Search

In this paper, we present the synthesis of four new complexes: the dimeric precursor [Ir(dmppz) 2 ( μ -Cl)] 2 (1) (Hdmppz - 3,5-dimethyl-1-phenyl-1 H -pyrazole) and heteroleptic bis-cyclometalated complexes: [Ir(dmppz) 2 (Py 2 CO)]PF 6 ·½CH 2 Cl 2 (2), [Ir(dmppz) 2 (H 2 biim)]PF 6 ·H 2 O (3), and [Ir(dmppz) 2 (PyBIm)]PF 6 (4), with auxiliary N,N-donor ligands: 2-di(pyridyl)ketone (Py 2 CO), 2,2'-biimidazole (H 2 biim) and 2-(2'-pyridyl)benzimidazole (PyBIm). In the obtained complexes, SC-X-ray analysis revealed that Ir(III) has an octahedral coordination sphere with chromophores of the type {IrN 2 C 2 Cl 2 } (1) or {IrN 4 C 2 } (2-4). The complexes obtained, which have been fully characterised by physicochemical methods (CHN, TG, FTIR, UV-Vis, PL and 1 H, 13 C, 15 N NMR), were used to continue our studies on the factors influencing the cytotoxic properties of potential chemotherapeutic agents ( in vitro ). To this end, the following studies are presented: (i) comparative analysis of the effects on the biological properties of N,N-donor ligands and C,N-donor ligands in the studied complexes, (ii) studies of the interactions of the compounds with the selected molecular target: DNA and BSA (UV-Vis, CD and PL methods), (iii) and the reactivity towards redox molecules: GSH, NADH (UV-Vis and/or ESI-MS methods), (iv) cytotoxic activity (IC 50 ) of potential chemotherapeutics against MCF-7, K-562 and CCRF-CEM cell lines.

As the heterogeneity of symptoms is increasingly recognized among long-COVID patients, it appears highly relevant to study potential pathophysiological differences along the different subtypes. Preliminary evidence suggests distinct alterations in brain structure and systemic inflammatory patterns in specific groups of long-COVID patients. To this end, we analyzed differences in cortical thickness and peripheral immune signature between clinical subgroups based on 3 T-MRI scans and signature inflammatory markers in n = 120 participants comprising healthy never-infected controls (n = 30), healthy COVID-19 survivors (n = 29), and subgroups of long-COVID patients with (n = 26) and without (n = 35) cognitive impairment according to screening with Montreal Cognitive Assessment. Whole-brain comparison of cortical thickness between the 4 groups was conducted by surface-based morphometry. We identified distinct cortical areas showing a progressive increase in cortical thickness across different groups, starting from healthy individuals who had never been infected with COVID-19, followed by healthy COVID-19 survivors, long-COVID patients without cognitive deficits (MoCA ≥ 26), and finally, long-COVID patients exhibiting significant cognitive deficits (MoCA < 26). These findings highlight the continuum of cortical thickness alterations associated with COVID-19, with more pronounced changes observed in individuals experiencing cognitive impairment (p < 0.05, FWE-corrected). Affected cortical regions covered prefrontal and temporal gyri, insula, posterior cingulate, parahippocampal gyrus, and parietal areas. Additionally, we discovered a distinct immunophenotype, with elevated levels of IL-10, IFNγ, and sTREM2 in long-COVID patients, especially in the group suffering from cognitive impairment. We demonstrate lingering cortical and immunological alterations in healthy and impaired subgroups of COVID-19 survivors. This implies a complex underlying pathomechanism in long-COVID and emphasizes the necessity to investigate the whole spectrum of post-COVID biology to determine targeted treatment strategies targeting specific sub-groups., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

As a remedy against stress and anxiety, cannabidiol (CBD) products are of increasing interest in veterinary medicine. Limited data is available describing the actual effectiveness of CBD in horses. The aim of this study (part 2 of 2) was to analyze stress parameters via behavioral observation, heart rate monitoring and assessment of blood and saliva cortisol levels in healthy horses treated repeatedly with a CBD containing paste. Twelve horses were randomly assigned to a treatment or a control group. Two pastes were orally administered in a double-blinded study design, one paste containing CBD and one paste without active ingredient. Both pastes were administered twice daily over 15 days (dose: 3 mg CBD/kg). Behavioral observations were conducted daily using a sedation score and a rating of facial expressions, based on the previously described facial sedation scale for horses (FaceSed) and the Horse Grimace Scale. Blood and saliva samples were obtained regularly to determine cortisol levels throughout the study. Cortisol levels were analyzed by means of liquid chromatography/tandem mass spectrometry (LC/MS/MS). Behavioral observations and cortisol levels were compared between groups. Prior to paste administration, a novel object test was performed and the horses' reaction to loading on a trailer was recorded. Both tests were repeated after 13 days of paste application. Movement patterns such as different gaits during the novel object test were evaluated and an ethogram was designed to assess exhibited behavioral traits. Cardiac beat-to-beat (R-R) intervals were recorded throughout and evaluated using heart rate (HR) and heart rate variability (HRV) parameters. Blood and saliva samples for cortisol analysis were taken before and after the tests. Daily behavioral observations and cortisol levels did not differ between the treatment and the control group. Similarly, analysis of movement patterns, HR, HRV and cortisol levels during the novel object test and trailer test did not identify significant differences between the groups. Regularly administered oral CBD (3 mg/kg BID over 15 days) had no statistically significant effect on behavioral observations, cortisol levels, HR and HRV in horses. Further research is required to establish adequate doses and indications for the use of CBD in horses., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Eichler, Ehrle, Machnik, Jensen, Wagner, Baudisch, Bolk, Pötzsch, Thevis, Bäumer, Lischer and Wiegard.)

