Objective To investigate the expression of NKG2D in peripheral blood of patients with cervical cancer,CIN,hysteromyoma and health person,and the expression of the human MHC class I chain-related gene A(MICA)on the correspondent tumor tissues.To discuss the anti-cervical cancer mechanism of NKG2D-MICA and immune escaping of cancer.Methods Flow cytometry analysis was used to detect the expression of NKG2D in the peripheral blood of patients with cervical cancer,CIN,hysteromyoma and health person.The expressions of MICA in part of the correspondent tissues were examined by means of reverse transcription-polymerase chain relation(RT-PCR).Results The expression of NKG2D in the patients with cervical cancer,CIN,hysteromyoma and health person were(76.87?9.39)%、(81.84?7.94)%、(86.77?8.68)%、(93.968?4.9)%,respectively.Compared with the normal group,the NKG2D expression in the particular disease group was of statistical significance,however,it is not statistically significant in the comparison in the particular disease group.The rate of MICA mRNA expression in cervical cancer was significantly higher than that in hysteromyoma and health tissues,and its difference is of statistical significance.But it is not statistically significant for the normal group to compare with the other group.Conclusion The activity of NK cell and the anti-cancer cellular immunity level reduce in patients with cervical cancer.The decrease of the receptor NKG2D is a reason for the descend of the activity of NK cells.MICA mRNA expression increases in the cervical cancer,and it has the tendency of up-regulation with the progress of pathological changes.It is relative to malignant transformation from cervical squamous intraepithelial lesion to cervical cancer;the immune-escape of cervical cancer probably is relative to the down-regulation of NKG2D and the up-regulation of its ligand MICA.

Objective To investigate the variance and significance of Th1/Th2 type cytokines in plasma of ovarian cancer patients. Methods The levels of IL-2, IFN-?, TNF-?, IL-4, IL-6, IL-10 in 34 plasma samples of ovarian cancer patients and 14 normal women were detected by cytometric bead array. Results The levels of IL-2, IFN-?, TNF-? in ovarian cancer patients were obviously lower than those in normal womens but the levels of IL-4, IL-6, IL-10 were obviously higher. Furthermore, the variance changed along with clinic stage. Conclusion It is suggested that the imbalance of Th1/Th2 type cytokines in ovarian cancer patients may provide clinical index for the evaluation of progression and prognosis.

BACKGROUND: Placenta mesenchymal stem cells have become hot spots in stem cells study in recent years because of its advantages. OBJECTIVE:To analyze the biological characteristics of amniotic mesenchymal stem cells and amnion epithelial cells, and to explore the feasibility of amniotic mesenchymal stem cells and amnion epithelial cells applied as seed cells in three-dimensional liquid culture to construct the tissue-engineered skin. METHODS:The amniotic mesenchymal stem cells and amnion epithelial cells were obtained by using multi-step digestion with trypsin and col agenase;then the flow cytometry, reverse transcription-PCR and immunofluorescent staining techniques were adopted to identify the surface molecular markers, stem cellcharacteristics and keratinocytes similarity respectively. Based on these data, amniotic mesenchymal stem cells and amnion epithelial cells used as seed cells together with rat type Ⅰ col agen matrix were adopted for three-dimensional liquid culture. RESULTS AND CONCLUSION:Flow cytometry test showed that amniotic mesenchymal stem cells and amnion epithelial cells cultured in vitro could highly express CD90, CD73 and CD105, and could not express the hemopoietic stem cellmarker of CD34 and MHC-class Ⅱ molecular HLA-DR. Reverse transcription-PCR results detected that amniotic mesenchymal stem cells could express stem cellcharacteristic genes CMCY and NANOG;amnion epithelial cells could express stem cellcharacteristic genes CMCY and KLF4, showing that both amniotic mesenchymal stem cells and amnion epithelial cells have stem cellproperties. Reverse transcription-PCR results showed that amniotic mesenchymal stem cells could express keratinocytes characteristic genes K19,β1-integrin and K8;amnion epithelial cells could express K19,β1-integrin, K5 and K8. Immunofluorescence staining results showed amnion epithelial cells could express keratinocytes proliferation related protein K14, which revealed that there was certain similarity in the mRNA expression between keratinocytes and amnion epithelial cells, and indicating that it has the potential to differentiate into keratinocytes. Tissue-engineered skin was successful y constructed by using amniotic mesenchymal stem cells and amnion epithelial cells, hematoxylin-eosin staining section showed that it has certain skin structure, and amnion epithelial cells had a preliminary differentiation. Al these prove that it is feasible to construct human skin tissues with amniotic mesenchymal stem cells and amniotic epithelial cells through the three-dimensional culture.

