Human mesenchymal stem cells increase LLC metastasis and stimulate or decelerate tumor development depending on injection method and cell amount.

Mesenchymal stem cells (MSCs) being injected into the body can stimulate or decelerate carcinogenesis. Here, the direction of influence of human placenta-derived MSCs (P-MSCs) on the Lewis lung carcinoma (LLC) tumor development and metastatic potential is investigated in C57BL/6 mice depending on the injection method. After intramuscular co-inoculation of LLC and P-MSCs (LLC + P-MSCs), the growth of primary tumor and angiogenesis are slowed down compared to the control LLC on the 15th day. This is explained by the fact of a decrease in the secretion of proangiogenic factors during in vitro co-cultivation of an equal amount of LLC and P-MSCs. When P-MSCs are intravenously (i.v.) injected in the mice with developing LLC (LLC + P-MSCs(i.v.)), the tumor growth and angiogenesis are stimulated on the 15th day. A highly activated secretion of proangiogenic factors by P-MSCs in a similar in vitro model can explain this. In both the models compared to the control on the 23rd day, there is no significant difference in the tumor growth, while angiogenesis remains correspondingly decelerated or stimulated. However, in both the models, the total volume and number of lung metastases constantly increase compared to the control: it is mainly due to small-size metastases for LLC + P-MSCs(i.v.) and larger ones for LLC + P-MSCs. The increase in the rate of LLC cell dissemination after the injection of P-MSCs is explained by the disordered polyploidy and chromosomal instability, leading to an increase in migration and invasion of cancer cells. After LLC + P-MSCs co-inoculation, the tumor cell karyotype has the most complex and heterogeneous chromosomal structure. These findings indicate a bidirectional effect of P-MSCs on the growth of LLC in the early periods after injection, depending on the injection method, and, correspondingly, the number of contacting cells. However, regardless of the injection method, P-MSCs are shown to increase LLC aggressiveness related to cancer-associated angiogenesis and metastasis activation in the long term.

Anti-inflammatory effect of polyherbal composition with hepatoprotective and choleretic properties on LPS-stimulated murine macrophages.

OBJECTIVES: A polyherbal formulation with hepatoprotective and choleretic properties combining pharmacological potential of eight medicinal plants was developed in Nargiz Medical center (Republic of Azerbaijan) for the use as herbal tea. To explore the effect of polyherbal composition on the metabolism of LPS-stimulated macrophages in vitro. METHODS: The qualitative and quantitative phytochemical analysis was conducted using specific color reactions and gas chromatography-mass spectrometry (GC-MS). Nitric oxide (NO) assay was determined using the Griess reaction. Reactive oxygen species (ROS) generation was measured using ROS-sensitive fluorescence indicator, H2DCFDA, by flow cytometry. Arginase activity was examined by colorimetric method. RESULTS: The studied polyherbal formulation exerted anti-inflammatory activity in LPS-stimulated macrophages which was evidenced by dose-dependent decrease of ROS generation and by shift of arginine metabolism to the increase of arginase activity and decrease of NO release. CONCLUSIONS: Our findings suggest that the herbal tea reduces macrophage inflammatory activity, that provide an important rationale to utilize it for the attenuation of chronic inflammation typical of hepatobiliary disorders.

Towards in vivo photomediated delivery of anticancer peptides: Insights from pharmacokinetic and -dynamic data.

An in vivo study of a photoswitchable cytotoxic peptide LMB040 has been undertaken on a chemically induced hepatocellular carcinoma model in immunocompetent rats. We analysed the pharmacokinetic profile of the less toxic photoform ("ring-closed" dithienylethene) of the compound in tumors, plasma, and healthy liver. Accordingly, the peptide can reach a tumor concentration sufficiently high to exert a cytotoxic effect upon photoconversion into the more active ("ring-open") photoform. Tissue morphology, histology, redox state of the liver, and hepatic biochemical parameters in blood serum were analysed upon treatment with (i) the less active photoform, (ii) the in vivo light-activated alternative photoform, and (iii) compared with a reference chemotherapeutic 5-fluorouracil. We found that application of the less toxic form followed by a delayed in vivo photoconversion into the more toxic ring-open form of LMB040 led to a higher overall survival of the animals, and signs of enhanced immune response were observed compared to the untreated animals.

Near-infrared light reduces ß-amyloid-stimulated microglial toxicity and enhances survival of neurons: mechanisms of light therapy for Alzheimer's disease.

