Ultrasensitive electrochemical sensor based on synergistic effect of Ag@MXene and antifouling cyclic multifunctional peptide for PD-L1 detection in serum.

A sensing interface co-constructed from the two-dimensional conductive material (Ag@MXene) and an antifouling cyclic multifunctional peptide (CP) is described. While the large surface area of Ag@MXene loads more CP probes, CP binds to Ag@MXene to form a fouling barrier and ensure the structural rigidity of the targeting sequence. This strategy synergistically enhances the biosensor's sensitivity and resistance to contamination. The SPR results showed that the binding affinity of the CP to the target was 6.23 times higher than that of the antifouling straight-chain multifunctional peptide (SP) to the target. In the 10 mg/mL BSA electrochemical fouling test, the fouling resistance of Ag@MXene + CP (composite sensing interface of CP combined with Ag@MXene) was 30 times higher than that of the bare electrode. The designed electrochemical sensor exhibited good selectivity and wide dynamic response range at PD-L1 concentrations from 0.1 to 50 ng/mL. The lowest detection limit was 24.54 pg/mL (S/N = 3). Antifouling 2D materials with a substantial specific surface area, coupled with non-straight chain antifouling multifunctional peptides, offer a wide scope for investigating the sensitivity and antifouling properties of electrochemical sensors.

Bioanalysis of lanreotide in dog plasma using liquid chromatography-tandem mass spectrometry and its application in a pharmacokinetic study of beagle dogs after single subcutaneous injection.

Lanreotide is similar to a naturally occurring hormone, somatostatin; thus, it may be used to treat acromegaly or metastatic gastroenteropancreatic neuroendocrine tumours. Here, a bioanalytical method coupling ultra-performance liquid chromatography with tandem mass spectrometry to quantify lanreotide and an internal standard (IS) was developed and validated in dog plasma. The plasma samples were extracted using typical protein precipitation processes. The analyte and internal standard were separated on Phenomenex Kinetex® C18 with 0.1% formic acid and acetonitrile in the mobile phase at a flow rate of 0.4 mL/min. The fragmentation of precursor ions to product ions was optimized at m/z 548.8 â†’ 170.0 for lanreotide [M + 2H]2+ and 472.2 â†’ 436.2 for IS [M + H]+. The peak retention times of lanreotide and IS were 1.09 min and 1.22 min, respectively. The calibration curve samples in dog plasma ranged from 0.3 to 1000 ng/mL and showed good linearity, with a correlation coefficient of r2=0.9996. The lower limit of quantitation was 0.3 ng/mL. The intra- and inter-day precision (relative standard deviation) values for each quality control level were < 9.7 % and < 9.3 %, respectively; intra- and inter-day accuracy were < 109.3% and < 110.4%, respectively. Lanreotide in dog plasma was stable in various conditions. The maximum plasma concentration of lanreotide in male beagle dogs after subcutaneous injection of Somatuline® (lanreotide) Autogel 120 mg was 88.1 ng/mL. The half-life (T1/2) of lanreotide in beagle dogs was long, approximately 198.6 h; the area under the plasma-concentration curve from 0 to 840 h (day 35) was 6,995 ng⋅h/mL. This novel quantification method using UPLC-MS/MS was successfully applied to the pharmacokinetic analysis of lanreotide in dog plasma. The results will assist future studies of drug formulation and repurposing.

Quality of life in patients with acromegaly receiving lanreotide autogel: a real-world observational study.

