This study aims to investigate the potential of oncolytic nanoparticles encapsulating Coxsackievirus A21 (CVA21) full-genome mRNA (CVA21@ONP) to resurrect CVA21 and induce apoptosis in host cells, as well as the antitumor immune effects of CVA21@ONP in immunocompetent tumor-bearing BALB/c mice. We used lipid nanoparticles (LNPs) to encapsulate CVA21 full-genome mRNA, thus preparing CVA21@ONP. The killing efficacy of CVA21@ONP was determined by the plaque assay and cell counting kit-8 (CCK-8), and the apoptosis in HT29 and CT26-iRFP cells was evaluated by flow cytometry. Mice were administrated with CVA21@ONP at high and low doses intratumorally, and the growth of tumors expressing infra-red fluorescent protein (iRFP) was monitored. Additionally, the types and changes of immune cells in the spleen were analyzed by flow cytometry. The results demonstrated that CVA21@ONP successfully resurrected CVA21 in both HT29 and U87MG cells. The plaque assay revealed robust killing effects of CVA21@ONP against both human and murine cell lines, and flow cytometry results showed increased early and late apoptotic cells. Notably, intratumoral detection revealed significantly down-regulated expression of iRFP in both high- and low-dose CVA21@ONP groups. Flow cytometry results further indicated that CVA21@ONP treatment effectively reduced the levels of immunosuppressive cells, including myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), in the spleen, while enhancing T cell-dependent antitumor immune responses. These findings suggest that CVA21@ONP can replicate and survive extensively both in vitro and in vivo, activating the immune system of mice administrated with CVA21@ONP to target cells at the tumor site, thereby remodeling the tumor immune microenvironment and accelerating the suppression or even complete regression of tumors. The oncolytic performance of CVA21@ONP has been verified through intratumoral injection administration in this study, aimed at further exploring its therapeutic potential and promoting the development of the field of tumor treatment.
Animals ; Nanoparticles/chemistry* ; Mice ; Mice, Inbred BALB C ; Humans ; Apoptosis ; Oncolytic Viruses/genetics* ; Oncolytic Virotherapy/methods* ; Cell Line, Tumor ; RNA, Messenger/genetics* ; HT29 Cells

In this study, we investigated the effects of Panax notoginseng saponins (PNS) on pulmonary vascular remodeling and ADAM10/Notch3 pathway in pulmonary arterial hypertension (PAH). PAH rat model was established, and male Sprague Dawley (SD) rats were randomly divided into control group, monocrotaline (MCT) group and MCT+PNS group, with 10 rats in each group. Rats in the control group were intraperitoneally injected with equal volume of normal saline. Rats in the MCT group was injected intraperitoneally with 60 mg/kg MCT on the first day, and then with the same volume of normal saline every day. Rats in the MCT+PNS group was injected intraperitoneally with 60 mg/kg MCT on the first day, and then with 50 mg/kg PNS every day. The modeling time of each group lasted for 21 days. After the model was established, the mean pulmonary artery pressure (mPAP) was measured by right heart catheterization technique, the right ventricular hypertrophy index (RVHI) was calculated, the microscopic morphology and changes of pulmonary vascular wall were observed by HE and Masson staining, and the expressions of ADAM10, Notch3, Hes-1, P27, PCNA, Caspase-3 proteins and mRNA in pulmonary vascular tissue of rats were detected by Western blot and qPCR. The expression and localization of Notch3 and α-SMA were detected by immunofluorescence staining. The protein expression of ADAM10 was detected by immunohistochemical staining. The results showed that compared with the control group, mPAP, RVHI, pulmonary vessels and collagen fibers in the MCT group were significantly increased, the expressions of ADAM10, Notch3, Hes-1, and PCNA protein and mRNA were significantly increased, while the expressions of P27 and Caspase-3 protein and mRNA were decreased significantly. Compared with the MCT group, mPAP and RVHI were significantly decreased, pulmonary vessels were significantly improved and collagen fibers were significantly reduced, the expressions of protein and mRNA of ADAM10, Notch3, Hes-1, and PCNA were decreased in MCT+PNS group, but the expressions of protein and mRNA of P27 and Caspase-3 were increased slightly. The results of immunofluorescence showed that Notch3 and α-SMA staining could overlap, which proved that Notch3 was expressed in smooth muscle cells. The expression of Notch3 in the MCT group was increased significantly compared with that in the control group, while PNS intervention decreased the expression of Notch3. Immunohistochemical staining showed that compared with the control group, the amount of ADAM10 in the MCT group was increased significantly, and the expression of ADAM10 in the MCT+PNS group was decreased compared with the MCT group. These results indicate that PNS can improve the PAH induced by MCT in rats by inhibiting ADAM10/Notch3 signaling pathway.
Animals ; Male ; Rats ; Caspase 3/metabolism* ; Collagen ; Disease Models, Animal ; Hypertension, Pulmonary/drug therapy* ; Monocrotaline/adverse effects* ; Panax notoginseng/chemistry* ; Proliferating Cell Nuclear Antigen/pharmacology* ; Pulmonary Arterial Hypertension ; Pulmonary Artery/metabolism* ; Rats, Sprague-Dawley ; Receptor, Notch3/genetics* ; RNA, Messenger ; Saline Solution ; Signal Transduction ; Saponins/pharmacology*

