Integrative proteome analysis of bone marrow interstitial fluid and serum reveals candidate signature for acute myeloid leukemia.

Acute myeloid leukemia (AML) is an aggressive form of blood cancer and clinically highly heterogeneous characterized by the accumulation of clonally proliferative immature precursors of myeloid lineage leading to bone marrow failure. Although, the current diagnostic methods for AML consist of cytogenetic and molecular assessment, there is a need for new markers that can serve as useful candidates in diagnosis, prognosis and understanding the pathophysiology of the disease. This study involves the investigation of alterations in the bone marrow interstitial fluid and serum proteome of AML patients compared to controls using label-free quantitative proteomic approach. A total of 201 differentially abundant proteins were identified in AML BMIF, while in the case of serum 123 differentially abundant proteins were identified. The bioinformatics analysis performed using IPA revealed several altered pathways including FAK signalling, IL-12 signalling and production of macrophages etc. Verification experiments were performed in a fresh independent cohort of samples using MRM assays led to the identification of a panel of three proteins viz., PPBP, APOH, ENOA which were further validated in a new cohort of serum samples by ELISA. The three-protein panel could be helpful in the diagnosis, prognosis and understanding of the pathophysiology of AML in the future. BIOLOGICAL SIGNIFICANCE: Acute Myeloid Leukemia (AML) is a type haematological malignancy which constitute one third of total leukemias and it is the most common acute leukemia in adults. In the current clinical practice, the evaluation of diagnosis and progression of AML is largely based on morphologic, immunophenotypic, cytogenetic and molecular assessment. There is a need for new markers/signatures which can serve as useful candidates in diagnosis and prognosis. The present study aims to identify and validate candidate biosignature for AML which can be useful in diagnosis, prognosis and understand the pathophysiology of the disease. Here, we identified 201 altered proteins in AML BMIF and 123 in serum. Among these altered proteins, a set of three proteins viz., pro-platelet basic protein (CXCL7), enolase 1 (ENO1) and beta-2-glycoprotein 1 (APOH) were significantly increased in AML BMIF and serum suggest that this panel of proteins could help in future AML disease management and thereby improving the survival expectancy of AML patients.

Serum proteome signatures associated with ileal and colonic ulcers in Crohn's disease.

At a clinical level, ileal and colonic Crohn's disease (CD) are considered as separate entities. These subphenotypes need to be better supported by biological data to develop personalised medicine in CD. To this end, we combined different technologies (proximity extension assay, selected reaction monitoring, and high-sensitivity turbidimetric immunoassay (hsCRP)) to measure 207 immune-related serum proteins in CD patients presenting no endoscopic lesions (endoscopic remission) (n = 23), isolated ileal ulcers (n = 17), or isolated colonic ulcers (n = 16). We showed that isolated ileal ulcers and isolated colonic ulcers were specifically associated with 6 and 18 serum proteins, respectively: (high level: JUN, CNTNAP2; low level: FCRL6, LTA, CLEC4A, NTF4); (high level: hsCRP, IL6, APCS, CFB, MBL2, IL7, IL17A, CCL19, CXCL10, CSF3, IL10, CLEC4G, MMP12, VEGFA; low level: CLEC3B, GSN, TNFSF12, TPSAB1). Isolated ileal ulcers and isolated colonic ulcers were detected by hsCRP with an area under the receiver operating characteristics curve of 0.64 (p-value = 0.07) and 0.77 (p-value = 0.001), respectively. We highlighted distinct serum proteome profiles associated with ileal and colonic ulcers in CD, this finding might support the development of therapeutics and biomarkers tailored to disease location. SIGNIFICANCE: Although ileal and colonic Crohn's disease present important clinical differences (eg, progression, response to treatment and reliability of biomarkers), these two entities are managed with the same therapeutic strategy. The biological specificities of ileal and colonic Crohn's disease need to be better characterised to develop more personalised approaches. The present study used robust technologies (selected reaction monitoring, proximity extension assays and turbidimetric immunoassay) to quantify precisely 207 serum immune-related proteins in three groups of Crohn's disease patients presenting: 1) no endoscopic lesions (endoscopic remission) (n = 23); 2) isolated ileal ulcers (n = 17); 3) isolated colonic ulcers (n = 16). We found distinct serum proteome signatures associated with ileal and colonic ulcers. Our findings could foster the development of biomarkers and treatments tailored to Crohn's disease location.