INTRODUCTION: Patients with acromegaly have substantially reduced quality of life (QoL). This study evaluated QoL in patients with acromegaly treated with lanreotide autogel. MATERIAL AND METHODS: This was a prospective, non-interventional, observational, multi-centre study conducted in Poland (NCT02396966). We included patients with acromegaly, who received treatment with lanreotide autogel 120mg for ≥ 3 months and < 3 years. Patients were assessed approximately every 4-5months for twoyears (six visits). QoL was measured with the Acromegaly Quality of Life Questionnaire (AcroQoL). RESULTS: Of 152 patients enrolled from November 2014 to May 2018 in 37 centres, 24 were excluded due to major protocol deviations. The results are reported for the study population (n = 128). At baseline, the median [95% confidence interval (CI)] time from diagnosis was 3.3 (2.8, 4.2)years, and the median time since lanreotide initiation was 13.4 (9.9, 17.3) months. Symptoms of acromegaly were present at baseline in 86% of patients (headache, 57%; sweating, 58%; joint symptoms, 64%); symptoms remained unchanged at two years in 82% of patients. At baseline, 27% of patients had hormonal control (growth hormone < 2.5 µg/L and insulin-like growth factor-1 within the normal range); hormonal control status did not change during the study period in over 81% of patients. At baseline, 88% of patients were either very satisfied or satisfied with treatment; treatment satisfaction was unchanged in 62% of patients over the study period. Mean (95% CI) AcroQoL scores at baseline were as follows: total, 50.3 (47.3, 53.3); physical dimension, 48.8 (45.2, 52.4); psychological dimension, 51.3 (48.2, 54.4); appearance subdimension, 40.7 (37.5, 43.8); and personal relations subdimension, 62.5 (58.8, 66.2). The psychological appearance subscore improved by 3.8 points (1.2, 6.5) over the two years; scores in the remaining dimensions and subdimensions did not change substantially. The total AcroQoL score remained unchanged over the twoyears, regardless of prior acromegaly treatment, surgery or radiotherapy, hormonal control, or lanreotide dosing interval. No new safety findings were identified. CONCLUSIONS: AcroQoL total scores and physical and psychological subscores remained stable but impaired among patients with long-lasting acromegaly treated with lanreotide autogel for two years. The psychological appearance subdimension improved numerically.

The Chemical Synthesis of Knob Domain Antibody Fragments.

Cysteine-rich knob domains found in the ultralong complementarity determining regions of a subset of bovine antibodies are capable of functioning autonomously as 3-6 kDa peptides. While they can be expressed recombinantly in cellular systems, in this paper we show that knob domains are also readily amenable to a chemical synthesis, with a co-crystal structure of a chemically synthesized knob domain in complex with an antigen showing structural equivalence to the biological product. For drug discovery, following the immunization of cattle, knob domain peptides can be synthesized directly from antibody sequence data, combining the power and diversity of the bovine immune repertoire with the ability to rapidly incorporate nonbiological modifications. We demonstrate that, through rational design with non-natural amino acids, a paratope diversity can be massively expanded, in this case improving the efficacy of an allosteric peptide. As a potential route to further improve stability, we also performed head-to-tail cyclizations, exploiting the proximity of the N and C termini to synthesize functional, fully cyclic antibody fragments. Lastly, we highlight the stability of knob domains in plasma and, through pharmacokinetic studies, use palmitoylation as a route to extend the plasma half-life of knob domains in vivo. This study presents an antibody-derived medicinal chemistry platform, with protocols for solid-phase synthesis of knob domains, together with the characterization of their molecular structures, in vitro pharmacology, and pharmacokinetics.

Pathogenic Role of Circulating Citrullinated Antigens and Anti-Cyclic Monoclonal Citrullinated Peptide Antibodies in Rheumatoid Arthritis.

