OBJECTIVE: To investigate the mechanism by which Chinese medicine Shengmai Yin (SMY) reverses epithelial-mesenchymal transition (EMT) through lipocalin-2 (LCN2) in nasopharyngeal carcinoma (NPC) cells CNE-2R. METHODS: Morphological changes in EMT in CNE-2R cells were observed under a microscope, and the expressions of EMT markers were detected using quantitative real-time PCR (RT-qPCR) and Western blot assays. Through the Gene Expression Omnibus dataset and text mining, LCN2 was found to be highly related to radiation resistance and EMT in NPC. The expressions of LCN2 and EMT markers following SMY treatment (50 and 100 µ g/mL) were detected by RT-qPCR and Western blot assays in vitro. Cell proliferation, migration, and invasion abilities were measured using colony formation, wound healing, and transwell invasion assays, respectively. The inhibitory effect of SMY in vivo was determined by observing a zebrafish xenograft model with a fluorescent label. RESULTS: The CNE-2R cells showed EMT transition and high expression of LCN2, and the use of SMY (5, 10 and 20 µ g/mL) reduced the expression of LCN2 and reversed the EMT in the CNE-2R cells. Compared to that of the CNE-2R group, the proliferation, migration, and invasion abilities of SMY high-concentration group were weakened (P<0.05). Moreover, SMY mediated tumor growth and metastasis in a dose-dependent manner in a zebrafish xenograft model, which was consistent with the in vitro results. CONCLUSIONS SMY can reverse the EMT process of CNE-2R cells, which may be related to its inhibition of LCN2 expression. Therefore, LCN2 may be a potential diagnostic marker and therapeutic target in patients with NPC.
Animals ; Humans ; Nasopharyngeal Carcinoma/genetics* ; Epithelial-Mesenchymal Transition ; Zebrafish ; Cell Proliferation ; Cell Line, Tumor ; Nasopharyngeal Neoplasms/radiotherapy* ; Cell Movement ; Gene Expression Regulation, Neoplastic

OBJECTIVE: To observe the effect of exosomes secreted by lipopolysaccharides (LPS)-stimulated macrophages on hepatic stellate cell activation and migration and explore the underlying molecular mechanism. METHODS: Human monocyte THP-1 cells were induced to differentiate into macrophages using propylene glycol methyl ether acetic acid (PMA, 100 ng/mL, 24 h) followed by stimulation with LPS, and the culture supernatant of macrophages was collected for extraction of the exosomes by ultracentrifugation. The expression of miR-155-5p in the exosomes was detected using qRT-PCR. A Transwell co-culture system was used to observe the effects of the macrophage-derived exosomes on LX2 cell (a hepatic stellate cell line) proliferation, migration, oxidative stress and the expression of fibrosis biomarkers, which were also observed in LX2 cells transfected with miR-155-5p-mimics or miR-155-5p-inhibitors. Western blotting was used to detect the expressions of SOCS1 and its downstream signal pathway proteins. RESULTS: Treatment with the exosomes from LPS-stimulated macrophages significantly enhanced the proliferation and migration ability of LX2 cells and increased the levels of oxidative stress and expressions of the fibrosis markers such as type Ⅰ collagen (P < 0.05). The expression of miR-155-5p in the exosomes secreted by macrophages was significantly increased after LPS treatment (P < 0.01). LX2 cells overexpressing miR-155-5p also exhibited significantly enhanced proliferation and migration with increased oxidative stress levels and expression of type Ⅰ collagen (P < 0.05), and interference of miR-155-5p expression produced the opposite effects. Western blotting showed that miR-155-5p overexpression obviously inhibited SOCS1 expression and promoted p-Smad2/3, Smad2/3 and RhoA protein expressions in LX2 cells (P < 0.05). CONCLUSION LPS stimulation of the macrophages increases miR-155-5p expression in the exosomes to promote activation and migration and increase oxidative stress and collagen production in hepatic stellate cells.
Humans ; Hepatic Stellate Cells ; Lipopolysaccharides/pharmacology* ; Collagen Type I ; Exosomes ; Macrophages ; MicroRNAs

