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Cellular and Molecular Gastroenterology... 2023Phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme of the de novo serine synthesis pathway (SSP), has been implicated in the carcinogenesis and metastasis...
BACKGROUND & AIMS
Phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme of the de novo serine synthesis pathway (SSP), has been implicated in the carcinogenesis and metastasis of hepatocellular carcinoma (HCC) because of its excessive expression and promotion of SSP. In previous experiments we found that SSP flux was diminished by knockdown of zinc finger E-box binding homeobox 1 (ZEB1), a stimulator of HCC metastasis, but the underlying mechanism remains largely unknown. Here, we aimed to determine how SSP flux is regulated by ZEB1 and the contribution of such regulation to carcinogenesis and progression of HCC.
METHODS
We used genetic mice with Zeb1 knockout in liver specifically to determine whether Zeb1 deficiency impacts HCC induced by the carcinogen diethylnitrosamine plus CCl. We explored the regulatory mechanism of ZEB1 in SSP flux using uniformly-labeled [C]-glucose tracing analyses, liquid chromatography-mass spectrometry, real-time quantitative polymerase chain reaction, luciferase report assay, and chromatin immunoprecipitation assay. We determined the contribution of the ZEB1-PHGDH regulatory axis to carcinogenesis and metastasis of HCC by cell counting assay, methyl thiazolyl tetrazolium (MTT) assay, scratch wound assay, Transwell assay, and soft agar assay in vitro, orthotopic xenograft, bioluminescence, and H&E assays in vivo. We investigated the clinical relevance of ZEB1 and PHGDH by analyzing publicly available data sets and 48 pairs of HCC clinical specimens.
RESULTS
We identified that ZEB1 activates PHGDH transcription by binding to a nonclassic binding site within its promoter region. Up-regulated PHGDH augments SSP flux to enable HCC cells to be more invasive, proliferative, and resistant to reactive oxygen species and sorafenib. Orthotopic xenograft and bioluminescence assays have shown that ZEB1 deficiency significantly impairs the tumorigenesis and metastasis of HCC, and such impairments can be rescued to a large extent by exogenous expression of PHGDH. These results were confirmed by the observation that conditional knockout of ZEB1 in mouse liver dramatically impedes carcinogenesis and progression of HCC induced by diethylnitrosamine/CCl, as well as PHGDH expression. In addition, analysis of The Cancer Genome Atlas database and clinical HCC samples showed that the ZEB1-PHGDH regulatory axis predicts poor prognosis of HCC.
CONCLUSIONS
ZEB1 plays a crucial role in stimulating carcinogenesis and progression of HCC by activating PHGDH transcription and subsequent SSP flux, deepening our knowledge of ZEB1 as a transcriptional factor in fostering the development of HCC via reprogramming the metabolic pathway.
Topics: Humans; Animals; Mice; Carcinoma, Hepatocellular; Liver Neoplasms; Phosphoglycerate Dehydrogenase; Diethylnitrosamine; Cell Line, Tumor; Carcinogenesis; Zinc Finger E-box-Binding Homeobox 1
PubMed: 37331567
DOI: 10.1016/j.jcmgh.2023.06.006 -
Zhongguo Ying Yong Sheng Li Xue Za Zhi... Nov 2022Three modeling methods were used to establish a mouse primary liver cancer model, and compared them to find a more optimal modeling method. Forty 15-day-old C3H/HeN...