The global cancer burden remains high; thus, a better understanding of the molecular mechanisms driving carcinogenesis is needed to improve current prevention and treatment options. We previously detected the ZNF643/ZFP69B gene upregulated in multiple tumors, and we speculated it may play a role in tumor biology. To test this hypothesis, we employed TCGA-centered databases to correlate ZNF643 status with various clinicopathological parameters. We also performed RNA-seq analysis and in vitro studies assessing cancer cell phenotypes, and we searched for ZNF643-bound genomic loci. Our data indicated higher levels of ZNF643 in most analyzed tumors compared to normal samples, possibly due to copy number variations. ZNF643 mRNA correlated with diverse molecular and immune subtypes and clinicopathological features (tumor stage, grade, patient survival). RNA-seq analysis revealed that ZNF643 silencing triggers the deregulation of the genes implicated in various cancer-related processes, such as growth, adhesion, and immune system. Moreover, we observed that ZNF643 positively influences cell cycle, migration, and invasion. Finally, our ChIP-seq analysis indicated that the genes associated with ZNF643 binding are linked to adhesion and immune signaling. In conclusion, our data confirm the oncogenic properties of ZNF643 and pinpoint its impact on cell adhesion and immune processes.

Despite the ongoing progress in diagnosis and treatments, cancer remains a threat to more than one-third of the human population. The emerging data indicate that many Krüppel-associated box zinc finger proteins (KRAB-ZNF) belonging to a large gene family may be involved in carcinogenesis. Our previous study identified Zinc Finger Protein 714 (ZNF714), a KRAB-ZNF gene of unknown function, as being commonly overexpressed in many tumors, pointing to its hypothetical oncogenic role. Here, we harnessed The Cancer Genome Atlas (TCGA)-centered databases and performed functional studies with transcriptomic and methylomic profiling to explore ZNF714 function in cancer. Our pan-cancer analyses confirmed frequent ZNF714 overexpression in multiple tumors, possibly due to regional amplification, promoter hypomethylation, and Nuclear Transcription Factor Y Subunit Beta (NFYB) signaling. We also showed that ZNF714 expression correlates with tumor immunosuppressive features. The in vitro studies indicated that ZNF714 expression positively associates with proliferation, migration, and invasion. The transcriptomic analysis of ZNF714 knocked-down cells demonstrated deregulation of cell adhesion, migration, proliferation, apoptosis, and differentiation. Importantly, we provided evidence that ZNF714 negatively regulates the expression of several known TSGs indirectly via promoter methylation. However, as ZNF714 did not show nuclear localization in our research model, the regulatory mechanisms exerted by ZNF714 require further investigation. In conclusion, our results reveal, for the first time, that ZNF714 may support pro-oncogenic features in lung cancer cells.

Cannabidiol (CBD) products gain increasing popularity amongst animal owners and veterinarians as an alternative remedy for treatment of stress, inflammation or pain in horses. Whilst the use of cannabinoids is banned in equine sports, there is limited information available concerning CBD detection times in blood or urine. The aim of this study was to determine the pharmacokinetic properties of CBD following oral administration in the horse to assist doping control laboratories with interpreting CBD analytical results. Part 1: dose escalation study: Single oral administration of three escalating doses of CBD paste (0.2 mg/kg, n  = 3 horses; 1 mg/kg, n  = 3; 3 mg/kg, n  = 5) with >7 days wash-out periods in between. Part 2: multiple dose study: oral administration of CBD paste (3 mg/kg, n  = 6) twice daily for 15 days. Multiple blood and urine samples were collected daily throughout both studies. Following study part 2, blood and urine samples were collected for 2 weeks to observe the elimination phase. Concentrations of CBD, its metabolites and further cannabinoids were evaluated using gas-chromatography/tandem-mass-spectrometry. Pharmacokinetic parameters were assessed via two approaches: population pharmacokinetic analysis using a nonlinear mixed-effects model and non-compartmental analysis. AUC 0-12 h and C max were tested for dose proportionality. During the elimination phase, the CBD steady-state urine to serum concentration ratio (Rss) was calculated. Oral CBD medication was well-tolerated in horses. Based on population pharmacokinetics, a three-compartment model with zero-order absorption most accurately described the pharmacokinetic properties of CBD. High volumes of distribution into peripheral compartments and high concentrations of 7-carboxy-CBD were observed in serum. Non-compartmental analysis identified a C max of 12.17 ± 2.08 ng/mL after single administration of CBD (dose: 3 mg/kg). AUC 0-12 h showed dose proportionality, increase for C max leveled off at higher doses. Following multiple doses, the CBD terminal half-life was 161.29 ± 43.65 h in serum. Rss was 4.45 ± 1.04. CBD is extensively metabolized and shows high volumes of tissue distribution with a resulting extended elimination phase. Further investigation of the potential calming and anti-inflammatory effects of CBD are required to determine cut-off values for medication control using the calculated Rss., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Eichler, Poźniak, Machnik, Schenk, Wingender, Baudisch, Thevis, Bäumer, Lischer and Ehrle.)

