Proinflammatory polarization strongly reduces human macrophage in vitro phagocytosis of tumor cells in response to CD47 blockade.

Antibody-based CD47 blockade aims to activate macrophage phagocytosis of tumor cells. However, macrophages possess a high degree of phenotype heterogeneity that likely influences phagocytic capacity. In murine models, proinflammatory (M1) activation increases macrophage phagocytosis of tumor cells, but in human models, results have been conflicting. Here, we investigated the effects of proinflammatory polarization on the phagocytic response of human monocyte-derived macrophages in an in vitro model. Using both flow cytometry-based and fluorescence live-cell imaging-based phagocytosis assays, we observed that mouse monoclonal anti-CD47 antibody (B6H12) induced monocyte-derived macrophage phagocytosis of cancer cells in vitro. Proinflammatory (M1) macrophage polarization with IFN-γ+LPS resulted in a severe reduction in phagocytic response to CD47 blockade. This reduction coincided with increased expression of the antiphagocytic membrane proteins LILRB1 and Siglec-10 but was not rescued by combination blockade of the corresponding ligands. However, matrix metalloproteinase inhibitors (TAPI-0 or GM6001) partly restored response to CD47 blockade in a dose-dependent manner. In summary, these data suggest that proinflammatory (M1) activation reduces phagocytic response to CD47 blockade in human monocyte-derived macrophages.

Circulating TREM2 as a noninvasive diagnostic biomarker for NASH in patients with elevated liver stiffness.

BACKGROUND AND AIMS: Reliable noninvasive biomarkers are an unmet clinical need for the diagnosis of NASH. This study investigates the diagnostic accuracy of the circulating triggering receptor expressed on myeloid cells 2 (plasma TREM2) as a biomarker for NASH in patients with NAFLD and elevated liver stiffness. APPROACH AND RESULTS: We collected cross-sectional, clinical data including liver biopsies from a derivation ( n = 48) and a validation cohort ( n = 170) of patients with elevated liver stiffness measurement (LSM ≥ 8.0 kPa). Patients with NAFLD activity scores (NAS) ≥4 were defined as having NASH. Plasma TREM2 levels were significantly elevated in patients with NASH of the derivation cohort, with an area under the receiver operating characteristics curve (AUROC) of 0.92 (95% confidence interval [CI], 0.84-0.99). In the validation cohort, plasma TREM2 level increased approximately two-fold in patients with NASH, and a strong diagnostic accuracy was confirmed (AUROC, 0.83; 95% CI, 0.77-0.89; p < 0.0001). Plasma TREM2 levels were associated with the individual histologic features of NAS: steatosis, lobular inflammation, and ballooning ( p < 0.0001), but only weakly with fibrosis stages. Dual cutoffs for rule-in and rule-out were explored: a plasma TREM2 level of ≤38 ng/ml was found to be an optimal NASH rule-out cutoff (sensitivity 90%; specificity 52%), whereas a plasma TREM2 level of ≥65 ng/ml was an optimal NASH rule-in cutoff (specificity 89%; sensitivity 54%). CONCLUSIONS: Plasma TREM2 is a plausible individual biomarker that can rule-in or rule-out the presence of NASH with high accuracy and thus has the potential to reduce the need for liver biopsies and to identify patients who are eligible for clinical trials in NASH.

Characterization of DNA-protein complexes by nanoparticle tracking analysis and their association with systemic lupus erythematosus.

Nanotechnology enables investigations of single biomacromolecules, but technical challenges have limited the application in liquid biopsies, for example, blood plasma. Nonetheless, tools to characterize single molecular species in such samples represent a significant unmet need with the increasing appreciation of the physiological importance of protein structural changes at nanometer scale. Mannose-binding lectin (MBL) is an oligomeric plasma protein and part of the innate immune system through its ability to activate complement. MBL also serves a role as a scavenger for cellular debris, especially DNA. This may link functions of MBL with several inflammatory diseases in which cell-free DNA now appears to play a role, but mechanistic insight has been lacking. By making nanoparticle tracking analysis possible in human plasma, we now show that superoligomeric structures of MBL form nanoparticles with DNA. These oligomers correlate with disease activity in systemic lupus erythematosus patients. With the direct quantification of the hydrodynamic radius, calculations following the principles of Taylor dispersion in the blood stream connect the size of these complexes to endothelial inflammation, which is among the most important morbidities in lupus. Mechanistic insight from an animal model of lupus supported that DNA-stabilized superoligomers stimulate the formation of germinal center B cells and drive loss of immunological tolerance. The formation involves an inverse relationship between the concentration of MBL superoligomers and antibodies to double-stranded DNA. Our approach implicates the structure of DNA-protein nanoparticulates in the pathobiology of autoimmune diseases.

