Cep120 is essential for kidney stromal progenitor cell growth and differentiation.

Mutations in genes that disrupt centrosome structure or function can cause congenital kidney developmental defects and lead to fibrocystic pathologies. Yet, it is unclear how defective centrosome biogenesis impacts renal progenitor cell physiology. Here, we examined the consequences of impaired centrosome duplication on kidney stromal progenitor cell growth, differentiation, and fate. Conditional deletion of the ciliopathy gene Cep120, which is essential for centrosome duplication, in the stromal mesenchyme resulted in reduced abundance of interstitial lineages including pericytes, fibroblasts and mesangial cells. These phenotypes were caused by a combination of delayed mitosis, activation of the mitotic surveillance pathway leading to apoptosis, and changes in both Wnt and Hedgehog signaling that are key for differentiation of stromal cells. Cep120 ablation resulted in small hypoplastic kidneys with medullary atrophy and delayed nephron maturation. Finally, Cep120 and centrosome loss in the interstitium sensitized kidneys of adult mice, causing rapid fibrosis after renal injury via enhanced TGF-ß/Smad3-Gli2 signaling. Our study defines the cellular and developmental defects caused by loss of Cep120 and aberrant centrosome biogenesis in the embryonic kidney stroma.

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.

Effect of Tryptophan-Derived AhR Ligands, Kynurenine, Kynurenic Acid and FICZ, on Proliferation, Cell Cycle Regulation and Cell Death of Melanoma Cells-In Vitro Studies.

Tryptophan metabolites: kynurenine (KYN), kynurenic acid (KYNA) and 6-formylindolo[3,2-b]carbazole (FICZ) are considered aryl hydrocarbon receptor (AhR) ligands. AhR is mainly expressed in barrier tissues, including skin, and is involved in various physiological and pathological processes in skin. We studied the effect of KYN, KYNA and FICZ on melanocyte and melanoma A375 and RPMI7951 cell toxicity, proliferation and cell death. KYN and FICZ inhibited DNA synthesis in both melanoma cell lines, but RPMI7951 cells were more resistant to pharmacological treatment. Tested compounds were toxic to melanoma cells but not to normal human adult melanocytes. Changes in the protein level of cyclin D1, CDK4 and retinoblastoma tumor suppressor protein (Rb) phosphorylation revealed different mechanisms of action of individual AhR ligands. Importantly, all tryptophan metabolites induced necrosis, but only KYNA and FICZ promoted apoptosis in melanoma A375 cells. This effect was not observed in RPMI7951 cells. KYN, KYNA and FICZ in higher concentrations inhibited the protein level of AhR but did not affect the gene expression. To conclude, despite belonging to the group of AhR ligands, KYN, KYNA and FICZ exerted different effects on proliferation, toxicity and induction of cell death in melanoma cells in vitro.

A Tryptophan Metabolite, 8-Hydroxyquinaldic Acid, Exerts Antiproliferative and Anti-Migratory Effects on Colorectal Cancer Cells.

8-Hydroxyquinaldic acid, the end-metabolite of tryptophan, is well-known metal chelator; however, its role in humans, especially in cancer promotion and progression, has not been fully revealed. Importantly, 8-hydroxyquinaldic acid is the analog of kynurenic acid with evidenced antiproliferative activity towards various cancer cells. In this study, we revealed that 8-hydroxyquinaldic acid inhibited not only proliferation and mitochondrial activity in colon cancer HT-29 and LS-180 cells, but it also decreased DNA synthesis up to 90.9% for HT-29 cells and 76.1% for LS-180 cells. 8-Hydroxyquinaldic acid induced changes in protein expression of cell cycle regulators (CDK4, CDK6, cyclin D1, cyclin E) and CDKs inhibitors (p21 Waf1/Cip1, p27 Kip1), but the effect was dependent on the tested cell line. Moreover, 8-hydroxyquinaldic acid inhibited migration of colon cancer HT-29 and LS-180 cells and increased the expression of ß-catenin and E-cadherin. Importantly, antiproliferative and anti-migratory concentrations of 8-hydroxyquinaldic acid were non-toxic in vitro and in vivo. We reported for the first time antiproliferative and anti-migratory activity of 8-hydroxyquinaldic acid against colon cancer HT-29 and LS-180 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.

Kynurenic Acid Induces Impairment of Oligodendrocyte Viability: On the Role of Glutamatergic Mechanisms.

