Proapoptotic and antimigration properties of osthole in combination with LY294002 against human glioma cells.

Anaplastic astrocytoma and glioblastoma multiforme are infiltrating and vascularized gliomas with a high degree of chemoresistance and metastasis. Our previous studies have shown that osthole may be of great importance in the treatment of gliomas. Therefore, in this work, for the first time, coumarin was used in combination with LY294002-an inhibitor of the PI3K-Akt/PKB-mTOR pathway, which is overly active in gliomas. MOGGCCM and T98G cells were incubated with osthole and LY294002, alone and in combination. Staining with specific fluorochromes was used to visualize cell death and the scratch test to assess the migration. The level of proteins was estimated by immunoblotting. Forming protrusions were visualized by SEM, and immunocytochemistry was used to determine the localization of proteins. Additionally, the expression of Bcl-2, beclin 1 and Raf kinase was silenced using specific siRNA. The obtained results showed that osthole in combination with LY294092 effectively inhibited the migration of glioma cells by reducing the level of metaloproteinases and Rho family proteins, as well as decreasing the level of N-cadherin. In addition, the combination of compounds induced apoptosis. New combination of compounds shows a high pro-apoptotic potential and also inhibits the migration of gliomas cells.

Furanocoumarins as Enhancers of Antitumor Potential of Sorafenib and LY294002 toward Human Glioma Cells In Vitro.

Furanocoumarins are naturally occurring compounds in the plant world, characterized by low molecular weight, simple chemical structure, and high solubility in most organic solvents. Additionally, they have a broad spectrum of activity, and their properties depend on the location and type of attached substituents. Therefore, the aim of our study was to investigate the anticancer activity of furanocoumarins (imperatorin, isoimperatorin, bergapten, and xanthotoxin) in relation to human glioblastoma multiforme (T98G) and anaplastic astrocytoma (MOGGCCM) cell lines. The tested compounds were used for the first time in combination with LY294002 (PI3K inhibitor) and sorafenib (Raf inhibitor). Apoptosis, autophagy, and necrosis were identified microscopically after straining with Hoechst 33342, acridine orange, and propidium iodide, respectively. The levels of caspase 3 and Beclin 1 were estimated by immunoblotting and for the blocking of Raf and PI3K kinases, the transfection with specific siRNA was used. The scratch test was used to assess the migration potential of glioma cells. Our studies showed that the anticancer activity of furanocoumarins strictly depended on the presence, type, and location of substituents. The obtained results suggest that achieving higher pro-apoptotic activity is determined by the presence of an isoprenyl moiety at the C8 position of the coumarin skeleton. In both anaplastic astrocytoma and glioblastoma, imperatorin was the most effective in induction apoptosis. Furthermore, the usage of imperatorin, alone and in combination with sorafenib or LY294002, decreased the migratory potential of MOGGCCM and T98G cells.

The Role of Bcl-2 and Beclin-1 Complex in "Switching" between Apoptosis and Autophagy in Human Glioma Cells upon LY294002 and Sorafenib Treatment.

BACKGROUND: Gliomas are the most malignant tumors of the central nervous system. One of the factors in their high drug resistance is avoiding programmed death (PCD) induction. This is related to the overexpression of intracellular survival pathways: PI3K-Akt/PKB-mTOR and Ras-Raf-MEK-ERK. Apoptosis and autophagy are co-existing processes due to the interactions between Bcl-2 and beclin-1 proteins. Their complex may be a molecular "toggle-switch" between PCD types. The aim of this research was to investigate the role of Bcl-2:beclin-1 complex in glioma cell elimination through the combined action of LY294002 and sorafenib. METHODS: Drug cytotoxicity was estimated with an MTT test. The type of cell death was evaluated using variant microscopy techniques (fluorochrome staining, immunocytochemistry, and transmission electron microscopy), as well as the Bcl-2:beclin-1 complex formation and protein localization. Molecular analysis of PCD indicators was conducted through immunoblotting, immunoprecipitation, and ELISA testing. SiRNA was used to block Bcl-2 and beclin-1 expression. RESULTS: The results showed the inhibitors used in simultaneous application resulted in Bcl-2:beclin-1 complex formation and apoptosis becoming dominant. This was accompanied by changes in the location of the tested proteins. CONCLUSIONS: "Switching" between apoptosis and autophagy using PI3K and Raf inhibitors with Bcl-2:beclin-1 complex formation opens new therapeutic perspectives against gliomas.

