Journal of Jilin University Medicine Edition

Objective: To investigate the effect of lidamycin (LDM) combined with bortezomid (BZM) against multiple myeloma and the mechanism of the combination treatment. Methods: The human multiple myeloma SKO-007 cells in logarithm growth phase were selected and randomly divided into control group, LDM group, BZM group, and BZM combined with LDM group. MTS assay was used to detect the survival rate of SKO-007 cells and flow cytometry was used to analyze the distribution of cell cycle and apoptotic rate of the proliferation cells in various groups; the expression levels of protein associated with apoptosis and endoplasmic reticulum stress (ERS) of SKO-007 cells in various groups were detected by Western blotting method. Results: After culture for 48 h, the survival rate of the cells in BZM combined with LDM group was lower than those in control, LDM and BZM groups (P<0.05). The percentage of cells at G 2/M phase, the apoptotic rate of cells, the expression levels of cleaved caspase-3 and poly ADP-ribose polymerase (PARP) of SKO-007 cells, the expression levels of GRP78/Bip, CHOP/GADD153, and phosphorylation of c-Jun NH2-terminal kinase (p-JNK) in BZM combined with LDM group were higher than those in control, LDM and BZM groups (all P<0.05). Conclusion: BZM can greatly enhance the efficacy of LDM against multiple myeloma by increasing the levels of cleaved caspase-3 and PARP, and remarkably increase the apoptosis induced by LDM through further activation of ERS.

Objective: To investigate the effects of prion protein 106-126 peptide on inducing apoptosis in differentiated PC12 cells. Methods: After differentiated by nerve growth factor (NGF), the PC12 cells were infected by prion protein 106-126 peptide. Cell viability was determined by using the MTT assay and the morphological changes were observed. Apoptotic was detected by morphological changes, DNA electrophoresis and flow cytometry. Results: Infected by this peptide, cell viability decreased 28.9%. Under light microscope, cells were shrinked and rounded, many cells were divorced from plate wall, some neuraxon shortened and broke. Apoptotic cells which nucleolus shrinked and rounded could be coloured orange by fluorescent colouration. Under electron microscope, chromatin gathered along the inside of the nuclear membrane, vacuole bodies appeared. Apoptotic peak was detected by flow cytometry and the ladder band appeared in DNA electrophoresis. Conclusion: Prion protein 106-126 peptide can induce apoptosis in differentiated PC12 cells and present cellular toxicity definitely. It might be a perfect model to study the cellular toxicity of prion protein.

Objective: To explore the correct test ways of the ductus venosus (DV) blood flow spectrum of the normal fetal and the rule of changes of the normal fetal DV blood flow spectrum along with gestational week, and to establish the reference ranges of the parameters of blood flow. Methods: A total of 320 cases of single fetal pregnant women within 11-40 weeks were selected as the objects. Using color doppler echocardiography, the ventricular systolic peak velocity (SV), atrial systolic trough (aV), pulse index (PD and resistance index (RI) were measured, and the ratio of ventricular systolic peak velocity trough ratio (S/a) from ultrasonic standard section was calculated. All of these were analysed statistically. Results: The doppler spectrum of normal fetal DV blood flow showed the same three-phase waves, SV was increased with the increase of gestational age (F=27. 00, P=0.000), and aV was elevated with the increase of gestational age (F=389. 81, P = 0.000), while PI (F= 65.41, P=0.000), RI(F=58.82, P=0. 000) and S/a ratio (F=47.79, P=0. 000) were decreased with the increase of gestational age. Conclusion: The performance of color Doppler flow imaging of normal fetal DV showed that the blood flow peak speed is increased along with the increase of gestational age and PI, RI and S/a are reduced with the increase of gestational age, and RI has the largest decline, which has important diagnosis value.

Objective: To investigate the induction of tanshinone [1 A (Tan fl A) on the differentiation of human placenta-derived mesenchymal stem cells (hPDMSCs) into cardiomyocytes, and to provide an experimental basis for Tan IT A as a cardiomyocytc differentiation inducer. McthodS; The hPDMSCs were treated with different concentrations of Tan Fl A (0.1.0.2.0.4.0.6.0.8.1.0. 2.0. 4.0. 6.0. 8.0. and 10. 0 mg • L ). and the nontoxic dose of Tan II A <0. 1 mg • L ) was scrccncd by MTT assay for experiment. The hPDMSCs were divided into control group. 5-aza induction (10 pmol • L ) group, and Tan Fl A induction (0. 1 mg • L ) group. After culture for 20 d. the expressions of o-sarcomeric acun (o-SCA) in the cells in various groups were detected with lmmunohistochcmastry; the positive expression rates of cardiac troponin 1 (cTnl) in the cells in various groups were detected with immunofluorescence, and the differentation rates of cardiomyocytes were calculated. The expression levels of GATA-binding protein 4 (GATA4). atnal natriuretic factor (ANF). cTnl. glycogen synthase kmase-30 (GSK-30 ) and 0-catemn in the cells were detected with Western blotting method. Results: The biological characteristics of hPDMSCs accorded with the mesenchymal stem cells. The MTT results showed that when the concentration of TanFl A was more than 0. 1 mg • L . the cell survival rates were decreased with the increase of concentrations the cells in control group showed a rapid growth trend before 12 d. and the proliferation activities of the cells began to decrease on the 12th day. Compared with control group, the cell activities in 5-aza induction group and TanFl A induction group were significantly decreased < P<0. 05). The immunohistochcmistry staining results showed that the cells in control group didn' t express a-SCA. and the cells in 5-aza induction group and TanFl A induction group expressed o-SCA. cspcaally in Tan Fl A induction group. Compared with control group, the expression levels of GAT A4 ( i - 2.937. P <0.05,4.769, P ,| , <0. 05) . ANF ( t -3.728. P <0.05 j r.-5.912, P ., ,<0.05). cTnl <f -3.623. P <0.05; f .-7.153, P.<0.05) and GSK-30 < t -2.995. P <0.05; tr.. i a - 5. 420. P <0.05) proteins in the cells in 5-aza induction group and Tan FI A induction group were significantly increased, and the expresson levels of (J-catcmn (t 2.985, P <0.05, l .1,-6.951, P .<0.05) protein were significantly decreased; compared with 5-aza induction group, the expresson levels of GATA4. ANF. and GSK-30 proteins in TanFl A induction group were increased < P<0. 05). Conclusion: Tan Fl A can induce the differentiation of hPDMSCs into cardiomyocytes. which has better effect than 5-aza. and its mechanism may be related to inhibiting the Wnt/|3-catcnin signaling pathway.

Objective: To analyze the genotypes of clinical isolates of Mycobacterium tuberculosis in Jilin Province with multiple locus variable numbers of tandem repeats(MLVA)genotyping. Methods: Mycobacterium tuberculosis strains were isolated from patient samples in Jilin Province. Genomic DNA was extracted and 12 variable numbers of tandem repeats(VNTR) loci were amplified. The gels were analyzed by Totalab software and the results were clustered with BioNumerics(6.0). Results: Analysis of a total of 110 isolates with the 12 VNTR loci showed great genetic polymorphism in these isolates. Nine of the 12 loci showed the Hunter-Gaston index(HGI) were higher than 0.5. The clustering results showed that total of 6 genotype groups could be defined, of which group 5 was the most represent one, accounting for 34% of the isolates. The less represented genotypes included group 1, accounting for 15%. Strains of group 5 mainly distributed in Tonghua and Songyuan. Conclusion: Mycobacterium tuberculosis isolates in Jilin Province shows great genetic polymorphisms. Different cities have different genotype profiles.

