Human Hepatoma Cell Line SMMC-7721 Research Articles

Three new metabolites from the endophyte Fusarium proliferatum T2-10

One new cyclopeptide, cyclo-(L-Trp-L-Phe-L-Phe) (1), one new 2-pyridone derivative, fusarone A (3), and one new natural indole derivative, ethyl 3-indoleacetate (4), along with six known compounds were isolated from the endophytic fungus Fusarium proliferatum T2-10. The planar structures of three new compounds were identified by spectral methods including 1D and 2D NMR techniques, and the absolute configuration of compound 1 was elucidated by Marfey-MS method. In addition, all compounds were evaluated for their cytotoxic and antibacterial activities in vitro. Compound 2 showed remarkable cytotoxic activities against two human hepatoma cell lines SMMC7721 and HepG2 with IC50 values of 5.89 ± 0.74 and 6.16 ± 0.52 μM, and showed moderate antibacterial activities against Staphylococcus aureus and Enterococcus faecalis with MIC values of 7.81 and 15.62 μg/mL, respectively.

Design, synthesis, antibacterial/antitumor activity and in vitro stability of novel cordycepin derivatives with unsaturated fatty acid chain

To overcome the metabolic instability of cordycepin (adenosine deaminase (ADA) metabolic deamination and plasma degradation) and obtain better bioactivity, three novel kinds of cordycepin derivatives 1a-1c containing unsaturated fatty acids including linoleic acid, arachidonic acid and α-linolenic acid, respectively, were designed and synthesized. In terms of antibacterial activity, the synthesized compounds 1a and 1c showed enhanced activity than cordycepin in the tested bacterial strains. 1a-1c also exhibited enhanced antitumor activity against four cancer cell lines (human cervical cancer cell line HeLa, human non-small cell lung cancer cell line A549, human breast cancer cell line MCF-7, and human hepatoma cell line SMMC-7721) compared with cordycepin. Notably, 1a and 1b showed better antitumor activity even compared with positive control 5-Fluorouracil (5-FU) in HeLa, MCF-7 and SMMC-7721. The cell cycle assay indicated that when compared with cordycepin, 1a and 1b could significantly inhibit the cell propagation trapped in S and G2/M phases and increase the percentage of cells trapped in G0/G1 in HeLa and A549, which might provide a synergistic antitumor mechanism evidence different from cordycepin. Last but not the least, 1a and 1b displayed improved stability both in ADA solution and mouse plasma compared with cordycepin and 1a owns a solubility of 130 μg/mL in PBS. These results offer a novel insight into the primary structure and activity relationship of how the unsaturated fatty acid chain could affect the bioactivity of cordycepin, which also represents a series of cordycepin analogs with obviously improved bioactivity and enhanced stability, therefore promoting its druggable enhancement.

Analysis of differentially expressed genes in bile acid-treated liver cancer cells

Liver cancer is one of the tumors of digestive system. Bile acids are derivatives of choline acids and play a regulatory role in tumors. However, the gene expression profiles in liver cancer cells after bile acid treatment remain unclear. Human hepatoma cell line SMMC7721 was herein cultured. mRNA expression profile was detected by mRNA suppression subtractive hybridization. SMMC7721 cells were divided into 3 groups: control group, DLC1 (deleted in hepatocellular carcinoma 1) group and DLC1 siRNA group. The expression of DLC1, cell proliferation, cell apoptosis and cell invasion were detected by Transwell chamber method. The expressions of VEGF, MMP-2 and DLC1 were detected by Western blot. After bile acid treatment, DLC1, B-cell receptor-associated protein 31 (BCAP31), extension factor 1-α1 (eEF1a1), cell division cycle 20 (CDC20), WD repeat containing protein 6 (WDR6), extension factor Tu and mitochondria (TUFM) were the most significantly increased genes. DLC1 gene was selected with most significant changes. Overexpression of DLC1 significantly decreased expression of VEGF, MMP-2, and ephrin type-A receptor 2 (EphA2) (P <0.05). Transfection of DLC1 siRNA significantly down-regulated DLC1, promoting cell proliferation, decreasing Caspase3 activity, and increasing cell invasion, expression of VEGF, MMP-2 and EphA2 (P <0.05). Bile acid can cause differential gene expressions in liver cancer cells with DLC1 changes being most significant. DLC1 can influence the invasion and of proliferation hepatoma cells by regulating the expression of VEGF, MMP-2 and EphA2.

