Alendronate-Anionic Clay Nanohybrid for Enhanced Osteogenic Proliferation and Differentiation.

BACKGROUND: Alendronate (AL), a drug for inhibiting osteoclast-mediated bone-resorption, was intercalated into an inorganic drug delivery nanovehicle, layered double hydroxide (LDH), to form a new nanohybrid, AL-LDH, with 1:1 heterostructure along the crystallographic C-axis. Based on the intercalation reaction strategy, the present AL-LDH drug delivery system (DDS) was realized with an enhanced drug efficacy of AL, which was confirmed by the improved proliferation and osteogenic differentiation of osteoblast-like cells (MG63). METHODS: The AL-LDH nanohybrid was synthesized by conventional ion-exchange reaction and characterized by powder X-ray diffraction (PXRD), high-resolution transmission electron microscopy (HR-TEM), and Fourier transform infrared (FT-IR) spectroscopy. Additionally, in vitro efficacy tests, such as cell proliferation and alkaline phosphatase (ALP) activity, were analyzed. RESULTS: The AL was successfully intercalated into LDH via ion-exchange reaction, and thus prepared AL-LDH DDS was X-ray single phasic and chemically well defined. The accumulated AL content in MG63 cells treated with the AL-LDH DDS nanoparticles was determined to be 10.6-fold higher than that within those treated with the intact AL after incubation for 1 hour, suggesting that intercellular permeation of AL was facilitated thanks to the hybridization with drug delivery vehicle, LDH. Furthermore, both in vitro proliferation level and ALP activity of MG63 treated with the present hybrid drug, AL-LDH, were found to be much more enhanced than those treated with the intact AL. This is surely due to the fact that LDH could deliver AL drug very efficiently, although LDH itself does not show any effect on proliferation and osteogenic differentiation of MG63 cells. CONCLUSION: The present AL-LDH could be considered as a promising DDS for improving efficacy of AL.

The suppressor of cytokine signaling 2 (SOCS2) inhibits tumor metastasis in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a common cause of cancer-related death worldwide, and its incidence continues to increase. However, the mechanism underlying the development and progression of HCC remains unknown. The suppressor of cytokine signaling 2 (SOCS2) is a member of the SOCS family and influences the carcinogenesis of multiple types of tumors, but the biological roles of SOCS2 in HCC remain unclear. In this study, we found that SOCS2 expression was reduced in HCC tissues compared with matched noncancerous liver tissues. Moreover, decreased SOCS2 expression was significantly associated with the presence of intrahepatic metastasis and high histological grade in HCC patients. Colony formation assays and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays demonstrated that overexpression of SOCS2 or knockdown of endogenous SOCS2 did not significantly affect cell proliferation and tumorigenicity in HCC cells in vitro and in vivo. However, SOCS2 overexpression significantly inhibited the migration and invasion of HCC cells in vitro and inhibited metastasis in vivo. Consistent with these findings, the knockdown of endogenous SOCS2 enhanced migration and invasion in HCC cells in vitro. Our study demonstrated that SOCS2 inhibited human HCC metastasis, and SOCS2 might provide a new potential therapeutic strategy for treating HCC.

Downregulation of FOXP1 Inhibits Cell Proliferation in Hepatocellular Carcinoma by Inducing G1/S Phase Cell Cycle Arrest.