Cathepsin S Is More Abundant in Serum of Mycobacterium avium subsp. paratuberculosis-Infected Dairy Cows.

Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of bovine paratuberculosis, a chronic granulomatous enteritis leading to economic losses and posing a risk to human health due to its zoonotic potential. The pathogen cannot reliably be detected by standard methods, and immunological procedures during the infection are not well understood. Therefore, the aim of our study was to explore host-pathogen interactions in MAP-infected dairy cows and to improve diagnostic tests. Serum proteomics analysis using quantitative label-free LC-MS/MS revealed 60 differentially abundant proteins in MAP-infected dairy cows compared to healthy controls from the same infected herd and 90 differentially abundant proteins in comparison to another control group from an uninfected herd. Pathway enrichment analysis provided new insights into the immune response to MAP and susceptibility to the infection. Furthermore, we found a higher abundance of Cathepsin S (CTSS) in the serum of MAP-infected dairy cows, which is involved in multiple enriched pathways associated with the immune system. Confirmed with Western blotting, we identified CTSS as a potential biomarker for bovine paratuberculosis. This study enabled a better understanding of procedures in the host-pathogen response to MAP and improved detection of paratuberculosis-diseased cattle.

Cross-platform proteomics signatures of extreme old age.

In previous work we used a Somalogic platform targeting approximately 5000 proteins to generate a serum protein signature of centenarians that we validated in independent studies that used the same technology. We set here to validate and possibly expand the results by profiling the serum proteome of a subset of individuals included in the original study using liquid chromatography tandem mass spectrometry (LC-MS/MS). Following pre-processing, the LC-MS/MS data provided quantification of 398 proteins, with only 266 proteins shared by both platforms. At 1% FDR statistical significance threshold, the analysis of LC-MS/MS data detected 44 proteins associated with extreme old age, including 23 of the original analysis. To identify proteins for which associations between expression and extreme-old age were conserved across platforms, we performed inter-study conservation testing of the 266 proteins quantified by both platforms using a method that accounts for the correlation between the results. From these tests, a total of 80 proteins reached 5% FDR statistical significance, and 26 of these proteins had concordant pattern of gene expression in whole blood. This signature of 80 proteins points to blood coagulation, IGF signaling, extracellular matrix (ECM) organization, and complement cascade as important pathways whose protein level changes provide evidence for age-related adjustments that distinguish centenarians from younger individuals.

Inflammatory and subtype-dependent serum protein signatures predict survival beyond the ctDNA in aggressive B cell lymphomas.

BACKGROUND: Biological heterogeneity of large B cell lymphomas (LBCLs) is poorly captured by current prognostic tools, hampering optimal treatment decisions. METHODS: We dissected the levels of 1,463 serum proteins in a uniformly treated trial cohort of 109 patients with high-risk primary LBCL (ClinicalTrials.gov: NCT01325194) and correlated the profiles with molecular data from tumor tissue and circulating tumor DNA (ctDNA) together with clinical data. FINDINGS: We discovered clinically and biologically relevant associations beyond established clinical estimates and ctDNA. We identified an inflamed serum protein profile, which reflected host response to lymphoma, associated with inflamed and exhausted tumor microenvironment features and high ctDNA burden, and translated to poor outcome. We composed an inflammation score based on the identified inflammatory proteins and used the score to predict survival in an independent LBCL trial cohort (ClinicalTrials.gov: NCT03293173). Furthermore, joint analyses with ctDNA uncovered multiple serum proteins that correlate with tumor burden. We found that SERPINA9, TACI, and TARC complement minimally invasive subtype profiling and that TACI and TARC can be used to evaluate treatment response in a subtype-dependent manner in the liquid biopsy. CONCLUSIONS: Altogether, we discovered distinct serum protein landscapes that dissect the heterogeneity of LBCLs and provide agile, minimally invasive tools for precision oncology. FUNDING: This research was funded by grants from the Research Council of Finland, Finnish Cancer Organizations, Sigrid Juselius Foundation, University of Helsinki, iCAN Digital Precision Cancer Medicine Flagship, Orion Research Foundation sr, and Helsinki University Hospital.

Protein-Centric Analysis of Personalized Antibody Repertoires Using LC-MS-Based Fab-Profiling on a timsTOF.