Bigelovin inhibits hepatocellular carcinoma cell growth and metastasis by regulating the MAPT-mediated Fas/FasL pathway.

Bigelovin (BigV), as traditional Chinese medicine, has been shown to inhibit the malignant progression of hepatocellular carcinoma (HCC). This study aimed to investigate whether BigV affects the development of HCC by targeting the MAPT and Fas/FasL pathway. Human HCC cell lines HepG2 and SMMC-7721 were used for this study. Cells were treated with BigV, sh-MAPT, and MAPT. The viability, migration, and apoptosis of HCC cells were detected by CCK-8, Transwell, and flow cytometry assays, respectively. Immunofluorescence and immunoprecipitation were used to verify the relationship between MAPT and Fas. Subcutaneous xenograft tumor and tail vein-injected lung metastases mouse models were constructed for histological observation. Hematoxylin-eosin staining was used to assess lung metastases in HCC. Western blotting was used to measure the expression of migration, apoptosis, and epithelial-mesenchymal transition (EMT) marker proteins, as well as Fas/FasL pathway-related proteins. BigV treatment inhibited the proliferation, migration, and EMT of HCC cells, whereas enhanced cell apoptosis. Moreover, BigV downregulated MAPT expression. The negative effects of sh-MAPT on HCC cell proliferation, migration, and EMT were enhanced by BigV treatment. Conversely, BigV addition attenuated the positive effects of MAPT overexpression on the malignant progression of HCC. In vivo experiments showed that BigV and/or sh-MAPT reduced tumor growth and lung metastasis while promoting tumor cell apoptosis. Furthermore, MAPT could act with Fas and inhibit its expression. sh-MAPT upregulated the expression of Fas/FasL pathway-associated proteins, which were enhanced by BigV administration. BigV suppressed the malignant progression of HCC via activating the MAPT-mediated Fas/FasL pathway.

Analysis of Differential Expression Profiles of Liver Cancer Cell Proteins After Treatment with Bile Acid

The pathogenesis of liver cancer has not been fully elucidated yet. Bile acids are components of bile, which are inorganic substances and regulate tumor progression. However, the differential expression profile of liver cancer cell proteins after bile acid treatment remains unclear. Human hepatoma cell line SMMC7721 was cultured and randomly assigned into control group and bile acid group followed by measuring the protein expression profile by protein fingerprinting. SMMC7721 cells were transfected with UGT2B or UGT2B, followed by analysis of UGT2B expression, cell proliferation, apoptosis, migration and PI3K/AKT signaling protein expression. The most obvious proteins with an increased expression after bile acid treatment were UGT2B, AAP, APLP2, LAPTM4B, NCOA4 with UGT2B being the most significant one. Overexpression of UGT2B decreased cell proliferation, promoted cell apoptosis, downregulated migration ability and AKT phosphorylation (P <0.05). UGT2B siRNA transfection significantly down-regulated UGT2B expression, promoted cell proliferation, decreased apoptosis rate, increased migration ability and AKT phosphorylation (P <0.05). In conclusion, bile acid can alter the protein expressions of liver cancer cells, with UGT2B being changed most obviously. UGT2B can affect liver cancer cell behaviors via modulating PI3K/AKT signaling.