Modern pharmacological studies have shown that Shengmai San has the effects of enhancing immunity and improving blood circulation, and Curcumae Longae Rhizoma(Jianghuang) has anti-inflammatory, anti-cancer, anti-oxidation and other functions. Shengmai San combined with Jianghuang is a new research direction in the study of anti-tumor of traditional Chinese medicines. The main treatment for nasopharyngeal carcinoma is radiation therapy, but radiation therapy can cause a variety of side effects, and it also changes the composition of the intestinal flora. In this study, the 16 s rDNA sequencing platform was used to perform macro-sequence sequencing of the intestinal flora samples of nude mice bearing the veins of Shengmai Jianghuang San, and then the results of intestinal flora data were analyzed to investigate the effect of Shengmai Jianghuang San on tumors. The results showed that Shengmai Jianghuang San combined with irradiation could enhance the therapeutic effect of tumor treatment. Radiation therapy would reduce the total number and diversity of intestinal flora in nude mice, and also change the structure of the flora. Shengmai Jianghuang San could protect the diversity of colonies, and also partially restore the colony imbalance caused by irradiation. This study provides a research idea for Shengmai Jianghuang San as a sensitizing adjuvant for radiotherapy of nasopharyngeal carcinoma.
Animals ; Drugs, Chinese Herbal ; pharmacology ; Gastrointestinal Microbiome ; drug effects ; Mice ; Mice, Nude ; Nasopharyngeal Carcinoma ; radiotherapy ; Radiation Tolerance ; Radiation-Sensitizing Agents ; pharmacology

OBJECTIVETo develop a rapid preimplantation genetic diagnosis method for -thalassemia SEA deletion based on blastocyst cell whole genome amplification (WGA) combined with short fragment Gap-PCR.

METHODSUsing multiple displacement amplification (MDA) WGA technique, we established a double-fluorescent PCR system of the housekeeping genes GAPDH and β-actin for WGA quality testing, and a genotyping PCR system of mutant and normal short sequences for α-thalassemia SEA deletion. The sensitivity and accuracy of this method for diagnosis of -thalassemia SEA deletion were evaluated by detecting lymphocyte samples containing different cell numbers from carriers of SEA deletion. The applicability of this method was evaluated by testing of 12 blastocyst biopsy samples.

RESULTSDetection of lymphocyte samples with different cell numbers using the method developed in this study revealed no ADO in 3-cell samples, and the product quantity of WGA became stable for 4-cell samples. Genotyping of the 10 blastocyst biopsy samples with successful WGA showed a genotype of --/ in 5 samples and / in the other 5 samples, which were consistent with the verification results.

CONCLUSIONSThe method developed in this study is a complete testing process for 4-6 blastocyst biopsy cells to allow rapid, accurate, and cost-effective PGD genotyping of -thalassemia SEA deletion using short fragment gap-PCR.

Objective To study the difference between intensity-modulated radiation therapy (IMRT) and three dimensional conformal radiation therapy (3D-CRT) for pelvic radiation of post-operative treatment with gynecologic malignant tumor. Methods A prospective investigation study was conducted on 183 patients of post-operative patients with whole pelvic radiation therapy of cervical cancer or endometrial cancer in Zhejiang Cancer Hospital [IMRT group (n=85) and 3D-CRT group (n=98)] from Oct. 2015 to Oct. 2016. The two groups received same dose (45 Gy in 25 fractions). Comparison of two groups with radiation dosimetry:the score according to the Radiation Therapy Oncology Group (RTOG) acute radiation injury grading standards before and after radiotherapy reaction, the score from functional assessment of cancer therapy scale-cervix (FACT-Cx) scale and expanded prostate cancer index composite for clinical practice (EPIC-CP) scale were also analyzed. Results (1) There were no significant effect with age, culture level, family economic condition and ratio of radiochemotherapy between two groups (all P>0.05). (2) Dosimetric comparison for IMRT vs 3D-CRT:the average dose of planning target volume (PTV) decreased(46.1 ± 0.4) vs(46.4 ± 0.5)Gy, V45 dose percentage increased(95.2 ± 1.0)%vs (93.3 ± 2.0)%, intestinal bag dose of V40 decreased(24.4 ± 6.8)%vs (36.5 ± 15.9)%, rectal V40 dose percentage decreased(73.9 ± 12.3)%vs (85.4 ± 8.4)%, and lower rectal V45 dose percentage(32.8 ± 13.4)%vs (71.5 ± 13.7)%, bladder V40 dose percentage decreased(55.5 ± 13.0)% vs (84.4 ± 13.0)%. Bone marrow V20 lower:(67.9 ± 5.4)% vs (79.5 ± 6.6)%, V10 lower:(82.1 ± 6.0)% vs (86.3 ± 6.6)%; there were significant differences (all P<0.05). There was no significant difference between the dose of V45 in the intestinal pouch and bladder (P>0.05). (3) Acute radiation injury classification for IMRT vs 3D-CRT:big or small intestine:Ⅱ-Ⅲreaction [13%(11/85) vs 24% (24/98); χ2=3.925, P=0.048], there was significant difference. Bladder: Ⅲ reaction [19% (16/85) vs 26% (25/98); χ2=1.171, P=0.279], there was no significant difference. Radiochemotherapy of bone marrow suppression:Ⅲ-Ⅳreaction (14/20), the incidence rate [26%(14/54) vs 31%(20/65);χ2=0.339, P=0.562], the difference was not statistically significant. (4) Quality of life scale by FACT-Cx scale in IMRT vs 3D-CRT:there were no significant difference before radiotherapy (82 ± 16 vs 85 ± 16;t=1.279, P=0.203), while there was significant difference after radiotherapy (76 ± 14 vs 71 ± 18;t=-2.160, P=0.032). EPIC-CP scale score:before radiotherapy they were (16±7 vs 15±6;t=-0.174, P=0.862) ,but after radiotherapy (18±7 vs 22± 7; t=3.158, P=0.002), there was significant difference between them. Before and after radiotherapy, the increased EPIC-CP scale of the IMRT group vs 3D-CRT group were 3 ± 4 and 6 ± 4, the 3D-CRT group was significantly higher, the difference was statistically significant (t=5.500, P=0.000). Conclusion IMRT has shown that there are a significant benefit for the post-operative patients with cervical cancer and endometrial cancer compared to 3D-CRT.

