Proteolysis dysfunction in the process of aging and age-related diseases

In this review, we discuss in detail the most relevant proteolytic systems that together with chaperones contribute to creating the proteostasis network that is kept in dynamic balance to maintain overall functionality of cellular proteomes. Data accumulated over decades demonstrate that the effectiveness of elements of the proteostasis network declines with age. In this scenario, failure to degrade misfolded or faulty proteins increases the risk of protein aggregation, chronic inflammation, and the development of age-related diseases. This is especially important in the context of aging-related modification of functions of the immune system

Wpływ rekombinowanej ludzkiej erytropoetyny na układ odpornościowy

Erytropoetyna (EPO) jest hormonem produkowanym przez komórki śródmiąższowe nerki, który działa na komórki macierzyste erytrocytów, regulując ich proliferację i chroniąc przed apoptozą. Ze względu na budowę upodobniającą ją do czynników tkankowych o charakterze cytokin uważa się, że EPO poza wpływem na układ czerwonokrwinkowy może też regulować pracę układu odpornościowego. Przemawiają za tym wyniki licznych badań przeprowadzonych wśród chorych na przewlekłą chorobę nerek (PChN), którym podaje się rekombinowaną ludzką erytropoetynę (rhEPO) w celu wyrównania niedokrwistości, która jest nieodłącznym elementem tej choroby. W publikacji omówiono zaburzenia funkcjonowania układu odpornościowego z uwzględnieniem różnych rodzajów odpowiedzi immunologicznej. Przedstawiono też wyniki badań dotyczących pozytywnego wpływu leczenia za pomocą rhEPO u chorych hemodializowanych, które wskazują na istnienie wielu mechanizmów poprawy odpowiedzi immunologicznej u tych pacjentów. Artykuł ma na celu zwrócić uwagę na fakt, że poprawa funkcjonowania układu odpornościowego u pacjentów z PChN nie wydaje się być jedynie efektem korekcji niedokrwistości. Przemawia za tym zmiana ekspresji antygenów na limfocytach, dzięki którym możliwe są ich wzajemne relacje, poprawa proliferacji limfocytów T oraz zmiany w profilu cytokin u tych chorych. Owe zmiany mogą wynikać z naśladowania przez EPO działania innych cytokin bądź bezpośredniego wpływu na limfocyty i monocyty poprzez receptor dla EPO lub inne receptory z rodziny receptorów hematopoetyn, które charakteryzują się podobną budową i przekazują sygnał przez podobny system sygnalizacyjny w komórce

Rola limfocytów B w mechanizmach patogenezy wybranych chorób

Układ immunologiczny zdrowego człowieka ma za zadanie obronę oraz utrzymywanie homeostazy organizmu poprzez kontrolowanie antygenów zarówno wnikających do ustroju, jak i tych, które są w nim produkowane. Do najważniejszych komórek pełniących taką funkcję należą ściśle ze sobą współistniejące różne populacje limfocytów. To od nich zależą rozpoznanie antygenu i reakcja mająca na celu jego neutralizację. Jedną z form tej aktywności neutralizującej potencjalny patogen jest produkcja przeciwciał, których źródłem są limfocyty B. Zaburzenia funkcji tych tak zwanych efektorów odpowiedzi humoralnej (prowadzące w konsekwencji do zaburzeń liczby i swoistości przeciwciał) są podstawą (elementem patogenezy) wielu chorób i jako takie stały się celem licznych prac badawczych. Liczebność i proporcje klas limfocytów B, wielkość ich odpowiedzi na stymulację oraz poziom, klasy i swoistość wytwarzanych przeciwciał są bardzo ważnymi czynnikami diagnostycznymi i prognostycznymi w tak na pozór różnych chorobach, jak między innymi: przewlekła obturacyjna choroba płuc, astma oskrzelowa, śródmiąższowe zapalenie płuc, rak płuc czy, co oczywiste, liczne choroby autoimmunologiczne. Fakt ten podkreśla uniwersalność zastosowania obserwacji limfocytów B w kontroli przebiegu wielu schorzeń i ponownie wykazuje, że stan układu immunologicznego odzwierciedla stan zdrowia danej osoby. Forum Medycyny Rodzinnej 2011, tom 5, nr 1, 37-4

