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Acute kidney injury (AKI) is a common critical disease clinically with high morbility and mortality and some survival patients also progress to chronic kidney disease. Renal ischemia-reperfusion (IR) is one of the main causes of AKI, in which, its repair and potential fibrosis, apoptosis, inflammation and phagocytosis play important roles. During the progression of IR-induced AKI, the expression of erythropoietin homodimer receptor (EPOR)2 and EPOR and β common receptor formed heterodimer receptor (EPOR/βcR) is changed dynamically. Moreover, (EPOR)2 and EPOR/βcR may synergistically participate in renoprotection at the stage of AKI and early repair, whereas at the late stage of AKI, the (EPOR)2 induces renal fibrosis and the EPOR/βcR facilitates repair and remodelling. The underlying mechanism, signaling pathways and the different effect turning point of (EPOR)2 and EPOR/βcR have not been well defined. It has been reported that EPO, according to its 3D structure, derived helix B surface peptide (HBSP) and cyclic HBSP (CHBP) only bind to EPOR/βcR. Synthesized HBSP, therefore, provides an effective tool to distinguish the different roles and mechanisms of both receptors, with the (EPOR)2 promoting fibrosis or the EPOR/βcR leading to repair/remodelling at the late stage of AKI. This review discusses the similarities and differences of (EPOR)2 and EPOR/βcR in their impacts on apoptosis, inflammation and phagocytosis in AKI, repair and fibrosis post IR, associated mechanisms, signaling pathways and outcomes.
Subject(s)
Humans , Receptors, Erythropoietin , Acute Kidney Injury , Apoptosis , Inflammation , Phagocytosis , Reperfusion InjuryABSTRACT
AIM:To investigate the effect of bromodomain-containing protein 4(BRD4)inhibitors on the via-bility and apoptosis of activated B cell-like diffuse large B-cell lymphoma(ABC-DLBCL)cells and the molecular mecha-nism. METHODS:The ABC-DLBCL cells were treated with BRD4 inhibitors JQ1 and I-BET-762,and Bruton tyrosine kinase(BTK)inhibitor ibrutinib. The viability and death of the cells were determined by CCK-8 assay and PI staining,re-spectively. The mRNA levels of BTK,phospholipase Cγ(PLCγ),LYN,SYK,interleukin-6(IL-6),MYC,protein ki-nase Cβ(PKCβ),mucosa-associated lymphoid tissue lymphoma translocation protein 1(MALT1),MYC and RELA were detected by real-time PCR. The protein levels of BTK,PLCγ,MYC and RELA were determined by Western blot. Super-enhancer around BTK gene was revealed by bioinformatics analysis. RESULTS:The ABC-DLBCL cells were sensitive to BRD4/super-enhancer inhibitors such as JQ1 and I-BET-762. Both JQ1 and I-BET-762 inhibited the chronic active B-cell receptor(BCR)/nuclear factorκB(NFκB)signaling through reducing the transcription of BTK,but they had minimal ef-fect on other components in BCR/NFκB signaling. Interestingly,there was no super-enhancer around BTK gene,and the inhibitory effect of JQ1 was likely due to disruption of BRD4 binding within BTK gene. Inhibition of BRD4 had synergic ef-fect with BTK inhibitor ibrutinib. Moreover,inhibition of BRD4 induced significant cell death in ibrutinib-resistant ABC-DLBCL cells. CONCLUSION:Inhibitors of BRD4 induce ABC-DLBCL cell death via blocking BCR/NFκB signaling and has synergic effect with BTK inhibitor. Inhibition of BRD4 might be a promising strategy for treatment of ABC-DLBCL,es-pecially ibrutinib-resistant ABC-DLBCL.