Comparison of culture media reveals that non-essential amino acids strongly affect the phenotype of human monocyte-derived macrophages.

Macrophages are important innate immune cells with the ability to adapt their phenotype to environmental cues. Research on human macrophages often uses monocyte-derived macrophages cultured in vitro, but it is unclear if culture medium affects macrophage phenotype. The objective of this study was to determine the impact of culture medium composition on monocyte-derived macrophage phenotype. Monocyte-derived macrophages were generated in different formulations of culture media (RPMI 1640, DMEM, MEM, McCoy's 5a and IMDM). Viability, yield and cell size were monitored, and RT-qPCR, flow cytometry or ELISA was used to compare levels of phenotype markers (CD163, CD206, CD80, TNFα, IL-10, SIRPα, LILRB1 and Siglec-10). Yield, cell size, gene expression, membrane protein levels and release of soluble proteins were all affected by changes in culture medium composition. The most pronounced effects were observed after culture in DMEM, which lacks the non-essential amino acids asparagine, aspartic acid, glutamic acid and proline. Supplementation of DMEM with non-essential amino acids either fully or partly reversed most effects of DMEM on macrophage phenotype. The results suggest culture medium composition and amino acid availability affect the phenotype of human monocyte-derived macrophages cultured in vitro.

Soluble CD163: a novel independent prognostic biomarker in patients with metastatic renal cell carcinoma.

The hemoglobin-haptoglobin scavenger receptor CD163 is present in both a membrane-bound form on monocytes and macrophages (mCD163) and a shed soluble circulating form (sCD163). CD163 is a well-described marker of M2-like tumor-associated macrophages, but in patients with metastatic renal cell carcinoma (mRCC), monocyte mCD163 and serum sCD163 levels have not previously been investigated and associated with patient overall survival (OS). Here, we report mCD163 expression on peripheral blood monocytes, as well as sCD163 serum levels, in samples from 89 patients newly diagnosed with mRCC and 20 healthy controls. We found that in mRCC patients, compared to healthy controls, monocyte mCD163 levels were reduced (P < 0.001) whereas serum sCD163 levels were increased (P = 0.004). Moreover, an inverse correlation between mCD163 and sCD163 levels (P = 0.04) was shown. In survival analyses, intermediary levels of monocyte mCD163 were associated with longest OS, compared to both lower and higher mCD163 levels, which were both associated with worse outcomes (P < 0.01). Further, higher levels of sCD163 at diagnosis were associated with poor OS in both univariate (P < 0.001) and multivariate analysis (HR = 1.28; 95%CI 1.09-1.50, P = 0.002). Importantly, stratification by low vs. high sCD163 was able to separate patients with International Metastatic RCC Database Consortium (IMDC) intermediate risk (IMDCINT) into two subgroups with different OS (P = 0.03): IMDCINT-sCD163LOW showed survival similar to IMDCFAV patients, and IMDCINT-sCD163HIGH showed survival similar to IMDCPOOR patients. Thus, baseline sCD163 is a novel independent biomarker of OS in mRCC, and using sCD163 as an add-on biomarker may improve prognostic value for patients in the heterogenous IMDC intermediate group.

Neoehrlichia mikurensis-An emerging opportunistic tick-borne infection in immunosuppressed patients.