Heterogeneous Cellular Response of Primary and Metastatic Human Gastric Adenocarcinoma Cell Lines to Magnoflorine and Its Additive Interaction with Docetaxel.

Gastric cancer is the most common cancer and remains the leading cause of cancer death worldwide. In this study, the anticancer action of magnoflorine isolated via counter-current chromatography from the methanolic extract of Berberis vulgaris root against gastric cancer in models of primary ACC-201 and AGS and metastatic MKN-74 and NCI-N87 cell lines was analyzed. Cell viability and proliferation were tested through the use of MTT and BrdU tests, respectively. Cell cycle progression and apoptosis were evaluated using flow cytometry. The interaction of magnoflorine and docetaxel has been examined through isobolographic analysis. Moreover, potential toxicity was verified in zebrafish in an in vivo model. Gastric cancer cell lines revealed different responses to magnoflorine treatment with regard to viability/proliferation, apoptosis induction and cell cycle inhibition without any undesirable changes in the development of larval zebrafish at the tested concentrations. What is more, magnoflorine in combination with docetaxel produced an additive pharmacological interaction in all studied gastric cancer cell lines, which may suggest a complementary mechanism of action of both compounds. Taken together, these findings provide a foundation for the possibility of magnoflorine as a potential therapeutic approach for gastric cancer and merits further investigation, which may pave the way for clinical uses of magnoflorine.

Synergism of antiproliferative effects of young green barley and chlorella water extracts against human breast cancer cells.

INTRODUCTION AND OBJECTIVE: Breast cancer is the most common type of tumour in terms of incidence and mortality among women. In the light of recent data that has revealed the beneficial impact of increasing plant-based food consumption on the risk of breast cancer, the use of young green barley and chlorella, the chemopreventive properties of which have been previously reported, seems to be a reasonable therapeutic strategy in this type of cancer. Nevertheless, there are only a few scientific reports focused on the influence of the mentioned products on breast cancer development; thus, the aim of the study was to enrich knowledge resources in this area. MATERIAL AND METHODS: The chemopreventive effect of water extracts of chlorella (CH) and young green barley (YGB) and their mixture (MIX) was investigated in human breast adenocarcinoma T47D cells and human skin fibroblasts HSF by LDH, MTT and BrdU assays. Changes in cell morphology in response to tested extracts were examined in light microscopy. RESULTS: Tested extracts were not toxic against HSF and did not affect their proliferation and morphology. Simultaneously, extracts increased the permeability of T47D cell membranes and inhibited their proliferation. Microscopic observation confirmed the results of biochemical assays and additionally suggested necrosis induction in T47D cells in response to tested compounds. Obtained results demonstrated that MIX induced stronger beneficial changes than their components. CONCLUSIONS: The study revealed the chemopreventive properties of the investigated green food products against breast cancer cells, without any side-effects in human skin fibroblasts. The beneficial properties discovered of the tested extracts on cancer cells enhanced by their concomitant administration, and in the case of antiproliferative effects YGB and CH, revealed synergism of action.

(1→3)-α-d-Glucooligosaccharides Increase the Killing Capacity of NK Cells against Selected Human Colon Cancer Cells.