Endogenous antibodies, or immunoglobulins (Igs), abundantly present in body fluids, represent some of the most challenging samples to analyze, largely due to the immense variability in their sequences and concentrations. It has been estimated that our body can produce billions of different Ig proteins with different isotypes, making their individual analysis seemingly impossible. However, recent advances in protein-centric proteomics using LC-MS coupled to Orbitrap mass analyzers to profile intact Fab fragments formed by selective cleavage at the IgG-hinge revealed that IgG repertoires may be less diverse, albeit unique for each donor. Serum repertoires seem to be dominated by a few hundred clones that cumulatively make up 50-95% of the total IgG content. Enabling such analyses required careful optimization of the chromatography and mass analysis, as all Fab analytes are highly alike in mass (46-51 kDa) and sequence. To extend the opportunities of this mass-spectrometry-based profiling of antibody repertoires, we here report the optimization and evaluation of an alternative MS platform, namely, the timsTOF, for antibody repertoire profiling. The timsTOF mass analyzer has gained traction in recent years for peptide-centric proteomics and found wide applicability in plasma proteomics, affinity proteomics, and HLA peptidomics, to name a few. However, for protein-centric analysis, this platform has been less explored. Here, we demonstrate that the timsTOF platform can be adapted to perform protein-centric LC-MS-based profiling of antibody repertoires. In a side-by-side comparison of the timsTOF and the Orbitrap we demonstrate that the extracted serum antibody repertoires are alike qualitatively and quantitatively, whereby in particular the sensitivity of the timsTOF platform excels. Future incorporation of advanced top-down capabilities on the timsTOF may make this platform a very valuable alternative for protein-centric proteomics and top-down proteomics and thus also for personalized antibody repertoire profiling.

Synaptic injury in the inner plexiform layer of the retina is associated with progression in multiple sclerosis.

While neurodegeneration underlies the pathological basis for permanent disability in multiple sclerosis (MS), predictive biomarkers for progression are lacking. Using an animal model of chronic MS, we find that synaptic injury precedes neuronal loss and identify thinning of the inner plexiform layer (IPL) as an early feature of inflammatory demyelination-prior to symptom onset. As neuronal domains are anatomically segregated in the retina and can be monitored longitudinally, we hypothesize that thinning of the IPL could represent a biomarker for progression in MS. Leveraging our dataset with over 800 participants enrolled for more than 12 years, we find that IPL atrophy directly precedes progression and propose that synaptic loss is predictive of functional decline. Using a blood proteome-wide analysis, we demonstrate a strong correlation between demyelination, glial activation, and synapse loss independent of neuroaxonal injury. In summary, monitoring synaptic injury is a biologically relevant approach that reflects a potential driver of progression.

CD109 identified in circulating proteomics mitigates postoperative recurrence in chronic rhinosinusitis with nasal polyps by suppressing TGF-ß1-induced epithelial-mesenchymal transition.

BACKGROUND: Chronic rhinosinusitis with nasal polyps (CRSwNP) is a common inflammatory disorder with a high rate of recurrence. This study aimed to explore biomarkers for identifying patients with recurrent CRSwNP (rCRSwNP). METHODS: We recruited two independent cohorts. In the discovery cohort, rCRSwNP patients and non-recurrent CRSwNP (non-rCRSwNP) patients were recruited, and the serum proteomic profile was characterized. The top 5 upregulated and downregulated proteins were confirmed in the validation cohort by ELISA, WB, and qRT-PCR, and their predictive values for postoperative recurrence were assessed. In vitro, human nasal epithelial cells (HNEpCs) were employed to assess the ability of candidate proteins to induce epithelial-mesenchymal transition (EMT). RESULTS: Serum proteomics identified 53 different proteins, including 30 increased and 23 decreased, between the rCRSwNP and non-rCRSwNP groups. ELISA results revealed that serum levels of CD163 and TGF-ß1 were elevated, CD109 and PRDX2 were decreased in the rCRSwNP group compared to the non-rCRSwNP group, and serum CD163, TGF-ß1, and CD109 levels were proved to be associated with the risk of postoperative recurrence. In addition, qRT-PCR and WB revealed that tissue CD163, TGF-ß1, and CD109 expressions in rCRSwNP patients were enhanced compared to those non-rCRSwNP patients. Kaplan-Meier analysis showed that increased CD163 and TGF-ß1 expression and decreased CD109 expression are associated with the risk of recurrence in CRSwNP patients. Receiver operating characteristic curves showed that TGF-ß1 and CD109 had superior diagnostic performances for rCRSwNP. In vitro experiments showed that TGF-ß1 promoted EMT in HNEpCs, and overexpression of CD109 reversed this effect. Functional recovery experiments confirmed that CD109 could attenuate EMT in HNEpCs by inhibiting the TGF-ß1/Smad signaling pathway, attenuating EMT in epithelial cells. CONCLUSION: Our data suggested that TGF-ß1 and CD109 might serve as promising predictors of rCRSwNP. The TGF-ß1/Smad pathway was implicated in fostering EMT in epithelial cells, particularly those exhibiting low expression of CD109. Consequently, the absence of CD109 expression in epithelial cells could be a potential mechanism underlying rCRSwNP.