Three modeling methods were used to establish a mouse primary liver cancer model, and compared them to find a more optimal modeling method. Forty 15-day-old C3H/HeN male mice were randomly divided into groups I-IV, 10 mice in each group. Group Ⅰ were not treated; Group Ⅱ were intraperitoneally injected with 25 mg/kg diethylnitrosamine (DEN) once; Group Ⅲ were intraperitoneally injected with 100 mg/kg DEN once; Group Ⅳ were intraperitoneally injected with 25 mg/kg DEN once and followed by another intraperitoneal injection of 100 mg/kg DEN at 42 days of age. The mortality of mice in each group was analyzed. At the 18th week of modeling, blood was collected from eyeballs after anesthesia, and liver was taken from abdominal cavity after neck was broken. The appearance of liver, the number of cancer nodules and the incidence of liver tumor were observed. The histopathological changes of liver were observed by HE staining. The serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were detected. At the 18th week of modeling, compared with the group I, serum levels of ALT and AST in groups II-IV were increased significantly (<0.05); The number of cancer nodules and the incidence of tumors in the surviving mice of groups III and IV were also increased significantly (<0.05). At the 18th week of modeling, no mice died in both groups I and II, and the incidence of liver cancer was 0%; The incidence of liver cancer in surviving mice in both groups III and IV was 100%, but the mortality rate of mice in group III was as high as 50%, and that in group IV was only 20%. C3H/HeN male mice can successfully establish a mouse liver cancer model by intraperitoneal injection of 25 mg/kg of DEN once at the age of 15 days and another intraperitoneal injection of 100 mg/kg of DEN once at the age of 42 days with short cycle and low mortality, which is an ideal method to establish a primary liver cancer model.
Topics: Male; Mice; Animals; Mice, Inbred C3H; Liver Neoplasms; Injections, Intraperitoneal; Alanine Transaminase; Disease Models, Animal
PubMed: 37308442
DOI: 10.12047/j.cjap.6367.2022.149 -
Cancer Cell International Jun 2023Malignant transformation from hepatic fibrosis to carcinogenesis may be a therapeutic target for hepatocellular carcinoma (HCC). The aim of this study was to evaluate...
BACKGROUND
Malignant transformation from hepatic fibrosis to carcinogenesis may be a therapeutic target for hepatocellular carcinoma (HCC). The aim of this study was to evaluate anti-cancer efficacy of Pien-Tze-Huang (PZH), and to investigate the underlying mechanisms by integrating transcriptional regulatory network analysis and experimental validation.
METHODS
A diethylnitrosamine (DEN)-induced HCC model in rats was established and used to evaluate the anti-cancer efficacy of PZH. After detecting a transcriptomic profiling, the "disease-related gene-drug effective target" interaction network was constructed, and the candidate targets of PZH against malignant transformation from hepatic fibrosis to HCC were identified and verified in vitro.
RESULTS
PZH effectively alleviated the pathological changes of hepatic fibrosis and cirrhosis, and inhibited tumor formation and growth in DEN-induced HCC rats. Additionally, the administration of PZH reduced the levels of various hepatic function-related serological indicators significantly. Mechanically, a ferroptosis-related SLC7A11-GSH-GPX4 axis might be one of potential targets of PZH against malignant transformation from hepatic fibrosis to HCC. Especially, high SLC7A11 expression may be associated with poor prognosis of HCC patients. Experimentally, the administration of PZH markedly increased the trivalent iron and ferrous ion, suppressed the expression levels of SLC7A11 and GPX4 proteins, and reduced the GSH/GSSG ratio in the liver tissues of DEN-induced HCC rats.
CONCLUSIONS
Our data offer an evidence that PZH may effectively improve the hepatic fibrosis microenvironment and prevent the occurrence of HCC through promoting ferroptosis in tumor cells via inhibiting the SLC7A11-GSH-GPX4 axis, implying that PZH may be a potential candidate drug for prevention and treatment of HCC at an early stage.
PubMed: 37280673
DOI: 10.1186/s12935-023-02946-2 -
Oncology Reports Jul 2023Lentinan (LNT) isolated from is a vital host defense potentiator previously utilized as an adjuvant in cancer therapy. The present study investigated the effect of LNT...