We examined whether it is possible to directly detect citrullinated antigens in the serum of rheumatoid arthritis (RA) patients using a monoclonal antibody (mAb) designed to be specific for citrullinated peptides. In order to confirm the potential of the mAb as a direct arthritis-inducing substance through experimental model of RA, a monoclonal antibody (mAb) 12G1 was generated using by immunization of mice with a challenging cyclic citrullinated peptide. Immunohistochemical analysis of RA-affected synovial tissue showed that our mAb 12G1 could indeed detect citrullinated proteins in target tissues. Subsequently, serum levels of citrullinated type II collagen and filaggrin were measured in healthy volunteers, patients with RA, ankylosing spondylitis (AS), and systemic lupus erythematosus (SLE) using a 12G1-based sandwich ELISA. This showed that citrullinated filaggrin showed 78.9% sensitivity and 85.9% specificity for RA diagnosis with a cutoff optical density (OD) value of 1.013, comparable with the results from a second-generation anti-citrullinated protein antibody (ACPA) test. Circulating citrullinated collagen and filaggrin were detected even in sera of RA patients who were negative for both rheumatoid factor (RF) and ACPA. ELISA results also showed that RF and ACPA titers showed significantly positive correlation with both citrullinated collagen and filaggrin OD values in sera of RA patients. 12G1 challenging aggravated the severity of murine arthritis. In summary, mAb 12G1 can directly detect citrullinated proteins in RA target tissue and in sera of RA patients and 12G1 showed direct arthritogenic potential in vivo. This, 12G1 might be useful for diagnosis of RA including seronegative RA and may help to elucidate the pathophysiological role of citrullination in RA.

Increased Gene Copy Number of DEFA1A3 Is Associated With the Severity of Ulcerative Colitis.

INTRODUCTION: DEFA1A3 encodes human neutrophil peptides (HNPs) 1-3 and has multiple copy number variations (CNVs). HNPs are associated with innate immunity. Ulcerative colitis (UC), a chronic inflammatory gastrointestinal disorder, is a life-threatening condition, and predictive markers of UC severity are needed. This study investigated the relationship between DEFA1A3 CNV and UC severity. METHODS: This study enrolled 165 patients with UC. The relationship between DEFA1A3 CNV and disease severity was analyzed based on Mayo score, patient characteristics, and treatment methods. In addition, serum and stimulated neutrophil-derived HNP concentrations were also measured in patients with high and low DEFA1A3 CNV. RESULTS: DEFA1A3 CNV was significantly correlated with Mayo score and white blood cell count (R = 0.46, P < 0.0001; R = 0.29, P = 0.003, respectively), and only high copy numbers of DEFA1A3 were independent factors for severe UC (P < 0.001, odds ratio: 1.88, 95% confidence interval, 1.34-2.61). The number of severe UC patients with high DEFA1A3 CNV was significantly greater than those with low CNV. We confirmed the associations between DEFA1A3 and UC severity using a validation cohort. In addition, the HNP concentration in high-copy number patients was significantly higher after neutrophil stimulation than that in low-copy number patients. DISCUSSION: This study demonstrated that there is a correlation between DEFA1A3 copy number and severity in patients with UC. In addition, neutrophils from UC patients with higher DEFA1A3 CNV had high reactivity of secretion of HNPs after stimulation. DEFA1A3 CNV may be a novel severity marker and a potential therapeutic target for UC.

In Vitro Selection of Macrocyclic d/l-Hybrid Peptides against Human EGFR.

d/l-Hybrid peptides are an attractive class of molecular modality because they are able to exhibit high proteolytic stability and unique structural diversity which cannot be accessed by those consisting of only proteinogenic l-amino acids. Despite such an expectation, it has not been possible to devise de novo d/l-hybrid peptides capable of disrupting the function of a protein target(s) due to the lack of an effective method that reliably constructs a highly diverse library and screens active species. Here we report for the first time construction of a library consisting of 1012 members of macrocyclic d/l-hybrid peptides containing five kinds of d-amino acids and performance of the RaPID selection against human EGFR as a showcase to uncover PPI (protein-protein interaction) inhibitors.

A MIF-Derived Cyclopeptide that Inhibits MIF Binding and Atherogenic Signaling via the Chemokine Receptor CXCR2.

Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine and atypical chemokine with a key role in inflammatory diseases including atherosclerosis. Key atherogenic functions of MIF are mediated by noncognate interaction with the chemokine receptor CXCR2. The MIF N-like loop comprising the sequence 47-56 is an important structural determinant of the MIF/CXCR2 interface and MIF(47-56) blocks atherogenic MIF activities. However, the mechanism and critical structure-activity information within this sequence have remained elusive. Here, we show that MIF(47-56) directly binds to CXCR2 to compete with MIF receptor activation. By using alanine scanning, essential and dispensable residues were identified. Moreover, MIF(cyclo10), a designed cyclized variant of MIF(47-56), inhibited key inflammatory and atherogenic MIF activities in vitro and in vivo/ex vivo, and exhibited strongly improved resistance to proteolytic degradation in human plasma in vitro, thus suggesting that it could serve as a promising basis for MIF-derived anti-atherosclerotic peptides.

Improved classification of rheumatoid arthritis with a score including anti-acetylated ornithine antibodies.

The presence of rheumatoid factor (RF) or anti-cyclic citrullinated peptide (anti-CCP) autoantibodies contributes to the current rheumatoid arthritis (RA) classification criteria. These criteria involve stratification on antibody levels, which limits reproducibility, and underperform in the RA patients without RF and anti-CCP. Here, we have explored if two anti-acetylated peptide antibodies (AAPA), anti-acetylated lysine (AcLys) and anti-acetylated ornithine (AcOrn), could improve the performance of the current criteria. The analysis was done in 1062 prospectively-followed early arthritis (EA) patients. The anti-AcOrn were more informative than the anti-AcLys, the conventional RA antibodies and the anti-carbamylated protein antibodies. The anti-AcOrn produced a classification that did not require antibody levels and showed improved specificity (77.6% vs. 72.6%, p = 0.003) and accuracy (79.0% vs. 75.8%, p = 0.002) over the current criteria. These improvements were obtained with a scoring system that values concordance between anti-AcOrn, RF and anti-CCP. No significant gain was obtained in sensitivity (80.2% vs. 78.8%, p = 0.25) or in improving the classification of the RA patients lacking RF and anti-CCP, although the anti-AcOrn ranked first among the analysed new antibodies. Therefore, the anti-AcOrn antibodies could contribute to the improvement of RA classification criteria by exploiting antibody concordance.

Ultra-sensitive quantification of the therapeutic cyclic peptide bremelanotide utilizing UHPLC-MS/MS for evaluation of its oral plasma pharmacokinetics.

Bremelanotide (Vyleesi®), a cyclic heptapeptide, was recently approved for the subcutaneous treatment of premenopausal hypoactive sexual desire disorder. To foster the development of alternative routes of administration, we aimed at determining the oral plasma pharmacokinetics of bremelanotide in beagle dogs. Therefore, we established a UHPLC-MS/MS assay with an LLOQ of 10 pg/mL (9.8 pM) using 100 µL of plasma and validated it according to the guidelines of the US Food and Drug Administration and the European Medicines Agency. Bremelanotide was isolated from plasma by protein precipitation and quantification was performed with positive heated ESI MS/MS in the SRM mode. The calibrated concentration range of 10-10,000 pg/mL was linear showing correlation coefficients > 0.99. In the calibrated range, interday and intraday accuracy ranged from 88.9-100.0 % with corresponding precision < 8 %. Accuracy at the LLOQ ranged from 93.6-100.8 % with corresponding precision < 11 %. Because of the validity of a dilution QC that showed accurate quantification of 10-fold diluted plasma samples (accuracy 99.4 %, precision < 6 %), the assay is suitable for bremelanotide quantification in its effective concentration range up to 100,000 pg/mL. The ultra-sensitive assay was applied to the quantification of bremelanotide plasma concentrations after oral administration to beagle dogs, which indicated minimal oral absorption.

Predicting factors for disappearance of anti-mutated citrullinated vimentin antibodies in sera of patients with rheumatoid arthritis.