Proteomic Analysis Identifies GSN as a Noninvasive Circulating Serum Biomarker for Predicting Early Recurrence of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is susceptible to early recurrence, but it lacks effective predictive biomarkers. In this study, we retrospectively selected 179 individuals as a discovery cohort (126 HCC patients and 53 liver cirrhosis (LC) patients) for screening candidate serum biomarkers of early recurrence based on data independent acquisition-mass spectrometry strategy. And then, the candidate biomarkers were validated in an additional independent cohort with 192 individuals (142 HCC patients and 50 LC patients) using parallel reaction monitoring targeted quantitative techniques (PXD047852). Eventually, we validated that gelsolin (GSN) concentrations were significantly lower in HCC than in LC (p < 0.0001), patients with low GSN concentrations had a poor prognosis (p < 0.0001), and GSN concentrations were significantly lower in early recurrence HCC than in late recurrence HCC (p < 0.0001). These trends were also observed in alpha-fetoprotein (AFP)-negative HCC patients. The area under the curve of machine-learning-based predictive model (GSN and microvascular invasion) for predicting early recurrence risk reached 0.803 (95% confidence interval (CI): 0.786-0.820) and maintained the same efficacy in AFP-negative patients. In conclusion, GSN is a novel serum biomarker for early recurrence of HCC. The model could provide timely warning to HCC patients at high risk of recurrence.

Serum proteomics identifies novel diagnostic biomarkers for asthma in preschool children.

Background: Asthma is characterized by airway hyperresponsiveness, reversible airway obstruction, and chronic airway inflammation. It is the most common chronic disease in childhood. However, the diagnosis of childhood asthma remains challenging, and there is an urgent need to develop new diagnostic methods. Methods: To identify biomarkers of asthma in children, we adopted the Orbitrap-based data-independent acquisition (DIA) mass spectrometry proteomics method to analyze the serum proteomic signatures of children with acute asthma and convalescent children. Results: We identified 747 proteins in 46 serum samples and 50 differentially expressed proteins (DEPs) that distinguished between asthmatic and healthy children. Next, functional enrichment analysis of the DEPs was conducted, it was indicated that the DEPs were significantly enriched in immune-related and function terms and pathways. Furthermore, we performed statistical analysis and identified MMP14, ABHD12B, PCYOX1, LTBP1, CFHR4, APOA1, IGHG4, ANG and IGFALS proteins as the diagnostic biomarker candidates. Ultimately, a promising asthma diagnostic model for preschool children based on IGFALS was built and evaluated. The area under the curve (AUC) of the IGFALS model was 0.959. Conclusions: In this study, the DIA proteome strategy was used and the largest number of proteins of asthmatic children serum proteomics was identified. The proteomics results showed that the DEPs play the central role of the inflammation-immune mechanism in asthma pathogenesis, suggesting that these proteins may be used in asthma diagnosis, prognosis, or therapy, and suggested biomarkers for asthma of preschool children. In conclusion, our results provide insight into the pathophysiology of asthma. We believe that the diagnostic model will facilitate clinical decision-making regarding asthma in preschool children.

Proteomic prediction of incident heart failure and its main subtypes.