Kynurenic acid (KYNA) is an end stage product of tryptophan metabolism with a variety of functions in the human body, both in the central nervous system (CNS) and in other organs. Although its activity in the human brain has been widely studied and effects on neural cells were emphasized, the effect of KYNA on oligodendroglial cells remains unknown. Present study aims at describing the activity of high concentration of KYNA in OLN-93 cells. The inhibition of OLN-93 oligodendrocytes viability by KYNA in a medium with reduced serum concentration has been demonstrated. Although decreased metabolic activity of KYNA treated OLN-93 cells was shown, the cells proliferation was not altered. KYNA treatment did not alter morphology as well as expression level of cell cycle and proliferation regulating proteins. Furthermore, glutamate receptor antagonists and agonists did not alter the inhibitory effect of KYNA on viability of OLN-93 oligodendrocytes. This study contributes to the elucidation of effects of KYNA on oligodendrocytes in vitro, yet further analyses are necessary to explain the mechanisms behind the damage and loss of myelin sheaths.

New derivative of 2-(2,4-dihydroxyphenyl)thieno-1,3-thiazin-4-one (BChTT) elicits antiproliferative effect via p38-mediated cell cycle arrest in cancer cells.

2-(2,4-Dihydroxyphenyl)thieno-1,3-thiazin-4-ones are a group of new compounds with potential anticancer activity. This type of derivatives was poorly investigated in the area of synthesis and biological activities. In the present study the antiproliferative action of the most active derivative BChTT was described. The aim of biological evaluation was to investigate the ability of the compound to inhibit cancer cell proliferation and identify mechanism involved in its action on the molecular level. BChTT inhibited the proliferation of lung cancer A549, colon cancer HT-29 and glioma C6 cells in the concentration-dependent manner. It was not toxic to normal cells including skin fibroblasts, hepatocytes and oligodendrocytes in the antiproliferative concentrations. BChTT decreased the DNA synthesis in the treated cancer cells and induced cell cycle arrest in the G0/G1 phase. Moreover, the ability of the compound to activate p38 kinase and decrease cyclin D1 expression was estimated. Participation of p38 kinase in the antiproliferative action of the compound was confirmed by the analysis of BChTT activity in the cells with the p38 silenced gene. The obtained results may suggest the ability of the tested derivative to inhibit cancer cells proliferation by induction of p38-mediated cyclin D1 downregulation.

Synthesis of 2-(2,4-dihydroxyphenyl)thieno-1,3-thiazin-4-ones, their lipophilicity and anticancer activity in vitro.

A new one-step synthesis of novel biologically active 2-substituted 2,4-dihydroxyphenyl-4[Formula: see text]-thieno[3,2-[Formula: see text]][1,3]thiazin-4-ones and 4[Formula: see text]-thieno[2,3-[Formula: see text]][1,3]thiazin-4-ones has been elaborated and described. The compounds were prepared by the reaction of aryl-modified sulfinylbis [(2,4-dihydroxyphenyl)methanethione]s and the corresponding aminothiophenecarboxamides. The derivatives showed anticancer activity in vitro. These compounds inhibited the proliferation and viability of lung cancer A549, colon cancer HT-29 and glioma C6 cells in a concentration-dependent manner. Some of the derivatives had no influence on normal skin fibroblasts culture viability. Moreover, one compound (1b) showed the ability to inhibit DNA synthesis in cancer cells, especially in C6 cells, and was not toxic for normal oligodendrocytes and hepatocytes. Using reversed phase RP 18 HPLC and immobilised artificial membrane (IAM) chromatography the phase affinity of the compounds was determined. The influence of lipophilicity on the activity of compounds has been discussed.

Inhibiting centrosome clustering reduces cystogenesis and improves kidney function in autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is a monogenic disorder accounting for approximately 5% of patients with renal failure, yet therapeutics for the treatment of ADPKD remain limited. ADPKD tissues display abnormalities in the biogenesis of the centrosome, a defect that can cause genome instability, aberrant ciliary signaling, and secretion of pro-inflammatory factors. Cystic cells form excess centrosomes via a process termed centrosome amplification (CA), which causes abnormal multipolar spindle configurations, mitotic catastrophe, and reduced cell viability. However, cells with CA can suppress multipolarity via "centrosome clustering," a key mechanism by which cells circumvent apoptosis. Here, we demonstrate that inhibiting centrosome clustering can counteract the proliferation of renal cystic cells with high incidences of CA. Using ADPKD human cells and mouse models, we show that preventing centrosome clustering with 2 inhibitors, CCB02 and PJ34, blocks cyst initiation and growth in vitro and in vivo. Inhibiting centrosome clustering activates a p53-mediated surveillance mechanism leading to apoptosis, reduced cyst expansion, decreased interstitial fibrosis, and improved kidney function. Transcriptional analysis of kidneys from treated mice identified pro-inflammatory signaling pathways implicated in CA-mediated cystogenesis and fibrosis. Our results demonstrate that centrosome clustering is a cyst-selective target for the improvement of renal morphology and function in ADPKD.

