The results of a close follow-up of trainees to gain a good blood collection practice

BackgroundPhlebotomy is one of the most important steps in the preanalytical phase of a clinical laboratory process. In order to decrease phlebotomy errors, this specific procedure should be taught in detail by laboratory organizations. Our study aims to practice the training program on venous blood sampling and observe the close follow-up results.MethodsIn this observational study, 127 students who started their summer internship in Antalya Education and Research Hospital were given a one-day theoretical phlebotomy training in accordance with the Venous Blood Sampling Guidelines. After the theoretical training, phlebotomy applications of 10 students who were working in the field of out-patient blood sampling were observed both with and without their knowledge. A comprehensive checklist related to phlebotomy was created by the trainers in Antalya Education and Research Hospital and the observers answered each question as yes or no. For the statistical analysis, IBM SPSS Statistics 21.0 was used.ResultsAfter the theoretical education, the trainees were observed but no significant difference was found between the first and the second informed observations (p = 0.125). The students were observed three times more in the following week without their knowledge. There was a statistically significant difference between the first and the third unannounced observations (p=0.001).ConclusionsIn order to perform phlebotomy correctly, apart from theoretical education, a close follow-up is necessary too.     

Lipid peroxidation and scavenging enzyme levels in the liver of streptozotocin-induced diabetic rats

In this study, alterations in the liver antioxidant enzymes status and lipid peroxidation in short-term (8-weeks) and long-term (24-weeks) diabetic rats were examined. Glutathione peroxidase (GSH-Px) activity and malondialdehyde (MDA) levels were significantly increased, but superoxide dismutase (SOD) activity was significantly reduced in 8-weeks diabetic rats, compared to control. Catalase (CAT) activity, however, was found unchanged. In 24-weeks diabetic rats, while GSH-Px activity was unchanged, but SOD and CAT activities and MDA levels were significantly increased, compared to control. These results suggest that diabetes-induced alterations in tissue antioxidant system may reflect a generalized increase in tissue oxidative stress. It can be concluded that lipid peroxidation and antioxidant enzyme levels are elevated in diabetic condition. Hence, diabetes mellitus, if left untreated, may increase degenerative processes due to accumulation of oxidative free radicals.     

β-Arrestin-1 links mitogenic sonic hedgehog signaling to the cell cycle exit machinery in neural precursors

Development of the cerebellum, a brain region regulating posture and coordination, occurs post-natally and is marked by rapid proliferation of granule neuron precursors (CGNPs), stimulated by mitogenic Sonic hedgehog (Shh) signaling. β-Arrestin (βArr) proteins play important roles downstream of Smoothened, the Shh signal transducer. However, whether Shh regulates βArrs and what role it plays in Shh-driven CGNP proliferation remains to be determined. Here, we report that Shh induces βArr1 accumulation and localization to the nucleus, where it participates in enhancing expression of the cyclin dependent kinase (cdk) inhibitor p27, whose accumulation eventually drives CGNP cell cycle exit. βArr1 knockdown enhances CGNP proliferation and reduces p27 expression. Thus, Shh-mediated βArr1 induction represents a novel negative feedback loop within the Shh mitogenic pathway, such that ongoing Shh signaling, while required for CGNPs to proliferate, also sets up a cell-intrinsic clock programming their ultimate exit from the cell cycle.Key words: sonic hedgehog, cerebellum, neural precursor, β-arrestin 1, p27, differentiation     

Cytoplasmic thioredoxin reductase is essential for embryogenesis but dispensable for cardiac development

Two distinct thioredoxin/thioredoxin reductase systems are present in the cytosol and the mitochondria of mammalian cells. Thioredoxins (Txn), the main substrates of thioredoxin reductases (Txnrd), are involved in numerous physiological processes, including cell-cell communication, redox metabolism, proliferation, and apoptosis. To investigate the individual contribution of mitochondrial (Txnrd2) and cytoplasmic (Txnrd1) thioredoxin reductases in vivo, we generated a mouse strain with a conditionally targeted deletion of Txnrd1. We show here that the ubiquitous Cre-mediated inactivation of Txnrd1 leads to early embryonic lethality. Homozygous mutant embryos display severe growth retardation and fail to turn. In accordance with the observed growth impairment in vivo, Txnrd1-deficient embryonic fibroblasts do not proliferate in vitro. In contrast, ex vivo-cultured embryonic Txnrd1-deficient cardiomyocytes are not affected, and mice with a heart-specific inactivation of Txnrd1 develop normally and appear healthy. Our results indicate that Txnrd1 plays an essential role during embryogenesis in most developing tissues except the heart.     