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Immune checkpoints (ICs) are immunosuppressive molecules expressed on immune cells, which can regulate immune cells' activation. Immune checkpoint inhibitors (ICIs) which can block the interaction of immune checkpoints and their ligands, improve the cytotoxic effect of the immune system on tumor cells. Immunotherapy such as employing ICIs has gradually become a conventional therapeutic strategy for cancer treatment. However, the low response rate and the emergence of drug resistance have seriously affected the clinical efficacy of ICIs. Reactive oxygen species (ROS) are electronic reduction products of active oxygen, as well as natural by-products of cell metabolism, which can be used as regulators of intercellular signals. Tumor microenvironment (TME) is often in the state of oxidative stress (OS), which is the imbalance between oxidative system and antioxidant system. ROS can affect the interaction with its ligands by regulating the expression and activity of immune checkpoints in TME, thus affecting the anti-tumor effect of immune cells. Accumulating studies have shown that ROS could regulate tumor immune checkpoints through several pathways. Due to different types and stages of tumor, it would be clinical beneficial to understand the mechanistic link of ROS on tumor immune checkpoint, and choose appropriate ROS regulators combined with immune checkpoint inhibitors to maximize anti-tumor effects. This article reviews the common metabolic sources and characteristics of ROS, the regulatory effect and mechanism of ROS on tumor immune checkpoints and its therapeutic application.
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To explore the effect of tanshinone IIA (TanIIA) on the occurrence and development of breast cancer, we employed the mouse mammary tumor virus-polyomavirus middle T antigen (MMTV-PyMT) transgenic mice as a spontaneous breast cancer mouse model. Animal welfare and experimental procedures were in accordance with the regulations of the Animal Ethics Committee of Nanjing University of Chinese Medicine. The animals were divided into control group, low-dose TanIIA treatment group (30 mg·kg-1·day-1), and high-dose TanIIA treatment group (60 mg·kg-1·day-1). The treatment was administered orally and daily for 5 weeks. The mice were sacrificed after final treatment. Mammary gland and lung were collected for histopathology studies. We evaluated the chemoprophylaxis effect of TanIIA on breast cancer in mice according to the pathological characteristics of breast cancer at different stages of development. Immunofluorescence staining were employed for blood vessel analysis. The expression levels of E-cadherin, proliferating nuclear antigen (PCNA), and oncogene c-Myc were detected by immunohistochemistry. Flow cytometry was used to analyze cell cycle and Cytoscape was used to construct drug-disease protein-protein interaction (PPI) network. Our results showed that TanIIA inhibits breast tumor progression by delaying malignancy from adenoma to early carcinoma, and inhibits blood vessel formation during tumor development. TanIIA (60 mg·kg-1·day-1) inhibits the expression levels of PCNA and c-Myc, upregulates the expression of E-cadherin. In addition, cell cycle experiments showed that the cell cycle of PyMT primary mammary cells in the high-dose TanIIA group was arrested in the G0/G1 phase. Our study demonstrated that TanIIA can significantly inhibit breast tumor progression in MMTV-PyMT mouse model, which may be related to the inhibition of angiogenic switch and cell cycle arrest.
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The elements of ethical review related to clinical research of acupuncture and moxibustion is discussed to provide ideas for various institutions to carry out relevant ethical review. It is believed that the ethical review of clinical research of acupuncture and moxibustion needs to focus on the specificity of acupuncture and moxibustion. Starting from the basic theory of traditional Chinese medicine, the theory of meridians and acupoints and the theory of syndrome differentiation along meridians, the key contents of ethical review such as intervention methods, grouping design and placebo control should be considered, so as to standardize the clinical research of acupuncture and moxibustion and protect the health and rights and interests of participants.
