A comparison of qPCR and ddPCR used for quantification of the "JAK2 V617F" allele burden in Ph negative MPNs

Philadelphia-negative myeloproliferative neoplasms (MPNs) are a diverse group of diseases whose common feature is the presence of V617F mutation of the JAK2 gene. In the era of novel therapeutic strategies in MPNs, such as JAK-inhibitor therapy, there is a growing need for establishing high sensitive quantitativemethods, which can be useful not only at diagnosis but also for monitoring therapeutic outcomes, such as minimal residual disease (MRD). In this study, we compared the qPCR and ddPCR methods and their clinical utility for diagnosis, prognostication, and treatment monitoring of MPNs with JAK2 V617F mutation in 63 MPN patients of which 6 were subjected to ruxolitinib treatment.We show a high conformance between the two methods (correlation coefficient r = 0.998 (p < 0.0001)). Our experiments revealed high analytical sensitivity for both tests, suggesting that they are capable of detecting the JAK2 V617F mutation at diagnosis of MPN with a limit of detection (LoD) of 0.12% for qPCR and 0.01% for ddPCR. The alterations of JAK2 V617F allele burden in patients treated with ruxolitinib were measured by both methods with equal accuracy. The results suggest an advantage of ddPCR in monitoring MRD because of allele burdens below the LoD of qPCR. Overall, the clinical utility of qPCR and ddPCR is very high, and both methods could be recommended for the routine detection of the V617F mutation at diagnosis, though ddPCR will probably supersede qPCR in the future due to costeffectiveness

Spinal cord tethering in children with myelomeningocele Zakotwiczenie rdzenia u dzieci z przepukliną oponowo-rdzeniową

STRESZCZENIE Zespół zakotwiczenia rdzenia kręgowego (TCS, ang. tethered cord syndrome) jest zróżnicowaną jednostką chorobową, spowodowaną przez nadmierne naciąganie rdzenia kręgowego. Może występować w każdej grupie wiekowej, a objawia się w zależności od stanu patologicznego i wieku bólem, zmianami skórnymi, deformacjami ortopedycznymi, neurologicznymi i urologicznymi. Wczesna interwencja chirurgiczna jest związana z poprawą obrazu klinicznego, głównie w ustąpieniu dolegliwości bólowych, stabilizacji funkcji neurologicznych i postępu skoliozy. Zespół rehabilitacyjny prowadzący terapię dziecka z przepukliną oponowo-rdzeniową powinien pamiętać, że TSC jest istotnym powodem pogorszenia stanu neurologicznego u dziecka z rozszczepem kręgosłupa. Słowa kluczowe: zakotwiczenie rdzenia, przepuklina oponowo-rdzeniowa, rehabilitacja ABSTRACT We retrospectively assessed tethered cord syndrome among patients with myelomeningocele in the Department of Pediatric Rehabilitation in the years 2004-2011. Fifteen (16.5% of 91 subjects with myelomeningocele) patients with tethered cord syndrome were diagnosed, aged 4-17 years (mean 11.4 ± 4.4), 11 girls and 4 boys. Reduced muscle strength and increased spasticity within the lower limbs, pain in the thoracolumbar region, and deterioration of bladder function were symptoms of tethered cord syndrome. The level of spinal cord injury in Th8-L3 prevented 9 patients from walking (use of a wheelchair); 6 children -level of spinal cord injury L4 and below -were able to walk with orthopedic equipment (crutches, walker, severely handicapped). MRI revealed the various pathologic etiologies of tethering included: lipomyelomeningocele, diastematomyelia, syringohydromyelia, and hydrosyringomyelia. A neurological or rehabilitation team working with children with myelomeningocele should remember that tethered cord syndrome is a significant cause of neurological deterioration

Early hematopoietic chimerism monitoring and molecular engraftment characteristic by STR-PCR method in patients after alloHSCT