Effects of sinomenine on apoptosis of human hepatoma cell line SMMC-7721 and its molecular mechanism in vitro

Evidence suggests that Sinomenine (SIN) has an excellent anticancer potential against hepatocellular carcinomas (HCCs), the apoptotic pathway genes affected by SIN and the mechanism of SIN that inhibits HCC are still unclear. Thus, the expression of apoptotic RNA and the early apoptosis in human HCC cells (SMMC-7721) caused by SIN need to be further investigated. In this study, to study the anti-tumor effect of SIN in vitro, human hepatoma cell line SMMC-7721 was treated with 0, 0.5, 1, 2 mM of SIN and 6.5 μM hydroxycamptothecin (HCPT) for 48 h. In addition, MTT assay, morphological observation and flow cytometry were used to detect the apoptosis of tumor cells, the changes of [Ca2+] and mitochondrial membrane potential (MMP) were detected by fluorescence microscopy. Moreover, the expression of tumor cell apoptosis-related genes was detected by real-time PCR, the anti-tumor mechanism of SIN was analyzed by bioinformatics methods including KEGG pathway graph was rendered by Pathview and heat map of three apoptosis pathways, including mitochondrial pathway, endoplasmic reticulum pathway (ER pathway), and death receptor pathway (DR pathway). In the results, significant inhibition of SMMC-7721 cell proliferation was found after SIN treatment. Under inverted microscopy, SIN-treated cells showed typical morphological alterations associated with apoptosis. A dose-dependent increase was observed in cytoplasmic Ca2+ in SIN-treated SMMC-7721 cells, while MMP levels decreased. The up-regulation and down-regulation of 47 apoptotic genes related to three apoptotic pathways were more clearly expressed by heat map analysis. Then the main pathways were analyzed, and the KEGG pathway graph was used to clearly express the expression of 92 apoptosis-related genes. Bid and Caspase-12 may be the main regulatory genes through mRNA expression. According to the previous changes in [Ca2+], SIN may affect the expression of caspase-12 through the change of [Ca2+], and then affect the expression of other related genes of Caspase family and Bcl-2 family. Finally, it affects the change of MMP of SMMC-7721 cell, leading to apoptosis of tumor cells.

Two new polycyclic polyprenylated acylphloroglucinols from Hypericum curvisepalum N. Robson

• Two new polycyclic polyprenylated acylphloroglucinols and seven known analogues were isolated. • Structure elucidation was performed by spectroscopic measurements including 2D-NMR spectroscopy. • Some compounds exhibited cytotoxic and antibacterial activities. Phytochemical investigation of the whole plant of Hypericum curvisepalum led to the isolation of nine polycyclic polyprenylated acylphloroglucinols ( 1 – 9 ). Among them, curvisepalumiones A ( 1 ) and B ( 2 ) are two new compounds and their structures were elucidated based on extensive spectroscopic data analysis combined with Rh 2 (OCOCF 3 ) 4 -induced electronic circular dichroism experiment. Cytotoxic assay showed that all the tested compounds except for 1 was moderately cytotoxic against human hepatocarcinoma cell line SMMC-7721 cell line, while compounds 3 and 4 also exhibited cytotoxicity toward MGC-803 cells. In addition, compound 2 exhibited weak inhibitory activity against Bacillus subtilis .

HMGB1 Promotes Resistance to Doxorubicin in Human Hepatocellular Carcinoma Cells by Inducing Autophagy via the AMPK/mTOR Signaling Pathway.

Chemoresistance remains as a major hindrance in the treatment of hepatocellular carcinoma (HCC). High mobility group box protein 1 (HMGB1) enhances autophagic flux and protects tumor cells from apoptosis, which results in acquired drug resistance. However, the exact mechanisms underlying HMGB1-modulated autophagy in HCC chemoresistance remain to be defined. In the present study, we found that administration of doxorubicin (DOX) significantly promoted HMGB1 expression and induced HMGB1 cytoplasmic translocation in human HCC cell lines BEL7402 and SMMC7721, which enhanced autophagy that contributes to protecting HCC cells from apoptosis and increasing drug resistance. Moreover, we observed HMGB1 translocation and elevation of autophagy in DOX-resistant BEL7402 and SMMC7721 cells. Additionally, inhibition of HMGB1 and autophagy increased the sensitivities of BEL-7402 and SMMC-7721 cells to DOX and re-sensitized their DOX-resistant cells. Subsequently, we confirmed with HMGB1 regulated autophagy by activating the 5ʹ adenosine monophosphate-activated protein kinase (AMPK)/mTOR pathway. In summary, our results indicate that HMGB1 promotes acquired DOX resistance in DOX-treated BEL7402 and SMMC7721 cells by enhancing autophagy through the AMPK/mTOR signaling pathway. These findings provide the proof-of-concept that HMGB1 inhibitors might be an important targeted treatment strategy for HCC.

