Nanna Larsen

Amniotic fluid inflammatory cytokines: Potential markers of immunologic dysfunction in autism spectrum disorders

Abstract Objectives. The aim of the study was to analyze cytokine profiles in amniotic fluid (AF) samples of children developing autism spectrum disorders (ASD) and controls, adjusting for maternal autoimmune disorders and maternal infections during pregnancy. Methods. AF samples of 331 ASD cases and 698 controls were analyzed for inflammatory cytokines using Luminex xMAP technology utilizing a historic birth cohort. Clinical data were retrieved from nationwide registers, and case-control differences in AF cytokine levels were assessed using chi-square tests, logistic and tobit regression models. Results. Overall, individuals with ASD had significantly elevated AF levels of TNF-α and TNF-β compared to controls. Analyzing individuals diagnosed only with ICD-10 codes yielded significantly elevated levels of IL-4, IL-10, TNF-α and TNF-β in ASD patients. Restricting analysis to infantile autism cases showed significantly elevated levels of IL-4, TNF-α and TNF-β compared to controls with no psychiatric comorbidities. Elevated levels of IL-6 and IL-5 were found in individuals with other childhood psychiatric disorders (OCPD) when compared to controls with no psychiatric comorbidities. Conclusions. AF samples of individuals with ASD or OCPD showed differential cytokine profiles compared to frequency-matched controls. Further studies to examine the specificity of the reported cytokine profiles in ASD and OCPD are required.

Amniotic fluid chemokines and autism spectrum disorders: An exploratory study utilizing a Danish Historic Birth Cohort

INTRODUCTION Elevated levels of chemokines have been reported in plasma and brain tissue of individuals with Autism Spectrum Disorders (ASD). The aim of this study was to examine chemokine levels in amniotic fluid (AF) samples of individuals diagnosed with ASD and their controls. MATERIAL AND METHODS A Danish Historic Birth Cohort (HBC) kept at Statens Serum Institute, Copenhagen was utilized. Using data from Danish nation-wide health registers, a case-control study design of 414 cases and 820 controls was adopted. Levels of MCP-1, MIP-1α and RANTES were analyzed using Luminex xMAP technology. Case-control differences were assessed as dichotomized at below the 10th percentile or above the 90th percentile cut-off points derived from the control biomarker distributions (logistic regression) or continuous measures (tobit regression). RESULTS AND CONCLUSION AF volume for 331 cases and 698 controls was sufficient for Luminex analysis. Including all individuals in the cohort yielded no significant differences in chemokine levels in cases versus controls. Logistic regression analyses, performed on individuals diagnosed using ICD-10 only, showed increased risk for ASD with elevated MCP-1 (elevated 90th percentile adjusted OR: 2.32 [95% CI: 1.17-4.61]) compared to controls. An increased risk for infantile autism with elevated MCP-1 was also found (adjusted OR: 2.28 [95% CI: 1.16-4.48]). Elevated levels of MCP-1 may decipher an etiologic immunologic dysfunction or play rather an indirect role in the pathophysiology of ASD. Further studies to confirm its role and to identify the potential pathways through which MCP-1 may contribute to the development of ASD are necessary.

Heterologous Immunological Effects of Early BCG Vaccination in Low-Birth-Weight Infants in Guinea-Bissau: A Randomized-controlled Trial

Background. Bacillus Calmette–Guérin (BCG) seems to have beneficial nonspecific effects; early BCG vaccination of low-birth-weight (LBW) newborns reduces neonatal mortality by >40% due to prevention of primarily septicemia and pneumonia. Methods. Within a randomized trial in LBW infants in Guinea-Bissau of early BCG vs the usual postponed BCG, a subgroup was bled 4 weeks after randomization. Levels of interleukin (IL)-1β, IL-5, IL-6, IL-10, IL-17, interferon (IFN)-γ and tumor necrosis factor (TNF)-α were measured from whole-blood assays stimulated with innate agonists to Toll-like receptor (TLR)-2, -4 or -7/8, or purified protein derivative (PPD). Results. Among 467 infants, BCG significantly increased the in vitro cytokine responses to purified protein derivative of Mycobacterium tuberculosis (PPD), as expected. BCG was also associated with increased responses to heterologous innate stimulation, particularly of the cytokines IL-1β, IL-6, TNF-α, and IFN-γ. Conclusion. Four weeks after immunization, BCG-vaccinated infants have a significantly increased production of cytokines upon heterologous challenge, particularly T helper cell type 1 polarizing and typically monocyte-derived pro-inflammatory cytokines. BCG may accelerate the development of the neonatal immune system, mediating comprehensive protection against infections and mortality.