Allogeniczna transplantacja komórek hematopoetycznych jest jednym ze sposobów leczenia nowotworowych i nienowotworowych schorzeń hematologicznych. Po przeszczepieniu komórki hematopoetyczne dawcy osiedlają się w niszach szpikowych biorcy, co skutkuje powstaniem prawidłowego układu krwiotwórczego i immunologicznego o genotypie dawcy - całkowity chimeryzm dawcy. Wszczepienie molekularne (ME) poprzedza wystąpienie wszczepienia hematologicznego. Wczesna ocena chimeryzmu może mieć istotne znaczenie dla dalszego przebiegu procesu przyjmowania się przeszczepienia i warunkuje możliwość podjęcia wczesnej interwencji leczniczej.Do badania włączono 38 pacjentów, u których wykonano 43 allogeniczne transplantacje (alloHSCT). Przyjmowanie się przeszczepu śledzono we krwi obwodowej od 2. do 14. doby po transplantacji, następnie w 21. i 28. dobie. W 30. dobie poziom chimeryzmu oznaczano we krwi obwodowej i w szpiku kostnym. Chimeryzm hematopoetyczny oceniano przy zastosowaniu metody molekularnej STR-PCR.Wykazano, że zmiany poziomu chimeryzmu we wczesnym okresie po transplantacji w grupie pacjentów poddanych alloSCT (alloHSCT po-przedzona kondycjonowaniem mieloablacyjnym) przebiegają zgodnie z zależnością liniową (R2=0,996), natomiast w grupie pacjentów poddanych alloNMSCT (alloHSCT poprzedzona kondycjonowaniem niemie-loablacyjnym) są zgodne z zależnością logarytmiczną (R2=0,959). Poziom chimeryzmu hematopoetycznego jest wyższy w grupie pacjentów poddanych alloSCT, w 2. dobie różnicę tę cechuje znamienność statystyczna (p=0,0048).Wszczepienie molekularne poprzedza wystąpienie wszczepienia hematologicznego (pacjenci poddani alloSCT p=1,44×10−12, pacjenci pod-dani alloNMSCT p=2,12×10−6).W grupach pacjentów poddanych alloSCT i alloHSCT, którzy otrzymali więcej niż 3×106 komórek CD34+/kg masy data różnica w czasie wystąpienia ME w porównaniu z grupą chorych, którzy otrzymali mniej niż 3×106 komórek CD34+/kg, była znamienna statystycznie (alloSCT p=0,0013, alloHSCT p = 0,021).Allogeneic hematopoietic stem cell transplantation (alloHSCT) is a curative treatment for proportion of patients suffering from malignant and non-malignant hematological disorders. Successful transplantation is a process that requires the engraftment of pluripotent hematopoietic stem cells which can re-establish normal hemopoesis and immune system. Distinguishing between donor and host origin of bone marrow and blood cells is crucial for monitoring of engraftment process. One of the most useful tools for engraftment monitoring is the assessment of hematopoietic chimerism after alloSCT that describes the percentage of donor hematopoietic in a transplant recipient.Thirty eight adult patients after alloHSCT were included into the study. In total 43 allogeneic stem cell transplantations were performed. Hematopoietic chimerism was assessed by STR-PCR technique. The analysis of early chimerism were performed starting from 2nd to 14th day on every 2 days than to 28 days weekly and on day +30 after alloHSCT.Early hematopoietic chimerism assessment demonstrated that the kinetics of chimerism in patients after alloSCT was compatible with linear trend (R2=0.996) and in patients after alloNMSCT was compatible with logarithmic trend (R2=0.959). The hematopoietic chimerism level was higher in alloSCT on day 2 the difference was statistically significant (p=0.0048).Molecular engraftment preceded hematological engraftment in patients after either myeloablative or non-myeloablative conditioning regiments (alloSCT patients p=1.44×10−12, alloNMSCT p=2.12×10−6).Earlier ME was observed in patients after alloHSCT and alloSCT who received more than 3×106 CD34+ cells/kg (alloSCT p=0.0013, alloHSCT p=0.021). The difference was statistically significant

Wytyczne dla laboratoriów genetyki nowotworów litych

Niniejsze wytyczne skierowane są do laboratoriów wykonujących diagnostyczne badania genetyczne technikami biologii molekularnej i cytogenetyki molekularnej (FISH) w zakresie genetyki nowotworów litych zarówno w obszarze zaburzeń genetycznych dziedzicznych, jak i nabytych. Diagnostyczne badanie genetyczne jest wykonywane w celu identyfikacji zaburzeń w DNA komórek człowieka. Przestrzeganie niniejszych zasad ma na celu zapewnienie wysokiego poziomu usług medycznych świadczonych przez laboratoria

Interplay of cell-cell contacts and RhoA/MRTF-A signaling regulates cardiomyocyte identity.