AIM: To examine the ability of serum proteins in predicting future heart failure (HF) events, including HF with reduced or preserved ejection fraction (HFrEF or HFpEF), in relation to event time, and with or without considering established HF-associated clinical variables. METHODS AND RESULTS: In the prospective population-based Age, Gene/Environment Susceptibility Reykjavik Study (AGES-RS), 440 individuals developed HF after their first visit with a median follow-up of 5.45 years. Among them, 167 were diagnosed with HFrEF and 188 with HFpEF. A least absolute shrinkage and selection operator regression model with non-parametric bootstrap were used to select predictors from an analysis of 4782 serum proteins, and several pre-established clinical parameters linked to HF. A subset of 8-10 distinct or overlapping serum proteins predicted different future HF outcomes, and C-statistics were used to assess discrimination, revealing proteins combined with a C-index of 0.80 for all incident HF, 0.78 and 0.80 for incident HFpEF or HFrEF, respectively. In the AGES-RS, protein panels alone encompassed the risk contained in the clinical information and improved the performance characteristics of prediction models based on N-terminal pro-B-type natriuretic peptide and clinical risk factors. Finally, the protein predictors performed particularly well close to the time of an HF event, an outcome that was replicated in the Cardiovascular Health Study. CONCLUSION: A small number of circulating proteins accurately predicted future HF in the AGES-RS cohort of older adults, and they alone encompass the risk information found in a collection of clinical data. Incident HF events were predicted up to 8 years, with predictor performance significantly improving for events occurring less than 1 year ahead, a finding replicated in an external cohort study.

S-Ketamine Improves Slow Wave Sleep and the Associated Changes in Serum Protein Among Gynecological Abdominal Surgery Patients: A Randomized Controlled Trial.

Purpose: This study aims to evaluate the effect of S-ketamine on slow wave sleep (SWS) and the related changes in serum protein in gynecological patients after open abdomen surgery. Methods: This was a randomized controlled trial. One hundred gynecological patients undergoing open abdomen surgery were randomized into an S-ketamine group (group S) or placebo group (0.9% saline; group C). During operation, patients in group S received adjuvant S-ketamine infusion (0.2 mg·kg-1·h-1) while those in group C received 0.9% saline. All patients were connected to patient-controlled intravenous analgesia (PCIA) pump in the end of the surgery and the patients in group S with an additional S-ketamine in PCIA pump. Polysomnogram (PSG) was monitored during the next night after surgery with PCIA pump. Blood samples were collected for proteomic analysis at 6:00 AM after PSG monitoring. The primary outcome was the percentage of SWS (also known as stage 3 non-rapid eye movement sleep, stage N3) on the next night after surgery, and the secondary outcome was subjective sleep quality, pain scores, and the changes in serum proteomics. Results: Complete polysomnogram recordings were obtained from 64 study participants (31 in group C and 33 in group S). The administration of S-ketamine infusion resulted in a significant increase in the percentage of SWS/N3 compared to the control group (group C, median (IQR [range]), 8.9 (6.3, 12.5); group S, median (IQR [range]), 15.6 (12.4, 18.8), P<0.001). However, subjective evaluations of sleep quality revealed no significant variances between the two groups. The protein affected by S-ketamine was primarily associated with posttranslational modification, protein turnover, carbohydrate transport, and metabolism. Conclusion: In patients undergoing open gynecological surgery, S-ketamine enhanced the percentage of objective sleep of SWS during the next night after surgery. Additionally, there were differences observed in serum protein levels between the two groups. Trial Registration: ChiCTR2200055180. Registered on 02/01/2022.

Proteomic Profiling in Patients With Peripartum Cardiomyopathy: A Biomarker Study of the ESC EORP PPCM Registry.

BACKGROUND: Peripartum cardiomyopathy (PPCM) remains an important cause of maternal morbidity and mortality globally. The pathophysiology remains incompletely understood, and the diagnosis is often missed or delayed. OBJECTIVES: This study explored the serum proteome profile of patients with newly diagnosed PPCM, as compared with matched healthy postpartum mothers, to unravel novel protein biomarkers that would further an understanding of the pathogenesis of PPCM and improve diagnostic precision. METHODS: Study investigators performed untargeted serum proteome profiling using data-independent acquisition-based label-free quantitative liquid chromatography-tandem mass spectrometry on 84 patients with PPCM, as compared with 29 postpartum healthy controls (HCs). Significant changes in protein intensities were determined with nonpaired Student's t-tests and were further classified by using the Boruta algorithm. The proteins' diagnostic performance was evaluated by area under the curve (AUC) and validated using the 10-fold cross-validation. RESULTS: Patients with PPCM presented with a mean left ventricular ejection fraction of 33.5% ± 9.3% vs 57.0% ± 8.8% in HCs (P < 0.001). Study investigators identified 15 differentially up-regulated and 14 down-regulated proteins in patients with PPCM compared with HCs. Seven of these proteins were recognized as significant by the Boruta algorithm. The combination of adiponectin, quiescin sulfhydryl oxidase 1, inter-α-trypsin inhibitor heavy chain, and N-terminal pro-B-type natriuretic peptide had the best diagnostic precision (AUC: 0.90; 95% CI: 0.84-0.96) to distinguish patients with PPCM from HCs. CONCLUSIONS: Salient biologic themes related to immune response proteins, inflammation, fibrosis, angiogenesis, apoptosis, and coagulation were predominant in patients with PPCM compared with HCs. These newly identified proteins warrant further evaluation to establish their role in the pathogenesis of PPCM and potential use as diagnostic markers.

