Inhibitory immune checkpoint molecules and exhaustion of T cells in COVID-19.

COVID-19 (Coronavirus Disease) is an infectious disease caused by the coronavirus SARS-CoV-2 (Severe acute respiratory syndrome Coronavirus 2), which belongs to the genus Betacoronavirus. It was first identified in patients with severe respiratory disease in December 2019 in Wuhan, China. It mainly affects the respiratory system, and in severe cases causes serious lung infection or pneumonia, which can lead to the death of the patient. Clinical studies show that SARS-CoV-2 infection in critical cases causes acute tissue damage due to a pathological immune response. The immune response to a new coronavirus is complex and involves many processes of specific and non-specific immunity. Analysis of available studies has shown various changes, especially in the area of specific cellular immunity, including lymphopenia, decreased T cells (CD3+, CD4+ and CD8+), changes in the T cell compartment associated with symptom progression, deterioration of the condition and development of lung damage. We provide a detailed review of the analyses of immune checkpoint molecules PD-1, TIM-3, LAG-3 CTLA-4, TIGIT, BTLA, CD223, IDO-1 and VISTA on exhausted T cells in patients with asymptomatic to symptomatic stages of COVID-19 infection. Furthermore, this review may help to better understand the pathological T cell immune response and improve the design of therapeutic strategies for patients with SARS-CoV-2 infection.

The impact of external stress factors on hippocampus volume during antidepressant treatment

Background and objectives: Magnetic resonance imaging (MRI) studies suggest that depression is associated with volumetric hippocampal changes. Investigations of these structures during antidepressant therapy is therefore important, however, volumetric studies are rare in this case. We aimed to study the effect of AD treatment on volumetric changes in hippocampus depending on stress factors in depressive patients.MethodsThirty patients with major depressive disorder (MDD) underwent MRI of the brain on the day of admission and at the time of stabilization of acute depressive symptomatology by venlafaxine. The presence of long-lasting stress factors in these patients was investigated by the social readjustment rating scale questionnaire.ResultsNo significant differences were found in hippocampi volumes before and after venlafaxine treatment. However, regression analysis revealed significant positive relation between stress factors and volumetric hippocampus change during AD treatment.ConclusionIt seems that antidepressant treatment by venlafaxine could be more suitable in the MDD patients with presence of stress-factors. (AU)

The metabolism of 5-methylcytosine residues in DNA.

The fundamental biochemical processes of 5-methylcytosine (5-mC) synthesis, maintenance, conversion and removal determine the time and spatial pattern of DNA methylation. This has a strong effect on a plethora of physiological aspects of cellular metabolism. While the presence of 5-mC within the promoter region can silence gene expression, its derivative - 5-hydroxymethylcytosine exerts an opposite effect. Dysregulations in the metabolism of 5-mC lead to an altered DNA methylation pattern which is linked with a disrupted epigenome, and are considered to play a significant part in the etiology of several human diseases. A summary of recent knowledge about the molecular processes participating in DNA methylation pattern shaping is provided here.

The age at onset in Multiple Sclerosis is associated with patient's prognosis.

BACKGROUND: The causes of the individual differences in the rate of disability progression in multiple sclerosis (MS) are still not completely clear. According to the long-term prognosis of MS patients, the search for new valuable prognostic markers of "benign" or "malign" MS is necessary. OBJECTIVES: Our aim was to assess the possible association of MS onset age with the disease disability progression rate in Slovak patients with MS. METHODS: By the unique pattern of evaluation of disability progression rate using Multiple Sclerosis Severity Score (MSSS), each of 270 MS patients was defined as slow-progressing, mid-rate progressing or rapidly progressing. RESULTS: We found a significant differences in the age at onset between MS patients with different rate of disability progression (p(K-W)<0,00005). The faster was a disability progression assessed by MSSS score, the higher was the MS onset age. CONCLUSION: We showed for the first time in Central European Slovak population that MS onset age is an early marker that is in the positive correlation with disease disability progression rate, evaluated by MSSS score. We conclude that relapsing-remitting MS patients older at clinical onset have a higher risk of unfavorable prognosis (Tab. 2, Fig. 1, Ref. 21).

The Role of Heat Shock Proteins in Leukemia.