OBJECTIVETo investigate the effect of losartan in regulating oxidative stress and the underlying mechanism in mice with ventilator-induced lung injury.

METHODSThirty-six male C57 mice were randomly divided into control group, losartan treatment group, mechanical ventilation model group, and ventilation plus losartan treatment group. After the corresponding treatments, the lung injuries in each group were examined and the expressions of caveolin-1 and NOX4 in the lung tissues were detected.

RESULTSThe mean Smith score of lung injury was significantly higher in mechanical ventilation model group (3.3) than in the control group (0.4), and losartan treatment group (0.3); the mean score was significantly lowered in ventilation plus losartan treatment group (2.3) compared with that in the model group (P<0.05). The expressions of caveolin-1 and NOX4 were significantly higher in the model group than in the control and losartan treatment groups (P<0.05) but was obviously lowered after losartan treatment (P<0.05). Co-expression of caveolin-1 and NOX4 in the lungs was observed in the model group, and was significantly decreased after losartan treatment.

CONCLUSIONLosartan can alleviate ventilator-induced lung injury in mice and inhibit the expression of caveolin-1 and NOX4 and their interaction in the lungs.

Animals ; Caveolin 1 ; metabolism ; Losartan ; pharmacology ; Lung ; metabolism ; physiopathology ; Male ; Mice ; Mice, Inbred C57BL ; NADPH Oxidase 4 ; NADPH Oxidases ; metabolism ; Oxidative Stress ; Respiration, Artificial ; Ventilator-Induced Lung Injury ; drug therapy ; metabolism

OBJECTIVETo observe epithelial-mesenchymal phenotypes and oxidative stress related protein expressions of the liver cells in a rat model of liver fibrosis induced by bile duct ligation and recanalization.

METHODSTwenty-four male Wistar rats were randomized into 4 groups, including a sham-operated group, two bile duct ligation groups with ligation for 2 and 4 weeks, and a bile duct ligation group with a 2-week ligation followed by a 2-week recanalization. HE staining and Masson staining were used to assess liver fibrosis in the rats, and immunohistochemistry and Western blotting were employed to detect expressions of the epithelial and mesenchymal marker proteins and oxidative stress-related proteins.

RESULTSCompared with the sham-operated group, the rats with bile duct ligation showed obvious liver fibrosis, which worsened as the ligation time extended, accompanied by significantly increased expression of α-SMA, collagen I, NOX(4) and vimetin and reduced E-cadherin expression. Compared with the rats with bile duct ligation for 4 weeks, the rats in bile duct ligation-recanalization group showed obviously lessened liver fibrosis, significantly lowered expressions of NOX(4) and mesenchymal cell maker proteins, and enhanced expressions of epithelial cell marker proteins.

CONCLUSIONBile duct ligation up-regulates mesenchymal phenotype-related proteins and NOX(4) protein expression and down-regulates the expression of epithelial phenotype-related proteins, and these changes can be reversed by subsequent bile duct recanalization.