Neonatal levels of cytokines and risk of autism spectrum disorders: An exploratory register-based historic birth cohort study utilizing the Danish Newborn Screening Biobank

The aim of the study was to analyze cytokine profiles in neonatal dried blood samples (n-DBSS) retrieved from The Danish Newborn Screening Biobank of children developing Autism Spectrum Disorders (ASD) later in life and controls. Samples of 359 ASD cases and 741 controls were analyzed using Luminex xMAP technology and clinical data were retrieved from nationwide registers. Findings showed that children developing ASD were more likely to have decreased levels of both T helper-1(Th-1)-like cytokines (i.e. IFN-γ) and Th-2like cytokines (i.e. IL-4, IL-10) which may suggest a depressed or hypoactive immune cell activity during neonatal period in ASD.

Amniotic fluid MMP-9 and neurotrophins in autism spectrum disorders: an exploratory study.

Evidence suggests that some developmental disorders, such as autism spectrum disorders (ASDs), are caused by errors in brain plasticity. Given the important role of matrix metalloproteinases (MMPs) and neurotrophins (NTs) in neuroplasticity, amniotic fluid samples for 331 ASD cases and 698 frequency‐matched controls were analyzed for levels of MMP‐9, brain‐derived neurotrophic factor, NT‐4 and transforming growth factor‐β utilizing a Danish historic birth cohort and Danish nationwide health registers. Laboratory measurements were performed using an in‐house multiplex sandwich immunoassay Luminex xMAP method, and measurements were analyzed using tobit and logistic regression. Results showed elevated levels of MMP‐9 in ASD cases compared with controls (crude and adjusted tobit regression P‐values: 0.01 and 0.06). Our results highlight the importance of exploring the biologic impact of MMP‐9 and potential therapeutic roles of its inhibitors in ASD and may indicate that neuroplastic impairments in ASD may present during pregnancy. Autism Res 2012, 5: 428–433.

Biochemical screening of 504,049 newborns in Denmark, the Faroe Islands and Greenland — Experience and development of a routine program for expanded newborn screening

Expanded newborn screening for selected inborn errors of metabolism (IEM) in Denmark, the Faroe Islands and Greenland was introduced in 2002. We now present clinical, biochemical, and statistical results of expanded screening (excluding PKU) of 504,049 newborns during nine years as well as diagnoses and clinical findings in 82,930 unscreened newborns born in the same period. The frequencies of diagnoses made within the panel of disorders screened for are compared with the frequencies of the disorders in the decade preceding expanded newborn screening. The expanded screening was performed as a pilot study during the first seven years, and the experience obtained during these years was used in the development of the routine neonatal screening program introduced in 2009. Methods for screening included tandem mass spectrometry and an assay for determination of biotinidase activity. A total of 310 samples from 504,049 newborns gave positive screening results. Of the 310 results, 114 were true positive, including results from 12 newborns in which the disease in question was subsequently diagnosed in their mothers. Thus, the overall frequency of an IEM in the screening panel was 1:4942 (mothers excluded) or 1:4421 (mothers included). The false positive rate was 0.038% and positive predictive value 37%. Overall specificity was 99.99%. All patients with true positive results were followed in The Center for Inherited Metabolic Disorders in Copenhagen, and the mean follow-up period was 45 months (range 2109 months). There were no deaths among the 102 children, and 94% had no clinically significant sequelae at last follow-up. Our study confirms the higher frequency of selected IEM after implementation of expanded newborn screening and suggests an improved outcome for several disorders. We argue that newborn screening for these disorders should be standard of care, though unresolved issues remain, e.g. about newborns with a potential for remaining asymptomatic throughout life. Well organized logistics of the screening program from screening laboratory to centralized, clinical management is important.