Experimental Research on the Inhibitory Effect of IFN-Lambda 3 Combined with Sorafenib on the Growth of Liver Cancer Transplanted into Nude Mice

Objective: To investigate the effect of sorafenib combined with interferon-lambda 3 on the growth of liver cancer transplanted into nude mice. Methods: Female nude mice of 4-5 weeks of age that passed quarantine were selected and fed for 1-2 weeks before experimental operation. The cell suspension of human hepatoma cell line SMMC-7721 was inoculated into the right cervical axillary fossa with a syringe. The tumor-bearing mice were randomly divided into a control group and an experimental group. The control group received normal saline whereas the experimental group was further divided into three other groups: IFN-lambda 3 treatment group, sorafenib treatment group, and IFN-lambda 3 combined with sorafenib treatment group. The situation of nude mice was analyzed. At the end of the experiment, the volume of allogeneic transplanted tumor was measured, and the morphology of tumor cells, the expression of proliferating protein Ki-67, as well as the number of apoptotic cells were observed by hematoxylin and eosin (H&E) staining, immunohistochemistry (IHC), and TUNEL staining. Results: The tumor cell volume of the IFN-lambda 3 treatment group, sorafenib treatment group, and IFN-lambda 3 combined with sorafenib treatment group decreased, which was statistically significant compared with the control group (p < 0.05). The increment rate of proliferating protein Ki-67 in the transplanted tumor tissue of the three drug groups was significantly lower than that of the control group (p < 0.05). IFN-lambda 3 combined with sorafenib had the greatest effect on the expression level of Ki-67 protein. Compared with the control group, the expression rate was significantly lower (p < 0.05). In terms of cell apoptosis, IFN-lambda 3 and/or sorafenib, as well as the combination of the two, showed statistically significant differences compared with the control group (p < 0.05). The rate of cell apoptosis was the highest in the IFN-lambda 3 combined with sorafenib group. Conclusion: IFN-lambda 3 combined with sorafenib can inhibit the growth and proliferation of human hepatocellular carcinoma cells in nude mice and promote the apoptosis of hepatocellular carcinoma cells, which proves that IFN-lambda 3 combined with sorafenib can treat hepatocellular carcinoma in vivo.

Verification and Improvement of the MTT Method for in vitro HSS Bioactivity Activity Determination

To optimize the experimental conditions of MTT colorimetric assay for HSS bioactivity in vitro, we studied the optimal combination of the major conditions of the MTT assay by orthogonal test and other experiments, and compared HSS bioactivity in vitro measured by the improved MTT protocol and published MTT assay at serial protein doses. Results showed that the absorbance value (A value) of the MTT assay directly correlated with the number of human hepatoma cell lines SMMC7721. The result of orthogonal test was the number of 5×104 SMMC7721 cells/ml, culture period 6 h before adding HSS, concentration of HSS 100 ?g/ml, incubation time with HSS 36 h. Additionally, several experiments demonstrated the optimal combination of other conditions was 50 ?g MTT, incubation time for MTT 6 h, DMSO was used to dissolve the MTT formazan crystals and measured with ELISA scanner at 570 nm. The result of determining HSS bio-activity in vitro by optimized MTT protocol showed that sHSS bio-activity increased with the growth of protein dose, but decreased when it beyond a certain dose. The optimized MTT protocol was a sensitive, convenient and stable quantitative method to evaluate HSS bio-activity.