Objective; To explore the effects of miR-124a on the expression levels of tumor necrosis factor alpha (TNF-a) and interleukin-6 (IL-6) and cell cycle in the macrophage J774. 1 cells of the rheumatoid arthritis (RA) model mice, and to elucidate the mechanism of miR-124a in the pathogenesis of RA. Methods: The J774. 1 cells in logarithmic growth phage were obtained and uniformly inoculated in the petri dish with 1 × 10⁶ mL⁻¹, and there were 3 multiple holes in each group; transfection was carried out when the fusion degree of the cells reached 60%. The cells were divided into blank control group (without any treatment), empty vector group (transfected with adenovirus negative fluid) and miR-124a overexpression group (transfected with miR-124a adenovirus vector). The expression levels of TNF-α and IL-6 in supernatant of the cells in three groups were detected by ELISA 48 h after transfection. RT-PCR was used to detect the relative expression levels of TNF-α and IL-6 mRNA in J774. 1 cells 48 h after transfection and the changes of cell cycle were detected using flow cytometry. Results: The expression levels of TNF-α and IL-6 in supernatant of the cells in miR-124a overexpression group were lower than those in blank control group and empty vector group 48 h after transfection (P=0. 038, P=0. 042; P=0. 043, P=0. 044). The expression levels of TNF-α mRNA and IL-6 mRNA in miR-124a group were significantly lower than those in blank control group and empty vector group (P = 0. 001, P=0.002; P=0. 001, P=0. 003). The pecentage of cells in Gi phase in miR-124a overexpression group was higher than those in blank control group and empty vector group (P<0. 01); the percentages of cells in S phase and G2 phase were lower than those in blank control group and empty vector group (P<0.01). Conclusion: MiR-124a can decrease the expression levels of inflammatory cytokines after transfecting the macrophages J774. 1 cells, and inhibit the proliferation of cells. MiR-124a is a potential therapeutic target for the treatment of RA.

Objective: To discuss the clinical effects of implantation of radioative ¹²⁵T seeds by the way of percutaneous puncture and laparotomy under the guidance of ultrasound in the treatment of locally advanced pancreatic cancer, and to provide the basis for choosing surgical methods in treating advanced pancreatic cancer. Methods: The clinical materials of 73 patients with advanced pancreatic cancer were collected, including 42 patients who underwent implantation of radioactive ¹²⁵I seeds by percutaneous puncture (group A) and 31 patients who underwent impantation of radioactive ¹²⁵I seeds by laparotomy (group B). The pain relief, local control of tumor, postoperative survival time and complications of the patients were compared between two groups. Results: The rates of pain relief of the patients in group A and group B were 91.89% and 86.40%, and there was no significant difference (P = 0.815). The local control rates of the patients in group A and group B were 71.43% and 77.42%, and there was no significant difference (P = 0.564). The medium survival time of the patients in group A and group B were 11 months and 12 months; the one-year survival rates were 36.9% and 35.8%, and there was no significant difference (P = 0.664). Seven patients in group A got fever; in group B, six patients got fever, two got calf muscle venous thrombosis, one got gastric retention, one got bilioentric anastomosis, one got abdominal distension and one got intestinal obstruction in the early stage after operation. The incidence rates of complications of the patients in two groups were 16.67% and 38.71%, and there was significant difference (P = 0.034). Conclusion: Percutaneous implantation of radioactive ¹²⁵T particles guided by ultrasound causes less complications in the treatment of locally advanced pancreatic cancer. Moreover, the percutaneous way reaches the same effect as the intraoperative way does on the pain relief, local control of tumor and survival time prolonged.

Objective: To investigate the possibility and mechanism of 125 I in treatment of glioma. Methods: SHG-44 glioma cells were cultivated in vitro, the inhibitory effect of 125 I on SHG-44 cell proliferation was determined by MTT method. The stereotactic method was used to establish the rat intracranial glioma model. The MRI was performed at 1st week after implantation and 125 I was implanted in the glioma area, the MRI was performed to measure the diameter of tumor 2 weeks after implantation. The rats were killed after 2 weeks, PCNA gene experession was determined with immunohistological method both in control and experiment group. Results: one week after implantation the glioma grew, the results of MTT method showed the growth of the SHG-44 was inhibited, 125 I inhibited the expression of PCNA gene and enlonged the rat survival period. Conclusion: 125 I can inhibit the growth of glioma, the mechanism may be concerned with its inhibitory effect on PCNA gene expression.

Objective: To investigate the effects of 125I seed on the growth and programmed cell death of human glioma cell line SHG-44, and clarify the action mechanism of the related genes in this process. Methods: SHG-44 glioma cells were cultivated in vitro, and divided into control group and treatment group. The inhibitory effect of 1251 on SHG-44 cell proliferation was determined by MTT method. The morphological changes of SHG-44 cells were examined by electron microscopy, TUNEL method and Acridine Orange/Ethidium bromide double fluorescent staining. The gene expresion profile was established by using gene chip technique. Results: 125I seed had inhibitory effect on the proliferation of SHG-44 cells cultivated in vitro in a dose and time-dependent manner. The result of MTT showed that the A value of SHG-44 cells treated with 125I seed was significantly decreased with the increasing of radiation dose and prolongation of time. The inhibitory rate of SHG-44 cells after treated with 125I seed for 3 d was 50%, there was significant difference compared with control group (P<0.05). Autophagy was frequently observed under transmission electron microscope in SHG-44 cells. The number of cells at S phase was reduced while the number of cells at G 1 and G2/M phase was increased. Although the apoptosis in SHG-44 cells was increased, but not more than 2%. There were 56 differential expression genes of SHG-44 cells after exposure to 125I including 36 up-regulation genes and 20 down-regulation genes. Conclusion: 125I seed can inhibit the proliferation of SHG-44 glioma cells in a dose-dependent manner by inducing the programmed cell death. There may be non-apoptotic programmed cell death (autophagy) in SHG-44 glioma cell line after induction by 125I at relatively low concentration. These results suggest that the mechanism of 125I-induced proliferation inhibitory effect and apoptosis in SHG-44 cells may be related to the genes of p53 and ATM pathway, C-myc family, p16 family, Bcl-2 family, TNF ligand family and TNF receptor family genes.

Objective; To evaluate the clinical effect of pereutaneous vertbroplasty (PVP) combined with implantation of iodine-125 0"T) radioactive particle in the treatment of vertebral metastasis, and to provide basis for the treatment of vertebral metastasis. Methods: A total of 69 patients with vertebral metastasis were divided into test group (n=32) and control group (n=37); the patients in test group were treated with PVP comined with implantation l2;T radioactive particle and the patients in control group were treated with PVP only. The heights of anterior and posterior vertebral bodies of the patients before and after treatment were detected by X-ray. The numerical rating scale (NRS) scores, pain relief rate and the incidence of surgical complications of the patients were recorded before operation and Id, 1 week, 1 month, 3 months, and 6 months after operation. Results: The operation was successfully performed in all the patients without local bleeding; there were no movement dysfunction and nerve compression phenomenon. There was no leakage of bone cement. All the 12 T radioactive particles located well and there was no particle obscission. The heights of vertebral bodies of the patients in two groups after operation were increased compared with before operation (P<0. 05). The NRS scores of the patients in two groups s at 1 d, 1 week, 1 month, 3 months, 6 months after operation were significantly decreased compared with before operation (P<C0. 05); compared with control group, the NRS scores of the patients in test group at 1 d, 1 week, 1 month, 3 months, 6 months after operation were decreased (P<C0. 05). The incidence of pulmonary embolism or radiation myelitis complications was about 4. 3% in 69 patients. Compared with control group, the difference in the incidence of complications of the patients in test group was not significant (P<0.05). Conclusion: PVP combined with '"'I radioactive particle implantation is a safe and effective method in the treatment of vertebral metastasis, which can relieve the pain of the patients obviously compared with PVP.

Objective: To observe the cell count changes of A549 cell line and human embryonic lung diploid cell line 2BS after irradiated by 125I seeds with different doses, and to study the growth inhibition of 125I on this two kinds of cell lines, and to determine its clinical safety dose in treatment of non-small cell lung. Methods: 125I seeds with different doses (low dose: 0.2 mCi, mediate dose: 0.4 mCi, high dose: 0.8 mCi) were chosen and put into A549 cells and human embryonic lung diploid cell line 2BS in vitro, the cells on the 2nd, 4th, 6th and 8th days after irradiation were collected, the alive cells were counted by cells dyeing experiments, then the growth curves were drawn, and the IC 50 of the radioactive 125I seeds to both two cell lines were calculated. Results: Compared with blank and control groups, the cell proliferation trend of A549 cells in low dose group was not significantly influenced(P>0.05), but the growth of A549 cells in mediate and high dose groups were inhibited in a time-dependent manner, there were significant differences(P<0.05), the most obvious change was on the 6th day. The IC 50 of the radioactive 125I seeds to A549 cells was about 0.4 mCi. While the growth inhibition of 125I 2BS had no statistically significant differences between various dose groups(P>0.05), and the IC 50 of the radioactive 125I seeds to 2BS cell line was about 1.65 mCi. Conclusion: 0.4 mCi of radioactive 125I seeds has already had the obvious damage effect on A549 cells, 0.8 mCi of radioactive 125I seeds has the stronger effect. The IC 50 of the radioactive 125I seeds to 2BS cells is about 1.65 mCi, so the clinical safety dosage is 0.4 -0. 8 mCi.

