Prolonged Remission Induced by FENofibrate in children with newly diagnosed type 1 diabetes (PRIFEN): protocol of a randomised controlled trial.

INTRODUCTION: Sphingolipids regulate proinsulin folding, insulin secretion and control beta cells apoptosis. Recent evidence has demonstrated that, among other factors, reduced amounts of sulfatide may be relevant in the development of type 1 diabetes (T1D). Thus, fenofibrate, which activates sulfatide biosynthesis, may prolong remission in subjects with T1D. The aim of the study is to evaluate clinical efficacy of fenofibrate on the maintenance of residual beta-cell function in children with newly diagnosed T1D. METHODS AND ANALYSIS: A total of 102 children aged 10-17 years with newly diagnosed T1D will be enrolled in a double-blind, two-centre randomised, non-commercial, placebo-controlled trial. Subjects who will meet all inclusion criteria will be randomly assigned to receive fenofibrate at a dose of 160 mg or an identically appearing placebo, orally, once daily, for 12 months. The primary endpoint will be the area under the curve of the C-peptide level during 2-hour responses to a mixed-meal tolerance test (MMTT). Secondary endpoints include fasting and maximum C-peptide concentration in the MMTT, parameters of diabetes control and glucose fluctuations, daily insulin requirement, inflammation markers, genetic analysis, safety and tolerance of the fenofibrate ETHICS AND DISSEMINATION: The study protocol was approved by the Bioethics Committee. The results of this study will be submitted to a peer-reviewed diabetic journal. Abstracts will be submitted to international and national conferences. TRIAL REGISTRATION NUMBER: EnduraCT 2020-003916-28.

Central Nervous System Lymphoma: The Great Mimicker-A Single-Institution Retrospective Study.

Background: Primary central nervous system lymphoma (PCNSL) is a rare, aggressive form of non-Hodgkin lymphoma contained in the brain and the spinal cord as well as the meninges, cranial nerves, eyes, and cerebrospinal fluid (CSF). Due to its variable presentation and lack of associated B-symptoms, it is quite challenging to diagnose PCNSL, if there is not a high level of suspicion. Methods: This is a retrospective case series examining 13 human immunodeficiency virus- (HIV-) negative patients with PCNSL and DLBCL type, with a median age of 75 years old. Results: The most common presenting symptom was altered mental status. The frontal lobes, basal ganglia, cerebellum, and corpus callosum were most affected. Prior to brain biopsy, 4/13 patients were on steroids, which did not affect biopsy results and the average time to diagnosis was 1 month. 9/13 patients who did not receive steroids had an average time to diagnosis of less than 1 month. Conclusion: Although steroid administration did not appear to diminish the yield of the biopsy, it is a best practice to withhold steroids prior to biopsy to decrease the time to diagnose PCNSL.

Suspicion of Recurrent Osteosarcoma in a Patient with Pseudotumour Adjacent To the Endoprosthesis.

Purpose: Pseudotumor is a rare complication after arthroplasty, most often of the hip joint, in response to metal particles present in the implant. There are merely sporadic reports of pseudotumor in patients with bone sarcoma after sparing surgery with endoprosthesis implant. The aim of this study is to present the characteristic imaging features of pseudotumor. Case report: We present a case of a 21-year-old male patient in whom a scheduled follow-up ultrasound revealed a painless lesion suspected of local recurrence at the border of the endoprosthesis and the bone stump 3.5 years after the end of treatment for osteosarcoma of the femur. Histopathology of the biopsy specimen revealed that the lesion was a pseudotumor. Conclusions: Although pseudotumor is sporadic in patients treated with endoprosthesis for bone sarcoma, their prolonged survival could bear the risk of such a complication. Imaging studies, in particular ultrasound, may be helpful in differentiating from local recurrence of sarcoma, however, the histopathology of the specimen obtained by open biopsy at a reference center is crucial for the final diagnosis.

Secretome of endothelial progenitor cells from stroke patients promotes endothelial barrier tightness and protects against hypoxia-induced vascular leakage.