Clozapine is a frequently used antipsychotic that, in case of overdose, can cause severe adverse side effects, such as hematological, cardiovascular, and neurological complications. As there is no specific antidote or reversal agent available, extracorporeal techniques such as CytoSorb hemoadsorption might represent a viable option, having already been used in a variety of intoxication scenarios with favorable rates of success. A 56-year-old male was admitted with generalized epileptic seizures and arrhythmias following ingestion of clozapine in a suicide attempt (5,000 mg). Subsequently, conventional supportive care was initiated. To accelerate drug removal, continuous veno-venous hemodiafiltration including the application of CytoSorb hemoadsorption therapy was started. Serial measurements confirmed rapid reduction of clozapine plasma levels. The patient remained hemodynamically stable throughout this period. Furthermore, there were no cardiac arrhythmias detected and liver values were normal. The patient improved and was successfully extubated 3 days after admission with good vigilance and no residual neurological abnormalities. This is the first clinical case report on the use of CytoSorb hemoadsorption in severe clozapine intoxication which helped quickly and efficiently reduce clozapine levels to nontoxic serum levels while preserving organ function. Therefore, CytoSorb might represent an alternative treatment modality to be considered for potentially lethal clozapine intoxications., Competing Interests: The authors have no conflicts of interest to declare., (© 2023 The Author(s). Published by S. Karger AG, Basel.)

The combination of sulfadoxine (SDO) with trimethoprim (TMP) is widely used in veterinarian medicine. The aim of the present study was to compare excretion profiles and detection time windows of SDO and TMP in plasma and urine by means of a validated quantitative method. Eight horses received a single intravenous (i.v.) dose of 2.7 mg TMP and 13.4 mg SDO per kg bodyweight. Plasma and urine samples were collected up to 15 and 70 days post-administration, respectively. While urine samples underwent an enzymatic hydrolysis, plasma samples were proteolysed before further analysis. After solid-phase extraction, samples were analysed by liquid chromatography/electrospray ionisation tandem mass spectrometry in positive ionisation mode. The applied multiple reaction monitoring (MRM) method allowed the detection of SDO and TMP with a lower limit of detection of 0.03 ng/mL in plasma and 0.2 (SDO) and 0.4 ng/mL (TMP) in urine, respectively. In the present study, detection times for SDO were 15 days in plasma and 49 days in urine, respectively. TMP was detected for up to 7 days in plasma and up to 50 days in urine, respectively. The detection via the TMP metabolite 3-desmethyl-trimethoprim was possible for 70 days in urine. Detection times of the other confirmed metabolites N 4 -acetylated sulfadoxine, hydroxytrimethoprim, trimethoprim-1-oxide and trimethoprim-3-oxide were significantly lower. In order to postulate reasonable screening limits (SLs) to control specific withdrawal times, a Monte Carlo simulation was performed for SDO. The proposed SL of 10 ng/mL SDO in blood and 300 ng/mL urine corresponds to a detection time of 4 days., (© 2023 The Authors. Drug Testing and Analysis published by John Wiley & Sons Ltd.)

Introduction: Post-COVID syndrome is increasingly recognized as a new clinical entity after SARS-CoV-2 infection. Patients living in rural areas may have to travel long with subjectively great effort to be examined using all necessary interdisciplinary tools. This problem could be addressed with mobile outpatient clinics., Methods: In this prospective observational study, we investigated physical fitness, fatigue, depression, cognitive dysfunction, and dyspnea in patients with post-COVID syndrome in a mobile interdisciplinary post-COVID outpatient clinic. Upon referral from their primary care physician, patients were offered an appointment at a mobile post-COVID outpatient clinic close to their home., Results: We studied 125 patients (female, n = 79; 63.2%) in our mobile unit. All patients reported symptoms lasting for more than 12 weeks after acute infection. 88.3% and 64.1% of patients reported significant impairment in physical and mental quality of life. Patients reported a median of three symptoms. The most frequently reported symptoms were fatigue (86.4%), cognitive dysfunction (85.6%), and dyspnea (37.6%). 56.0% of patients performed at < 2.5th percentile at the 1 min sit-to-stand test compared to age- and sex-matched healthy controls, and 25 patients (20.0%) exhibited a drop in oxygen saturation. A questionnaire given to each patient regarding the mobile unit revealed a very high level of patient satisfaction., Conclusion: There is an increasing need for high-quality and locally available care for patients with post-COVID syndrome. A mobile post-COVID outpatient clinic is a new concept that may be particularly suitable for use in rural regions. Patients' satisfaction following visits in such units is very high., (© 2022. The Author(s).)

Head and neck squamous cell carcinoma (HNSCC) is one of the ten most common cancers. Most cancer cases originate from alcohol and tobacco consumption. However, studies have demonstrated that human papillomavirus ( HPV ) infection, particularly HPV-16 , may also significantly influence disease progression. The KRAB-ZNF family of genes is involved in epigenetic suppression, and its involvement in carcinogenesis is the subject of extensive studies. The available literature data demonstrate that they may play different roles, both as tumor suppressors and oncogenes. In this study, six ZNF genes, ZFP28 , ZNF132 , ZNF418 , ZNF426 , ZNF540 , and ZNF880 , were tested using several in silico approaches based on the TCGA and GEO datasets. Our analyses indicate that the expression of the analyzed ZNFs was significantly downregulated in tumor tissues and depended on tumor localization. The expression levels of ZNFs differed between HPV -positive vs. HPV -negative patients depending on the clinical-pathological parameters. More specifically, the patients with higher levels of ZNF418 and ZNF540 showed better survival rates than those with a lower expression. In addition, the level of ZNF540 expression in HPV -positive ( HPV(+) ) patients was higher than in HPV -negative ( HPV(-) ) patients ( p < 0.0001) and was associated with better overall survival (OS). In conclusion, we demonstrate that ZNF540 expression highly correlates with HPV infection, which renders ZNF540 a potential biomarker for HNSCC prognosis and treatment.