Subject(s)
Humans , Acupuncture Points , Acupuncture Therapy , Biomedical Research , Ethical Review , Meridians , MoxibustionABSTRACT
Objective:To explore the effect of Chaihu Jia Longgu Muli Tang on the hippocampus of rats with chronic stress depression based on the brain-derived neurotrophic factor (BDNF)/tyrosine kinase B (TrkB)/cyclic adenosine phosphate response element-binding protein (CREB) pathway. Method:Sixty SD rats were divided into a blank group (<italic>n</italic>=10) and an experimental group (<italic>n</italic>=50) for the induction of the chronic stress depression model. The rats in the experimental group were further divided into the following five groups: a model group， a fluoxetine hydrochloride group (0.003 g·kg<sup>-1</sup>), and low-(1.625 g·kg<sup>-1</sup>), medium-(3.25 g·kg<sup>-1</sup>), and high-dose (6.5 g·kg<sup>-1</sup>) Chaihu Jia Longgu Muli Tang groups. The rats were administered correspondingly by gavage once a day for eight weeks. Behavioral tests were performed to evaluate the depression state of the rats before modeling， after modeling， and after drug administration. Hematoxylin-eosin (HE) staining was used to observe the morphological changes in the hippocampus of rats. The immunohistochemical (IHC) staining was used to quantitatively detect BDNF protein expression in the rat hippocampus. The mRNA and protein expression of BDNF， TrkB， and CREB in the rat hippocampus was detected by the real-time fluorescence-based quantitative PCR (Real-time PCR) and Western blot， respectively. Result:Compared with the blank group， the model group showed decreased sucrose preference rate (<italic>P</italic><0.05)， declining horizontal and vertical scores (<italic>P</italic><0.05)， and prolonged immobility time and floating time (<italic>P</italic><0.05). Additionally， HE staining results revealed that hippocampal neuron structure was damaged. IHC staining showed that the mRNA and protein expression of BDNF， TrkB， and CREB was significantly decreased (<italic>P</italic><0.05). Compared with the model group， the fluoxetine hydrochloride group and the Chaihu Jia Longgu Muli Tang groups displayed elevated sucrose preference rate (<italic>P</italic><0.05)， increased horizontal and vertical scores (<italic>P</italic><0.05)， and shortened immobility time and floating time (<italic>P</italic><0.05). Furthermore, the hippocampal neuron structure was significantly repaired. IHC staining showed that the mRNA and protein expression of BDNF， TrkB， and CREB was significantly increased (<italic>P</italic><0.05). Conclusion:Chaihu Jia Longgu Muli Tang can significantly improve the depression-like behaviors of rats after chronic stress stimulation and enhance the regeneration and repair of neurons in the hippocampus. The underlying mechanism may be related to the up-regulation of the BDNF/TrkB/CREB signaling pathway in the hippocampus of rats.
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The tumor contains abundant new vessels, which are unevenly distributed, irregular, and branch-disordered. Angiopoietin (Ang) and tyrosine kinase receptor Tie mediate stable maturation of angiogenesis. Ang1 mainly plays a role in promoting vascular stabilization, and Ang2 is highly expressed in vessels, which makes the structure and function of vessels abnormal. Leaked vessels provide opportunities for invasion and metastasis of circulating tumor cells. Targeting the Ang/Tie axis to correct the abnormal state of vessels and promote its normalization, combined with chemotherapy drugs or immunotherapy, play a synergistic effect against tumors. This article summarizes the role of Ang/Tie axis in tumor angiogenesis and metastasis, and it aims to provide new ideas and strategies for clinical treatment of tumors.
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@#AIM: To investigate the correlation between retinal microstructural changes and visual function after scleral buckling and vitrectomy with rhegmatogenous retinal detachment.<p>METHODS: Prospective clinical study was conducted. With PVR grade B-C1, 75 patients with 75 eyes of aged 15-30 years who were macular-off retinal detachment were selected. PPV was performed for posterior equatorial retinal hiatus(35 eyes, PPV group), and SB was performed for anterior equatorial retinal hiatus(40 eyes, SB group).The patients in the two groups were followed up for 6mo to observe the changes in the best corrected visual acuity(BCVA)and retinal microstructures.<p>RESULTS: Compared with SB group, the probability of inner sensory layer/outer sensory layer and inner boundary membrane fracture after operation in PPV group was 2.812 times, with a statistically significant difference(<i>P</i>=0.020), and the probability of subretinal effusion was 0.115 times, with a statistically significant difference(<i>P</i><0.001). There was no statistically significant difference(<i>P</i>=0.700)between the two groups. And with the extension of observation time, the possibility of inner sensory layer/outer sensory layer and inner boundary membrane, persistent subretinal fluid and interlayer effusion in the two groups of patients gradually decreased. After the operation, the thickness of the neurosensory retinal and outer nuclear layer in the two groups gradually increased, and BCVA gradually improved. With the prolonging of observation time, the thickness of the neurosensory retinal in SB group increased greatly, and BCVA recovered better.<p>CONCLUSION: The integrity of the inner sensory layer/outer sensory layer and inner boundary membrane, subretinal effusion, interlaminar effusion, and the thickness of the neurosensory retinal are the key factors affecting the recovery of visual function after SB or PPV.