Long noncoding RNA DUXAP8 contributes to the progression of hepatocellular carcinoma via regulating miR‐422a/PDK2 axis

BackgroundHepatocellular carcinoma (HCC) is one of the most deadly cancer worldwide. Multiple long noncoding RNAs (lncRNAs) are recently identified as crucial oncogenic factors or tumor suppressors. In this study, we explored the functon and mechanism of lncRNA double homeobox A pseudogene 8 (DUXAP8) in the progression of HCC.MethodsExpression levels of DUXAP8 in HCC tissue samples were measured using qRT‐PCR. The association between pathological indexes and the expression of DUXAP8 was also analyzed. Human HCC cell lines SMMC‐7721 and QSG‐7701 were used in in vitro studies. CCK‐8 assay was used to assess the effect of DUXAP8 on HCC cell line proliferation. Scratch healing assay and Transwell assay were conducted to detect the effect of DUXAP8 on migration and invasion. Furthermore, dual‐luciferase reporter assay was used to confirm targeting relationship between miR‐422a and DUXAP8. Additionally, Western blot was used to detect the regulatory function of DUXAP8 on pyruvate dehydrogenase kinase 2 (PDK2).ResultsDUXAP8 expression HCC clinical samples was significantly increased and this was correlated with unfavorable pathological indexes. High expression of DUXAP8 was associated with shorter overall survival time of patients. Its overexpression remarkably facilitated the proliferation, metastasis, and epithelial‐mesenchymal transition of HCC cells. Accordingly, knockdown of it suppressed the malignant phenotypes of HCC cells. Overexpression of DUXAP8 significantly reduced the expression of miR‐422a by sponging it, but enhanced the expression of PDK2.ConclusionsDUXAP8 was a sponge of tumor suppressor miR‐422a in HCC, enhanced the expression of PDK2 indirectly, and functioned as an oncogenic lncRNA.

A dual cancer-specific recombinant adenovirus suppresses the growth of liver cancer cells in vivo and in vitro

Oncolytic virus therapy is emerging as important means in cancer treatment. In a previous study, we constructed a dual cancer-specific antitumor recombinant adenovirus, designating it Ad-apoptin-hTERTp-E1a (Ad-VT). This study aimed to investigate the anticancer potential of recombinant adenovirus Ad-apoptin-hTERTp-E1a (Ad-VT) in liver cancer. Crystal Violet staining and CCK-8 assays were used to analyse the inhibitory effect of recombinant adenovirus on human hepatoma cell line QGY-7703 and SMMC-7721. Ad-VT had a significant tumour killing inhibitory effect on QGY-7703 and SMMC-7721 cells that was both dose and a time dependent. Ad-VT-induced apoptosis of QGY-7703 cells was detected using Hoechst, Annexin V, and JC-1 staining, as well as western blotting. Recombinant adenovirus had a strong apoptosis-inducing effect on QGY-7703 cells, and killed QGY-7703 cells mainly through the mitochondrial apoptotic pathway. QGY-7703 cells invasion were detected using cell-scratch and Transwell assays. Recombinant adenovirus could significantly inhibit the invasion of QGY-7703 cells over a short period of time. The pGL4.51 plasmid was used to transfect QGY-7703 cells to construct tumour cells stably expressing luciferase (QGY-7703-LUC). The tumour inhibition effect of Ad-VT in vivo was subsequently confirmed by establishing a tumour-bearing nude mouse model. Ad-VT could effectively inhibit tumour growth and prolong survival of the mice. Recombinant adenovirus Ad-VT has the characteristics of tumour-specific replication and specific tumour killing, and could inhibit the growth of liver cancer QGY-7703 cells and promote their apoptosis.

Apatinib potentiates irradiation effect via suppressing PI3K/AKT signaling pathway in hepatocellular carcinoma