BACKGROUND: Cell-based therapeutic strategies have been proposed as an alternative for brain repair after stroke, but their clinical application has been hampered by potential adverse effects in the long term. The present study was designed to test the effect of the secretome of endothelial progenitor cells (EPCs) from stroke patients (scCM) on in vitro human models of angiogenesis and vascular barrier. METHODS: Two different scCM batches were analysed by mass spectrometry and a proteome profiler. Human primary CD34+-derived endothelial cells (CD34+-ECs) were used for designing angiogenesis studies (proliferation, migration, and tubulogenesis) or in vitro models of EC monolayer (confluent monolayer ECs-CMECs) and blood-brain barrier (BBB; brain-like ECs-BLECs). Cells were treated with scCM (5 µg/mL) or protein-free endothelial basal medium (scEBM-control). CMECs or BLECs were exposed (6 h) to oxygen-glucose deprivation (OGD) conditions (1% oxygen and glucose-free medium) or normoxia (control-5% oxygen, 1 g/L of glucose) and treated with scCM or scEBM during reoxygenation (24 h). RESULTS: The analysis of different scCM batches showed a good reproducibility in terms of protein yield and composition. scCM increased CD34+-EC proliferation, tubulogenesis, and migration compared to the control (scEBM). The proteomic analysis of scCM revealed the presence of growth factors and molecules modulating cell metabolism and inflammatory pathways. Further, scCM decreased the permeability of CMECs and upregulated the expression of the junctional proteins such as occludin, VE-cadherin, and ZO-1. Such effects were possibly mediated through the activation of the interferon pathway and a moderate downregulation of Wnt signalling. Furthermore, OGD increased the permeability of both CMECs and BLECs, while scCM prevented the OGD-induced vascular leakage in both models. These effects were possibly mediated through the upregulation of junctional proteins and the regulation of MAPK/VEGFR2 activity. CONCLUSION: Our results suggest that scCM promotes angiogenesis and the maturation of newly formed vessels while restoring the BBB function in ischemic conditions. In conclusion, our results highlight the possibility of using EPC-secretome as a therapeutic alternative to promote brain angiogenesis and protect from ischemia-induced vascular leakage.

Increased frequency of severe diabetic ketoacidosis at type 1 diabetes onset among children during COVID-19 pandemic lockdown: an observational cohort study.

INTRODUCTION: On March 11, 2020 the WHO announced a coronavirus disease 2019 (COVID-19) pandemic. Lockdown restrictions, compromised access to medical care and fear of potential exposure to SARS-CoV-2 have forced patients with non-COVID-19 illnesses such as type 1 diabetes (T1D) to stay home. This situation can lead to delay in T1D diagnosis and insulin treatment resulting in rapid progression to diabetic ketoacidosis (DKA) and therefore increased risk of complications and death.  . AIM: The aim of this study was to evaluate the frequency and severity of DKA at the onset of T1D in children diagnosed in our department during COVID-19 pandemic lockdown from March 2020 till May 2020 in comparison to corresponding period of the previous year. . MATERIAL AND METHODS: We collected data of children with newly diagnosed T1D. DKA was defined according to ISPAD guidelines. . RESULTS: The study cohort comprised 34 children in group 2020 and 52 in group 2019 with an average age 9.90 ±4.9 vs. 9.59±4.7 years with mean HbA1c 12.9 ±2.4 vs. 11.5 ±2.2%, respectively. The incidence of DKA was higher by 12% in group 2020 vs. 2019 (52.94% vs 40.38%; p = 0.276).  Regarding the DKA severity (2020 vs. 2019) 32.35% vs. 11.54% were severe (p = 0.026), 17.65 vs. 13% were moderate (p = 0.759), and 2.94 vs. 15.38% were mild (p = 0.081). None of the analyzed patients were COVID-19 positive. CONCLUSIONS: During the COVID-19 pandemic lockdown changes in society and health care system, the DKA rate has increased by 12 percentage points with more severe cases noted in children with newly diagnosed T1D. Regular education of the whole society about the symptoms of diabetes could contribute to faster diagnosis of T1D and reduction of DKA prevalence. .

Progesterone impairs Herceptin effect on breast cancer cells.

