Current methods for measuring wheat quality and dough rheology in the later stages of wheat breeding programs, including extensographs and farinographs, are costly and time-consuming. There is a significant interest in the Australian wheat industry for developing non-destructive, field-based, rapid dough-making quality assessment methods for Australian wheat varieties throughout earlier and later stages of the wheat breeding process. Fourier transform infrared (FTIR) spectroscopy is a valuable tool for analysis and quality control in the food industry as it is a simple and rapid technique requiring no sample pre-treatment before analysis. We aimed to investigate the application of FTIR spectroscopy coupled with partial least squares (PLSR) regression data analysis to rapidly assess wheat flour's dough-making quality. Results indicated that using FTIR data, PLSR could be applied to accurately predict multiple dough-making qualities, including protein content, extensibility, water absorption, dough development time (DDT), dough stability, and maximum resistance to tension (R max ). FTIR spectroscopy could not only be used to accurately predict the dough making quality of wheat lines from an in-sample test dataset, but this method also outperformed genetic predictive analysis, an established quality-prediction method in wheat breeding, in predicting dough making quality using out-of-sample data. • FTIR spectroscopy outperformed genomic analysis to predict dough-making quality. • PLSR accurately predicted dough making qualities using FTIR spectroscopy data. • Rapid predictions of protein content, extensibility, DDT, water absorption, R max . • More informed decisions on high-quality wheat lines early in the breeding process.
The intake of excess dietary fructose most often leads to non-alcoholic fatty liver disease (NAFLD). Fructose is metabolized mainly in the liver and its chronic consumption results in lipogenic gene expression in this organ. However, precisely how fructose is involved in NAFLD progression is still not fully understood, limiting therapy. Lipocalin-2 (LCN2) is a small secreted transport protein that binds to fatty acids, phospholipids, steroids, retinol, and pheromones. LCN2 regulates lipid and energy metabolism in obesity and is upregulated in response to insulin. We previously discovered that LCN2 has a hepatoprotective effect during hepatic insult, and that its upregulation is a marker of liver damage and inflammation. To investigate if LCN2 has impact on the metabolism of fructose and thereby arising liver damage, we fed wild type and Lcn2-/- mice for 4 or 8 weeks on diets that were enriched in fructose either by adding this sugar to the drinking water (30% w/v), or by feeding a chow containing 60% (w/w) fructose. Body weight and daily intake of food and water of these mice was then measured. Fat content in liver sections was visualized using Oil Red O stain, and expression levels of genes involved in fat and sugar metabolism were measured by qRT-PCR and Western blot analysis. We found that fructose-induced steatosis and liver damage was more prominent in female than in male mice, but that the most severe hepatic damage occurred in female mice lacking LCN2. Unexpectedly, consumption of elevated fructose did not induce de novo lipogenesis or fat accumulation. We conclude that LCN2 acts in a lipid-independent manner to protect the liver against fructose-induced damage.
In the October 2010 issue of the Journal of Clinical Investigation, Viau et al. [1] provided primary evidence for lipocalin-2 ([Lcn2; also known as neutrophil gelatinaseassociated lipocalin [(NGAL)], 24p3 protein, α1-microglobulin-related protein, or uterocalin]) as a central effector of progressive renal tissue damage upon acute kidney injury. Their studies are based on two experimental mouse strains which differed profoundly in their responses to 75% nephrectomy: whereas FVB/N mice develop severe renal lesions resembling features of human chronic kidney disease (CKD), B6D2F1mice are protected from early deterioration and instead exhibit compensatory alterations only. Post-surgical microarray analyses of the remnant renal tissues unravelled Lcn2 as the most markedly up-regulated gene in the FVB/N mice when compared to the B6D2F1 strain. Moreover, renal expression levels and urinary excretion of Lcn2 highly reflected the degree of tubular damage in the FVB/N mice, quite similar to humans with various forms of chronic kidney disease (CKD). Introduction of a homozygous disruption of the Lcn2 gene (i) largely abolished tubular cell proliferation, (ii) prevented the development of chronic renal lesions and (iii) preserved kidney function in FVB/N mice. The authors also identified Lcn2 as a downstreammediator following epidermal growth factor receptor (EGFR) activation. Genetically engineered mice with impaired EGF signalling did not up-regulate their Lcn2 levels and developed less severe renal damage in the remaining tissue after nephrectomy. Furthermore, EGFR activation mediated protein stabilization of the hypoxia-inducible factor (HIF)-1α, which accounted for increased Lcn2 expression.
Read moreHaemochromatosis is an iron overload disorder that can be inherited or acquired and when diagnosis is delayed, disease progression and death can occur. Iron overload was first described by the French internist Armand Trousseau in 1865 in an article on diabetes in which alterations in skin pigmentations were reported. Some years later, the German pathologist Friedrich Daniel von Recklinghausen coined the term ‘haemochromatosis’ for a metabolic disorder characterised by excess deposition of iron in the tissue. This disorder affects 1 in 200 subjects of Caucasians of Northern European descent. The systemic excess iron build-up condition quickly gained an intense clinical interest. Haemochromatosis can lead to severe pathological symptoms in multiple organs, including the liver, bones, spleen, heart, pancreas, joints, and reproductive organs. With the progress of the disease, hepatic damage predominates. Polymorphisms in several independent genes can lead to haemochromatosis. However, the most widely known haemochromatosis-associated and studied ones are genetic variants in the HFE gene, located on the short arm of human chromosome 6. Early detection and phlebotomy prior to the onset of fibrosis/cirrhosis can reduce morbidity and normalise life expectancy. Consequently, phlebotomy has been accepted for decades as a standard treatment for the reduction of iron load. Nowadays, other methods, such as erythrocytapheresis, therapeutic application of iron chelators and proton pump inhibitors, or hepcidin-targeted therapy, are discussed as alternative personalised treatments of hereditary haemochromatosis. This review focusses on the pathogenesis, diagnosis, and therapy of haemochromatosis.
