Establishment of a hydrodynamic delivery system in ducks.

Chronic hepatitis B virus (HBV) poses a significant global health challenge as it can lead to acute or chronic liver disease and hepatocellular carcinoma (HCC). To establish a safety experimental model, a homolog of HBV-duck HBV (DHBV) is often used for HBV research. Hydrodynamic-based gene delivery (HGD) is an efficient method to introduce exogenous genes into the liver, making it suitable for basic research. In this study, a duck HGD system was first constructed by injecting the reporter plasmid pLIVE-SEAP via the ankle vein. The highest expression of SEAP occurred when ducks were injected with 5 µg/mL plasmid pLIVE-SEAP in 10% bodyweight volume of physiological saline for 6 s. To verify the distribution and expression of exogenous genes in multiple tissues, the relative level of foreign gene DNA and ß-galactosidase staining of LacZ were evaluated, which showed the plasmids and their products were located mainly in the liver. Additionally, ß-galactosidase staining and fluorescence imaging indicated the delivered exogenous genes could be expressed in a short time. Further, the application of the duck HGD model on DHBV treatment was investigated by transferring representative anti-HBV genes IFNα and IFNγ into DHBV-infected ducks. Delivery of plasmids expressing IFNα and IFNγ inhibited DHBV infection and we established a novel efficient HGD method in ducks, which could be useful for drug screening of new genes, mRNAs and proteins for anti-HBV treatment.

CTRP1 prevents high fat diet-induced obesity and improves glucose homeostasis in obese and STZ-induced diabetic mice.

BACKGROUND: C1q/tumor necrosis factor-related protein 1 (CTRP1) is an adipokine secreted by adipose tissue, related to chondrocyte proliferation, inflammation, and glucose homeostasis. However, the therapeutic effects on metabolic disorders and the underlying mechanism were unclear. Here, we investigated the functions and mechanisms of CTRP1 in treating obesity and diabetes. METHODS: The plasmid containing human CTRP1 was delivered to mice by hydrodynamic injection, which sustained expression of CTRP1 in the liver and high protein level in the blood. High-fat diet (HFD) fed mice and STZ-induced diabetes model were used to study the effects of CTRP1 on obesity, glucose homeostasis, insulin resistance, and hepatic lipid accumulation. The lipid accumulation in liver and adipose tissue, glucose tolerance, insulin sensitivity, food intake, and energy expenditure were detected by H&E staining, Oil-Red O staining, glucose tolerance test, insulin tolerance test, and metabolic cage, respectively. The metabolic-related genes and signal pathways were determined using qPCR and western blotting. RESULTS: With high blood circulation, CTRP1 prevented obesity, hyperglycemia, insulin resistance, and fatty liver in HFD-fed mice. CTRP1 also improved glucose metabolism and insulin resistance in obese and STZ-induced diabetic mice. The metabolic cage study revealed that CTRP1 reduced food intake and enhanced energy expenditure. The mechanistic study demonstrated that CTRP1 upregulated the protein level of leptin in blood, thermogenic gene expression in brown adipose tissue, and the gene expression responsible for lipolysis and glycolysis in white adipose tissue (WAT). CTRP1 also downregulated the expression of inflammatory genes in WAT. Overexpression of CTRP1 activated AMPK and PI3K/Akt signaling pathways and inhibited ERK signaling pathway. CONCLUSION: These results demonstrate that CTRP1 could improve glucose homeostasis and prevent HFD-induced obesity and fatty liver through upregulating the energy expenditure and reducing food intake, suggesting CTRP1 may serve as a promising target for treating metabolic diseases.

Gdf11 gene transfer prevents high fat diet-induced obesity and improves metabolic homeostasis in obese and STZ-induced diabetic mice.

BACKGROUND: The growth differentiation factor 11 (GDF11) was shown to reverse age-related hypertrophy on cardiomyocytes and considered as anti-aging rejuvenation factor. The role of GDF11 in regulating metabolic homeostasis is unclear. In this study, we investigated the functions of GDF11 in regulating metabolic homeostasis and energy balance. METHODS: Using a hydrodynamic injection approach, plasmids carrying a mouse Gdf11 gene were delivered into mice and generated the sustained Gdf11 expression in the liver and its protein level in the blood. High fat diet (HFD)-induced obesity was employed to examine the impacts of Gdf11 gene transfer on HFD-induced adiposity, hyperglycemia, insulin resistance, and hepatic lipid accumulation. The impacts of GDF11 on metabolic homeostasis of obese and diabetic mice were examined using HFD-induced obese and STZ-induced diabetic models. RESULTS: Gdf11 gene transfer alleviates HFD-induced obesity, hyperglycemia, insulin resistance, and fatty liver development. In obese and STZ-induced diabetic mice, Gdf11 gene transfer restores glucose metabolism and improves insulin resistance. Mechanism study reveals that Gdf11 gene transfer increases the energy expenditure of mice, upregulates the expression of genes responsible for thermoregulation in brown adipose tissue, downregulates the expression of inflammatory genes in white adipose tissue and those involved in hepatic lipid and glucose metabolism. Overexpression of GDF11 also activates TGF-ß/Smad2, PI3K/AKT/FoxO1, and AMPK signaling pathways in white adipose tissue. CONCLUSIONS: These results demonstrate that GDF11 plays an important role in regulating metabolic homeostasis and energy balance and could be a target for pharmacological intervention to treat metabolic disease.