Heat shock proteins (HSPs) HSP27, HSP70 and HSP90 are molecular chaperones; their expression is increased after exposure of cells to conditions of environmental stress, including heat shock, heavy metals, oxidative stress, or pathologic conditions, such as ischemia, infection, and inflammation. Their protective function is to help the cell cope with lethal conditions. The HSPs are a class of proteins which, in normal cells, are responsible for maintaining homeostasis, interacting with diverse protein substrates to assist in their folding, and preventing the appearance of folding intermediates that lead to misfolded or damaged molecules. They have been shown to interact with different key apoptotic proteins and play a crucial role in regulating apoptosis. Several HSPs have been demonstrated to directly interact with various components of tightly regulated caspase-dependent programmed cell death. These proteins also affect caspase-independent apoptosis by interacting with apoptogenic factors. Heat shock proteins are aberrantly expressed in hematological malignancies. Because of their prognostic implications and functional role in leukemias, HSPs represent an interesting target for antileukemic therapy. This review will describe different molecules interacting with anti-apoptotic proteins HSP70 and HSP90, which can be used in cancer therapy based on their inhibition.

Type and occurrence of serious complications in patients after mild traumatic brain injury.

Traumatic brain injury (TBI) remains a major public health and socio-economic problem, and 70-90% of all TBIs are classified as mild. Mild TBIs and concussions are mostly considered to be non-serious conditions with symptoms subsiding within a few days or weeks. However in 10-15% of patients, the symptoms persist one year after concussion and mostly include headache, fatigue, irritability, and cognitive problems (e.g. memory, concentration). These persisting symptoms negatively influence patient daily activities as postconcussion syndrome (PCS). Second-impact syndrome (SIS) is a very rare but usually fatal condition and occurs when repeated brain injuries lead to a catastrophic diffuse brain swelling. There is no scientific evidence on the incidence and risk of SIS. Chronic traumatic encephalopathy (CTE) is a progressive degenerative disease of the brain found in patients with a history of repetitive brain trauma. CTE presents with behavioural, cognitive, and motor symptoms. The literature to date lacks prospective epidemiological studies of the incidence of CTE. In recent medical literature, there is a description of 110 athletes with postmortem diagnosis of CTE (Tab. 1, Ref. 37).

Application of atomic force microscopy for investigation of Na(+),K(+)-ATPase signal-transducing function.

The Young's modulus of 10-12-day-old chick embryos' sensory neurons cultivated in dissociated cell culture was measured using a PeakForce Quantitative Nanomechanical Mapping atomic force microscopy. The native cells were tested in control experiments and after application of ouabain. At low "endogenous" concentration of 10⁻¹° M, ouabain tended to increase the rigidity of sensory neurons. We hypothesize that this trend resulted from activation of Na⁺,K⁺-ATPase signal-transducing function.

A new approach in DCE MRI data analysis for differentiating benign and malignant breast lesions.

BACKGROUND: Dynamic contrast enhanced MRI (DCE MRI) is able to reflect changes in vascularity, vessel permeability and extracellular diffusion space of tissues. The goal of this study was to investigate the use of DCE MRI to differentiate benign and malignant breast lesions. PATIENTS AND METHODS: From a database, five patients with malignant and five patients with benign lesions were randomly chosen. All patients underwent measurement in a 3T MR scanner using a breast coil. A series of T1-weighted MRI were performed using an intravenously delivered contrast agent. Then, 17 postcontrast sets were acquired within a timeframe of 13 seconds. All DCE MRI data were evaluated using the JIM image analysis package. We observed changes in signal intensity over the acquisition time -  curves of dynamic contrast enhancement. CONCLUSION: We investigated parts of the curves with the largest increase in signal intensity during the timeframe. For further comparison, we used values of the highest signal intensity increases between the timeframes. Analysis of these results led to the proposal that the threshold between benign and malignant lesion had a relative value of 100. Furthermore, there was a significant difference between these two types of lesions.

Hyperhomocysteinemia as a risk factor for the neuronal system disorders.