The present study established the spectrum-effect relationship model of flavonoids in Citri Reticulatae Pericarpium(CRP) from 15 batches of Liujunzi Decoction and statistically analyzed the correlation between chemical peaks and efficacy to identify the main effective components. HPLC fingerprints of flavonoids in CRP from 15 batches of Liujunzi Decoction were established. HPLC analysis was carried out on the Venusil XBP C_(18)(L) column(4.6 mm×250 mm, 5 μm) at 30 ℃ with acetonitrile-water(containing 0.1% formic acid) as mobile phase for gradient elution, a flow rate of 1.0 mL·min~(-1), and detection wavelength of 300 nm to obtain chemical fingerprints. Additionally, the effects of flavonoids from CRP in 15 batches of Liujunzi Decoction on the content of GAS, MTL, and VIP, TFF3 mRNA expression, and percentage of CD3~+ T-cells of model rats with spleen deficiency were determined. The spectrum-effect relationship model was established by gray correlation analysis. The results showed that the main characteristic peaks with great contribution to the regulation of gastrointestinal tract were peak 16(vicenin-2), peak 63(sinensetin), peak 64(isosinensetin), peak 65(nobiletin), peak 67(3,5,6,7,8,3',4'-heptemthoxyflavone), peak 68(tangeretin), and peak 69(5-desmethylnobiletin). Therefore, there was a linear correlation between flavonoids from CRP in Liujunzi Decoction and the efficacy, and the medicinal effect was achieved by multi-component action. This study is expected to provide a new idea for exploring the material basis of the effect, i.e., regulating qi prior to replenishing qi, of CRP in Liujunzi Decoction.
Animals ; Citrus/chemistry* ; Drugs, Chinese Herbal ; Flavonoids/pharmacology* ; Hormones ; RNA, Messenger/genetics* ; Rats ; Spleen