BACKGROUND: Neoehrlichia mikurensis (N. mikurensis) is a newly discovered tick-borne pathogen that can inflict life-threatening illness in immunocompromised patients. N. mikurensis infection is only detectable by polymerase chain reaction (PCR)-based methodologies. We describe three distinct clinical manifestations of N. mikurensis infection (neoehrlichiosis) in Danish patients receiving B-lymphocyte-depleting therapy, rituximab, for underlying hematological, rheumatological, or neurological disorders. All three patients went through a protracted pre-diagnostic period. METHODS: N. mikurensis DNA was detected and confirmed using two methods. Blood was tested by specific real-time PCR targeting the groEL gene and by 16S and 18S profiling followed by sequencing. Bone marrow was analyzed by 16S and 18S profiling. RESULTS: N. mikurensis was detected in blood samples in all three cases and in bone marrow from one of the three. The severity of the symptoms ranged from prolonged fever lasting more than 6 months to life-threatening hyperinflammation in the form of hemophagocytic lymphohistiocytosis (HLH). Interestingly, all patients presented with splenomegaly and two with hepatomegaly. After starting doxycycline therapy, symptoms were relieved within a few days, and biochemistry and organomegaly quickly normalized. CONCLUSION: We present three Danish patients recognized by the same clinician over a period of 6 months, strongly suggesting that many cases are going unrecognized. Second, we describe the first case of N. mikurensis-induced HLH and emphasize the potential severity of undetected neoehrlichiosis.

Fungal translocation measured by serum 1,3-ß-D-glucan correlates with severity and outcome of liver cirrhosis-A pilot study.

BACKGROUND & AIMS: On a global scale, liver cirrhosis is attributable to ~1 million deaths per year. This systemic disease comes along with diverse sequelae, including microbiota alterations, increased gut permeability and translocation of microbial components into the systemic circulation. Alongside the extensively studied influence of bacterial translocation and its host-pathogen interactions, far less is known about the role and impact of fungal components once having crossed the intestinal barrier. METHODS: Including 70 patients with different aetiologies of liver cirrhosis, we investigated the relationship between fungal translocation, measured by 1,3-ß-D-glucan (BDG), and biomarkers of gut integrity, inflammation and severity/outcome of liver disease. RESULTS: Patients with cirrhosis Child-Pugh class (CPC)-B were more likely to have positive serum BDG (aOR 5.4, 95% CI 1.2-25.2) compared to patients with cirrhosis CPC-A. BDG showed a moderate positive correlation with several markers of inflammation (sCD206, sCD163, Interleukin 8, interferon-gamma-induced protein). Mortality differed significantly between patients with positive versus negative BDG (log-rank test, p = 0.015). The multivariable Cox regression model yielded an aHR of 6.8 (95% CI 1.8-26.3). DISCUSSION: We observed trends for increased fungal translocation depending on the severity of liver cirrhosis, an association of BDG with an inflammatory environment and the adverse effects of BDG on disease outcome. In order to gain more in-depth knowledge about (fungal-)dysbiosis and its detrimental consequences in the setting of liver cirrhosis, these trends need to be studied in more detail including prospective sequential testing in larger cohorts together with mycobiome analyses. This will further elucidate complex host-pathogen interactions and potentially introduce points of application for therapeutic interventions.

Acculturation and risk of traffic crashes in young Asian-born Australian drivers.

The study examines changes over time in crash risk differences between young Australian drivers born in Asia and those born in Australia.Data from the 2003 baseline survey of the DRIVE cohort of 20 806 young drivers aged 17-24 years were linked to police, hospital and death data up until 2016. The association between country of birth and crash was investigated using flexible parametric survival models adjusted for confounders.Six months after baseline, the crash risk in Asian-born drivers was less than half that of their Australian-born counterparts (mean HR, MHR 0.41; 95% CI 0.29 to 0.57), only to increase steadily over time to resemble that of Australian-born drivers 13 years later (MHR 0.94; 95% CI 0.66 to 1.36).This is likely to be associated with acculturation and the adoption by young Asian-born Australian drivers of driving behaviour patterns akin to those born locally. This needs to be considered in future road safety campaigns.

Self-harm in adolescence and risk of crash: a 13-year cohort study of novice drivers in New South Wales, Australia.

