Laboratory Genetic Testing in Clinical Practice 2016.

WOS: 000392526600001PubMed ID: 2813360

Evolution of Genetic Techniques: Past, Present, and Beyond.

Genetics is the study of heredity, which means the study of genes and factors related to all aspects of genes. The scientific history of genetics began with the works of Gregor Mendel in the mid-19th century. Prior to Mendel, genetics was primarily theoretical whilst, after Mendel, the science of genetics was broadened to include experimental genetics. Developments in all fields of genetics and genetic technology in the first half of the 20th century provided a basis for the later developments. In the second half of the 20th century, the molecular background of genetics has become more understandable. Rapid technological advancements, followed by the completion of Human Genome Project, have contributed a great deal to the knowledge of genetic factors and their impact on human life and diseases. Currently, more than 1800 disease genes have been identified, more than 2000 genetic tests have become available, and in conjunction with this at least 350 biotechnology-based products have been released onto the market. Novel technologies, particularly next generation sequencing, have dramatically accelerated the pace of biological research, while at the same time increasing expectations. In this paper, a brief summary of genetic history with short explanations of most popular genetic techniques is given

Laboratory genetic testing in clinical practice 2014.

WOS: 000352434200001PubMed ID: 2587902

Human isotype‐dependent inhibitory antibody responses against Mycobacterium tuberculosis

Accumulating evidence from experimental animal models suggests that antibodies play a protective role against tuberculosis (TB). However, little is known about the antibodies generated upon Mycobacterium tuberculosis (MTB) exposure in humans. Here, we performed a molecular and functional characterization of the human B‐cell response to MTB by generating recombinant monoclonal antibodies from single isolated B cells of untreated adult patients with acute pulmonary TB and from MTB‐exposed healthcare workers. The data suggest that the acute plasmablast response to MTB originates from reactivated memory B cells and indicates a mucosal origin. Through functional analyses, we identified MTB inhibitory antibodies against mycobacterial antigens including virulence factors that play important roles in host cell infection. The inhibitory activity of anti‐MTB antibodies was directly linked to their isotype. Monoclonal as well as purified serum IgA antibodies showed MTB blocking activity independently of Fc alpha receptor expression, whereas IgG antibodies promoted the host cell infection. Together, the data provide molecular insights into the human antibody response to MTB and may thereby facilitate the design of protective vaccination strategies

Evaluation of the role of downregulation of SNF5/INI1 core subunit of SWI/SNF complex in clear cell renal cell carcinoma development

Clear cell renal cell carcinoma (ccRCC) is characterized by stabilization of hypoxia-inducible factor (HIF1), and mutations in von Hippel-Lindau (VHL) gene. Additionally, in about 40% of ccRCC cases the mutation in PBRM1 (POLYBROMO1) gene coding for a non-core subunit of SWI/SNF chromatin remodeling complex was found suggesting potential impairment of this complex function in ccRCC. In this study we assessed the extent to which the core SWI/SNF complex subunit - INI1 (hSNF5/SMARCB1) is affected in ccRCC and whether it has any consequences on the development of this type of cancer. The evaluation of INI1 protein level in samples from 50 patients with diagnosed ccRCC, including three displaying rhabdoid features, showed the INI1 positive staining in rhabdoid cells while the conventional ccRCC cells exhibited reduced INI1 level. This indicated the rhabdoid component of ccRCC as distinct from other known rhabdoid tumors. The reduced INI1 protein level observed in all conventional ccRCC cases used in this study correlated with decreased SMARCB1 gene expression at the transcript level. Consistently, the overexpression of INI1 protein in A498 ccRCC cell line resulted in the elevation of endogenous SMARCB1 transcript level indicating that the INI1-dependent regulatory feedback loop controlling expression of this gene is affected in ccRCC Moreover, the set of INI1 target genes including i.e. CXCL12/CXCR7/CXCR4 chemokine axis was identified to be affected in ccRCC. In summary, we demonstrated that the inactivation of INI1 may be of high importance for ccRCC development and aggressiveness

Two novel C-terminal frameshift mutations in the β-globin gene lead to rapid mRNA decay

