Alzheimer's disease (AD) is associated with the deposition of β-amyloid in the brain. AD accounts for over 50% of cases of dementia which results from disturbances in redox homeostasis. Indeed, ...increased intensity of protein oxidation and nitration as well as lipid peroxidation is observed in brain areas with considerable amounts of amyloid plaques and neurofibrillary tangles. However, little is known about the oxidoreductive balance of salivary glands in AD patients. Therefore, the aim of this study was to evaluate the antioxidant barrier and oxidative/nitrosative stress biomarkers in stimulated saliva and blood of AD patients. The study was participated by 25 AD patients and 25 non-demented controls without neurological diseases or cognitive impairment, matched by age and gender to the study group. The number of patients was determined based on a previous pilot study (test power = 0.9). We found a significant decrease in the activity of erythrocyte superoxide dismutase (SOD) and glutathione peroxidase (GPx), increased activity of catalase (CAT) and reduced concentration of plasma non-enzymatic antioxidants (uric acid, UA and reduced glutathione, GSH). In contrast, in the stimulated saliva of AD patients we observed significantly decreased activity of all antioxidant enzymes (SOD, CAT and GPx) as well as concentration of GSH compared to the control group. The content of lipid (malondialdehyde, MDA) and protein (advanced oxidation protein products, AOPP; advanced glycation end-products, AGE) oxidation products as well as biomarkers of nitrosative stress (peroxynitrite, nitrotyrosine) was significantly higher in both saliva and plasma of AD patients compared to the controls. In AD patients, we also observed a considerable decrease in stimulated saliva secretion and salivary total protein content, and an increase in salivary β-amyloid concentration. In conclusion, AD results in redox imbalance towards oxidative reactions, both at the level of the oral cavity and the entire body. General redox balance disturbances do not coincide with salivary redox balance disturbances. Reduction in stimulated saliva secretion in AD patients reflects secretory dysfunction of the parotid glands.
Neutrophils (PMN) play a key role in eliciting congenital immune response. These cells are equipped with specific receptors that are located on the surface of their cell membrane. These receptors ...produce various signals which in turn help in the effective functioning of PMN. The activity of these cells may be modified by factors of endo- and exogenous origin, including xenoestrogens such as bisphenol A (BPA). The aim of this study was to evaluate the effect of BPA on the expression of CD11c, CD14, CD15, CD16, CD62L and CD284 compounds on the surface of neutrophils in women and men. The study material included PMN isolated from the whole blood. The cells were incubated in the presence of BPA and/or LPS. Flow cytometry technique was used to evaluate the expression of CD antigens. Studies of these receptors indicate that BPA, at a concentration corresponding to the serum level of this compound in healthy subjects as well as at higher doses, induces changes in the immunophenotype of PMN, which may lead to immunity disorders associated with the dysfunction of these cells. Moreover, the observed effects of xenoestrogen on the expression of CD11c, CD14, CD15, CD16, CD62L and CD284 differentiation markers on these cells are sex-independent.
Ovarian cancer (OC) is one of the deadliest gynecological cancers, largely due to the fast development of metastasis and drug resistance. The immune system is a critical component of the OC tumor ...microenvironment (TME) and immune cells such as T cells, NK cells, and dendritic cells (DC) play a key role in anti-tumor immunity. However, OC tumor cells are well known for evading immune surveillance by modulating the immune response through various mechanisms. Recruiting immune-suppressive cells such as regulatory T cells (Treg cells), macrophages, or myeloid-derived suppressor cells (MDSC) inhibit the anti-tumor immune response and promote the development and progression of OC. Platelets are also involved in immune evasion by interaction with tumor cells or through the secretion of a variety of growth factors and cytokines to promote tumor growth and angiogenesis. In this review, we discuss the role and contribution of immune cells and platelets in TME. Furthermore, we discuss their potential prognostic significance to help in the early detection of OC and to predict disease outcome.
Background and Purpose
Bruton's tyrosine kinase (Btk) is a non‐receptor tyrosine kinase involved in the activation of signalling pathways responsible for cell maturation and viability. Btk has ...previously been reported to be overexpressed in colon cancers. This kind of cancer is often accompanied by anaemia, which is treated with an erythropoietin supplement. The goal of the present study was to assess the effects of combination therapy with erythropoietin β (Epo) and LFM‐A13 (Btk inhibitor) on colon cancer in in vitro and in vivo models.
Experimental Approach
DLD‐1 and HT‐29 human colon adenocarcinoma cells were cultured with Epo and LFM‐A13. Cell number and viability, and mRNA and protein levels of Epo receptors, Btk and Akt were assessed. Nude mice were inoculated with adenocarcinoma cells and treated with Epo and LFM‐A13.