INTRODUCTION: Self-harm and suicide are leading causes of morbidity and death for young people, worldwide. Previous research has identified self-harm is a risk factor for vehicle crashes, however, there is a lack of long-term crash data post licensing that investigates this relationship. We aimed to determine whether adolescent self-harm persists as crash risk factor in adulthood. METHODS: We followed 20 806 newly licensed adolescent and young adult drivers in the DRIVE prospective cohort for 13 years to examine whether self-harm was a risk factor for vehicle crashes. The association between self-harm and crash was analysed using cumulative incidence curves investigating time to first crash and quantified using negative binominal regression models adjusted for driver demographics and conventional crash risk factors. RESULTS: Adolescents who reported self-harm at baseline were at increased risk of crashes 13 years later than those reporting no self-harm (relative risk (RR) 1.29: 95% CI 1.14 to 1.47). This risk remained after controlling for driver experience, demographic characteristics and known risk factors for crashes, including alcohol use and risk taking behaviour (RR 1.23: 95% CI 1.08 to 1.39). Sensation seeking had an additive effect on the association between self-harm and single-vehicle crashes (relative excess risk due to interaction 0.87: 95% CI 0.07 to 1.67), but not for other types of crashes. DISCUSSION: Our findings add to the growing body of evidence that self-harm during adolescence predicts a range of poorer health outcomes, including motor vehicle crash risks that warrant further investigation and consideration in road safety interventions. Complex interventions addressing self-harm in adolescence, as well as road safety and substance use, are critical for preventing health harming behaviours across the life course.

STAT3 is over-activated within CD163pos bone marrow macrophages in both Multiple Myeloma and the benign pre-condition MGUS.

Tumour-associated macrophages (TAMs) support cancer cell survival and suppress anti-tumour immunity. Tumour infiltration by CD163pos TAMs is associated with poor outcome in several human malignancies, including multiple myeloma (MM). Signal transducer and activator of transcription 3 (STAT3) is over-activated in human cancers, and specifically within TAMs activation of STAT3 may induce an immunosuppressive (M2-like) phenotype. Therefore, STAT3-inhibition in TAMs may be a future therapeutic strategy.We investigated TAM markers CD163, CD206, and activated STAT3 (pSTAT3) in patients with MGUS (n = 32) and MM (n = 45), as well as healthy controls (HCs, n = 13).Blood levels of the macrophage biomarkers sCD163 and sCD206, and circulating cytokines, as well as bone marrow mRNA expression of CD163 and CD206, were generally increased in MGUS and MM patients, compared to HCs, but to highly similar levels. By immunohistochemistry, bone marrow levels of pSTAT3 were increased specifically within CD163pos cells in both MGUS and MM patients.In conclusion, macrophage-related inflammatory changes, including activation of STAT3, were present already at the MGUS stage, at similar levels as in MM. Specific increase in pSTAT3 levels within CD163pos cells supports that the CD163 scavenger receptor may be a useful target for future delivery of STAT3-inhibitory drugs to TAMs in MM patients.

Cytokine profiles and the dynamic of gingivitis development in humans.

AIM: To investigate the relationship between cytokine profiles and "fast" and "slow" patterns of gingival inflammation development. MATERIALS AND METHODS: Forty-two adults participated in an experimental gingivitis study, comprising a 2-week hygiene phase (clinical examination and professional cleaning); a 3-week induction phase (absence of oral hygiene); and a 2-week resolution phase (re-establishment of oral hygiene). Plaque and gingival inflammation scores were assessed. Interferon-gamma (IFN-γ), interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, and tumour necrosis factor-alpha (TNF-α) from gingival crevicular fluid were collected and measured by multiplex ELISA. Group-based-trajectory-modelling (GBTM) was used to model cytokine profiles over the induction phase. The effect of gingival inflammation on cytokine levels over time was estimated with mixed-effects modelling. RESULTS: GBTM analysis revealed two cytokine profiles, "non-organized response" (IL-4, IL-6, IL-8, IL-12, and IL-13) and "organized response" (IL-2, IL-10, and TNF-α). Among the "slow" responders, neither cytokine profile was associated with gingivitis. In contrast, a "fast" response was associated with a higher "non-organized response" factor (coef. 0.14) and a lower "organized response" factor (coef. -0.03). CONCLUSION: A "fast" gingivitis development was associated with a higher "non-organized response" and a lower "organized response", which may elucidate the role of individual variability in gingivitis susceptibility.

Haptoglobin-related protein in human plasma correlates to haptoglobin concentrations and phenotypes.

