepresenceofabnormalphosphorylatedmicrotubule- associated protein tau aggregates as neurofibrillary tangles in both neurons and glia in specific cortical and subcortical regions [3]. Very little is known about the aetiology and pathogenesis of PSP, although genetic and epigenetic factors, oxidative stress and inflammation are thought to contribute to their development and progression [3]. Even though it remains unclear whether neuroinflammation is the cause or rather an effect of pathological tau accumulation and neuronal loss, the presence of this process is definite [4]. Microglial activation has been demonstrated in the brain of patients with PSP in in vivo and postmortem studies [5–7]. Microglia activation has been shown to correlate with tau burden and cell loss, at least in some areas. Different patterns of cytokine expression have been identified in the brains of PSP patients compared tohealthycontrols(HCs),andtheyhavealsobeensuggestedtobe related to neurodegeneration [4,8].Besides,apro-inflammatory cytokine profile has also been reported in the cerebrospinal fluid (CSF)[9, 10] and serum [10] of patients with PSP. In this line, current evidence suggests an involvement of peripheral inflammation in the pathogenesis of PSP and suggests that it is a common featurewithParkinson'sdisease(PD).However,previousstudieson leukocytes and subpopulations in peripheral blood in PSP have shown controversial results [10–12].Inthiscontext,theneutrophil- to-lymphocyteratio(NLR)inperipheralbloodhasbeenprovento beawell-establishedbiomarkerofsystemicinflammationinother neurodegenerative diseases such as PD [13]. In PSP, the role of the NLRisnotclear[10–12], nor is it clear whether it differs from PD [11].PreviousstudieshaveshownthatdefiningtheroleoftheNLR in PSP is probably affected by obstacles such as small numbers of patients in the studies and overlapping clinical manifestations. Besides, little is known about the role of lymphocytes and neutrophils in this scenario. Hence, the peripheral immune profile in PSP has not been clearly defined, nor whether the inflammatory pathways differ significantly from those in PD. Therefore, this study aimed to evaluate whether there was a peripheral inflammation in patients with PSP. Thus, the peripheral immuneprofilewasfirstinvestigated,measuredbytheNLR,lymphocytes and neutrophils, in patients with PSP compared with patientswithPDandHCsthroughacross-sectionalstudyinourcohort.Subsequently,asystematicreviewandameta-analysiswere performed to clarify the peripheral immune profile in PSP. PATIENTS AND METHODS Observational case–control study Participants and clinical assessment In all, 121 patients with PSP were included from the Movement Disorders Clinic at the Hospital Universitario Virgen del Rocio in Seville(Spain),recruitedfrom1999to2023.Allthemedicalrecords were carefully reviewed and patients with PSP were diagnosed following the currently accepted Movement Disorder Society Clinical Diagnostic Criteria [1, 2]. Also 127 patients with sporadic PD diagnosed following the Movement Disorder Society Clinical Diagnostic Criteria [14] were included,and266HCswhowerenon-bloodrelativesofpatientsfrom the same geographical area. Allsubjectsunderwentaclinicalassessmentatourcentreand the demographic and clinical data were retrospectively obtained by consultingtheirmedicalrecords.Allparticipantswereexaminedfor exclusion criteria that could influence the immune profile at the time of the blood extraction or any other relevant neurological disease. Therefore, participants with cancer, active or chronic autoimmune, inflammatoryorinfectiousdiseases(includingthosewithleukocyte count >11.000 cells/μL),recent vaccination(<3 months)and those receiving immunosuppressive therapy were not considered for the study. See the study flowchart, exclusion criteria and resultant PSP cohort in Figure S1. Total leukocyte count and subpopulations (neutrophils and lymphocytes) in peripheral blood were measured in the Central LaboratoryofourcentreusingaSysmexXNautomatedhaematology analyser. It uses fluorescence flow cytometry with forward- scattered, side-scattered and side-fluorescence lights for cell