Despite the progress of medicine, colorectal cancer has occupied one of the highest positions in the rankings of cancer morbidity and mortality for many years. Thus, alternative methods of its treatment are sought. One of the newer therapeutic strategies is immunotherapy based on NK cells (natural killers), which are the body's first line of defense against cancer. The aim of the study was to verify the possibility of using (1→3)-α-d-glucooligosaccharides (GOSs) obtained via acid hydrolysis of (1→3)-α-d-glucan from the fruiting body of Laetiporus sulphureus to improve the anticancer effect of NK-92 cells, with proven clinical utility, against selected human colon adenocarcinoma cell lines LS180 and HT-29. The study revealed that the investigated oligosaccharides significantly enhanced the ability of NK-92 cells to eliminate the examined colon cancer cells, mostly by an increase in their cytotoxic activity. The most significant effect was observed in LS180 and HT-29 cells exposed to a two-times higher quantity of NK cells activated by 500 µg/mL GOS, wherein NK-92 killing properties increased for 20.5% (p < 0.001) and 24.8% (p < 0.001), respectively. The beneficial impact of (1→3)-α-d-glucooligosaccharides on the anticancer properties of NK-92 suggests their use in colon cancer immunotherapy as adjuvants; however, the obtained data require further investigation and confirmation.

Binge-like mephedrone treatment induces memory impairment concomitant with brain kynurenic acid reduction in mice.

While mephedrone (4-methylmethcathinone), a synthetic cathinone derivative, is widely abused by adolescents and young adults, the knowledge about its long-term effects on memory processes is limited. Kynurenic acid (KYNA) is a neuroactive metabolite of the kynurenine pathway of tryptophan degradation. KYNA is considered an important endogenous modulator influencing physiological and pathological processes, including learning and memory processes. The aim of this study was to determine whether (A) binge-like mephedrone administration (10.0 and 30.0 mg/kg, intraperitoneally, in 4 doses separated by 2 h) induces memory impairments, assessed 2, 8 and 15 days after mephedrone cessation in the passive avoidance test in mice, and whether (B) KYNA is involved in these memory processes. To clarify the role of KYNA in the mephedrone effects, its production in the murine brain in vivo, and in cortical slices in vitro, as well as the activities of kynurenine aminotransferases (KATs) I and II were assessed. Furthermore, cell line experiments were conducted to investigate the effects of mephedrone on normal human brain cells. Our results showed memory impairments 8 and 15 days after binge-like mephedrone administration. At the same time, reduction in the KYNA level in the murine brain was noted. In vitro studies showed no effect of mephedrone on the production of KYNA in cortical slices or on the activity of the KAT I and II enzymes. Finally, exposure of normal cells to mephedrone in vitro resulted in a modest reduction of cell viability and proliferation.

Immunomodulatory Properties of Polysaccharide-Rich Young Green Barley (Hordeum vulgare) Extract and Its Structural Characterization.

Young green barley (YGB) water extract has revealed a beneficial impact on natural killer (NK) cells' ability to recognize and eliminate human colon cancer cells, without any side effects for normal colon epithelial cells. The direct anticancer effect of the tested compounds has been also shown. The mixture of oligosaccharides found in this extract was characterized by chemical analyses and via FT-IR spectroscopy and MALDI-TOF MS techniques. The YGB preparation contained 26.9% of proteins and 64.2% of sugars, mostly glucose (54.7%) and fructose (42.7%), with a small amount of mannose (2.6%) and galactose (less than 0.5%). Mass spectrometry analysis of YGB has shown that fructose oligomers contained from 3 to 19 sugar units. The number of fructans was estimated to be about 10.2% of the dry weight basis of YGB. The presented results suggest the beneficial effect of the consumption of preparations based on young barley on the human body, in the field of colon cancer prevention.

Impact of phytochemicals and plant extracts on viability and proliferation of NK cell line NK-92 - a closer look at immunomodulatory properties of goji berries extract in human colon cancer cells.