Elevated concentration of the homocysteine (Hcy) in human tissues, resulting either from mutations in genes enconding Hcy-metabolizing enzymes, or from deficiences of folic acid has recognized cytotoxic effect. Even a mild Hcy level increase is a risk factor for cardiovascular diseases and stroke in humans and also a risk factor for neurodegenerative disordes, such as dementia, or Alzheimer's disease. However, it is not yet clear whether homocysteine is a marker, or a causative agent. We present here an overview of recent data on the homocysteine metabolism and on the genetic and the metabolic causes of hyperhomocysteinemia-related pathologies in humans. In context of our results which detected an increased oxidative stress in hyperhomocysteinemic rats we discuss here the role of free radicals in this disorder. Imbalance between homocysteine auto-oxidation, production of reactive metabolites and cellular antioxidant defence induced by hyperhomocysteinemia results to cytotoxicity by oxidizing membrane lipids and proteins. Consequently, protein thiolation and homocysteinylation results in the structural and functional modifications of cells, including neuronal ones. It is our hope that identification of prophylacting factors effective in the prevention of toxic effect of Hcy would lead to improved therapeutics, especially the brain tissue.

Clinical correlations of proton magnetic resonance spectroscopy findings in acute phase after mild traumatic brain injury.

INTRODUCTION: Standard brain magnetic resonance imaging (MRI) is typically normal in most patients after mild traumatic brain injury (MTBI). Proton magnetic resonance spectroscopy (¹H-MRS) is more sensitive to detect subtle post-traumatic changes. The aim of the study was to evaluate the clinical correlations of these changes in the acute phase (within 3 days) after MTBI. METHODS: Twenty-one patients with MTBI and 22 controls were studied. Both groups underwent neuropsychological testing and single-voxel ¹H-MRS examination of both frontal lobes and upper brainstem. RESULTS: Significant decrease in NAA was found in both frontal lobes and in NAA/Cre ratio in the right frontal lobe (p < 0.05). Correlation analysis showed a correlation of NAA in the left frontal lobe with Backward Digit Span (p = 0.022) and Stroop test A (p = 0.0034) and a weak correlation with TMT B time (p = 0.046). The NAA/Cre in the right frontal lobe correlated with Stroop test A (p = 0.007) and with the total score of Digit Span (p = 0.016). Lower NAA was found in the upper brainstem (p = 0.0157) in the sub-group of patients with post-traumatic unconsciousness. CONCLUSIONS: This study found a correlation of ¹H-MRS metabolite changes with cognitive decline and presence or absence of loss of consciousness in the acute phase after MTBI.

Correlations of ADHD symptoms with neurometabolites measured by 1H magnetic resonance spectroscopy.

OBJECTIVES: Despite the number of studies on neurometabolite changes in ADHD (Attention deficit/hyperactivity disorder), there is lack of evidence on neurometabolite associations with ADHD symptoms. BACKGROUND: We aimed to find the correlations of neurometabolites with ADHD symptoms. METHODS: Twenty ADHD children were examined by means of 1H-MRS. The spectra were taken from dorsolateral prefrontal cortex (DLPFC) and white matter behind DLPFC, bilaterally. Neurometabolites were correlated with ADHD-RS-IV (ADHD-Rating Scales IV), CPRS (Conners Parent rating Scale) and DPREMB (Daily Parent Rating of Evening and Morning Behavior) scores. RESULTS: NAA/Cr (N-acetylaspartate/creatine) in the right DLPFC positively correlated with CPRS subscale IV learning problems and negatively correlated in the left white matter with DPREMB morning behavior subscale and ADHD-RS-IV score. Glx/Cr (glutamate + glutamine/creatine) positively correlated in the right white matter with ADHD-RS-IV and negatively correlated in the left white matter with DPREMB morning behavior subscale score. Cho/Cr (choline/creatine) in the left white matter negatively correlated with DPREMB morning behavior subscale and ADHD-RS-IV score. CONCLUSION: ADHD symptoms could result from different activities of the left- and right-hemisphere prefrontal circuits. In consequence to impulses to novel task searching the increased right prefrontal circuit activity could be mediated by different motivational control (Fig. 9, Ref. 73).

Expression of Ki-67, Bcl-2, Survivin and p53 Proteins in Patients with Pulmonary Carcinoma.