MicroRNA-1254 exerts oncogenic effects by directly targeting RASSF9 in human breast cancer.

MicroRNAs (miRNAs or miRs) play crucial roles in human breast cancer. Although miR-1254 has been shown to have oncogenic activity in several cancer types, its biological function in breast cancer and its mechanisms of action remain unclear. In this study, we investigated the role of miR-1254 in human breast cancer and sought to elucidate the relevant underlying mechanisms. We found that miR-1254 expression was markedly increased in breast cancer tissues and cell lines. Additionally, miR-1254 overexpression accelerated breast cancer cell proliferation, cell cycle G1-S phase transition and inhibited apoptosis. Nevertheless, the inhibition of miR-1254 suppressed cell proliferation and induced apoptosis. Further analyses revealed that miR-1254 expression negatively correlated with RASSF9 expression in breast cancer tissues. We verified that RASSF9 was a direct target of miR-1254 using a luciferase reporter assay. The overexpression of miR-1254 reduced the RASSF9 mRNA and protein levels, and the suppression of miR-1254 promoted RASSF9 expression. Notably, the knockdown or overexpression of RASSF9 corroborated the biological effects observed upon miR-1254 overexpression or inhibition. Taken together, these results demonstrate that miR-1254 accelerates breast cancer cell growth by activating the AKT signaling pathway and suppresses apoptosis by inhibiting p53 expression through the targeting of RASSF9. The data indicate that miR-1254 plays a crucial role in human breast cancer, and may represent a novel therapeutic target for this malignancy.

Combination of S-1 and gefitinib increases the sensitivity to radiotherapy in lung cancer cells.

OBJECTIVE: To investigate the potential radiosensitization of S-1 and gefitinib in human non-small cell lung cancer (NSCLC) in vitro and in vivo. METHODS: The impact of radiation, 5-fluorouracil (5-Fu), and gefitinib on the proliferation and apoptosis of human NSCLC A549, H1299, H1975, and HCC827 cells was examined by MTT and flow cytometry. The effect of radiation, 5-Fu, and gefitinib on the clonogenicity of H1975 and HCC827 cells was determined by colony formation assay. The effect of radiation, 5-fluorouracil (5-Fu), and gefitinib on the EGFR, AKT, and ERK1/2 activation in H1975 cells was determined by Western blot. The therapeutic efficacy of radiation, S-1, and gefitinib in the growth of implanted H1975 tumors and the AKT activation in the tumors were examined in vivo and immunohistochemistry, respectively. RESULTS: Combination of radiation, 5-Fu, and gefitinib significantly inhibited the proliferation of H1975 cells and triggered their apoptosis, but not other NSCLC cells tested. The combination therapy significantly mitigated the clonogenicity and attenuated the activation of EGFR and AKT signaling in H1975 cells. Furthermore, combination of S-1, gefitinib, and radiation significantly inhibited the growth of implanted H1975 tumors in mice and remarkably reduced the AKT phosphorylation in the tumors. CONCLUSIONS: Our data indicated that combination of S-1 and gefitinib significantly increased radiosensitivity of H1975 cells. The triple combination therapies may benefit patients with the EGFR T790M mutant NSCLC.

[Effect of zu' ai weitai granule on substance P, vasoactive intestinal peptide, calcitonin gene-related peptide of precancerous lesion of gastric antrum in rats].

OBJECTIVE: To explore the relationship between precancerous lesions of gastric antrum and substance P (SP) , vasoactive intestinal peptide ( VIP) , calcitonin gene-related peptide (CGRP), and the therapeutic mechanism of Zu' ai Weitai Granule (ZWG) , a TCM preparation. METHODS: The rat model of precancerous lesions of gastric carcinoma was induced by the combined method of N-methyl N' -nitrosoguani-dine (MNNG) and mechanical injury on gastric mucosa. The pathologic morphological changes of gastric mucosa were observed after prophylactic and therapeutic administration of ZWG. In the meantime,the changes in SP, VIP and CGRP contents were determined by immunohistochemical staining. RESULTS: The contents of SP and CGRP in gastric antrum were obviously improved in the ZWG group when compared with those in the control group (P <0. 05). There was no significant difference in VIP content between the two groups (P >0. 05). CONCLUSION: ZWG could improve SP, VIP, and CGRP contents in rats' gastric antrum either as prophylactic administration or therapeutic administration.