Influence of oxygen concentration on T cell proliferation and susceptibility to apoptosis in healthy men and women

Introduction. Much of what we know about the functioning of human T lymphocytes is based on the experiments carried out in atmospheric oxygen (O2) concentrations, which are significantly higher than those maintained in blood. Interestingly, the gender differences in the activity of T cells and their susceptibility to apoptosis under different O2 conditions have not yet been described. The aim of the study was to compare two main markers of lymphocyte function: proliferation capacity and ability to produce cytokines as well as their susceptibility to apoptosis under two different O2 concentrations, between men and women. Materials and methods. 25 healthy volunteers, both males (13) and females (12) were recruited to the study (mean age 25.48 ± 5.51). By using cytometry proliferation parameters of human CD4+ CD28+ cells or CD8+CD28+ cells in response to polyclonal stimulation of the TCR/CD3 complex at atmospheric (21%) and physiological (10%) O2 concentrations using our modified dividing cell tracking technique (DCT) were analyzed as well as the percentages of apoptotic cells. We also determined the levels of IFN-γ, IL-2, IL-10 and IL-17A using Cytometric Bead Array Flex system in cell culture supernatants. Results. CD4+CD28+ and CD8+CD28+ cells from the whole study group were characterized by shorter time required to enter the first (G1) phase of the first cell cycle at 21% compared to 10% O2. Both T cell populations performed significantly more divisions at 21% O2. The percentages of dividing cells were also significantly higher at atmospheric O2. Interestingly, data analysis by gender showed that male lymphocytes had similar proliferative parameters at both O2 concentrations while female lymphocytes proliferate more efficiently (note from the author: we cannot say that lymphocytes proliferate faster, rather more effectively, because cells perform more divisions, which gives more percentage of offspring cells) at 21% oxygen. Compared to males, the female CD4+ cells showed increased susceptibility to apoptosis at both O2 concentrations. No differences in the levels of cytokines regardless of gender and oxygen conditions were found. Conclusions. We showed that in vitro female T cells (both CD4+ and CD8+ cells) are more sensitive than male lymphocytes to low O2 concentration as demonstrated by the decrease in their proliferation dynamics. The effect does not depend on increased apoptosis of female T cells under low O2 because percentage of apoptotic cells was similar at both O2 concentrations

Hemodialysis Affects Phenotype and Proliferation of CD4-Positive T Lymphocytes

CD4+ T lymphocytes of patients with chronic kidney disease (CKD) are characterized by reduced levels of crucial surface antigens and changes in the cell cycle parameters. Recombinant human erythropoietin (rhEPO) normalizes their altered phenotype and proliferative capacity. Mechanisms leading to the deficient responses of T lymphocytes are still not clear but it is postulated that immunological changes are deepened by hemodialysis (HD). Study of activation parameters of CD4+ T lymphocytes in hemodialyzed and predialysis CKD patients could bring insight into this problem. Two groups of patients, treated conservatively (predialysis, PD) and hemodialyzed (HD), as well as healthy controls, were included into the study; neither had received rhEPO. Proportions of main CD4+CD28+, CD4+CD25+, CD4+CD69+, CD4+CD95+, and CD4+HLA-DR+ lymphocyte subpopulations and proliferation kinetic parameters were measured with flow cytometry, both ex vivo and in vitro. No differences were seen in the proportions of main CD4+ lymphocyte subpopulations (CD4+CD28+, CD4+CD25+, CD4+HLA-DR+, CD4+CD69+, CD4+CD95+) between all examined groups ex vivo. CD4+ T lymphocytes of HD patients exhibited significantly decreased expression of co-stimulatory molecule CD28 and activation markers CD25 and CD69 after stimulation in vitro when compared with PD patients and healthy controls. HD patients showed also decreased percentage of CD4+CD28+ lymphocytes proliferating in vitro; these cells presented decreased numbers of finished divisions after 72 h of stimulation in vitro and had longer G0→G1 time when compared to healthy controls. CD4+ T lymphocytes of PD patients and healthy controls were characterized by similar cell cycle parameters. Our study shows that repeated hemodialysis procedure influences phenotype and proliferation parameters of CD4+ T lymphocytes

An Ultrasensitive Biosensor for Detection of Femtogram Levels of the Cancer Antigen AGR2 Using Monoclonal Antibody Modified Screen-Printed Gold Electrodes.