Lentinan (LNT) isolated from is a vital host defense potentiator previously utilized as an adjuvant in cancer therapy. The present study investigated the effect of LNT on the mouse hepatocellular carcinoma (HCC) cell line Hepa1‑6 and its possible mechanism. Mouse HCC apoptosis and its potential associated mechanism were then explored using and approaches. For approaches, the effect of LNT on the proliferation of Hepa1‑6 cells was investigated by Cell Counting Kit‑8 assay. Annexin V‑FITC staining and flow cytometry were applied to explore HCC apoptosis. Western blotting was used to analyze related proteins, such as EGR1, phosphatase and tensin homolog (PTEN), phosphorylated protein kinase B (p‑Akt), protein kinase B (Akt), B lymphocyte‑2 (Bcl‑2), Bcl2 family‑associated X protein (Bax), etc. Cellular immunofluorescence staining was employed to assess the localization and expression of EGR1 and PTEN in nuclear and cytoplasmic fractions of Hepa1‑6 cells. The association between and was explored by overexpression in cell lines. For methods, a mouse model of diethylnitrosamine (DEN)‑induced primary liver cancer was established using C57BL/6 mice to investigate the inhibitory effect of LNT on liver cancer. Histopathology of liver tissue from mice was detected by hematoxylin‑eosin staining and immunohistochemical assay. and results showed that LNT can inhibit the proliferation and promote the apoptosis of mouse HCC cells. Besides, LNT increased the expression of EGR1 in Hepa1‑6 cells, which is translocated to the nucleus to function as a transcriptional factor. EGR1 then activates the expression of the tumor suppressor PTEN, thereby inhibiting the activation of the AKT signaling pathway. These data revealed a novel anti‑tumor mechanism by which LNT can induce apoptosis to inhibit mouse HCC progression through the EGR1/PTEN/AKT axis. These results provide a scientific basis for the potential use of LNT in drug development and clinical applications associated with primary liver cancer.
Topics: Mice; Animals; Proto-Oncogene Proteins c-akt; Carcinoma, Hepatocellular; Lentinan; Liver Neoplasms; Cell Line, Tumor; Mice, Inbred C57BL; Mice, Inbred Strains; Signal Transduction; Apoptosis; PTEN Phosphohydrolase; Cell Proliferation; Early Growth Response Protein 1
PubMed: 37264970
DOI: 10.3892/or.2023.8579 -
Sorafenib Alleviates Inflammatory Signaling of Tumor Microenvironment in Precancerous Lung Injuries.Pharmaceuticals (Basel, Switzerland) Feb 2023According to population-based studies, lung cancer is the prominent reason for cancer-related mortality worldwide in males and is also rising in females at an alarming...
According to population-based studies, lung cancer is the prominent reason for cancer-related mortality worldwide in males and is also rising in females at an alarming rate. Sorafenib (SOR), which is approved for the treatment of hepatocellular carcinoma and renal cell carcinoma, is a multitargeted protein kinase inhibitor. Additionally, SOR is the subject of interest for preclinical and clinical trials in lung cancer. This study was designed to assess in vivo the possible effects of sorafenib (SOR) in diethylnitrosamine (DEN)-induced lung carcinogenesis and examine its probable mechanisms of action. A total of 30 adult male rats were divided into three groups (1) control, (2) DEN, and (3) DEN + SOR. The chemical induction of lung carcinogenesis was performed by injection of DEN intraperitoneally at 150 mg/kg once a week for two weeks. The DEN-administered rats were co-treated with SOR of 10 mg/kg by oral gavage for 42 alternate days. Serum and lung tissue samples were analyzed to determine SRY-box transcription factor 2 (SOX-2) levels. The tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) levels were measured in lung tissue supernatants. Lung sections were analyzed for cyclooxygenase-2 (COX-2) and c-Jun N-terminal kinase (JNK) histopathologically. In addition, cyclooxygenase-2 (COX-2) and c-Jun N-terminal kinase (JNK) were analyzed by immunohistochemistry and immunofluorescence methods, respectively. SOR reduced the level of SOX-2 that maintenance of cancer stemness and tumorigenicity, and TNF-α and IL-1β levels. Histopathological analysis demonstrated widespread inflammatory cell infiltration, disorganized alveolar structure, hyperemia in the vessels, and thickened alveolar walls in DEN-induced rats. The damage was markedly reduced upon SOR treatment. Further, immunohistochemical and immunofluorescence analysis also revealed increased expression of COX-2 and JNK expression in DEN-intoxicated rats. However, SOR treatment alleviated the expression of these inflammatory markers in DEN-induced lung carcinogenesis. These findings suggested that SOR inhibits DEN-induced lung precancerous lesions through decreased inflammation with concomitant in reduced SOX-2 levels, which enables the maintenance of cancer stem cell properties.