OBJECTIVE: To determine the effects of hawthorn extract on serum lipid levels, pathological changes in aortic atherosclerosis plaque, inflammatory factors, and apoptosis-related protein and mRNA expression in apolipoprotein E gene knockout (ApoE) mice. METHODS: Thirty-six ApoE mice were fed with a high-fat diet starting at the age of 8 weeks. Mice were randomly divided into 3 groups by a random number table including model group, hawthorn extract group, and simvastatin group, 12 mice in each group. Twelve 8-week-old C57BL/6 mice were fed a basic diet and served as control. The mice in the control and model groups were administered 0.2 mL saline daily, the mice in the hawthorn extract and simvastatin groups were administered with 50 mg/kg hawthorn extract or 5 mg/kg simvastatin daily for 16 weeks. After 16 weeks, plasma lipids including total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were determined by an enzymatic assay. Aortic atherosclerotic lesions were observed by light microscopy, scanning and transmission electron microscopy, respectively. Plasma levels of monocyte chemoattractant protein-1 (MCP-1), interleukin-1β (IL-1β), adiponectin (APN), and hypersensitive C-reactive protein (hs-CRP) were measured by enzyme-linked immunosorbent assay (ELISA). Protein and mRNA expressions of Bax and Bcl-2 in the aorta were assessed by Western blotting and quantitative real-time polymerase chain reaction (qRT-PCR), respectively. RESULTS: Compared to the control group, the plasma levels of TC, TG and LDL-C were significantly increased and HDL-C were significantly decreased in the model group (P<0.01). Compared to the model group, treatment with hawthorn extract significantly decreased the plasma levels of TC, TG, and LDL-C and increased the plasma level of HDL-C in ApoE mice (P<0.01). The levels of MCP-1, IL-1ß, and hs-CRP in the model group were significantly increased and APN was significantly decreased compared with the control group (P<0.01). Compared to the model group, treatment with hawthorn extract decreased the levels of MCP-1, IL-1ß, and hs-CRP and increased the APN level (P<0.01). Compared to the control group, the protein and mRNA expression of Bax in the model group were significantly increased and the expression of Bcl-2 was significantly decreased (P<0.01). Hawthorn extract also reduced the protein and mRNA expression of Bax and increased the Bcl-2 expression in the aorta (P<0.01). CONCLUSION Hawthorn extract has anti-atherosclerosis and stabilizing unstable plaque effects. The mechanism may be related to the inflflammation and apoptosis signaling pathways.
Animals ; Aorta ; pathology ; ultrastructure ; Apoptosis ; drug effects ; Atherosclerosis ; blood ; complications ; drug therapy ; Crataegus ; chemistry ; Inflammation ; blood ; complications ; drug therapy ; Inflammation Mediators ; metabolism ; Lipids ; blood ; Male ; Mice, Inbred C57BL ; Plant Extracts ; pharmacology ; therapeutic use ; RNA, Messenger ; genetics ; metabolism ; bcl-2-Associated X Protein ; metabolism

OBJECTIVETo investigate the effect of total flavone of haw leaves (TFHL) on the expression of nuclear factor erythroid-2 related factor (Nrf2) and other related factors in nonalcoholic steatohepatitis (NASH) rats induced by high-fat diet and then to further discuss the mechanism of TFHL's prevention against NASH.

METHODSHigh-fat diet was fed to 40 rats to establish the NASH model. Then model rats were intragastrically administrated with 40, 80, 160 mg/(kg•day) TFHL, respectively. The pathological changes of liver tissues in NASH rats were detected by oil red O and hematoxylin-eosin (HE) stainings. The expression of Nrf2 in rat liver was examined through immunohistochemistry. The level of 8-iso-prostaglandin F2α in serum was detected through enzyme linked immunosorbent assay (ELISA). The mRNA and protein levels of Nrf2 and other related factors in liver tissue were measured by real-time reverse transcriptionpolymerase chain reaction and western blot.

RESULTSLipid deposition, hepatic steatosis, focal necrosis in lobular inflammation and ballooning degeneration were emerged in livers of NASH rats. The 8-iso-prostaglandin F2α in the serum of NASH rats increased significantly compared with the control group (P<0.05). The mRNA of Nrf2, hemeoxyenase1 (HO-1) and the mRNA and protein levels of quinine oxidoreductase (NQO1) in NASH rats liver tissue showed a striking increase, while the mRNA levels of Keap1, r-glutamylcysteine synthethase (rGCS) and glutathione S-transferase (GST) were significantly decreased compared with the control group (P<0.05). After TFHL treatment, 8-iso-prostaglandin F2α level in serum significantly decreased, and Nrf2 mRNA and protein levels in hepatocytes nucleus enhanced compared with the model group (P<0.05 or 0.01). Meanwhile the Keap1 mRNA, the mRNA and protein levels of HO-1, NQO1 antibody, rGCS antibody, GST increased after TFHL treatment (P<0.05 or 0.01).

CONCLUSIONSNrf2 and other related factors were involved in development of NASH, and they also served as an important part in its occurrence. By regulating expression of Nrf2 and other related factors, TFHL may play a role in antioxidative stress and prevention of NASH.