Neuropsychiatric symptoms are the most common sequelae of long-COVID. As accumulating evidence suggests an impact of survived SARS-CoV-2-infection on brain physiology, it is necessary to further investigate brain structural changes in relation to course and neuropsychiatric symptom burden in long-COVID. To this end, the present study investigated 3T-MRI scans from long-COVID patients suffering from neuropsychiatric symptoms (n = 30), and healthy controls (n = 20). Whole-brain comparison of gray matter volume (GMV) was conducted by voxel-based morphometry. To determine whether changes in GMV are predicted by neuropsychiatric symptom burden and/or initial severity of symptoms of COVID-19 and time since onset of COVID-19 stepwise linear regression analysis was performed. Significantly enlarged GMV in long-COVID patients was present in several clusters (spanning fronto-temporal areas, insula, hippocampus, amygdala, basal ganglia, and thalamus in both hemispheres) when compared to controls. Time since onset of COVID-19 was a significant regressor in four of these clusters with an inverse relationship. No associations with clinical symptom burden were found. GMV alterations in limbic and secondary olfactory areas are present in long-COVID patients and might be dynamic over time. Larger samples and longitudinal data in long-COVID patients are required to further clarify the mediating mechanisms between COVID-19, GMV and neuropsychiatric symptoms., Competing Interests: Declaration of Competing Interest Outside of the study, PAR received consulting and lecture fees from CSL Behring, Dr. Wilmar Schwabe, Boston Scientific, Pfizer, Bristol Myers Squibb and travel grants from Merz Pharma. All other authors state, that they have no conflict of interests., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Somatic mutations accumulating over a patient's lifetime are well-defined causative factors that fuel carcinogenesis. It is now clear, however, that epigenomic signature is also largely perturbed in many malignancies. These alterations support the transcriptional program crucial for the acquisition and maintenance of cancer hallmarks. Epigenetic instability may arise due to the genetic mutations or transcriptional deregulation of the proteins implicated in epigenetic signaling. Moreover, external stimulation and physiological aging may also participate in this phenomenon. The epigenomic signature is frequently associated with a cell of origin, as well as with tumor stage and differentiation, which all reflect its high heterogeneity across and within various tumors. Here, we will overview the current understanding of the causes and effects of the altered and heterogeneous epigenomic landscape in cancer. We will focus mainly on DNA methylation and post-translational histone modifications as the key regulatory epigenetic signaling marks. In addition, we will describe how this knowledge is translated into the clinic. We will particularly concentrate on the applicability of epigenetic alterations as biomarkers for improved diagnosis, prognosis, and prediction. Finally, we will also review current developments regarding epi-drug usage in clinical and experimental settings., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Studies on carcinogenesis continue to provide new information about different disease-related processes. Among others, much research has focused on the involvement of cancer stem cells (CSCs) in tumor initiation and progression. Studying the similarities and differences between CSCs and physiological stem cells (SCs) allows for a better understanding of cancer biology. Recently, it was shown that stem cell identity is partially governed by the Krϋppel-associated box domain zinc finger proteins (KRAB-ZFPs), the biggest family of transcription regulators. Several KRAB-ZFP factors exert a known effect in tumor cells, acting as tumor suppressor genes (TSGs) or oncogenes, yet their role in CSCs is still poorly characterized. Here, we review recent studies regarding the influence of KRAB-ZFPs and their cofactor protein TRIM28 on CSCs phenotype, stemness features, migration and invasion potential, metastasis, and expression of parental markers., (© 2022 The Authors. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co., Ltd.)

In horses, the benzodiazepine diazepam (DIA) is used as sedative for pre-medication or as an anxiolytic to facilitate horse examinations. As the sedative effects can also be abused for doping purposes, DIA is prohibited in equine sports. DIA is extensively metabolized to several active metabolites such as nordazepam, temazepam and oxazepam (OXA). For veterinarians, taking into account the detection times of DIA and its active metabolites is needed for minimizing the risk of an anti-doping rule violation. Therefore, a pharmacokinetic study on 6 horses was conducted using a single intravenous (IV) dose of 0.2 mg/kg DIA Plasma and urine samples were collected at specified intervals until 16 and 26 days post-administration, respectively. Samples were analysed by a sensitive liquid chromatography-electrospray ionization/tandem mass spectrometry method. DIA showed a triphasic elimination pattern in the horse. The mean plasma clearance of DIA was 5.9 ml/min/kg, and the plasma elimination half-life in the terminal phase was 19.9 h. Applying the Toutain model approach, an effective plasma concentration of DIA was estimated at 24 ng/ml, and irrelevant plasma concentration (IPC) and irrelevant urine concentration (IUC) were computed to 0.047 and 0.1 ng/ml, respectively. The detection time according to the European Horserace Scientific Liaison Committee (EHSLC), that is the time for which observed DIA plasma concentrations of all investigated horses were below the IPC was 10 days. Using Monte Carlo Simulations, it was estimated that concentrations of DIA in plasma would fall below the IPC 18 days after the DIA administration for 90% of horses. However, in the present study, a single administration of DIA could be detected for 24 days in urine via the presence of OXA, its dominant metabolite., (© 2021 The Authors. Journal of Veterinary Pharmacology and Therapeutics published by John Wiley & Sons Ltd.)

