A Potential Combination of Targeting HSP90 and mTOR in Breast Cancer Cell Growth, Migration, and Invasion Through Inhibiting AKT Phosphorylation and F-actin Organization.

BACKGROUND/AIM: Breast cancer is the most prevalent form of cancer among women worldwide, with a high mortality rate. While the most common cause of breast cancer death is metastasis, there is currently no potential treatment for patients at the metastatic stage. The present study investigated the potential of using a combination of HSP90 and mTOR inhibitor in the treatment of breast cancer cell growth, migration, and invasion. MATERIALS AND METHODS: Gene Expression Profiling Interactive Analysis (GEPIA) was used to investigate the gene expression profiles. Western blot analysis and fluorescence staining were used for protein expression and localization, respectively. MTT, wound healing, and transwell invasion assays were used for cell proliferation, migration, and invasion, respectively. RESULTS: GEPIA demonstrated that HSP90 expression was significantly higher in breast invasive carcinoma compared to other tumor types, and this expression correlated with mTOR levels. Treatment with 17-AAG, an HSP90 inhibitor, and Torkinib, an mTORC1/2 inhibitor, significantly inhibited cell proliferation. Moreover, combination treatment led to down-regulation of AKT. Morphological changes revealed a reduction in F-actin intensity, a marked reduction of YAP, with interference in nuclear localization. CONCLUSION: Targeting HSP90 and mTOR has the potential to suppress breast cancer cell growth and progression by disrupting AKT signaling and inhibiting F-actin polymerization. This combination treatment may hold promise as a therapeutic strategy for breast cancer treatment that ameliorates adverse effects of a single treatment.

Role of Oxidative Stress-Dependent C/EBPß Expression on CAF Transformation Inducing HCT116 Colorectal Cancer Cell Progression; Migration and Invasion.

OBJECTIVE: To investigate oxidative stress-related CAF transformation through C/EBPß, which affects CRC progression and may have a potential implication for CRC treatment. METHODS: The conditioned media (CM) from HCT116, CRC cells, was used to activate CCD-18Co, colon fibroblasts, then the ability of activated FBs to induce HCT116 growth and progression was assessed using MTT assay, transwell migration, and matrix invasion assay. Alteration of the cytokine profile and oxidative stress of the activated FBs were studied with cytokine arrays and DCFH-DA assay, respectively. The protein expressions of the CAF markers (α-SMA and FAP) and C/EBPß were investigated with immunofluorescence and western blotting. RESULT: It was found that CM from HCT116 cells induced oxidative stress, change of cytokine profile, CAF markers, and the C/EBPß expression of activated FBs. Furthermore, when the oxidative stress of the activated FBs was suppressed, FAP and C/EBPß expression were downregulated, correlating with the disabling of their capability to support the cancer progression. The C/EBPß and prognosis for CRC patients were accessed using the GEPIA dataset, in which high C/EBPß expression was associated with a poor prognosis. CONCLUSION: These findings suggest that C/EBPß expression has a role in CAF transformation in an oxidative stress-related manner and might be used as a target to improve aggressive CRC treatment outcomes.

RED RICE BRAN EXTRACT SUPPRESSES COLON CANCER CELLS VIA APOPTOSIS INDUCTION/CELL CYCLE ARREST AND EXERTS ANTIMUTAGENIC ACTIVITY.

BACKGROUND: Red rice bran extract (RRBE) contains many biologically active substances exerting antioxidant and anti-inflammatory effects. AIM: To evaluate the anticancer potential of RRBE in human colon cancer cells and its mutagenic/antimutagenic effects on nonmalignant cells. MATERIALS AND METHODS: The cytotoxic effect of RRBE was determined by trypan blue exclusion in HCT116, HT29 cell lines and a non-cancerous HEK293 cell line, and its antiproliferative effect using MTS and colony formation assay. The apoptosis induction was evaluated using ELISA, and the apoptotic rate and cell cycle progression were assessed by flow cytometry. The mutagenic/ antimutagenic potential of RRBE was analyzed by micronucleus assay in the V79 cell line. RESULTS: RRBE caused a dose-dependent reduction of cell viability in colon cancer cells and showed a limited cytotoxicity against HEK293 cells. The treatment with RRBE suppressed proliferation of HCT116 and HT29 cells and induced apoptosis as evidenced by the increased DNA fragmentation and the apoptotic cell counts. Furthermore, RRBE treatment significantly increased the number of cells at the G2/M phase triggering the arrest of the cell cycle in colon cancer cells. Interestingly, RRBE did not increase the micronucleus frequency in V79 cells but reduced the micronucleus formation caused by mitomycin C. CONCLUSION: RRBE effectively suppressed proliferation, induced apoptosis, and caused a cell cycle arrest in human colon cancer cells while being non-mutagenic and exerting antimutagenic effects in vitro.