Cell-cell and cell-matrix interactions guide organ development and homeostasis by controlling lineage specification and maintenance, but the underlying molecular principles are largely unknown. Here, we show that in human developing cardiomyocytes cell-cell contacts at the intercalated disk connect to remodeling of the actin cytoskeleton by regulating the RhoA-ROCK signaling to maintain an active MRTF/SRF transcriptional program essential for cardiomyocyte identity. Genetic perturbation of this mechanosensory pathway activates an ectopic fat gene program during cardiomyocyte differentiation, which ultimately primes the cells to switch to the brown/beige adipocyte lineage in response to adipogenesis-inducing signals. We also demonstrate by in vivo fate mapping and clonal analysis of cardiac progenitors that cardiac fat and a subset of cardiac muscle arise from a common precursor expressing Isl1 and Wt1 during heart development, suggesting related mechanisms of determination between the two lineages

An analysis of certification processes for Good Clinical Practice and project management competencies

Aim/purpose - The purpose of this paper is to clarify the certification process of the Good Clinical Practice (GCP) competencies based on a comparison with the project management (PM) certification process. This aim was accomplished by model development. Originality/value/contribution - The paper raises research topics at the crossroads of project management, clinical trials, and GCP, topics that have been underdeveloped so far. The results might be significant for all organizations involved in conducting clinical research projects. The findings contribute to the quality of clinical trials and provide public assurance that the rights, safety, and well-being of trial subjects are protected and the clinical trial data are credible. Design/methodology/approach - The study was divided into three main phases: 1st phase - the identification of key characteristics of PM and GCP certification processes, 2nd phase - the development of certification models for PM and GCP, 3rd phase - the conclusions from in-depth interviews. Findings - As a result of the research, key characteristics of PM and GCP certification processes were identified, certification models for PM and GCP were developed. In addition, based on conclusions from in-depth interviews, solutions for organizing the way of confirming knowledge of GCP guidelines were proposed. Research implications/limitations - The proposed rules may be too complex and may exceed the needs and expectations of the clinical trial environment. The models focus on stakeholder relations, without the rules of certification granting procedures, to enable broader contextualization of the issues discussed in the paper. The analysis might be fragmented as it regards the sole certification process. The research refers to Poland only, hence it will be valuable to identify whether foreign authorities have the same attitude to the GCP competency certification model. It is recommended to increase the number of respondents' interviews to obtain results of higher validity and reliability

A Report from the Interdisciplinary Academic Conference Old Polish Literature. Reading Texts, History and Culture (Pobierowo, 19–21 October 2017)

A Report from the Conference 'Old Polish Literature. Reading Texts, History and Culture' (October 2017).Sprawozdanie z konferencji 'Literatura staropolska. Czytanie tekstów, historia i kultura' (październik 2017)

Phytochemical Profiling, Antioxidant and Anti-Inflammatory Activity of Plants Belonging to the Lavandula Genus

Lavender is a valuable medicinal plant belonging to the Lamiaceae family. Currently 39 species are known, but only Lavandula angustifolia is a pharmacopoeial raw material. Lavender has a long history of medicinal use and mainly exhibits antioxidant, anti-inflammatory, sedative, antidepressant, spasmolytic, anticholinesterases, antifungal and antibacterial properties. Used internally, it relieves symptoms of mental stress and insomnia and supports digestion. Topical use of lavender in aromatherapy, neuralgia and antiseptics is also known. The constant interest in lavender, and in particular in Lavandula angustifolia, in the field of medicine and pharmacy is evidenced by the growing number of publications. In view of so many studies, it seems important to review traditional and modern extraction techniques that determine the chemical composition responsible for the antioxidant and anti-inflammatory effects of various extracts from the species of the Lavandula genus

Profile of Bioactive Components and Antioxidant Activity of <i>Aronia melanocarpa</i> Fruits at Various Stages of Their Growth, Using Chemometric Methods

Aronia (chokeberry, Aronia melanocarpa) is a valuable fruit that combines the health benefits of green tea and red wine and is gaining popularity worldwide. Aronia has a unique chemical composition with strong antioxidant properties, including anthocyanins and chlorogenic acids (CGAs). However, it remains unclear which specific compounds or groups are primarily responsible for the antioxidant properties of chokeberry. Therefore, an analysis of the antioxidant properties of aronia berries based on geographical region and their variability during ripening (from green to frostbitten fruit) was conducted. The fruits were collected from three locations for two years. The aim of our work was to identify the compounds responsible for the antioxidant properties of aronia berry extracts by using chemometric methods. The analyses of anthocyanins and CGAs were performed using HPLC-DAD, and the antioxidant capacity was assessed by FRAP and DPPH methods. The PCA analysis also considered variations in temperature and precipitation. The chemometric analysis revealed a strong correlation between radical-scavenging properties and the content levels of chlorogenic acids. The results obtained in this study show that unripe green chokeberry fruits exhibit the highest antioxidant properties, which can be attributed to the high content of CGAs at this stage