Description of an Institutional Cohort of Myeloid Neoplasms Carrying ETV6-Locus Deletions or ETV6 Rearrangements.

The gene encoding for transcription factor ETV6 presents recurrent lesions in hematologic neoplasms, most notably the ETV6-RUNX1 rearrangement in childhood B-ALL. The role of ETV6 for normal hematopoiesis is unknown, but loss of its function probably participates in oncogenic procedures. In myeloid neoplasms, ETV6-locus (12p13) deletions are rare but recurrent; ETV6 translocations are even rarer, but those reported seem to have phenotype-defining consequences. We herein describe the genetic and hematologic profile of myeloid neoplasms with ETV6 deletions (10 cases), or translocations (4 cases) diagnosed in the last 10 years in our institution. We find complex caryotype to be the most prevalent cytogenetics among patients with 12p13 deletion (8/10 patients), with most frequent coexisting anomalies being monosomy 7 or deletion 7q32 (5/10), monosomy 5 or del5q14-15 (5/10), and deletion/inversion of chromosome 20 (5/10), and most frequent point mutation being TP53 mutation (6/10 patients). Mechanisms of synergy of these lesions are unknown. We describe the entire genetic profile and hematologic phenotype of cases with extremely rare ETV6 translocations, confirming the biphenotypic T/myeloid nature of acute leukemia associated to ETV6-NCOA2 rearrangement, the association of t (1;12) (p36; p13) and of the CHIC2-ETV6 fusion with MDS/AML, and the association of the ETV6-ACSL6 rearrangement with myeloproliferative neoplasm with eosinophilia. Mutation of the intact ETV6 allele was present in two cases and seems to be subclonal to the chromosomal lesions. Decoding the mechanisms of disease related to ETV6 haploinsufficiency or rearrangements is important for the understanding of pathogenesis of myeloid neoplasms and fundamental research must be guided by observational cues.

Association of Gestational Hypertension with Sociodemographic and Anthropometric Factors, Perinatal Outcomes, Breastfeeding Practices, and Mediterranean Diet Adherence: A Cross-Sectional Study.

Background and Objectives: Gestational hypertension has been associated with several pregnancy short-term and long-term complications, affecting both the mother and her infant's health. The present study aims to assess the potential association of gestational hypertension with sociodemographic and anthropometry factors, perinatal outcomes, breastfeeding habits, and Mediterranean diet (MD) compliance. Materials and Methods: This is a cross-sectional study conducted on 5271 mothers that was carried out after delivery. The anthropometry characteristics and perinatal outcomes were retrieved from the mothers' medical records. Sociodemographic characteristics, MD adherence, and breastfeeding habits were assessed via one-to-one interviews of the assigned women with qualified staff. Results: Maternal older age, being employed, family history of gestational hypertension, overweight/obesity before gestation, and abnormal gestational weight gain (GWG) independently increased the risk of developing gestational hypertension. Moreover, gestational hypertension was independently related with a greater incidence of abnormal childbirth body weight and preterm birth, not exclusively breastfeeding, and lower levels of MD adherence. Conclusions: This study highlights the importance of informing future mothers about the risk factors of gestational hypertension, underlining also that a healthy lifestyle, which simultaneously includes a healthy nutritional pattern such as MD, may decrease the risk of developing gestational hypertension and the subsequent pregnancy complications.

Systemic mastocytosis accompanied by a non-secretory plasma cell dyscrasia and nephrotic syndrome-level proteinuria in a 76-year-old patient.

We report here the interesting case of a 76-year-old man with severe proteinuria who was diagnosed with systemic mastocytosis accompanied by a clonal non-mast-cell lineage haematological disorder (a non-secretory plasma cell dyscrasia). This is a unique report of systemic mastocytosis with a non-secretory plasma cell dyscrasia and nephrotic syndrome. The pathophysiological relevance between these entities along with the probability of occult amyloidosis is discussed.

POZ-, AT-hook-, and zinc finger-containing protein (PATZ) interacts with human oncogene B cell lymphoma 6 (BCL6) and is required for its negative autoregulation.