INTRODUCTION: Due to the fact that lymphocytes NK (natural killer cells) are the first line of defence of the body against cancer, one of the goals of modern immunotherapy is the enhancement of their natural activities for the effective recognition, detection, and elimination of cancer cells. OBJECTIVE: The aim of the study was to evaluate the influence of selected phytochemicals (curcumin and resveratrol) and plant extracts (chlorella and goji berries) on NK cells viability and proliferation, as well as cytotoxic activity against colon cancer - one of the most common cancer worldwide. MATERIAL AND METHODS: The impact of phytochemicals, viability and proliferation of plant extracts on NK cells was examined in NK-92 cells using both LDH and MTT assays. The immunomodulatory properties of selected compounds were tested against human colon cancer cell line LS180 using the MTT test. RESULTS: Extracts of chlorella and goji berries significantly increased NK cell proliferation, while curcumin and resveratrol did not affect this process. Curcumin, as well as extracts of chlorella and goji berries, did not impact NK viability, while resveratrol significantly increased it. LDH test revealed the cytotoxic effect of chlorella extract and curcumin in NK-92 cell cultures. On the contrary, goji berries extract significantly decreased LDH level, while resveratrol did not affect the integrity of NK cell membranes. Studies conducted in co-cultures NK cells, also directly eliminated colon cancer cells. CONCLUSIONS: Performed studies revealed immunomodulatory properties of goji berries extract, which improved viability and proliferation of NK cells, and above all, significantly increased their ability to recognize and eliminate colon cancer cells.

Lensoside Aß as an Adjuvant to the Anti-Glioma Potential of Sorafenib.

AIM: The anti-glioma effect of lensoside Aß alone and in combination with sorafenib (pro-survival Raf kinase inhibitor) was evaluated for the first time in terms of programmed cell death induction in anaplastic astrocytoma and glioblastoma multiforme cell lines as an experimental model. Apoptosis, autophagy, and necrosis were identified microscopically (fluorescence and scanning microscopes) and confirmed by flow cytometry (mitochondrial membrane potential MMP and cell death). The expression of apoptotic (caspase 3) and autophagic markers (beclin 1) as well as Raf kinase were estimated by immunoblotting. The FTIR method was used to determine the interaction of the studied drugs with lipid and protein groups within cells, while the modes of drug action within the cells were assessed with the FLIM technique. RESULTS: Lensoside Aß itself does not exhibit anti-glioma activity but significantly enhances the anti-cancer potential of sorafenib, initiating mainly apoptosis of up to 90% of cells. It was correlated with an increased level of active caspase 3, a reduced MMP value, and a lower level of Raf kinase. The interaction with membrane structures led to morphological changes typical of programmed death. CONCLUSIONS: Our results indicate that lensoside Aß plays an important role as an adjuvant in chemotherapy with sorafenib and may be a potential candidate in anti-glioma combination therapy.

Involvement of PI3K Pathway in Glioma Cell Resistance to Temozolomide Treatment.

The aim of the study was to investigate the anticancer potential of LY294002 (PI3K inhibitor) and temozolomide using glioblastoma multiforme (T98G) and anaplastic astrocytoma (MOGGCCM) cells. Apoptosis, autophagy, necrosis, and granules in the cytoplasm were identified microscopically (fluorescence and electron microscopes). The mitochondrial membrane potential was studied by flow cytometry. The activity of caspases 3, 8, and 9 and Akt was evaluated fluorometrically, while the expression of Beclin 1, PI3K, Akt, mTOR, caspase 12, and Hsp27 was determined by immunoblotting. SiRNA was used to block Hsp27 and PI3K expression. Cell migration and localization of Hsp27 were tested with the wound healing assay and immunocytochemistry, respectively. LY294002 effectively diminished the migratory potential and increased programmed death of T98G and MOGGCCM. Autophagy was dominant in MOGGCCM, while apoptosis was dominant in T98G. LY294002 with temozolomide did not potentiate cell death but redirected autophagy toward apoptosis, which was correlated with ER stress. A similar effect was observed after blocking PI3K expression with siRNA. Transfection with Hsp27 siRNA significantly increased apoptosis related to ER stress. Our results indicate that inhibition of the PI3K/Akt/mTOR pathway sensitizes glioma cells to apoptosis upon temozolomide treatment, which was correlated with ER stress. Hsp27 increases the resistance of glioma cells to cell death upon temozolomide treatment.