Objective:To construct a RNAi lentiviral vector targeting rat CD80 gene and detect its effect of gene silencing in NRK and IEC6 cells.Methods:The effective sequence of siRNA targeting rat CD80 gene was confirmed in our previous work.Oligo-DNA fragment containing short hairpin frame was synthesized and reannealed,and then cloned into pGCSIL-GFP lentiviral expression vector.PCR and sequencing analysis were made for verifying the positive clones.The virus packaging plasmids were transfected into 293T cells to harvest shRNA lentivirus.After infection in NRK and IEC6 cells,Real-time PCR was performed to determine the expressing level of CD80.Results:PCR and sequencing revealed that shRNA plasmids was correctly constructed.Virus with a titer of 4×10~8 TU/ml was successfully packaged.CD80 expression in NRK and IEC6 cells could be knockdown by virus infection as characterized by 66.9% and 63.5% decrease of CD80 mRNA in NRK and IEC6 cells respectively,compared with negative control lentivirus.Conclusion:The recombinant lentiviral shRNA expressing vector targeting rat CD80 gene has been successfully constructed and packaged.CD80 mRNA could be down-regulated availably in NRK and IEC6 cells.

BACKGROUND: It has been reported that human adipose-derived mesenchymal stem cells can be induced to differentiate into hepatocyte-like cells with the function of albumin synthesis and urea secretion in vitro.OBJECTIVE: To investigate the potential of adipose-derived mesenchymal stem cells differentiating into hepatocyte-like cells in vitro.METHODS: Adipose-derived mesenchymal stem cells were isolated from the subcutaneous fat of hepatitis B virus infection patients by collagenase digestion and adherent method. Adipose-derived mesenchymal stem cells were induced by three-phase induction method and observed morphologically. The expression levels of alpha-fetoprotein, albumin and cytokeratin 18 were detected by immunohistochemical staining and glycogen synthesis function was detected by glycogen staining method.RESULTS AND CONCLUSION: Most of adipose-derived mesenchymal stem cells induced by three-phase induction method were differentiated into hepatocyte-like cells with polygonal morphology. Immunohistochemistry staining results showed that hepatocyte-like cells expressed alpha-fetoprotein, albumin and cytokeratin 18, and the expression levels of albumin and cytokeratin 18 increased with the culture time. The induced cells had the function of glycogen synthesis and were positive for periodic acid Schiff staining. These results showed that the subcutaneous adipose-derived mesenchymal stem cells could be induced into functional hepatocyte-like cells in hepatitis B virus infection patients.

Objective:To investigate the effect of pidotimod in reducing pulmonary infection in patients with lung cancer undergoing chemotherapy.Methods:One hundred and twenty patients with lung cancer in the Fifth People′s Hospital of Dalian City from July 2017 to July 2018 were selected. The patients were divided into control group and pidotimod group by random digits table method with 60 cases each. The patients were treated with standard two drugs chemotherapy containing platinum drug according to the pathological type, and the patients in pidotimod group were combined with pidotimod. The number of pulmonary infections during chemotherapy, number of completed scheduled chemotherapy and adverse reaction were observed. The correlation between pulmonary infection and pidotimod was analyzed by multivariate orderly Logistic regression.Results:The incidence of pulmonary infection in pidotimod group was significantly lower than that in control group: 18.33% (11/60) vs. 40.00% (24/60), and there was statistical difference ( χ2 = 6.845, P<0.01). The rate of completed scheduled chemotherapy in pidotimod group was significantly higher than that in control group: 55.00% (33/60) vs. 36.67% (22/60), and there was statistical difference ( χ2 = 4.062, P<0.05). Multivariate orderly Logistic regression analysis result showed that pidotimod could reduce the risk of pulmonary infection ( OR = 0.210, 95% CI 0.072 to 0.606, P = 0.004), and help to complete the scheduled chemotherapy ( OR = 2.323, 95% CI 1.080 to 5.003, P = 0.031). In pidotimod group, no obvious adverse reaction related to pidotimod application was detected, and chemotherapy was not affected. Conclusions:Application of pidotimod can reduce the chance of pulmonary infection in patients with lung cancer undergoing chemotherapy and help patients complete scheduled chemotherapy.