Objective: To study the anti-tumor effects of 125I radioactive particles implantation on transplantated tumor model of human breast cancer cells in nude mice and clarify their anti-tumor mechanisms. Methods: 120 nude mice transplantated with human breast cancer cells MCF-7 were randomly devided into 3 groups (n = 40) : 125 I radioactive particles implanted group, non-radioactive particles implanted group and non-particles implanted group. The articles were implanted into mice according to Pairs system principle. The expressions of Fas mRNA and protein and the activaties of caspase-3 and caspase-8 enzyme were detected by RT-PCR and Western blotting. The changes of cell cycle were detected by flow cytometry. Results: Compared with non-radioactive particles implanted group and non-particles implanted group, the size of cancer tisses in 125I radioactive particles implanted group was reduced significantly(P< 0.05). The expressions of Fas mRNA, Fas protein, caspase-3 and caspase-8 were incresed remarkably (P< 0.05). The expressions of caspase-3 and caspase-8 were more than 0.6. The flow cytometry results showed that the number of cells in G0/G1 phase was significatly increased(P<0.05) while the number of cells in S phase was decreased remarkably (P<0.05). Conclusion: The implantation of 125I radioactive particles into transplantated tumor model of human breast cancer cells can kill tumor cells, inhibit the growth cycle of tumor cells and induce the apoptosis of tumor cells in nude mice.

Objective: To evaluate the clinical efficacy of thermal-therapy combined with ¹²⁵I radioactive particles by comparing the results of preoperative and postoperative images, serum carcino-embryonic antigen (CEA) and following-up after treatment of the patients with recurrent rectal cancer. Methods: Nineteen rectal cancer patients with postoperative recurrence after radiotherapy were treated with ¹²⁵1 implantation guided by CT (20 cases were treated with operation, but 1 patient was exited who couldn' t tolerate the thermal-therapy). Radioactive particle treatment planning system (TPS) was used to make the plan before the particle implantation, and the distance of the particle was 1.0 cm; all patients received CT scanning and were quantificationally evaluatated after implantation; the number of seeded particles was about 12 - 58, the radioactivity of the ¹²⁵I particle was 0. 5 mCi, and the matching dose surrounding the tumor was 90 - 140 Gy. All patients were treated with microwave thermal-therapy after particle implantation, 60 min per time, lasting the temprature at 43'C , twice per week, for three weeks. And the patients were followed up after treatment for 3 months, the efficacy was evaluated by image and CEA results, and the urinary frequency, dysuria, hematuria and rectal bleeding were evaluated. Results: Compred with before treatment, after following up for 6 months, the image results showed that the tumor volume was reduced, and the blood CEA level of all patients were decreased from (30.25±8. 32) mg • L⁻¹ to (11. 89±5.22) mg • L⁻¹ (i = 13.158, P<0.01)i the local efficacy was 94.7% (18/19), and the pain relief rate was 94.4% (17/18), the NRS median was 6 (4, 7) before treatment, and it was 1 (0, 3) after treatment, there was significant difference (P<0.01). There were no frequent urination, urinary pain, hematuria, rectal bleeding and other complications in all patients. Conclusion: Thermal-therapy combined with ¹²⁵I radioactive particle implantation has good curative effects on the recurrent rectum cancer, and is effective means for recurrent rectum cancer treatment.

Objective: To establish the method of using the 12th vertebrae for sex determination of adult Chinese and evaluate its effect. Methods: The 12 th thoracic vertebrae were developed by the clinic abdomen CT images. A total of 25 linear measurements on 7 aspects of the vertebrae were measured and 4 ratios were calculated. The items were selected which had the significant difference to establish the sex determination equation and its effect was evaluated. Results: Of the total 29 traits, 27 were sexually dimorphic (P< 0.05), the accuracy was 56.3%-89.2%, 8 traits had the accuracies mean or over 80.0%. The trait iVL had the highest accuracy of 89.2%. A function with four variables predicting sex with 90.8% accuracy was derived by using stepwise method of discriminant function analysis: Y= 2.98 X iBDsm + 1.97 X PH + 3.37 X BHp + 3.27 X sVL/BHa - 32.80 (mean centroids =-7.69). Conclusion: The method of using the selected traits for sex determination of adult Chinese is practicable and it has a relatively high accuracy.

Objective: To detect the expression levels of urokinase-type plasminogen activator (uPA), matrix metalloproteinase-3 (MMP-3), MMP-9, MMP-13 and MMP-14 in the patients with osteoarthritis(OA) before and after arthroscopic debridement, and to explore the influence of arthroscopic debridement in the expressions of uPA, MMP-3, MMP-9, MMP-13, and MMP-14. Methods: 420 cases of synovial fluid from knee OA patients undergoing arthroscopic debridement were obtained before operation. After six months follow-up, 350 cases of synovial fluid samples were obtained and according to inclusion and exclusion criteria, 228 synovial fluid were selected to analyze. The expression levels of uPA, MMP-3, MMP-9, MMP-13, and MMP-14 were measured by ELISA assay. Pain intensity of these patients before operation and six months after operation were recorded using the Visual Analogue Scale/Score(VAS). The differences of the expression levels of uPA, MMP-3, MMP-9, MMP-13, and MMP-14 between before operation and after operation were compared. The relationship between the expression levels of uPA, and MMP-3, MMP-9, MMP-13, MMP-14 and VAS was analyzed with Spearman analysis. Results: All the patients were followed up for 36.5 months. Compared with before operation, the expression levels of uPA and MMP-3 in the synovial fluid of the patients after arthroscopic debridement were significantly decreased(P<0.01), the expression levels of MMP-9 and MMP-13 were also decreased (P<0.05), but the MMP-14 expression level showed no significant change. The expression levels of uPA, MMP-3, MMP-9, MMP-13, MMP-14 were positively associated with VAS before arthroscopic debridement (r=0.361, r=0.417, r=0. 136, r=0.514, r=0.156, P<0.05); uPA and MMP-3 were positively correlated with VAS after arthroscopic debridement(r=0.981, r=0.831, P<0.01), as well as the expression level of MMP-13 and VAS, but there were no significant differences between the expression levels of MMP-9, MMP-14 and VAS. Conclusion The decreased levels of uPA, MMP-3 and MMP-13 in synovial fluid may contribute to the pain-relief effects of arthroscopic debridement.

Objective: To investigate the effect of IL-12 on mediator release from mast cells and the potential signal transduction pathways correspondingly. Methods: P815 cells were challenged with various concentrations of IL-12. The supernatants were collected and analyzed by enzyme-linked immunosorbent assay (ELISA) to detect the quantity of released IL-13, IL-6 and histamine. The cells were treated and analyzed by cellular activation of signal ELISA (CASE) to detect phosphorylation of ERK and P38. Results: After incubated with different doses of IL-12 (0, 1.0, 10.0 and 100.0 μg · L-1 the IL-13 release of P815 cells increased with the concentration of IL-12 compared with medium alone control (P<0.05); but there were no significant differences of IL-6 release and histamine release compared with mediun alone control. After P815 cells were incubated with PD98059 and U0126 prior to IL-12, the percentage of intracellular phosphorylation of ERK and IL-13 release decreased significantly compared with groups without inhibitor (P<0.05). After P815 cells were incubated with SB203580 prior to IL-12, the percentage of intracellular phosphorylation of P38 and IL-13 release didn't change compared with groups without inhibitor. Conclusion: IL-12 induced IL-13 release from P815 cells is likely through activation of ERK signalling pathway.

Objective: To investigate and evaluate the curative effect of radioimmunological targeting drug on nude mice bearing breast cancer. Methods: The anti-CEA monoclonal antibody C50 was combined with 131 I to produce radioimmunological targeting drug. 16 nude mice inoculated subcutaneously with breast cancer cell MCF-7 with tumor diameter about 0.5 cm were randomly into 4 groups (n=4): group I, injected in part with 131 I-C50 18.5 MBq; group II, injected in part with 131 I-C50 3.7 MBq; group III, injected in part with 131 I-mIgG 18.5 MBq; group IV, injected in part with C50 0.75 μg. The size of tumor volume and inhibitory rate UR) after treatment for six weeks were calculated and compared with the control group. Results: The tumor volume and curves for tumor growth and tumor weight had significant differences between group I and the group III as well as group II (P<0.01); and there was significant difference between group I and group II (P<0.05), there was no significant difference between group III and group IV (P>0.05). Conclusion: Anti-CEA monoclonal antibody C50 labeled with radionuclide 131 I could inhibit the growth of the tumor when given locally. 131 I-C50 has a potential value of clinical application.