PROGRESS STUDY: Progression of chronic kidney disease in children and heat shock proteins

Various molecular and cellular processes are involved in renal fibrosis, such as oxidative stress, inflammation, endothelial cell injury, and apoptosis. Heat shock proteins (HSPs) are implicated in the progression of chronic kidney disease (CKD). Our aim was to evaluate changes in urine and serum HSP levels over time and their relationships with the clinical parameters of CKD in children. In total, 117 children with CKD and 56 healthy children were examined. The CKD group was followed up prospectively for 24 months. Serum and urine HSP27, HSP40, HSP47, HSP60, HSP70, HSP72, and HSP90 levels and serum anti-HSP60 and anti-HSP70 levels were measured by ELISA at baseline, 12 months, and 24 months. The urine levels of all HSPs and the serum levels of HSP40, HSP47, HSP60, HSP70, anti-HSP60, and anti-HSP70 were higher at baseline in the CKD group than in the control group. Over the months, serum HSP47 and HSP60 levels steadily decreased, whereas HSP90 and anti-HSP60 levels steadily increased. Urine HSP levels were elevated in children with CKD; however, with the exception of HSP90, they decreased over time. In conclusion, our study demonstrates that CKD progression is a complicated process that involves HSPs, but they do not predict CKD progression. The protective role of HSPs against CKD may weaken over time, and HSP90 may have a detrimental effect on the disease course.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12192-021-01239-9.     

Monoallelic and Biallelic Variants in EMC1 Identified in Individuals with Global Developmental Delay,Hypotonia, Scoliosis,and Cerebellar Atrophy

The paradigm of a single gene associated with one specific phenotype and mode of inheritance has been repeatedly challenged. Genotype-phenotype correlations can often be traced to different mutation types, localization of the variants in distinct protein domains, or the trigger of or escape from nonsense-mediated decay. Using whole-exome sequencing, we identified homozygous variants in EMC1 that segregated with a phenotype of developmental delay, hypotonia, scoliosis, and cerebellar atrophy in three families. In addition, a de novo heterozygous EMC1 variant was seen in an individual with a similar clinical and MRI imaging phenotype. EMC1 encodes a member of the endoplasmic reticulum (ER)-membrane protein complex (EMC), an evolutionarily conserved complex that has been proposed to have multiple roles in ER-associated degradation, ER-mitochondria tethering, and proper assembly of multi-pass transmembrane proteins. Perturbations of protein folding and organelle crosstalk have been implicated in neurodegenerative processes including cerebellar atrophy. We propose EMC1 as a gene in which either biallelic or monoallelic variants might lead to a syndrome including intellectual disability and preferential degeneration of the cerebellum.     

Assay for adhesion and agar invasion in S. cerevisiae

Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, biofilms can reduce industrial productivity, destroy structures, and threaten human life. 1-3 On the other hand, harnessing the power of biofilms can help clean the environment and generate sustainable energy. 4-8 The ability of S. cerevisiae to colonize surfaces and participate in complex biofilms was mostly ignored until the rediscovery of the differentiation programs triggered by various signaling pathways and environmental cues in this organism. 9, 10 The continuing interest in using S. cerevisiae as a model organism to understand the interaction and convergence of signaling pathways, such as the Ras-PKA, Kss1 MAPK, and Hog1 osmolarity pathways, quickly placed S. cerevisiae in the junction of biofilm biology and signal transduction research. 11-20 To this end, differentiation of yeast cells into long, adhesive, pseudohyphal filaments became a convenient readout for the activation of signal transduction pathways upon various environmental changes. However, filamentation is a complex collection of phenotypes, which makes assaying for it as if it were a simple phenotype misleading. In the past decade, several assays were successfully adopted from bacterial biofilm studies to yeast research, such as MAT formation assays to measure colony spread on soft agar and crystal violet staining to quantitatively measure cell-surface adherence. 12, 21 However, there has been some confusion in assays developed to qualitatively assess the adhesive and invasive phenotypes of yeast in agar. Here, we present a simple and reliable method for assessing the adhesive and invasive quality of yeast strains with easy-to-understand steps to isolate the adhesion assessment from invasion assessment. Our method, adopted from previous studies, 10, 16 involves growing cells in liquid media and plating on differential nutrient conditions for growth of large spots, which we then wash with water to assess adhesion and rub cells completely off the agar surface to assess invasion into the agar. We eliminate the need for streaking cells onto agar, which affects the invasion of cells into the agar. In general, we observed that haploid strains that invade agar are always adhesive, yet not all adhesive strains can invade agar medium. Our approach can be used in conjunction with other assays to carefully dissect the differentiation steps and requirements of yeast signal transduction, differentiation, quorum sensing, and biofilm formation.     

Essential role for mitochondrial thioredoxin reductase in hematopoiesis, heart development, and heart function

Oxygen radicals regulate many physiological processes, such as signaling, proliferation, and apoptosis, and thus play a pivotal role in pathophysiology and disease development. There are at least two thioredoxin reductase/thioredoxin/peroxiredoxin systems participating in the cellular defense against oxygen radicals. At present, relatively little is known about the contribution of individual enzymes to the redox metabolism in different cell types. To begin to address this question, we generated and characterized mice lacking functional mitochondrial thioredoxin reductase (TrxR2). Ubiquitous Cre-mediated inactivation of TrxR2 is associated with embryonic death at embryonic day 13. TrxR2(TrxR2(-/-)minus;/TrxR2(-/-)minus;) embryos are smaller and severely anemic and show increased apoptosis in the liver. The size of hematopoietic colonies cultured ex vivo is dramatically reduced. TrxR2-deficient embryonic fibroblasts are highly sensitive to endogenous oxygen radicals when glutathione synthesis is inhibited. Besides the defect in hematopoiesis, the ventricular heart wall of TrxR2(TrxR2(-/-)minus;/TrxR2(-/-)minus;) embryos is thinned and proliferation of cardiomyocytes is decreased. Cardiac tissue-restricted ablation of TrxR2 results in fatal dilated cardiomyopathy, a condition reminiscent of that in Keshan disease and Friedreich's ataxia. We conclude that TrxR2 plays a pivotal role in both hematopoiesis and heart function.     