The detection of cancer antigens is a major aim of cancer research in order to develop better patient management through early disease detection. Many cancers including prostate, lung, and ovarian secrete a protein disulfide isomerase protein named AGR2 that has been previously detected in urine and plasma using mass spectrometry. Here we determine whether a previously developed monoclonal antibody targeting AGR2 can be adapted from an indirect two-site ELISA format into a direct detector using solid-phase printed gold electrodes. The screen-printed gold electrode was surface functionalized with the anti-AGR2 specific monoclonal antibody. The interaction of the recombinant AGR2 protein and the anti-AGR2 monoclonal antibody functionalized electrode changed its electrochemical impedance spectra. Nyquist diagrams were obtained after incubation in an increasing concentration of purified AGR2 protein with a range of concentrations from 0.01 fg/mL to 10 fg/mL. In addition, detection of the AGR2 antigen can be achieved from cell lysates in medium or artificial buffer. These data highlight the utility of an AGR2-specific monoclonal antibody that can be functionalized onto a gold printed electrode for a one-step capture and quantitation of the target antigen. These platforms have the potential for supporting methodologies using more complex bodily fluids including plasma and urine for improved cancer diagnostics

Changes in the Expression of Transcription Factors Involved in Modulating the Expression of EPO-R in Activated Human CD4-Positive Lymphocytes.

We have recently described the presence of the erythropoietin receptor (EPO-R) on CD4(+) lymphocytes and demonstrated that its expression increases during their activation, reaching a level reported to be typical for erythroid progenitors. This observation suggests that EPO-R expression is modulated during lymphocyte activation, which may be important for the cells' function. Here we investigated whether the expression of GATA1, GATA3 and Sp1 transcription factors is correlated with the expression of EPO-R in human CD4(+) lymphocytes stimulated with monoclonal anti-CD3 antibody. The expression of GATA1, GATA3 and Sp1 transcription factors in CD4(+) cells was estimated before and after stimulation with anti-CD3 antibody by Western Blot and flow cytometry. The expression of EPO-R was measured using real-time PCR and flow cytometry. There was no change in the expression of GATA1 and GATA3 in CD4(+) lymphocytes after stimulation with anti-CD3 antibody. However, stimulation resulted in the significantly increased expression of the Sp1 factor. CD4(+) lymphocytes stimulated with anti-CD3 antibody exhibited an increase in both the expression level of EPOR gene and the number of EPO-R molecules on the cells' surface, the latter being significantly correlated with the increased expression of Sp1. Sp1 is noted to be the single transcription factor among the ones studied whose level changes as a result of CD4(+) lymphocyte stimulation. It seems that Sp1 may significantly affect the number of EPO-R molecules present on the surface of activated CD4(+) lymphocytes

Associated Effects of Cadmium and Copper Alter the Heavy Metals Uptake by Melissa Officinalis

Lemon balm (Melissa officinalis) is a popular herb widely used in medicine. It is often cultivated in soils with substantial heavy metal content. Here we investigate the associated effects of cadmium and copper on the plant growth parameters augmented by the manganese, zinc, and lead uptake indicators. The concentration of all elements in soil and plants was determined by the HR-CS FAAS with the ContrAA 300 Analytik Jena spectrometer. Bioavailable and total forms calculated for all examined metals were augmented by the soil analyses. The index of chlorophyll content in leaves, the activity of net photosynthesis, stomatal conductance, transpiration rate, and intercellular concentration of CO2 were also investigated. Either Cd or Cu acting alone at high concentrations in soil are toxic to plants as indicated by chlorophyll indices and gas exchange parameters. Surprisingly, this effect was not observed when both metals were administered together. The sole cadmium or copper supplementations hampered the plant’s growth, lowered the leaf area, and altered the plant’s stem elongation. Analysis of variance showed that cadmium and copper treatments of lemon balm significantly influenced manganese, lead, and zinc concentration in roots and above ground parts