Key Results
The combination of Epo and LFM‐A13 mostly exerted a synergistic inhibitory effect on colon cancer cell growth. The therapeutic scheme used effectively killed the cancer cells and attenuated the Btk signalling pathways. Epo + LFM‐A13 also prevented the normal process of microtubule assembly during mitosis by down‐regulating the expression of Polo‐like kinase 1. The combination of Epo and LFM‐A13 significantly reduced the growth rate of tumour cells, while it showed high safety profile, inducing no nephrotoxicity, hepatotoxicity or changes in the haematological parameters.
Conclusion and Implications
Epo significantly enhances the antitumour activity of LFM‐A13, indicating that a combination of Epo and LFM‐A13 has potential as an effective therapeutic approach for patients with colorectal cancer.
Mucoadhesive gelling systems based on chitosan and chitosan/β-glycerophosphate (β-GP) were developed in order to increase clotrimazole residence time in the vaginal cavity. Ex vivo mucoadhesiveness ...using porcine vaginal mucosa followed with mechanical, viscoelastic, and swelling properties of prepared hydrogels were evaluated. Drug-free, sterile, unmodified, and β-GP crosslinked chitosan were investigated for the in vitro cytotoxicity in CRL 2616 human vaginal mucosa cells using MTT assay, fluorescent microscopy, and flow cytometry analysis. Chitosan/β-GP hydrogels exhibited pseudoplastic and thixotropic properties. Ionic interaction between β-GP and chitosan improved mechanical properties of hydrogels in terms of hardness, cohesiveness, and compressibility. The hydrogels’ ability to interact with porcine vaginal mucosa (measured as force of detachment and work of adhesion) was comparable to those obtained with reference mucoadhesive gel Replens™. Surprisingly, greater mucoadhesive properties were noticed for chitosan/β-GP hydrogels. The cytotoxic effect of unmodified and β-GP crosslinked chitosan was hardly affected by chitosan molecular weight, exhibited mainly through inducing apoptosis, and was found to be significantly lower in the presence of chitosan/β-GP. Furthermore, the higher amount of β-GP was used to crosslink chitosan, the lower cytotoxic effect was observed.
The aim of this study was to examine the viability of neurons and the putative neuroprotective effects of second-generation antiepileptic drug, levetiracetam (LEV), on cultured hippocampal neurons ...injured by hyperthermia.
Primary cultures of rat's hippocampal neurons at 7day in vitro (DIV) were incubated in the presence or absence of LEV in varied concentrations under hyperthermic conditions. Cultures were heated in a temperature of 40°C for 24h or in a temperature of 41°C for 6h. Flow cytometry with Annexin V/PI staining as well as fluorescent microscopy assay were used for counting and establishing neurons as viable, necrotic or apoptotic. Additionally, the release of lactate dehydrogenase (LDH) to the culture medium, as a marker of cell death, was evaluated. Assessment was performed after 9DIV and 10 DIV.
Incubation of hippocampal cultures in hyperthermic conditions resulted in statistically significant increase in the number of injured neurons when compared with non-heated control cultures. Intensity of neuronal destruction was dependent on temperature-value. When incubation temperature 40°C was used, over 80% of the population of neurons remained viable after 10 DIV. Under higher temperature 41°C, only less than 60% of neurons were viable after 10 DIV. Both apoptotic and necrotic pathways of neuronal death induced by hyperthermia were confirmed by Annexin V/PI staining.
LEV showed no neuroprotective effects in the current model of hyperthermia in vitro. Moreover, drug, especially when used in higher concentrations, exerted unfavorable intensification of aponecrosis of cultured hippocampal neurons.
Osteochondral defects of the knee are common in orthopaedic patients. They are challenging to treat, especially in young, highly demanding patients who do not qualify for arthroplasty. Among the many ...possibilities to treat osteochondral lesions presented so far, none is ideal. Because of the poor healing potential of cartilage, treatment outcomes significantly worsen with larger lesions. The treatment of large defects usually requires expensive solutions, sometimes including second-stage surgery. Using mesenchymal stem cell transplantation and cancellous bone autografts, the technique presented here for osteochondral lesion reconstruction can be effectively used to treat large osteochondral lesions in a single-stage procedure.
Video 1
Osteochondral lesion reconstruction using mesenchymal stem cell transplantation and cancellous bone autografts; right knee. Patients are considered candidates after careful selection. To promote mesenchymal stem cell growth, 4 days before surgery, patients are administered granulocyte colony-stimulating factor. On the day of surgery, the mesenchymal stem cell suspension is obtained, using an MSC blood separator, under the control of the blood bank. A cytometric test is performed for the quantitative evaluation of cell lines, as well as a histopathological evaluation of stained preparations. The cell suspension is then transferred to the operating theater. The diagnosis is confirmed arthroscopically through a standard anterolateral portal, and concomitant lesions are excluded. Mini-open access is created next to the site of the lesion. After the lesion is exposed, delaminated cartilage or bony-cartilage sequestrum is removed. Using a bone spoon, the bony edges of the cavity are resected to healthy cartilaginous tissue. The next step is decortication and debridement of the bottom. To ensure the integration of bone grafts, it is necessary to remove the sclerotic subchondral bone layer until bleeding from the bone occurs. An additional factor improving the blood supply is the microfracturing the bottom of the cavity. From a separate site above the iliac crest, autologous cancellous bone grafts are harvested. The bottom of the lesion is filled with the debris of the cancellous bone. Restoration with a bone graft is performed up to the height of the surrounding healthy bone margin. During this time, the previously sized collagen sponge Tachosil is soaked in MSC suspension. The sponge is placed on the lesion and adjusted to the edges of the lesion using tweezers. The soaked implant should not cover surrounding healthy cartilage edges, but only fill the cavity. The entirety of the damaged fragment is covered with a 2-component fibrin glue. After binding, a suspension of mesenchymal cells, is administered under the fibrin layer using a no. 12 needle, and the application site is sealed with the rest of the glue. The wound is closed with layered sutures and sealed with sterile dressing. The operated joint is protected by cocoon dressing to limit mobility in the early postoperative period.