Ultrastructure expansion microscopy (U-ExM) of mouse and human kidneys for analysis of subcellular structures.

Ultrastructure expansion microscopy (U-ExM) involves the physical magnification of specimens embedded in hydrogels, which allows for super-resolution imaging of subcellular structures using a conventional diffraction-limited microscope. Methods for expansion microscopy exist for several organisms, organs, and cell types, and used to analyze cellular organelles and substructures in nanoscale resolution. Here, we describe a simple step-by-step U-ExM protocol for the expansion, immunostaining, imaging, and analysis of cytoskeletal and organellar structures in kidney tissue. We detail the critical modified steps to optimize isotropic kidney tissue expansion, and preservation of the renal cell structures of interest. We demonstrate the utility of the approach using several markers of renal cell types, centrioles, cilia, the extracellular matrix, and other cytoskeletal elements. Finally, we show that the approach works well on mouse and human kidney samples that were preserved using different fixation and embedding conditions. Overall, this protocol provides a simple and cost-effective approach to analyze both preclinical and clinical renal samples in high detail, using conventional lab supplies and standard widefield or confocal microscopy.

Ultrastructure expansion microscopy (U-ExM) of mouse and human kidneys for analysis of subcellular structures.

Ultrastructure expansion microscopy (U-ExM) involves the physical magnification of specimens embedded in hydrogels, which allows for super-resolution imaging of subcellular structures using a conventional diffraction-limited microscope. Methods for expansion microscopy exist for several organisms, organs, and cell types, and used to analyze cellular organelles and substructures in nanoscale resolution. Here, we describe a simple step-by-step U-ExM protocol for the expansion, immunostaining, imaging, and analysis of cytoskeletal and organellar structures in kidney tissue. We detail the critical modified steps to optimize isotropic kidney tissue expansion, and preservation of the renal cell structures of interest. We demonstrate the utility of the approach using several markers of renal cell types, centrioles, cilia, the extracellular matrix, and other cytoskeletal elements. Finally, we show that the approach works well on mouse and human kidney samples that were preserved using different fixation and storage conditions. Overall, this protocol provides a simple and cost-effective approach to analyze both pre-clinical and clinical renal samples in high detail, using conventional lab supplies and standard widefield or confocal microscopy.

Silencing of Hsp27 and Hsp72 in glioma cells as a tool for programmed cell death induction upon temozolomide and quercetin treatment.

The aim of the present study was to investigate whether silencing of Hsp27 or Hsp72 expression in glioblastoma multiforme T98G and anaplastic astrocytoma MOGGCCM cells increases their sensitivity to programmed cell death induction upon temozolomide and/or quercetin treatment. Transfection with specific siRNA was performed for the Hsp gene silencing. As revealed by microscopic observation and flow cytometry, the inhibition of Hsp expression was correlated with severe apoptosis induction upon the drug treatment studied. No signs of autophagy were detected. This was correlated with a decreased mitochondrial membrane potential, increased level of cytochrome c in the cytoplasm, and activation of caspase 3 and caspase 9. All these results suggest that the apoptotic signal was mediated by an internal pathway. Additionally, in a large percentage of cells treated with temozolomide, with or without quercetin, granules within the ER system were found, which was accompanied by an increased level of caspase 12 expression. This might be correlated with ER stress. Quercetin and temozolomide also changed the shape of nuclei from circular to "croissant like" in both transfected cell lines. Our results indicate that blocking of Hsp27 and Hsp72 expression makes T98G cells and MOGGCCM cells extremely vulnerable to apoptosis induction upon temozolomide and quercetin treatment and that programmed cell death is initiated by an internal signal.

Apoptosis induction in human glioblastoma multiforme T98G cells upon temozolomide and quercetin treatment.

Glioblastoma multiforme is the most aggressive primary brain tumour. At the cellular and molecular levels, several mechanisms responsible for apoptosis or autophagy induction are blocked. Identification of molecular targets stimulating cells to initiate programmed cell death should be performed for therapeutic purposes. A promising solution is the combination of temozolomide and quercetin. The aim of our study was to evaluate the effect of both drugs, applied alone and in combinations, on apoptosis and autophagy induction in human glioblastoma multiforme T98G cells. Our results clearly indicate that quercetin and temozolomide induce apoptosis very significantly, having no effect on autophagy induction. At the molecular level, it was correlated with caspase 3 and 9 activation, cytochrome c release from the mitochondrium and a decrease in the mitochondrial membrane potential. Both drugs are also potent Hsp27 and Hsp72 inhibitors. This suggests that the apoptotic signal goes through an internal pathway. Increased expression of caspase 12 and the presence of several granules in the cytoplasm after temozolomide treatment with or without quercetin preceding appearance of apoptosis may suggest that apoptosis is initiated by ER stress. Additionally, it was accompanied by changes in the nuclear morphology from circular to 'croissant like'.

