AERI 2019 – Archival Education and Research Institute. Liverpool, 8–12 VII 2019 r. Impresje

AERI 2019 – Archival Education and Research Institute. Liverpool, 8–12 VII 2019 r. Impresj

The alterations in SATB1 and nuclear F-actin expression affect apoptotic response of the MCF-7 cells to geldanamycin

Introduction. The function and localization of actin in the nucleus have not yet been fully described. However, actin seems to be a key protein in nuclear processes interacting with chromatin and matrix proteins. The aim of the study was to evaluate the effect of controlled expression of nuclear pool of F-actin and special AT-rich sequence-binding protein 1 (SATB1) on the in vitro induction of active cell death by geldanamycin (GA). Material and methods. The expression of SATB1 was regulated by the transfection of non-aggressive breast cancer MCF-7 cells with siRNA against SATB1 or expression plasmid with cloned cDNA of SATB1. The altered expression of cofilin-1 in these cells was used to regulate the nuclear expression and localization of F-actin. The effect of GA was analyzed in the context of cell death induction and cell cycle alterations. Results. Our studies revealed that the targeted regulation of SATB1 and cofilin-1 expression changed the apoptotic response of the MCF-7 cells to GA. The overexpression of these proteins potentiated GA-induced arrest of the cells in the G1 phase of cell cycle and increased the population of the hypodiploid cells. Conclusion. The alterations in the nuclear expression of SATB1 and F-actin in MCF-7 cells may affect their active cell death in response to GA

The interactions between SATB1 and F-actin are important for mechanisms of active cell death

Introduction. The direct involvement of nuclear actin filaments in gene transcription and remodeling of chromatin is still debatable. However, nuclear localization of F-actin and its interactions with other nuclear matrix proteins have been reported. The aim of the study was to estimate the interactions between nuclear F-actin and one of the matrix proteins, special AT-rich sequence-binding protein 1 (SATB1), during active cell death induced in vitro by geldanamycin (GA). Material and methods. The expression of SATB1 was modified by the transfection of non-aggressive breast cancer MCF-7 cells with siRNA against SATB1 or expression plasmid with cloned cDNA of SATB1. The amount and localization of F-actin were altered by changes of cofilin-1 (CFL1) expression in MCF-7 cells. The association between SATB1 and F-actin during GA-induced cell death was analyzed using confocal and transmission electron microscopy. Results. Our studies revealed the colocalization between nuclear F-actin and SATB1 protein, during GA-induced death of breast cancer MCF-7 cells. The colocalization was enhanced in cells with overexpressed SATB1 and cofilin-1. At the ultrastructural level the SATB1 and F-actin complexes were seen at the border of condensed and decondensed chromatin. The presence of SATB1/F-actin molecular complexes was confirmed by magnetic separation of F-actin and interacting proteins. Conclusion. We suggest that the molecular interactions between SATB1 and F-actin are necessary for active cell death to occur