Vitamin C Improves the Inhibitory Effects of Oxaliplatin on HCT-116 Colorectal Cancer Growth and Progression Through Cellular Oxidant Function-associated Cadherin Molecules.

BACKGROUND/AIM: Colorectal cancer (CRC) is strongly associated with altered cadherin adhesion molecules. Oxaliplatin is a standard treatment for CRC, yet high-doses have concerning side effects. In this study, the effects of oxaliplatin and the combination of oxaliplatin with vitamin C on HCT-116 CRC cell migration and invasion were studied through the roles of cellular oxidative stress associated with cadherin molecules. MATERIALS AND METHODS: The cellular assays used in this research were MTT, DCFH-DA, immunofluorescence, and western blotting. Cancer progression was examined using wound healing and Boyden chamber techniques. RESULTS: The results indicate that hydrogen peroxide-induced cellular oxidative stress induced cancer cell migration and invasion. The combined treatment of oxaliplatin with a pro-oxidant concentration of vitamin C resulted in higher toxicity than treatment with oxaliplatin alone. However, treatment with the combination of oxaliplatin and antioxidant concentrations of vitamin C suppressed cancer migration and invasion. Furthermore, the combination treatment increased E-cadherin expression, whereas decreased that of N-cadherin. CONCLUSION: Treatment with the combination of oxaliplatin with vitamin C can inhibit CRC cell growth and decrease cancer cell migration and invasion, via oxidative stress and cadherins.

Anticancer and Antimutagenic Properties of Pogonatherum paniceum on Colorectal Cancer Cells.

Background: Pogonatherum paniceum (P. paniceum) (Lam.) Hack. plays an important role in detoxification. However, its anticancer activity has not yet been elucidated. The aim of our study was to examine the suppressive proliferation, anti-migration and mutagenic/antimutagenic properties of P. paniceum. Moreover, we set out to determine the cellular mechanism underlying its antiproliferation. Methods: To investigate P. paniceum's anticancer ability, HCT116 and HT29 cell lines were treated with a water extract containing P. paniceum, and then the cell viability was examined using the trypan blue exclusion method which were compared to HEK293 (non-cancerous cells). The anticancer effects were investigated by MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) and colony formation assay. Apoptosis induction, cell cycle distribution, and migration abilities were assessed by cell death detection enzyme-linked immunoassay (ELISA), flow cytometry, and wound healing assay. Finally, the mutagenicity and antimutagenicity were evaluated using the micronucleus assay. Results: Treatment with P. paniceum caused a loss of cell viability in HCT116 and HT29 cells (not found in HEK293), which had an IC50 (half-maximal inhibitory concentration) of 1,156.2 and 1,207.0 µg/mL, respectively. We found that P. paniceum significantly inhibited the proliferative function of HCT116 and HT29 cells. To find the mechanism that exerts a suppressive proliferation effect on P. paniceum, we determined the DNA fragmentation and cell cycle distribution. We also found that P. paniceum treatment increased apoptosis and arrested of the cell cycle at G0/G1 remarkably when compared with the control group. Moreover, P. paniceum could decrease the migration of HCT116 and HT29 cancer cells. Finally, the treatment of P. paniceum did not induce micronucleus formation but did decrease the micronucleus frequency against mutagen-mitomycin C. Conclusions: P. paniceum did not possess any toxicity (cytotoxic and mutagenic) but has the potential for anticancer activity against human colorectal cells by increasing apoptosis, which leads to the suppression of cell proliferation. P. paniceum also inhibits cell migration and exerts antimutagenicity, thereby suggesting that P. paniceum might be useful for colorectal cancer treatment.

N-acetylcysteine improves the inhibitory effect of Quercetin-rich onion extract on HT-29 and HCT-116 colorectal cancer migration and invasion through iNOS suppression.

