TY - JOUR A1 - Enssle, Jörg A1 - Weylandt, Karsten-Henrich T1 - Secure and optimized detection of PNPLA3 rs738409 genotype by an improved PCR-restriction fragment length polymorphism method JF - BioTechniques : the international journal of life science methods N2 - The PNPLA3 reference single-nucleotide polymorphism rs738409 has been identified as a predisposing factor for nonalcoholic fatty liver disease. A simple method based on PCR and restriction fragment length polymorphism (RFLP) analysis had been published to detect the nonpathogenic allele PNPLA3 rs738409 variant. The presence of the pathogenic variant was deduced by the indigestibility of the corresponding PCR product with BtsCI recognizing the nonpathogenic allele. However, one cannot exclude that an enzymatic reaction does not occur for other, more trivial, reasons. For safe and secure detection of the pathogenic PNPLA3 rs738409, we have further developed the PCR-restriction fragment length polymorphism method by adding a second restriction enzyme digest, clearly identifying the correct PNPLA3 alleles and in particular the pathogenic variant.
METHOD SUMMARY
The method presented here represents an improved genetic diagnosis of the PNPLA3 rs738409 alleles based on conventional and inexpensive molecular biological methods. We used methodology based on PCR and restriction fragment length polymorphisms and clearly identified both described alleles by the use of two restriction enzymes. Digestion of individuals' specific PNPLA3 PCR fragments with both enzymes in independent reactions clearly showed the PNPLA3 rs738409 genotype. KW - n-3 polyunsaturated fatty acid therapies KW - nonalcoholic fatty liver KW - disease KW - PCR– RFLP KW - PNPLA3 KW - rs738409 Y1 - 2021 U6 - https://doi.org/10.2144/btn-2020-0163 SN - 0736-6205 SN - 1940-9818 VL - 70 IS - 6 SP - 345 EP - 349 PB - Future Science Ltd. CY - London ER - TY - JOUR A1 - He, Yangyang A1 - Würtz-Kozak, Karin A1 - Kühl, Linn Kristina A1 - Wippert, Pia-Maria T1 - Extracellular vesicles BT - Potential mediators of psychosocial stress contribution to osteoporosis? JF - International journal of molecular sciences N2 - Osteoporosis is characterized by low bone mass and damage to the bone tissue’s microarchitecture, leading to increased fracture risk. Several studies have provided evidence for associations between psychosocial stress and osteoporosis through various pathways, including the hypothalamic-pituitary-adrenocortical axis, the sympathetic nervous system, and other endocrine factors. As psychosocial stress provokes oxidative cellular stress with consequences for mitochondrial function and cell signaling (e.g., gene expression, inflammation), it is of interest whether extracellular vesicles (EVs) may be a relevant biomarker in this context or act by transporting substances. EVs are intercellular communicators, transfer substances encapsulated in them, modify the phenotype and function of target cells, mediate cell-cell communication, and, therefore, have critical applications in disease progression and clinical diagnosis and therapy. This review summarizes the characteristics of EVs, their role in stress and osteoporosis, and their benefit as biological markers. We demonstrate that EVs are potential mediators of psychosocial stress and osteoporosis and may be beneficial in innovative research settings. KW - allostatic load KW - bone remodeling KW - microRNA KW - osteoblast KW - osteoclast Y1 - 2021 U6 - https://doi.org/10.3390/ijms22115846 SN - 1422-0067 VL - 22 IS - 11 PB - Molecular Diversity Preservation International CY - Basel ER - TY - JOUR A1 - Henkel, Janin A1 - Klauder, Julia A1 - Statz, Meike A1 - Wohlenberg, Anne-Sophie A1 - Kuipers, Sonja A1 - Vahrenbrink, Madita A1 - Püschel, Gerhard Paul T1 - Enhanced Palmitate-Induced Interleukin-8 Formation in Human Macrophages by Insulin or Prostaglandin E-2 JF - Biomedicines N2 - Macrophages in pathologically expanded dysfunctional white adipose tissue are exposed to a mix of potential modulators of inflammatory response, including fatty acids released from insulin-resistant adipocytes, increased levels of insulin produced to compensate insulin resistance, and prostaglandin E-2 (PGE(2)) released from activated macrophages. The current study addressed the question of how palmitate might interact with insulin or PGE(2) to induce the formation of the chemotactic pro-inflammatory cytokine interleukin-8 (IL-8). Human THP-1 cells were differentiated into macrophages. In these macrophages, palmitate induced IL-8 formation. Insulin enhanced the induction of IL-8 formation by palmitate as well as the palmitate-dependent stimulation of PGE(2) synthesis. PGE(2) in turn elicited IL-8 formation on its own and enhanced the induction of IL-8 release by palmitate, most likely by activating the EP4 receptor. Since IL-8 causes insulin resistance and fosters inflammation, the increase in palmitate-induced IL-8 formation that is caused by hyperinsulinemia and locally produced PGE(2) in chronically inflamed adipose tissue might favor disease progression in a vicious feed-forward cycle. KW - macrophages KW - insulin KW - prostaglandin E-2 KW - interleukin-8 KW - inflammation Y1 - 2021 U6 - https://doi.org/10.3390/biomedicines9050449 SN - 2227-9059 VL - 9 IS - 5 PB - MDPI CY - Basel ER - TY - JOUR A1 - Ranisch, Robert T1 - Consultation with Doctor Twitter BT - consent Fatigue, and the role of developers in digital medical ethics JF - The American journal of bioethics : ajob Y1 - 2021 U6 - https://doi.org/10.1080/15265161.2021.1926595 SN - 1526-5161 SN - 1536-0075 VL - 21 IS - 7 SP - 24 EP - 25 PB - Routledge, Taylor & Francis Group CY - Philadelphia, Pa. ER - TY - JOUR A1 - Witte, Leonie A1 - Linnemannstoens, Karen A1 - Honemann-Capito, Mona A1 - Groß, Julia Christina T1 - Visualization and quantitation of Wg trafficking in the Drosophila wing imaginal epithelium JF - Bio-protocol N2 - Secretory Wnt trafficking can be studied in the polarized epithelial monolayer of Drosophila wing imaginal discs (WID). In this tissue, Wg (Drosophila Wnt-I) is presented on the apical surface of its source cells before being internalized into the endosomal pathway. Long-range Wg secretion and spread depend on secondary secretion from endosomal compartments, but the exact post-endocytic fate of Wg is poorly understood. Here, we summarize and present three protocols for the immunofluorescencebased visualization and quantitation of different pools of intracellular and extracellular Wg in WID: (1) steady-state extracellular Wg; (2) dynamic Wg trafficking inside endosomal compartments; and (3) dynamic Wg release to the cell surface. Using a genetic driver system for gene manipulation specifically at the posterior part of the WID (EnGal4) provides a robust internal control that allows for direct comparison of signal intensities of control and manipulated compartments of the same WID. Therefore, it also circumvents the high degree of staining variability usually associated with whole-tissue samples. In combination with the genetic manipulation of Wg pathway components that is easily feasible in Drosophila, these methods provide a tool-set for the dissection of secretory Wg trafficking and can help us to understand how Wnt proteins travel along endosomal compartments for short-and long-range signal secretion. KW - Wingless/Wnt secretion KW - Morphogen signaling KW - Drosophila wing imaginal disc KW - Recycling assay KW - Extracelluar wingless KW - Imaginal disc dissection Y1 - 2021 U6 - https://doi.org/10.21769/BioProtoc.4040 SN - 2331-8325 VL - 11 IS - 11 PB - bio-protocol.org CY - Sunnyvale, CA ER -