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<p><b>OBJECTIVE</b>To study the clinical and laboratory characteristics of juvenile myelomonocytic leukemia (JMML).</p><p><b>METHODS</b>The clinical characteristics and laboratory results were retrospectively analyzed in 10 children with newly diagnosed JMML. They were compared with those of 28 children with myelodysplastic syndrome (MDS) and 44 children with chronic myeloid leukemia (CML).</p><p><b>RESULTS</b>Compared with the children with CML or MDS, the children with JMML had significantly higher rates of skin rashes, ecchymosis, and lymphadenectasis, a significantly lower serum cholinesterase (ChE) level, and a significantly higher fetal hemoglobin level (P<0.05). The white blood cell count of children with JMML was significantly higher than that of children with MDS, but significantly lower than that of children with CML (P<0.05). In addition, the myeloid/erythroid ratio and rate of dyshaematopoiesis were significantly lower in children with JMML than those in children with CML or MDS. The children with JMML had a significantly higher expression of mature monocyte marker CD14 than those with CML or MDS (P<0.05). The levels of myeloid markers CD33, CD11b, CD13, and CD15 in children with JMML were significantly higher than those in children with MDS, but significantly lower than those in children with CML (P<0.05). The levels of CD2 and CD7 in children with JMML were higher than those in children with CML, but lower than those in children with MDS (P<0.05).</p><p><b>CONCLUSIONS</b>Skin rashes, ecchymosis, lymphadenectasis, and ChE reduction are more common in children with JMML than in those with CML or MDS, while dyshaematopoiesis is less common. In addition, CD14 level increases significantly in children with JMML.</p>
Subject(s)
Child , Child, Preschool , Female , Humans , Infant , Male , Leukemia, Myelomonocytic, Juvenile , Genetics , Allergy and Immunology , Lipopolysaccharide ReceptorsABSTRACT
Angiogenesis inhibitors can make tumor cells in a harsh environment by inhibiting tumor angiogenesis and effectively blocking the tumor progression.However,anti-angiogenic drugs have shown lots of limitations,such as short-term duration,numerous adverse reactions,benefiting only a minority of tumor types and so on.These limitations restrain the development of new drugs and limit the cancer therapies.Many studies have revealed that tumor cells can escape from anti-angiogenic treatments through a variety of ways and mechanisms.In this review,we focus on the reasons behind the failure in treatments,so as to propose solving strategies to improve the current anti-angiogenic drug efficacy and provide reference for new angiogenesis inhibitors and clinical medication.