As colorectal cancer (CRC) usually presents at an advanced stage, it responds poorly to traditional surgery and chemoradiotherapy. Reactive oxygen species (ROSs) are a critical factor in cancer progression. Quercetin, a bioflavonoid derived from onion peel extract, provides great anti-oxidant and anti-cancer potential. Therefore, quercetin in combination with N-Acetylcysteine (NAC), a well-known anti-oxidant and adjuvant agent in cancer-chemotherapeutic drugs, was considered as a way of increasing treatment efficacy. Thus, this study aimed to evaluate the improvement effect of quercetin in combination with NAC in human CRC (HT-29 and HCT-116) cell progression, migration and invasion. Firstly, the effects of quercetin, NAC, and the combination of quercetin and NAC on cellular oxidants and glutathione levels were evaluated. Cell viability, anti-migrative activity and invasive activity were determined by MTT, wound healing, and Matrigel invasion tests, respectively. Then, the proteins involved in cell migration, invasion, and cellular oxidants were investigated. Moreover, the gene expression and overall survival were further validated by the GEPIA2 database. The results reveal that the combination was most effective in decreasing cellular oxidants and increasing glutathione levels, while there was a significant decrease in cancer cell migration and invasion involved in the suppression of iNOS, ICAM-1, and MMP-2 proteins. Furthermore, bioinformatic analysis verified that iNOS, ICAM-1, and MMP-2 were highly expressed in CRC tissue and also associated with a poor prognosis. This study demonstrated that Quercetin has higher efficacy when used in combination with NAC, representing a potential combination agent for anti-cancer drug development.

Increased platelet activation and lower platelet-monocyte aggregates in COVID-19 patients with severe pneumonia.

BACKGROUND: The increased procoagulant platelets and platelet activation are associated with thrombosis in COVID-19. In this study, we investigated platelet activation in COVID-19 patients and their association with other disease markers. METHODS: COVID-19 patients were classified into three severity groups: no pneumonia, mild-to-moderate pneumonia, and severe pneumonia. The expression of P-selectin and activated glycoprotein (aGP) IIb/IIIa on the platelet surface and platelet-leukocyte aggregates were measured prospectively on admission days 1, 7, and 10 by flow cytometry. RESULTS: P-selectin expression, platelet-neutrophil, platelet-lymphocyte, and platelet-monocyte aggregates were higher in COVID-19 patients than in uninfected control individuals. In contrast, aGPIIb/IIIa expression was not different between patients and controls. Severe pneumonia patients had lower platelet-monocyte aggregates than patients without pneumonia and patients with mild-to-moderate pneumonia. Platelet-neutrophil and platelet-lymphocyte aggregates were not different among groups. There was no change in platelet-leukocyte aggregates and P-selectin expression on days 1, 7, and 10. aGPIIb/IIIa expression was not different among patient groups. Still, adenosine diphosphate (ADP)-induced aGPIIb/IIIa expression was lower in severe pneumonia than in patients without and with mild-to-moderate pneumonia. Platelet-monocyte aggregates exhibited a weak positive correlation with lymphocyte count and weak negative correlations with interleukin-6, D-dimer, lactate dehydrogenase, and nitrite. CONCLUSION: COVID-19 patients have higher platelet-leukocyte aggregates and P-selectin expression than controls, indicating increased platelet activation. Compared within patient groups, platelet-monocyte aggregates were lower in severe pneumonia patients.

Riceberry Rice Germination and UVB Radiation Enhance Protocatechuic Acid and Vanillic Acid to Reduce Cellular Oxidative Stress and Suppress B16F10 Melanogenesis Relating to F-Actin Rearrangement.

Ultraviolet type B (UVB) radiation plays an important role in hyperpigmentation disorder, which induces cellular oxidative stress and causes abnormal melanin production and secretion. The stress condition plays an essential role in actin polymerization relating to F-actin rearrangement and forms dendrite to send melanin pigment to the uppermost layer of the skin. Phenolic compounds are secondary metabolites that mainly synthesize under stress conditions to protect plants from harmful environments and have been reported as effective agents in anti-oxidant and anti-melanogenesis. However, the influence of phenolic compounds on F-actin rearrangement-associated dendrite formation has not been studied so far. Hence, this study aimed to investigate the enhancing phytophenolic targets in riceberry rice (Oryza sativa L.) germination and UVB radiation (RR-GR) to suppress melanogenesis relating to F-rearrangement. As a result, the RR-GR had the potential to enhance phenolic acids such as protocatechuic and vanillic acid, which have been proven to possess anti-oxidant activity and anti-tyrosinase properties. Riceberry rice's modification showed the potential to reduce cellular oxidative stress and suppress B16F10 melanogenesis relating to F-actin rearrangement that is associated with dendrite formation.