Comparison of dose distributions and organs at risk (OAR) doses in conventional tangential technique (CTT) and IMRT plans with different numbers of beam in left-sided breast cancer

Aim

Our aim was to improve dose distribution to the left breast and to determine the dose received by the ipsilateral lung, heart, contralateral lung and contralateral breast during primary left-sided breast irradiation by using intensity modulated radiotherapy (IMRT) techniques compared to conventional tangential techniques (CTT). At the same time, different beams of IMRT plans were compared to each other in respect to CI, HI and organs at risk (OAR) dose.

Background

Conventional early breast cancer treatment consists of lumpectomy followed by whole breast radiation therapy. CTT is a traditional method used for whole breast radiotherapy and includes standard wedged tangents (two opposed wedged tangential photon beams). The IMRT technique has been widely used for many treatment sites, allowing both improved sparing of normal tissues and more conformal dose distributions. IMRT is a new technique for whole breast radiotherapy. IMRT is used to improve conformity and homogeneity and used to reduce OAR doses.

Materials and methods

Thirty patients with left-sided breast carcinoma were treated between 2005 and 2008 using 6, 18 or mixed 6/18 MV photons for primary breast irradiation following breast conserving surgery (BCS). The clinical target volume [CTV] was contoured as a target volume and the contralateral breast, ipsilateral lung, contralateral lung and heart tissues as organs at risk (OAR). IMRT with seven beams (IMRT7), nine beams (IMRT9) and 11 beams (IMRT11) plans were developed and compared with CTT and among each other. The conformity index (CI), homogeneity index (HI), and doses to OAR were compared to each other.

Results

All of IMRT plans significantly improved CI (CTT: 0.76; IMRT7: 0.84; IMRT9: 0.84; IMRT11: 0.85), HI (CTT: 1.16; IMRT7: 1.12; IMRT9: 1.11; IMRT11: 1.11), volume of the ipsilateral lung receiving more than 20 Gy (>V_20 Gy) (CTT: 14.6; IMRT7: 9.08; IMRT9: 8.10; IMRT11: 8.60), and volume of the heart receiving more than 30 Gy (>V_30 Gy) (CTT: 6.7; IMRT7: 4.04; IMRT9: 2.80; IMRT11: 2.98) compared to CTT. All IMRT plans were found to significantly decrease >V_20 Gy and >V_30 Gy volumes compared to conformal plans. But IMRT plans increased the volume of OAR receiving low dose radiotherapy: volume of contralateral lung receiving 5 and 10 Gy (CTT: 0.0–0.0; IMRT7: 19.0–0.7; IMRT9: 17.2–0.66; IMRT11: 18.7–0.58, respectively) and volume of contralateral breast receiving 10 Gy (CTT: 0.03; IMRT7: 0.38; IMRT9: 0.60; IMRT11: 0.68). The differences among IMRT plans with increased number of beams were not statistically significant.

Conclusion

IMRT significantly improved conformity and homogeneity index for plans. Heart and lung volumes receiving high doses were decreased, but OAR receiving low doses was increased.     

The drug-transporter gene MDR1 C3435T and G2677T/A polymorphisms and the risk of multidrug-resistant epilepsy in Turkish children

One-third of all individuals with epilepsy are resistant to antiepileptic drug (AED) treatment. Antiepileptic treatment response has been suggested to be modulated by genetic polymorphisms of drug efflux transporters. Several polymorphic variants within the multidrug resistance 1 (MDR1) gene, which encodes the major transmembrane efflux transporter P-glycoprotein, have been proposed to be associated with AED resistance in epilepsy patients. The aim of this study was to evaluate the effect of C3435T and G2677T/A polymorphisms of MDR1 on AED resistance in Turkish children with epilepsy. MDR1 C3435T and G2677T/A were genotyped in 152 patients with epilepsy, classified as drug-resistant in 69 and drug-responsive in 83. Genotypes of the C3435T and G2677T/A polymorphisms were determined by polymerase chain reaction followed by restriction fragment length polymorphism. Genotype and allele frequencies of C3435T and G2677T/A polymorphisms of the MDR1 gene did not differ between drug-resistant and drug-responsive epilepsy patients. Our results suggest that MDR1 C3435T and G2677T/A polymorphisms are not associated with AED resistance in Turkish epileptic patients. To clarify the exact clinical implication of the MDR1 polymorphisms on the multidrug resistance in epilepsy, further investigations in various ethnic populations would be necessary.