determination.TheNLRwascalculatedasabsoluteneutrophilcount divided by absolute lymphocyte count. Statistical analysis Continuous and categorical variables were compared using analysis ofcovariance(ANCOVA)orWelch'stwo-samplet tests and Fisher's orthechi-squaredtest,asappropriate.Correlationanalyseswereassessed by Pearson's or Spearman's correlation, as appropriate according to data distribution. Multivariate linear regression analyses and posthoctestswereappliedtodeterminethedifferencesintheNLR, lymphocytesandneutrophilsamongstgroups.Sex-stratifiedanalyses werealsoperformed.Allstatisticalanalyseswereperformedusingthe 14681331, 2024, 12, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/ene.16451 by Readcube (Labtiva Inc.), Wiley Online Library on [14/04/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License | 3 of 9 PERIPHERAL INFLAMMATION IN PROGRESSIVE SUPRANUCLEAR PALSY Rversion3.5.1andGraphPadPrism8(GraphPadSoftware).Ap value <0.05 was considered statistically significant. Metaanalysis This meta-analysis was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA)statement[15]. Search strategy and eligibility criteria A comprehensive literature search was conducted in electronic scientificdatabases(PubMed,Scopus,Embase,WebofScienceand Cochrane)inNovember2023toidentifyrelevantcase–controlstudies ontheassociationbetweentheNLR,lymphocytesand/orneutrophils andPSP.Thefollowingsearchstringwasused:‘PSP’OR‘progressive supranuclearpalsy’AND‘neutrophil’OR‘lymphocyte’OR‘neutrophil lymphocyte ratio’ OR ‘NLR’. Articles were included if the following inclusion criteria were met: (1) case–control studies describing the associationbetweentheNLR,lymphocytesorneutrophils,andPSP; (2)studieshadanHCgroup;and(3)studieshadtoincludebothmeans andstandarddeviationsorsufficientstatisticalinformation(t, F, χ, Z scores, or p values). No restrictions were applied regarding sample size, language or time of publication. To assess additional studies not indexed in common databases, all retrieved references of the enrolled studies and recent published review articles were also checked. Data extraction Two investigators (LMD and ALA) independently extracted the data. Discrepancies were resolved by a third investigator (DMG). The following data were collected: (1) general information—first author, publication year,study design and cohort selected;(2)individualcharacteristics—samplesize,age,sex,medicaldiseasesand dopaminergictreatment;and(3)meanandstandarddeviationofthe NLR,thelymphocyteandtheneutrophilcountsinperipheralblood. The corresponding author of the studies was contacted for further information when required. Statistical analysis The‘metaphor’packageinRwasused.Effectsizeswerecalculated asstandardizedmeandifferencesofNLRbetweenpatientswithPSP and HCs using the Hedges' g statistic, which provides an unbiased effectsizeadjustedforsamplesize.Between-studyheterogeneity was assessed using the Cochrane Q test and I2 statistic. When heterogeneitywassignificant(p < 0.05orI2 >50%),therandomeffects model was used to evaluate the overall effect size; otherwise, the fixed effects model was used. Publication bias was evaluated by a funnel plot and Egger's test. Sensitivity analyses were undertaken to assess the influence of individual studies on the overall effect estimate. RESULTS Observational case–control study The demographic and clinical characteristics of the different groups (PSP,PDandHCs)fromourcohortareshowninTable 1. Patients withPSPwereolderthanpatientswithPDandHCs(p < 0.001).In patients with PSP there was a similar male/female ratio, in contrast tothemalepredominanceinthePDgroup(63.78%).Atthetimeof blood extraction, patients with PSP had a mean disease duration of Patient with PSP (n = 121) Patients with PD (n = 127) HCs (n = 266) p value Age(years)mean ± SD 70.95 ± 7.52 64.73 ± 10.61 62.76 ± 11.96 <0.001a Sex(male/female) 60/61 81/46 135/131 <0.03b Diseaseduration(years), mean ± SD 3.5 ± 2.27 5.04 ± 3.14 –<0.001a Ageofonset(years),mean ± SD 67.45 ± 7.64 59.69 ± 10.10 –<0.001a Neutrophils(×103 cells/μL), mean ± SD 4.64 ± 1.44 4.32 ± 1.27 3.94 ± 1.21 <0.001c Lymphocytes(×103 cells/μL), mean ± SD 2.05 ± 0.69 1.81 ± 0.59 2.12 ± 0.64 <0.001c NLR,mean ± SD 2.50 ± 1.16 2.59 ± 1.03 2.01 ± 0.91 <0.001c Abbreviations:HCs,healthycontrols;n,totalnumberofsubjects;NLR,neutrophil-to-lymphocyte ratio; PD, Parkinson's disease; PSP, progressive supranuclear palsy; SD, standard deviation. aBasedonlinearregressionandanalysisofcovariance(ANCOVA). bBased on χ2 test. cBased on multivariate linear regression, adjusting for age and sex. TABLE 1 Clinicalanddemographic features as well as peripheral immune profile of patients with PSP, patients with PD and HCs from our local cohort. 