YAP, a novel target regulates F-actin rearrangement-associated CAFs transformation and promotes colorectal cancer cell progression.

Colorectal cancer (CRC) progression is strongly influenced by the tumor microenvironment (TME) in which cancer-associated fibroblasts (CAFs) are the major components influencing CRC growth and progression. The present study aimed to investigate the effect of YAP on F-actin arrangement in CAF transformation and the possibility of using YAP as a target for inhibiting CRC growth and progression. Conditioned media were collected from direct interaction between CRC cells and fibroblasts. CAF markers were investigated by flow cytometry, western blot analysis, and immunofluorescence assay in CM-treated fibroblasts. Promoting the CRC progression of conditioned media was determined in CRC cells by using MTT assay, fluorescence assay, wound healing assay, transwell migration assay, and tubulogenesis. The results showed that the conditioned media induced the expression of CAF markers associated with the central rearrangement of F-actin in colon fibroblasts, upregulating and promoting the nuclear translocation of YAP. The conditioned media also significantly promoted the proliferation, migration, invasion, and angiogenesis of CRC cells. Interestingly, Verteporfin, a YAP inhibitor during cocultivation, abolished the conversion of CAFs and inhibited proliferation, migration, invasion, and angiogenesis in CRC cells. Moreover, bioinformatics analysis was employed to determine the potential role of YAP as a prognostic marker in CRC patients from databases. The results suggested that YAP has higher expression in CRC patients and is associated with a poor prognosis. In conclusion, these findings demonstrate that YAP-related F-actin rearrangement may be a potential new target of combination therapy with a focus on targeting TME.

Aqueous Thunbergia laurifolia leaf extract alleviates paraquat-induced lung injury in rats by inhibiting oxidative stress and inflammation.

BACKGROUND: Paraquat (PQ) has been reported to have a high mortality rate. The major target organ of PQ poisoning is the lungs. The pathogenesis of PQ-induced lung injury involves oxidative stress and inflammation. Unfortunately, there is still no effective antidote for PQ poisoning. We hypothesized that aqueous Thunbergia laurifolia (TL) leaf extract is a possible antidote for PQ-induced lung injury. METHODS: The total phenolic content and caffeic acid content of an aqueous extract of TL leaves were analyzed. Male Wistar rats were randomly divided into four groups (n = 4 per group): the control group (administered normal saline), the PQ group (administered 18 mg/kg body weight (BW) PQ dichloride subcutaneously), the PQ + TL-low-dose (LD) group (administered PQ dichloride subcutaneously and 100 mg/kg BW aqueous TL leaf extract by oral gavage) and the PQ + TL-high-dose (HD) group (administered PQ dichloride subcutaneously and 200 mg/kg BW aqueous TL leaf extract by oral gavage). Malondialdehyde (MDA) levels and lung histopathology were analyzed. In addition, the mRNA expression of NADPH oxidase (NOX), interleukin 1 beta (IL-1ß), and tumor necrosis factor alpha (TNF-α) was assessed using reverse transcription-polymerase chain reaction (RT-PCR), and the protein expression of IL-1ß and TNF-α was analyzed using immunohistochemistry. RESULTS: The total phenolic content of the extract was 20.1 ± 0.39 µg gallic acid equivalents (Eq)/mg extract, and the caffeic acid content was 0.31 ± 0.01 µg/mg. The PQ group showed significantly higher MDA levels and NOX, IL-1ß and TNF-α mRNA expression than the control group. Significant pathological changes, including alveolar edema, diffuse alveolar collapse, hemorrhage, leukocyte infiltration, alveolar septal thickening and vascular congestion, were observed in the PQ group compared with the control group. However, the aqueous TL leaf extract significantly attenuated the PQ-induced increases in MDA levels and NOX, IL-1ß and TNF-α expressions. Moreover, the aqueous TL leaf extract ameliorated PQ-induced lung pathology. CONCLUSION: This study indicates that aqueous TL leaf extract can ameliorate PQ-induced lung pathology by modulating oxidative stress through inhibition of NOX and by regulating inflammation through inhibition of IL-1ß and TNF-α expressions. We suggest that aqueous TL leaf extract can be used as an antidote for PQ-induced lung injury.