Objective: To research the relationship between hypoxia-inducible factor-1 alpha (HIF-1α) expression and the recurrence of differentiated thyroid carcinoma (DTC) after complete thyroidectomy combined with 131I treatment, and explore its possible mechanism of recurrence. Methods: The recurrence rate of 173 cases of DTC after complete thyroidectomy within 3 years was analyzed, according to the follow-up detecting data of Tg, TgAb, B-mode ultrasonography, X-ray scanning and global 131I imaging, the cases of recurrence were used as recurrence group, the same number cases without recurrence as control group which were chosen randomly. The HIF-1α expressions in recurrence cancer tissues and un-recurrence cancer tissues were detected with immunohistochemical staining and RT-PCR technique. Results: The 3-year recurrence rate after thyroidectomy was 5.20% (9/ 173), among them, the recurrence rate of papillary carcinoma was; 4.90% (5/102), while the recurrence rate of follicle carcinoma was 5.63% (4/ 71), there was no significant difference between two groups (P>0.05). The HIF-1α expression levels in recurrence tissues of papillary carcinoma and follicle carcinoma were significantly higher than those in unrecurrence tissues (P<0.05). Conclusion: Several recurrence cases can be still found in DTC cases after complete thyroidectomy eombined with 131I treatment, one of its mechanisms may be related to significant increase of the HIF-1α expression level in cancer tissue.

Objective: To label anti- c-erbB-2 monoclonal antibody with 131I, and investigate its inhibitory effect on the growth of ovarian cancer in mude mice. Methods: Anti-c-erbB2 monoclonal antibody was combined with 131I to produce radioimmunological targeting drug. The nude mice which had been inoculated subcutaneously with ovarian cancer SKOV3 cells and the diameter of tumor mass was about 1.0 cm were randomly divided into 3 groups: G1, G2 and control groups. The mice in G1 and G2 groups were treated with 11.1 MBq and 3.7 MBq anti-c-erbB2 monoclonal antibody labeled with 131I respectively. The mice in control group was treated with mIgG labeled with 131I. The size and weight of tumor were determined once every week during 6 weeks after administration. Results: Compared with control group, the size and weight of tumor in G1 and G2 groups were significantly decreased, and the inhibitory rate was increased (P<0.05). The size and weight of tumor in G1 group were lower than those in G2 group (P<0.05), and the inhibirory rate in G1 group was higher than that in G 2 group (P< 0.05). Conclusion: Anti-c-erbB-2 monoclonal antibody labeled with radionuclide 131I could inhibit the growth of ovarian cancer in nude mice and the effect of 11.1 MBq c-erbB-2 monoclonal antibody labeled with 131I is better than 3.7 MBq 131I-c-erbB-2.

Objective: To discuss the relationship between coagulation function and gastrointestinal hemorrhage in premature infants. Methods: The prothrombin time (PT), part-time activation thromboplastin (APTT), thrombin time (TT), fibrinogen (Fib), D-dimer (D-D) in 132 hospitalized premature infants with gastrointestinal hemorrhage (observation group) and 156 non-hemorrhage premature infants (control group) were detected. The difference of coagulation function between two groups was compared. Results: Compared with control group, the PT, APTT, and TT in observation group were significantly increased (P<0.01), Fib was decreased (P<0.01) and D-D was obviously increased (P<0.01). Conclusion: The changes of coagulation function are closely related to gastrointestinal hemorrhage in premature infants and may be involved in the pathogenesis of gastrointestinal hemorrhage in premature infants.

Objective: To investigate the inhibition of proliferation and induction of apoptosis by proteasome inhibitor MG-132 on C6 glioma cell in vitro. Methods: Rat glioma C6 cells were cultured with different concentrations of proteasome inhibitor MG-132 (10, 20 and 50 μmol • L-1). Cell viability was determined by MTT assay at different cultured periods. Flow cytometry was used to detect apoptosis change and HE and AO/EB staining and electronic microscope were used to detect the morphological changes of apoptotic cells. Results: Compared with normal control, MG-132 significantly reduced the viability of C6 cells in 10, 20 and 50 μmol • L-1 MG-132 groups (P< 0.01). Moreover, the inhibitory effect was higher on high concentration MG-132 compared with low dose MG-132. After treatment with 10 μmol • L-1 MG-132 for 24 h, the apoptosis characteristic C6 cells were detected by AO/EB and HE staining. Apoptotic sub-G was detected by flow cytometry in C6 cells incubated with 10 μmol • L-1 MG-132 for 12 h and displayed a time-dependent manner, the apoptotic percentages in 10, 20 and 50 μmol • L-1 MG-132 groups were significantly higher than that in control group (P<0.01). Conclusion: Proteasome inhibitor MG-132 can inhibit C6 cell proliferation and induce C6 cell apoptosis.

Objective: To construct lentiviral vector which can overexpression miR-137 and produce lentivirus by lentivirus packaging system, and to explore its infection efficiency and expression in HEK293T cells. Methods: miR-137 sequence was chemically synthesized and cloned into lentiviral vector GV209, and the recombinant plasmid containing human miR-137 was obtained and identified. Then miR-137 recombinant plasmid together with two helper plasmids were transfected into HEK293T cells using Lipofectamine 2000. After the HEK293T cells were infected in multiplicity of infection (MOD 40 for 48 h, the expression of green fluorescent protein (GFP) was observed by fluorescence microscope and the expression level of miR-137 was detected by fluorescence quantitative PCR. Results: The sequencing results showed that the inserted gene sequence was completely consistent with the published human miR-137 gene sequence in GenBank. The GFP was observed in the HEK293T cells infected with miR-137 overexpression lentivirus under fluorescence microscope. The fluorescence quantitative PCR results showed that the expression level of miR-137 in the cells infected with overexpression lentivirus was 12.74 times higher than that in the control cells. Conclusion: The lentivirus containing miR-137 gene is successful packaged, and it could efficiently infect the HEK293T cells.

Objective：To investigate the effect of miRNA-138-5p overexpression on cisplatin（DDP）resistance of the human breast cancer MCF-7 cells ， and to elucidate its possible mechanism. Methods ：The MCF-7 cells were induced by concentration gradient increasing method to establish the DDP cell strain MCF-7/DDP. Then miR-138-5p mimics or hypoxia inducible factor-1α（HIF-1α）overexpression plasmid were transfected into the MCF-7/DDP cells separately or simultaneously. The MCF/DDP cells were divided into negative control group（NC group，transfected with miR-138-5p mimics negative control），miR-138-5p mimics group（transfected with mir-138-4p mimics），miR-138-5p mimics+ Vector group（transfected with miR-138-5p mimics and empty plasmid）and miR-138-5p mimics+HIF-1α group（tranfected with miR-138-5p mimics and HIF-1α overexpression plasmid）；and the other MCF-7/ DDP cells without transfection were used as blank control group. The cells were treated with different concentrations（0，10，20，40，80 and 100 μmol·L－¹）of DDP for 24 h and the proliferation activity of cells was detected by MTT，and half inhibitory concentration （IC50） and drug resistance index （RI） were calculated. The expression levels of miR-138-5p and HIF-1α mRNA in the cells were detected by Real-time fluorescence quantitative polymerase chain reaction（qRT-PCR）；flow cytometry was used to detect the apoptotic rate of cells；Western blotting method was used to detect the expression levels of HIF-1 α，P-gp and MRP1 proteins in the cells. TargetScan software was used to predict the target binding sites of miR-138-5p and HIF-1α，and the dual luciferase reporter system was used to verify the targeting relationship between miR-138-5p and HIF-1α. Results：The IC50 of drug-resistant cell strain MCF-7/ DDP for DDP was significantly higher than that of its parent MCF-7 cells（P<0. 01），and the RI value was 5. 72. Compared with parental MCF-7 cells，the miR-138-5p expression level in drug-resistant cell strain MCF-7/DDP was markedly reduced（P<0. 01），while the HIF-1α mRNA and protein expression levels were significantly increased（P<0. 01）. Compared with blank control group and NC group，the miR-138-5p expression level and apoptotic rate in miR-138-5p mimics group were significantly increased（P<0. 01），while the HIF-1α protein expression level and the proliferation activity of cells were significantly decreased （P<0. 01）. Compared with NC group，the luciferase activity of wild-type HIF-1α cells in miR-138-5p mimics group was markedly decreased（P<0. 01）. Compared with miR-138-5p mimics+Vector group，the proliferation activity of cells in miR-138-5p mimics+HIF-1α group was significantly increased（P< 0. 05），the apoptotic rate of cells was significantly decreased（P<0. 01），and the expression levels of HIF-1α，P-gp and MRP1 proteins in the cells were significantly increased（P<0. 05）. Conclusion：miRNA-138-5p inhibits the expression of HIF-1α and down-regulates the expression levels of resistance-related proteins P-gp and MRP1，thereby enhances the sensitivity of MCF-7/DDP cells to DDP.