14681331, 2024, 12, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/ene.16451 by Readcube (Labtiva Inc.), Wiley Online Library on [14/04/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 4 of 9 | MUÑOZ-DELGADO et al. 3.5 ± 2.27 yearsandameanlevodopaequivalentdailydose(LEDD) of352.03 ± 376.85 mg(range0–1614). ThePSPcohortincluded94patientswithprobablePSP-RS,15 patientswithprobablePSP-P,fivepatientswithprobablePSPwith predominantfrontalpresentation(PSP-F),twopatientswithprobablePSPwithprogressivegaitfreezing(PSP-PFG),twopatientswith possiblePSPwithpredominantspeech/languagedisorder(PSP-SL) and three patients that could not be classified into PSP-RS and PSP-Pduetoinsufficientdatafromdiseaseonset. When the leukocyte subpopulations amongst the different groups(PSP,PDandHC)werecompared,statisticallysignificantdifferencesintheNLR(F[4,509] = 12.83,p < 0.001),aswellasinthe neutrophil (F[4,509] = 9.25, p < 0.001) and the lymphocyte counts (F[4,509] = 6.48,p < 0.001),werefoundafteradjustingforsexand age (Table 1). Multiple comparisons were subsequently accomplished to determine the differences in the peripheral immune responsebetweengroups(Figure 1a–c).Thecompleteresultsofpost hoc tests are described in Tables S1–S3. Inourcohort,theNLRwassignificantlyhigherinbothpatientswith PSPandpatientswithPDcomparedwithHCs(p < 0.001).Nosignificant differenceswerefoundintheNLRbetweenPSPandPD(p = 0.41). Patients with PSP had a significantly higher neutrophil count comparedwithHCs(p < 0.001).Ahigherneutrophilcountwasalso found in patients with PD compared to HCs but it did not reach statisticalsignificance(p = 0.06).Interestingly,whilstasignificantlower lymphocyte count was found in patients with PD compared with HCs(p = 0.001),thelymphocytecountdidnotdifferbetweenpatientswithPSPandHCs(p = 0.97). In patients with PSP, no correlation was found between disease durationandtheNLR(p = 0.55),theneutrophilcount(p = 0.54)or thelymphocytecount(p = 0.10)(Table S4).Besides,nosignificant correlationwasfoundbetweentheNLRorthelymphocytecount andthedopaminergictreatment ofpatientswithPSP, butanon- significant weak correlation was found between the neutrophil count andtheLEDDinpatientswithPSP(ρ = 0.177,p = 0.05)(Table S5). In addition, sex-stratifiedanalyseswereperformed to explore whether the peripheral immune profile could be influenced by sex. Detailed features of the male and female subcohorts are shown in Table S6. In both the male and female cohorts, there were differencesintheNLR,theneutrophilandthelymphocytecountbetween patients with PSP and patients with PD compared with HCs, and afteradjustingforage.Aftermultiplecomparisonanalyses,itwas demonstratedthattheNLRwashigherinmaleandfemalepatients with PSP and patients with PD compared with HCs, and without differences between PSP and PD. In male and female patients with PSP, a higher neutrophil count was observed compared to HCs, but the lymphocyte count did not differ between patients with PSP and HCs (Tables S7–S12). Finally, the aim was to explore the peripheral immune profile amongst the different PSP phenotypes. The detailed features of each PSP phenotype can be found in Table S13.Analyseswerefirst performed in a homogeneous subcohort including only patients withPSP-RS,andtheresultswerethesameasthoseobtainedinthe totalPSPcohort(Tables S14–S17).TheNLRwassignificantlyhigher in patients with PSP-RS and PD compared with HCs (p < 0.005). PatientswithPSP-RShadahigherneutrophilcountcomparedwith HCs(p < 0.001).Again,thelymphocytecountdidnotdifferbetween patientswithPSP-RSandHCs(p = 0.97).Finally,nosignificantdifferencesineithertheRNL(F[3,105] = 1.395,p = 0.25),theneutrophil (F[3,105] = 0.359,p = 0.78)orthelymphocytecount(F[3,105] = 1.523, FIGURE 1 Peripheralinflammatoryimmuneprofileamongstpatientswithprogressivesupranuclearpalsy,patientswithParkinson's diseaseandhealthycontrolsofthelocalcohort.Multiplecomparisonsof(a)neutrophil-to-lymphocyteratio(NLR),(b)neutrophilsand(c) lymphocytesamongstpatientswithprogressivesupranuclearpalsy(PSP),patientswithParkinson'sdisease(PD)andhealthycontrols(HCs). Numbersshownasmeans± SD with multivariate linear regression and post hoc testing for multiple comparisons as appropriate. *p < 0.05. 