To evaluate the antioxidant activity of potential synthetic enzyme mimetics, we prepared new five copper(II) complexes via a self-assembly method and named them [Cu(2-(HOCH 2 )py) 3 ](ClO 4 ) 2 ( 1 ), [Cu(2-(HOCH 2 )py) 2 (H 2 O) 2 ]SiF 6 ( 2 ), [Cu 2 (2-(HOCH 2 CH 2 )py) 2 (2-(OCH 2 CH 2 )py) 2 ](ClO 4 ) 2 ( 3 ), [Cu(pyBIm) 3 ](BF 4 ) 2 ·1.5H 2 O ( 4 ) and [Cu(py 2 C(OH) 2 ) 2 ](ClO 4 ) 2 ( 5 ). The synthetic protocol involved N,O- or N,N-donors: 2-(hydroxymethyl)pyridine (2-(HOCH 2 )py), 2-(hydroxyethyl)pyridine (2-(HOCH 2 CH 2 )py), 2-(2-pyridyl)benzimidazole (pyBIm), di(2-pyridyl)ketone (py 2 CO). The obtained Cu(II) complexes were fully characterised by elemental analysis, FTIR, EPR, UV-Vis, single-crystal X-ray diffraction and Hirshfeld surface analysis. Crystallographic and spectroscopic analyses confirmed chromophores of both monomeric ({CuN 3 O 3 } ( 1 ), {CuN 2 O 4 } ( 2 ), {CuN 6 } ( 4 ), {CuN 4 O 2 } ( 5 )) and dimeric complex ({CuN 2 O 3 } ( 3 )). Most of the obtained species possessed a distorted octahedral environment, except dimer 3 , which consisted of two copper centres with square pyramidal geometries. The water-soluble compounds ( 1 , 3 and 5 ) were selected for biological testing. The results of the study revealed that complex 1 in solutions displayed better radical scavenging activity than complexes 3 , 5 and free ligands. Therefore, complex 1 has been selected for further studies to test its activity as an enzyme mimetic. The chosen compound was tested on the erythrocyte lysate of two groups of patients after undergoing chemotherapy and chemoradiotherapy. The effect of the tested compound ( 1 ) on enzyme activity levels (TAS, SOD and CAT) suggests that the selected complex can be treated as a functional mimetic of the enzymes.

Telomerase is known to contribute to telomere maintenance and to provide cancer cell immortality. However, numerous reports are showing that the function of the enzyme goes far beyond chromosome ends. The study aimed to explore how telomerase downregulation in MCF7 and MDA-MB-231 breast cancer cells affects their ability to survive. Consequently, sensitivity to drug resistance, proliferation, and adhesion were assessed. The lentiviral-mediated human telomerase reverse transcriptase (hTERT) downregulation efficiency was performed at gene expression and protein level using qPCR and Western blot, respectively. Telomerase activity was evaluated using the Telomeric Repeat Amplification Protocol (TRAP) assay. The study revealed that hTERT downregulation led to an increased sensitivity of breast cancer cells to doxorubicin which was demonstrated in MTT and clonogenic assays. During a long-term doubling time assessment, a decreased population doubling level was observed. Interestingly, it did not dramatically affect cell cycle distribution. hTERT downregulation was accompanied by an alteration in β1-integrin- and by focal adhesion kinase (FAK)-driven pathways together with the reduction of target proteins phosphorylation, i.e., paxillin and c-Src. Additionally, autophagy activation was observed in MDA-MB-231 cells manifested by alternations in Atg5, Beclin 1, LC3II/I ratio, and p62. These results provide new evidence supporting the possible therapeutic potential of telomerase downregulation leading to induction of autophagy and cancer cells elimination.

Following the outbreak of severe acute respiratory syndrome coronavirus (SARS-CoV-2), airborne water droplets have been identified as the main transmission route. Identifying and breaking all viable transmission routes are critical to stop future outbreaks, and the potential of transmission by water has been highlighted. By modifying established approaches, we provide a method for the rapid assessment of the risk of transmission posed by fecally contaminated river water and give example results for 39 countries. The country relative risk of transmission posed by fecally contaminated river water is related to the environment and the populations' infection rate and water usage. On the basis of in vitro data and using temperature as the primary controller of survival, we then demonstrate how viral loads likely decrease after a spill. These methods using readily available data suggest that sewage spills into rivers within countries with high infection rates could provide infectious doses of >40 copies per 100 mL of water. The approach, implemented in the supplementary spreadsheet, can provide a fast estimate of the upper and lower viral load ranges following a riverine spill. The results enable evidence-based research recommendations for wastewater epidemiology and could be used to evaluate the significance of fecal-oral transmission within freshwater systems., Competing Interests: The authors declare no competing financial interest., (© 2021 The Authors. Published by American Chemical Society.)

Krüppel-associated box zinc finger proteins (KRAB-ZFPs) constitute the largest family of transcriptional factors exerting co-repressor functions in mammalian cells. In general, KRAB-ZFPs have a dual structure. They may bind to specific DNA sequences via zinc finger motifs and recruit a repressive complex through the KRAB domain. Such a complex mediates histone deacetylation, trimethylation of histone 3 at lysine 9 (H3K9me3), and subsequent heterochromatization. Nevertheless, apart from their repressive role, KRAB-ZFPs may also co-activate gene transcription, likely through interaction with other factors implicated in transcriptional control. KRAB-ZFPs play essential roles in various biological processes, including development, imprinting, retroelement silencing, and carcinogenesis. Cancer cells possess multiple genomic, epigenomic, and transcriptomic aberrations. A growing number of data indicates that the expression of many KRAB-ZFPs is altered in several tumor types, in which they may act as oncogenes or tumor suppressors. Hereby, we review the available literature describing the oncogenic and suppressive roles of various KRAB-ZFPs in cancer. We focused on their association with the clinicopathological features and treatment response, as well as their influence on the cancer cell phenotype. Moreover, we summarized the identified upstream and downstream molecular mechanisms that may govern the functioning of KRAB-ZFPs in a cancer setting.