Forkhead box P1 (FOXP1) belongs to a family of winged-helix transcription factors that are involved in the processes of cellular proliferation, differentiation, metabolism, and longevity. FOXP1 can affect cell proliferation and migratory ability in hepatocellular carcinoma (HCC) in vitro. However, little is known about the mechanism of FOXP1 in the proliferation of HCC cells. This study aimed to further explore the function of FOXP1 on the proliferation of HCC cells as well as the relevant mechanism involved. Western blot analysis, tumor xenograft models, and flow cytometry analysis were performed to elucidate the function of FOXP1 in the regulation of cell proliferation in human HCC. We observed that silencing FOXP1 significantly suppressed the growth ability of HCC cells both in vitro and in vivo. In addition, knockdown of FOXP1 induced G1/S phase arrest, and the expression of total and phosphorylated Rb (active type) as well as the levels of E2F1 were markedly decreased at 24 h; however, other proteins, including cyclin-dependent kinase (CDK) 4 and 6 and cyclin D1 did not show noticeable changes. In conclusion, downregulation of FOXP1 inhibits cell proliferation in hepatocellular carcinoma by inducing G1/S phase cell cycle arrest, and the decrease in phosphorylated Rb is the main contributor to this G1/S phase arrest.

Serum DLK1 is a potential prognostic biomarker in patients with hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the sixth most prevalent cancer and the third most frequent cause of cancer-related death in developing countries, especially in East Asia and South Africa, and the identification of new biomarkers for early diagnosis and prognosis is needed. Delta-like 1 homologue (Drosophila) (DLK1) is expressed in malignancies and promotes cancer cell stemness and tumourigenicity, which makes this molecule a potential target for therapies directed against cancer stem/progenitor cells. Here, we aimed to assess the predictive value of DLK1 as a diagnostic and prognostic biomarker in HCC. With this purpose, serum DLK1 levels were detected using an enzyme-linked immunosorbent assay (ELISA) in serum specimens from 397 HCC patients, 114 healthy individuals, 43 patients with chronic hepatitis B virus (HBV) infection and 24 cirrhotic liver patients with HBV infection, and the correlation between DLK1 levels and clinical features was evaluated. Our data showed that the serum DLK1 level was significantly higher in HCC patients than in healthy individuals or patients with chronic HBV infection (HBV carriers) (P < 0.05). Moreover, the serum DLK1 levels were positively correlated with tumour size and α-fetoprotein (AFP) levels, but not with gender, age, histological grade, HBV infection, intrahepatic metastasis or cirrhosis in HCC patients. Kaplan-Meier analysis indicated that higher DLK1 levels were associated with shorter survival in HCC patients. These results suggest that the serum levels of DLK1 may serve as a prognostic biomarker for HCC patients.

Farmland transfer contract performance and green production technology adoption among farmers: evidence from a survey of four provinces in China.

The popularization and use of green production technology is the key to promoting the sustainable development of agriculture. Exploring the path of promoting farmers' adoption of green production technology has important theoretical and practical significance for promoting the green transformation of agriculture and realizing the high-quality development of agriculture. This study examined the impact of contract performance on the adoption of green production technology in the farmland transfer market based on the characteristics of green production technology. An empirical test was conducted using survey data from Heilongjiang, Henan, Zhejiang, and Sichuan Provinces, which differ in agricultural resource endowment and economic development. The results demonstrated that farmland area was positively associated with the number of green production technologies adopted by farmers and use of straw returning, subsoiling, and soil testing for formula fertilization technology. Moreover, improving the performance rates of farmland transfer contracts strengthened the promotional effect of operation scale on green production technology adoption. Using either contract execution or dispute resolution rates to measure contract performance promoted the adoption of green production technology, particularly for large-scale farmers. Therefore, in the publicity and promotion of agricultural green production technology, the government should not only formulate differentiated promotion strategies and support policies based on resource endowment, but also focus on the construction of rural legal systems, which are helpful for building and maintaining a good farmland market order and performance environment, and create conditions for the durability and stability of technology adoption.

Changes in the composition of the fecal metabolome and gut microbiota contribute to intervertebral disk degeneration in a rabbit model.