14681331, 2024, 12, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/ene.16451 by Readcube (Labtiva Inc.), Wiley Online Library on [14/04/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License | 5 of 9 PERIPHERAL INFLAMMATION IN PROGRESSIVE SUPRANUCLEAR PALSY p = 0.22)werefoundbetweenpatientswithPSP-RS(n = 94)andpatientswithPSP-P(n = 15). Metaanalyses A systematic review and three different meta-analyses were conducted to assess the differences between patients with PSP and HCs on the NLR, lymphocytes and neutrophils, respectively (PRISMAflowchart,Figure 2).Afterapplyinginclusionandexclusion criteria,threearticleswithacase–controldesignwereselectedfor theNLRmeta-analysis[10–12].Alongwithourstudy,fourNLRmean differences with a case–control designwere included, comprising 189patientswithPSPand350HCs.However,onlytwoarticles[10, 11]wereselectedfortheneutrophilandlymphocytemeta-analyses as there was no information about these leukocyte subpopulations inthestudyofAlsteretal.So,alongwithourstudy,thelasttwo meta-analysescomprisedthreecase–controlstudiesinvolving166 patientswithPSPand318HCs. Overallheterogeneitywashighinthethreemeta-analysesconducted; hence the random effects model was applied. The standardizedmeandifference(SMD)foreachindividualstudyandtheoverall effect size are shown in Figures 3–5. Patients with PSP had a higher NLR compared with HCs (SMD = 0.45; 95% confidence interval [CI] −0.07–0.98) (Figure 3). Whilst patients with PSP had a higher neutrophil count compared with HCs (SMD = 0.44; 95% CI −0.16–0.76) (Figure 4), no differences were found in the lymphocyte count between PSP and HCs (SMD = 0.01;95%CI−1.18–1.20)(Figure 5). No significant risk of publication bias was detected in any of the three meta-analyses (Figures S2–S4). The sensitivity analyses showed that our study had little influence on the global heterogeneityinthethreemeta-analyses.Besides,ourstudyhadamajorinfluenceontheglobaleffectsizeinthemeta-analysisoftheneutrophil countinPSP(Figures S5–S7). DISCUSSION This study specifically investigated the peripheral immune profile in patients with PSP, and whether it could be different from patients with PD. Inourcohort,patientswithPSPshowedahigherNLRanda higher neutrophil count in peripheral blood than HCs, supporting the presence of a peripheral inflammatory immune response. However, whereas patients with PD had a lower lymphocyte count compared with HCs, no differences were found in the lymphocyte count between patients with PSP and HCs. Besides, the differential immune profile reported did not differ according to sex. Remarkably, the peripheral inflammatory immune profile found in our cohort was supported by the subsequent meta-analyses performed. FIGURE 2 PRISMAflowchartofthe literature search and study selection process. 14681331, 2024, 12, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/ene.16451 by Readcube (Labtiva Inc.), Wiley Online Library on [14/04/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 6 of 9 | MUÑOZ-DELGADO et al. Peripheral inflammation seems to be present in patients with PSP, as shown by a higher NLR compared with HCs, and regardlessofdiseaseduration.Tothebestofourknowledge,ourcross- sectional study included the largest sample size of patients with PSPpublishedtodate(n = 121)andthiswasthefirstmeta-analysis conductedtoclarifytheroleoftheNLRinPSP.Itshouldbenoted that a substantial heterogeneity was observed amongst all the includedstudiesinthemeta-analysisthatcouldnotbeclarified.First, PSP encompasses different clinical phenotypes in which different pathogenic mechanisms are likely to be involved. In the four studiesincludedinthemeta-analysis,PSPwasdiagnosedaccordingto themostrecentclinicalcriteriapublishedin2017.However,itcould be hypothesized that the heterogeneity found could be explained by phenotype heterogeneity amongst PSP patients. Even though Alsteretal.