Thioredoxin Reductase-1 as a Potential Biomarker in Fibroblast-Associated HCT116 Cancer Cell Progression and Dissemination in a Zebrafish Model.

The tumor microenvironment, especially that of fibroblasts, strongly promotes colorectal cancer (CRC) progression. Progressive cancers usually accumulate high reactive oxygen species (ROS), leading to oxidative stress. The stress relates to the expression of thioredoxin reductase-1 (TrxR-1), which is an oxidative stress sensitivity molecule. This study aimed to investigate TrxR-1 expression as an indication of colon-fibroblast-inducing colorectal cancer progression and metastasis. We found that the high proliferative fibroblast-cultured media (FCM) contained pro-inflammatory cytokines that have a high ability to influence HCT116 and CRC cell progression, when compared with complete media (CM) as a control in terms of growth (CM = 100.00%, FCM = 165.96%), migration (CM = 32.22%, FCM = 83.07%), invasion (CM = 130 cells/field, FCM = 449 cells/field), and EMT transformation while decreasing E-cadherin expression (CM = 1.00, FCM = 0.69) and shape factor (CM = 0.94, FCM = 0.61). In addition, the overexpression of TrxR-1 is associated with cellular oxidant enchantment in FCM-treated cells. A dot plot analysis showed a strong relation between the EMT process and the overexpression of TrxR-1 in FCM-treated cells (CM = 13/100 cells, FCM = 45/100 cells). The cancer transplantation of the adult zebrafish model illustrated a significantly higher number of microtumors in FCM-treated cells (CM = 4.33 ± 1.51/HPF, FCM = 25.00 ± 13.18/HPF) disseminated in the intraperitoneal cavity with TrxR-1 positive cells. The overexpression of TrxR-1 indicated fibroblast-associated CRC progression in HCT116 cells and the zebrafish model. Therefore, TrxR-1 could be applied as a novel biomarker for colorectal cancer progression and prognostic evaluation.

Onion Peel Extract Inhibits Cancer Cell Growth and Progression through the Roles of L1CAM, NF-κB, and Angiogenesis in HT-29 Colorectal Cancer Cells.

Colorectal cancer (CRC) is an aggressive malignancy. Critical mechanisms that support CRC progression include cell migration, invasion, metastasis, and angiogenesis, which is associated with L1 cell adhesion molecule (L1CAM) and nuclear factor-kappa B (NF-κB) signaling pathways. In this study, viability of HT-29 cells and human umbilical vein endothelial cells (HUVECs) was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, and cell apoptosis was investigated by flow cytometry assays. HT-29 cell migration and invasion were observed by wound healing and Transwell invasion assays, respectively, and tube formation of HUVECs was observed by tubulogenesis assays. L1CAM and NF-κB protein expressions in HT-29 cells treated with onion peel extract were determined by indirect immunofluorescence. Results showed that high dose treatments of onion peel extract inhibited cell viability of both HT-29 cells and HUVECs, induced HT-29 cell apoptosis, and inhibited HT-29 cell migration and invasion. Moreover, onion peel extract decreased total HUVEC tube length and, at a concentration of 10 µg/mL, showed potential to downregulate L1CAM and NF-κB. In conclusion, onion peel extract inhibits HT-29 cell growth, migration, and invasion through suppressing pathways related to angiogenesis downstream of L1CAM-activated NF-κB.

Additive Effect of a Combination of Artocarpus lakoocha and Glycyrrhiza glabra Extracts on Tyrosinase Inhibition in Melanoma B16 Cells.

