Identification of a rare p.G320R alpha-1-antitrypsin variant in emphysema and lung cancer patients

The alpha-1-antitrypsin (A1AT) gene is highly polymorphic, with more than 100 genetic variants identified of which some can affect A1AT protein concentration and/or function and lead to pulmonary and/or liver disease. This study reports on the characterization of a p.G320R variant found in two patients, one with emphysema and the other with lung cancer. This variant results from a single base-pair substitution in exon 4 of the A1AT gene, and has been characterized as P by isoelectric focusing. Functional evaluation of the A1AT p.G320R variant was through comparing specific trypsin inhibitory activity in two patients with pulmonary disorders, carriers of the p.G320R variant, and 19 healthy individuals, carriers of normal A1AT M variants. Results showed that specific trypsin inhibitory activity was lower in both emphysema (2.45 mU/g) and lung cancer (2.07 mU/g) patients than in carriers of the normal variants (range 2.51-3.71 mU/g). This rare A1AT variant is associated with reduced functional activity of A1AT protein. Considering that it was found in patients with severe pulmonary disorders, this variant could be of clinical significance

Fluorescence Quantum Yield of Thioflavin T in Rigid Isotropic Solution and Incorporated into the Amyloid Fibrils

In this work, the fluorescence of thioflavin T (ThT) was studied in a wide range of viscosity and temperature. It was shown that ThT fluorescence quantum yield varies from 0.0001 in water at room temperature to 0.28 in rigid isotropic solution (T/η→0). The deviation of the fluorescence quantum yield from unity in rigid isotropic solution suggests that fluorescence quantum yield depends not only on the ultra-fast oscillation of ThT fragments relative to each other in an excited state as was suggested earlier, but also depends on the molecular configuration in the ground state. This means that the fluorescence quantum yield of the dye incorporated into amyloid fibrils must depend on its conformation, which, in turn, depends on the ThT environment. Therefore, the fluorescence quantum yield of ThT incorporated into amyloid fibrils can differ from that in the rigid isotropic solution. In particular, the fluorescence quantum yield of ThT incorporated into insulin fibrils was determined to be 0.43. Consequently, the ThT fluorescence quantum yield could be used to characterize the peculiarities of the fibrillar structure, which opens some new possibilities in the ThT use for structural characterization of the amyloid fibrils

Assessment of the Red Cell Proteome of Young Patients with Unexplained Hemolytic Anemia by Two-Dimensional Differential In-Gel Electrophoresis (DIGE)

Erythrocyte cytosolic protein expression profiles of children with unexplained hemolytic anemia were compared with profiles of close relatives and controls by two-dimensional differential in-gel electrophoresis (2D-DIGE). The severity of anemia in the patients varied from compensated (i.e., no medical intervention required) to chronic transfusion dependence. Common characteristics of all patients included chronic elevation of reticulocyte count and a negative workup for anemia focusing on hemoglobinopathies, morphologic abnormalities that would suggest a membrane defect, immune-mediated red cell destruction, and evaluation of the most common red cell enzyme defects, glucose-6-phosphate dehydrogenase and pyruvate kinase deficiency. Based upon this initial workup and presentation during infancy or early childhood, four patients classified as hereditary nonspherocytic hemolytic anemia (HNSHA) of unknown etiology were selected for proteomic analysis. DIGE analysis of red cell cytosolic proteins clearly discriminated each anemic patient from both familial and unrelated controls, revealing both patient-specific and shared patterns of differential protein expression. Changes in expression pattern shared among the four patients were identified in several protein classes including chaperons, cytoskeletal and proteasome proteins. Elevated expression in patient samples of some proteins correlated with high reticulocyte count, likely identifying a subset of proteins that are normally lost during erythroid maturation, including proteins involved in mitochondrial metabolism and protein synthesis. Proteins identified with patient-specific decreased expression included components of the glutathione synthetic pathway, antioxidant pathways, and proteins involved in signal transduction and nucleotide metabolism. Among the more than 200 proteins identified in this study are 21 proteins not previously described as part of the erythrocyte proteome. These results demonstrate the feasibility of applying a global proteomic approach to aid characterization of red cells from patients with hereditary anemia of unknown cause, including the identification of differentially expressed proteins as potential candidates with a role in disease pathogenesis

Revisiting the mechanism of coagulation factor XIII activation and regulation from a structure/functional perspective