Breast cancer (BCa) is the most common cancer affecting women worldwide. Overexpression of human epidermal growth factor receptor 2 (HER2) occurs in ~20-25% of invasive ductal breast carcinomas and is associated with the more aggressive phenotype. Herceptin, a humanized antibody against HER2, is a standard therapy in HER2-overexpressing cases. Approximately one-third of patients relapse despite treatment. Therefore numerous studies have investigated the molecular mechanisms associated with Herceptin resistance. An interaction between HER2 signalling and steroid hormone receptor signalling pathways has been previously investigated, but the effect of this relationship on Herceptin resistance has never been studied. The present study analysed an impact of the steroid hormone, progesterone (PG), on Herceptin-dependent cell growth inhibition. Results indicated that Herceptin-inhibited proliferation of breast cancer cell lines overexpressing HER2 (BT474 and MCF/HER2) in 3D culture is abolished by PG. Furthermore, results demonstrated that PG led to the activation of HER2/HER3-mediated signalling. Moreover, PG treatment induced a shift of Herceptin-dependent cell cycle arrest in G1 phase towards S and G2 phases with concomitant upregulation of cyclin-dependent kinase 2 (CDK2) and downregulation of CDK inhibitor p27Kip1. These results demonstrate for the first time PG involvement in the failure of Herceptin treatment in vitro. The present observations suggest that cross-talk between PG- and HRG/HER2-initiated signalling pathways may lead to the acquisition of resistance to Herceptin in patients with BCa.

Impact of ELKa, the Electronic Device for Prandial Insulin Dose Calculation, on Metabolic Control in Children and Adolescents with Type 1 Diabetes Mellitus: A Randomized Controlled Trial.

Background. The ELKa system is composed of computer software, with a database of nutrients, and a dedicated USB kitchen scale. It was designed to automatize the everyday calculations of food exchanges and prandial insulin doses. Aim. To investigate the influence of the ELKa on metabolic control in children with type 1 diabetes mellitus (T1DM). Methods. A randomized, parallel, open-label clinical trial involved 106 patients aged <18 years with T1DM, HbA1C ≤ 10%, undergoing intensive insulin therapy, allocated to the intervention group, who used the ELKa (n = 53), or the control group (n = 53), who used conventional calculation methods. Results. After the 26-week follow-up, the intention-to-treat analysis showed no differences to all endpoints. In per protocol analysis, 22/53 (41.5%) patients reporting ELKa usage for >50% of meals achieved lower HbA1C levels (P = 0.002), lower basal insulin amounts (P = 0.049), and lower intrasubject standard deviation of blood glucose levels (P = 0.023) in comparison with the control. Moreover, in the intervention group, significant reduction of HbA1C level, by 0.55% point (P = 0.002), was noted. No intergroup differences were found in the hypoglycemic episodes, BMI-SDS, bolus insulin dosage, and total daily insulin dosage. Conclusions. The ELKa system improves metabolic control in children with T1DM under regular usage. The trial is registered at ClinicalTrials.gov, number NCT02194517.

Analysis of the doctor-patient communication if breast cancer is diagnosed / Análisis de la comunicación médico-paciente en el diagnóstico de cáncer de mama / Análise da comunicação médico-paciente em caso de diagnóstico de câncer de mama

The aim of the paper is the recognition and evaluation of demand for medical information among patients suffering from breast cancer. The research was conducted among 120 women with diagnosed breast cancer in the Oncological Surgery Clinic of the Independent Public Research Hospital Nº 1 in Lublin, the Oncological Clinic of the Independent Public Research Hospital Nº 1 in Lublin and the Rehabilitation Centre with the Rehabilitation Clinic at the Lublin Oncology Centre. The research included women from the Club of Women after Mastectomy “Amazons”, the Club “Amazons” at the Complex of Specialist Clinics in Ostrowiec Swietokrzyski and the Club “Amazons” in Radom. Research showed that the demand for medical information among women with diagnosed breast cancer is very high. Respondents want to know all the information concerning the diagnosis, treatment and prognosis (93 percent). They also expect that the doctor will give them medical information concerning further consequences of cancer and its influence on future health and life (78 percent). Most of the respondents expect to receive information concerning medicaments which should be taken (77 percent) and the essence of the conducted treatment (93 percent). The research analysis showed that women with diagnosed breast cancer expect that the doctor will give them reliable and real medical information. Patients want the doctor to present them the probable course of the cancer (85 percent) and all the possible side effects connected with it (89 percent).