Serum Protein Profiling Reveals a Decrease in Apolipoprotein A-IV During a Clinical Depressive Mood State.

Purpose: Depressive mood is a major psychiatric symptom that causes serious disturbances in daily life. Unlike physical symptoms, psychiatric symptoms are more difficult to evaluate objectively. Therefore, we aimed to discover biomarkers that reflect changes in serum protein metabolism during a clinical depressive mood. Methods: Serum protein profiling was conducted in participants who were not experiencing a current depressive episode (healthy individuals and patients in remission). Serum proteins were identified and quantified using liquid chromatography-tandem mass spectrometry. Differentially expressed proteins with a p-value <0.05 were selected, and candidate biomarkers were verified using multiple reaction monitoring analysis for absolute quantification. Results: Apolipoprotein A-IV levels were lower in the group with a current episode of depression than in the remission and healthy control groups. Further, fibronectin levels were also lower in the group with a current episode of depression than in the healthy control group but not in the remission group. Conclusion: We found that apolipoprotein A-IV-mediated inflammation is involved in clinical depressive moods, possibly by inducing neurological changes in the brain. Therefore, apolipoprotein A-IV and fibronectin levels may be explored as potentially novel biomarkers for detecting a current episode of depression.

Potential biomarkers for psoriasis topical treatment by in-depth serum proteomics.

BACKGROUND: Psoriasis is a chronic skin disease, and topical sequential therapy with a combination of calcipotriol and calcipotriol betamethasone is currently approved topical treatment. However, the exact mechanism by which this treatment regimen relieves psoriasis is unknown. METHOD: We assembled a cohort of 65 psoriasis patients and divided post-treatment cohort into responder group and non-responder group according to the Psoriasis Area Severity Index (PASI) score after 12-week treatment. We measured the expression levels of proteins in collected 130 serum samples using our in-depth proteomics platform with a data-independent acquisition mass spectrometer and antibody microarray. We performed bioinformatics analyses of the biologic processes and signaling pathways that were changed in the responder group and constructed a proteomics landscape of psoriasis pathogenesis response to treatment. We then validated the biomarkers of disease severity in an independent cohort of 88 samples using an enzyme-linked immunosorbent assay. RESULTS: We first identified 174 differentially expressed proteins (DEPs) for comparative analysis of proteins between responders and non-responders at baseline (p < 0.05). Then pathway analysis showed that the responders focused more on signaling molecules and interaction, complement and coagulation cascades, whereas the non-responders more on signal transduction and IL-17 signaling pathways. We further identified four candidate biomarkers (COLEC11, C1QA, BNC2, ITIH4) response to treatment. We also found 125 DEPs (p < 0.05) after treatment compared with before treatment in responder group. Pathway analysis showed an enrichment in pathways related to complement and coagulation cascades, phagosome, ECM-receptor interaction, cholesterol metabolism, vitamin digestion and absorption. CD14 was validated as potential biomarkers for the disease severity of psoriasis and treatment targets. CONCLUSION: In this work, we analyzed the response to topical sequential therapy and finally identified four biomarkers. Additionally, we found that topical sequential therapy may alleviate psoriasis by regulating lipid metabolism and modulating the immune response by affecting the complement activation process.

Targeted serum proteomics of longitudinal samples from newly diagnosed youth with type 1 diabetes distinguishes markers of disease and C-peptide trajectory.

