Objective:To explore the effects of neutrophil extracellular traps (NET) on the proliferation and migration of breast cancer cells and the related mechanisms.Methods:The peripheral blood neutrophils were separated using density gradient centrifugation, 100 nmol/L phorbol 12-myristate 13-acetate (PMA) was used to stimulate the formation of NET. Breast cancer cell line MDA-MB-231 was selected and treated with NET (NET group) or deoxyribonucleaseⅠ (DNase Ⅰ) digested NET (NET+DNase Ⅰ group),normal cultured MDA-MB-231 cells were used as the control group. In MDA-MB-231 cells of each group; cell proliferation ability was detected using CCK-8 assay; cell migration was detected by cell scratch assay and Transwell assay; real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expressions of frizzled homolog 10 (FZD10), cyclin D1, E-cadherin and Vimentin immunofluorescence assay was performed to detect the formation of NET and the expressions of E-cadherin and Vimentin; Western blotting was used to detected the expression of FZD10-Wnt-β-catenin signaling pathway-related proteins.Results:Neutrophils were stimulated with PMA for 4 hours, and immunofluorescence assay results showed that NET exhibited fibrous reticular structure with high expression of citrullinated histone (citH3) and myeloperoxidase (MPO). The CCK-8 assay results showed that the absorbance values of cells in the control group, NET group and NET+DNase Ⅰ group were 1.25±0.06, 2.14±0.15 and 1.47±0.11, respectively, and the difference was statistically significant ( F = 80.60, P < 0.001). The results of scratch assay displayed that the percentage of wound closure in the control group, NET group and NET+DNase Ⅰ group were (36.2±1.3)%, (67.2±2.0)% and (46.3±3.2)%, respectively, and the difference was statistically significant ( F = 140.50, P < 0.001). Transwell assay indicated that the number of migrating cells in the control group, NET group and NET+DNase Ⅰ group were 317±18, 512±23 and 356±23, respectively, and the difference was statistically significant ( F = 75.39, P < 0.001). qRT-PCR assay revealed that the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET group were higher than those in the control group, while the relative expression of E-cadherin mRNA was lower than that in the control group, and the differences were statistically significant (all P < 0.001); the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin mRNA was higher than that in the NET group, and the differences were statistically significant (all P < 0.001). Immunofluorescence assay indicated that the number of cells expressing Vimentin in the control group, NET group and NET+DNase Ⅰ group were 35±6, 86±13 and 51±6, respectively, and the difference was statistically significant ( F = 24.65, P = 0.001); the number of cells expressing E-cadherin were 31±4, 11±3 and 24±2, respectively, and the difference was statistically significant ( F = 38.36, P < 0.001). Western blotting results demonstrated that the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET group were higher than those in the control group, while the relative expression of E-cadherin protein was lower than that in the control group; the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin protein was higher than that in the NET group. Conclusions:NET may promote the proliferation and migration of breast cancer cells by upregulating the expression of FZD10 and subsequently activating the Wnt-β-catenin signal pathway.