Artocarpus lakoocha (Al) and Glycyrrhiza glabra (Gg) extracts have been reported to show tyrosinase inhibitory activity and melanin pigment reduction. This is the first study to assess the combination of Al and Gg extracts in enhancing inhibition of tyrosinase and reduction of melanin pigments. Al and Gg extracted by maceration in 70% and 95% ethanol were analyzed for oxyresveratrol and glabridin using Ultra High Performance Liquid Chromatography. Extracts of Al and Gg singly and combinations of Al95 and Gg95 were tested for cytotoxicity, tyrosinase inhibitory activity, and reduction of melanin pigments in melanoma B16 cells. Al95 had higher antioxidant, tyrosinase inhibitory activity and reduced more melanin pigments in B16 cells compared to Al70, and exhibited higher levels of oxyresveratrol. Gg95 inhibited oxidative stress and mushroom tyrosinase better than Gg70, and exhibited higher levels of glabridin. Combinations of Al95 and Gg95 at various ratios (concentration of 0.1 mg/mL) were not cytotoxic to B16 cells. Interestingly, Al95 and Gg95 combined at a ratio 9:1 reduced melanin pigment up to 53% in B16 cells. This combination of Al95 and Gg95 extracts exhibited the additive effect of reducing melanin pigments by suppressing the expression of microphthalmia-associated transcription factor (MITF), tyrosinase (TYR) and tyrosinase-related protein-2 (TRP-2) in B16 cells. The combination of Al and Gg extracts could be developed as skin care products for hyperpigmentation treatment.

Germinated Riceberry Rice Enhanced Protocatechuic Acid and Vanillic Acid to Suppress Melanogenesis through Cellular Oxidant-Related Tyrosinase Activity in B16 Cells.

The anti-melanogenic bioactivities of phytophenolic compounds have been well recognized. Riceberry rice contains a rich source of phenolic compounds that act as melanin inhibitors through their antioxidant and anti-tyrosinase properties. Germination has been shown to be an effective process to improve targeted phenolic compounds. In this study, germinated riceberry rice extract was tested for antioxidant activity. Total phenolic content was determined while the tyrosinase inhibitory effect was screened by the in vitro mushroom tyrosinase assay. Cytotoxicity of germinated riceberry rice extract was investigated in B16 cells before evaluating its activities on cellular tyrosinase, melanogenesis, melanin excretion, morphological appearance, and cellular oxidants. Germinated riceberry rice extract showed increased potency of antioxidants and was also twice as effective for mushroom tyrosinase inhibition when compared with ungerminated riceberry rice extract. In B16 cells, the extract inhibited cellular tyrosinase, melanogenesis, and cellular oxidants in a dose-dependent manner when compared with untreated cells. Germinated riceberry rice extract also displayed an effect on B16 cells morphology by reducing the number of melanin- containing cells and their dendriticity. Additionally, the germination of riceberry rice dominantly enhanced two phenolic acids, protocatechuic acid and vanillic acid, which have the potential for antioxidant-associated hyperpigmentation control. Thus, the restricted germination of riceberry rice tended to promote protocatechuic acid and vanillic acid, which dominantly displayed antioxidants and tyrosinase-related melanogenic inhibition.

Modified Riceberry rice extract suppresses melanogenesis-associated cell differentiation through tyrosinase-mediated MITF downregulation on B16 cells and in vivo zebrafish embryos.

BACKGROUND AND PURPOSE: Excessive melanin production caused by overactive tyrosinase (TYR) enzyme results in several dermatological problems. The TYR inhibitor, derived from metabolite changes during fermentation, has been well recognized for pigmentation control. EXPERIMENTAL APPROACH: This study is interested in alternative anti-melanogenic agents from bio-modified Riceberry rice through fermentation. Modified Riceberry rice extract (MRB) was evaluated for its cytotoxicity, melanin content, melanin excretion, and TYR activity in B16 cells. TYR and their melanogenesis-related molecules such as TYR-related proteins-1 and -2, and microphthalmia-associated transcription factor (MITF) were determined. The anti-melanogenic activity and toxicity were also tested using the embryonic zebrafish model. Furthermore, comprehensive genotoxicity testing was verified by cytokinesis-block micronucleus cytome assay. FINDINGS/RESULTS: The study found that non-cytotoxic concentrations of MRB at 20 and 40 mg/mL inhibited melanogenesis and melanin excretion by interfering B16 cell morphology. Cellular TYR enzymatic activity was also suppressed in the treated cells. The mRNA transcription and protein expression levels of TYR and MITF decreased by dose-dependent and time-dependent manners with MRB treatment. In the animal model, MRB was found to be safe and potent for melanogenesis-related TYR inhibition in embryonic zebrafish at 20 and 30 mg/mL. The toxicity of effective doses of MRB showed no genotoxicity and mutagenicity. CONCLUSION AND IMPLICATIONS: This study suggests that MRB has anti-melanogenesis potential through TYR and its-related protein inhibitions. MRB is also safe for applications and maybe a promising anti-melanogenic agent for hyperpigmentation control.