Apoptosis is the fundamental process necessary for eliminating damaged or mutated cells. Alterations in the apoptotic pathway appear to be key events in cancer development and progression. Bcl-2 is the key member of the Bcl-2 family of apoptosis regulator proteins with anti-apoptotic effects. Survivin acts as an inhibitor of apoptosis as well and has been implicated in both inhibition of apoptosis and mitosis regulation. p53 is one of the tumor suppressor proteins, prevents tumor formation through cell cycle blocking and eliminates damaged cells via the activation of apoptosis. The Ki-67 protein is a cellular marker for proliferation. To investigate the possible interactions of the aforementioned proteins, we examined their expression in 76 patients with diagnosed lung cancer using immunohistochemical visualisation. Ki-67 protein was expressed in the cancer cells of all patients with small cell lung cancer (SCLC). We found a negative correlation between survivin and p53 expression. A decreased intensity of survivin expression and fewer cells positive for survivin (66.7%) in SCLC in comparison with other lung cancer types (98.0%) was detected. Reversely, expression of Bcl-2 was found in more than 90% of cases with SCLC. We hypothesize that high expression and intensity of Bcl-2 protein could be a factor behind a bad prognosis in SCLC.

Age-associated changes in Ca(2+)-ATPase and oxidative damage in sarcoplasmic reticulum of rat heart.

Altered Ca(2+) handling may be responsible for the development of cardiac contractile dysfunctions with advanced age. In the present study, we investigated the roles of oxidative damage to sarcoplasmic reticulum (SR) and expression of Ca(2+)-ATPase (SERCA 2a) and phospholamban in age-associated dysfunction of cardiac SR. SR vesicles were prepared from hearts of 2-, 6-, 15-, and 26-month-old Wistar rats. Although activity of Ca(2+)-ATPase decreased with advancing age, no differences in relative amounts of SERCA 2a and phospholamban protein were observed. On the other hand, significant accumulation of protein oxidative damage occurred with aging. The results of this study suggest that age-related alteration in Ca(2+)-ATPase activity in the rat heart is not a consequence of decreased protein levels of SERCA 2a and phospholamban, but could arise from oxidative modifications of SR proteins. Cellular oxidative damage caused by reactive oxygen species could contribute to age-related alternations in myocardial relaxation.

Radiation-induced long-term alterations in hippocampus under experimental conditions.

BACKGROUNDS: The aim of the present study was to investigate the effect of ionizing radiation on the cell population that co-forms hippocampal formation in an adult rat brain. MATERIALS AND METHODS: Adult male Wistar rats were exposed to whole-body irradiation with fractionated doses of gamma rays (the total dose of 4 Gy). Thirty, 60 and 90 days after irradiation the cell-specific types housed in the CA1, CA3 subregions and adjacent layers were labelled using immunohistochemistry for specific cell phenotypes; Ki-67 marker was used for proliferating cells and GFAP for detection of astrocytes. RESULTS: During the 30th day post-exposure, a considerable increase in the numbers of Ki-67-positive cells was seen. Moreover, significant decline in the density of neurons, mostly in the CA1 subregion, was observed on the 60th day. Slight overaccumulation of Ki-67-positive cells was seen in CA1 area 90 days after radiation treatment. Temporary decrease of GFAP-positive astrocytes was seen thirty days after irradiation, followed by their subsequent increase 60 days after exposure. Secondary decrease of GFAP-positive cells in both of regions was found in the group surviving 90 days post-irradiation. CONCLUSION: Results showed that radiation response of neurons and astrocytes that form the adult hippocampus may play contributory role in the development of prognostically unfavourable adverse radiation-induced late effect.

Lung adenocarcinoma and squamous cell carcinoma in association with genetic polymorphisms of GSTs in Slovak population.

Slovak Republic belongs to the countries with high incidence of lung cancer. Gene polymorphisms of the glutathione S-transferases (GSTs) may play a role in individual lung cancer susceptibility. In presented case-control study we investigate the incidence of polymorphism of GSTT1, GSTM1, GSTP1 genes and their combinations as possible predictive factors for identification of individuals with increased risk of formation and development of adenocarcinoma (AC) and squamous cell carcinoma (SCC) of lung in Slovak population. The study was conducted on 520 individuals consisting of 118 patients with adenocarcinoma, 112 patients with squamous cell carcinoma and 290 control individuals. GSTT1, GSTM1, GSTP1 gene polymorphisms were assayed by standard PCR and PCR-RFLP technique. The results of this study indicate that the GSTT1null-genotype and combination GSTT1 null and Ile/Val or Val/Val are associated with increased risk of lung adenocarcinoma. A significant association with 2.13 - fold increased risk was observed between lung adenocarcinoma and GSTT1 null genotype (95% CI = 1.29 - 3.51; p= 0.004). Also it was proved 2.83 times statistically higher risk for development of this histological type of lung cancer (95% CI = 1.34 - 6.01; P= 0.005) in combination of GSTT1null and Ile/Val or Val/Val genotypes. GSTT1, GSTM1, GSTP1 polymorphism did not show any significant association with SCC. Our study suggests that genetic make-up in metabolizing genes may increase susceptibility towards lung cancer development.