Objective To analyze the expression levels of natural cytotoxicity receptor(NCR)NKp46,NKp30,NKG2A and NKG2D in peripheral blood of non-small cell lung cancer(NSCLC)patients and its clinical diagnostic value.Methods A total of 50 NSCLC patients admitted to Affiliated Shijiazhuang Ping'an Hospital of Hebei University of Chinese Medicine from October 2021 to August 2022 were selected as the study group(NSCLC group).Meanwhile,20 healthy individuals who underwent physical examinations during the same period were selected as the control group.Peripheral blood samples were collected and the positive expression rate of NKp4,NKp30,NKG2A and NKG2D on NK cell surface was detected by flow cytometry.Clinical data from patients was collected and the correlation between the positive expression rate of NKp46,NKp30,NKG2A and NKG2D and pathological type,TNM staging and survival time were analyzed by One-way ANOVA/t test.Results Compared to control group,the positive expression rate of NKp46 on NK cells in peripheral blood of NSCLC patients were significantly elevated(42.09%±18.55%vs 25.92%±14.03%),the difference was statistically significant(t=3.511,P＜0.01).Comparison of the positive expression rates of NKG2A,NKG2D and NKp30 between the two groups and the differences were not statisticany significant(t=0.447,0.536,1.941,all P＞0.05).Subgroup analysis of NK cells revealed a significant increase in the proportion of CD56brightCD16+NK subgroups in NSCLC group compared to control group(3.76%±1.91%vs 2.42%±0.85%),the difference was statistically significant(t=3.017,P＜0.01).The positive expression rate of NKp46 in the CD56brightCD16+NK subgroup and CD56dimCD16+NK subgroup significantly increased(59.64%±21.12%vs 43.91%±16.04%,51.32%±19.84%vs 38.69%±15.12%),the differences were statistically significant(t=2.999,2.562,all P＜0.01).There was no significant difference in the positive expression rate of NKp46,NKG2A,NKG2D and NKp30 among squamous cell carcinoma and adenocarcinoma in lung(t=0.188～0.600,all P＞0.05).As TNM progresses,the positive expression rate of NKp46 on peripheral blood NK cells in NSCLC patients decreases,but the trend is of no statistical difference(F=2.381,P=0.10).Compared to patients with survival period≤12 months,the positive expression rate of NKp46 in patients with survival period>12 months was significantly higher(49.77%±17.52%vs 36.71%±15.41%),the difference was statistically significant(t=2.716,P＜0.01).Conclusion The positive expression rate of NKp46 in peripheral blood NK cells of NSCLC patients is increased,mainly in the CD56dimCD16+NK subgroup and CD56brightCD16+NK subgroup.The expression of NKp46 is related to the survival time of NSCLC patients,but not to the pathological type.Peripheral blood NCR NKp46 can serve as a potential biomarker for the diagnosis and prognostic evaluation of NSCLC.

Objective To analyze the expression levels of natural cytotoxicity receptor(NCR)NKp46,NKp30,NKG2A and NKG2D in peripheral blood of non-small cell lung cancer(NSCLC)patients and its clinical diagnostic value.Methods A total of 50 NSCLC patients admitted to Affiliated Shijiazhuang Ping'an Hospital of Hebei University of Chinese Medicine from October 2021 to August 2022 were selected as the study group(NSCLC group).Meanwhile,20 healthy individuals who underwent physical examinations during the same period were selected as the control group.Peripheral blood samples were collected and the positive expression rate of NKp4,NKp30,NKG2A and NKG2D on NK cell surface was detected by flow cytometry.Clinical data from patients was collected and the correlation between the positive expression rate of NKp46,NKp30,NKG2A and NKG2D and pathological type,TNM staging and survival time were analyzed by One-way ANOVA/t test.Results Compared to control group,the positive expression rate of NKp46 on NK cells in peripheral blood of NSCLC patients were significantly elevated(42.09%±18.55%vs 25.92%±14.03%),the difference was statistically significant(t=3.511,P＜0.01).Comparison of the positive expression rates of NKG2A,NKG2D and NKp30 between the two groups and the differences were not statisticany significant(t=0.447,0.536,1.941,all P＞0.05).Subgroup analysis of NK cells revealed a significant increase in the proportion of CD56brightCD16+NK subgroups in NSCLC group compared to control group(3.76%±1.91%vs 2.42%±0.85%),the difference was statistically significant(t=3.017,P＜0.01).The positive expression rate of NKp46 in the CD56brightCD16+NK subgroup and CD56dimCD16+NK subgroup significantly increased(59.64%±21.12%vs 43.91%±16.04%,51.32%±19.84%vs 38.69%±15.12%),the differences were statistically significant(t=2.999,2.562,all P＜0.01).There was no significant difference in the positive expression rate of NKp46,NKG2A,NKG2D and NKp30 among squamous cell carcinoma and adenocarcinoma in lung(t=0.188～0.600,all P＞0.05).As TNM progresses,the positive expression rate of NKp46 on peripheral blood NK cells in NSCLC patients decreases,but the trend is of no statistical difference(F=2.381,P=0.10).Compared to patients with survival period≤12 months,the positive expression rate of NKp46 in patients with survival period>12 months was significantly higher(49.77%±17.52%vs 36.71%±15.41%),the difference was statistically significant(t=2.716,P＜0.01).Conclusion The positive expression rate of NKp46 in peripheral blood NK cells of NSCLC patients is increased,mainly in the CD56dimCD16+NK subgroup and CD56brightCD16+NK subgroup.The expression of NKp46 is related to the survival time of NSCLC patients,but not to the pathological type.Peripheral blood NCR NKp46 can serve as a potential biomarker for the diagnosis and prognostic evaluation of NSCLC.