BACKGROUND: Low-intensity light can decelerate neurodegenerative disease progression and reduce amyloid ß (Aß) levels in the cortex, though the cellular and molecular mechanisms by which photobiomodulation (PBM) protects against neurodegeneration are still in the early stages. Microglia cells play a key role in the pathology of Alzheimer's disease by causing chronic inflammation. We present new results concerning the PBM of both oxidative stress and microglia metabolism associated with the activation of metabolic processes by 808 nm near-infrared light. METHODS: The studies were carried out using healthy male mice to obtain the microglial cell suspension from the hippocampus. Oligomeric ß-amyloid (1-42) was prepared and used to treat microglia cells. Light irradiation of cells was performed using diode lasers emitting at 808 nm (30 mW/cm2 for 5 min, resulting in a dose of 10 J/cm2). Mitochondrial membrane potential, ROS level studies, cell viability, apoptosis, and necrosis assays were performed using epifluorescence microscopy. Phagocytosis, nitric oxide and H2O2 production, arginase, and glucose 6-phosphate dehydrogenase activities were measured using standard assays. Cytokines, glucose, lactate, and ATP were measurements with ELISA. As our data were normally distributed, two-way ANOVA test was used. RESULTS: The light induces a metabolic shift from glycolysis to mitochondrial activity in pro-inflammatory microglia affected by oligomeric Aß. Thereby, the level of anti-inflammatory microglia increases. This process is accompanied by a decrease in pro-inflammatory cytokines and an activation of phagocytosis. Light exposure decreases the Aß-induced activity of glucose-6-phosphate dehydrogenase, an enzyme that regulates the rate of the pentose phosphate pathway, which activates nicotinamide adenine dinucleotide phosphate oxidases to further produce ROS. During co-cultivation of neurons with microglia, light prevents the death of neurons, which is caused by ROS produced by Aß-altered microglia. CONCLUSIONS: These original data clarify reasons for how PBM protects against neurodegeneration and support the use of light for therapeutic research in the treatment of Alzheimer's disease.

Red and near infrared light-stimulated angiogenesis mediated via Ca2+ influx, VEGF production and NO synthesis in endothelial cells in macrophage or malignant environments.

Irradiation with red or near-infrared (NIR) light in low level light therapy (LLLT) is found to stimulate cellular processes and bioenergetics, resulting in enhanced wound healing, pain control, neurodegenerative diseases treatment, etc. During light irradiation of tissues and organs, different cells are affected, though the connection between photostimulation of cells and their environmental conditions remains poorly understood. In this report, red/NIR light-stimulated angiogenesis is investigated using endothelial cells in vitro, with a focus on the capillary-like structure (CLS) formation and the respective biochemical processes in cells under conditions proximate to a healthy or malignant environment, which strongly defines angiogenesis. To model environmental conditions for endotheliocytes in vitro, the cell culture environment was supplemented by an augmented conditioned medium from macrophages or cancer cells. The biochemical processes in endothelial cell cultures were investigated with and without irradiation by red (650 nm) and near-infrared (808 nm) laser diodes and under normoxia or hypoxia conditions. A light-stimulated angiogenesis has been found, with a more efficient stimulation by 650 nm light compared to 808 nm light. It was shown that the irradiation with light promoted extracellular Ca2+ influx, fostered cell cycle progression, proliferation and NO generation in endothelial cells, and caused an increase in vascular endothelial growth factor (VEGF) production by endothelial cells and M2 macrophages under hypoxia conditions. The activation of VEGF production by macrophages was found to be associated with an increase in the number of M2 macrophages after light irradiation under hypoxia conditions. Thus, a new pathway of an activation of the endothelial cell metabolism, which is related with the extracellular Ca2+ influx after light irradiation, has been revealed. STATEMENT OF SIGNIFICANCE: Red/NIR light-stimulated angiogenesis has been studied using endothelial cells in vitro, with focus on CLS formation and the respective biochemical processes in cell models proximate to a healthy or malignant environment. A light-stimulated angiogenesis has been found, stimulated via extracellular Ca2+ influx, cell cycle progression, proliferation and NO generation, VEGF production increase by endothelial cells under hypoxia conditions.

Immunohistochemical and genetic prognostic factors of neoadjuvant chemoradiotherapy efficаcy in personalized treatment of local advanced rectal cancer.