Haptoglobin-related protein (Hpr) is a plasma protein with high sequence similarity to haptoglobin (Hp). Like Hp, Hpr also binds hemoglobin (Hb) with high affinity, but it does not bind to the Hb-Hp receptor CD163 on macrophages. The Hpr concentration is markedly lower than Hp in plasma and its regulation is not understood. In the present study, we have developed non-crossreactive antibodies to Hpr to analyze the Hpr concentration in 112 plasma samples from anonymized individuals and compared it to Hp. The results show that plasma Hpr correlated with Hp concentrations (rho = 0.46, p = .0001). Hpr accounts for on average 0.35% of the Hp/Hpr pool but up to 29% at low Hp levels. Furthermore, the Hpr concentrations were significantly lower in individuals with the Hp2-2 phenotype compared to those with the Hp2-1 or Hp1-1 phenotypes. Experimental binding analysis did not provide evidence that Hpr associates with Hp and in this way is removed via CD163. In conclusion, the Hpr concentration correlates to Hp concentrations and Hp-phenotypes by yet unknown mechanisms independent of CD163-mediated removal of Hb-Hp complexes.

Re-evalution of the measurement of haptoglobin in human plasma samples.

Haptoglobin (Hp) is an abundant plasma protein scavenging hemoglobin (Hb) via CD163 on macrophages. This process consumes Hp, which therefore negatively correlates to hemolysis. However, exact measurements of Hp plasma levels are complicated by different phenotypes (Hp1-1, Hp2-1, and Hp2-2) forming different oligomeric states with differences in immunoreactivity. In addition, humans have an immune-cross-reactive Hp-related protein. In the present study, we developed Hp-specific monoclonal antibodies for an accurate Hp analysis of the different Hp phenotypes in a panel of 112 anonymous samples from hospitalized individuals subjected to routine Hp immunoturbidimetric measurements. The data revealed immunoturbidimetry as a reliable method in most cases but also that the use of non-phenotype-specific calibrators leads to substantial bias in the measurement of the Hp-concentration, non at least in Hp1-1 individuals. Furthermore, analysis of the Hb-dependence of the CD163 interaction with Hp1-1 and Hp2-2 showed that a higher 'cost-effectiveness' in the consumption of dimeric Hp1-1 versus multimeric Hp phenotypes is a likely contribution to the observed differences in the plasma levels of the Hp phenotypes. In conclusion, the determination of Hp phenotype and the use of phenotype-specific calibrators are essential to obtain a precise estimate of the Hp level in healthy and diseased individuals.

Driving offences and risk of subsequent crash in novice drivers: the DRIVE cohort study 12-year follow-up.

BACKGROUND: Penalties are a key component to improve road user safety, but previous studies suggested that they might not be successful in reducing crashing in offending drivers. However, these studies were not able to consider important crash risk factors in the analysis that might confound the results. Using data from a large prospective cohort study of young drivers in New South Wales, Australia, we explored if novice drivers with driving offences have a higher rates of car crash and if these differences are explained by established crash risk factors. METHODS: We used data from a 2003/2004 Australian survey of young drivers, linked to police reported offence and crash data, hospital data and deaths data up to 2016. We used Poisson regression models adjusted for confounders to estimate the association between driving offences during 2003-2006 with car crash during 2007-2016. RESULTS: The study cohort comprised 20 781 young drivers of whom 7860 drivers (37.8%) had at least one driving offence and 2487 (12.0%) were involved in at least one crash. After adjusting for confounders in the regression model, drivers with three or more driving offences had 2.25 (95% CI 1.98 to 2.57), 2.87 (95% CI 1.60 to 5.17) and 3.28 (95% CI 2.28 to 4.72) times higher rates of any crash, crashes that resulted in hospital admission or death and single vehicle crashes compared with drivers with no driving offences. CONCLUSION: Measures that successfully mitigate the underlying risk factors for both, crashes and offences, have the potential to improve road safety.

Metabolism of 25-Hydroxy-Vitamin D in Human Macrophages Is Highly Dependent on Macrophage Polarization.

Macrophages synthesize active vitamin D (1,25-dihydroxy-vitamin D) and express the vitamin D receptor in the nucleus; however, vitamin D metabolism in relation to macrophage polarization and function is not well understood. We studied monocyte-derived macrophages (MDMs) from human buffy coats polarized into M0, M1 (LPS + IFNγ), M2a (IL4 + IL13) and M2c (IL10) macrophage subtypes stimulated with 25-hydroxy-vitamin D (1000 and 10,000 nanomolar). We measured vitamin D metabolites (25-hydroxy-vitamin D, 1,25-dihydroxy-vitamin D, 24,25-dihydroxy-vitamin D and 3-epi-25-hydroxy-vitamin D) in cell media with liquid chromatography-mass spectrometry-mass spectrometry. The mRNA expression (CYP27B1, CYP24A1 and CYP24A1-SV) was measured with qPCR. We found that reparative MDMs (M2a) had significantly more 1,25-dihydroxy-vitamin D compared to the other MDMs (M0, M1 and M2c). All MDMs were able to produce 3-epi-25-hydroxy-vitamin D, but this pathway was almost completely attenuated in inflammatory M1 MDMs. All MDM subtypes degraded vitamin D through the 24-hydroxylase pathway, although M1 MDMs mainly expressed an inactive splice variant of CYP24A1, coding the degrading enzyme. In conclusion, this study shows that vitamin D metabolism is highly dependent on macrophage polarization and that the C3-epimerase pathway for vitamin D is active in macrophages.