Promising Potential of Crude Polysaccharides from Sparassis crispa against Colon Cancer: An In Vitro Study.

The aim of the present study was to evaluate in vitro the beneficial potential of crude polysaccharides from S. crispa (CPS) in one of the most common cancer types-colon cancer. The determination of the chemical composition of CPS has revealed that it contains mostly carbohydrates, while proteins or phenolics are present only in trace amounts. 1H NMR and GC-MS methods were used for the structural analysis of CPS. Biological activity including anticancer, anti-inflammatory and antioxidant properties of CPS was investigated. CPS was found to be non-toxic to normal human colon epithelial CCD841 CoN cells. Simultaneously, they destroyed membrane integrity as well as inhibited the proliferation of human colon cancer cell lines: Caco-2, LS180 and HT-29. Antioxidant activity was determined by various methods and revealed the moderate potential of CPS. The enzymatic assays revealed no influence of CPS on xanthine oxidase and the inhibition of catalase activity. Moreover, pro-inflammatory enzymes such as cyclooxygenase-2 or lipooxygenase were inhibited by CPS. Therefore, it may be suggested that S. crispa is a valuable part of the regular human diet, which may contribute to a reduction in the risk of colon cancer, and possess promising activities encouraging further studies regarding its potential use as chemopreventive and therapeutic agent in more invasive stages of this type of cancer.

Enhancement of chemopreventive properties of young green barley and chlorella extracts used together against colon cancer cells.

INTRODUCTION AND OBJECTIVE: Young green barley and chlorella are listed as a panacea for a whole range of disorders, including diabetes, cardiovascular disease, infections, and hypercholesterolaemia. The first reports have appeared presenting the anticancer properties of these products. The present study is an attempt to extend this knowledge with particular emphasis on the possibility of using young green barley, chlorella, and their combination in colon cancer chemoprevention. MATERIAL AND METHODS: Extracts of young green barley (YB) and chlorella (CH), as well as their combination MIX (YB+CH; 1:1), were examined. The influence of the extracts on viability and proliferation of human colon epithelial CCD841 CoN cells was analyzed by LDH and MTT assays. Anticancer properties of extracts were screened on human colon adenocarcinoma cell line HT-29 by MTT and BrdU assays. Changes in cells morphology in response to extracts were investigated after May-Grünwald-Giemsa staining. RESULTS: Extracts used together or separately did not affect the viability and proliferation of CCD841 CoN cells. Simultaneously, YB, CH and MIX inhibited proliferation of HT-29 cells in a dose-dependent manner. Furthermore changes in the morphology of HT-29 cells treated with YB, CH and MIX suggested necrosis induction. Performed studies also revealed that MIX induced a stronger antiproliferative effect than their individual ingredients; however, the discovered enhancement of anticancer properties was weaker than the sum of the effects of YB and CH creating MIX. CONCLUSIONS: The study demonstrates great chemopreventive properties of young green barley and chlorella water extracts and their combination in in vitro model of colon cancer. The anticancer properties of the investigated extracts were significantly enhanced after combining, without an increase of their toxicity against normal cells.

Antiglioma Potential of Coumarins Combined with Sorafenib.