[12]includedonlypatientswithPSP-RSprobableand Madetko-Alsteretal.'sstudy[10]includedpatientswithPSP-RSand PSP-P,Incietal.'sstudy[11]assessedpatientswithprobablePSP without specifying the subtypes. Likewise, our study cohort also included different PSP phenotypes. For this reason, the peripheral immune profile in a homogeneous subgroup including only patients withPSP-RSprobablewasalsostudied.PatientswithPSP-RSalso showedahigherNLRcomparedtoHCs,reinforcingthepresenceof peripheral inflammation at least in the most frequent PSP phenotype.Nodifferenceswerefoundintheperipheralimmuneprofile FIGURE 3 Forestplotdisplaysrandomeffectsmeta-analysisresultsoftheassociationbetweentheneutrophil-to-lymphocyteratio(NLR) andprogressivesupranuclearpalsy.Theoverallstandardmeandifferencebetweengroupsandits95%confidenceintervalisrepresentedby a black diamond. FIGURE 4 Forestplotdisplaysrandomeffectsmeta-analysisresultsoftheassociationbetweentheneutrophilcountandprogressive supranuclearpalsy.Theoverallstandardmeandifferencebetweengroupsandits95%confidenceintervalisrepresentedbyablack diamond. FIGURE 5 Forestplotdisplaysrandomeffectsmeta-analysisresultsoftheassociationbetweenlymphocytecountandprogressive supranuclearpalsy.Theoverallstandardmeandifferencebetweengroupsandits95%confidenceintervalisrepresentedbyablack diamond. 14681331, 2024, 12, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/ene.16451 by Readcube (Labtiva Inc.), Wiley Online Library on [14/04/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License | 7 of 9 PERIPHERAL INFLAMMATION IN PROGRESSIVE SUPRANUCLEAR PALSY betweenthePSP-RSandthePSP-Pphenotypes.However,dueto thesmallsamplesizeofpatientswithPSP-Pincluded,resultsshould be interpreted with caution and further specific studies are needed. Unfortunately, the peripheral immune profile could not be studied in the other PSP phenotypes also due to the small sample size. But, considering that different PSP phenotypes have different clinical features and disease progression [16], further research should be conducted to assess whether different inflammatory mechanisms also exist amongst PSP phenotypes. Secondly, dopaminergic treatment should be considered since dopaminergic receptors are expressed in immune cells [17].TheInci,AlsterandMadetko-Alster studies did not report whether patients with PSP were under dopaminergic treatment. In our cohort, although most of the patients with PSP were under dopaminergic treatment, no correlation was foundwiththeNLR. The presence of a higher peripheral inflammation present in PSP is also consistent with the scarce data reported in the literature [8, 10].AnalteredinflammatoryperipheralcytokineprofileinPSP-RS andPSP-PwasalsosuggestedbyMadetko-Alsteretal.However, they did not find any correlation between serum or CSF cytokines andtheNLR.StudiesthatcorrelatetheNLRwithotherinflammatory or neurodegeneration biomarkers are lacking to date. So, whether theNLRisatriggeroranoutcomeofPSPpathogenesisisstillunclear. Furthermore, both patients with PSP and patients with PD showed a higher NLR compared to HCs, with no statistically significant differences between the two parkinsonisms. According to our results, peripheral inflammation certainly has a role in both PSP and PD [13].Besides,ahigherNLRhasconsistentlybeenreportedinothertauopathiessuchasAlzheimer'sdisease(AD)[18, 19].Altogether,althoughtheNLRisprobablyagoodbiomarkerof peripheral inflammation, it does not seem to be a useful biomarker for differential diagnosis. Remarkably,theNLRisthoughttoreflectthebalancebetween the innate and adaptive immune responses, as represented by the neutrophils and lymphocytes. It could be hypothesized that different inflammatory processes may be contributing to neurodegeneration in both conditions. To the best of our knowledge, little is known about the contribution of lymphocytes and neutrophils in PSP. Noteworthy,ourstudytogetherwiththemeta-analysesperformed showed that the altered peripheral immune