Severe Acute Respiratory Syndrome Coronavirus 2 Seroprevalence Survey Among 17 971 Healthcare and Administrative Personnel at Hospitals, Prehospital Services, and Specialist Practitioners in the Central Denmark Region.

BACKGROUND: The objective of this study was to perform a seroprevalence survey on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) among Danish healthcare workers to identify high-risk groups. METHODS: All healthcare workers and administrative personnel at the 7 hospitals, prehospital services, and specialist practitioner clinics in the Central Denmark Region were invited to be tested by a commercial SARS-CoV-2 total antibody enzyme-linked immunosorbent assay (Wantai Biological Pharmacy Enterprise Co, Ltd, Beijing, China). RESULTS: A total of 25 950 participants were invited. Of these, 17 971 had samples available for SARS-CoV-2 antibody testing. After adjustment for assay sensitivity and specificity, the overall seroprevalence was 3.4% (95% confidence interval [CI], 2.5%-3.8%). The seroprevalence was higher in the western part of the region than in the eastern part (11.9% vs 1.2%; difference: 10.7 percentage points [95% CI, 9.5-12.2]). In the high-prevalence area, the emergency departments had the highest seroprevalence (29.7%), whereas departments without patients or with limited patient contact had the lowest seroprevalence (2.2%). Among the total 668 seropositive participants, 433 (64.8%) had previously been tested for SARS-CoV-2 RNA, and 50.0% had a positive reverse-transcription polymerase chain reaction (PCR) result. CONCLUSIONS: We found large differences in the prevalence of SARS-CoV-2 antibodies in staff working in the healthcare sector within a small geographical area of Denmark. Half of all seropositive staff had been tested positive by PCR prior to this survey. This study raises awareness of precautions that should be taken to avoid in-hospital transmission. Regular testing of healthcare workers for SARS-CoV-2 should be considered to identify areas with increased transmission.

Soluble PD-1 (sPD-1) is expressed in human macrophages.

The PD-1/PD-L1 axis plays a crucial role in regulating the anti-tumour immune response. A soluble PD-1 protein (sPD-1) has previously been observed, which could block the binding of PD-L1 to PD-1. Tumour associated macrophages are abundant in tumours, and evidence suggest they express PD-1. However, whether they also express sPD-1 remains unclear. The objective of this study was to investigate expression of sPD-1 in two in vitro models of human macrophages: THP-1 cells and monocyte-derived macrophages (MDM). Cells were polarised with either LPS + IFN-γ or IL-4 + IL-13 or left unpolarised. PD-1 and sPD-1 mRNAs were measured using droplet digital PCR, sPD-1 protein by electrochemiluminescence immunoassay and PD-1 by flow cytometry. sPD-1 mRNA was induced in both THP-1 cells and MDM after polarisation with LPS + IFN-γ, while IL-4 + IL-13 induced sPD-1 mRNA in MDM only. sPD-1 protein was measurable in culture supernatants. These findings show that macrophages can be induced to express sPD-1.

Soluble CD163 Changes Indicate Monocyte Association With Cognitive Deficits in Parkinson's Disease.