Introduction: Dental caries is a transmissible chronic disease of dental hard tissues. To monitor the course of carious process, age reference groups were designated which are subject to regular dental check-ups. The groups include children aged 6, 12, and 18 years as well as adults at the age of 35-44 and 55-64 years., Aim: The aim of the study was to evaluate the state of dentition and effectiveness of treatment among children and adolescents in the reference groups aged 6, 12, and 18 years from the town of Skierniewice and its region in the years 2017-2020., Material and Methods: The study included 385 children and adolescents from the reference groups. Dental examination was carried out in the dental office of the Ogrodowa Medical Centre in Skierniewice. Dental caries intensity expressed by the the DMFT index, and its particular components, D, M, and F (D- the mean number of teeth with caries (decayed), M- teeth extracted (missing), and F- filled) as well as the dental caries treatment index DTI (F/(D+F)) were determined according to sex., Results: Distribution of the DMFT index components was very similar in children aged 6 years and in 18-year-old adolescents. The D component constituted half the DMF index (50.7% in 6-year-olds and 52.4% in 18-year-olds), the M component values were lowest (16.1% and 12.3%, respectively) while the F component accounted for slightly more than 1/3 of the whole value (33.2% and 35.3%, respectively). In 12-year-olds, the D component value was 68.4% of the DMF index, the M component accounted for only a few percent (4.5%) and the F component was 27.1%. The DTI value was 0.40 for 6-year-olds and 18-year-olds and appeared to be significantly higher than in the group of 12-year-olds (0.3)., Conclusions: There is an urgent need for monitoring the dental status, professional caries prevention, and conservative treatment of teeth in children and adolescents of the Skierniewice region., (© National Institute of Public Health – National Institute of Hygiene.)

Even though chemotherapy and immunotherapy emerged to limit continual and unregulated proliferation of cancer cells, currently available therapeutic agents are associated with high toxicity levels and low success rates. Additionally, ongoing multi-targeted therapies are limited only for few carcinogenesis pathways, due to continually emerging and evolving mutations of proto-oncogenes and tumor-suppressive genes. CRISPR/Cas9, as a specific gene-editing tool, is used to correct causative mutations with minimal toxicity, but is also employed as an adjuvant to immunotherapy to achieve a more robust immunological response. Some of the most critical limitations of the CRISPR/Cas9 technology include off-target mutations, resulting in nonspecific restrictions of DNA upstream of the Protospacer Adjacent Motifs (PAM), ethical agreements, and the lack of a scientific consensus aiming at risk evaluation. Currently, CRISPR/Cas9 is tested on animal models to enhance genome editing specificity and induce a stronger anti-tumor response. Moreover, ongoing clinical trials use the CRISPR/Cas9 system in immune cells to modify genomes in a target-specific manner. Recently, error-free in vitro systems have been engineered to overcome limitations of this gene-editing system. The aim of the article is to present the knowledge concerning the use of CRISPR Cas9 technique in targeting treatment-resistant cancers. Additionally, the use of CRISPR/Cas9 is aided as an emerging supplementation of immunotherapy, currently used in experimental oncology. Demonstrating further, applications and advances of the CRISPR/Cas9 technique are presented in animal models and human clinical trials. Concluding, an overview of the limitations of the gene-editing tool is proffered.

Highly dynamic epigenetic signaling is influenced mainly by (micro)environmental stimuli and genetic factors. The exact mechanisms affecting particular epigenomic patterns differ dependently on the context. In the current review, we focus on the causes and effects of the dynamic signatures of the human epigenome as evaluated with the high-throughput profiling data and single-gene approaches. We will discuss three different aspects of phenotypic outcomes occurring as a consequence of epigenetics interplaying with genotype and environment. The first issue is related to the cases of environmental impacts on epigenetic profile, and its adverse and advantageous effects related to human health and evolutionary adaptation. The next topic will present a model of the interwoven co-evolution of genetic and epigenetic patterns exemplified with transposable elements (TEs) and their epigenetic repressors Krüppel-associated box zinc finger proteins (KRAB-ZNFs). The third aspect concentrates on the mitosis-based microevolution that takes place during carcinogenesis, leading to clonal diversity and expansion of tumor cells. The whole picture of epigenome plasticity and its role in distinct biological processes is still incomplete. However, accumulating data define epigenomic dynamics as an essential co-factor driving adaptation at the cellular and inter-species levels with a benefit or disadvantage to the host.

In equine and racing practice, detomidine and butorphanol are commonly used in combination for their sedative properties. The aim of the study was to produce detection times to better inform European veterinary surgeons, so that both drugs can be used appropriately under regulatory rules. Three independent groups of 7, 8 and 6 horses, respectively, were given either a single intravenous administration of butorphanol (100 µg/kg), a single intravenous administration of detomidine (10 µg/kg) or a combination of both at 25 (butorphanol) and 10 (detomidine) µg/kg. Plasma and urine concentrations of butorphanol, detomidine and 3-hydroxydetomidine at predetermined time points were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The intravenous pharmacokinetics of butorphanol dosed individually compared with co-administration with detomidine had approximately a twofold larger clearance (646 ± 137 vs. 380 ± 86 ml hr -1  kg -1 ) but similar terminal half-life (5.21 ± 1.56 vs. 5.43 ± 0.44 hr). Pseudo-steady-state urine to plasma butorphanol concentration ratios were 730 and 560, respectively. The intravenous pharmacokinetics of detomidine dosed as a single administration compared with co-administration with butorphanol had similar clearance (3,278 ± 1,412 vs. 2,519 ± 630 ml hr -1  kg -1 ) but a slightly shorter terminal half-life (0.57 ± 0.06 vs. 0.70 ± 0.11 hr). Pseudo-steady-state urine to plasma detomidine concentration ratios are 4 and 8, respectively. The 3-hydroxy metabolite of detomidine was detected for at least 35 hr in urine from both the single and co-administrations. Detection times of 72 and 48 hr are recommended for the control of butorphanol and detomidine, respectively, in horseracing and equestrian competitions., (© 2020 John Wiley & Sons Ltd.)