BACKGROUND: The thalassemia syndromes are classified according to the globin chain or chains whose production is affected. β-thalassemias are caused by point mutations or, more rarely, deletions or insertions of a few nucleotides in the β-globin gene or its immediate flanking sequences. These mutations interfere with the gene function either at the transcriptional, translational or posttranslational level. METHODS: Two cases of Polish patients with hereditary hemolytic anemia suspected of thalassemia were studied. DNA sequencing and mRNA quantification were performed. Stable human cell lines which express wild-type HBB and mutated versions were used to verify that detected mutation are responsible for mRNA degradation. RESULTS: We identified two different frameshift mutations positioned in the third exon of HBB. Both patients harboring these mutations present the clinical phenotype of thalassemia intermedia and showed dominant pattern of inheritance. In both cases the mutations do not generate premature stop codon. Instead, slightly longer protein with unnatural C-terminus could be produced. Interestingly, although detected mutations are not expected to induce NMD, the mutant version of mRNA is not detectable. Restoring of the open reading frame brought back the RNA to that of the wild-type level. CONCLUSION: Our results show that a lack of natural stop codon due to the frameshift in exon 3 of β-globin gene causes rapid degradation of its mRNA and indicate existence of novel surveillance pathway

Immunopathogenesis of idiopathic pulmonary fibrosis

Praca jest przeglądem nowoczesnej wiedzy dotyczącej patogenezy samoistnego włóknienia płuc (IPF). Zarówno przyczyna jak i patogeneza IPF są niewyjaśnione, nie istnieje spójna teoria tłumacząca przyczynę choroby. Kluczowymi elementami patogenezy IPF są: uszkodzenie komórek nabłonka pęcherzyków płucnych i dysregulacja lub zmiana fenotypu fibroblastów Obecnie uznana hipoteza głosi, że samoistne włóknienie płuc jest niezależne od zjawiska zapalenia i jest wynikiem nieprawidłowej interakcji pomiędzy komórkami nabłonka a mezenchymą oraz zaburzonego gojenia przewlekłych uszkodzeń nabłonka pęcherzyków. Nieprawidłowa jest również funkcja oddechowych komórek macierzystych i innych komórek progenitorowych rezydujących w dystalnych częściach płuc. Sugeruje się, że u chorych na IPF istnieje defekt genetyczny powodujący szybsze skracanie długości telomerów, co jest przyczyną ograniczonych zdolności tkanki płucnej do odnowy. W myśl współczesnej wiedzy IPF jest chorobą o bardzo złożonej, wieloczynnikowej etiopatogenezie i jest zainicjowane przez cykl urazów i patologicznej odnowy powtarzających się wielokrotnie.The paper presents the state of the art in the pathogenesis of idiopathic pulmonary fibrosis (IPF). Both, etiology and pathogenesis of IPF are unclear. The key elements in the pathogenesis of the disease are epithelial destruction and dysregulation of the phenotype of lung fibroblasts. Currently accepted hypothesis claims that IPF is not related to underlying inflammatory state but it is rather a result of pathological interaction between pulmonary epithelium and mesenchyme followed by disturbed healing of damaged alveolar epithelial cells. The function of lung progenitor cells residing in distant lung structures is also impaired. Some scientists claim that genetic defect causes fast shortening of telomeres reducing lung properties of regeneration. According to the current knowledge, IFP is a multifactorial disease resulting from a repeated cycle of injuries followed by pathological regeneration

The immunological and molecular diagnostics of respiratory tract infections

Diagnostyka laboratoryjna zakażeń górnych i dolnych dróg oddechowych opiera się na metodach konwencjonalnych oraz nowoczesnych metodach biologii molekularnej. Serologia odgrywa ważną rolę w rozpoznawaniu niektórych zakażeń o etiologii wirusowej i w diagnostyce zakażeń drobnoustrojami atypowymi. Metody serologiczne nie są jednak przydatne w diagnostyce zakażeń u małych dzieci oraz u osób w immunosupresji. W przypadku patogenów niewykrywalnych metodami konwencjonalnymi oraz zakażeń o ciężkim przebiegu, zagrażających życiu, metody molekularne mają duże znaczenie. Pneumonol. Alergol. Pol. 2011; 79, 6: 446–453Diagnostic strategies in upper and lower respiratory tract infections include conventional procedures and the new molecular tools. Serology plays a role in certain viral and atypical infections of lower respiratory track but is not reliable in small children and in immunocompromised subjects. Antigen detection can be also carried out. If a potential pathogen is non detectable by simply assay or in severe or complicated lower respiratory tract infections, the use of molecular tools is highly indicated. Pneumonol. Alergol. Pol. 2011; 79, 6: 446–45