AIMS/HYPOTHESIS: There is a growing need for markers that could help indicate the decline in beta cell function and recognise the need and efficacy of intervention in type 1 diabetes. Measurements of suitably selected serum markers could potentially provide a non-invasive and easily applicable solution to this challenge. Accordingly, we evaluated a broad panel of proteins previously associated with type 1 diabetes in serum from newly diagnosed individuals during the first year from diagnosis. To uncover associations with beta cell function, comparisons were made between these targeted proteomics measurements and changes in fasting C-peptide levels. To further distinguish proteins linked with the disease status, comparisons were made with measurements of the protein targets in age- and sex-matched autoantibody-negative unaffected family members (UFMs). METHODS: Selected reaction monitoring (SRM) mass spectrometry analyses of serum, targeting 85 type 1 diabetes-associated proteins, were made. Sera from individuals diagnosed under 18 years (n=86) were drawn within 6 weeks of diagnosis and at 3, 6 and 12 months afterwards (288 samples in total). The SRM data were compared with fasting C-peptide/glucose data, which was interpreted as a measure of beta cell function. The protein data were further compared with cross-sectional SRM measurements from UFMs (n=194). RESULTS: Eleven proteins had statistically significant associations with fasting C-peptide/glucose. Of these, apolipoprotein L1 and glutathione peroxidase 3 (GPX3) displayed the strongest positive and inverse associations, respectively. Changes in GPX3 levels during the first year after diagnosis indicated future fasting C-peptide/glucose levels. In addition, differences in the levels of 13 proteins were observed between the individuals with type 1 diabetes and the matched UFMs. These included GPX3, transthyretin, prothrombin, apolipoprotein C1 and members of the IGF family. CONCLUSIONS/INTERPRETATION: The association of several targeted proteins with fasting C-peptide/glucose levels in the first year after diagnosis suggests their connection with the underlying changes accompanying alterations in beta cell function in type 1 diabetes. Moreover, the direction of change in GPX3 during the first year was indicative of subsequent fasting C-peptide/glucose levels, and supports further investigation of this and other serum protein measurements in future studies of beta cell function in type 1 diabetes.

Serum Proteomic Analysis of Peripartum Cardiomyopathy Reveals Distinctive Dysregulation of Inflammatory and Cholesterol Metabolism Pathways.

BACKGROUND: The pathophysiology of peripartum cardiomyopathy (PPCM) and its distinctive biological features remain incompletely understood. High-throughput serum proteomic profiling, a powerful tool to gain insights into the pathophysiology of diseases at a systems biology level, has never been used to investigate PPCM relative to nonischemic cardiomyopathy. OBJECTIVES: The aim of this study was to characterize the pathophysiology of PPCM through serum proteomic analysis. METHODS: Aptamer-based proteomic analysis (SomaScan 7K) was performed on serum samples from women with PPCM (n = 67), women with nonischemic nonperipartum cardiomyopathy (NPCM) (n = 31), and age-matched healthy peripartum and nonperipartum women (n = 10 each). Serum samples were obtained from the IPAC (Investigation of Pregnancy-Associated Cardiomyopathy) and IMAC2 (Intervention in Myocarditis and Acute Cardiomyopathy) studies. RESULTS: Principal component analysis revealed unique clustering of each patient group (P for difference <0.001). Biological pathway analyses of differentially measured proteins in PPCM relative to NPCM, before and after normalization to pertinent healthy controls, highlighted specific dysregulation of inflammatory pathways in PPCM, including the upregulation of the cholesterol metabolism-related anti-inflammatory pathway liver-X receptor/retinoid-X receptor (LXR/RXR) (P < 0.01, Z-score 1.9-2.1). Cardiac recovery by 12 months in PPCM was associated with the downregulation of pro-inflammatory pathways and the upregulation of LXR/RXR, and an additional RXR-dependent pathway involved in the regulation of inflammation and metabolism, peroxisome proliferator-activated receptor α/RXRα signaling. CONCLUSIONS: Serum proteomic profiling of PPCM relative to NPCM and healthy controls indicated that PPCM is a distinct disease entity characterized by the unique dysregulation of inflammation-related pathways and cholesterol metabolism-related anti-inflammatory pathways. These findings provide insight into the pathophysiology of PPCM and point to novel potential therapeutic targets.

Characterization of the serum and skin inflammatory profile in canine pemphigus foliaceus using multiplex assay and quantitative real-time polymerase chain reaction (qRT-PCR).