PubMed: 37259369
DOI: 10.3390/ph16020221 -
European Journal of Nuclear Medicine... Aug 2023Hepatocellular carcinoma (HCC) is a highly vascularized solid carcinoma and tumor vessel-targeted molecular imaging might be effective for early diagnosis of HCC....
PURPOSE
Hepatocellular carcinoma (HCC) is a highly vascularized solid carcinoma and tumor vessel-targeted molecular imaging might be effective for early diagnosis of HCC. Herein, we developed a novel trimeric affibody (Z) with highly specific binding to the platelet-derived growth factor receptor beta (PDGFRβ). The aim of this study is to evaluate the feasibility of Ga-radiolabeled Z ([Ga]Ga-DOTA-Z) as PET tracer for diagnosis of HCC.
METHODS
The bioinformatics analysis of clinical database and immunoblotting of clinical specimens were performed to validate the potential of PDGFRβ as HCC biomarker. The trimeric affibody Z was conjugated with DOTA-NHS-ester and radiolabeled with Ga to produce [Ga]Ga-DOTA-Z conjugate. Immunoreactivity and specific uptake of [Ga]Ga-DOTA-Z were assessed by dose-dependent cell binding, autoradiography, and biodistribution analysis. [Ga]Ga-DOTA-Z PET/CT scanning of diethylnitrosamine (DEN)-induced primary HCC rats and a rare case of idiopathical HCC rhesus monkey was performed to evaluate the imaging capability and radiation dosimetry of [Ga]Ga-DOTA-Z in vivo.
RESULTS
Excessive PDGFRβ was validated as a representative biomarker of HCC neovascularization. The radiolabeling of [Ga]Ga-DOTA-Z was achieved at more than 95% radiochemical yield. In vitro assays showed specific uptake of [Ga]Ga-DOTA-Z in HCC tumor vessels by autoradiography. Animal PET/CT imaging with [Ga]Ga-DOTA-Z successfully visualized the tumor lesions in primary HCC rats and rhesus monkey, and indicated radiation absorbed dose of 2.03E-02 mSv/MBq for each scanning.
CONCLUSIONS
Our results demonstrated that [Ga]Ga-DOTA-Z conjugate could be applied as a promising PET tracer for early diagnosis of hepatocellular carcinoma.
Topics: Rats; Animals; Positron Emission Tomography Computed Tomography; Carcinoma, Hepatocellular; Gallium Radioisotopes; Tissue Distribution; Macaca mulatta; Cell Line, Tumor; Liver Neoplasms; Positron-Emission Tomography; Biomarkers
PubMed: 37256321
DOI: 10.1007/s00259-023-06260-x -
Frontiers in Oncology 2023Babao Dan (BBD) is a traditional Chinese medicine that has been widely used as a complementary and alternative medicine to treat chronic liver diseases. In this study,...
BACKGROUND
Babao Dan (BBD) is a traditional Chinese medicine that has been widely used as a complementary and alternative medicine to treat chronic liver diseases. In this study, we aimed to observe the effect of BBD on the incidence of diethylnitrosamine (DEN)-initiated hepatocellular carcinoma formation in rats and explored its possible mechanism.
METHODS
To verify this hypothesis, BBD was administrated to rats at a dose of 0.5g/kg body weight per two days from the 9th to 12th week in HCC-induced by DEN. Liver injury biomarkers and hepatic inflammatory parameters were evaluated by histopathology as well as serum and hepatic content analysis. We applied immunohistochemical analysis to investigate the expression of CK-19 and SOX-9 in liver tissues. The expression of TLR4 was determined by immunohistochemical, RT-PCR, and western blot analysis. Furthermore, we also detected the efficacy of BBD against primary HPCs neoplastic transformation induced by LPS.
RESULTS
We observed that DEN could induce hepatocarcinogenesis, and BBD could obviously decrease the incidence. The biochemical and histopathological examination results confirmed that BBD could protect against liver injury and decrease inflammatory infiltration. Immunohistochemistry staining results showed that BBD could effectively inhibit the ductal reaction and the expression of TLR4. The results showed that BBD-serumcould obviously inhibit primary HPCs neoplastic transformation induced by regulating the TLR4/Ras/ERK signaling pathway.