Objective: To isolate and identify the proteins responsible for aggressive character of giant cell tumor of bone (GCTB) with proteomic techniques. Methods: 5 cases of aggressive GCTB and 4 cases of benign G CTB were selected as experimental and control groups, respectively, the differential protein expression were determined by isoelectric focusing / SDS acrylamide gel two-dimensional electrophoresis and Coomassie brilliant blue staining, then the differentially expressed proteins were identified by mass spectrographic analysis. Results: (506 ± 23) protei n stains were found in experimental group, (468 ± 28) protein stains were found in control group, there was significant difference between the two groups on two-dimensional electrophoresis (P<0.05), up-regulation of Annexin A1 and 14-3-3 protein E, down- regulation of Peroxiredoxin 1 were found in experimental group. Conclusion: The down-regulation of Peroxiredoxin 1 may be responsible for the aggression of GCTB, the up-regulation of Annexin A1 and 14-3-3 protein E may be due to the result of reaction of organism, these three proteins may be selected as tumor markers of the aggressive GCTB.

Objective: To explore the clinical efficacy of digital arteria axial flap augment volume in the treatment of Wassel type III- V congenti tal thumb duplication, and to provide the basis for its clinical treatment. Methods: The thumb augment volume surgery was performed in 14 children with Wassel type III-V congenital thumb dupliation. A narrow and long flap with vascular vessel and soft tissue was designed with microsurgical technique to augment the soft tissue of the reconstructed thumb and reconstruct the perimeter and appearances of the reconstructed thumb. The perimeters and appearences of the reconstructed thumb and healthy thumb were detected before and after surgery. Results: All the flaps were survived after the surgery. The average perimeter of the reconstructed thumb was 89% -104% relative to the healthy contralateral thumb based on the data detected from the tuberositas unguicularis, proximal interphalangeal joint, proximal phalanx midshaft, metacarpophalangeal joint and metacarpal midshaft. The appearance of reconstructed thumb was more similar to that of healthy thumb. Conclusion: The digital arteria axial vascular flaps which are used to augment the volume of reconstructed thumb of Wassel type IH - V congenital thumb duplication can be easily obtained and the reconstructed thumbs have better appearences. The surgery is an effective way to improve the thin and small appearences of reconstructed thumbs.

Objective: To detect the expressions of MMP-14 and TIMP-2 in colorectal carcinoma tissues, and investigate the relationship between MMP-14, TIMP-2 and clinicopathological features. Methods: 60 paraffin-embedded cancer specimens from operated patients with colorectal carcinoma were selected, and 20 normal mucous membranes of colon and rectum were selected as control. The expressions of MMP-14 and TIMP-2 were detected by S-P immunohistochemistry. The relationship between the expressions of MMP-14, TIMP-2 and clinicopathological parameters were analyzed. Results: The expression rates of MMP-14 and TIMP-2 were 86.7% and 80.0%, respectively, in colorectal carcinoma tissues and were significantly higher than those in control group (P<0.01). The expression of MMP-14 was closely correlated with the depth of tumor invasion, lymph node metastasis and Dukes stage. The expression rates of MMP-14 in serosa group, lymph node metastasis group and Dukes C+D stage were 89.5%, 94.1%, and 94.3%, respectively, and were significantly higher than those in non-serosa group, no lymph node metastasis group and Dukes A+B stage (68.2%, 73.1%, and 76.0%)(P<0.05). The expression of TIMP-2 was closely correlated with the depth of tumor invasion, lymph node metastasis and Dukes stage. The expression rates of TIMP-2 in serosa group, lymph node metastasis group and Dukes C + D stage were 55.3%, 64.7%, and 62.9%, respectively, and were significantly lower than those in non-serosa group, no lymph node metastasis group and Dukes A + B stage (81.8%, 88.5%, and 92.0%)(P<0.05). There was negative correlation between the expressions of MMP-14 and TIMP-2 in colorectal carcinoma tissues (r, =-1.0, P < 0.05). Conclusion: The high expression of MMP-14 in colorectal carcinoma tissues may contribute to tumor invasion and metastasis and TIMP-2 may play an inhibitory effect in the development of colorectal carcinoma. The imbanlance of MMP-14 and TIMP- 2 may be one of the mechanisms of tumor invasion and metastasis.

Objective: To explore the effect of 14-3-3ϵ protein on the localization of Cdc25B protein during the meiotic resumption of mouse oocytes, and to pay foundation for the further study on the molecular mechanism of 14-3-3ϵ protein in regulating the development of mouse oocytes. Methods: The Kunming genealology female mice aged 3 weeks were used to obtain the germinal vesicle (GV) -stage oocytes after superovulation. The GV-stage oocytes were divided into non-injection group, control siRNA injection group and 14-3-3s siRNA injection group. The pmax-FP-Red-HA-14-3-3ϵ expression vector was constructed. Indirect immunofluorescence was used to observe the colocalization of 14-3-3s protein and Cdc25B protein in the mouse oocytes; direct immunofluorescence was used to observe the subcellular localization of 14-3-3ϵ protein and Cdc25B protein in the mouse oocytes; 14-3-3ϵ siRNA was microinjected into the GV-stage oocytes; the morphology was observed under phase-contrast microscope; the germinal vesicle breakdown ( GVDB) rates of the mouse oocytes were calculated; the expression level of 14-3-3ϵ protein and the relative expression level of Cdc2-pTyrl5 protein were observed by Western blotting method; the matuation-promoting factor (MPF) activity in the oocytes was measured by autoradiography. Results: The indirect immunofluorescence and direct immunofluorescence results showed that the 14-3-3s protein and wild Cdc25B protein were co-localized in the cytoplasm; Cdc25B was translocated from the cytoplasm to the nucleus shortly before GVBD. When the Ser321 of Cdc25B protein turned into Ala, the expression level of 14-3-3ϵ protein was decreased. None of the oocytes in non-injection group and control siRNA injection group were able to undergo GVBD until at least 24 h after injection, there was no significant differences in the rate of GVBD between non-injection group and control siRNA injection group (P>0. 05) ; the GVBD rates of oocytes in 14-3-3ϵ siRNA injection group at 22 and 24 h after injection were significantly higher than those in non-injection group and control siRNA injection group (P<0. 01) ; the rate of oocytes progressed to metaphase II (MID in 14-3-3ϵ siRNA injection group at 24 h after injection was significantly higher than those in non-injection group and control siRNA injection group (P<0. 01). Conclusion: Ser321 might be involved in the process of regulating the subcellular localization of Cdc25B by 14-3-3ϵ protein in the meiotic resumption of mouse oocytes.

