Relationship between apoptosis and invasive and metastatic potential of hepatocellular carcinoma

Objective: To study relationship between apoptosis and invasive and metastatic potential of hepatocellu- lar carcinoma (HCC). Methods: Apoptotic rate (AR), proliferative index (PI) and S-phase fraction (SPF) were measured by flow cytometry, and p170, p21 and nucleoside diphosphate kinase (ndpk) by strept avidin-biotin complex immunohistochemical technique in 57 pa- tients with HCC. Results: In this group, AR was 1.77% ±0.19%, SPF 12.55% ±0.68%, and PI 20.91% ±1.12% (r =-0.173). p170, p21 and ndpk positive rates were 61.36%, 68, 18%, 52.27% respectively in patients with a mean AR of ≤1.77%, and 23.08%, 38.46%, 84.62% respectively in patients with a mean AR of >1.77% (all P<0.05). In patients with positive tumor invasiveness and metastasis, nd- pk (+) was 43.75%, p21 (+) 75.00%, p170 (+) 65.63%, AR 1.12% ±0. 16%, PI 23.78% ±1.48%, and SPF 13.90 % ±0.99 %. In patients with negative invasiveness and metastasis, however, ndpk (+) was 80.00%, p21 (+) 44.00%, p170 (+) 36.00%, AR 2,32%±0.52%, PI 18.53% ±0.82% and SPF 11.43% ±0.70%. Conclusion: Apoptosis of HCC is negatively correla- ted with its invasive and metastatic potential or other factors as proliferative activity, p21, p170 and ndpk.

Magnetic Resonance Gd？RGD Imaging Study of Hepatocellular Carcinoma with High and Low Metastatic Potential before and after Human Bone Marrow？derived Mesenchymal Stem Cell Intervention

Background： Biotherapy based on human bone marrow-derived mesenchymal stem cells （BMSCs） is currently the focus of research, especially in the field of autologous stem cell transplantation. A novel type of metastasis-associated magnetic resonance （MR） molecular imaging probe was constructed, and the changes in metastasis and proliferation of hepatocellular carcinoma （HCC） before and after BMSC intervention were observed through MR imaging （MRI）. Methods：Metastasis-associatedMRmolecularimagingprobe,integrin αvβ3 ligandcRGD-PEG-DGL-DTPA-Gd （Gd-RGD）,wereconstructed. After human BMSC intervention was performed for 6weeks, tumor weight inhibition rates were calculated, and the RGD molecular probe was imaged through MRI with molecular imaging agent Gd-DTPAas control.The signal-to-noise ratio （SNR） and contrast-to-noise ratio （CNR） in the MRI experiment were used as semi-quantitative indicators. Polymerase chain reaction method was performed to detect proliferation- and metastasis-associated indicators, transforming growth factor β-1 （TGFβ1）, osteopontin （OPN）, and integrin subunit αv and β3. Results： The highest tumor weight inhibition rates were observed 3 weeks after the BMSC transplantation. The MR Gd-RGD in the HCC tissues after the BMSC intervention showed less enhancement than Gd-DTPA. The Gd-DTPAMRI of control group had higher SNR and CNR than Gd-RGD MRI in the experimental groups （P 〈 0.05）. For high metastatic potential hepatocellular carcinoma （MHCC97-H）, significant differenceswereobservedintheSNRsandCNRsofGd-RGDMRIbeforeandaftertheBMSCintervention（P〈0.05）.Forlowmetastaticpotential hepatocellular carcinoma （MHCC97-L）, the CNRs of Gd-RGD MRI were statistically different before and after BMSC intervention （P 〈 0.05）. With regard to MHCC97-H, OPN, β3, and TGFβ1 expression significantly decreased after BMSC intervention （P 〈 0.05）. In MHCC97-L and OPN, β3, TGFβ1, and αv expression after BMSC intervention decreased, and the difference was statistically significant （P 〈 0.05）. Conclusions： The CNR index of MRI is a good indicator for distinguishing high- and low-metastatic potential HCC tissues.After BMSC transplantation of MRI through the two kinds of tracer, the SNR and CNR indexes can distinguish two kinds of high and low metastatic potential HCC tissues, and Gd-RGD imaging is more suitable in distinguishing the metastatic potential changes through BMSC intervention.

Differential thymosin β10 expression levels and actin filament organization in tumor cell lines with different metastatic potential

Background To investigate the differential expression levels of thymosin β10 (Tβ10) and the corresponding changes of actin filament organization in human tumor cell lines with different metastatic potential.Methods Four groups of nine human tumor cell lines with different metastatic potential were analyzed for the amount of Tβ10 mRNAs by Northern blot and for their peptide expression levels by immunohistochemistry. The filamentous actin (F-actin) was observed by staining of TRITC-phalloidin to detect changes in actin organization.Results In comparison with non-/weakly metastatic counterparts, Tβ10 was upregulated in highly metastatic human lung cancer, malignant melanoma and breast cancer cell lines. Staining of TRITC-phalloidin revealed less actin bundles, a fuzzy network of shorter filaments and some F-actin aggregates in the highly metastatic tumor cells. Meanwhile, the actin filaments were robust and orderly arranged in the non-/weakly metastatic cancer cell lines.Conclusion Tβ10 levels correlate positively with the metastatic capacity in human tumors currently examined. The increasing metastatic potential of tumor cells is accompanied by a loss of F-actin, poorly arranged actin skeleton organizations and presence of F-actin aggregates. There is a consistent correlation between the elevated Tβ10 expression and the disrupted actin skeleton.

Whole-exome mutational landscape of metastasis in patient-derived hepatocellular carcinoma cells

In order to explore the genomic basis for liver cancer metastasis,whole-exome sequencing(WES)was performed on patient-derived hepatocellular carcinoma(HCC)cell lines with differential metastatic potentials and analyzed their clonal evolution relationships.An evolutionary tree based on genomic single nucleotide polymorphism(SNP)was constructed in MegaX software.The WES data showed that the average percentage of heterogeneous mutations in each HCC cell lines was 16.55%(range,15.38%e18.17%).C:G>T:A and T:A>C:G somatic transitions were the two most frequent substitutions.In these metastatic HCC cell lines,non-silent gene mutations were found in 21.88%of known driver genes and 10 classical signaling pathways.The protein interaction network was constructed by STRING,and hub genes were found in the shared trunk mutation genes and the heterogeneous branch mutations respectively.In cBioPortal database,some of the selected hub genes were found to be associated with poor overall survival(OS)of HCC patients.Among the mutated HCC driver genes,a novel KEAP1 mutation with a homozygous frameshift truncation at the c-terminal Nrf2 binding region was detected and verified in MHCC97-H and HCC97LM3 cells.In conclusion,WES data demonstrate that HCC cell lines from tumor biopsy specimens of the same patient have obtained different metastatic potentials through repeated selection in rodents in vivo,and they do indeed have a genetic relationship at the genomic level.