The PATZ1 gene encoding a POZ/AT-hook/Kruppel zinc finger (PATZ) transcription factor, is considered a cancer-related gene because of its loss or misexpression in human neoplasias. As for other POZ/domain and Kruppel zinc finger (POK) family members, the transcriptional activity of PATZ is due to the POZ-mediated oligomer formation, suggesting that it might be not a typical transactivator but an architectural transcription factor, thus functioning either as activator or as repressor depending on the presence of proteins able to interact with it. Therefore, to better elucidate PATZ function, we searched for its molecular partners. By yeast two-hybrid screenings, we found a specific interaction between PATZ and BCL6, a human oncogene that plays a key role in germinal center (GC) derived neoplasias. We demonstrate that PATZ and BCL6 interact in germinal center-derived B lymphoma cells, through the POZ domain of PATZ. Moreover, we show that PATZ is able to bind the BCL6 regulatory region, where BCL6 itself acts as a negative regulator, and to contribute to negatively modulate its activity. Consistently, disruption of one or both Patz1 alleles in mice causes focal expansion of thymus B cells, in which BCL6 is up-regulated. This phenotype was almost completely rescued by crossing Patz1(+/-) with Bcl6(+/-) mice, indicating a key role for Bcl6 expression in its development. Finally, a significant number of Patz1 knock-out mice (both heterozygous and homozygous) also develop BCL6-expressing lymphomas. Therefore, the disruption of one or both Patz1 alleles may favor lymphomagenesis by activating the BCL6 pathway.

Acquired Clotting Factor Deficits During Treatment with Asparaginase in an Institutional Cohort.

We invariably see prolongation of activated partial thromboplastin time in patients treated with asparaginase in our clinical practice, but have noted that, contrary to hypofibrinogenemia and low antithrombin, clotting times' prolongation by asparaginase is largely unreported in the literature and guidelines and is not widely known to clinicians. We report on aPTT prolongations in a small cohort of patients, and on their origin, as investigated by measurements of clotting factors, fibrinogen, and D-dimers before and after asparaginase administration. We observed significant reductions in FIX and FXI (median post-treatment values of 27 IU/dl and 52 IU/dl, respectively), confirming one previous observation. A decrease in FXII was less pronounced but contributed to the prolonged aPTTs (FXII has no effect on in vivo haemostasis). The factor deficits are not due to consumption, as evidenced by unchanged D-dimer levels, and are, therefore, probably caused by disturbed factor synthesis. Our observations and insights contribute to elucidation of the profile of clotting assays during asparaginase treatment, and thus, to optimally monitor for undesirable events or steer situations of therapeutic anticoagulation without the risk of suboptimal or excessive anticoagulation.

Deciphering the induction of Listeria monocytogenes into sublethal injury using fluorescence microscopy and RT-qPCR.

The adaptive response of bacterial cells to changing environmental conditions depends on the behavior of single cells within the population. Exposure of Listeria monocytogenes to sublethal acidic conditions in foods or in the gastrointestinal track of the host may induce injuries relevant to difficult physiological states within the dormancy continuum. In this study, exposure to acidic conditions (acetic-AA and hydrochloric acid-HCl adjusted to pH 3.0, 2.7, 2.5 at 20 °C for 5 h) was used to evaluate injury of L. monocytogenes, Scott A strain. To differentiate the resistant sub-population from the total, Tryptic Soy Agar with 0.6 % Yeast Extract (TSAYE) supplemented or not with 5 % NaCl were comparatively used. Sublethally injured cells were detected by comparing plate counts with fluorescence microscopy, using combinations of CFDA (viability) and Propidium-Iodide (death). Effect of acid stress on the relative transcription of clpP, mazE, mazF, relA, gadC, gadD, gadB, sigB, inlA and prfA upon transition of total population into different physiological stages was evaluated through RT-qPCR. AA treated cells showed measurable logarithmic reduction at pH 2.7 and 2.5, while there was a significant percentage of CFDA-/PI+ cells. Evaluation of the potentially culturable population on TSAYE, from the percentage of CFDA/PI-stained cells, revealed that unstained cells represented a non-culturable sub-population. Exposure to Ringer's solution pH 2.7, adjusted with AA, resulted in higher percentages of non-esterase active with membrane integrity cells (CFDA-/PI-) compared to the percentages of the enumerated culturable cells on TSAYE after 4 and 5 h. Under the same conditions, after 1 h of exposure macroscopic observation revealed size colony variations (SCVs) of the total population (CFU on TSAYE). L. monocytogenes retained its culturability after hydrochloric acid exposure, while cells remained metabolically active (CFDA+). However, a stochastic change in cell's shape, was detected after exposure to pH 3.0 and 2.5, adjusted with HCl, for 2 h at 20 °C. A pattern of gene up-regulation was observed during treatment with AA pH 2.7 and HCl pH 3.0 at the 3rd h of exposure. Deciphering L. monocytogenes sublethal injury sheds light into the physiological and molecular characteristics of this state and provides the food science community with quantitative data to improve risk assessment.