Objective: To explore the influence of hypoxia and transforming growth factor β (TGF-β) on the differentiation of umbilical cord mesenchymal stem cells (UCMSC) into the type II alverolar epithelial cells (AT II) and the regulatory effect of miR-145 in this process, and to illuminate the differentiation mechanism of UCMSC into AT II under the double stimulation of both hypoxia and TGF-β in the damaged lung tissue microenvironment. Methods: The UCMSC were isolated in vitro and co-cultured with human lung cancer cell line A549 to induce the differentiation of UCMSC into AT II. Cobaltous chloride (C0CI2) was used to mimic the hypoxia condition, and the induced cells were divided into normoxia group and hypoxia group. In hypoxia group, phase contrast microscope was used to observe the changes in cell morphology and flow cytometry was used to detect the percentage of AT II in the induced cells. qPCR and Western blotting methods were applied to test the expression levels of AT II specific genes, fibrosis-related genes and miR-145 in normoxia group and hypoxia group. In hypoxia group, the cells were divided into inh-145 group, inh-145 scramble group and control group, the expression levels of fibrosis-related genes in the cells in three groups were tested by qPCR and Western blotting methods. The pre-145 and pre-145 scramble were transfected into the 293T cells, and then the dual luciferase reporter gene system was used to check the relative luciferase unit (RLU) of TGF-βRII 37-UTR wild type and mutants. Results: In hypoxia group, the fibroblast-like UCMSC became flat and spindle, and finally showed a morphology of cobblestone-like epithelial cells 8 d later, and the percentage of the induced cells with high expression of AT II surface marker SpC was up to (94. 50 + 3. 37) %. The expression levels of specific genes of AT II (KGF, CK18, SpA, SpB and SpC) and miR-145 were obviously upregulated in hypoxia group compared with normoxia group (P<0.05). After stimulating the UCMSC-AT II differentiation with TGF-fil, the expression levels of fibrosis-related genes Col-I, TGF-βSR II and its downstream signaling factors p-Smad2 and p-Smad3 in hypoxia group were significantly down-regulated compared with normoxia group (P<0. 05). The expression levels of Col-I and TGF-βR II in inh-145 group were significantly raised compared with inh-145 scramble group and control group under hypoxia induction (P<0. 05). The RLU of TGF-βSRII 3'-UTR wild type was decreased after treated with pre-145 compared with TGF-βR II mutants (P< 0. 05). Conclusion: Hypoxia can promote the differentiation of UCMSC into the AT II and inhibit the TGF-β1-induced fibrosis. Its mechanism may be related to the inhibition of TGF-β1/TGFβR II signaling pathway through the down-regulation of TGF-βRII expression by hypoxia-induced miR-145.

Objective: To investigate the expression of microRNA-155 (miRNA-155) in the cerebral cortex tissue of the rats with cerebral ischemia-reperfusion injury, and to clarify the effect of miRNA-155 on cerebral ischemiareperfusion injury. Methods: A total of 48 healthy adult male SD rats were randomly divided into sham operation group (n=24) and cerebral ischemia reperfusion group (I/R group, n=24). The cerebral ischemia-reperfusion models in I/R group were established by Longa modified suture occlusion in the right middle cerebral artery of the rats. The rats in sham operation group only received blood vessel isolation. TTC staining was used to calculate the ischemic infarction volume of the rats in various groups. qRT-PCR method was used to detect the expression levels of miR-155 in cerebral cortex tissue of the rats in various groups. Results; Compared with sham operation group, the ischemic infarction volume of the rats in I/R group was significantly increased at 24, 48, and 72 h after reperfusion (P<0. 05); the ischemic infarction volumes of the rats in I/R group were decreased gradually with the prolongation of reperfusion time. Compared with sham operation group, the expression levels of miR-155 in cerebral cortex tissue of the rats in I/R group were significantly increased at 24, 48, and 72 h after reperfusion (P<0.05); the expression levels of miR-155 were gradually decreased with the prolongation of reperfusion time. Conclusion: The expression levels of miR-155 in cerebral cortex tissue of the rats with cerebral ischemia-reperfusion injury are high, and miR-155 may participate in the process of cerebral ischemia-reperfusion injury.

Objective: To construct prokaryotic expression vector harboring human papillomavirus-16 E7 (HPV-16) gene and observe the E7 protein expression. Methods: Synthetic E7 gene was amplified using polymerase chain reaction and subcloned into prokaryotic expressing vector pGEX-4T1. The recombinant plasmids, pGEX-4T1-HPV16-E7, were transferred into engineering bacteria Rosetta, and the fusion proteins E7-GST were analyzed on SDS-PAGE and purified by Ni 2+-NTA affinity chromatography under induction with IPTG. Results: The target gene fragment at a length of 300 bp was observed on the agarose gel electrophoresis pattern and PCR. SDS-PAGE analysis showed that the expressed recombinant protein, with a relative molecular mass of about 36 000, could be found. Conclusion: The recombinant protein, E7-GST, is effectively expressed in prokaryotic expression system.

Objective: To observe the clinical efficacy and safety of tacrolimus (TAC) combined with low dose of corticosteroids in the treatment of the children with refractory nephrotic syndrome (RNS), and to investigate the effect of diltiazem on the blood concentration of TAC. Methods: A total of 16 hospitalized children with refractory nephrotic syndrome (RNS) were selected. All patients were treated with TAC (0. 10-0. 15 mg · kg⁻¹ · d⁻¹) combined with low dose of corticosteroids (1. 0-2. 0 mg · kg⁻¹ · d⁻¹). The concentrations of TAC of the children during the treatment process were monitored. The 24 h urine protein quantitative (UP), plasma albumin (Alb), creatinine and other 6 indexes were recorded before treatment and 2, 4, 8, 12 and 24 weeks after treatment. Results: In 16 cases, 9 cases were completely relieved, 5 cases were partially relieved, and 2 cases were invalid; the total efficiency was 94%. The 24 h UP of 16 children at 24 weeks after treatment was significantly decreased compared with before treatment (Z= - 3. 516, P<0. 01); the plasma Alb level of the children was significantly increased compared with before treatment (Z= - 3.516, P<0. 01). The total cholesterol and triglyceride levels at 2 weeks after treatment were decreased significantly compared with before treatment (Z = - 2. 223, P<0. 05; Z= - 3. 464, P<0. 01). Eight patients had low blood concentration of TAC, and it was increased to an effective concentration after treated with diltiazem. In the course of treatment, creatinine, urea nitrogen and other indexes of the patients were fluctuated in a normal range. Conclusion: TAC combined with low dose of corticosteroids for treatment of RNS is safe and effective, and diltiazem can effectively increase the blood concentration of TAC.

Objective To explore the protective effects of 17-β estradiol(E2) on the apoptosis of cultured retinal ganglion cells(RGC-5 cells) induced by pressure, and to clarify its mechanism. Methods The RGC-5 cells were divided into control group, 80 mmHg pressure group and 80 mmHg pressure combined with E2 group. The cells at logarithmic growth phase were used in the experiment. The cell viabilities in various groups were determined by MTT assay. The apoptotic morphology of the cells in various groups was determined by Hoe33342 staining. The expression levels of bax, bcl-2, P53, iASPP, and cleaved-caspase-3 were determined by Western blotting method. Results The MTT assay showed that compared with control group, the viabilities of the RGC-5 cells in pressure group and pressure combined with E2 group were decreased (P < 0. 05); compared with pressure group, the viability of the RGC-5 cells in pressure combined with E2 was increased (P<0. 05). The results of cell staining showed that compared with pressure group, the chromatin condensation and formation of apoptotic bodies in the RGC-5 cells in pressure combined with E2 group were decreased. The Western blotting results showed that compared with control group, the expression levels of bax, P53 and cleaved-caspase-3 were increased(P<0. 05), the expression levels of bcl-2 and iASPP were decreased (P < 0. 05) in the RGC-5 cells in pressure group and pressure combined with E2 group; compared with pressure group, the expression levels of bax, P53 and cleaved- caspase-3 were decreased(P<0. 05), and the expression levels of bcl-2 and iASPP were increased(P<0. 05) in the RGC-5 cells in pressure combined with E2 group. Conclusion E2 has the protective effects on the RGC-5 cells and could decrease the apoptosis of RGC-5 cells under pressure loading condition.

Objective: To observe and compare the photochemotherapeutic effects of BCPD-17 (hematoporphyrin derivative) and PSD-007 on osteosarcoma LM-8 transplanted in mice and to explore a novel photosenitizer for photodynamic therapy of osteosarcoma Methods The tumor model was made by subcutaneous injection of osteosarcoma LM-8 on C3H mice. When the tumors reached about 6-8 mm in diameter, the tested mice were randomly divided into control group, PSD-007 group (5 mg·kg-1), BCPD-17 groups (5 and 10 mg·kg-1). After received intravenous injection of 5 or 10 mg·kg-1 of PSD-007 and BCPD-17, respectively, the tumors were irradiated vertically with 630 nm or 670 nm laser (power density 200 mW·cm-2, energy density 240 J·cm-2) for 20 min. The size of osteosarcoma was measured 7 d after treatment. And all the tumors were extirpated, weighted and taken for pathological examination. The inhibitory rate of tumor was calculated. Results The results of pathohistological observation showed that in control group the size of mouse osteosarcoma was big and hyperchromatic with obvious nuclear atypia; in PDT treatment group, the turmor cells presented apoptosis, vacuolar degeneration and karyopycnosis, especially in BCPD-17 group. Compared with control group, the volumes and the weights of the tumors in PDT treatment groups were obviously decreased, there were significant differences (P<0.01). Compared with PSD-007 group, the inhibitory rates of tumor in BCPD-17 groups irradiated by the laser with wavelength 670 nm were increased, there were significant differences(P<0. 01). Conclusion Both of the photosenitizers BCPD-17 and PSD-007 could obviously inhibit the growth of the tumors. Photosenitizer BCPD-17 has more anti-tumor effect.