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Objective To investigate the intervening effect of low-frequency electroacupuncture on rat neuralgia and its regulating effect on substance P (SP) in the spinal dorsal horn. Method SD rats were randomized to normal, sham operation (sham SNI), operation (SNI) and electroacupuncture (SNI+EA) groups, 8 rats each. A rat model of neuralgia was made by spared sciatic nerve injury (SNI). Points Zusanli(ST36) and Kunlun(BL60) on the operation side were given 2 Hz electroacupuncture once daily for 14 days. The rat hind paw withdrawal threshold (PWT) was measured on the operation side to observe its pain hypersensitivity. SP-positive expression in the spinal dorsal horn on the operation side was determined by immunofluorescence. Result Operative side PWT decreased significantly in the SNI group of rats (P<0.01). Electroacupuncture increased operative side PWT in the SNI neuralgia rats (P<0.01). Pain threshold onthe healthy side had no marked change in the SNI group of rats (P>0.05). SP-positive expression in the spinal dorsal horn increased on the operation side (P<0.01) and also on the healthy side (P<0.05) in the SNI group of rats. Electroacupuncture decreased SP-positive expression in the spinal dorsal horn on the operation side in the SNI rats (P<0.01). Electroacupuncture did not significantly change SP-positive expression in the spinal dorsal horn on the healthy side (P>0.05). PWT on the operation and healthy sides and SP-positive cell expression in the spinal dorsal horn on the operation and healthy sides had no marked changes in the SNI group of rats (P>0.05). Conclusion Low-frequency electroacupuncture can relieve rat neuralgia. Its mechanism may be related to it inhibiting SP-positive expression in the spinal dorsal horn on the operation side.
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This paper describes a study exploring the interaction between gomizine D and α-glucosidase. The inhibitory activity of α-glucosidase by gomizine D was determined using PNPG as substrates Gomizine D gave the IC₅₀ value of 0.59 mmol•L⁻¹, which was higher than that of acarbose (1.95 mmol•L⁻¹). Gomizine D was a reversible and non-competitiveα-glucosidase inhibitor with an inhibition constant Ki=4.026 g•L⁻¹. The binding mode between gomizine D and α-glucosidase was analyzed by AutoDock Vina molecular docking software. The lowest energy of Gomizine D binding to α-glucosidase was -7.7 kcal•mol⁻¹, which was lower than that of acarbose (-6.6 kcal•mol⁻¹). After binding with gomizine D, UV spectroscopy analysis displayed that the microenvironment of aromatic residue in the secondary structure of α-glucosidase was changed, and the polarity of protein was reduced.
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p53 gene plays an important role in apoptosis, which is necessary for successful invasion of trophoblast cells. The change from an arginine (Arg) to a proline (Pro) at codon 72 can influence the biological activity of p53, which predisposes to an increased risk of recurrent spontaneous abortion (RSA). In order to investigate the association between p53 polymorphism at codon 72 and RSA, we conducted this meta-analysis. Pubmed, Embase and Web of science were used to identify the eligible studies. Odds ratio (OR) with 95% confidence interval (CI) was used to evaluate the strength of the association. Six studies containing 937 cases of RSA and 830 controls were included, and there was one study deviated from Hardy-Weinberg equilibrium (HWE). There was a significant association between p53 polymorphism at codon 72 and RSA in recessive model (Pro/Pro vs. Pro/Arg+Arg/Arg; OR=1.60, 95% CI: 1.14-2.24) and co-dominant model (Pro/Pro vs. Arg/Arg; OR=1.47, 95% CI: 1.02-2.12) whether the study that was deviated from HWE was eliminated or not. A significant association was observed in allelic model (Pro vs. Arg; OR=1.28, 95% CI: 1.04-1.57) after exclusion of the study that was deviated from HWE. No association was noted in recessive model (Pro/Pro+Pro/Arg vs. Arg/Arg; OR=1.05, 95% CI: 0.86-1.30) and co-dominant model (Pro/Arg vs. Arg/Arg; OR=0.96, 95% CI: 0.77-1.19). Subgroup analysis by ethnicity also indicated a significant association between p53 polymorphism at codon 72 and RSA in Caucasian group. No heterogeneity and publication bias were found. Our meta-analysis implied that p53 polymorphism at codon 72 carries high maternal risk of RSA.