The low number of circulating endothelial progenitor cells (EPCs) has emerged as a biomarker of cardiovascular (CV) risk in adults. Data regarding EPCs in paediatric populations with CV risk factors ...are limited. The aim of the study was to estimate the EPC number and its relationship with vascular function and structure in children with type 1 diabetes mellitus (T1DM).
We performed a comparative analysis of 52 children with T1DM (mean age 14.5 years; diabetes duration, 6.0 years; HbA1c level, 8.5%) and 36 healthy age- and gender-matched control children. EPCs were identified and analysed by flow cytometry with the use of MABs directed against CD34, CD144 (VE-cadherin) and CD309 (VEGFR-2). sICAM-1, hsCRP, thrombomodulin and adiponectin levels were also assessed. We evaluated vascular function (flow-mediated dilation (FMD)) and structure (carotid intima-media thickness (IMT)) ultrasonographically.
Frequencies of CD34+ cells were similar in both groups (P=0.30). In contrast, frequencies of CD34+VE-cadherin+ cells were significantly higher in diabetic children compared with the healthy group (P=0.003). Similarly, diabetic patients tended to present with higher frequencies of CD34+VEGFR+ cells (P=0.06). FMD was lower (6.9 vs 10.5%, P=0.002) and IMT was higher (0.50 vs 0.44 mm, P=0.0006) in diabetic children. We demonstrated a significant relationship between CD34+VEGFR-2+ cells and BMI (r=0.3, P=0.014), HDL (r=-0.27, P=0.04), sICAM-1 (r=0.47, P=0.023) and FMD (r=-0.45, P<0.001). Similarly, frequencies of CD34+VE-cadherin+ cells were significantly correlated with BMI (r=0.32, P=0.02) and FMD (r=-0.31, P=0.03).
We demonstrated here that increased frequencies of EPCs observed in diabetic children are negatively correlated with endothelial function. Further studies are warranted to assess whether this phenomenon might result from effective mobilisation of EPCs in order to repair damaged endothelium in children at increased risk for atherosclerosis.
We aimed to analyse whether quantitative assessment of peripheral blood lymphocyte CD19+CD20+CD22+CD79a+ B cells, CD3+CD4+CD5+CD8+ T cells and CD4+CD25+++Foxp3high Treg can improve prognostication in ...DLBCL patients.
The absolute count of lymphocytes, B-cells, T-cells and Treg-cells as well as the percentage of apoptotic cells were assessed by means of flow cytometry in all studied subjects.
Significantly lower level of ALC and the percentage of apoptotic cells have been observed exclusively in DLBCL patients with HR. We also showed, that in comparison with LR, in HR and MR groups, there is a significant decrease in the absolute number of T-cells and Tregs. The applied treatment does no normalize the number of B-cells, Tregs and apoptotic cells only in the case of HR patients.
Lymphopenia, the decreased absolute number of T cells, Tregs, and a percentage of apoptotic cells, correlates with clinical staging in DLBCL patients. The increased number of B cells and the decreased level of Tregs and apoptotic cells after treatment might predict a poor clinical outcome in patients treated with RCHOP. Thereby, flow-cytometry-based evaluation of peripheral blood lymphocytes may be useful in prognostication of newly diagnosed DLBCL patients.
The evaluation of a new group of distamycin analogues 1-6 as potential minor groove binders for the treatment of cancer were investigated. The activity of the new compounds against several ...restriction enzymes was examined. The studied compounds did not block GC-rich sequences regions of DNA but inhibited catalytic action of endonucleases in AA, AT, TT and AG restriction sites. Determination of association constants using calf thymus DNA, T4 coliphage DNA, poly(dA-dT)₂ and poly(dG-dC)₂ have confirmed that the tested compounds bind within minor groove of B-DNA. All of the compounds demonstrated activity against DNA topoisomerases II at the concentration 10 µM, but they did not inhibit activity of topoisomerase I. The studied derivatives were evaluated in human MCF-7 breast cancer cells and showed antiproliferative and cytotoxic effects in the range of 81.70 µM and 200.00 µM.
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