profile in patients with PSP is mainly driven by neutrophils. Previous studies provided inconclusive results on the role of neutrophils in PSP [10, 11]. In contrast to our study, the small size oftheotherstudies(<24patientswithPSP)couldhavelimited thestatisticalpowertoachievesignificantdifferences.Although in our study patients with PSP were under dopaminergic treatment and a weak correlation trend between neutrophils and the LEDD was found, it should be noted that dopamine has been associated with inhibitory effects on neutrophils [17]. Consequently, the higher neutrophil count found in our cohort would not be explainedbydopaminergictreatment.Neutrophilshaveemergedto strongly contribute to chronic inflammation and neurodegenerative diseases,includingother tauopathiessuchasAD[19, 20]. Interestingly, many studies support that neutrophils are increased andhyperactivatedinthebloodandbrainofpatientswithADand it is widely accepted that neutrophils would be contributing to neurotoxicity [19, 21].Inthiscontext,pro-inflammatorycytokines such as interleukin 1β ( I L - 1 β)regulateneutrophillifespanatsitesof inflammationandcandelayneutrophilapoptosis.NotablyinPSP, ithasbeenreportedthatanincreaseofIL-1β has been found in the substantia nigra and in the CSF of patients with PSP [4, 9]. However, no studies have specifically addressed the relationship between neutrophils and neurodegeneration in PSP. Whether neutrophils are the cause or consequence of neurodegeneration requires further research. On the other hand, the lymphocyte count did not differ between patients with PSP and HCs. However, a significantly lower lymphocyte count was found in patients with PD compared with HCs, suggesting that inflammatory pathogenic mechanisms might be different between these two degenerative parkinsonisms. This finding is in line with a previous study [8] which did not see any differences in the levels of total T cells in blood in patients with PSP. However, these authors reported alterations in the lymphocyte subpopulations and the natural killer cells suggesting the presence of an active and dysregulated adaptive and innate immune response in PSP. In addition, this would also be in line with data reported in othertauopathieswhereaCNSlymphocyteinfiltrationaswellasa dysregulation in T lymphocyte subpopulations have been reported [22–24], without changes in the total lymphocyte count, in contrast to PD [13]. These data need validation in other cohorts, but they warrant future research into the possible role of the immune system in PSP pathophysiology. The major limitations of our study were its retrospective nature and that the recruitment was based on clinical diagnosis without pathological confirmation. However, the diagnosis was confirmed ateachfollow-upvisitandrigorousexclusioncriteriawereapplied to avoid confounding factors. Other inflammatory markers were not available to support systemic inflammation and correlations with clinical features or neurodegenerative markers could not be assessed.Eventhoughitwasasingle-centrestudyandtheresults could not be validated in another cohort, the meta-analyses performed reinforce the results. CONCLUSIONS Taken together, our findings indicate an increased peripheral inflammationpresentinpatientswithPSPcomparedwithHCs.Ahigher NLRwasfoundinpatientswithPSP,regardlessofdiseaseduration. However, different pathogenic inflammatory mechanisms are probably involved in PSP and PD, since in patients with PSP this altered peripheral immune profile is mainly driven by neutrophils. By contrast, our results do not suggest a primary role of lymphocytes in PSP.Furtherprospectivemulti-centrestudiesarerequiredtoconfirmourresultsandtoelucidatethecause-and-effectrelationship. Understanding the role of the peripheral inflammatory immune 14681331, 2024, 12, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/ene.16451 by Readcube (Labtiva Inc.), Wiley Online Library on [14/04/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 8 of 9 | MUÑOZ-DELGADO et al. responses and the neutrophils in the complex pathophysiological mechanisms in PSP is also likely to have therapeutic implications in the future. AUTHOR CONTRIBUTIONS