Objectives: We aimed to determine the predicting factors for disappearance of anti-mutated citrullinated vimentin antibody (anti-MCV Ab) in sera from rheumatoid arthritis (RA) patients.Methods: In 2013, 95 RA patients whose Disease Activity Score with erythrocyte sedimentation rate were moderate to severe (DAS28-ESR ≥3.2) at baseline were enrolled. Titers of anti-MCV Ab and anti-cyclic citrullinated peptide (anti-CCP) Ab for 2013 and 2014 were measured. The association of anti-MCV disappearance with disease activity, treatment, interstitial lung disease (ILD), and serum markers of ILD were retrospectively examined. Predicting factors of anti-MCV disappearance were determined by multivariable analysis.Results: While anti-CCP positivity rate did not change during the year, anti-MCV Ab changed from positive to negative in 18 patients (=19.0%). Continuous biological disease-modifying anti-rheumatic drug use, prednisolone dose (≥5.0 mg daily), and low KL-6 level (<191 U/mL) were determined as predicting factors of anti-MCV disappearance by multivariable analysis. In our cohort, anti-MCV Ab disappearance was not linked to clinical and radiological improvement.Conclusion: Different from anti-CCP Ab, anti-MCV Ab in sera from RA patients can disappear in a year. Some predicting factors for such negative seroconversion were found, whereas clinical significance of anti-MCV Ab disappearance was undetermined.

Rheumatoid factor versus anti - cyclic citrullinated peptide antibody as screening tool for rheumatoid arthritis in an ophthalmic clinic.

Patients with moderate to severe dry eyes are often screened at the Dry Eye Clinic to rule out connective tissue diseases. Rheumatoid factor (RF) is one of the screening tools to rule out rheumatoid arthritis (RA). Patients who turn out positive for the RF are often subjected to anti-CCP antibody evaluation for confirmation of disease. This article tries to highlight 3 cases of negative and anti-CCP antibody positive cases which presented to the ophthalmic clinic, unaware of their systemic status. Though RF is the cheapest modality to screen for RA, it is not always a reliable marker. One should order anti-CCP antibody for patients where suspicion is high, despite RF being normal.

Diagnostic performance of 14-3-3η and anti-carbamylated protein antibodies in Rheumatoid Arthritis in Han population of Northern China.

OBJECTIVES: As we already know, Rheumatoid arthritis (RA) cannot be excluded when the rheumatoid factor (RF) or anti-cyclic citrullinated peptide antibody (anti-CCP) is negative. Here, we determined the application value of 14-3-3η protein, anti-carbamylated proteins antibodies (anti-CarP), as well as their potential role to diagnose RA together with RF or anti-CCP. METHOD: Serum levels of anti-CCP, RF, 14-3-3η and anti-CarP antibodies were detected in 291 RA patients, 223 patients with autoimmune diseases except RA, and 156 healthy subjects recruited from Han population of Northern China. We examined the differences in the levels of these indicators among groups and compared the correlations between any two of the indicators. At the same time, a total of 12 testing strategies were established for comparison to maximize the diagnostic value. RESULT: The levels of RF, anti-CCP, anti-CarP and 14-3-3η were significantly higher in RA patients (12.5;[9.36-15.7], 30.7;[25.7-35.6], 1.90;[1.70-2.01], 15.8;[10.8-20.8], respectively) compared with either interference-control group (1.24;[1.07-1.41], 0.64;[0.42-0.86], 0.51;[0.46-0.57], 0.33;[0.23-0.44], respectively) (p < 0.0001) or healthy-control group (1.03;[0.99-1.08], 0.49;[0.38-0.59], 0.28;[0.21-0.35], 0.55;[0.27-0.85], respectively) (p < 0.0001). Among all 12 detection strategies, the YI and κ value of a novel strategy that either double-positive of any 2 markers or single-positive of anti-CCP can be diagnosed as RA had the highest diagnostic value. CONCLUSION: The results of our study demonstrated that in Han population of Northern China, anti-CarP antibodies and 14-3-3η protein can be treated as valuable indicators of RA, especially when combined with RF and anti-CCP, the detection value is maximized.