Animals ; Cell Nucleus ; drug effects ; metabolism ; Crataegus ; chemistry ; Dinoprost ; metabolism ; Flavones ; pharmacology ; therapeutic use ; Lipids ; chemistry ; Liver ; drug effects ; metabolism ; pathology ; NF-E2-Related Factor 2 ; genetics ; metabolism ; Non-alcoholic Fatty Liver Disease ; drug therapy ; genetics ; pathology ; Phytotherapy ; Plant Leaves ; chemistry ; RNA, Messenger ; genetics ; metabolism ; Rats, Sprague-Dawley

OBJECTIVETo examine the effect of icariin (ICA) on the cognitive impairment induced by traumatic brain injury (TBI) in mice and the underlying mechanisms related to changes in hippocampal acetylation level.

METHODSThe modifified free-fall method was used to establish the TBI mouse model. Mice with post-TBI cognitive impairment were randomly divided into 3 groups using the randomised block method (n=7): TBI (vehicle-treated), low-dose (75 mg/kg) and high-dose (150 mg/kg) of ICA groups. An additional sham-operated group (vehicle-treated) was employed. The vehicle or ICA was administrated by gavage for 28 consecutive days. The Morris water maze (MWM) test was conducted. Acetylcholine (ACh) content, mRNA and protein levels of choline acetyltransferase (ChAT), and protein levels of acetylated H3 (Ac-H3) and Ac-H4 were detected in the hippocampus.

RESULTSCompared with the sham-operated group, the MWM performance, hippocampal ACh content, mRNA and protein levels of ChAT, and protein levels of Ac-H3 and Ac-H4 were signifificantly decreased in the TBI group (P<0.05). High-dose of ICA signifificantly ameliorated the TBI-induced weak MWM performance, increased hippocampal ACh content, and mRNA and protein levels of ChAT, as well as Ac-H3 protein level compared with the TBI group (P<0.05).

CONCLUSIONICA improved post-TBI cognitive impairment in mice by enhancing hippocampal acetylation, which improved hippocampal cholinergic function and ultimately improved cognition.

Acetylation ; Acetylcholine ; metabolism ; Animals ; Brain Injuries, Traumatic ; complications ; Choline O-Acetyltransferase ; genetics ; metabolism ; Cognitive Dysfunction ; drug therapy ; etiology ; Flavonoids ; chemistry ; pharmacology ; therapeutic use ; Hippocampus ; pathology ; Histones ; metabolism ; Homeostasis ; drug effects ; Male ; Maze Learning ; drug effects ; Mice ; RNA, Messenger ; genetics ; metabolism

OBJECTIVESTo investigate the hair growth-promoting effect of Miscanthus sinensis var. purpurascens (MSP) flower extracton on in vitro and in vivo models.

METHODSMSP flower extract was extracted in 99.9% methanol and applied to examine the proliferation of human dermal papilla cells (hDPCs) in vitro at the dose of 3.92-62.50 μg/mL and hair growth of C57BL/6 mice in vivo at the dose of 1000 μg/mL. The expression of transforming growth factor β1 (TGF-β1), hepatocyte growth factor (HGF), β-catenin, substance P was measured by relative quantitative realtime polymerase chain reaction. Histopathological and immunohistochemical analysis were performed.

RESULTSMSP (7.81 μg/mL) down-regulated TGF-β1 and up-regulated HGF and β-catenin in hDPCs (P<0.01). MSP (1000 μg/mL)-treated mice showed the earlier transition of hair follicles from the telogen to the anagen phase. The number of mast cells was lower in the MSP-treated mice than in other groups (P<0.05 vs. NCS group). Substance P and TGF-β1 were expressed in hair follicles and skin of the MSP group lower than that in negative control. Stem cell factor in hair follicles was up-regulated in the MSP-treated mice (P<0.01).

CONCLUSIONSThe MSP flower extract may have hair growth-promotion activities.