Objective To investigate the relationship between the marrow edema and general clinical index,quadriceps muscle area,and the meniscus grade of knee osteoarthritis(OA).Methods 72 patients were collected with knee OA in our hospital, underwent X-ray and routine MRI examination of knee, and the same X-ray and MRI were reviewed at 12 months later in different time point.The K-L grading, bone marrow edema score, meniscal grading and VAS score of each knee joint were evaluated.The t-test and Rank-sum test were used to compare the two groups of general data, Spearman was used to perform bivariate correlation analysis.Results The age and VAS score of bone marrow edema group at the initial follow-up were significantly lower than those without edema group(P<0.05),the degree of marrow edema was moderately correlated with age and VAS score, and was not significantly correlated with other indexes.12 months later, BMI and the area of quadriceps femoris in the group with marrow edema were different from those in the group without bone marrow edema(P<0.05), the degree of marrow edema was moderately correlated with age, quadriceps area and VAS.Conclusion The range of marrow edema was related to age, quadriceps area, BMI index and VAS score in MRI.The MRI measurement could reflect the progression of knee OA more than that of X-ray.It also revealed some factors related to the progression of knee OA.

Objective To apply intervention of integrated pharmaceutical care (IPC) for asthma-COPD overlap syn-drome patients,so as to reduce the side effects of drugs,enhance medication compliance,promote reasonable drug application,cut down the medical expenses in ACOS patients. Methods A total of 60 ACOS patients were randomly divided into IPC group (group A) (n=34) and contrast group(group B) (n=26).The patients in group A were given IPC measurements such as noso-comial guidance,classroom teaching,regular follow-up,life coaching and psychological advice.While the patients in group B were not given any intervention measures. Results In group A,patients' awareness rate of action and side-effects of drugs were ob-viously increased;Knowledges of inhalation preparation were greatly improved;the ratio of ADRs was significantly reduced;The FEV1 and the value added of FEV1 was dramatically improved.Furthermore,the differences showed statistical significance as com-pared with group B(P<0.05).Total medical costs and anti-bacterial drug costs per year were significantly lower in group A than group B. Conclusion IPC is beneficial to enhance drug compliance,promote reasonable drug application and bring down the medical expenses in ACOS patients.

Objective:To observe the efficacy of apatinib combined with first-line chemotherapy and maintenance therapy of only apatinib in patients with extensive small-cell lung cancer.Methods:The clinical data of 56 newly diagnosed patients with extensive small-cell lung cancer in the Fifth People′s Hospital of Dalian City from January 2018 to June 2019 were retrospectively analyzed. Among them, 27 patients (experimental group) were treated with first-line chemotherapy combined with apatinib, and 29 patients (control group) were treated with first-line chemotherapy alone. In experimental group, the expression levels of vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptor (VEGFR)-2 1 day before chemotherapy and 1 day after chemotherapy were detected by enzyme linked immunosorbent assay method. Response evaluation criteria in solid tumor (RECIST) was used to evaluate the efficacy. The occurrence of adverse reaction was recorded. The patients were followed up for 12 to 24 months, and progression-free survival and 1-year survival were recorded.Results:The objective response rate, median progression-free survival time and 1-year survival rate in experimental group were significantly higher than those in control group: 81.5% (22/27) vs. 55.2% (16/29), 10.5 months vs. 8.5 months and 81.5% (22/27) vs. 55.2% (16/29), and there were statistical differences ( P<0.05); there was no statistical difference in disease control rate between 2 groups ( P>0.05). In experimental group, the patients with complete response and partial response after chemotherapy were classified as effective subgroup (22 cases), and the patients with stationary disease and progressive disease were classified as ineffective subgroup (5 cases). There were no statistical difference in VEGF and VEGFR-2 before chemotherapy between 2 subgroups ( P>0.05). The VEGF and VEGFR-2 in effective subgroup were significantly lower than those in ineffective subgroup: (275.34 ± 16.15) ng/L vs. (330.24 ± 23.21) ng/L and (89.35 ± 4.34) ng/L vs. (112.34 ± 5.45) ng/L, and there were statistical differences ( P<0.01). There were no uncontrollable adverse reactions in 2 groups, and there was no statistical difference in the incidence of adverse reactions between 2 groups ( P>0.05). Conclusions:Application of apatinib in first-line therapy and maintenance therapy for patients with extensive small-cell lung cancer can improve clinical efficacy and survival benefit with controllable adverse reactions.