Outcome of Involuntary Mental Health Assessment in a Psychiatric Department in Greece.

Despite their controversiality, involuntary admissions in psychiatric departments remain a central issue in mental health care. The present study aims to identify demographic and clinical factors possibly associated with emergency involuntary psychiatric assessment and its outcome in Greece. This study was carried out in the psychiatric department of the University General Hospital of Alexandroupolis (UGHA) from 1 March 2018 to 28 February 2019. The sample included 191 individuals who had been psychiatrically assessed without their consent following a prosecutorial order. The majority of the involuntary assessments resulted in hospitalization (71%), with 51% of them resulting in involuntary hospitalization. Almost all patients diagnosed with "F20-29 schizophrenia, schizotypal and delusional disorders" were subsequently admitted to the psychiatric department of the UGHA (77 of 81, 66 of them involuntarily). Higher admission rates were recorded among those who had been referred from the Prosecutor's Office of regions that are located far from the psychiatric department of UGHA (Fisher's exact test, p-value = 0.045). In multivariate logistic regression, prior contact with psychiatric services and having an "F20-29 schizophrenia, schizotypal and delusional disorders" diagnosis was statistically significant with admission to the hospital as an outcome variable. Our study suggests an increased risk of involuntary admission among patients with psychosis, patients who had visited a psychiatric service prior to their assessment as well as those living further away from the main psychiatric services of the hospital. Better organization of community psychiatric services in remote places from hospital central services may lead to fewer prosecutorial referrals and coercive measures.

Single-center, observational study of AML/MDS-EB with IDH1/2 mutations: genetic profile, immunophenotypes, mutational kinetics and outcomes.

OBJECTIVE: IDH1/2 mutations, intervening in epigenetic procedures, are frequently encountered in acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). Knowledge of the genetics, immunophenotypes, and mutational kinetics of IDH1/2-mutated AML can contribute to the understanding of AML clonal architecture and inform therapeutics and monitoring. METHODS: We retrospectively analyzed 50 IDH1/2-mutated AML/MDS-EB cases of our institution, to identify recurrent co-mutations, immunophenotypes, patterns of co-variance of IDH1/2 allele burdens with those of recurrent co-mutations, frequency of persistent IDH1/2 mutation as clonal hematopoiesis of indeterminate potential (CHIP) in remission and response to hypomethylating agents. RESULTS: Most frequently co-mutated genes were DNMT3A, SRSF2 and NPM1. Most cases with co-existent IDH1/2 and NPM1 mutations (11/13) showed an 'APL-like' immunophenotype (CD34-HLADR-). Allele burdens of mutated IDH1/2 were identical to mutated SRSF2 allele burdens at diagnosis and remission, but not always to mutated NPM1 allele burden in remission. We show persistence of significant mutIDH1/2 allele burden in approximately one-fourth of patients with deep remissions. IDH1/2 mutations were significantly more frequent among responders to first-line HMA-based regimens than among non-responders, in patients treated for myeloid neoplasms with excess blasts. CONCLUSIONS: IDH1/2 mutations are most frequently accompanied by DNMT3A, SRSF2 and NPM1 mutations. NPM1-IDH1/2 mutated AML has a mature phenotype possibly amenable to differentiation therapies. IDH1/2 and SRSF2 mutations probably arise at the same developmental stage of the disease, as their allele burdens covariate. IDH1/2 mutation represents CHIP in a substantial proportion of cases and is therefore no reliable residual disease marker. The preferential presence of IDH1/2 mutations among HMA-responders could inform therapeutic decisions if confirmed in larger series.