Animals ; Antioxidants ; pharmacology ; Cell Count ; Cell Proliferation ; drug effects ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Female ; Flowers ; chemistry ; Hair Follicle ; cytology ; drug effects ; growth & development ; Hepatocyte Growth Factor ; metabolism ; Humans ; Mast Cells ; cytology ; Mice, Inbred C57BL ; Phosphorylation ; drug effects ; Plant Extracts ; pharmacology ; Poaceae ; chemistry ; RNA, Messenger ; genetics ; metabolism ; Skin ; metabolism ; Stem Cell Factor ; metabolism ; Stress, Psychological ; pathology ; Substance P ; metabolism ; Transforming Growth Factor beta ; genetics ; metabolism ; Vascular Endothelial Growth Factor A ; genetics ; metabolism ; beta Catenin ; metabolism

OBJECTIVETo investigate the changes in the expression of EphA5 and its ligand ephrinA5 in the hippocampus of rats with epilepsy and their role in the pathogenesis of temporal lobe epilepsy (TLE).

METHODSA total of 240 Sprague-Dawley rats were randomly divided into control group and TLE group, with 120 rats in each group. A rat model of lithium-pilocarpine TLE was established, and then the rats were divided into subgroups at 12 and 24 hours and 7, 15, 30, and 60 days after epilepsy was induced. In-situ hybridization was used to measure the mRNA expression of ephrinA5 in the CA3 region and the dentate gyrus of the hippocampus in 9 rats; immunohistochemistry was used to measure the protein expression of EphA5 in the CA3 region and the dentate gyrus of the hippocampus in 9 rats; Neo-Timm silver staining was used to observe mossy fiber sprouting in the CA3 region of the hippocampus in 2 rats.

RESULTSIn-situ hybridization showed mRNA expression of ephrinA5 in the CA3 region of the hippocampus, but this was not found in the dentate gyrus. Compared with the control group at the same time point, the TLE group had a significant reduction in the mRNA expression of ephrinA5 in the CA3 region of the hippocampus at 7 and 15 days after epilepsy was induced (P<0.05); at 30 and 60 days after epilepsy was induced, the TLE group had a gradual increase in the mRNA expression of ephrinA5 in the CA3 region of the hippocampus, and there was no significant difference between the TLE and control groups (P>0.05). Immunohistochemistry showed that EphA5 protein was expressed in the CA3 region and the dentate gyrus of the hippocampus and had a similar trend of change as ephrinA5 mRNA. Neo-Timm silver staining showed that the TLE group developed marked mossy fiber sprouting in the CA3 region of the hippocampus at 7 and 15 days after epilepsy was induced.

CONCLUSIONSDownregulation of ephrinA5 and EphA5 in the CA3 region of the hippocampus may participate in the mechanism of mossy fiber sprouting and is closely associated with the development and progression of epilepsy.

Animals ; Ephrin-A5 ; analysis ; genetics ; physiology ; Epilepsy, Temporal Lobe ; etiology ; metabolism ; Hippocampus ; chemistry ; Male ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Receptor, EphA5 ; analysis ; genetics ; physiology

To investigate the effect of a novel EZH2 inhibitor GSK126 on cell growth, apoptosis and migration of prostate cancer cells.Prostate cancer PC-3 and DU145 cells were treated with GSK126 at different doses. Cell growth was detected by sulforhodamine assay. Cell apoptosis was assayed by Annexin V-/PI kit. Transwell chamber and wound healing assays were conducted to detect cell migration. The mRNA level was detected by quantitative PCR, and protein expression was detected by Western blot analysis.GSK126 showed significant effect on cell growth and apoptosis when the dose was higher than 50 μmol/L. Wound healing assay revealed that scratch space in PC-3 cells was significantly increased in a dose-dependent manner in GSK126-treated groups[(247.2±24.4),(347.2±19.2) and (410.5±18.1) μm in low, medium and high dose (5.0, 20.0, 50.0 μmol/L), respectively] as compared with the control group[(171.3±17.8) μm](all<0.05). Transwell assay showed that migrated PC-3 cells in control group was 322.0±17.9,while those in GSK126-treated groups were 198.3±15.4 (low),82.7±6.2 (medium) and 30.2±4.1 (high), and the differences between the control group and GSK126-treated groups were significant(all<0.05). In addition, GSK126 up-regulated E-cadherin mRNA expression and down-regulated N-cadherin and Vimentin mRNA expression, whereas had no significant effect on Snail, Fibronectin and VEGF-A mRNA expression. The protein expression of E-cadherin was elevated but VEGF-A protein did not change in GSK126-treated groups. Similar results were exhibited in DU145 cell.GSK126 can significantly inhibit cell migration and invasion in prostate cancer PC-3 and DU145 cells, which may be resulted from its effect on epithelial-mesenchymal transition. GSK126 may be used as a potential anti-prostate cancer dug in clinic.
Apoptosis ; drug effects ; Cadherins ; analysis ; drug effects ; metabolism ; Cell Line, Tumor ; drug effects ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Down-Regulation ; drug effects ; Drug Screening Assays, Antitumor ; methods ; Enhancer of Zeste Homolog 2 Protein ; analysis ; drug effects ; metabolism ; Fibronectins ; analysis ; drug effects ; metabolism ; Humans ; Indoles ; pharmacology ; Male ; Prostatic Neoplasms ; chemistry ; genetics ; physiopathology ; Pyridones ; pharmacology ; RNA, Messenger ; Up-Regulation ; drug effects ; Vascular Endothelial Growth Factor A ; analysis ; drug effects ; Vimentin ; analysis ; drug effects ; metabolism