Observations of water intake by sucking piglets during various seasons

Celem pracy były obserwacje i ocena ilościowa pobierania wody przez prosięta w okresie ssania do 28 dnia życia. Podjęto próbę porównania ilości pobieranej z poideł wody w czterech różnych okresach, zbliżonych do występowania kalendarzowych pór roku w warunkach klimatycznych Polski. Badania prowadzono w warunkach chlewni towarowej, produkującej tuczniki w cyklu zamkniętym. Obserwacjami objęto 17 miotów pozyskanych w wyniku krzyżowania loch rasy polskiej białej zwisłouchej (pbz) z czterema knurami rasy wielkiej białej polskiej (wbp). Prowadzono je w czasie pełnego roku kalendarzowego w czterech grupach, w zależności od terminu porodu i odchowu: od 1 grudnia do 28 lutego (grupa I), od 1 marca do 31 maja (grupa II), od 1 czerwca do 31 sierpnia (grupa III) i od 1 września do 30 listopada (grupa IV). Liczebność rodzonych miotów, ich masa całkowita i indywidualna masa ciała prosiąt były między grupami zbliżone. Niektóre prosięta (grupa I) rozpoczęły pobieranie wody już od pierwszego dnia życia aczkolwiek były to nieznaczne ilości. Regularne pobieranie przez wszystkie prosięta rozpoczęło się w 3 dniu życia. Największe pobranie wody odnotowano w okresie wiosenno-letnim (grupa II I III), a w przeliczeniu na przyrost 1 kg masy ciała prosiąt największe zużycie wystąpiło w grupie II (marzec-maj). Wysunięto hipotezę, że zwiększone zapotrzebowanie na wodę w miesiącach wiosenno-letnich może wynikać ze zróżnicowania warunków związanych z temperaturą zewnętrzną, długością dnia świetlnego, a przez to zróżnicowaną aktywnością życiową prosiąt. Być może, ma to także związek z objawami atawistycznymi u świń. Wymaga to jednak dalszych badań.The aim of the study was to observe and quantify the intake of water by piglets in the sucking period up to 28 days. An attempt was made to compare the quantity of taken water sinks in four different periods, similar to the occurrence of calendar seasons in Poland\u27s climatic conditions. The research was conducted in the conditions of a pig farm, producing fatteners in a closed cycle. The observations included 17 litters obtained from Polish Landrace (PL) sows mated by four Polish Large White (PLW) boars. They were conducted during the full calendar year in four groups, depending on the date of birth and rearing: from 1 December to 28 February (Group I), from 1 March to 31 May (Group II), from 1 June to 31 August (Group III) and from 1 September to 30 November (Group IV). The number piglets in litters, total weight of litter and individual weight of piglets were similar among groups. Some piglets (Group I) started taking water from the first day of life, but it was a small amount. The regular intake of all piglets started at 3 days of age. The highest water intake was recorded in spring-summer (Group II and III), and as a percentage of 1 kg body weight of piglets, the highest consumption occurred in group II (March-May). It was hypothesized that the increased water requirement in the spring and summer months may be due to the variability of the conditions related to the outdoor temperature, the length of the light day and thus the varied life activities of the piglets. Perhaps, it is also related to atavistic symptoms in pigs. However, this requires further research

Frequency of 22q11.2 microdeletion in children with congenital heart defects in western poland

<p>Abstract</p> <p>Background</p> <p>The 22q11.2 microdeletion syndrome (22q11.2 deletion syndrome -22q11.2DS) refers to congenital abnormalities, including primarily heart defects and facial dysmorphy, thymic hypoplasia, cleft palate and hypocalcaemia. Microdeletion within chromosomal region 22q11.2 constitutes the molecular basis of this syndrome. The 22q11.2 microdeletion syndrome occurs in 1/4000 births. The aim of this study was to determine the frequency of 22q11.2 microdeletion in 87 children suffering from a congenital heart defect (conotruncal or non-conotruncal) coexisting with at least one additional 22q11.2DS feature and to carry out 22q11.2 microdeletion testing of the deleted children's parents. We also attempted to identify the most frequent heart defects in both groups and phenotypic traits of patients with microdeletion to determine selection criteria for at risk patients.</p> <p>Methods</p> <p>The analysis of microdeletions was conducted using fluorescence <it>in situ </it>hybridization (FISH) on metaphase chromosomes and interphase nuclei isolated from venous peripheral blood cultures. A molecular probe (Tuple) specific to the <it>HIRA (TUPLE1, DGCR1</it>) region at 22q11 was used for the hybridisation.</p> <p>Results</p> <p>Microdeletions of 22q11.2 region were detected in 13 children with a congenital heart defect (14.94% of the examined group). Microdeletion of 22q11.2 occurred in 20% and 11.54% of the conotruncal and non-conotruncal groups respectively. Tetralogy of Fallot was the most frequent heart defect in the first group of children with 22q11.2 microdeletion, while ventricular septal defect and atrial septal defect/ventricular septal defect were most frequent in the second group. The microdeletion was also detected in one of the parents of the deleted child (6.25%) without congenital heart defect, but with slight dysmorphism. In the remaining children, 22q11.2 microdeletion originated <it>de novo</it>.</p> <p>Conclusions</p> <p>Patients with 22q11.2DS exhibit wide spectrum of phenotypic characteristics, ranging from discreet to quite strong. The deletion was inherited by one child. Our study suggests that screening for 22q11.2 microdeletion should be performed in children with conotruncal and non-conotruncal heart defects and with at least one typical feature of 22q11.2DS as well as in the deleted children's parents.</p