BACKGROUND: Parkinson's disease (PD) is a neurodegenerative disorder with a significant immune component, as demonstrated by changes in immune biomarkers in patients' biofluids. However, which specific cells are responsible for those changes is unclear because most immune biomarkers can be produced by various cell types. OBJECTIVES: The aim of this study was to explore monocyte involvement in PD. METHODS: We investigated the monocyte-specific biomarker sCD163, the soluble form of the receptor CD163, in cerebrospinal fluid (CSF) and serum in two experiments, and compared it with other biomarkers and clinical data. Potential connections between CD163 and alpha-synuclein were studied in vitro. RESULTS: CSF-sCD163 increased in late-stage PD and correlated with the PD biomarkers alpha-synuclein, Tau, and phosphorylated Tau, whereas it inversely correlated with the patients' cognitive scores, supporting monocyte involvement in neurodegeneration and cognition in PD. Serum-sCD163 increased only in female patients, suggesting a sex-distinctive monocyte response. CSF-sCD163 also correlated with molecules associated with adaptive and innate immune system activation and with immune cell recruitment to the brain. Serum-sCD163 correlated with proinflammatory cytokines and acute-phase proteins, suggesting a relation to chronic systemic inflammation. Our in vitro study showed that alpha-synuclein activates macrophages and induces shedding of sCD163, which in turn enhances alpha-synuclein uptake by myeloid cells, potentially participating in its clearance. CONCLUSIONS: Our data present sCD163 as a potential cognition-related biomarker in PD and suggest a role for monocytes in both peripheral and brain immune responses. This may be directly related to alpha-synuclein's proinflammatory capacity but could also have consequences for alpha-synuclein processing. © 2020 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Risky youth to risky adults: Sustained increased risk of crash in the DRIVE study 13 years on.

The objective of this study was to investigate if drivers who exhibit risky driving behaviours during youth (aged 17-24 years) have an increased risk of car crash up to 13 years later. We used data from the DRIVE study, a 2003/04 survey of 20,806 young novice drivers in New South Wales, Australia. The data were linked with police crash, hospital and deaths data up to 2016. We analysed differences in crash associated with 13 items of risky driving behaviours using negative binominal regression models adjusted for driver demographics, driving exposure and known crash risk factors. The items were summarised in one index and grouped into quintiles for the analysis. After adjusting for confounding, drivers of the third (RR 1.16, 95% CI 1.05-1.30), fourth (RR1.22, 95% CI1.09-1.36) and fifth quintile (RR 1.36, 95% CI 1.21-1.53) had higher crash rates compared to the lowest risk-takers. Drivers with the highest scores on the risky driving measure had higher rates of crash related hospital admission or death (RR 1.92, 95% CI 1.13-3.27), crashes in wet conditions (RR 1.35,95% CI 1.05-1.73), crashes in darkness (RR 1.55, 95% CI 1.25-1.93) and head-on crashes (RR 2.14, 95% CI 1.07-4.28), compared with drivers with the lowest scores. Novice adolescent drivers who reported high levels of risky driving when they first obtained a driver licence remained at increased risk of crash well into adulthood. Measures that successfully reduce early risky driving, have the potential to substantially reduce road crashes and transport related injuries and deaths over the lifespan.

Urine soluble CD163 (sCD163) as biomarker in glomerulonephritis: stability, reference interval and diagnostic performance.

OBJECTIVES: Soluble (s) CD163 is a well-established macrophage biomarker, and recent data suggests urine sCD163 to reflect disease activity in crescentic glomerulonephritis (GN). Other types of GN may also be associated with glomerular inflammation but the potential usefulness of urine sCD163 as a general biomarker of GN remains unaddressed. METHODS: An in-house sCD163 enzyme-linked immunosorbent assay (ELISA) was validated for urinary use and compared to a frequently used commercial ELISA. The pre-analytical stability of urine sCD163 was assessed and a reference interval was established according to the CLSI guidelines using specimens from 253 healthy individuals. Urine samples from 64 patients with different types of renal disorders were also analysed. RESULTS: Urine sCD163 was highly stable during storage. An upper reference limit of 5.1 µg/L (1.9 µg/mmol, normalised to creatinine) was established using the in-house ELISA. Urine sCD163 was generally increased in GN patients (3.9 µg/mmol, p<0.0001, AUROC=0.97) and decreased upon treatment, but did not perform better than urine albumin (AUROC=1.00). Patients with proliferative GN had higher urine sCD163/albumin (p=0.0001) ratio. The commercial assay had a higher detection limit, and patient levels were 4-6 times lower than in the in-house assay. CONCLUSIONS: Urine sCD163 is a stable biomarker that can be measured with acceptable accuracy using our in-house ELISA. Its pre-analytical characteristics makes it a reliable biomarker and our findings point towards the use of urine sCD163 as a biomarker of specific subtypes of GN.