Summary: Krüppel-associated box domain zinc finger protein (KRAB ZNF) explorer is a web-based tool for comprehensive characterization of the mRNA expression status of KRAB-ZNF transcription factors in the data from The Cancer Genome Atlas study. Key functionalities cover a comparative analysis of KRAB-ZNF expression between normal and cancer tissues, an association of KRAB-ZNF expression with various pathological features, including survival analysis, association with global DNA methylation status, analyses of KRAB-ZNF isoform expressions and analysis of KRAB-ZNF levels in normal tissues., Availability and Implementation: KRAB ZNF explorer is available at http://mi2.mini.pw.edu.pl: 8080/KRAB&#95;ZNF/. The source code for shiny application is available at: https://github.com/MI2DataLab/KRAB&#95;ZNF/tree/master/app and the source code for analyses and precalculations are available at: https://github.com/MI2DataLab/KRAB&#95;ZNF/tree/master/work., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Health promoting behaviours have a decisive effect on the development and cours of caries at any age., Objectives: The aim of the study was to investigate the correlation between health promoting behaviours and the state of dentition in children aged 1-3 years by determining the child's age, the cause of the first visit at the dental office and evaluating the relationship between the intensity of dental caries and the frequency of oral hygiene practices as well as the consumption of sweet foods., Material and Methods: The study was conducted among 204 children aged 1-3 years from the town of Skierniewice and its vicinity, who visited the Dental Clinic at the Ogrodowa Medical Centre in Skierniewice in the years 2019 and 2020. The study consisted of two parts: a questionnaire and clinical examination., Results: In the entire study group, active caries was found in 65.2% of children. Most of the children came for the first time to the dentist at the age of 3 years (37.5%), and the least number of them in the first year of age (22.6%). There was no significant correlation observed between the purpose of the visit (adaptation, check-up and treatment, pain) and the child's age (p> 0.05). The majority of children (67.6%) cleaned their teeth twice a day; and the value of their dmf index (2.00 ± 0.88) was significantly lower compared to those brushing teeth once a day: in the morning (2.82 ± 1.01) or in the evening (2.89 ± 0.93) (p <0.001). Only 8.8% of children did not eat sweets at all, while more than half (58.8%) consumed them more than once a day. The dmf index value significantly (p<0.001) increased with the frequency of sweets consumption from 0.37 ± 0.08 - never, through 1.73 ± 0.86 - once a day, to 2.99 ± 1.03 - often., Conclusions: The results of our study showed that health-promoting measures have a significant impact on the dental state of children aged 1-3 years., (© National Institute of Public Health – National Institute of Hygiene.)

The KRAB-ZNF (Krüppel-associated box domain zinc finger) gene family is composed of a large number of highly homologous genes, gene isoforms, and pseudogenes. The proteins encoded by these genes, whose expression is often tissue-specific, act as epigenetic suppressors contributing to the addition of repressive chromatin marks and DNA methylation. Due to its high complexity, the KRAB-ZNF family has not been studied in sufficient detail, and the involvement of its members in carcinogenesis remains mostly unexplored. In this study, we aimed to provide a comprehensive description of cancer-associated KRAB-ZNFs using publicly available The Cancer Genome Atlas pan-cancer datasets. We analyzed 6727 tumor and normal tissue samples from 16 cancer types. Here, we showed that a small but distinctive cluster of 16 KRAB-ZNFs is commonly upregulated across multiple cancer cohorts in comparison to normal samples. We confirmed these observations in the independent panels of lung and breast cancer cell lines and tissues. This upregulation was also observed for most of the KRAB-ZNF splicing variants, whose expression is simultaneously upregulated in tumors compared to normal tissues. Finally, by analyzing the clinicopathological data for breast and lung cancers, we demonstrated that the expression of cancer-associated KRAB-ZNFs correlates with patient survival, tumor histology, and molecular subtyping. Altogether, our study allowed the identification and characterization of KRAB-ZNF factors that may have an essential function in cancer biology and thus potential to become novel oncologic biomarkers and treatment targets., (© 2018 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.)

Aim: The aim of the study was to evaluate the results of the preventive and therapeutic programme addressed to adolescents aged 17 and 18, based on a comparison of the state of dentition prior to and after the completion of the programme., Material and Methods: All 44 adolescents from the City of Skierniewice and the surrounding area who applied for enrolment were included in the programme. Based on clinical examination, the prevalence of active caries, caries intensity expressed by mean values of the DMFT (decayed, missing, filled teeth) Index including D, M, and F components and the Dental Treatment (DT) Index F/(D + F) were assessed. All parameters were presented according to sex., Results: Active caries was observed in 34 (77.3%) patients. Caries was more frequently detected in boys (82.6%) than in girls (71.4%). The value of the Caries Intensity Index, DMF, equalled 12.05 and was significantly higher (p<0.01) in boys than in girls: 13.73 ± 3.63 and 10.06 ± 3.42. In the examined group of 17- and 18-year-olds, the mean number of decayed teeth D (6.31) had the highest mean value, followed by the mean number of filled teeth F (4.26), the mean number of extracted (missing) teeth M (1.48) being the lowest. The values of the F Index were statistically significantly higher (t=2.570; p=0.0195) in boys as compared to girls (5.26 ± 2.69 vs. 3.13 ± 2.83). Mean value the Dental Treatment (DT) Index (F/D+F) appeared to be higher in the group of boys than in girls: 0.431 ± 0.182 and 0.358 ± 0.126, respectively. After the completion of the programme, all adolescents studied achieved the value of the Dental Treatment Index equalling 1, Conclusions: Dental examination qualifying 17- and 18-year old adolescents in Skierniewice to the preventive and therapeutic programme detected numerous foci of active caries. Implementation of the programme allowed to effectively treat the teeth of the adolescents participating in the project and to develop proper hygiene and dietary habits., (© National Institute of Public Health – National Institute of Hygiene)