Subject(s)
Adult , Female , Humans , Pregnancy , Abortion, Spontaneous , Diagnosis , Ethnology , Genetics , Alleles , Asian People , Case-Control Studies , Codon , White People , Gene Frequency , Genetic Predisposition to Disease , Odds Ratio , Polymorphism, Single Nucleotide , Recurrence , Risk Factors , Tumor Suppressor Protein p53 , Genetics , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To verify the clinical efficacy of acupuncture on motor dysfunction in ischemic stroke of subacute stage.</p><p><b>METHODS</b>The multi-central randomized controlled trial was adopted. One hundred and twenty-six cases of ischemic stroke of subacute stage were randomized into an acupuncture group (61 cases) and a conventional treatment group (65 cases). The basic treatment of western internal medicine and rehabilitation training were applied to the patients of the two groups. In the acupuncture group, acupuncture was supplemented at the body points located on the extensor of the upper limbs and the flexor of the lower limbs. In combination, scalp acupuncture was applied to NS5, MS6 and MS6 on the affected side. The treatment was given 5 times a week and totally 8 weeks were required. The follow-up observation lasted for 3 months. The scores in Fugl-Meyer scale and NIHSS scale and Barthel index were compared between the two groups before treatment, in 4 and 8 weeks of treatment and the 3-month follow-up observation after treatment separately.</p><p><b>RESULTS</b>In 4 and 8 weeks of treatment and the follow-up observation, Fugl-Meyer scale score was improved obviously in the patients of the two groups (all P<0. 01). In 8 weeks of treatment and the follow-up observation, Fugl-Meyer scale score in the acupuncture groupwas im proved much apparently as compared with that in the conventional treatment group [68. 0 (43. 0,86. 5) vs 52. 5 (30.3, 77.0), 77.0 (49.5, 89.0) vs 63. 0 (33.0, 84.0), both P<0. 05]. Except that NIHSS scale score was not reduced apparently in 4 weeks of treatment in the conventional treatment group (P>0.05), the results of NIHSS scale at the other time points were all decreased obviously as compared with those before treatment in the patients of the two groups (all P<0. 01). In 8 weeks of treatment and the follow-up observation, the results in the acupuncture group were reduced much apparently as compared with those in the conventional treatment group [5. 0 (3.0,8.0) vs 7. 0 (3.0,13.8), 4. 0 (1.5,7.0) vs 6.0 (2.0,11.7) ,both P<0. 05]. In 8 weeks of treatment and the follow-up observation, Barthel index was improved obviously as compared with that before treatment in the patients of the two groups (all P<0. 05). The improvement in the acupuncture group was much more significant as compared with the conventional treatment group [75. 0 (60. 0,87. 5) vs 65. O (36. 3, 87. 5), P<0. 051.</p><p><b>CONCLUSION</b>Based on the conventional treatment, Acupuncture achieves the satisfactory clinical efficacy on motor dysfunction in ischemic stroke of subacute stage.</p>
Subject(s)
Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Acupuncture Therapy , Motor Activity , Stroke , Therapeutics , Treatment OutcomeABSTRACT
Recent clinical trials have shown that electrical stimulation has beneficial effects in obstructive sleep apnea syndrome (OSAS). The purpose of this study was to evaluate the efficacy of electrical stimulation therapy for OSAS with a meta-analysis. The meta-analysis of all relative studies was performed through searching international literature, including PUBMED, CNKI, and EMBASE databases. This literature analysis compared all patients undergoing electrical stimulation therapy with respect to the respiratory disturbance index (RDI) and changes in sleep structure. Six studies were selected involving a total of 91 patients. The meta-analysis indicated that electrical stimulation therapy reduced RDI, longest apnea time, and improved the minimum SaO2. Based on the evidence found, electrical stimulation may be a potential therapy for OSAS, warranting further clinical trials.
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Recent clinical trials have shown that electrical stimulation has beneficial effects in obstructive sleep apnea syndrome (OSAS). The purpose of this study was to evaluate the efficacy of electrical stimulation therapy for OSAS with a meta-analysis. The meta-analysis of all relative studies was performed through searching international literature, including PUBMED, CNKI, and EMBASE databases. This literature analysis compared all patients undergoing electrical stimulation therapy with respect to the respiratory disturbance index (RDI) and changes in sleep structure. Six studies were selected involving a total of 91 patients. The meta-analysis indicated that electrical stimulation therapy reduced RDI, longest apnea time, and improved the minimum SaO2. Based on the evidence found, electrical stimulation may be a potential therapy for OSAS, warranting further clinical trials.