Objective:To observe and analyze depression-like behavioral performances of mouse models of vitiligo.Methods:Fifteen female C57BL/6 mice aged about 9 weeks were modeled for vitiligo. Whether the mouse models of vitiligo were successfully constructed or not was determined by macroscopy and full-thickness epidermal immunofluorescence staining of mouse tail tissues on day 23 after the start of the experiment; on day 8 (pre-modeling stage) and day 21 (early modeling stage), the elevated plus maze test and the open field test were used to evaluate the behavioral performances of the mice, including the number of entry into the open arms, percentages of time spent in the open arms, percentages of time spent in the central area and total distance traveled, aiming to assess whether depression-like behaviors were exhibited in the mouse models of vitiligo. To further clarify the degree of the impact of vitiligo modeling on the depression-like state in mice, 20 female C57BL/6 mice were equally divided into 2 groups: vitiligo modeling group and vitiligo modeling + chronic restraint stress group; the mice in the vitiligo modeling + chronic restraint stress group were subjected to chronic restraint stress on day 9, that is, these mice were placed in centrifuge tubes and restrained for about 6 hours every day for 28 consecutive days; on days 7, 22, 29 and 38 after the start of vitiligo modeling, the above-mentioned behavioral indicators were determined by the elevated plus maze test and open field test in the 2 groups. Repeated measurement data in a single group were compared before and after treatment by using paired t-test, and repeated measurement data at multiple time points were compared by using two-way repeated measures analysis of variance. Results:By macroscopy, the mice gradually developed well-defined white patches on the tail skin during vitiligo modeling, which were similar to the clinical manifestations of vitiligo patients; on day 23, full-thickness epidermal immunofluorescence staining of the mouse tail tissues was conducted and showed obvious infiltration of CD8 + T cells and a decrease in the number of Melan-A-positive epidermal melanocytes under a laser confocal microscope, which were consistent with typical pathological characteristics of vitiligo; based on the macroscopic results and immunofluorescence findings, a total of 12 mouse models of vitiligo were successfully constructed on day 23. The elevated plus maze test showed that the number of entry into the open arms and the percentages of time spent in the open arms were significantly lower in the 12 mouse models of vitiligo on day 21 (2.33 ± 1.78 times, 5.01% ± 5.27%, respectively) than in those on day 8 (10.75 ± 2.30 times, 29.20% ± 12.48%, t = 9.63, 6.36, respectively, both P < 0.001) ; the open field test showed that the percentages of time spent in the central area and total distance traveled were also significantly lower in the mouse models on day 21 (2.31% ± 1.53%, 2 518.31 ± 528.38 cm, respectively) than in those on day 8 (4.47% ± 2.65%, 3 533.45 ± 465.47 cm, t = 2.40, 5.47, P = 0.036, < 0.001, respectively). In the chronic restraint stress test, a total of 14 mouse models of vitiligo were successfully constructed on day 23, including 5 in the vitiligo modeling group and 9 in the vitiligo modeling + chronic restraint stress group. There were no significant differences in the number of entry into the open arms, percentages of time spent in the open arms, percentages of time spent in the central area, and total distance traveled between the vitiligo modeling group and the vitiligo modeling + chronic restraint stress group on days 7, 22, 29, and 38 ( F = 0.21, 0.20, 0.46, 2.35, P = 0.889, 0.893, 0.719, 0.134, respectively) ; moreover, all the above indicators significantly changed over time (all P < 0.001), except for the total distance traveled ( P = 0.422) . Conclusion:The mouse models of vitiligo developed depression-like behavior at the early modeling stage, and the degree of depression could not be further deepened by chronic restraint stress on the basis of vitiligo modeling.