Laura Muñoz- Delgado: Conceptualization; funding acquisition; writing–originaldraft;methodology;writing–reviewandediting; visualization; project administration; resources. Antonio Luque- Ambrosiani:Resources;writing–reviewandediting.Belén Benítez Zamora:Formalanalysis;methodology;visualization;writing–review and editing. Daniel Macías- García:Resources;writing–review and editing. SILVIA JESUS: Resources;writing – review and editing. Astrid Adarmes- Gómez:Writing–reviewandediting;resources. Elena Ojeda- Lepe:Resources;writing–reviewandediting. Fátima Carrillo:Resources;writing–reviewandediting.Pablo Mir: Conceptualization; methodology; resources; writing – review and editing; supervision. ACKNOWLEDGEMENTS The authors thank the donors and the Hospital Universitario Virgen delRocío-InstitutodeBiomedicinadeSevillaBiobank(Andalusian Public Health System Biobank and ISCIII-Red de Biobancos y Biomodelos-ISCIII-PT20/00069)forthehumanspecimensusedin this study. FUNDING INFORMATION ThisworkwassupportedbytheConsejeríadeSaludyConsumode la Junta de Andalucía (NEU-0005-2022), the Spanish Ministry of ScienceandInnovation(RTC2019-007150-1),theInstitutodeSalud Carlos III-Fondo Europeo de Desarrollo Regional (ISCIII-FEDER) (PI16/01575, PI18/01898, PI19/01576, PI20/00613, PI21/01875, PI22/01704,PI23/00512),theConsejeríadeEconomía,Innovación, CienciayEmpleodelaJuntadeAndalucía(CVI-02526,CTS-7685), theConsejeríadeSaludyBienestarSocialdelaJuntadeAndalucía ( P I - 0 4 7 1 - 2 0 1 3 ,  P E - 0 2 1 0 - 2 0 1 8 ,  P I - 0 4 5 9 - 2 0 1 8 ,  P E - 0 1 8 6 - 2 0 1 9 ) ,  t h e  ConsejeríadeTransformaciónEconómica,Industria,Conocimiento y Universidades de la Junta de Andalucía (PY20_00896). Several authors of this publication are members of the European Reference NetworkforRareNeurologicalDiseases—ProjectIDNo.739510and L.M.D.andP.M.aremembersoftheCOSTActionIMMUPARKNET (CA21117).L.Muñoz-Delgadowassupportedbythe‘RíoHortega’ programme (CM21/00051) from the Instituto de Salud Carlos III, and has received support for attending meetings and/or travel or honorarium for lecturing from Abbvie, Merz and Italfarmaco. A. Luque-Ambrosiani and B. Benítez-Zamora report no disclosures. D. Macías-Garcia was supported by the ‘Juan Rodés’ programme (JR22/00073)fromtheInstitutodeSaludCarlosIII,andhasreceived support for attending meetings and/or travel or honorarium for lecturingfromPalex,AbbvieandItalfarmaco.S.Jesúshasreceived support for attending meetings and/or travel or honorarium for lecturingfromAbbott,Abbvie,Bial,Italfarmaco,MerzandZambon. ADAdarmes-Gómezhasreceivedsupportforattendingmeetings and/or travel or honorarium for lecturing from Abbvie, Bial, Italfarmaco,Merz,UCBandZambon.F.Carrillohasreceivedsupport for attending meetings and/or travel or honorarium for lecturing fromAbbvieandMedtronic.E.Ojeda-Lepewassupportedbythe ‘RíoHortega’programme(CM23/00288)fromtheInstitutodeSalud Carlos III (ISCIII-FEDER), and has received support for attending meetings and/or travel or honorarium for lecturing from Abbvie, Bial, Esteve, Merz and Zambon. P. Mir has received support for attending meetings and/or travel or honorarium for lecturing from Abbott, Allergan, Abbvie, Bial, Britannia, Italfarmaco, Merz, UCB, TevaandZambon. CONFLICT OF INTEREST STATEMENT The authors declare no competing interests. DATA AVAILABILITY STATEMENT The data that support the findings of this study are available from the corresponding author upon reasonable request. ETHICS STATEMENT The local ethics committee approved the study (application code S2300212),andwritteninformedconsentforresearchwasobtained from study participants. ORCID Laura Muñoz- Delgado https://orcid.org/0000-0002-8818-1824 Daniel Macías- García https://orcid.org/0000-0002-4822-1529 Pablo Mir https://orcid.org/0000-0003-1656-302X REFERENCES 1. Höglinger GU, Respondek G, Stamelou M, et al. Clinical diagnosis of progressive supranuclear palsy: the Movement Disorder Society criteria. Mov Disord.2017;32(6):853-864.doi:10.1002/mds.26987 2. Grimm MJ, Respondek G, Stamelou M, et al. How to apply the Movement Disorder Society criteria for diagnosis of progressive supranuclear palsy. 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