Impaired centrosome biogenesis in kidney stromal progenitors reduces abundance of interstitial lineages and accelerates injury-induced fibrosis.

Defective centrosome function can disrupt embryonic kidney development, by causing changes to the renal interstitium that leads to fibrocystic disease pathologies. Yet, it remains unknown how mutations in centrosome genes impact kidney interstitial cells. Here, we examined the consequences of defective centrosome biogenesis on stromal progenitor cell growth, differentiation and fate. Conditional deletion of Cep120 , a ciliopathy gene essential for centrosome duplication, in the stromal mesenchyme resulted in reduced abundance of pericytes, interstitial fibroblasts and mesangial cells. This was due to delayed mitosis, increased apoptosis, and changes in Wnt and Hedgehog signaling essential for differentiation of stromal lineages. Cep120 ablation resulted in hypoplastic kidneys with medullary atrophy and delayed nephron maturation. Finally, centrosome loss in the interstitium sensitized kidneys of adult mice, causing rapid fibrosis via enhanced TGF-ß/Smad3-Gli2 signaling after renal injury. Our study defines the cellular and developmental defects caused by centrosome dysfunction in embryonic kidney stroma. Highlights: Defective centrosome biogenesis in kidney stroma causes:Reduced abundance of stromal progenitors, interstitial and mesangial cell populationsDefects in cell-autonomous and paracrine signalingAbnormal/delayed nephrogenesis and tubular dilationsAccelerates injury-induced fibrosis via defective TGF-ß/Smad3-Gli2 signaling axis.

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.

Identification of an Hsp90 mutation that selectively disrupts cAMP/PKA signaling in Saccharomyces cerevisiae.

The molecular chaperone Hsp90 cooperates with multiple cochaperone proteins as it promotes the folding and activation of diverse client proteins. Some cochaperones regulate the ATPase activity of Hsp90, while others appear to promote Hsp90 interaction with specific types of client proteins. Through its interaction with the adenylate cyclase Cyr1, the Sgt1 cochaperone modulates the activity of the cAMP pathway in Saccharomyces cerevisiae. A specific mutation in yeast Hsp90, hsc82-W296A, or a mutation in Sgt1, sgt1-K360E, resulted in altered transcription patterns genetically linked to the cAMP pathway. Hsp90 interacted with Cyr1 in vivo and the hsc82-W296A mutation resulted in reduced accumulation of Cyr1. Hsp90-Sgt1 interaction was altered by either the hsc82-W296A or sgt1-K360E mutation, suggesting defective Hsp90-Sgt1 cooperation leads to reduced Cyr1 activity. Microarray analysis of hsc82-W296A cells indicated that over 80 % of all transcriptional changes in this strain may be attributed to altered cAMP signaling. This suggests that a majority of the cellular defects observed in hsc82-W296A cells are due to altered interaction with one specific essential cochaperone, Sgt1 and one essential client, Cyr1. Together our results indicate that specific interaction of Hsp90 and Sgt1 with Cyr1 plays a key role in regulating gene expression, including genes involved in polarized morphogenesis.

Alpha-ketoglutarate (AKG) inhibits proliferation of colon adenocarcinoma cells in normoxic conditions.

BACKGROUND AND OBJECTIVE: Alpha-ketoglutarate (AKG), a key intermediate in Krebs cycle, is an important biological compound involved in the formation of amino acids, nitrogen transport, and oxidation reactions. AKG is already commercially available as a dietary supplement and its supplementation with glutamine, arginine, or ornithine alpha-ketoglutarate has been recently considered to improve anticancer immune functions. It is well documented that AKG treatment of Hep3B hepatoma cells in hypoxia induced HIF-alpha (hypoxia-inducible factor) degradation and reduced vascular endothelial growth factor (VEGF) synthesis. Moreover, AKG showed potent antitumor effects in murine tumor xenograft model, inhibiting tumor growth, angiogenesis, and VEGF gene expression. However, the mechanisms of its anticancer activity in normoxia have not been examined so far. RESULTS: Here, we report that in normoxia, AKG inhibited proliferation of colon adenocarcinoma cell lines: Caco-2, HT-29, and LS-180, representing different stages of colon carcinogenesis. Furthermore, AKG influenced the cell cycle, enhancing the expression of the inhibitors of cyclin-dependent kinases p21 Waf1/Cip1 and p27 Kip1. Moreover, expression of cyclin D1, required in G1/S transmission, was decreased, which accompanied with the significant increase in cell number in G1 phase. AKG affected also one the key cell cycle regulator, Rb, and reduced its activation status. CONCLUSION: In this study for the first time, the antiproliferative activity of AKG on colon adenocarcinoma Caco-2, HT-29, and LS-180 cells in normoxic conditions was revealed. Taking into consideration an anticancer activity both in hypoxic and normoxic conditions, AKG may be considered as a new potent chemopreventive agent.