PURPOSE: Lower back pain (LBP), mainly caused by intervertebral disk (IVD) degeneration (IDD), is widely prevalent worldwide and is a serious socioeconomic burden. Numerous factors may trigger this degenerative process, and microbial dysbiosis has recently been implicated as one of the likely causes. However, the exact relationship between IDD and the microbiome remains obscure. In this study, we investigated the gut microbiota composition and fecal metabolic phenotype and discussed the possible influences of microbiome dysbiosis on IDD. METHODS: Fecal DNA was extracted from 16 fecal samples (eight rabbit models with IDD and eight sex- and age-matched healthy controls) and analyzed by high-throughput 16S rDNA sequencing. The fecal samples were also analyzed by liquid chromatography-mass spectrometry-based metabolomics. Multivariate analyses were conducted for the relationship between the omics data and IDD, linear discriminant analysis effect size was employed for biomarker discovery. Moreover, the Kyoto Encyclopedia of Genes and Genomes (KEGG) database was used to annotate the differential metabolites. The potential correlation between differential gut microbiota and metabolites was then assessed. RESULTS: The 16S rDNA sequencing results showed that the ß-diversity of the gut microbiota was significantly different between the IDD and control groups, with distinct abundance levels of dominant genera. Moreover, 59 metabolites were significantly upregulated and 91 were downregulated in IDD rabbits versus the controls. The KEGG enrichment analysis revealed that the top pathways remarkably impacted by IDD were tyrosine metabolism, amino sugar and nucleotide sugar metabolism, benzoate degradation, ABC transporters, ascorbate and aldarate metabolism, pantothenate and CoA biosynthesis, and pyrimidine metabolism. The correlation analysis revealed that DL-tyrosine and N-acetylmuramic acid were associated with multiple differential bacterial genera, including Helicobacter and Vibrio, which may play important roles in the process of IVD degeneration. CONCLUSION: Our findings revealed that IDD altered gut microbiota and fecal metabolites in a rabbit model. The correlation analysis of microbiota and metabolome provides a deeper understanding of IDD and its possible etiopathogenesis. These results also provide a direction and theoretical basis for the clinical application of fecal transplantation, probiotics, and other methods to regulate gut microbiota in the treatment of LBP caused by IDD.

Pigment epithelium-derived factor promotes tumor metastasis through an interaction with laminin receptor in hepatocellular carcinomas.

Pigment epithelium-derived factor (PEDF) has complex functions in tumor metastasis, but little is known about the roles of PEDF and its receptors in hepatocellular carcinoma (HCC). Here we found that high expression of PEDF is associated with shorter overall survival in HCC patients. Forced expression of PEDF enhanced HCC cell aggressive behavior in vitro and in vivo, whereas silencing PEDF expression reduced migration and invasion. Furthermore, PEDF expression led to changes in cell morphology and the expression of epithelial-mesenchymal transition (EMT)-related markers via ERK1/2 signaling pathway, including the upregulation of N-cadherin and slug, and the downregulation of E-cadherin in HCC cells. Our results further showed that PEDF could interact with laminin receptor (LR) and LR knockdown attenuated PEDF-induced migration, invasion and the change of EMT-related markers. More importantly, in clinical HCC specimens, we found that PEDF expression was correlated with subcellular localization of LR, and that high expression of PEDF and positive expression of LR predicted a poor prognosis. In conclusion, our results demonstrate a novel functional role of PEDF/LR axis in driving metastasis through ERK1/2-mediated EMT in HCC and provided a promising prognostic marker in HCC.

Dysfunction of IKZF1/MYC/MDIG axis contributes to liver cancer progression through regulating H3K9me3/p21 activity.

MDIG is known to be overexpressed in many types of human cancers and has demonstrated predictive power in the prognosis of cancer, although the functions and mechanisms of MDIG in liver cancer, especially in hepatocellular carcinoma (HCC), are still unknown. In this study, we report that MDIG and MYC were negatively regulated by IKZF1. MDIG overexpression substantially promoted HCC cell proliferation, cell migration and spreading, whereas knockdown of MDIG would reverse above-mentioned effect. MDIG effects on tumour cell growth were further demonstrated in a tumour xenograft model. Moreover, MDIG had effects on the level of p21(CIP1/WAF1) via H3K9me3 expression in HCC. MDIG was also found to be closely related to the sorafenib resistance of HCC cells in vitro. Clinically, we found that MDIG was frequently overexpressed in human HCCs (69.7%; n=155) and was significantly associated with histological grade and hepatitis B virus infection. Our findings indicate that MDIG plays an important role in HCC progression via MDIG/H3K9me3/p21(CIP1/WAF1) signalling and serves as a potential therapeutic target.