CONCLUSION
In summary, our results indicate that BBD has potential applications in the prevention and treatment of HCC, which may be related to its effect on hepatic progenitor cells malignant transformation via inhibiting the TLR4/Ras/ERK signaling pathway.
PubMed: 37251918
DOI: 10.3389/fonc.2023.1073859 -
Genomics Jul 2023Differences in the gut microbiota and metabolic processes between males and females may explain differences in the risk of liver injury; however, the sex-specific...
Differences in the gut microbiota and metabolic processes between males and females may explain differences in the risk of liver injury; however, the sex-specific effects of antibiotics and probiotics on these relationships are not clear. We evaluated differences in the gut microbiota and the risk of liver injury between male and female rats after the oral administration of antibiotics or probiotics followed by a period of diethylnitrosamine treatment to chemically induce liver injuryusing high-throughput sequencing of fecal microbiota combined with histological analyses of liver and colon tissues. Our results suggest that the ratio of gram-positive to gram-negative bacteria in kanamycin-treated rats was significantly higher than that of other groups, and this difference persisted for the duration of the experiment. Antibiotics significantly changed the composition of the gut microbiota of experimental rats. Clindamycin caused more diethylnitrosamine-induced damage to livers of male rats. Probiotics did not influencethe gut microbiota; however, they hadprotective effects against liver injury induced by diethylnitrosamine, especially in female rats. These results strengthen our understanding of sex differences in the indirect effects of antibiotics or probiotics on metabolism and liver injury in hosts via the gut microbiota.
Topics: Female; Male; Rats; Animals; Anti-Bacterial Agents; Gastrointestinal Microbiome; Chemical and Drug Induced Liver Injury, Chronic; Diethylnitrosamine; Sex Characteristics; Probiotics
PubMed: 37217087
DOI: 10.1016/j.ygeno.2023.110647 -
Intestinal microecology in mice bearing diethylnitrosamine-induced primary hepatocellular carcinoma.Zhejiang Da Xue Xue Bao. Yi Xue Ban =... Aug 2022To explore the characteristics of intestinal microecology in hepatocellular carcinoma (HCC) model mice.
OBJECTIVE
To explore the characteristics of intestinal microecology in hepatocellular carcinoma (HCC) model mice.
METHODS
C57BL/6 male mice aged 2 weeks were divided into normal control group and HCC model group. Mice in HCC model group were exposed to a single intraperitoneal injection of diethylnitrosamine (DEN) 2 weeks after birth; the surviving mice were intraperitoneally injected with 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP), once every 2 weeks for 8 times starting from the 4 week after birth. Mice in each group were randomly selected and sacrificed at 10 , 18 and 32 weeks after birth, respectively, the liver tissue samples were obtained for histopathological examination. At the 32 week, all mice in both groups were sacrificed and the feces samples were collected under sterile conditions right before the sacrifice. The feces samples were sequenced for the V3-V4 hypervariable regions of the 16S rRNA gene, and the species abundance, flora diversity and phenotype, as well as flora correlation and functional prediction were analyzed.