Intratumoral distribution of YSNSG cyclopeptide in a mouse melanoma model using microdialysis.

The YSNSG peptide is a synthetic cyclopeptide targeting αvß3 integrin with antitumor activity. Previous study has determined main pharmacokinetic parameters in plasma and in tissue in healthy animals using microdialysis. First we aim to assess the impact of a 20 mg/kg dosage instead of 10 mg/kg in tumor growth inhibition. Secondly we aim to investigate the YSNSG peptide distribution in two different tumor regions in animals with melanoma. C57BL/6 mice were exposed at Days 8, 10 and 12 after melanoma cells implantation (B16F1) to different dosage of YSNSG peptide or control, respectively (n = 10 per group). Data analysis was performed at D16, 20 and 24 with a Nonlinear Mixed-Effects (NLME) approach. For pharmacokinetic study n = 8 mice (same disease condition) received YSNSG peptide by intravenous after insertion of two microdialysis probes in central peripheral region of tumor, respectively. Plasma and tissue samples were collected during 2 h. A non-compartmental analysis was performed to determine main pharmacokinetic parameters. There was a significant tumor growth inhibition in mice receiving 20 mg/kg vs Control (p < 0.02). Main plasma parameters were half-life elimination 25.8 ± 8.2 min, volume of distribution 11.9 ± 0.4 mL, clearance 19.8 ± 9.4 mL/h and area under the curve 1,173.6 µg.min/mL. Penetration rate of the YSNSG peptide from plasma to tumor tissue were 3.3 ± 2.1% and 3.4 ± 2.7% in central and peripheral, respectively. Contrary to subcutaneous distribution in healthy animals the distribution of the YSNSG peptide into tumoral tissue is low but seems non-heterogeneous between central and peripheral tumor region.

Autoantibodies as Diagnostic Markers and Mediator of Joint Inflammation in Arthritis.

Rheumatoid arthritis is a systemic, polygenic, and multifactorial syndrome characterized by erosive polyarthritis, damage to joint architecture, and presence of autoantibodies against several self-structures in the serum and synovial fluid. These autoantibodies (anticitrullinated protein/peptide antibodies (ACPAs), rheumatoid factors (RF), anticollagen type II antibodies, antiglucose-6 phosphate isomerase antibodies, anticarbamylated protein antibodies, and antiacetylated protein antibodies) have different characteristics, diagnostic/prognostic value, and pathological significance in RA patients. Some of these antibodies are present in the patients' serum several years before the onset of clinical disease. Various genetic and environmental factors are associated with autoantibody production against different autoantigenic targets. Both the activating and inhibitory FcγRs and the activation of different complement cascades contribute to the downstream effector functions in the antibody-mediated disease pathology. Interplay between several molecules (cytokines, chemokines, proteases, and inflammatory mediators) culminates in causing damage to the articular cartilage and bones. In addition, autoantibodies are proven to be useful disease markers for RA, and different diagnostic tools are being developed for early diagnosis of the clinical disease. Recently, a direct link was proposed between the presence of autoantibodies and bone erosion as well as in the induction of pain. In this review, the diagnostic value of autoantibodies, their synthesis and function as a mediator of joint inflammation, and the significance of IgG-Fc glycosylation are discussed.

Population Pharmacokinetic Modeling of VL-2397, a Novel Systemic Antifungal Agent: Analysis of a Single- and Multiple-Ascending-Dose Study in Healthy Subjects.