Objective: To investigate the relationships between the levels of serum interleukin-17 (IL-17), interleukin-33 (IL-33) and the concentration of fractional exhaled nitric oxide (FeNO) in the patients with chronic cough, and to further evaluate their effects in the pathogenesis of chronic cough. Methods: A total of 160 patients diagnosed with chronic cough for more than 8 weeks were chosen and used as chronic cough group. At the same time, 60 healthy controls received physical examination were selected as healthy control group. The levels of serum IL-17 and IL-33 of the subjects were examined and pulmonary function test, FeNO concentration test and comprehensive allergen test were performed. All data were analyzed by GraphPad Prism statistical software. The levels of serum IL-17 and IL-33 were compared between chronic cough group and healthy control groups. The correlations between the levels of serum IL-17 or IL-33 and lung function or FeNO in the patients with chronic cough were further analyzed. Results: The serum IL-17 and IL-33 levels of the patients in chronic cough groups were higher than those in healthy control group (P<0. 05). The serum IL-17 and IL-33 levels in the patients with chronic cough showed a significant negative correlations with the percentage of the first second force expiratory volume to the estimated value (FEV1%) of the patients (r=-0. 624 5, r=- 0. 672 2), and the level of serum IL-33 in the patients with chronic cough was positively correlated with the concentration of FeNO; the higher the concentration of FeNO, the higher the level of serum IL-33 (rs = 0. 758, P<0. 05). The FeNO concentration geometric mean of the patients with the serum total IgE<C100 IU · mL⁻¹ (33 ppb) was significantly lower than those with the serum total IgE>200 IU · ml⁻¹ (78 ppb) and IgE 100-200 IU · mL⁻¹ (69 ppb) among the chronic cough patients (P<0. 01), but there was no significant difference between the later two groups (P = 0. 082 4). Conclusion: The serum IL-17 and IL-33 may play an important role in the pathogenesis of chronic cough as proinflammatory factors. The levels of serum IL-17 or IL-33 have negative correlations with the pulmonary ventilation function in the patients with chronic cough. The increasing of serum IL-33 level may predict the formation of eosinophilic airway inflammation and reflect the severity of eosinophilic airway inflammation.

Objective: To investigate the feasibility of detection of serum pepsinogen (PG), gastrin-17 (G-17) and Helicobacter pylori antibody (anti-HP) in the gastric cancer screening, and to elucidate its clinical value. Methods: A total of 208 patients with early gastric cancer were selected as observation group; at the same time, 208 healthy examinees were regarded as blank control group. The levels of PG I, PGJJ and G-17 of the subjects in two groups were detected by enzyme-linked immunosorbent assay and latex-enhanced immunoturbidimetry ; the positive rates of anti-HP were detected by l-, C urea breath test; the serum levels of PG I, PG JJ and PG I / PG TI ratio of the anti-HP positive subjects (anti-HP positive group) and anti-HP negative subjects (anti-HP negative group) were compared and analyzed. Results: Compared with blank control group, the serum PG I / PGJJ ratio and PG I level of the subjects in observation group were significantly decreased (P<0. 01), and the levels of PG TI, G-17 and the positive rate of anti-HP were increased (P<C0. 01). Compared with anti-HP negative group, the serum PG I / PG II ratio and PG I level of the subjects in anti-HP positive group were significantly decreased (P<0. 01). The levels of PG II and G-17 were significantly increased (P<0. 01). Conclusion: The detection of positive rate of anti-HP combined with the ratio of PG I / PG II has important clinical significance in gastric cancer screening. The levels of G-17 and PG II in the epithelial neoplasia lesion tissue can be used as indicators of gastric precancerous lesions.

Objective: To explore the expression level of serum interleukin-17 (IL-17) in the patients with psoriasis vulgaris and the changes of interleukin-6 (IL-6) and interleukin-23 (IL-23) levels secrered by the HaCaT cells after stimulated with IL-17, and to clarify its clinical significance and the intervention effect of shikonin. Methods: Twenty-five normal controls, 29 patients with psoriasis vulgaris and different groups of HaCaT cells (blank control group, IL-17-24 h group, IL-17-36 h group, IL-17-48 h group, shikonin + IL-17 group, CsA + IL-17 group and IL-17 group) were used as the subjects. The levels of serum IL-17 in the patients with psoriasis vulgaris and the levels of IL-6 and IL-23 in supernatant of HaCaT cells in various groups were measured by double antibody sandwich enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction RT-PCR was used to detect the expression levels of IL-6 and IL-23 pl9 mRNA in HaCaT cells in various groups. At the same time, Cell Counting Kit-8 (CCK-8) method was used to detect the viabilities of HaCaT cells in various groups. Results: The level of serum IL-17 of the patients in psoriasis vulgaris group was increased compared with nomral control group, especially in the severe skin lesion group (P<0. 05). The levels of IL-6 and IL-23 in HaCaT cells and supernatant and their mRNA expression levels in IL-17-24 h, IL-17-36 h and IL-17-48 h groups were significantly higher than those in blank control group (P<0.01). The expression levels of IL-6 and IL-23 in HaCaT cells and supernatant and their mRNA expression levels in shikonin+IL-17 and CsA-|-IL-17 groups were lower than those in IL-17 groups (P<0.05). No statistical differences were found in the cell viabilities between each drug treatment group and blank control group (P>0. 05). Conclusion: The expression level of IL-17 in the patients with psoriasis vulgaris is significantly increased, especially in the patients with severe psoriasis vulgaris. IL-17 can promote the secretion of IL-6 and IL-23 in HaCaT cells in a time-dependent manner. Shikonin can inhibit the proinflammatory effect of IL-17.

Objective: To explore the effect of recombinant human interleukin-17AI (IL-17A) on the growth of colon cancer cells, and to investigate the related mechanism. Methods: The colon cancer SW480 cells were divided into control group and experimental group. The SW480 cells in control group were untreated and the SW480 cells in experimental group were added with 50 fig • L _1 IL-17A. The proliferation abilities of SW480 cells in two groups were detected by CKK-8 method. The levels of IL-17A in the SW480 cells in two groups were detected by ELISA, and the expression levels of signal transducers and activators of transcription 3 (STAT3) and p-signal transducers and activators of transcriptions 3 (p-STAT3) were examined by Western blotting methed. Results: Compared with control group, the proliferation ability of the SW480 cells in experimental group was increased (P < 0 . 05). The level of IL-17A in the SW480 cells in experimental group was significantly higher than that in control group (P< 0. 01). Compared with control group, the expression levels of STAT3 and p-STAT3 proteins in the SW480 cells in experimental group were significantly higher than those in control group (P < 0 . 01). Conclusion: Recombinant protein IL-17A can stimulate the growth of colon cancer SW480 cells, which may be related to the activation of STAT3 signaling pathway.

Objective: To explore the expressions of interleukin-17E (IL-17E), interleukin-17F (IL-17F) and their receptors interleukin -17RB (IL-17RB), interleukin -17RC (IL-17RC) in enteritis, intestinal polyps and colorectal carcinoma tissues, and to analyze their relationships with the malignancy of colorectal carcinoma. Methods: Immunohistochemistry staining was used to detect the expressions of IL-17E, IL-17RB, IL-17F, and IL-17RC in the tissues of enteritis (n=15), intestinal polyps (n=5) and colorectal carcinoma (n=30). The relationships between IL-17E and IL-17RB, IL-17F and IL-17RC, and the relationships between IL-17E, IL-17F and malignancy of colorectal carcinoma were analyzed. Results: IL-17E, IL-17F, IL-17RB and IL-17RC mainly expressed in glandular epithelial cells, mononuclear cells and vascular endothelial cells as well as partial carcinoma cells. The positive expression rates of IL-17E in enteritis and intestinal polyps tissues were higher than that in colorectal carcinoma tissue (P<0. 05). Compared with enteritis tissue, the positive expression rates of IL-17F in intestinal polyps and colorectal carcinoma tissues were increased (P < 0. 05), which was increased with the increasing of malignancy of coloretal carcinoma. Compared with colorectal carcinoma tissue, the positive expression rates of IL-17RB in enteritis and intestinal polyps tissues were increased (P<C0. 05). Compared with enteritis and intestinal polyps tissues, the positive expression rate of IL-17RC in colorectal carcinoma tissue was increased (P< 0. 05). There was positive correlation between the positive expression rate of IL-17F and the positive expression rate of IL-17RC in colorectal carcinoma tissue (r=0. 667, P=0. 001). Conclusion: IL-17F, IL-17RC, IL-17E, and IL-17RB signals may take part in the incidence and progression of colorectal carcinoma.