El objetivo de este trabajo es el reconocimiento y la evaluación de la demanda de información médica en pacientes con cáncer de mama. Se realizó la investigación con 120 mujeres diagnosticadas con cáncer de mama en la Clínica de Cirugía Oncológica y la Clínica Oncológica del Hospital de Investigación Público Independiente Nº 1 en Lublin, y el Centro de Rehabilitación con la Clínica de Rehabilitación del Centro Oncológico de Lublin. La investigación incluyó mujeres del Club “Amazons” de Mujeres que han sufrido Mastectomía, el Club “Amazons” del Complejo de Clínicas Especializadas en Ostrowiec Swietokrzyski y el Club “Amazons” en Radom. La investigación mostró que la demanda de información médica de mujeres con diagnóstico de cáncer de mama es muy alta. Aquellas que respondieron quieren saber todo acerca del diagnóstico, tratamiento y pronóstico (93 por ciento). También esperan que el médico les dé información médica respecto de posteriores consecuencias del cáncer y su influencia en su salud futura y su vida (78 por ciento). La mayoría de las que respondieron esperan recibir información sobre medicamentos que deberían tomar (77 por ciento) y lo esencial sobre el tratamiento realizado (93 por ciento). El análisis de la investigación muestra que las mujeres diagnosticadas con cáncer de mama esperan que el médico les dé información médica confiable y verdadera, les presente el curso probable del cáncer (85 por ciento) y todos los posibles efectos secundarios conectados (89 por ciento).

O objetivo deste artigo é o reconhecimento e a avaliação da demanda por informação médica entre pacientes que sofrem de câncer de mama. A investigação foi conduzida entre 120 mulheres diagnosticadas com câncer de mama na Oncological Surgery Clinic of the Independent Public Research Hospital Nº 1 de Lublin, a Oncological Clinic of the Independent Public Research Hospital Nº 1 de Lublin e a Rehabilitation Centre with the Rehabilitation Clinic do Lublin Oncology Centre. A pesquisa incluiu mulheres do Club of Women after Mastectomy “Amazons”, o Club “Amazons” do Complex of Specialist Clinic sem Ostrowiec Swietokrzyskie o Club “Amazons”em Radom. Pesquisa revelou que a demanda por informação médica entre mulheres com diagnóstico de câncer mamário é muito alta. As respondentes queriam saber todas as informações concernentes ao diagnóstico, tratamento e prognóstico (93 por cento). Elas também esperavam que o médico pudesse dar-lhes informação sobre consequências tardias do câncer e a influência sobre a sua saúde e vida futuras (78 por cento). A maioria dos respondentes tinham a expectativa de receber informação sobre medicamentos que deveriam tomar(77 por cento) e a essência do tratamento realizado(93 por cento). A análise da pesquisa demonstrou que as mulheres com diagnóstico de câncer de mama esperavam que o médico pudesse fornecer-lhes informação confiável e honesta. Pacientes queriam que o médico lhes apresentasse o provável curso do câncer (85 por cento) e todos os possíveis efeitos relacionados a ele (89 por cento).

Gender differences in exercise--induced intravascular haemolysis during race training in thoroughbred horses.

Exercise-induced intravascular haemolysis and "sport anemia" are widely reported in human sports medicine. It has been recognized also in horses, however, the clinical importance and the onset of this condition seem different than in human. In this study we investigated the episodes of intravascular haemolysis, indicated by the increase in plasma haemoglobin and the decrease in serum haptoglobin levels, after routine training sessions in race horses. Heart rate and changes in haematological parameters confirmed, that the exertion was relatively high. Intravascular haemolysis did not appear in stallions but was detected in mares after two training sessions. It has been determined that serum haptoglobin levels were higher in mares than in stallions before and after all training sessions. It is postulated that intravascular haemolysis induced by training is of limited clinical importance because it occurred only in mares which are better adapted due to higher haptoglobin level at rest, and it had no cumulative effect. Therefore gender differences should be taken into consideration in experiments with athletic horses.

A study of the intermolecular interactions of tolmetin/N-acetyl-L-tyrosine ethyl ester complex.