Canine pemphigus foliaceus (PF) is a common autoimmune skin disease characterized by autoantibodies binding to epithelial adhesion molecules resulting inflammatory response. The immune network of cytokine and chemokine abnormalities that characterize the immune response in canine PF are poorly explored. This study evaluated serum and lesional skin cytokine and chemokine profiles of dogs diagnosed with PF compared to healthy control dogs. Serum samples obtained from 11 PF dogs and 16 healthy control dogs were analyzed using commercially available canine multiplex assay for 13 biomarkers (Canine Milliplex assay). Eight lesional skin samples from seven PF dogs and five healthy site-matched samples from five healthy dogs were evaluated for 20 immune markers using quantitative real-time PCR. Immunomodulating medications were suspended for at least four weeks in all dogs before obtaining serum and skin samples. PF patients showed significantly higher serum concentrations of tumor necrosis factor-α, interleukin (IL)- 6, IL-8, IL-18, CCL2, KC-like, and granulocyte-macrophages colony-stimulating factor when compared to healthy controls (Mann-Whitney U test; p < 0.05 for all). Lesional PF skin exhibited significant expression and upregulation of pro-inflammatory/T helper (Th1) 1 markers IL-1ß, MX1, GZMB, OAS1, and IFN-γ as well as Th2 cytokines IL-13, IL-33, TSLP, IL-31 and Th17/22 markers IL-17A and IL-22 (Mann-Whitney U test; p < 0.05 for all). Taken together, the findings from this study describe the role of numerous cytokines and chemokines associated with immune response in the skin and serum of canine PF patients. Further larger-sample proteomics and RNA-sequencing transcriptomics studies are needed to understand the immune pathogenesis of canine PF skin lesions.

Serum proteomics analysis for differentiation among Mycobacterium tuberculosis infection categories.

Inhalation of Mycobacterium tuberculosis (Mtb) bacilli can lead to a range of TB categories including early clearance (EC), latent TB infection (LTBI) and active TB (ATB). There are few biomarkers available to differentiate among these TB categories: effective new biomarkers are badly needed. Here, we analyzed the serum proteins from 26 ATB cases, 20 LTBI cases, 34 EC cases and 38 healthy controls (HC) using label-free LC-MS/MS. The results were analyzed using MaxQuant software and matched to three different bacterial proteomics databases, including Mtb, Mycobacterium spp. and normal lung flora. PCA of protein candidates using the three proteomics databases revealed 44.5% differentiation power to differentiate among four TB categories. There were 289 proteins that showed potential for distinguishing between each pair of groups among TB categories. There were 50 candidate protein markers specifically found in ATB and LTBI but not in HC and EC groups. Decision trees using the top five candidate biomarkers (A0A1A2RWZ9, A0A1A3FMY8, A0A1A3KIY2, A0A5C7MJH5 and A0A1X0XYR3) had 92.31% accuracy to differentiate among TB categories and the accuracy was increased to 100% when using 10 candidate biomarkers. Our study shows that proteins expressed from Mycobacterium spp. have the potential to be used to differentiate among TB categories.

Mapping early serum proteome signatures of liver regeneration in living donor liver transplant cases.

The liver is the only solid organ capable of regenerating itself to regain 100% of its mass and function after liver injury and/or partial hepatectomy (PH). This exceptional property represents a therapeutic opportunity for severe liver disease patients. However, liver regeneration (LR) might fail due to poorly understood causes. Here, we have investigated the regulation of liver proteome and phosphoproteome at a short time after PH (9 h), to depict a detailed mechanistic background of the early LR phase. Furthermore, we analyzed the dynamic changes of the serum proteome and metabolome of healthy living donor liver transplant (LDLT) donors at different time points after surgery. The molecular profiles from both analyses were then correlated. Insulin and FXR-FGF15/19 signaling were stimulated in mouse liver after PH, leading to the activation of the main intermediary kinases (AKT and ERK). Besides, inhibition of the hippo pathway led to an increased expression of its target genes and of one of its intermediary proteins (14-3-3 protein), contributing to cell proliferation. In association with these processes, metabolic reprogramming coupled to enhanced mitochondrial activity cope for the energy and biosynthetic requirements of LR. In human serum of LDLT donors, we identified 56 proteins and 13 metabolites statistically differential which recapitulate some of the main cellular processes orchestrating LR in its early phase. These results provide mechanisms and protein mediators of LR that might prove useful for the follow-up of the regenerative process in the liver after PH as well as preventing the occurrence of complications associated with liver resection.