Utility investigation of automated techniques in hematopoietic progenitor cell count and viability assessment in the Good Manufacturing Practice (GMP) settingg

Aim: To compare our parameters as regards: i) cell count via two different automated cell count techniques, and ii) viability via automated trypan blue exclusion and 7-aminoactinomycin D (7-AAD) staining. Method: We used the trypan blue exclusion technique and an automated cell counter and for viability testing, and the trypan blue exclusion technique and the 7-AAD evaluation by flow cytometry. The trypan blue exclusion and the radio frequency techniques were used for automated cell counting. Flow cytometric analysis was performed by evaluating the yielded cellular products for 7-AAD uptake during the cell count of CD34+ cells. Results: The mean values for cell count were estimated as 3.44±1.22x106/ml (range, 2.48-5.71x106/ml) and 4.14±1.94x106/ml (range, 1.77-7.43x106/ml) for the trypan blue exclusion and radio frequency techniques, respectively. Additionally, the mean values for viability analyses via the automated trypan blue exclusion and 7-AAD were 93.38±6.09% (range, 79.00-98.00%) and 99.49±0.60% (range, 98.40-100.00%), respectively. Conclusions: Our study has responded to two fundamental questions: whether the results of both of the automated techniques for cell count correspond with each other, and whether the results of the automated viability assessment conform those of the 7-AAD technique during the manufacturing processes of cellular therapy products intended for clinical use. Even though we have the opportunity to use the hemocytometer in our laboratory setting, the automated trypan blue exclusion technique gives cell count results in concordance within the range of the expectations of our Quality Management System (QMS)

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Relationship between catheter related cerebrospinal fluid infections and systemic immune-inflammation index

Background This study investigated the relationship between the systemic immune inflammation index (SII) and catheter-related infections and their effects on prognosis in pediatric patients. Methods A total of 56 pediatric patients diagnosed with ventriculoperitoneal (V-P) shunt infection between January 2017 and October 2019 were included. V-P shunt infection diagnosis was made based on clinical findings. All cerebrospinal fluid (CSF) samples were subjected to direct microscopic examination and culture. Protein, glucose, and sodium levels in CSF, CSF leukocytes, and hematological and biochemical parameters were measured. Results Fifty-six patients with growth in CSF culture were included in this study. 55.4% of the cases were female and 44.6% male. V-P shunt was detected in 82.1% of the cases and external ventricular drainage (EVD) catheter-related infection in 17.9%. The CSF/blood glucose ratio was significantly lower (p = 0.046), and SII was significantly increased (p = 0.002) in non-coagulase-negative staphylococci. Conclusions Early and appropriate antibiotic therapy reduces morbidity and mortality in catheter-related infections. However, it is important to start empirical antibiotherapy until culture results are expected. Therefore, further research on the estimation of possible factors is needed

IDENTIFICATION OF GENETIC DIVERSITY IN WILD PEAR (Pyrus Elaeagrifolia Pall.) GENOTYPES COLLECTED FROM DIFFERENT REGIONS OF TURKEY WITH SSR MARKER SYSTEM

Uzun A., H. Pinar, M. Yaman, M. A. Yigit, Y. Cakiroglu, A. Karakaya, M. Uysal, G. wild pear (Pyrus elaeagrifolia pall.) genotypes collected from different regions of Turkey with SSR marker system. - Genetika, Vol 54, No.1, 109-118. Turkey with diverse ecologies is among the unique countries in terms of plant species and diversity. Among these plant species, naturally growing wild pears (Pyrus elaeagrifolia Pall.) are resistant to chlorosis and drought and could be used in rootstock development programs. In present study, genetic diversity in 96 wild pear genotypes collected from 11 investigated with the use of SSR (Simple Sequence Repeat) molecular marker system. Present analyses carried out in ABI (Applied Biosystem) 3500 capillary electrophoresis system revealed 93 scorable and all polymorphic bands, thus polymorphism rate was 100%. In UPGMA (Unweighted pair group method with arithmetic mean) dendrogram of wild pear genotypes, similarity index values varied between 0.20-0.83 and a large variation was observed among the genotypes. Present finding may have significant contributions to further studies to be conducted for preservation of gene sources and breeding of wild pear genotypes

Effects of personality traits on severity of sepsis

Introduction: The aim of this study was to reveal the effect of the individual's lifestyle and personality traits on the disease process in patients with sepsis and to have clinical predictions about these patients

Tocilizumab treatment in severe COVID-19: a multicenter retrospective study with matched controls

Coronavirus disease-2019 (COVID-19) associated pneumonia may progress into acute respiratory distress syndrome (ARDS). Some patients develop features of macrophage activation syndrome (MAS). Elevated levels of IL-6 were reported to be associated with severe disease, and anti-IL-6R tocilizumab has been shown to be effective in some patients. This retrospective multicenter case-control study aimed to evaluate the efficacy of tocilizumab in hospitalized COVID-19 patients, who received standard of care with or without tocilizumab. Primary outcome was the progression to intubation or death. PSMATCH (SAS) procedure was used to achieve exact propensity score (PS) matching. Data from 1289 patients were collected, and study population was reduced to 1073 based on inclusion-exclusion criteria. The composite outcome was observed more frequently in tocilizumab-users, but there was a significant imbalance between arms in all critical parameters. Primary analyses were carried out in 348 patients (174 in each arm) after exact PS matching according to gender, ferritin, and procalcitonin. Logistic regression models revealed that tocilizumab significantly reduced the intubation or death (OR 0.40, p = 0.0017). When intubation is considered alone, tocilizumab-users had > 60% reduction in odds of intubation. Multiple imputation approach, which increased the size of the matched patients up to 506, provided no significant difference between arms despite a similar trend for intubation alone group. Analysis of this retrospective cohort showed more frequent intubation or death in tocilizumab-users, but PS-matched analyses revealed significant results for supporting tocilizumab use overall in a subset of patients matched according to gender, ferritin and procalcitonin levels

What should be the appropriate minimal duration for patient examination and evaluation in pulmonary outpatient clinics?

INTRODUCTION: Patient examinations performed in a limited time period may lead to impairment in patient and physician relationship, defective and erroneous diagnosis, inappropriate prescriptions, less common use of preventive medicine practices, poor patient satisfaction, and increased violent acts against health-care staff