Coumarins, which occur naturally in the plant kingdom, are diverse class of secondary metabolites. With their antiproliferative, chemopreventive and antiangiogenetic properties, they can be used in the treatment of cancer. Their therapeutic potential depends on the type and location of the attachment of substituents to the ring. Therefore, the aim of our study was to investigate the effect of simple coumarins (osthole, umbelliferone, esculin, and 4-hydroxycoumarin) combined with sorafenib (specific inhibitor of Raf (Rapidly Accelerated Fibrosarcoma) kinase) in programmed death induction in human glioblastoma multiforme (T98G) and anaplastic astrocytoma (MOGGCCM) cells lines. Osthole and umbelliferone were isolated from fruits: Mutellina purpurea L. and Heracleum leskowii L., respectively, while esculin and 4-hydroxycoumarin were purchased from Sigma Aldrich (St. Louis, MO, USA). Apoptosis, autophagy and necrosis were identified microscopically after straining with specific fluorochromes. The level of caspase 3, Beclin 1, PI3K (Phosphoinositide 3-kinase), and Raf kinases were estimated by immunoblotting. Transfection with specific siRNA (small interfering RNA) was used to block Bcl-2 (B-cell lymphoma 2), Raf, and PI3K expression. Cell migration was tested with the wound healing assay. The present study has shown that all the coumarins eliminated the MOGGCCM and T98G tumor cells mainly via apoptosis and, to a lesser extent, via autophagy. Osthole, which has an isoprenyl moiety, was shown to be the most effective compound. Sorafenib did not change the proapoptotic activity of this coumarin; however, it reduced the level of autophagy. At the molecular level, the induction of apoptosis was associated with a decrease in the expression of PI3K and Raf kinases, whereas an increase in the level of Beclin 1 was observed in the case of autophagy. Inhibition of the expression of this protein by specific siRNA eliminated autophagy. Moreover, the blocking of the expression of Bcl-2 and PI3K significantly increased the level of apoptosis. Osthole and sorafenib successfully inhibited the migration of the MOGGCCM and T98G cells.

Coumarins modulate the anti-glioma properties of temozolomide.

In the recent years, coumarin bioactive compounds have been identified to posess anticancer properties. Therefore, the aim of the present study was to investigate for the first time the efficacy of osthole, umbelliferone, esculin, and 4-hydroxycoumarin, alone and in combination with Temozolomide, in the elimination of deadly brain tumors, anaplastic astrocytoma (AA) and glioblastoma multiforme (GBM) cells via programmed death. Our results indicated that osthole, umbelliferone, esculin, and 4-hydroxycoumarin initiated mainly apoptosis in the T98G and MOGGCCM cells. Osthole was the most effective. It also initiated autophagy in a small percentage of the cell population. The co-incubation with Temozolomide did not increase the pro-apoptotic potential of natural compounds but decreased the level of autophagy in the T98G cells. Apoptosis was associated with reduced mitochondrial membrane potential, activation of caspase 3, inhibition of Bcl-2 expression and the presence of a Bcl-2/Beclin 1. Blocking of Bcl-2 expression resulted in promotion of apoptosis, but not autophagy, in the MOGGCCM and T98G lines. It also sensitized astrocytoma cells, but not GBM, to the combined osthole and TMZ treatment, which was correlated with a reduced level of Beclin 1 and increased expression of caspase 3. Osthole and TMZ, alone and in combination, inhibited the migratory phenotype of the GBM and AA cells. In summary, our results indicated that osthole effectively eliminated glioma cells via apoptosis, what was correlated with Bcl-2/Beclin 1 complex formation. Considering the anti-migratory effect, osthole and Temozolomide display antiglioma potential but it needs further extensive studies.

Pro-apoptotic action of protein-carbohydrate fraction isolated from coelomic fluid of the earthworm Dendrobaena veneta against human colon adenocarcinoma cells.