Effect of the HBV whole-X gene on the expression of hepatocellular carcinoma associated proteins.

BACKGROUND: The hepatitis B virus (HBV) pre-X gene resides upstream of the HBV X gene, and together they form the HBV whole-X gene. Although it has been evident that the HBV whole-X protein is involved in the development of hepatocellular carcinoma, its biological role and molecular mechanism remain largely unknown. METHODS: In this study, we subcloned the HBV whole-X gene and constructed a HBV whole-X expressing vector. After transfection of the HBV whole-X gene into HL-7702 cells, the profile of the differential cellular protein composition in the cells was analyzed by using two-dimensional electrophoresis coupled to matrix-assisted laser desorption/ionization-time of flight mass spectrometry. RESULTS: The results showed that 18 major proteins were differentially expressed in the cells transfected with or without the HBV whole-X gene. The expression of these genes was further confirmed by reverse transcription-polymerase chain reaction and Western blot analysis. CONCLUSION: Our findings provide a new insight into the investigation of the pathological role that the HBV whole-X gene plays in the development of hepatocellular carcinoma and may lead to the design of novel strategies for the treatment of this disease.

The transcription factor FOXN3 inhibits cell proliferation by downregulating E2F5 expression in hepatocellular carcinoma cells.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death worldwide, and the mechanisms underlying the development of HCC remain to be elucidated. Forkhead box N3 (FOXN3) is an important member of the FOX family of transcription factors that plays an essential role in several cancers but has not been investigated in HCC. In this study, we demonstrate that FOXN3 is downregulated in human primary HCC tissues compared with their matched adjacent liver tissues. Functional tests of FOXN3 demonstrated that FOXN3 inhibits the proliferation of HCC cells in vitro and in vivo. Additionally, FOXN3 repressed the mRNA and protein expression of E2F5, a reported potential oncogene, by inhibiting the promoter activity of E2F5. Collectively, our findings indicate that FOXN3 functions as a tumor suppressor in HCC by downregulating the expression of E2F5.

Orosomucoid 2 inhibits tumor metastasis and is upregulated by CCAAT/enhancer binding protein ß in hepatocellular carcinomas.

Cancer metastasis is a complex process, and the incidence of metastasis is influenced by many biological factors. Orosomucoid 2 (ORM2) is an important glycoprotein that is mainly biosynthesized and secreted by hepatocytes. As an acute-phase protein, ORM2 likely plays important roles in anti-inflammation, immunomodulation and drug delivery. However, little is known regarding the function of ORM2 in hepatocellular carcinoma (HCC). In this study, we determined that ORM2 expression in HCC tissues was negatively associated with intrahepatic metastasis and histological grade. Moreover, the ectopic overexpression of ORM2 decreased HCC cell migration and invasion in vitro and intrahepatic metastasis in vivo, whereas silencing ORM2 expression resulted in increased tumor cell migration and invasion in vitro. The CCAAT/enhancer binding protein ß (C/EBPß) upregulated ORM2 expression, while only the LAP1/2 (C/EBPß isoforms) possessed transcription-promoting activity on the ORM2 promoter. Subsequently, we found that LAP1 repressed HCC cell migration and invasion via the induction of ORM2 expression. Consistently, the protein expression of C/EBPß was negatively associated with histological grade and positively correlated with ORM2 protein expression in HCC tissues. Collectively, our findings indicate that ORM2 is a functional downstream target of C/EBPß and functions as a tumor suppressor in HCC.