Actins ; metabolism ; Animals ; Bile Ducts ; surgery ; Cadherins ; metabolism ; Collagen Type I ; metabolism ; Disease Models, Animal ; Epithelial Cells ; cytology ; metabolism ; Hepatocytes ; cytology ; metabolism ; Ligation ; Liver Cirrhosis ; metabolism ; Male ; NADPH Oxidase 4 ; NADPH Oxidases ; metabolism ; Oxidative Stress ; Phenotype ; Rats ; Rats, Wistar ; Vimentin ; metabolism

Objective To observe the clinical role of Qiliqiangxin capsules uniting conventional western medicine interfering with chronic heart failure(CHF).Methods 61 cases of CHF were randomly recruited into a control group(31 cases)and a treatment group(30 cases).The control group was treated with conventional anti-hean failure treatment,and the treatment group was added Qiliqiangxin capsules on the treatment of the control group.After having been treated for 2 courses (fonr weeks),such indexes as clinical efficacy,body weigbt,urine volume,6 min walking distance(6MWD),electrocardiogram,and cardiac ultrasound were measured for both groups.Results The overall efficiency of cardiac function was 93.33%for the treatment group and 74.19%for the control group.There was significant difierence between the two groups(χ~2=4.07985,P＜0.05).The enhance rate of left ventricular ejection fraction(LVEF)was(16±4)%in the treatment group as compared to(11±3)%in the control group,also showing statistical difierence between the two groups(t=5.509,P＜0.001).Conclusion Qiliqiangxin capsules combined with conventional western medicine could effectively improve the cardiac function,LVEE clinical symptoms,and quality of life in the patients with ehronic heart failure.

OBJECTIVETo establish immortalized B lymphoblast cell lines (B-LCLs) from healthy anti-HBs antibody (anti-HBs)-positive volunteers and screen for human anti-HBs and the antibody-secreting cells.

METHODSThe peripheral blood mononuclear cells (PBMC) isolated from 3 healthy volunteers positive for anti-HBs with hepatitis B vaccine boost vaccination were infected with Epstein-Barr virus (EBV) and incubated in the presence of CpG DNA motifs and cyclosporin A (CyA). The anti-HBs in the culture supernatant of the immortalized B-cells was quantified by Architect anti-HBs assay with chemiluminescent microparticle technique. Immunocytochemistry was performed to identify the differentiation of the cell clones expressing anti-HBs.

RESULTSImmortalized B-cell culture was successfully established from the cell clones secreting anti-HBs with EBV infection and CpG DNA stimulation. The titer of anti-HBs in the culture supernatant was at its peak at 3 weeks of cell culture and then decreased gradually. At 3 months of cell culture, the cells still retained the capacity of anti-HBs production as verified by the results of immunocytochemistry for CD20 and CD138.

CONCLUSIONImmortalized B-cell culture secreting anti-HBs from volunteers receiving boost hepatitis B vaccination has been successfully established by modified EBV immortalization technique.

B-Lymphocytes ; immunology ; Cell Line ; Cell Transformation, Viral ; Hepatitis B ; prevention & control ; Hepatitis B Antibodies ; immunology ; Hepatitis B Surface Antigens ; immunology ; Hepatitis B Vaccines ; immunology ; Herpesvirus 4, Human ; immunology ; Humans ; Immunization, Secondary ; Vaccination

OBJECTIVETo investigate the expression of programmed cell death 5 (PDCD5) in tissues of normal human prostate (NP), benign prostatic hyperplasia (BPH), and prostate cancer (PCa) in order to assess the clinical role of PDCD5 in PCa.

METHODSPDCD5 expression was determined by EnVision immunohistochemical staining in formalin-fixed and paraffin-embedded specimens obtained from 12 subjects with NP, 22 with BPH, and 22 with PCa. In addition, PCa cases were classified as low/middle-risk (Gleason sum < or = 7) and high-risk (Gleason sum >7) on the basis of Gleason grade. Positive expression rates and intensity of PDCD5 protein were observed under light microscope and analyzed with computer imaging technique. Expression of PDCD5 was compared among different prostatic tissues.

RESULTSThe expression of PDCD5 was significantly lower in tissue of PCa than in tissues of NP and BPH (P<0.01). However, there was no significant difference in PDCD5 expression between tissues of NP and BPH. In addition, the expression of PDCD5 was further downregulated with the increase of Gleason sum in PCa.

CONCLUSIONSBy downregulating apoptosis, low PDCD5 expression may play an important role in the occurrence and development of PCa. PDCD5 is supposed to have a potential clinical value to be a new predictor of progression and target of gene therapy in PCa.

Aged ; Apoptosis ; Apoptosis Regulatory Proteins ; analysis ; physiology ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Neoplasm Proteins ; analysis ; physiology ; Prostate ; chemistry ; Prostatic Neoplasms ; chemistry ; etiology ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; analysis