The formation of tolmetin/N-acetyl-l-tyrosine ethyl ester (ATEE) complex has been reported by means of both theoretical and experimental studies, including quantum mechanical calculations as well as UV-vis absorption, fluorescence and time-resolved spectroscopy measurements. It has been found that the fluorescence of ATEE is quenched due to the formation of a non-fluorescent complex between ATEE and tolmetin in the ground state. The geometrical parameters of ATEE/tolmetin complex have been determined with the use of the DFT method applying the B3LYP correlation-exchange functional and 6-31G(d) basis set. The results of experiments indicated the static ATEE quenching by tolmetin. Additionally, the experimental and theoretically predicted Gibbs free energy of complexation has been calculated.

Protoporphyrin IX induces apoptosis in HeLa cells prior to photodynamic treatment.

Photodynamic therapy (PDT) combining treatment with a light-excited compound and laser light induction, via cellular ROS generation, kills cancer cells by damaging organelles and impairing metabolic pathways. As the exact mechanisms underlying cancer cell death due to PDT treatment remain controversial, the influence of photosensitizer itself, protoporphyrin IX (PpIX) on cancer cells was investigated. The concentration-dependent viability of HeLa cells was estimated after PpIX-treatment. Microscopic analyses revealed that treated cells exhibited apoptosis-like morphology: blebbing, chromatin condensation, nuclear fragmentation, asymmetry of cellular membrane. These results shed a new light on cancer cell death due to PDT because they showed that PpIX can induce apoptosis without light excitation.

Novel DNA-binding element within the C-terminal extension of the nuclear receptor DNA-binding domain.

The heterodimer of the ecdysone receptor (EcR) and ultraspiracle (Usp), members of the nuclear receptors superfamily, is considered as the functional receptor for ecdysteroids initiating molting and metamorphosis in insects. Here we report the 1.95 A structure of the complex formed by the DNA-binding domains (DBDs) the EcR and the Usp, bound to the natural pseudopalindromic response element. Comparison of the structure with that obtained previously, using an idealized response element, shows how the EcRDBD, which has been previously reported to possess extraordinary flexibility, accommodates DNA-induced structural changes. Part of the C-terminal extension (CTE) of the EcRDBD folds into an alpha-helix whose location in the minor groove does not match any of the locations previously observed for nuclear receptors. Mutational analyses suggest that the alpha-helix is a component of EcR-box, a novel element indispensable for DNA-binding and located within the nuclear receptor CTE. This element seems to be a general feature of all known EcRs.

Tumor suppressor Fhit protein interacts with protoporphyrin IX in vitro and enhances the response of HeLa cells to photodynamic therapy.

Fhit, the product of tumor suppressor fragile histidine triad (FHIT) gene, exhibits antitumor activity of still largely unknown cellular background. However, it is believed that Fhit-Ap(3)A or Fhit-AMP complex might act as a second class messenger in cellular signal transduction pathway involved in cell proliferation and apoptosis. We demonstrate here for the first time that the photosensitizer, protoporphyrin IX (which is a natural precursor of heme) binds to Fhit protein and its mutants in the active site in vitro. Furthermore, PpIX inhibits the enzymatic activity of Fhit. Simultaneously, PpIX shows lower binding capacity to mutant Fhit-H96N of highly reduced hydrolase activity. In cell-based assay PpIX induced HeLa cell death in Fhit and Fhit-H96N-dependent manner which was measured by means of MTT assay. Moreover, HeLa cells stably expressing Fhit or mutant Fhit-H96N were more susceptible to protoporphyrin IX-mediated photodynamic therapy (2J/cm(2)) than parental cells.

EcR and Usp, components of the ecdysteroid nuclear receptor complex, exhibit differential distribution of molecular determinants directing subcellular trafficking.