Mediterranean Diet and Sarcopenia Features in Apparently Healthy Adults over 65 Years: A Systematic Review.

Low muscle mass combined with changes in physical function and muscle quality is defined as sarcopenia. In people > 60 years, sarcopenia reaches 10% and tends to increase with age. Individual nutrients, such as protein, may have a protective role against sarcopenia, but recent evidence suggests that protein alone has been ineffective in increasing muscle strength. Dietary patterns, instead, with a high "anti-inflammatory" potential, such as the Mediterranean dietary pattern, have been considered as an emerging dietary remedy against sarcopenia. The aim of this systematic review was to summarize the evidence of the role of Mediterranean diet in sarcopenia prevention and/or improvement, including recent data, in healthy elders. We searched published studies about sarcopenia and the Mediterranean diet until December 2022 in Pubmed, Cochrane, Scopus search engine and grey literature. In total, ten articles were identified as relevant: four cross-sectional studies and six prospective. No clinical trial was identified. Only three studies assessed sarcopenia presence and four measured muscle mass, which is an essential criterion in sarcopenia diagnosis. Mediterranean diet adherence had, in general, a positive role in muscle mass and muscle function, while the results were less clear with regard to muscle strength. Additionally, there was no evidence of a positive effect of the Mediterranean diet on sarcopenia. There is a need for conduction of clinical trials in order to reach cause-effects conclusions regarding the importance of the Mediterranean diet in sarcopenia prevention and management in Mediterranean and non-Mediterranean populations.

The Effect of Various, Everyday Practices on Glucose Levels in Critically Ill Children.

BACKGROUND: To evaluate the effect of various, everyday intensive care unit (ICU) practices on glucose levels in critically ill pediatric patients with the use of a continuous glucose monitoring system. METHODS: Seventeen sensors were placed in 16 pediatric patients (8 male). All therapeutic and diagnostic interventions were recorded and 15 minutes later, a flash glucose measurement was obtained by swiping the sensor with a reader. Glucose difference was calculated as the glucose value 15 minutes after the intervention minus the mean daily glucose value for each individual patient. Additionally, the consciousness status of the patient (awake or sedated) was recorded. RESULTS: Two hundred and five painful skin interventions were recorded. The mean difference of glucose values was higher by 1.84 ± 14.76 mg/dL (95% CI: -0.19 to 3.87 mg/dL, P = .076). However, when patients were categorized regarding their consciousness level, mean glucose difference was significantly higher in awake state than in sedated patients (4.76 ± 28.07 vs -2.21 ± 15.77 mg/dL, P < .001). Six hundred forty-nine interventions involving the respiratory system were recorded. Glucose difference during washings proved to be significantly higher than the ones during simple suctions (4.74 ± 14.18 mg/dL vs 0.32 ± 18.22 mg/dL, P = .016). Finally, glucose difference in awake patients was higher by 3.66 ± 13.91 mg/dL compared to glucose difference of -2.25 ± 21.07 mg/dL obtained during respiratory intervention in sedated patients. CONCLUSIONS: Diagnostic and therapeutic procedures in the ICU, especially when performed in an awake state, exacerbate the stress and lead to a significant rise in glucose levels.

ZEB1 and CtBP form a repressive complex at a distal promoter element of the BCL6 locus.