BackgroundLimited effective intervention for advanced hepatocellular carcinoma (HCC) is available. This study aimed to investigate the potential clinical utility of apatinib, a highly selective inhibitor of the vascular endothelial growth factor receptor-2 (VEGFR2) tyrosine kinase, as a radiosensitizer in the treatment of HCC.MethodsFour human HCC cell lines SMMC-7721, MHCC-97H, HCCLM3 and Hep-3B were treated with apatinib, irradiation or combination treatment. Colony formation assay, flow cytometry and nuclear γ-H2AX foci immunofluorescence staining were performed to evaluate the efficacy of combination treatment. RNA sequencing was conducted to explore the potential mechanism. The impact of combination treatment on tumor growth was assessed by xenograft mice models.ResultsColony formation assay revealed that apatinib enhanced the radiosensitivity of HCC cell lines. Apatinib suppressed repair of radiation-induced DNA double-strand breaks. Flow cytometry analysis showed that apatinib increased radiation-induced apoptosis. Apatinib radiosensitized HCC via suppression of radiation-induced PI3K/AKT pathway. Moreover, an in vivo study indicated apatinib combined with irradiation significantly decreased xenograft tumor growth.ConclusionsOur results indicate that apatinib has therapeutic potential as a radiosensitizer in HCC, and PI3K/AKT signaling pathway plays a critical role in mediating radiosensitization of apatinib.

MicroRNA-16 Via Twist1 Inhibits EMT Induced by PM2.5 Exposure in Human Hepatocellular Carcinoma.

Epidemiological study has confirmed that PM2.5 (particulate matter with an aerodynamic diameter less than 2.5 μm) is associated with the incidence and progression of human hepatocellular carcinoma (HCC). Accordingly, this study was undertaken to investigate the pro-metastatic effects of PM2.5 on human HCC cell line SMMC-7721 in vitro and to explore the underlying mechanisms. CCK-8 assay was performed to examine the effect of PM2.5 on the proliferation of SMMC-7721 cells; scratch wound assay and transwell matrigel system has been used to examine the effect of PM2.5 on the migration and invasion ability of SMMC-7721 cells; furthermore, effect of PM2.5 on epithelial mesenchymal transition (EMT) of SMMC-7721 cells were examined by examining the EMT markers vimentin, ɑ-smooth muscle actin (ɑ-SMA), and E-cadherin; furthermore, the roles of microRNA-16 (miR-16) and its target Twist1 in PM2.5 induced carcinogenic effects were also examined. Results of CCK-8 assay suggested that PM2.5 promoted the proliferation of SMMC-7721 cells in a dose and time dependent manner. PM2.5 also markedly promoted the migration and invasion ability of SMMC-7721 cells. Moreover, epithelial mesenchymal transition (EMT) was also triggered by PM2.5. On the other hand, microRNA-16 (miR-16) and its target Twist1 was found to be mediated by PM2.5, and miR-16 mimic could suppress the metastatic ability of SMMC-7721 cells exposure to PM2.5 via inversely regulating the expression of Twist1. Furthermore, dual Luciferase reporter assay confirmed the specifically binding of miR-16 to the predicted 3′-UTR of Twist1. The present study confirmed the pro-proliferative and pro-metastatic effect of PM2.5 on HCC cell line SMMC-7721. The possible mechanisms were EMT process induced by PM2.5 in SMMC-7721 cells, which was accompanied by a decrease in miR-16 and increase in Twist1 expression.

Knockdown of the Sonic Hedgehog (SHH) Gene Inhibits Proliferation of Hep3B and SMMC-7721 Hepatocellular Carcinoma Cells via the PI3K/Akt/PCK1 Signaling Pathway.

BackgroundThe PCK1 gene encodes phosphoenolpyruvate carboxykinase (PEPCK), which has been shown have a role in metabolic events in hepatocellular carcinoma (HCC). This study aimed to investigate the role of the SHH gene and its encoded protein, sonic hedgehog (SHH), in two human hepatocellular carcinoma (HCC) cell lines.Material/MethodsThe human HCC cell lines Hep3B and SMMC-7721 were cultured. Cells were transfected with plasmids carrying specific SHH gene short-hairpin RNA (shRNA) and negative control (NC) shRNA. The effects of knockdown of expression levels of the SHH gene were studied on cell survival, cell apoptosis, the cell cycle, gluconeogenesis, and the expression of PCK1. Anchorage-independent growth, a characteristic of transformed cells, was detected by the colony formation assay. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot were performed 24 h after transfection.ResultsKnockdown of expression levels of the SHH gene reduced cell proliferation and growth of HCC cells and induced cell apoptosis and G1 cell cycle arrest in both HCC cell lines. Knockdown of the SHH gene decreased the levels of glycolysis products and increased the production of glucose and reduced the phosphorylation of PI3K and Akt but induced the expression of PCK1.ConclusionsKnockdown of the SHH gene reduced cell survival of HCC cells by increasing apoptosis, reducing cell proliferation, inducing G1 cell cycle arrest, and restoring gluconeogenesis, and was associated with the inhibition of the PI3K/Akt axis and induced the expression of PCK1.