VL-2397, a novel, systemic antifungal agent, has potent in vitro and in vivo fungicidal activity against Aspergillus species. Plasma concentrations from a phase 1 study were used to construct a population pharmacokinetic (PPK) model for VL-2397. Healthy subjects aged 18 to 55 years received single doses of VL-2397, ranging from 3 to 1,200 mg, multiple daily doses of 300, 600, or 1,200 mg for 7 days, or 300 mg three times/day for 7 days followed by 600 mg daily for 21 days. Plasma samples were collected throughout the dosing intervals. Sixty-six subjects provided 1,908 concentrations. Drug concentrations over time were increased less than dose proportionally for doses above 30 mg. Dose-normalized concentrations plotted over time did not overlap. A 3-compartment nonlinear saturable binding model fit the data well. Clearance increased with dose, and mean values ranged from 0.4 liters/h at 3 mg to 8.5 liters/h at 1,200 mg. Mean volume in the central compartment ranged from 4.8 to 6.9 liters across doses. In the first 24 h, once-daily dosing results in a rapid decrease in concentrations by hour 16 to approximately 1 mg/liter, regardless of dose, with slow clearance over time. Administration of 300 mg every 8 h achieved concentrations above 1 mg/liter over an entire 24-h period. There was a significant relationship between body surface area and clearance. The data suggest that VL-2397 has nonlinear saturable binding kinetics. Protein binding is the likely primary source of the nonlinearity. The PPK model can now be used to optimize dosing by bridging the kinetics to efficacious pharmacodynamic targets.

Plasma cyclic glycine proline/IGF-1 ratio predicts clinical outcome and recovery in stroke patients.

OBJECTIVE: Many stroke patients make a partial recovery in function during the first 3 months, partially through promoting insulin-like growth factor-1 (IGF-1) function. A prognostic biomarker that associates with IGF-1 function may predict clinical outcome and recovery of stroke. This study evaluated plasma concentrations of IGF-1, IGF binding protein (IGFBP)-3 and cyclic-glycine-proline (cGP) and their associations with clinical outcome in stroke patients. METHODS: Thirty-four patients were recruited within 3 days of stroke. Clinical assessments included the National Institutes of Health Stroke Scale (NIHSS) within 3 days (baseline), and at days 7 and 90; the modified Rankin Scale (mRS) and Fugl-Meyer Upper-Limb Assessment Scale (FM-UL) at days 7 and 90. Plasma samples were collected from the patients at the baseline, days 7 and 90. Fifty age-matched control participants with no history of stroke were also recruited and provided plasma samples. IGF-1, IGFBP-3, and cGP concentrations were analyzed using ELISA or HPLC-MS. RESULTS: Baseline concentrations of IGFBP-3, cGP, and cGP/IGF-1 ratio were lower in stroke patients than the control group. The neurological scores of stroke patients were improved and plasma cGP and cGP/IGF-1 ratio increased over time. Baseline cGP/IGF-1 ratio was correlated with the NIHSS scores at day 90 and the changes in NIHSS scores from the baseline to 90 days. INTERPRETATION: Low cGP concentrations and cGP/IGF-1 ratio in stroke patients suggest an impaired IGF-1 function. The cGP/IGF-1 ratio at admission maybe further developed as a prognostic biomarker for stroke recovery.

Investigation of the Potential Use of Sialic Acid as a Biomarker for Rheumatoid Arthritis.

BACKGROUND: Sialic acids are a diverse family of sugar units attached to the outermost ends of sugar chains, which were demonstrated to be related to many diseases. We hypothesized sialic acids could be used as biomarkers for diagnosis of rheumatoid arthritis (RA). METHODS: The serum level of the main sialic acids, N-acetyl-D-neuraminic acid (Neu5Ac) in 163 RA patients and 50 healthy individuals were detected by pre-column derivatization-high performance liquid chromatography method. And three biomarkers to diagnose RA in clinic, C-reactive protein (CRP), rheumatoid factor (RF), and cyclic citrullinated peptides (CCP) in serum of these subjects were analyzed using enzyme-linked immunosorbent assay (ELISA). The data was analyzed using the receiver operating characteristic (ROC) curve and the Youden index. RESULTS: We found the concentration of Neu5Ac in RA group was significantly higher than healthy group. There was a positive correlation between the concentration of Neu5Ac and the RA disease activity score (DAS). The sensitivity and specificity of Neu5Ac were significantly greater than CRP, RF, CCP. CONCLUSIONS: Higher serum levels of Neu5Ac were significantly associated with the presence as well as severity of RA. Neu5Ac may be a potential biomarker for prediction and severity of RA in clinical practice.