To investigate the effect of silencing DJ-1 on xenografted human laryngeal squamous cell carcinoma (LSCC) Hep-2 cells in nude mice.Xenograft model of human LSCC was established by subcutaneous transplantation of Hep-2 cells in 24 nude mice. The LSCC-bearing nude mice were randomly divided into 3 groups (=8 in each):DJ-1 siRNA low dose group and DJ-1 siRNA high dose group were injected in tumors with 20 μg of DJ-1 siRNA or 40 μg of DJ-1 siRNA in 50 μL, respectively; control group was injected with 5% glucose solution in 50 μL, twice a week for 3 weeks. The weight and size of tumors were measured before injection. The animals were sacrificed 48 h after the final treatment, and the tumors were harvested and weighed. The apoptosis and proliferation of tumor cells were determined; the expressions of Caspase-3 and Ki-67 in tumor specimens were detected with immunohistochemistry. The expression of DJ-1, PTEN, survivin mRNA and protein in tumor tissues were detected by RT-PCR and Western blotting, respectively.Tumor weight in low dose group[(0.66±0.15)g] and high dose group[(0.48±0.11)g] were significantly lower than that in control group[(0.83±0.16)g, all<0.05]. The inhibition rates of low dose group and high dose group were (20.48±0.18)% and (42.16±0.13)%, respectively. Immunohistochemistry showed that the expression of Caspase-3 was increased and Ki-67 was reduced in tumor specimens, compared with the control group (all<0.05). RT-PCR and Western blot results showed that in low dose group and high dose group the mRNA and protein expression of DJ-1 and survivin significantly decreased (all<0.05), while PTEN mRNA and protein content increased (all<0.05).High dose DJ-1 siRNA can inhibit the tumor growth in human LSCC xenograft nude mouse model, which indicates that down-regulating DJ-1 and survivin, and up-regulating PTEN expression may lead to blockage of PI3K-PKB/Akt signaling pathway and promoting tumor cell apoptosis.
Animals ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; genetics ; Carcinoma, Squamous Cell ; chemistry ; genetics ; physiopathology ; Caspase 3 ; analysis ; drug effects ; Cell Line, Tumor ; chemistry ; drug effects ; physiology ; transplantation ; Cell Proliferation ; drug effects ; Down-Regulation ; Gene Expression Regulation ; drug effects ; genetics ; physiology ; Head and Neck Neoplasms ; chemistry ; genetics ; physiopathology ; Heterografts ; drug effects ; physiology ; Humans ; Inhibitor of Apoptosis Proteins ; analysis ; drug effects ; Ki-67 Antigen ; analysis ; drug effects ; Laryngeal Neoplasms ; chemistry ; genetics ; physiopathology ; Mice, Nude ; PTEN Phosphohydrolase ; analysis ; drug effects ; Phosphatidylinositol 3-Kinases ; drug effects ; Protein Deglycase DJ-1 ; pharmacology ; Proto-Oncogene Proteins c-akt ; drug effects ; RNA Interference ; physiology ; RNA, Messenger ; pharmacology ; RNA, Small Interfering ; physiology ; Signal Transduction ; drug effects ; genetics ; physiology