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Brandenburger, T.

Publications and source records attributed to Brandenburger, T..

2 recordsLinked to original sources
bioRxiv2023

An optimized workflow for analyzing extracellular vesicles as biomarkers in liver diseases.

Background & AimsExtracellular vesicles (EVs) play an important role in intercellular communication, serving as vehicles for the exchange of biological materials and being involved in the regulation of physiological processes. EVs and their associated cargoes are considered a promising source of disease-associated biomarkers. The purpose of this study was to establish an easy-to-use, reproducible, and scalable workflow to efficiently analyze EVs in the context of liver disease. MethodsAn optimized workflow was established for the pre-analytical processing and isolation of EVs from plasma and serum. Nanoparticle Tracking Analysis (NTA) was used to characterize circulating EVs in the serum of patients with nonalcoholic fatty liver disease (NAFLD), autoimmune liver disease (AIH), and animal models with impaired liver function. EVs were separated from soluble proteins by an optimized, polyethylene glycol (PEG)-based enrichment protocol. Enriched EVs were either labeled and functionally characterized by monitoring cellular uptake or lysed for biomarker identification. ResultsCirculating EVs in the serum of patients with NAFLD or AIH and in different animal models have been characterized by NTA. Here we show that both the quantity and size of EVs in the serum of patients/animal models are significantly different from those of healthy individuals. We show that isolated EVs are functional, and their uptake by acceptor cells can be quantified after fluorescence labelling. Enriched EVs were directly used to analyze RNA biomarkers. Several microRNAs, including miR-15b, -16, -21, -122 and -223, were found to be significantly up-regulated in EVs isolated from the sera of patients with NAFLD and AIH. We show that EVs transport cytokines, and that IL-2, IL-6 and IL-8 were significantly up-regulated in EVs enriched from patients with cholangiocarcinoma (CCA) compared to healthy controls. ConclusionsThe workflow presented here represents an accessible and easy-to-use approach that enables the analysis and enrichment of EVs from complex biological fluids and their preparation for functional characterization or downstream analysis. In this study, the levels of several miRNAs were found to be significantly increased in EVs isolated from AIH and NAFLD patients compared with healthy controls. HighlightsO_LIEVs circulating in crude serum reflect the diseased stage of the donors. C_LIO_LIEnrichment of EVs with the approach presented here efficiently separates soluble proteins from EVs, providing optimal material for further characterization. C_LIO_LIExosomal markers are present in the EVs-enriched fraction. C_LIO_LIEnriched EVs are intact and are functionally taken up by acceptor cells. C_LIO_LIEnriched EVs are suitable, and have been used for, biomarkers identification both at RNA and protein level. C_LI

molecular biology↗
bioRxiv2023

Cytokines in the Urine of AKI patients regulate TP53 and SIRT1 and can be used as biomarkers for the early detection of AKI

Acute kidney injury (AKI) is a major kidney disease with a poor clinical outcome. It is a common complication with an incidence of 10-15% of patients admitted to hospital. This rate even increases for patients who are admitted to the intensive care unit with an incidence of >50%. AKI is characterized by a rapid increase in serum creatinine, decrease in urine output, or both. Associated Symptoms include feeling sick or being sick, diarrhoea, dehydration, decreased urine output-although occasionally urine output remains normal, fluid retention-causing swelling in the legs or ankles, shortness of breath, fatigue and nausea. However, sometimes acute kidney injury causes no signs or symptoms and is detected by lab tests. Therefore, an urgent demand for non-invasive biomarkers for early detection of AKI are highly desirable. This might enable the prevention of the progression from AKI to CKD. In this study, we analysed the secretome of urine of an AKI patient cohort employing a kidney-biomarker cytokine assay. Based on these results we suggest, ADIPOQ, EGF and SERPIN3A as potential biomarkers, which might be able to detect AKI as soon as 24 h post-surgery. For the later stages, common biomarkers for the detection of AKI in both male and female patients we suggest, VEGF, SERPIN3A, TNFSF12, ANPEP, CXCL1, REN, CLU and PLAU. These markers in combination might present a robust strategy to identify the development of AKI as early as 24h or 72h post-surgery. Furthermore, we evaluated the effect of patient and healthy urine on human podocyte cells. We conclude that cytokines in the urine of AKI patients trigger processes which are needed to repair the damaged nephron and activate TP53 and SIRT1 to maintain the balance between proliferation, angiogenesis, and cell cycle arrest. In conclusion, the Renin-Angiotensin pathway seems to have major implications.

molecular biology↗