Association of XPO1 Overexpression with NF-κB and Ki67 in Colorectal Cancer.

OBJECTIVES: Exportin 1(XPO1), a nuclear exporter protein, has been gaining recognition in cancer progression and treatment. This study aimed to evaluate the association between the overexpression of XPO1 with NF-κB, Ki67 and clinicopathological characteristics in colorectal cancer (CRC) tissue samples and to explore the anti-proliferative effect of KPT-330, as XPO1 inhibitor, in colorectal cancer cell line. METHODS: Forty CRC tissue samples were analyzed by immunostaining for the expressions of XPO1, NF-κB and Ki67 and then the anti-proliferative effect of the KPT-330 was also evaluated in HT29 colorectal cancer cell line. RESULTS: XPO1 overexpression was observed in 52.5% of CRC and significantly apparent with strong intensity in tumor cells compared to the normal adjacent epithelium (P<0.001). Regarding to the histopathological characteristics, the XPO1 overexpression significantly associated with advanced tumor stages (P=0.049) and has great tendency towards moderate/poorly differentiated tumors. Although the XPO1 overexpression was strongly associated with high Ki67 expression (P=0.001), only Ki67 expression showed significant association with tumor size (P=0.012). No significant association was detected between the XPO1 overexpression and NF-κB, while the NF-κB positive expression was significantly associated with lymph node metastasis and Ki67 expression at P=0.027 and P= 0.007, respectively. The in vitro experiments showed a great impact of KPT-330, as XPO1 inhibitor, to inhibit cancer growth in dose and time dependent manner and significantly diminished the colony formation (P<0.001) of HT29 cells- associated with the expression of Ki67 (P<0.001). CONCLUSION: XPO1 overexpression and NF-κB expression may serve as potential biomarker associated with CRC pathogenesis and proliferation, while the KPT-330 is effectively inhibited-colon cancer growth in vitro. Further studies considering the prognostication role of XPO1 overexpression in CRC are required.
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The roles of p53 and XPO1 on colorectal cancer progression in Yemeni patients.

BACKGROUND: The colorectal cancer (CRC) tumorigenesis is driving by genetic alterations leading to changes in protein expression such as p53. The p53 is frequently expressed in CRC and its association with clinicopathological features is still controversial. Moreover, accumulated evidence suggests that both p53 and nuclear exporter protein, exportin 1 (XPO1), are working in reciprocal manner may lead to loss of p53 nuclear localization and enhance cancer progression through hyperactive nuclear export. Accordingly, the present study aimed to evaluate the expression of p53 in CRC Yemeni patients and to explore the association between the p53 and XPO1 coexpression in relation to clinicopathological features. METHODS: A series of 40 formalin fixed paraffin embedded (FFPE) tissue blocks taken from CRC patients that diagnosed as adenocarcinoma were prospectively collected and then analyzed for p53 and XPO1 expression by immunohistochemistry (IHC). The patients and tumor clinicopathological characteristics were retrieved from the histopathology reports and the P value <0.05 were considered statistically significant. RESULTS: The p53 expression was observed in 60% (24/40) of CRC tumor samples. Significantly, the p53 expression was noted in 72.4% (21/29) of the left side compared to 27.3% (3/11) of the right side colon tumors (P=0.014). Furthermore, p53 expression was positively and significantly correlated with well-but not moderate- or poorly-differentiated tumors (P=0.023). No significant difference was observed between the p53 expression and age, gender and tumor size. Regarding the XPO1 expression, the p53 expression didn't show an association with XPO1 expression. The coexpression of p53 and XPO1 analysis revealed that 100% (11/11) tumors with negative p53 and positive XPO1 coexpression was noted with lymph node metastasis with significant difference (P=0.003) and more frequently observed in moderate-or poorly- differentiated tumors. CONCLUSIONS: The loss of p53 accompanied with increased XPO1 expressions was associated with the progression of histopathological features of CRC Yemeni patients. Further studies are needed to elucidate the p53 genetic mutations in relation to the XPO1 coexpression in CRC prognosis.