Ecdysteroids coordinate development, reproduction and other essential biological processes in insects and other arthropods through the receptor which is a heterodimer of two members of the nuclear receptors superfamily, the ecdysteroid receptor (EcR) and the Ultraspiracle (Usp). Although the transcriptionally active EcR/Usp heterocomplex is believed to be the only functional form of the receptor, there are data indicating that EcR may be involved in the mediation of the non-genomic effects outside of the nucleus. Since the nucleocytoplasmic shuttling could be a key element determining participation of the single nuclear receptor molecule both in the genomic and non-genomic functions we have analyzed nuclear import and export properties of the EcR and Usp from Drosophila melanogaster. We show for the first time that both receptors exhibit differential distribution of the nuclear localization and nuclear export signals (NLSs and NESs). In particular, the Usp which exhibits exclusively nuclear localization in all cell types analyzed, contains apparently only NLS activity within the DNA-binding domain. In contrast, the three known EcR isoforms (A, B1 and B2) are mosaics of elements which can potentially mediate their nucleocytoplasmic shuttling. We have found two active NESs in ligand binding domain and NLS activity within the DNA-binding domain of all isoforms. Simultaneously we demonstrate that B1 and A isoforms possess an additional NLS activity localized in AB regions. We speculate that this characteristic, along with the previously reported structural pliability of the EcR molecule, allows the single receptor to evoke many different genomic as well as non-genomic ecdysteroid-dependent responses.

The DNA-binding domain of the ultraspiracle drives deformation of the response element whereas the DNA-binding domain of the ecdysone receptor is responsible for a slight additional change of the preformed structure.

Ecdysteroids control molting and metamorphosis in insects via a heterodimeric complex of two nuclear receptors, the ecdysone receptor (EcR) and ultraspiracle protein (Usp). We used fluorescence resonance energy transfer (FRET) to study the topology of the natural pseudopalindromic element from the hsp27 gene (hsp27pal) in complex with the DNA-binding domains of Usp and EcR (UspDBD and EcRDBD, respectively). Steady-state data revealed shortening of the end-to-end distance of the hsp27pal-derived probe. For the 70.8 +/- 0.6 A distance obtained for the UspDBD-complexed DNA a bend of about 23.1 +/- 2.9 degrees was measured. Nearly the same value (23.0 +/- 3.4 degrees) was obtained for the DNA complexed with the UspDBD/EcRDBD heterodimer. The respective bend angles estimated using fluorescence decay measurements were 19.0 +/- 2.1 degrees and 20.9 +/- 3.6 degrees . Thus, the FRET data suggest for the first time that the UspDBD defines the architecture of the UspDBD/EcRDBD heterocomplex due to the significant deformation of the hsp27pal. This suggestion has been further reinforced using gel retardation experiments, which, in conjunction with high-resolution DNase I footprinting, indicate that the main contribution to the observed bend is given by the UspDBD itself, while binding of the EcRDBD molecule brings on a slight additional change of the preformed structure.

Plasticity of the ecdysone receptor DNA binding domain.

Ecdysteroids coordinate molting and metamorphosis in insects via a heterodimer of two nuclear receptors, the ecdysone receptor (EcR) and the ultraspiracle (Usp) protein. Here we show how the DNA-recognition alpha-helix and the T box region of the EcR DNA-binding domain (EcRDBD) contribute to the specific interaction with the natural response element and to the stabilization of the EcRDBD molecule. The data indicate a remarkable mutational tolerance with respect to the DNA-binding function of the EcRDBD. This is particularly manifested in the heterocomplexes formed between the EcRDBD mutants and the wild-type Usp DNA-binding domain (UspDBD). Circular dichroism (CD) spectra and protein unfolding experiments indicate that, in contrast to the UspDBD, the EcRDBD is characterized by a lower alpha-helix content and a lower stability. As such, the EcRDBD appears to be an intrinsically unstructured protein-like molecule with a high degree of intramolecular plasticity. Because recently published crystal structures indicate that the ligand binding domain of the EcR is also characterized by the extreme adaptability, we suggest that plasticity of the EcR domains may be a key factor that allows a single EcR molecule to mediate diverse biological effects.

Application of electrophoretic methods for detection of protein-porphyrin complexes.

Simple methods for detection and isolation of protein-porphyrin complexes were elaborated in our laboratory. They are based on the separation of protein-porphyrin complexes in native polyacrylamide gel and measurement of their fluorescence, with the use of two detection systems: the commercially available Gel Doc(TM) 2000 system, and a system specially designed for the purpose of these investigations, concerning protein-porphyrin interactions. The fluorescent complexes can be electro-transferred from the gel onto PVDF membrane, eluted and analyzed in order to identify the protein interacting with porphyrins.