Objective: To study the expression of 17β-hydroxysteroid dehydrogenase (17β-HSD) type 2 in breast cancer and adjacent non-malignant tissue, and the correlations between the results and clinical factors. Methods: The expressions of the oxidative enzyme 17β-HSD type 2 in 76 specimens of female human breast cancer and adjacent non-malignant tissues were determined by immunohistochemistry. The relationships between the clinical parameters including patient's age, ER, PR, P185, the stage of breast cancer, tumor size and the percentage of involved lymph nodes were analyzed. Results: 17β-HSD type 2 was found in the cytoplasm of tumor cells in 5.3% of cases. In adjacent non-malignant tissues, 82.9% of cases were positive and the enzyme was detected in the cytoplasm of epithelial cells of the acini and ducts. There was a significant difference between the malignant tissue and the adjacent non-malignant tissue in the percentage of positive cells (χ2=92.908, P<0.001). In adjacent non-malignant tissues, the expression of 17β-HSD type 2 was negatively correlated with the tumor size (r=-0.341, P<0.05), the stage of the breast cancer (r=-0.706, P<0.01) and was positively correlated with the patient's age (r=0.677, P<0.01). For the other clinical parameters including ER, PR, P185 and the percentage of involved lymph nodes, there was no significant correlation. No significant correlation was found between the expression of 17β-HSD type 2 and all clinical parameters in breast cancer tissue. Conclusion: 17β-HSD type 2 can regulate the local estrogen level. It may play a significant role in the development and/or progression of breast cancer.

Objective: To explore the molecular mechanism of brain tissue injury induced by endotoxin and observe the effects of Dex on TLR4, IκBα and IL-18 mRNA expressions of cortex in the endotoxic shock rats. Methods: Eighteen Wistar rats were randomly divided into LPS, LPS+DEX and control group (n=6). Except control group, all the rats were administered with LPS (4 mg · kg-1) by intravenous injection. The rats in control group were given glucose solution at same volume. The TLR4, IκBα and IL-18 mRNA expressions were assayed 4 h after intravenous injection. Results: The mean arterial pressure (MAP) and the survival rate in LPS+DEX group were significantly higher than those in the LPS group (P<0.01). TLR4 and IL-18 mRNA expressions in LPS+DEX groups were obviously lower than those in LPS group (P<0.05). The IκBα mRNA expression in LPS +DEX group was significantly higher than that in LPS group (P<0.05). Conclusion: Dex may down-regulate TLR4 expression and up-regulate IκBα expression in the brain tissue, and has protective effect on central nervous system.

Objective: To evaluate the nested-reverse-transcription-polymerase chain reaction (nested-RT-PCR) for detection of cytokeratin 18 mRNA (CK18 mRNA) in diagnosis of micrometastasis in esophageal cancer and its clinical significance. Methods: Nested-RT-PCR was used to detect the expressions of CK18 mRNA in the rib marrow of 50 esophageal carcinoma patients without distal metastasis (experimental group), 10 esophageal carcinoma patients with distal metastasis, 10 benign esophageal disease patients. Results: CK18 mRNA was not detected in benign esophageal disease patients. In the rib marrow of 50 esophageal carcinoma patients without distal metastasis and 10 esophageal carcinoma patients with distal metastasis, the positive expression rates of CK18 mRNA were 42.0% (21/50) and 100% (10/10), respectively. The positive rate in stage IV of esophagenal cancer (100%) was significantly higher than those in stage II a (40%), stage II b (41.7%) and stage III (45.8%) (P<0.05). But there were no significant differences between stage II a, stage II b and stage III (P>0.05 ). The positive rates in pathological types I, II, and III were 41.7%, 41.7%, and 42.9%, there were no significant differences between various pathological types (P>0.05). Conclusion: In the early stage of esophageal cancer, the cancer cells have already metastasized. It is helpful to use Nested-RT-PCR to detect the expression of CK18 mRNA in the rib marrow in early diagnosis of tumor micrometastasis.

Objective To investigate the effects of different doses and infusion methods of recombinant mouse interleukin-18 (rmIL-18) on the survival time and tumor diameter of the mice with hepatocellular carcinoma, and to elucidate the rational application of rmIL-18 in vivo. Methods: A total of 60 Babl/C mice were randomly divided into 5 fig rmIL-18 intraperitoneal injection group, 0. 5 fxg rmIL-18 intraperitoneal injection group, 0. 5 μg rmIL-18 tumor injection group, cytotoxic T lymphocyte (CTL) intraperitoneal injection group, CTL tumor injection group and saline control group; there were 10 mice in each group. From the 10th day of inoculation, the mice in different rmIL-18 groups were injected with the corresponding doses and methods. The mice in different CTL groups were injected with tumor-specific CTL (1 X 10s /mouse) by intraperitoneal and intratumoral injection. The mice in saline control group were injected with an equal volume (100 jiL) of saline, the injections were performed 10 times. The diameters of mice were measured weekly and the survival time was recorded. Results: Compared with 5 pg rmIL-18 intraperitoneal injection group, 0. 5 jig rmIL-18 intraperitoneal injection group and saline control group, the tumor growth rate of the mice in 0. 5 fig rmIL-18 tumor injection group was decreased (P<0. 01) and the survival rate of the mice was increased (P<0.01); compared with 0.5 jig rmIL-18 intraperitoneal injection group, the tumor growth rate and the survival rate of the mice in CTL intraperitoneal injection group were decreased (P<0. 01)} compared with 0. 5 /ig rmIL-18 tumor injection group, the tumor growth rate and the survival rate of the mice in CTL tumor injection group were decreased (P<0. 01). Conclusion: The best way for rmlL-18 anti-Tumor effect is tumor injection and the effect has a dose-dependent manner.

Objective: To construct an eukaryotic expression plasmid containing the chimeric gene coding for the hemagglutinin-neuraminidase (HN) of newcastle disease virus (NDV) and human IL-18. Methods: The enzymolysis and ligation were used to joint HN cDNA to IL-18 cDNA of pVAX1 IL-18 plasmid and at the same time the terminator of IL-18 cDNA was replaced, an eukaryotic expression vector of chimeric gene for HN and IL-18 was constructed. The recombinant plasmid was transfected into HeLa cells with liposome and the expression products were examined by hemagglutination test and Western blotting. Results: The expression plasmid of pVAX1 IL-18 HN was contructed successfully confirmed by enzyme digestion identification. The expressions of IL-18 and HN chimeric genes were confirmed, and the expressed HN protein had higher hemagglutinative titer. Conclusion: The constructed eukaryotic expression plasmid can express in vitro, and the expression products of the chimeric gene have good reactivity and specificity.

Objective: To compare the efficiency of IL-18 and IL-12 in inducing the tumor specific CTL in vitro, and test the cytotoxicities of tumor specific CTL induced in the same culture system with IL-18 or IL-12. Methods: The NK cells, T cells and dendritic cells were separated from fresh PBMNCs by using the Stem Sep™ immunomagnetic beads. Cell phenotypes of the purified cell populations were identified with FCM technique. The cell co-culture system was established as follows: the cell preparations in which definite cell subset had been deleted was co-cultivated with mitotic-inactivated tumor cells in the presence of rhIL-18 and/or rhIL-12. Cytotoxicities of the various effector cell preparations to a series of tumor cell lines were examined by the isotope releasing assay. Results: In the cell co-culture system in vitro, rhIL-18 or rhIL-12 alone and dose-dependently induced tumor specific cytotoxicities. The level of cytolytic activity of the tumor specific CTL induced by IL-12 was substantially inferior to that induced by IL-18 (P<0.01). Conclusion: These results suggest that IL-12 does not induce tumor-specific CTL as efficiently as IL-18 does in this co-culture system under these conditions, but the synergistic effect could be observed during administration of IL-18 and rIL-12.