OBJECTIVE: The aim: The aim of our study was to define the factors that can robustly predict a response to neoadjuvant chemoradiotherapy (NCRT) in patients with local advanced rectal cancer (LARC) and prognosis factors of progression free survival (PFS) using molecular (8-oxodGu), immunohystochemical (Ki-67) and genetic (GSTP1 and MTHFR genes polymorphism) markers. PATIENTS AND METHODS: Materials and methods: GSTP1 and MTHFR polymorphisms were studied by real-time PCR on tumour material from 110 patients with LARC. Ki-67 protein expression was assessed using rabbit monoclonal antibodies to Ki-67 (Dako, Denmark) on EnVisionTM FLEX detection system (Dako, Denmark). 8-oxodGu level in eluate was measured by spectrophotometry. RESULTS: Results: Patients from both groups showed significant pathomorphological response to NCRT. It is robust correlation between 8-oxodGu levels in patients' blood and their response to CRT (mrTRG scale) in MG was determined. Oxaliplatin-containing chemotherapy promotes statistically significant decrease of 8-oxodGu levels. With the decrease of Ki-67 protein expression level the probability of tumour relapse increases. It is determined that critical value of Ki-67 protein expression level makes less than 27 and tumour relapse probability in this case makes 50%. Tumour relapse risk in patients with GSTP1 and MTHFR polymorphism is 12.3 and 16.3 times higher than in patients who do not carry such polymorphism, respectively. Combination of GSTP1, МTНFR polymorphisms and Кі-67 protein expression factors determines prognostic probability of tumour relapse within 51-99%. CONCLUSION: Conclusions: 8-oxodGu level can serve as independent prognostic factor of NCRT efficacy in patients with LARC. Combination of GSTP1, МTНFR genes polymorphism with Кі-67 protein expression decrease enables monitoring and robust prognosis of LARC relapse.

A Ukrainian multicenter prospective study of the value of PET/ CT prognostic role in primary patients with Hodgkins lymphoma in a real-life cohort.

INTRODUCTION: In recent years, the positron emission tomography combined with computed tomography (PET/CT) has changed and the treatment approaches in Hodgkins lymphoma (HL) patients have entirely improved. The main idea in several studies is the use of PET/CT and the International Prognostic Score (IPS) protocols in identification of patients within a high-risk group and potential early relapse/refractory disease. MATERIALS AND METHODS: This study was based on PET/CT evaluation and treatment strategies of patients from eight Centers of Hematology in Ukraine. The patients included were newly dia-gnosed with HL and were aged 67 years or younger. They received a treatment with ABVD or BEACOPP-14/esc or “switched-regimens” (ABVD + BEACOPP-esc/14, BEACOPP-esc/14 + ABVD). The primary endpoints were to assess a correlation between PET/CT findings at the time of dia-gnosis, response to the therapy and clinical outcome (relapse/death) for patients with early and advanced stages of HL. The secondary endpoints were to evaluate the relationship between IPS and PET/CT findings. RESULTS: The study group included 106 patients. The overall response rate (ORR) was 90.5%. The ORR for patients with stages I-II was 96.5% (55/57) vs. 91% (41/45) for stage III-IV patients. In total, the disease progression occurred in 58.3% (7/12) of PET2+ patients and in 13.3% (12/90) of PET2 patients (P < 0.05). No significant difference was found between the event free survival (EFS) rate and IPS for patients with PET2+ vs. PET2, (log-rank test; P = 0.4). The PET3 status was found in 88.8% (79/89) of the study group patients and 1.2% (10/89) had a PET3+ status (P < 0.05). Using the Cox regression, we confirmed a significant correlation between EFS with PET3 Deauville scale (DS) and IPS. Patients with DS 1-2, DS 3 and DS 4-5 had a 1-year event-free survival of 94.4%, 100% and 33%, respectively (HR 0.56; 95% CI 1.07-2.8; P < 0.02). Our multivariable analysis showed no statistically significant correlation between PET2+ and PET3+ status and extranodal involvement or large tumor burden. CONCLUSION: The results of using PET/CT in patients with primary HL demonstrated a high prognostic value of PET at the end of the treatment. In addition, we confirmed the predictive role of IPS prognostic model in the treatment outcome depending on PET status.

Effect of cryopreservation on viability and growth efficiency of stromal-epithelial cells derived from neonatal human thymus.

The thymus is the major site of T lymphocyte generation and so is critical for a functional adaptive immune system. Since, thymectomy is a component of neonatal surgery for congenital heart diseases, it provides great potential for collection and storage of thymic tissue for autologous transplantation. However, specific investigation into the optimum parameters for thymic tissue cryopreservation have not been conducted. In this research, we evaluated the effect of different cryoprotective media compositions, which included penetrating (Me2SO, glycerol) and non-penetrating (dextran-40, sucrose, hydroxyethyl starch) components, on the viability and functionality of frozen-thawed human thymic samples to select an optimal cryoprotective medium suitable for long-term storage of thymic tissue and a stromal-epithelial enriched population. Our primary focus was on receiving, low-temperature storage, culturing and evaluation of thymic tissue samples from newborns and infants with congenital heart diseases, who had undergone thymectomy as a part of standard surgical procedure. Thus, this work builds the platform for autologous clinical intervention into the thymus-deficient patients with congenital heart diseases. From our data, we conclude that although there were no significant differences in efficiency of tested cryoprotective media compositions, the combination of Me2SO and dextran-40 compounds was the most suitable for long-term storage both thymic cell suspensions and thymic fragments based on the viability of CD326+ epithelial cells and stromal-epithelial cell monolayer formation.