Earthworm coelomic fluid (CF) is known as a rich source of various bioactive compounds with promising anticancer features. However, it has been demonstrated that CF affects functionality of both, cancer and normal cells. This non-selective activity causes a major problem for medical application of CF. In this study, we present the anticancer activity of the active protein-carbohydrate fraction (AF) isolated from thermally treated CF of earthworm Dendrobaena veneta. The in vitro effect of the AF was examined in human colon model including normal human colon epithelium (CCD 841 CoTr) and human colon adenocarcinoma (HT-29 and LS180) cell lines. We investigated the impact of AF on cell viability neutral red and lactate dehydrogenase assays, morphology May-Grünwald-Giemsa staining assay proliferation MTT tetrazolium salt and BrdU incorporation assays as well as cell cycle progression propidium iodide/RNase staining and the activity of human 20S proteasome the hydrolysis of AMC from a Suc-LLVY-AMC peptide substrate. Additionally, the influence of AF on apoptosis was examined in HT-29 cells by Annexin V/PI, Hoechst 33342 staining and active caspase-3 assays. Our investigation demonstrated that AF at the tested concentration range does not affect the viability and morphology of CCD 841 CoTr cells. Simultaneously, AF inhibits human 20S proteasome activity as well as significantly decreases mitochondrial metabolism, disturbs cell cycle and induces apoptosis via activation of procaspase-3 in HT-29 cancer cells. Obtained results demonstrate the antiproliferative and proapoptotic activity of AF that can be useful in developing therapeutic strategies to treat human colon cancer.

Pantoea agglomerans chronic exposure induces epithelial-mesenchymal transition in human lung epithelial cells and mice lungs.

Pantoea agglomerans is gram-negative bacteria widely distributed in nature. It predominates in inhalable dust from grain, herbs, and flax, and was identified as the most important cause of hypersensitivity pneumonitis (HP) in eastern Poland. To better understand the molecular mechanism of HP development studies focused on the interactions between P. agglomerans and alveolar epithelial cells as well as lung tissue with particular emphasis on the epithelial-mesenchymal transition (EMT). The studies were conducted on human normal lung epithelial NL20 cells and mice strain C57BL/6J. Cells and mice underwent chronic exposure to saline extract of P. agglomerans (SE-PA). Morphological changes were evaluated under light microscopy, the concentration of fibrosis markers was examined by the ELISA method, while the expression of genes involved in EMT was evaluated by RealTime PCR. During incubation with SE-PA epithelial cells underwent conversion and assumed fibroblast phenotype characterized by a decrease in epithelial cells markers (CDH1, CLDN1, JUP) and increase in mesenchymal cells markers (FN1, VIM, CDH2). Mice lungs collected after 14 days of SE-PA treatment revealed inflammation with marked lymphocytes infiltration. The intensified inflammatory process accompanied by increased proliferation of fibrous connective tissue was noted in mice lungs after 28 days of SE-PA exposure. Histological changes correlated with an increase of fibrosis markers (hydroxyproline, collagens), downregulation of epithelial markers (Cdh1, Cldn1, Jup, Ocln) and upregulation of myofibroblasts markers (Acta2, Cdh2, Fn1, Vim). Obtained results revealed SE-PA ability to induce EMT in human lung epithelial cells and mice lung tissue, with the scale of changes proportional to the time of treatment.

Prostate and breast cancer cells death induced by xanthohumol investigated with Fourier transform infrared spectroscopy.

Fourier Transform Infrared spectroscopy was applied to detect in vitro cell death induced in prostate (PC-3) and breast (T47D) cancer cell lines treated with xanthohumol (XN). After incubation of the cancer cells with XN, specific spectral shifts in the infrared spectra arising from selected cellular components were identified that reflected biochemical changes characteristic for apoptosis and necrosis. Detailed analysis of specific absorbance intensity ratios revealed the compositional changes in the secondary structure of proteins and membrane lipids. In this study, for the first time we examined the changes in these molecular components and linked them to deduce the involvement of molecular mechanisms in the XN-induced death of the selected cancer cells. We showed that XN concentration-dependent changes were attributed to phospholipid ester carbonyl groups, especially in the case of T47D cells, suggesting that XN acts as an inhibitor of cell proliferation. Additionally, we observed distinct changes in the region assigned to the absorption of DNA, which were correlated with a specific marker of cell death and dependent on the XN dose and the type of cancer cells. The microscopic observation and flow cytometry analysis revealed that the decrease in cancer cell viability was mainly related to the induction of necrotic cell death. Moreover, the T47D cells were slightly more sensitive to XN than the PC-3 cells. Considering the results obtained, it can be assumed that apoptosis and necrosis induced by XN may contribute to the anti-proliferative and cytotoxic properties of this flavonoid against cancer cell lines PC-3 and T47D.