Why mitochondria are excellent targets for cancer therapy.

New insights into cancer cells - specific biological pathways are urgently needed to promote development of exactly targeted therapeutics. The role of oncoproteins and tumor suppressor proteins in proliferative signaling, cell cycle regulation and altered adhesion is well established. Chemicals, viruses and radiation are also generally accepted as agents that commonly induce mutations in genes encoding these cancer-inducing proteins, thereby giving rise to cancer. More recent evidence indicates the importance of two additional key factors imposed on proliferating cells - hypoxia and/or lack of glucose. These two additional triggers can initiate and promote the process of malignant transformation, when a low percentage of cells escape cellular senescence. Disregulated cell proliferation leads to formation of cellular masses that extend beyond the resting vasculature, resulting in oxygen and nutrient deprivation. Resulting hypoxia triggers a number of critical adaptations that enable cancer cell survival. The process of apoptosis is suppressed and glucose metabolism is altered. Recent investigations suggest that oxygen depletion stimulates mitochondria to compensate increased reactive oxygen species (ROS). It activates signaling pathways, such as hypoxia-inducible factor 1, that promote cancer cell survival and tumor growth. During the last decade, mitochondria have become key organelles involved in chemotherapy-induced apoptosis. Therefore, the relationship between mitochondria, ROS signaling and activation of survival pathways under hypoxic conditions has been the subject of increased study. Insights into mechanisms involved in ROS signaling may offer novel ways to facilitate discovery of cancer-specific therapies.

Microsomal epoxide hydrolase polymorphisms, cigarette smoking and prostate cancer risk in the Slovak population.

Polymorphisms in tobacco carcinogen metabolizing enzymes may generate interindividual variations towards the risk of developing prostate cancer. One of these enzymes is microsomal epoxide hydrolase (EPHX1) which metabolizes polycyclic aromatic hydrocarbons, or PAH, carcinogens found in cigarette smoke. The activity of this enzyme is affected by two polymorphisms, a substitution of Tyr113 by His in exon 3 and a substitution of His139 by Arg in exon 4. The aim of this study was to use a population-based case-control study to investigate whether or not such genetic polymorphisms in EPHX1 gene can modify the relationship between smoking status and the risk of developing prostate cancer. We used restriction fragment length polymorphism, or PCR-RFLP to determine EPHX1 genotypes in subjects comprising 194 patients with histologically verified prostate cancer and 305 healthy individuals as control. We found no overall association between prostate cancer risk and functional polymorphisms of EPHX1 gene in exon 3 and exon 4. We further analysed the association between the EPHX1 genotypes and smoking. Smokers carrying the exon 3 Tyr/Tyr and Tyr/His genotypes were at no significant risk compared to non-smokers with the "rapid" Tyr/Tyr genotype. By contrast, a significant interaction of smoking and the exon 4 polymorphism was present.

Effects of aging on activities of mitochondrial electron transport chain complexes and oxidative damage in rat heart.

Mitochondrial dysfunction and accumulation of oxidative damage have been implicated to be the major factors of aging. However, data on age-related changes in activities of mitochondrial electron transport chain (ETC) complexes remain controversial and molecular mechanisms responsible for ETC dysfunction are still largely unknown. In this study, we examined the effect of aging on activities of ETC complexes and oxidative damage to proteins and lipids in cardiac mitochondria from adult (6-month-old), old (15-month-old) and senescent (26-month-old) rats. ETC complexes I-IV displayed different extent of inhibition with age. The most significant decline occurred in complex IV activity, whereas complex II activity was unchanged in old rats and was only slightly reduced in senescent rats. Compared to adult, old and senescent rat hearts had significantly higher levels of malondialdehyde, 4-hydroxynonenal (HNE) and dityrosine, while thiol group content was reduced. Despite marked increase in HNE content with age (25 and 76 % for 15- and 26-month-old rats, respectively) Western blot analysis revealed only few HNE-protein adducts. The present study suggests that non-uniform decline in activities of ETC complexes is due, at least in part, to mitochondrial oxidative damage; however, lipid peroxidation products appear to have a limited impact on enzyme functions.