Crystal Structures of the ATPase Domains of Four Human Hsp70 Isoforms: HSPA1L/Hsp70-hom, HSPA2/Hsp70-2, HSPA6/Hsp70B', and HSPA5/BiP/GRP78

The 70-kDa heat shock proteins (Hsp70) are chaperones with central roles in processes that involve polypeptide remodeling events. Hsp70 proteins consist of two major functional domains: an N-terminal nucleotide binding domain (NBD) with ATPase activity, and a C-terminal substrate binding domain (SBD). We present the first crystal structures of four human Hsp70 isoforms, those of the NBDs of HSPA1L, HSPA2, HSPA5 and HSPA6. As previously with Hsp70 family members, all four proteins crystallized in a closed cleft conformation, although a slight cleft opening through rotation of subdomain IIB was observed for the HSPA5-ADP complex. The structures presented here support the view that the NBDs of human Hsp70 function by conserved mechanisms and contribute little to isoform specificity, which instead is brought about by the SBDs and by accessory proteins.This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1

Results of Polish Adult Leukemia Study Group (PALG) project assessing TP53 mutations with next-generation sequencing technology in relapsed and refractory chronic lymphocytic leukemia patients — an 18-month update

Indtroduction and methods: In chronic lymphocytic leukemia (CLL), molecular and cytogenetic diagnostics are crucial for the determination of accurate prognosis and treatment choice. Among different genetic aberrations, del(17p13) or TP53 mutations constitute high-risk factors, and early identification of such defects is a high priority for CLL patients. While cytogenetic diagnostics is well-established and accessible for the majority of CLL patients in Poland, molecular diagnostics of TP53 mutations is performed only in a few ERIC-certified centers (eight as of September 2020), and only two of these employ next-generation sequencing (NGS) for routine analysis of TP53 status in CLL patients. Here we report the interim results of a project assessing TP53 mutations with NGS technology in relapsed or refractory CLL patients with confirmed negative del(17p13) status. 249 patients from 32 clinical centers were included in the study. Results: NGS analysis revealed TP53 mutations in 42/249 (17%) patients, half of whom (21/249, 8.5%) had subclonal mutations (VAF ≤10%). These results are in line with published data in relapsed/refractory CLL patients. Conclusions: The results of the project demonstrated the feasibility and accuracy of NGS testing in CLL patients despite several initial logistical and technical obstacles. Our study also proved that, with appropriate funding, CLL patients from any hematological center in Poland can have access to state-of-the-art molecular diagnostic

Characterization and Whole Genome Analysis of Human Papillomavirus Type 16 E1-1374^63nt Variants

Background. The variation of the most common Human papillomavirus (HPV) type found in cervical cancer, the HPV16, has been extensively investigated in almost all viral genes. The E1 gene variation, however, has been rarely studied. The main objective of the present investigation was to analyze the variability of the E6 and E1 genes, focusing on the recently identified E1-1374^63nt variant. Methodology/Principal Findings. Variation within the E6 of 786 HPV16 positive cervical samples was analyzed using high-resolution melting, while the E1-1374^63nt duplication was assayed by PCR. Both techniques were supplemented with sequencing. The E1-1374^63nt duplication was linked with the E-G350 and the E-C109/G350 variants. In comparison to the referent HPV16, the E1-1374^63nt E-G350 variant was significantly associated with lower grade cervical lesions (p=0.029), while the E1-1374^63nt E-C109/G350 variant was equally distributed between high and low grade lesions. The E1-1374^63nt variants were phylogenetically closest to E-G350 variant lineage (A2 sub-lineage based on full genome classification). The major differences between E1-1374^63nt variants were within the LCR and the E6 region. On the other hand, changes within the E1 region were the major differences from the A2 sub-lineage, which has been historically but inconclusively associated with high grade cervical disease. Thus, the shared variations cannot explain the particular association of the E1-1374^63nt variant with lower grade cervical lesions. Conclusions/Significance. The E1 region has been thus far considered to be well conserved among all HPVs and therefore uninteresting for variability studies. However, this study shows that the variations within the E1 region could possibly affect cervical disease, since the E1-1374^63nt E-G350 variant is significantly associated with lower grade cervical lesions, in comparison to the A1 and A2 sub-lineage variants. Furthermore, it appears that the silent variation 109T&gt;C of the E-C109/G350 variant might have a significant role in the viral life cycle and warrants further study