Two new 8-isopentenyl isoflavane derivatives from Astragalus dolichochaete diels

Astragali Radix (Huangqi) is one of the well-known traditional Chinese medicines and has been used in China for more than two thousand years. As a close species, Astragalus dolichochaete Diels was found to possess cytotoxicity during our preliminary study of Astragalus plants from Yunnan Province, China. To better understand the chemical foundation of the cytotoxicity, the major constituents of A. dolichochaete were studied. As a result, two new 8-isopentenyl isoflavane derivatives, dolichochaeteins A and B (1 and 2), together with ten known constituents were isolated, and their structures were elucidated by means of spectroscopy, mainly 1D and 2D NMR techniques. The cytotoxic activities were evaluated for the raw extract, ethyl acetate fraction, compound 2 and glyasperin H (3) against human gastric carcinoma cell line SGC-7901, human hepatoma cell line SMMC-7721 and human leukemic cell line K562. All the samples exhibited significant cytotoxicity.

Screening and characterization of a novel high-efficiency tumor-homing cell-penetrating peptide from the buffalo cathelicidin family.

Targeted delivery of antitumor drugs is especially important for tumor therapy. Cell-penetrating peptides (CPPs) have been shown to be very effective drug carriers for tumor therapy. However, most CPPs lack tumor cell specificity. Here, we identified a highly efficient CPP, CAT, from the newly identified buffalo-derived cathelicidin family, which exhibits a preferential binding capacity for multiple tumor cell lines and delivers carried drug molecules into cells. CAT showed an approximately threefold to sixfold higher translocation efficiency than some reported cell-penetrating antimicrobial peptides, including the well-known classical CPP TAT. Moreover, the delivery efficiency of CAT was greater in a variety of tested tumor cells than in normal cells, especially for the human hepatoma cell line SMMC-7721, for which delivery was 7 times more efficient than the normal human embryonic lung cell line MRC-5, according to fluorescent labeling experiment results. CAT was conjugated to the Momordica charantia-derived type-I ribosome-inactivating protein MAP 30, and the cytotoxicity of the MAP 30-CAT fusion protein in the tumor cell line SMMC-7721 was significantly enhanced compared with that of the unconjugated MAP 30. The IC50 value of MAP 30-CAT was approximately 83 times lower than the IC50 value of the original MAP 30. Interestingly, the IC50 value of MAP 30 alone for MRC-5 was approximately twofold higher than the value for SMMC-7721, showing a small difference. However, when MAP 30 was conjugated to CAT, the difference in IC50 values between the two cell lines was significantly increased by 38-fold. The results of the flow cytometric detection of apoptosis revealed that the increase in cytotoxicity after CAT conjugation was mainly caused by the increased induction of apoptosis by the fusion protein. These results suggest that CAT, as a novel tumor-homing CPP, has great potential in drug delivery applications in vivo and will be beneficial to the development of tumor therapeutics.

Psoralen inhibits malignant proliferation and induces apoptosis through triggering endoplasmic reticulum stress in human SMMC7721 hepatoma cells