Read moreParietal epithelial cells (PECs) are crucially involved in the pathogenesis of rapidly progressive glomerulonephritis (RPGN) as well as in focal and segmental glomerulosclerosis (FSGS). In this study, transgenic mouse lines were used to isolate pure, genetically tagged primary cultures of PECs or podocytes using FACsorting. By this approach, the morphology of primary glomerular epithelial cells in culture could be resolved: Primary podocytes formed either large cells with intracytoplasmatic extensions or smaller spindle shaped cells, depending on specific culture conditions. Primary PECs were small and exhibited a spindle-shaped or polygonal morphology. In the very early phases of primary culture, rapid changes in gene expression (e.g. of WT-1 and Pax-2) were observed. However, after prolonged culture primary PECs and podocytes still segregated clearly in a transcriptome analysis--demonstrating that the origin of primary cell cultures is important. Of the classical markers, synaptopodin and podoplanin expression were differentially regulated the most in primary PEC and podocyte cultures. However, no expression of any endogenous gene allowed to differentiate between the two cell types in culture. Finally, we show that the transcription factor WT1 is also expressed by PECs. In summary, genetic tagging of PECs and podocytes is a novel and necessary tool to derive pure primary cultures with proven origin. These cultures will be a powerful tool for the emerging field of parietal epithelial cell biology.
Read moreAims: We have previously shown that Endoglin, an accessory receptor for TGF-β1, is highly expressed in hepatic stellate cells (HSC) and Myofibroblast-like cells (MFB) [1]. Additionally, we could show that endoglin is upregulated during transdifferentiation of HSC to MFB and that a soluble form of endoglin (solEng) is generated by shedding in a cultured HSC cellline (CFSC–2G) [2]. Since solEng is increasingly detected in the serum of patients with cirrhosis [3] we asked if solouble endoglin has an impact on signaling in isolated liver cells especially in HSC.Methods and Results: The rat extracellular domain of endoglin (solEng) was transiently or virally expressed in COS–7-, CHO- or HEK293-cells. SolEng was secreted as a dimer in all cells. In a model cell line for TGF-β1-signaling we found that solEng, differentially modulated TGF-β1-mediated Smad3- and Smad1/Smad5-activation as shown by the reporter gene construct (CAGA)12-MLP-Luc [4] and Western blot. In line, a target gene of Smad3, i.e. Collagen I, was reduced, and a target gene of Smad1/Smad5, i.e. Id1, was increased, respectively. Overexpression of solEng in a mouse hepatic stellate cellline caused very similar changes. The Collagen I expression was decreased and α-smooth muscle actin (α-SMA) as well as Id2 were upregulated. Conclusions: Soluble endoglin not only affects the fibrotic response (Smad3-dependent) but may also be capable of modulating differentiation processes of hepatic stellate cells as shown by the activation marker α-SMA and the Id2 protein.
Read moreUNLABELLED: Liver fibrogenesis is associated with the transition of quiescent hepatocytes and hepatic stellate cells (HSCs) into the cell cycle. Exit from quiescence is controlled by E-type cyclins (cyclin E1 [CcnE1] and cyclin E2 [CcnE2]). Thus, the aim of the current study was to investigate the contribution of E-type cyclins for liver fibrosis in man and mice. Expression of CcnE1, but not of its homolog, CcnE2, was induced in fibrotic and cirrhotic livers from human patients with different etiologies and in murine wild-type (WT) livers after periodical administration of the profibrotic toxin, CCl(4). To further evaluate the potential function of E-type cyclins for liver fibrogenesis, we repetitively treated constitutive CcnE1(-/-) and CcnE2(-/-) knock-out mice with CCl(4) to induce liver fibrosis. Interestingly, CcnE1(-/-) mice were protected against CCl(4)-mediated liver fibrogenesis, as evidenced by reduced collagen type I α1 expression and the lack of septum formation. In contrast, CcnE2(-/-) mice showed accelerated fibrogenesis after CCl(4) treatment. We isolated primary HSCs from WT, CcnE1(-/-), and CcnE2(-/-) mice and analyzed their activation, proliferation, and survival in vitro. CcnE1 expression in WT HSCs was maximal when they started to proliferate, but decreased after the cells transdifferentiated into myofibroblasts. CcnE1(-/-) HSCs showed dramatically impaired survival, cell-cycle arrest, and strongly reduced expression of alpha smooth muscle actin, indicating deficient HSC activation. In contrast, CcnE2-deficient HSCs expressed an elevated level of CcnE1 and showed enhanced cell-cycle activity and proliferation, compared to WT cells. CONCLUSIONS: CcnE1 and CcnE2 have antagonistic roles in liver fibrosis. CcnE1 is indispensable for the activation, proliferation, and survival of HSCs and thus promotes the synthesis of extracellular matrix and liver fibrogenesis.
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