The activation and regulation of coagulation Factor XIII (FXIII) protein has been the subject of active research for the past three decades. Although discrete evidence exists on various aspects of FXIII activation and regulation a combinatorial structure/functional view in this regard is lacking. In this study, we present results of a structure/function study of the functional chain of events for FXIII. Our study shows how subtle chronological submolecular changes within calcium binding sites can bring about the detailed transformation of the zymogenic FXIII to its activated form especially in the context of FXIIIA and FXIIIB subunit interactions. We demonstrate what aspects of FXIII are important for the stabilization (first calcium binding site) of its zymogenic form and the possible modes of deactivation (thrombin mediated secondary cleavage) of the activated form. Our study for the first time provides a structural outlook of the FXIIIA 2 B 2 heterotetramer assembly, its association and dissociation. The FXIIIB subunits regulatory role in the overall process has also been elaborated upon. In summary, this study provides detailed structural insight into the mechanisms of FXIII activation and regulation that can be used as a template for the development of future highly specific therapeutic inhibitors targeting FXIII in pathological conditions like thrombosis

The sperm factor: paternal impact beyond genes

The fact that sperm carry more than the paternal DNA has only been discovered just over a decade ago. With this discovery, the idea that the paternal condition may have direct implications for the fitness of the offspring had to be revisited. While this idea is still highly debated, empirical evidence for paternal effects is accumulating. Male condition not only affects male fertility but also offspring early development and performance later in life. Several factors have been identified as possible carriers of non-genetic information, but we still know little about their origin and function and even less about their causation. I consider four possible non-mutually exclusive adaptive and non-adaptive explanations for the existence of paternal effects in an evolutionary context. In addition, I provide a brief overview of the main non-genetic components found in sperm including DNA methylation, chromatin modifications, RNAs and proteins. I discuss their putative functions and present currently available examples for their role in transferring non-genetic information from the father to the offspring. Finally, I identify some of the most important open questions and present possible future research avenues

Expression of hereditary hemochromatosis C282Y HFE protein in HEK293 cells activates specific endoplasmic reticulum stress responses

<p>Abstract</p> <p>Background</p> <p>Hereditary Hemochromatosis (HH) is a genetic disease associated with iron overload, in which individuals homozygous for the mutant C282Y <it>HFE </it>associated allele are at risk for the development of a range of disorders particularly liver disease. Conformational diseases are a class of disorders associated with the expression of misfolded protein. HFE C282Y is a mutant protein that does not fold correctly and consequently is retained in the Endoplasmic Reticulum (ER). In this context, we sought to identify ER stress signals associated with mutant C282Y HFE protein expression, which may have a role in the molecular pathogenesis of HH.</p> <p>Results</p> <p>Vector constructs of Wild type HFE and Mutant C282Y HFE were made and transfected into HEK293 cell lines. We have shown that expression of C282Y HFE protein triggers both an unfolded protein response (UPR), as revealed by the increased GRP78, ATF6 and CHOP expression, and an ER overload response (EOR), as indicated by NF-κB activation. Furthermore, C282Y HFE protein induced apoptotic responses associated with activation of ER stress. Inhibition studies demonstrated that tauroursodeoxycholic acid, an endogenous bile acid, downregulates these events. Finally, we found that the co-existence of both C282Y HFE and Z alpha 1-antitrypsin protein (the protein associated with the liver disease of Z alpha 1-antitrypsin deficiency) expression on ER stress responses acted as potential disease modifiers with respect to each other.</p> <p>Conclusion</p> <p>Our novel observations suggest that both the ER overload response (EOR) and the unfolded protein response (UPR) are activated by mutant C282Y HFE protein.</p

Meta-Analysis of Genome-Wide Association Studies for Abdominal Aortic Aneurysm Identifies Four New Disease-Specific Risk Loci

Rationale: Abdominal aortic aneurysm (AAA) is a complex disease with both genetic and environmental risk factors. Together, 6 previously identified risk loci only explain a small proportion of the heritability of AAA. Objective: To identify additional AAA risk loci using data from all available genome-wide association studies (GWAS). Methods and Results: Through a meta-analysis of 6 GWAS datasets and a validation study totalling 10,204 cases and 107,766 controls we identified 4 new AAA risk loci: 1q32.3 (SMYD2), 13q12.11 (LINC00540), 20q13.12 (near PCIF1/MMP9/ZNF335), and 21q22.2 (ERG). In various database searches we observed no new associations between the lead AAA SNPs and coronary artery disease, blood pressure, lipids or diabetes. Network analyses identified ERG, IL6R and LDLR as modifiers of MMP9, with a direct interaction between ERG and MMP9. Conclusions: The 4 new risk loci for AAA appear to be specific for AAA compared with other cardiovascular diseases and related traits suggesting that traditional cardiovascular risk factor management may only have limited value in preventing the progression of aneurysmal disease

Standards in semen examination: publishing reproducible and reliable data based on high-quality methodology

Biomedical science is rapidly developing in terms of more transparency, openness and reproducibility of scientific publications. This is even more important for all studies that are based on results from basic semen examination. Recently two concordant documents have been published: the 6th edition of the WHO Laboratory Manual for the Examination and Processing of Human Semen, and the International Standard ISO 23162:2021. With these tools, we propose that authors should be instructed to follow these laboratory methods in order to publish studies in peer-reviewed journals, preferable by using a checklist as suggested in an Appendix to this article