BackgroundPsoralen is a coumarin-like and coumarin-related benzofuran glycoside, which is a commonly used traditional Chinese medicine to treat patients with kidney and spleen-yang deficiency symptom. Psoralen has been reported to show estrogen-like activity, antioxidant activity, osteoblastic proliferation accelerating activity, antitumor effects and antibacterial activity. However, the antitumor mechanism of psoralen is not fully understood. This study aimed to investigate the therapeutic efficacy of psoralen in human hepatoma cell line SMMC7721 and the mechanism of antitumor effects.ResultsPsoralen inhibited proliferation of SMMC7721 in a dose- and time-dependent manner, and promoted apoptosis. Further, psoralen activated the ER stress signal pathway, including the expansion of endoplasmic reticulum, increasing the mRNA levels of GRP78, DDIT3, ATF4, XBP1, GADD34 and the protein levels of GDF15, GRP78, IRE1α, XBP-1s in a time-dependent manner. Psoralen induces cell cycle arrest at G1 phase by enhancing CyclinD1 and reducing CyclinE1 expression. Moreover, TUDC couldn’t inhibit the psoralen-induced ER stress in SMMC7721 cells.ConclusionsPsoralen can inhibit the proliferation of SMMC7721 cells and induce ER stress response to induce cell apoptosis, suggesting that psoralen may represent a novel therapeutic option for the prevention and treatment hepatocellular carcinoma.

Mytoxin B and Myrothecine A Induce Apoptosis in Human Hepatocarcinoma Cell Line SMMC-7721 via PI3K/Akt Signaling Pathway.

Trichothecene macrolides comprise a class of valuable leading compounds in developing anticancer drugs, however, there are few reports concerning their anticancer mechanisms, especially the anticancer mechanism of the 10,13-cyclotrichothecane derivatives that are found mainly in symbiotic fungi. In vitro anticancer activity of two trichothecene macrolides mytoxin B and myrothecine A against the human hepatocarcinoma cell line SMMC-7721 was investigated in the present study. MTT assay showed that mytoxin B and myrothecine A inhibited the proliferation of SMMC-7721 cells in dose- and time-dependent manners. Annexin V-FITC/PI dual staining assay revealed that mytoxin B and myrothecine A both could induce SMMC-7721 cells apoptosis in a dose-dependent manner. The decreased expression level of anti-apoptotic protein Bcl-2 and the increased expression level of pro-apoptotic protein Bax were observed apparently in Western blot analysis. The reduced ratio of Bcl-2/Bax further confirmed the apoptosis-inducing effect of mytoxin B and myrothecine A on SMMC-7721 cells. Moreover, the expression levels of caspases-3, -8, and -9, and cleaved caspases-3, -8, and -9 were all upregulated in both mytoxin B and myrothecine A-treated cells in Western blot analysis, which indicated that both compounds might induce SMMC-7721 cells apoptosis through not only the death receptor pathway but also the mitochondrial pathway. Finally, mytoxin B and myrothecine A were found to reduce the activity of PI3K/Akt signaling pathway that was similar to the effect of LY294002 (a potent and specific PI3K inhibitor), suggesting that both mytoxin B and myrothecine A might induce SMMC-7721 cells apoptosis via PI3K/Akt pathway.

Identification and characterization of a T-2 toxin-producing Fusarium poae strain and the anti-tumor effect of the T-2 toxin on human hepatoma cell line SMMC-7721.

T-2 toxin, produced by Fusarium moulds, is a type A trichothecene mycotoxin which is known to inhibit protein synthesis and also reported to induce DNA lesions, potentially causing DNA fragmentation. T-2 toxin is a very potent cytotoxic toxin, which displays anti-tumor properties. Nevertheless, more studies are still needed to explore its antitumor mechanisms as well as its clinical application in cancer treatment. Here, we report the identification and characterization of a T-2 toxin produced by a Fusarium poae isolated from Jilin, Northeast China. 17 strains of Fusarium poae were screened for T-2 toxin-production and one strain with the highest yield was selected further studies. T-2 toxin produced by the selected Fusarium poae was isolated and purified by HPLC. Anticancer properties of the purified T-2 toxin were evaluated with human hepatoma cell SMMC-7721. The purified T-2 toxin inhibits the proliferation of SMMC-7721 cells and induces cell apoptosis. The mitochondrial membrane potential decreased and the intracellular ROS was up-regulated after T-2 treatment of the cells. Further studies revealed that T-2 treatment activates the intrinsic mitochondrial and MAPKs pathway. Our data provide insight into the promising application of the T-2 toxin in cancer treatment.