RAD51 gene polymorphisms and sporadic colorectal cancer risk in Poland.

BACKGROUND: DNA repair processes play an important role in protection against carcinogenic factors. Mutations in DNA repair genes, which code proteins engaged in repair processes, may lead to carcinogenesis and among others also to colorectal cancer (CRC) development. The genetic variability in RAD51 may contribute to the appearance and progression of various cancers including CRC. The aim of the study was to compare the distribution of genotypes of RAD51 135G>C and 172G>T polymorphism between colorectal cancer patients and controls. MATERIAL AND METHODS: Both polymorphisms were evaluated by PCR-RFLP methods in colorectal tissue of 320 colorectal cancer subjects and 320 healthy subjects who served as controls. RESULTS: In the present work we demonstrated a significant positive association between the RAD51 C/C genotype and colorectal carcinoma. Variant 135C allele of RAD51 increased the cancer risk. However, we did not observe any relationship between each polymorphism and colorectal cancer progression assessed by node metastasis, tumour size and Dukes' stage. CONCLUSIONS: Our results suggest that variant genotypes of the 135G>C of RAD51 polymorphism may be positively associated with colorectal carcinoma in the Polish population. Further studies conducted on a larger group are required to clarify this point.

Dietary derived compounds in cancer chemoprevention.

Cancer chemoprevention is defined as the application of natural or synthetic agents to suppress or reverse cancer development and progression. In this field especially diet derived compounds have recently attracted researchers' attention as potential therapeutics generally exerting low toxicity compared with regular drugs. This review presents a survey of recent findings concerning the most promising dietary chemopreventive agents such as green tea polyphenols (i.e. catechins), long-chain polyunsaturated fatty acids, carotenoids, glucosinolates/isothiocyanates, vitamins (i.e. vitamin D and folate) and minerals (i.e. calcium and selenium). Molecular targets involved in intrinsic pathways affected by these natural compounds are also shortly discussed.

Kynurenic acid synthesis and kynurenine aminotransferases expression in colon derived normal and cancer cells.

BACKGROUND: Kynurenic acid (KYNA), a tryptophan metabolite, was found in human saliva, gastric juice, bile, pancreatic juice and mucus of rat small intestine. METHODS: KYNA content in mucus aspirated from human caecum or colon ascendens and KYNA production in colon epithelial and cancer cells were determined using HPLC. Moreover, biological properties of KYNA and kynurenine aminotransferases (KATs) expression in colon epithelial and colon cancer cells were studied. RESULTS: Considerably higher KYNA concentration was detected in samples from patients diagnosed with colon carcinoma (269.40 ± 107.00 pmol/ml, N = 4), Adenoma tubulovillosum (200.50 ± 36.72, N = 10) or Adenoma tubulare (243.50 ± 38.09, N = 9) than in control group (82.22 ± 7.61 pmol/ml, N = 30). Moreover, colon epithelium CCD 841 CoTr cells actively synthesized KYNA in a concentration- and time-dependent manner. This process was decreased by aminooxyacetic acid and L-glutamate in opposite to 4-aminopyridine treatment. Interestingly, KYNA production in colon cancer cells (HT-29 1.39 ± 0.27, LS-180 1.18 ± 0.15 and Caco-2 4.21 ± 0.30 pmol/1 x 10(5) cells/2 h) was considerably higher in comparison to normal colon epithelial cells (0.70 ± 0.07 pmol/1 x 10(5) cells/2 h). However, KATs I and II were expressed at similar level in both colon epithelium and cancer cells. Furthermore, KYNA exerted an antiproliferative effect at higher micro- and millimolar concentrations against colon cancer cells with the IC(50) of 0.9, 0.2 and 1.2 mM for HT-29, LS-180 and Caco-2 cells, respectively. CONCLUSION: Summarizing, this is the first report presenting KYNA synthesis and KAT expression in colon derived normal and cancer cells.