Methylxanthines positives in competition samples have challenged doping control laboratories and racing jurisdictions since methylxanthines are naturally occurring prohibited substances and often constituents of feed. For theobromine, an international threshold (renamed in International Residue Limit, IRL) of 2 µg/mL in urine has been established. On the basis of the data presented herein, a threshold or rather an IRL for theobromine in plasma of 0.3 µg/mL was proposed and was thereupon approved by the International Federation of Horseracing Authorities (IFHA). Official recommendations for reporting caffeine and theophylline are still lacking. The aim of the study was to investigate IRLs for theobromine in blood and for caffeine and theophylline in blood and urine. Therefore, a set of six administrations were carried out including both single i.v. and single oral administrations of caffeine, theobromine and theophylline. Plasma and urine concentrations were determined using a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS). Applying the Toutain model approach an effective plasma concentration (EPC) of caffeine was estimated at 3.05 µg/mL, irrelevant concentrations in blood (IPC) and urine (IUC) approached 6 and 12 ng/mL, respectively. EPC of theobromine was calculated with 3.80 µg/mL, and irrelevant concentrations of theobromine were determined at 8 ng/mL in plasma and at 142 ng/mL in urine. Toutain modelling of the theophylline data produced an EPC, IPC, and IUC of 3.20 µg/mL, 6 ng/mL, and 75 ng/mL, respectively. The obtained irrelevant concentrations were used to postulate IRLs for theobromine in plasma and for caffeine and theophylline in plasma and urine. Copyright © 2016 John Wiley & Sons, Ltd., (Copyright © 2016 John Wiley & Sons, Ltd.)

OBJECTIVE To evaluate the influence of respiratory tract disease (ie, recurrent airway obstruction [RAO]) and mode of inhalation on detectability of inhaled budesonide in equine plasma and urine samples. ANIMALS 16 horses (8 healthy control horses and 8 horses affected by RAO, as determined by results of clinical examination, blood gas analysis, bronchoscopy, and cytologic examination of bronchoalveolar lavage fluid). PROCEDURES 4 horses of each group inhaled budesonide (3 μg/kg) twice daily for 10 days while at rest, and the remaining 4 horses of each group inhaled budesonide during lunging exercise. Plasma and urine samples were obtained 4 to 96 hours after inhalation and evaluated for budesonide and, in urine samples, the metabolites 6β-hydroxybudesonide and 16α-hydroxyprednisolone. RESULTS Detected concentrations of budesonide were significantly higher at all time points for RAO-affected horses, compared with concentrations for the control horses. All samples of RAO-affected horses contained budesonide concentrations above the limit of detection at 96 hours after inhalation, whereas this was found for only 2 control horses. Detected concentrations of budesonide were higher, but not significantly so, at all time points in horses that inhaled budesonide during exercise, compared with concentrations for inhalation at rest. CONCLUSIONS AND CLINICAL RELEVANCE Results of this study indicated that the time interval between inhalation of a glucocorticoid and participation in sporting events should be increased when inhalation treatment is administered during exercise to horses affected by respiratory tract disease.

Background: Although acetylsalicylic acid (ASA) is not frequently used as a therapeutic agent in horses, its metabolite SA is of special interest in equestrianism since it is a natural component of many plants used as horse feed. This led to the establishment of thresholds by horse sport organizations for SA in urine and plasma. The aim of this study was to investigate plasma and urine concentrations of salicylic acid (SA) after oral administration of three different single dosages (12.5 mg/kg, 25 mg/kg and 50 mg/kg) of acetylsalicylic acid (ASA) to eight horses in a cross-over designed study., Results: In the 12.5 mg/kg group, SA concentrations in urine peaked 2 h after oral administration (2675 μg/mL); plasma concentrations peaked at 1.5 h (17 μg/mL). In the 25 mg/kg group, maximum concentrations were detected after 2 h (urine, 2785 μg/mL) and 1.5 h (plasma, 23 μg/mL). In the 50 mg/kg group, maximum concentrations were observed after 5 h (urine, 3915 μg/mL) and 1.5 h (plasma, 45 μg/mL). The plasma half-life calculated for SA varied between 5.0 and 5.7 h. The urine concentration of SA fell below the threshold of 750 μg/mL (set by the International Equestrian Federation FEI and most of the horseracing authorities) between 7 and 26 h after administration of 12.5 and 25 mg/kg ASA and between 24 and 36 h after administration of 50 mg/kg ASA. For ASA, IC 50 were 0.50 μg/mL (COX-1) and 5.14 μg/mL (COX-2). For salicylic acid, it was not possible to calculate an IC 50 for either COX due to insufficient inhibition of both cyclooxygenases., Conclusion: The established SA thresholds of 750 μg//mL urine and 6.5 μg/mL plasma appear too generous and are leaving space for misuse of the anti-inflammatory and analgetic compound ASA in horses.