Objective:To explore the effects of neutrophil extracellular traps (NET) on the proliferation and migration of breast cancer cells and the related mechanisms.Methods:The peripheral blood neutrophils were separated using density gradient centrifugation, 100 nmol/L phorbol 12-myristate 13-acetate (PMA) was used to stimulate the formation of NET. Breast cancer cell line MDA-MB-231 was selected and treated with NET (NET group) or deoxyribonucleaseⅠ (DNase Ⅰ) digested NET (NET+DNase Ⅰ group),normal cultured MDA-MB-231 cells were used as the control group. In MDA-MB-231 cells of each group; cell proliferation ability was detected using CCK-8 assay; cell migration was detected by cell scratch assay and Transwell assay; real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expressions of frizzled homolog 10 (FZD10), cyclin D1, E-cadherin and Vimentin immunofluorescence assay was performed to detect the formation of NET and the expressions of E-cadherin and Vimentin; Western blotting was used to detected the expression of FZD10-Wnt-β-catenin signaling pathway-related proteins.Results:Neutrophils were stimulated with PMA for 4 hours, and immunofluorescence assay results showed that NET exhibited fibrous reticular structure with high expression of citrullinated histone (citH3) and myeloperoxidase (MPO). The CCK-8 assay results showed that the absorbance values of cells in the control group, NET group and NET+DNase Ⅰ group were 1.25±0.06, 2.14±0.15 and 1.47±0.11, respectively, and the difference was statistically significant ( F = 80.60, P < 0.001). The results of scratch assay displayed that the percentage of wound closure in the control group, NET group and NET+DNase Ⅰ group were (36.2±1.3)%, (67.2±2.0)% and (46.3±3.2)%, respectively, and the difference was statistically significant ( F = 140.50, P < 0.001). Transwell assay indicated that the number of migrating cells in the control group, NET group and NET+DNase Ⅰ group were 317±18, 512±23 and 356±23, respectively, and the difference was statistically significant ( F = 75.39, P < 0.001). qRT-PCR assay revealed that the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET group were higher than those in the control group, while the relative expression of E-cadherin mRNA was lower than that in the control group, and the differences were statistically significant (all P < 0.001); the relative expressions of FZD10, cyclin D1 and Vimentin mRNA in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin mRNA was higher than that in the NET group, and the differences were statistically significant (all P < 0.001). Immunofluorescence assay indicated that the number of cells expressing Vimentin in the control group, NET group and NET+DNase Ⅰ group were 35±6, 86±13 and 51±6, respectively, and the difference was statistically significant ( F = 24.65, P = 0.001); the number of cells expressing E-cadherin were 31±4, 11±3 and 24±2, respectively, and the difference was statistically significant ( F = 38.36, P < 0.001). Western blotting results demonstrated that the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET group were higher than those in the control group, while the relative expression of E-cadherin protein was lower than that in the control group; the relative expressions of FZD10, β-catenin and Vimentin proteins in the NET+DNase Ⅰ group were lower than those in the NET group, while the relative expression of E-cadherin protein was higher than that in the NET group. Conclusions:NET may promote the proliferation and migration of breast cancer cells by upregulating the expression of FZD10 and subsequently activating the Wnt-β-catenin signal pathway.