The Variability of Urbanization in Lower Silesian Voivodeship, Poland

Includes Tables, Figures, Maps and Bibliography.Most research studies examining Polish urban patterns at the national scale consider only one approach to quantify changes in the urbanization levels: measurement based on demographic data or measurement based on land-cover. With few exceptions, research on urbanization process utilizing multiple datasets in the individual voivodeships is limited. By applying the urban life cycle model using both demographic data, measured using enumeration units derived from Poland's administrative system, and land-cover data, derived from satellite imagery, this research can have a unique contribution to the Polish urbanization literature. Furthermore, the results of this work also have potential implications for urban planning and management at the provincial level by providing a more comprehensive approach to studying the spatial variation of the urban patterns of communes in a voivodeship. The goal of this research is to characterize the urbanization level of Lower Silesian Voivodeship between 1990 and 2010 using both demographic and land-cover data and apply the urban life cycle model to describe urban change within the province. My hypothesis is that a better understanding of urbanization patterns and trends in the province will result by examining urban change at multiple spatial scales and with multiple data sets. Specifically, I addressed the following research questions to evaluate my hypothesis: 1) what are the patterns and trends of urban change in Lower Silesia at each of the three administrative levels; 2) what is the potential spatial, temporal and typological variability in urbanization patterns within the region based on the two datasets; and 3) how does the inclusion of land-cover information better inform the identification of the stage of the urban life cycle model. Specifically, the urban life cycle model used to categorize the urban development is discussed, and previous urbanization studies in the context of population change as well as land-cover change are reviewed. The Background section introduces the Polish administrative system, which varies considerably from the system in the United States, and describes the study site of Lower Silesian Voivodeship. The Methodology section discusses the analyses needed to address each research question. Specifically, the datasets used in the research are discussed in detail, along with the methods used to quantify urban patterns. The Results of the analysis are then presented. The Discussion section synthesizes the results in order to assess variability of urban change in Lower Silesian Voivodeship at multiple scales and to evaluate the inclusion of land-cover information in the urban life cycle model. Finally, the Conclusion section highlights the major findings of the work

Worsening functional status in nephrogeriatrics needs to be accounted for when clinically assessing CKD advancement in addition to GFR; supporting evidence based on the practical application of theoretical modelling

Abstract The incidence of chronic kidney disease (CKD) has been found to increase with age. This has resulted in an increase in the number of elderly patients undergoing renal replacement therapy. There is a significant risk of error in making treatment decisions in patients with advanced CKD based solely on biochemical parameters of renal function, if the changes in the functional status of patients' health are not taken into account. Aim To determine the interrelated dependencies between chronic kidney disease with the functional status of patients aged over 65 years and to elucidate differences in functional status between CKD patients and controls. Methods Patient subjects were qualified according to their assessed outcomes from the study protocol, which were achieved by: geriatric interview, assessing functional status by the IADL, Barthel and Tinetti tests together with assessing kidney function by performing laboratory tests of glomerular filtration rate (GFR), creatinine and urea. Subjects were divided into two groups: method 1—according to GFR and method 2—according to GFR and functional test results. The data were statistically analysed by structural equation modelling and k-means. Results Positive relationships were found between the CKD stage and comorbidity (β = 0.55, p < 0.01), along with the number of medications taken and age (respectively β = 0.19, p = 0.001 and β = 0.30, p < 0.001). A highly negative relationship was observed between the CKD stage and the Tinetti test results (β = -0.71, p < 0.001), whilst more moderate ones were found with the IADL and Barthel scores (respectively β = -0.49, p < 0.001 and β = -0.40, p < 0.001). The patient groups demonstrated differences in health status when selected by method-2 for: age, comorbidity, number of medications taken, fitness test outcomes (Tinetti, Barthel and IADL tests at p < 0.005). Those groups divided according to GFR, however only showed differences in age, comorbidity and the number of medication taken (p < 0.005). Conclusions The functional status worsens in geriatric patients suffering from CKD. It may thus be important to also account for disruptions to functional status when assessing CKD advancement in the elderly in addition to the GFR. The biggest problems for the over 80 s suffering from CKD are gait and balance disorders, leading to a high risk of falls. Another common problem is polypharmacy, found in both the geriatric population and particularly in those suffering from CKD