BCL6 is essential for normal antibody responses and is highly expressed in germinal centre B-cells. Constitutive expression due to chromosomal translocations or mutations of cis-acting regulatory elements contributes to diffuse large B-cell lymphoma. BCL6 expression is therefore tightly regulated in a lineage- and developmental-stage-specific manner, and disruption of normal controls can contribute to lymphomagenesis. In order to discover potential cis-acting control regions we carried out DNase I-hypersensitive site mapping. Gel-shift assays and chromatin immunoprecipitation of the core region of a hypersensitive site 4.4 kb upstream of BCL6 transcription initiation (HSS-4.4) showed an E-box element-binding ZEB1 (zinc finger E-boxbinding homeobox 1) and the co-repressor CtBP (C-terminal binding protein). As compared with peripheral blood B-cells, ZEB1, a two-handed zinc finger transcriptional repressor, is expressed at relatively low levels in germinal centre cells, whereas BCL6 has the opposite pattern of expression. Transfection of ZEB1 cDNA caused a reduction in BCL6 expression and a mutated ZEB1, incapable of binding CtBP, lacked this effect. siRNA (small interfering RNA)-mediated knockdown of ZEB1 or CtBP produced an increase in BCL6 mRNA. We propose that HSS-4.4 is a distal promoter element binding a repressive complex consisting of ZEB1 and CtBP. CtBP is ubiquitously expressed and the results of the present study suggest that regulation of ZEB1 is required for control of BCL6 expression.

Listeria monocytogenes Sublethal Injury and Viable-but-Nonculturable State Induced by Acidic Conditions and Disinfectants.

The dormancy continuum hypothesis states that in response to stress, cells enter different stages of dormancy ranging from unstressed living cells to cell death, in order to ensure their long-term survival under adverse conditions. Exposure of Listeria monocytogenes cells to sublethal stressors related to food processing may induce sublethal injury and the viable-but-nonculturable (VBNC) state. In this study, exposure to acetic acid (AA), hydrochloric acid (HCl), and two disinfectants, peracetic acid (PAA) and sodium hypochlorite (SH), at 20°C and 4°C was used to evaluate the potential induction of L. monocytogenes strain Scott A into different stages of dormancy. To differentiate the noninjured subpopulation from the total population, tryptic soy agar with 0.6% yeast extract (TSAYE), supplemented or not with 5% NaCl, was used. Sublethally injured and VBNC cells were detected by comparing plate counts obtained with fluorescence microscopy and by using combinations of carboxyfluorescein and propidium iodide (viable/dead cells). Induction of sublethal injury was more intense after PAA treatment. Two subpopulations were detected, with phenotypes of untreated cells and small colony variants (SCVs). SCVs appeared as smaller colonies of various sizes and were first observed after 5 min of exposure to 5 ppm PAA at 20°C. Increasing the stress intensity from 5 to 40 ppm PAA led to earlier detection of SCVs. L. monocytogenes remained culturable after exposure to 20 and 30 ppm PAA for 3 h. At 40 ppm, after 3 h of exposure, the whole population was considered nonculturable, while cells remained metabolically active. These results corroborate the induction of the VBNC state. IMPORTANCE Sublethally injured and VBNC cells may evade detection, resulting in underestimation of a food product's microbial load. Under favorable conditions, cells may regain their growth capacity and acquire new resistant characteristics, posing a major threat for public health. Induction of the VBNC state is crucial for foodborne pathogens, such as L. monocytogenes, the detection of which relies almost exclusively on the use of culture recovery techniques. In the present study, we confirmed that sublethal injury is an initial stage of dormancy in L. monocytogenes that is followed by the VBNC state. Our results showed that PAA induced SCVs (a phenomenon potentially triggered by external factors) and the VBNC state in L. monocytogenes, indicating that tests of lethality based only on culturability may provide false-positive results regarding the effectiveness of an inactivation treatment.

Complex Modes of Inheritance in Hereditary Red Blood Cell Disorders: A Case Series Study of 155 Patients.

