Person:
Freire, Rachel

Profile Picture

Email Address

AA Acceptance Date

Birth Date

Research Projects

Organizational Units

Job Title

Last Name

Freire

First Name

Rachel

Name

Freire, Rachel
Author's Publications: search.filters.isAuthorOfPublication.0290c375-19ee-4827-8a3d-8d262b48db9a

Search Results

Now showing 1 - 2 of 2
  • Thumbnail Image
    Publication
    Widely Used Commercial ELISA Does Not Detect Precursor of Haptoglobin2, but Recognizes Properdin as a Potential Second Member of the Zonulin Family
    (Frontiers Media S.A., 2018) Scheffler, Lucas; Crane, Alyce; Heyne, Henrike; Tönjes, Anke; Schleinitz, Dorit; Ihling, Christian H.; Stumvoll, Michael; Freire, Rachel; Fiorentino, Maria; Fasano, Alessio; Kovacs, Peter; Heiker, John T.
    Background: There is increasing evidence for the role of impaired intestinal permeability in obesity and associated metabolic diseases. Zonulin is an established serum marker for intestinal permeability and identical to pre-haptoglobin2. Here, we aimed to investigate the relationship between circulating zonulin and metabolic traits related to obesity. Methods: Serum zonulin was measured by using a widely used commercial ELISA kit in 376 subjects from the metabolically well-characterized cohort of Sorbs from Germany. In addition, haptoglobin genotype was determined in DNA samples from all study subjects. Results: As zonulin concentrations did not correlate to the haptoglobin genotypes, we investigated the specificity of the zonulin ELISA assay using antibody capture experiments, mass spectrometry, and Western blot analysis. Using serum samples that gave the highest or lowest ELISA signals, we detected several proteins that are likely to be captured by the antibody in the present kit. However, none of these proteins corresponds to pre-haptoglobin2. We used increasing concentrations of recombinant pre-haptoglobin2 and complement C3 as one of the representative captured proteins and the ELISA kit did not detect either. Western blot analysis using both the polyclonal antibodies used in this kit and monoclonal antibodies rose against zonulin showed a similar protein recognition pattern but with different intensity of detection. The protein(s) measured using the ELISA kit was (were) significantly increased in patients with diabetes and obesity and correlated strongly with markers of the lipid and glucose metabolism. Combining mass spectrometry and Western blot analysis using the polyclonal antibodies used in the ELISA kit, we identified properdin as another member of the zonulin family. Conclusion: Our study suggests that the zonulin ELISA does not recognize pre-haptoglobin2, rather structural (and possibly functional) analog proteins belonging to the mannose-associated serine protease family, with properdin being the most likely possible candidate.
  • Thumbnail Image
    Publication
    Human Fetal-Derived Enterospheres Provide Insights on Intestinal Development and a Novel Model to Study Necrotizing Enterocolitis (NEC)
    (Elsevier, 2018) Senger, Stefania; Ingano, Laura; Freire, Rachel; Anselmo, Antony; Zhu, Weishu; Sadreyev, Ruslan; Walker, William Allan; Fasano, Alessio
    Background & Aims Untreated necrotizing enterocolitis (NEC) can lead to massive inflammation resulting in intestinal necrosis with a high mortality rate in preterm infants. Limited access to human samples and relevant experimental models have hampered progress in NEC pathogenesis. Earlier evidence has suggested that bacterial colonization of an immature and developing intestine can lead to an abnormally high inflammatory response to bacterial bioproducts. The aim of our study was to use human fetal organoids to gain insights into NEC pathogenesis. Methods: RNA sequencing analysis was performed to compare patterns of gene expression in human fetal-derived enterospheres (FEnS) and adult-derived enterospheres (AEnS). Differentially expressed genes were analyzed using computational techniques for dimensional reduction, clustering, and gene set enrichment. Unsupervised cluster analysis, Gene Ontology, and gene pathway analysis were used to predict differences between gene expression of samples. Cell monolayers derived from FEnS and AEnS were evaluated for epithelium function and responsiveness to lipopolysaccharide and commensal bacteria. Results: Based on gene expression patterns, FEnS clustered according to their developmental age in 2 distinct groups: early and late FEnS, with the latter more closely resembling AEnS. Genes involved in maturation, gut barrier function, and innate immunity were responsible for these differences. FEnS-derived monolayers exposed to either lipopolysaccharide or commensal Escherichia coli showed that late FEnS activated gene expression of key inflammatory cytokines, whereas early FEnS monolayers did not, owing to decreased expression of nuclear factor-κB–associated machinery. Conclusions: Our results provide insights into processes underlying human intestinal development and support the use of FEnS as a relevant human preclinical model for NEC. Accession number of repository for expression data: GSE101531.