[Five year-results of chemoresistance testing in cancer patients]. / Nase pät'rocné výsledky in vitro testovania chemorezistencie u onkologických pacientov.

BACKGROUNDS: Translational medicine is a medical field encompassing basic research and development of new diagnostic and therapeutic strategies for clinical practice. The present scientific paper focuses on our previous experience in the field of chemoresistance testing in patients with oncological diseases. MATERIAL AND METHODS: Since 2005, we sampled 71 patients with a leukaemia (AML, ALL and CML) and 92 patients with a solid tumour (lung and gastrointestinal tract cancer). Malignant cell in vitro drug resistance testing was carried out using cytotoxic methyl-thiazol tetrazolium (MTT) assay. RESULTS: Based on the LC50 (lethal concentration of a drug killing 50% of cell population), we found that patients with acute myeloblastic leukaemia exhibit a greater degree of resistance than patients with acute lymphoblastic leukaemia. In patients with bronchogenic carcinomas, primary resistance to cisplatin was identified in 28% of tested samples, paclitaxel 36%, vincristine 50%, etoposide 56%, vinorelbine 57%, topotecan 62%, gemcitabine 77% and dacarbazine 86%. CONCLUSION: In vitro tests with gastrointestinal tract cancers also suggested high effectiveness of cisplatin (with the exception of gastric carcinoma) that was comparable with 5-fluorouracil. Even though the MTT assay has some limitations (insufficient number of vital cells, possible contamination by non-malignant cells, etc.), this in vitro method proved very effective in testing malignant cell resistance to clinically used cytostatics.

Serum level of IGFBP3 and IGF1/IGFBP3 molar ratio in addition to PSA and single nucleotide polymorphism in PSA and CYP17 gene may contribute to early diagnostics of prostate cancer.

The aim of the paper is to determine whether IGF1, IGFBP3 and IGF1/IGFBP3 molar ratio in addition to PSA and one-nucleotide polymorphism in PSA and CYP17 gene might contribute to early diagnostics of prostate cancer (PCa). Serum level of PSA, IGF1 and IGFBP3 in the group of 158 individuals (92 PCa and 66 controls) was examined by RIA method and IGF1/IGFBP3 was calculated. PCR RLFP method was used to examine one- nucleotide polymorphism in PSA and CYP 17 gene. The results suggest that serum level of IGF1 over 95% CI did not increase relative risk of PCa development in overall group, not even regarding to particular investigated genotypes, not even if individuals with genotype AG+A1A1, AG+A1A2, GG+A1A1 and GG+A1A2 were evaluated. Serum level of IGFBP3 under 95% CI increased PCa relative risk in overall group(chi(2) = 10,03, p= 0,001, OR 3,12, 95% CI 1,44-6,93), as well as regarding to one-nucleotide polymorphism in individuals with PSA genotype AG(chi(2) = 4,72 p= 0,029, OR 2,87, 95% CI 01,09-7,49) and CYP 17 genotype A1A1(chi(2) = 3,76 p= 0,052, OR 2,57, 95% CI 0,97-6,75). The association between frequencies of occurrence of PCa and higher IGF1/IGFBP3 molar ratio was not confirmed, nor for gene polymorphism in PSA and CYP17, however OR (chi(2) = 1,58, p= 0,208, OR 1,67, 95% CI 0,75-3,71) was more than 1, nor in combination AG+A1A1,AG+ A1A2. Serum level of IGFBP3 and IGF1/IGFBP3 molar ratio in addition to PSA and gene polymorphism in PSA and CYP17 gene might contribute to early diagnostics of PCa. Further research is needed to prove, whether serum level of IGFBP3 in addition to PSA determines the prognosis and progression of PCa.