Thai Water Lily Extract Induces B16 Melanoma Cell Apoptosis and Inhibits Cellular Invasion Through the Role of Cellular Oxidants

Melanoma is a cancer that is associated with a high capacity of invasion. Oxidative stress is recognized as cancer growth and progression. The phytochemical pigments of natural products show either anti-oxidant or pro-oxidant activity from the redox system. In addition, the phytophenolics also prevent cancer cell proliferation and progression. Objective: This study aims to investigate the effects of Thai water lily on cell apoptosis and cellular invasion through the role of cellular oxidants in B16 melanoma cells. Methods: The cytotoxicity and cell apoptosis of Thai water lily extract treating B16 cells were performed by using the MTT and Annexin V/PI-flow cytometry methods, respectively. In addition, cellular oxidants and cancer cell invasion were also obtained by using DCFH-DA and Boyden chamber assays, respectively. Results: Thai water lily, Nymphaea stellate extract was shown to be markedly toxic to B16 melanoma cells with IC50 = 814 µg/ml. The extract at 800 and 1,000 µg/ml demonstrated pro-oxidant activity relating to the cell apoptosis. The low concentrations of the extract at 200 and 400 µg/ml showed the anti-oxidant function associated with the inhibitory effect of melanoma cell invasion. Conclusion: Thai water lily extract may play an important role in bioactive work as a chemo preventive agent on the modulation of cellular oxidative stress-induced apoptosis and suppressed cancer cell invasion.

Unpolished Thai Rice Prevents Aberrant Crypt Foci Formation through the Invovement of ?catenin and COX2 Expression in AzoxymethaneTreated Rats.

Colorectal cancer (CRC) is a major cause of morbidity and mortality throughout the world, with chronic inflammation and diet as major causes in its development. Chemopreventive effects of natural dietary products have been the focus of studies for prevention over the past decade. This study was conducted to determine the effects of unpolished Thai rice during precancerous stage through the involvement of ?catenin, cyclooxygenase2 (COX2) expression and inflammatory cytokines focusing on azoxymethane (AOM)induced aberrant crypt foci (ACF)related to CRC. Male Sprague Dawley rats received two injections of AOM (15 mg/kg body weight) at weeks 4 and 5 while rats were treated with 20% or 70% unpolished Thai rice. The rats were sacrificed at week 38 and the colons removed for aberrant crypt foci (ACF) identification. Histopathologic changes, immunohistochemical analysis of ?catenin and COX2 expression, and cytokine expression of proinflammatory and antiinflammatory markers were determined. The administration of unpolished Thai rice significantly and dose dependently decreased the total number of ACF and the percentages of ACF with highgrade dysplasia. Interestingly, unpolished Thai rice suppressed the expression of ßcatenin and COX2. In addition, it also altered proinflammatory (IL6 and IFNγ) and antiinflammatory (IL 10) markers. The results suggested that unpolished Thai rice may provide a promising dietary intake for prevention during precancerous stage of CRC development, through the involvement of ßcatenin and COX2 expression, and also modulate inflammatory cytokinesrelated to CRC.

Heme oxygenase-1 alleviates alcoholic liver steatosis: histopathological study.

Excessive alcohol consumption is one of the most important causes of hepatic steatosis, which involves oxidative stress. In particular, increased oxidative stress has been strongly linked to stimulation of the expression of heme oxygenase-1 (HO-1). This study aimed to investigate whether HO-1 could alleviates alcoholic steatosis in rats. Male Wistar rats were randomly divided into 4 groups: 1) the control group, 2) the EtOH group, 3) the EtOH + ZnPP-IX group and 4) the EtOH + Hemin group. Liver histopathology was investigated in weeks 1 and 4 after the start of the treatment period. Alcohol treatment significantly increased the hepatic malondialdehyde (MDA) levels, an oxidative stress marker. In addition, it increased the triglyceride, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in both weeks. Gross examination demonstrated a yellowish and slightly enlarged liver in the alcohol-treated rats. Hematoxylin and eosin (H&E) and Oil Red O staining indicated hepatic steatosis, which was characterized by diffuse, extensive fatty accumulation and discrete lipid droplets of variable size in hepatocytes of the alcohol-treated rats. Administration of the HO-1 inducer hemin resulted in upregulation of hepatic HO-1 gene expression, reduced the MDA, triglyceride, ALT and AST levels and alleviated alcoholic hepatic steatosis, whereas administration of the HO-1 inhibitor zinc protoporphyrin IX (ZnPP-IX) resulted in downregulation of hepatic HO-1 gene expression and could not alleviate alcoholic hepatic steatosis either week. In conclusion, HO-1 could alleviate alcoholic hepatic steatosis in male Wistar rats and may be useful in development of a new therapeutic approach.