Subject(s)
Humans , Electric Stimulation Therapy , Methods , Respiratory Mechanics , Physiology , Sleep , Physiology , Sleep Apnea, Obstructive , TherapeuticsABSTRACT
<p><b>OBJECTIVE</b>To optimize extraction and purification methods of acidic polysaccharide from Moerella iridescens (MIAP).</p><p><b>METHODS</b>With alkali extraction process and orthogonal experiment,the time consumption,temperature,pH value of the solution and alcohol concentration during the extraction were optimized. The crude products were deprived of protein,pigment and ion,then were purified with DEAE-cellulose ion-exchange chromatography and verified with Sephadex G-100 and cellulose acetate membrane electrophoresis,and examined with infrared spectrum.</p><p><b>RESULTS</b>The optimized extraction conditions were as follows: extraction time 6 h,extraction temperature 70 degree,the solution pH 8.0 and the concentration of alcohol precipitation 70%. Intuitive features showed that the MIAP was pure white crystalline granular with slight dark brown color. The purification results demonstrated that the target MIAP was eluted and identified as a homogeneous components by DEAE-cellulose ion exchange column,Sephadex G-100 and cellulose acetate membrane electrophoresis. Infrared spectral scanning suggested that MIAP was α-D-type terminated glucopyranose. Intuitive features showed that MIAP was soft and cottony white.</p><p><b>CONCLUSION</b>The extraction process with orthogonal test has been optimized and the acidic polysaccharide from Moerella iridescens is successfully isolated.</p>
Subject(s)
Animals , Bivalvia , Chemistry , Chromatography, DEAE-Cellulose , Methods , PolysaccharidesABSTRACT
<p><b>OBJECTIVE</b>To screen the gene expression profiles of IFN-alpha antiviral proteins based on a low-density cDNA Macroarray, and to explore the relationship between the expression of antiviral protein and the HBV replication.</p><p><b>METHODS</b>The HepG2 and HepG2.2.15 cells were treated with various concentrations of IFN-alpha (0 IU/ml, 100 IU/ml, 1000 IU/ml) of IFN-alpha for 6 h, and then the low-density cDNA Macroarray was used for analysing the expression profiles of antiviral genes and screening differential expressions of antiviral proteins. Meanwhile, the HepG2 cells were transiently transfected with HBV core protein-expressed plasmid pHBc-EGFP, and the expressions of antiviral proteins were analysed by RT-PCR assay. Moreover, the HepG2.2.15 cells were also transfected with the antiviral protein-expressed plasmid pcDNA3.1-Flag-MxA. ELISA was used for analysing the secreted HBV antigens, while dot blot and Southern blot were applied for analysing the extracellular HBV DNA and intracellular replicative intermediate HBV DNA in HepG2.2.15 cells. All data were presented as mean+/-SD and analyzed using the t-test and one-way analysis of variance (ANOVA) in the experiments.</p><p><b>RESULTS</b>The Macroarray results suggested that the expression of IFN-alpha antiviral genes like 6-16, IFITM1, IFITM2, IFITM3 and RING4 in HepG2.2.15 cells were partially inhibited. More importantly, it was found, in this research, the expression of antiviral protein MxA in HepG2.2.15 cells was completely suppressed. RT-PCR analysis indicated that the expression of MxA was also significantly decreased in HepG2 cells transfected with pHBc-EGFP plasmid. Although HepG2.2.15 cells transfected with pcDNA3.1-Flag-MxA plasmid could not inhibit extracellular HBV DNA and intracellular replicative intermediate HBV DNA, the MxA exerted some antiviral activities as it effectively suppressed the secretion of HBsAg and HBeAg in HepG2.2.15 cells.</p><p><b>CONCLUSIONS</b>HBV and its antigen components probably influence the expression of antiviral proteins. IFN- resistance may be related to the down-regulation of antiviral proteins expression.</p>
Subject(s)
Humans , Antiviral Agents , Pharmacology , Gene Expression Profiling , Gene Expression Regulation, Viral , Hep G2 Cells , Hepatitis B virus , Physiology , Interferon-alpha , Pharmacology , PlasmidsABSTRACT
The aim of this study was to investigate the expression of Tspan-1, Jab1 and p27 in human hepatocellular carcinoma (HCC) and their clinicopathological significance. The expression of Tspan-1, Jab1 and p27 was detected in HCC tissues, the tissues around cancer (76 cases), and the normal tissues around the liver hemangiomas (10 cases). The overexpression of Tspan-1 and Jab1 was found in HCC tissues, positively correlated with clinical stage and negatively correlated with survival rate. The expression of p27 was found inversely linked to which of Tspan-1 and Jab1. In conclusion, the expression of Tspan-1, Jab1 and p27 is significantly associated with development of HCC. Overexpression of Tspan-1 and Jab1 suggests poor prognosis but overexpression of p27 may expect good prognosis for patients with HCC.