Anticancer effects of sodium and potassium quercetin-5'-sulfonates through inhibition of proliferation, induction of apoptosis, and cell cycle arrest in the HT-29 human adenocarcinoma cell line.

In the present study, we compared the anticancer potential of quercetin (3,3',4',5,7-pentahydroxyflavone, I) and its sulfonic derivatives sodium/potassium quercetin-5'-sulfonates (described as II and III) against several human carcinoma cell lines. Quercetin (I) was used as a starting compound for synthesis of II and III. In this work, a modified and more efficient method of synthesizing derivatives II and III has been described. The molecular structures of the compounds were characterized in a solution and in the solid state using 1H NMR, 13C NMR, 2D NMR, and XPS spectroscopy, respectively. The stoichiometry of these complexes was determined by elemental analysis as well as thermogravimetric and X-ray fluorescence methods. The spectral data allowed complete characterization of the investigated compounds in the solution and in the solid state and unambiguous determination of the place of substitution of the sulfonic group in the phenyl ring in the C-5' position. Our in vitro studies revealed that II and III prominently reduced the viability of the HT-29 colon cancer cell line. Additionally, we observed that sulfonic derivatives decreased proliferation of colon (HT-29, LS180), lung (A549), and breast (T47D) cancer cell lines. Moreover, we detected a lower cytotoxic effect of II and III on several normal cell lines (colon epithelial CCD 841 CoTr, mouse subcutaneous connective tissue L-929, and human skin fibroblasts HSF cell lines) than that exerted by pure quercetin. The anticancer properties were especially evident in the HT-29 colon cancer cell line, where cell cycle inhibition in the G2-M phase and prominent apoptosis induced by II and III were observed. In conclusion, the sodium/potassium quercetin-5'-sulfonates prepared from quercetin showed promising anti-proliferative and pro-apoptotic activity against colon cancer cells. Therefore, we support the opinion that sodium/potassium quercetin-5'-sulfonates should be considered as promising organometallic compounds for possible clinical applications.

Branched mannans from the mushroom Cantharellus cibarius enhance the anticancer activity of natural killer cells against human cancers of lung and colon.

Several studies have shown that mushroom polysaccharides enhance the ability of natural killer (NK) cells to recognize cancer cells as foreign and thereby enhance the effectiveness of host immune defence mechanisms. Nevertheless, the use of NK cells in cancer treatment requires finding selective stimulators of their cytotoxicity without disturbing organism homeostasis. Our studies revealed that Cantharellus cibarius polysaccharides present in the CC2a fraction, mainly composed of an O-2 and O-3 branched (1→6)-linked mannan, not only beneficially influenced the viability and proliferation of the human natural killer cells NK92 but also enhanced their anticancer properties against the human lung and colon cancer cells A549 and LS180, and at the same time did not affect the human lung and colon epithelial cells NL20 and CCD841 CoN. Furthermore, the CC2a fraction used alone was also nontoxic to the normal epithelium, while it inhibited the viability of these cancer cells. Nevertheless, the therapeutic potential of NK92 cells was greatly enhanced after coincubation with these polysaccharides and the observed effect was dependent on the CC2a concentrations. The beneficial effect of CC2a on NK92 cells was associated with stimulation of p38 and Erk expression as well as induction of the transcription factor CREB. The discovered beneficial impact of the CC2a fraction on NK92 cells suggested the therapeutic use of the investigated compound especially as an adjuvant. Furthermore, taking into account the abundance of these water soluble mannans in C. cibarius, the results also suggest that an increase in the intake of C. cibarius may promote innate immunity response against cancer through the enhancement of NK cell activity.