Neonatal chemokine levels and risk of autism spectrum disorders: Findings from a Danish historic birth cohort follow-up study

A potential role of chemokines in the pathophysiology of Autism Spectrum Disorders (ASDs) has been previously suggested. In a recent study we examined levels of three inflammatory chemokines (MCP-1, MIP-1α and RANTES) in samples of amniotic fluid of children diagnosed later in life with ASD and controls frequency-matched to cases on gender and year of birth. In this follow-up study, levels of the same chemokines were analyzed postnatally in dried blood spot samples from the same subjects utilizing the Danish Newborn Screening Biobank. Crude estimates showed decreased levels of RANTES. In the adjusted estimates, no differences were found in levels of the three examined chemokines in ASD cases compared to controls. Our findings may cautiously suggest an altered cell-mediated immunity during the early neonatal period in ASD. Further research is needed to examine the relationship between maternal/fetal and neonatal chemokine levels and their role in ASD.

Small angle X-ray scattering study of calreticulin reveals conformational plasticity

Calreticulin plays a central role in vital cell processes such as protein folding, Ca(2+) homeostasis and immunogenicity. Even so, only limited three-dimensional structural information is presently available. We present a series of Small-Angle X-ray Scattering data on human placenta calreticulin. The data from the calreticulin monomer reveal the shape of calreticulin in solution: The previously structurally un-described C-terminal is seen as a globular domain, and the P-domain beta-hairpin extends from the N-domain in a spiral like conformation. In the calreticulin solution dimer, the N-, C-, and P-domains are easily identified, and the P-domain is in an extended conformation connecting to the second calreticulin molecule. The SAXS solution data enables the construction of a medium-resolution model of calreticulin. In the light of the unresolved chaperone mechanism of calreticulin and calnexin, we discuss the functional consequences of the conformational plasticity of the calreticulin P-domain.

Aquaporin-Based Biomimetic Polymeric Membranes: Approaches and Challenges

In recent years, aquaporin biomimetic membranes (ABMs) for water separation have gained considerable interest. Although the first ABMs are commercially available, there are still many challenges associated with further ABM development. Here, we discuss the interplay of the main components of ABMs: aquaporin proteins (AQPs), block copolymers for AQP reconstitution, and polymer-based supporting structures. First, we briefly cover challenges and review recent developments in understanding the interplay between AQP and block copolymers. Second, we review some experimental characterization methods for investigating AQP incorporation including freeze-fracture transmission electron microscopy, fluorescence correlation spectroscopy, stopped-flow light scattering, and small-angle X-ray scattering. Third, we focus on recent efforts in embedding reconstituted AQPs in membrane designs that are based on conventional thin film interfacial polymerization techniques. Finally, we describe some new developments in interfacial polymerization using polyhedral oligomeric silsesquioxane cages for increasing the physical and chemical durability of thin film composite membranes.

Cerebral autoregulation in the first day after preterm birth: no evidence of association with systemic inflammation

Introduction:Both systemic inflammation and impaired cerebral autoregulation (CA) have been associated with brain injury in preterm infants. We hypothesized that impaired CA represents a hemodynamic link between inflammation and brain injury.Results:Neither fetal vasculitis nor interleukin-6 (IL-6) affected CA significantly. A high level of IL-6 was associated with hypotension (P = 0.03) irrespective of dopamine therapy. The magnitude of impairment in CA increased with decreasing mean arterial blood pressure (MAP) (P = 0.02). No significant associations were found between these parameters and either intraventricular hemorrhage (IVH) (n = 10) or neonatal mortality (n = 8).Discussion:In conclusion, postnatal inflammation was weakly associated with arterial hypotension, and hypotension was weakly associated with impaired autoregulation. There was no direct association, however, between autoregulation and antenatal or postnatal signs of inflammation.Methods:In our study, 60 infants (mean (±SD) of gestational age (GA) 27 (±1.3) wk) underwent continuous recording of MAP and cerebral oxygenation index (OI) by means of near-infrared spectroscopy (NIRS) for 2.3 ± 0.5 h, starting 18 ± 9 h after birth. Coherence and transfer function gain between MAP and OI represented the presence and degree of impairment of CA, respectively. We considered fetal vasculitis (placenta histology) to be an antenatal marker of inflammation, and used the level of IL-6 in blood, measured at 18 ± 10 h after birth, as a postnatal marker of inflammation. Definition of hypotension was MAP (mm Hg) ≤ GA (wk).