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Carcinoma, Hepatocellular/metabolism , Cyclin-Dependent Kinase Inhibitor p27/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Liver Neoplasms/metabolism , Membrane Proteins/metabolism , Peptide Hydrolases/metabolism , Prognosis , Survival RateABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of NET-1 siRNA on NET-1 expression and on the proliferation and infiltration of skin squamous cell carcinoma cell line A431.</p><p><b>METHODS</b>Four recombinant vectors of pU6H1-GFP-siRNAs NET-1 were transfected into A431 cells. The levels of NET-1 mRNA expression were measured by semi-quantitative RT-PCR for selecting the most effective one in the four kind vectors of pU6H1-GFP-siRNA-NET-1. The controls consisted of pU6H1-GFP-siRNA-target off with random double-stranded RNA, pcDNA3.1 sense and other controls including the empty vectors (pU6H1-GFP and pcDNA3.1), NET-specific siRNA alone, lipofection reagent alone, target off siRNA alone, or untreated cells. After transfection, levels of NET-1 mRNA and protein expression were detected using semi-quantitive RT-PCR and Western blotting, respectively. The intracellular location of NET-1 protein was documented by immunofluorescence staining followed by laser scanning confocal microscopy. Proliferation rates of A431 cells were determined by MTT assay and flow cytometry. Abilities of migration and infiltration of A431 cells were determined by wound healing effect and transwell migration assay, respectively.</p><p><b>RESULTS</b>siRNAs coding NET-1 sequences (19-23nt) were confirmed between H1 and U6 promoters of pU6H1-GFP vector by sequencing. The transfection efficiency of pU6H1-GFP in A431 cells was about 80% as the percentage of GFP expression cells in total cells. Transfection with either pU6H1-GFP-siRNA NET-1 or pcDNA3.1 antisense NET-1 gave rise to an obvious reduction in the expression of NET-1 mRNA and protein in A431 cells. The abilities of proliferation, migration and infiltration of A431 cells were significantly reduced 48 hours after transfection with either pU6H1-GFP-siRNA NET-1 or pcDNA3.1 antisense NET-1, compared with the vector controls including pU6H1-GFP or pcDNA3.1 (P < 0.01, respectively). pU6H1-GFP-siRNA NET-1 showed a stronger inhibition effect on proliferation, migration and infiltration of A431 cells over pcDNA3.1 antisense NET-1 (P < 0.05). Transfection with pcDNA3.1 sense NET-1 resulted in an obvious increase in the expression level of NET-1 mRNA and protein, but without inhibition effect to proliferation, migration and infiltration of A431 cells.No obviously inhibition effects were observed when transfected with pU6H1-GFP-siRNA-target or the controls.</p><p><b>CONCLUSIONS</b>RNAi targeting NET-1 gene effectively down-regulates the expression of NET-1 in A431 cells, leading to an inhibition of proliferation, migration and infiltration. RNAi technique appears to be superior to antisense oligonucleotide technique in the suppression of gene expression.</p>