Clinical characteristics of rheumatoid arthritis patients with peripheral neuropathy and potential related risk factors.

OBJECTIVES: To investigate potential risk factors of peripheral neuropathy (PN) in rheumatoid arthritis patients (RA). METHODS: Eighty-eight patients with RA were enrolled in this study, including patients with PN (n = 44; 28 patients with multiple nerves (MN) involvement and 16 patients with single nerve (SN) involvement) and without (n = 44) peripheral neuropathy were enrolled. Their clinical features were comprehensively collected including symptoms/signs, lab results, electromyogram data. T test or chi-squared test and further binary regression analysis were used to explore risk factors based on analyzing these clinical features. RESULTS: There was no difference as regards patients' age (59.50 ± 8.11 vs 58.68 ± 11.44 years), gender ratio (female/male, 29:15 vs 29:15), and disease duration (6.34 ± 7.87 vs 8.13 ± 9.52 months) between patients with and without PN. RA patients with PN had lower total protein (61.13 ± 7.06 vs 66.06 ± 6.44 g/L), anti-CCP levels (239.13 ± 203.77 vs 361.41 ± 168.09 U/ml) compared with control patients, while patients with MN had higher inflammatory parameters (white blood cells, platelet, C-reactive protein (CRP), erythrocyte sedimentation rate, rheumatoid factor) than patients with SN (p < 0.05). Low total protein (< 63 g/L, 30/44 vs. 12/44) and anti-CCP (< 285.7 U/ml, 27/44 vs. 11/44) were risk factors for patients with PN, while CRP (> 6 mg/L, 26/28 vs. 6/16) and PLT (> 243 × 109/L, 25/28 vs.5/16) were related to the development of MN. CONCLUSIONS: RA patients with PN, especially MN can present various clinical symptoms, which will aggravate patients' anxiety and depression status. The increase of blood platelet, and CRP levels and decrease of blood albumin are probable risk factors for PN in RA patients.

Porphyromonas gingivalis experimentally induces periodontis and an anti-CCP2-associated arthritis in the rat.

OBJECTIVES: Association between periodontal disease (PD) and rheumatoid arthritis (RA) has been extensively described, but direct evidence of causal involvement of PD in RA is missing. We investigated the priming role of oral Porphyromonas gingivalis (P. gingivalis) in PD and subsequent RA and we assessed biomarkers of bone resorption and arthritis development in rats. METHODS: Lewis rats were orally exposed to either P. gingivalis, Prevotella intermedia or control gel for 1 month and then followed for 8 months. The onset and development of PD was assessed by serology, gingivitis severity and micro-CT (µCT). We investigated arthritis development using circulating proinflammatory markers, anticyclic citrullinated peptide (CCP), anticitrullinated protein antibody (ACPA), ankle histology and µCT. RESULTS: PD was only observed in the P. gingivalis treated rats, as early as 1 month postexposure. Joint and systemic inflammation were detected only in the P. gingivalis group after 4 and 8 months. At 8 months, inflammatory cell infiltrate was observed in ankle joints and paralleled cortical erosions and overall cortical bone reduction. Furthermore, anti-CCP2 correlated with local and systemic bone loss. CONCLUSIONS: In our long-term study, PD induced by oral exposure to P. gingivalis triggered seropositive arthritis, with systemic inflammation and bone erosions. This is the first in vivo demonstration of arthritis induced by oral priming with P. gingivalis.