RESULTS
Alpha diversity analysis showed that all Good's coverage reached the maximum value of 1.00, and the differences in the Observed features, Chao1 index, Shannon index and Simpson index of the intestinal flora of mice between normal control group and HCC model group were all statistically significant (all <0.05). Beta diversity analysis showed that PCoA based on weighted or unweighted Unifrac distances all yielded >0, confirming that the intra-group differences of the samples were less than the inter-group differences; the trend of separation between the two groups was significant ( <0.05). Bacteroidetes, Firmicutes, Actinobacteria and Patescibacteria were the dominant taxa at the phylum level in both normal control group and HCC model group. However, compared with normal control group, the abundance of Bacteroidetes in HCC model group was significantly decreased ( <0.01), while the abundance of Patescibacteria was significantly increased ( <0.05). Moreover, the dominant taxa at the genus level in normal control group mainly included , , , , . The dominant taxa at the genus level in HCC model group mainly included , , , , . There were 30 genera with statistically significant differences in relative abundance at the genus level between the two groups (all <0.05). LEfSe analysis of the intestinal flora of mice in the two groups revealed a total of 14 multi-level differential taxa (all <0.05, LDA score>4.0), which were mainly enriched in Bacteroidetes. The enrichment of 10 differential taxa including Bacteroidetes, Bacteroidia, Bacteroidales, Muribaculaceae, etc. were found in normal control group, and the enrichment of 4 differential taxa including , , etc. were found in HCC model group. There were both positive and negative correlations between the dominant intestinal genera in normal control group (|rho|>0.5, <0.05), while the correlations of the dominant intestinal genera in HCC model group, being less complex than that in normal control group, were all positive. The relative abundance of gram positive and mobile element containing in the intestinal flora of mice in HCC model group was significantly up-regulated compared with normal control group (both <0.05), while that of gram negative ( <0.05) and pathogenic potential ( <0.05) was significantly down-regulated. The metabolic pathways of the intestinal flora in the two groups were significantly different. For instance, 18 metabolic pathways were enriched in normal control group (all <0.005), including those related to energy metabolism, cell division, nucleotide metabolism, etc., while 12 metabolic pathways were enriched in HCC model group (all <0.005), including those related to energy metabolism, amino acid metabolism, carbohydrate metabolism, etc. Conclusions: The amount of intestinal flora in DEN-induced primary HCC model mice decreased, and the composition, correlation, phenotype and function of the intestinal flora in mice were significantly altered. Bacteroidetes at the phylum level, as well as several microbial taxa at the genus level such as , , and could be closely associated with DEN-induced primary HCC in mice.
Topics: Male; Animals; Mice; Carcinoma, Hepatocellular; Diethylnitrosamine; RNA, Ribosomal, 16S; Mice, Inbred C57BL; Liver Neoplasms; Bacteria
PubMed: 37202098
DOI: 10.3724/zdxbyxb-2022-0283 -
Biomedicines Mar 2023A safe and effective treatment for liver cancer is still elusive despite all attempts. Biomolecules produced from natural products and their derivatives are potential...
Streptomyces Bioactive Metabolites Prevent Liver Cancer through Apoptosis, Inhibiting Oxidative Stress and Inflammatory Markers in Diethylnitrosamine-Induced Hepatocellular Carcinoma.
A safe and effective treatment for liver cancer is still elusive despite all attempts. Biomolecules produced from natural products and their derivatives are potential sources of new anticancer medications. This study aimed to investigate the anticancer potential of a sp. bacterial extract against diethylnitrosamine (DEN)-induced liver cancer in Swiss albino mice and explore the underlying cellular and molecular mechanisms. The ethyl acetate extract of a sp. was screened for its potential anticancer activities against HepG-2 using the MTT assay, and the IC was also determined. Gas chromatography-mass spectrometric analysis was used to identify the chemical constituents of the extract. Mice were administered DEN at the age of 2 weeks, and from week 32 until week 36 (4 weeks), they received two doses of extract (25 and 50 mg/kg body weight) orally daily. The extract contains 29 different compounds, according to the GC-MS analysis. The rate of HepG-2 growth was dramatically reduced by the extract. In the mice model. extract considerably lessened the negative effects of DEN on liver functions at both doses. Alpha-fetoprotein (AFP) levels were significantly ( < 0.001) decreased, and P53 mRNA expression was increased, both of which were signs that extract was suppressing carcinogenesis. This anticancer effect was also supported by histological analysis. extract therapy additionally stopped DEN-induced alterations in hepatic oxidative stress and enhanced antioxidant activity. Additionally, extract reduced DEN-induced inflammation, as shown by the decline in interleukin-1 beta (IL-1β) and tumor necrosis factor-alpha (TNF-α) levels. Additionally, the extract administration dramatically boosted Bax and caspase-3 levels while decreasing Bcl-2 expressions in the liver according to the Immunohistochemistry examination. In summary, extract is reported here as a potent chemopreventive agent against hepatocellular carcinoma through multiple mechanisms, including inhibiting oxidative stress, cell apoptosis, and inflammation.
PubMed: 37189672
DOI: 10.3390/biomedicines11041054