Hereditary erythrocytes disorders include a large group of conditions with heterogeneous molecular bases and phenotypes. We analyzed here a case series of 155 consecutive patients with clinical suspicion of hereditary erythrocyte defects referred to the Medical Genetics Unit from 2018 to 2020. All of the cases followed a diagnostic workflow based on a targeted next-generation sequencing panel of 86 genes causative of hereditary red blood cell defects. We obtained an overall diagnostic yield of 84% of the tested patients. Monogenic inheritance was seen for 69% (107/155), and multi-locus inheritance for 15% (23/155). PIEZO1 and SPTA1 were the most mutated loci. Accordingly, 16/23 patients with multi-locus inheritance showed dual molecular diagnosis of dehydrated hereditary stomatocytosis/xerocytosis and hereditary spherocytosis. These dual inheritance cases were fully characterized and were clinically indistinguishable from patients with hereditary spherocytosis. Additionally, their ektacytometry curves highlighted alterations of dual inheritance patients compared to both dehydrated hereditary stomatocytosis and hereditary spherocytosis. Our findings expand the genotypic spectrum of red blood cell disorders and indicate that multi-locus inheritance should be considered for analysis and counseling of these patients. Of note, the genetic testing was crucial for diagnosis of patients with a complex mode of inheritance.

Biopharmaceutics classification systems for new molecular entities (BCS-NMEs) and marketed drugs (BCS-MD): theoretical basis and practical examples.

The aim of this work is to develop biopharmaceutics classification systems for new molecular entities (BCS-NMEs) and marketed drugs (BCS-MD). The kinetics of gastrointestinal (GI) wall permeation and dissolution were re-considered theoretically. The relationships between the solubility/dose ratio and the fractions of dose dissolved and absorbed, were also examined. Mean time calculations for drug dissolution (MDT) and permeation (MPT) of the GI wall were analyzed in respect to the mean intestinal transit time (MITT) to identify a cutoff point for drug dissolution and GI wall permeation. Dissolution experiments for marketed drugs were carried out. NMEs were classified into four classes of BCS-NMEs, based on solubility/dose ratio and apparent permeability estimates. A physiologically based cutoff time point for dissolution and permeation was used to differentiate rapidly from slowly dissolving-permeating marketed drugs, which were classified into four classes of BCS-MD using their dissolution index (DI=MITT/MDT) and permeation index (PI=MITT/MPT) values as follows: I (DI>or=3, PI>or=3), II (DI<3, PI>or=3), III (DI>or=3, PI<3) and IV (DI<3, PI<3). In conclusion, two classification systems were developed, one for NMEs based on solubility/dose ratio and permeability estimates and one for marketed drugs based on MDT and MPT estimates.

On the use of the Weibull function for the discernment of drug release mechanisms.

Previous findings from our group based on Monte Carlo simulations indicated that Fickian drug release from Euclidian or fractal matrices can be described with the Weibull function. In this study, the entire drug release kinetics of various published data and experimental data from commercial or prepared controlled release formulations of diltiazem and diclofenac are analyzed using the Weibull function. The exponent of time b of the Weibull function is linearly related to the exponent n of the power law derived from the analysis of the first 60% of the release curves. The value of the exponent b is an indicator of the mechanism of transport of a drug through the polymer matrix. Estimates for b< or =0.75 indicate Fickian diffusion in either fractal or Euclidian spaces while a combined mechanism (Fickian diffusion and Case II transport) is associated with b values in the range 0.75

Expression, prognostic significance and mutational analysis of protein tyrosine phosphatase SHP-1 in chronic myeloid leukemia.

The protein tyrosine phosphatase SHP-1 dephosphorylates BCR-ABL1, thereby serving as a potential control mechanism of BCR-ABL1 kinase activity. Pathways regulating SHP-1 expression, which could be exploited in the therapeutics of TKI-resistant chronic myeloid leukemia (CML), remain unknown. Moreover, the questions of whether there is any kind of SHP-1 deregulation in CML, contributing to disease initiation or evolution, as well as the question of prognostic significance of SHP-1, have not been definitively answered. This study shows moderately lower SHP-1 mRNA expression in chronic phase CML patients in comparison to healthy individuals and no change in SHP-1 mRNA levels after successful TKI treatment. Mutational analysis of the aminoterminal and phosphatase domains of SHP-1 in patients did not reveal genetic lesions. This study also found no correlation of SHP-1 expression at diagnosis with response to treatment, although a trend for lower SHP-1 expression was noted in the very small non-responders' group of the 3-month therapeutic milestone.