Objective:To compare the clinical outcomes of arthroscopic external tension band fixation versus open reduction and internal fixation in the treatment of greater tubercle fracture of the humerus.Methods:A retrospective cohort study was conducted on 55 patients with greater tubercle fracture of the humerus admitted to Taizhou Hospital of Zhejiang Province from September 2019 to June 2022, including 24 males and 31 females, aged 26-80 years [(61.7±10.5)years]. Out of them, 35 patients treated with open reduction and internal fixation (open reduction group), and 20 patients were treated with external anchor tension band under arthroscopy (arthroscopy group). The operation time, and the Visual Analogue Scale (VAS) score, American Shoulder and Elbow Surgeons (ASES) score, Constant-Murley score and shoulder active range of motion (anterior flexion, abduction and posterior extension) before operation, at 1 month after operation and at the last follow-up were compared between the two groups. Bone healing was observed in both groups at the last follow-up. Postoperative complications were compared between the two groups.Results:All the patients were followed up for 12-29 months [(16.9±4.0)months]. There was no significant difference in operation time between the two groups ( P>0.05). There were no significant differences in the VAS score, ASES score, Constant-Murley score and shoulder active range of motion between the two groups before operation ( P>0.05). The VAS score of the arthroscopy group was 3(2, 3)points at 1 month after operation, which was significantly lower than that of the open reduction group [4(3, 4) points] ( P<0.01). No significant difference was found in the VAS score at the last follow-up between the two groups ( P>0.05).The ASES scores of the arthroscopy group were (70.6±4.2)points and (90.2±3.7)points at 1 month after operation and at the last follow-up respectively, which were significantly higher than those of the open reduction group [(64.7±6.4)points and (87.5±4.9)points respectively] ( P<0.05 or 0.01). There was no significant difference in the Constant-Murley score between the arthroscopy group [(71.8±4.3)points] and the open reduction group [(70.9±5.3)points] at 1 month after operation ( P>0.05), while the Constant-Murley score of the arthroscopy group was (94.1±3.1)points at the last follow-up, which was significantly higher than that of the open reduction group [(89.2±4.7)points] ( P<0.01). At 1 month after operation and at the last follow-up, ranges of motion of the anterior flexion, abduction and posterior extension were (52.7±12.3)° and (140.0±16.9)°, (57.4±8.6)° and (125.0±14.3)°, and 16(15, 19)° and 25(20, 30)° in the arthroscopy group respectively, which were significantly higher than those in the open reduction group [(42.2±5.2)° and (110.9±14.0)°, (52.8±6.0)° and (103.7±11.7)°, and 10(10, 20)° and 16(15, 25)° respectively] ( P<0.05 or 0.01). At the last follow-up, it was found that bony union was achieved in both groups. There were no obvious complications such as incision infection or joint stiffnessin both groups. In the open reduction group, 2 patients had internal fixation failure within 1-3 months after operation but was treated with revision operation; 6 patients developed shoulder stiffness at 3-6 months after operation but had outpatient rehabilitation. The incidence rate of postoperative complications in the arthroscopy group [0%(0/20)] was significantly lower than that in the open reduction group [23%(8/35)] ( P<0.05). Conclusion:Compared with open reduction and internal fixation with plates and screws, arthroscopic external anchor tension band fixation in the treatment of greater tuberosity fracture of the humerus has the advantages of earlier pain relief, better shoulder functional improvement, better recovery of shoulder mobility, and fewer complications.

Objective:To explore the effects of breast cancer mesenchymal stem cells (BC-MSC) on the proliferation and migration of breast cancer MCF-7 cells and the related mechanisms.Methods:The resected cancer tissues and paracancerous tissues were taken from breast cancer patients after surgery, and the bone marrow samples of healthy people were selected. BC-MSC, breast cancer paracancerous mesenchymal stem cells (BCN-MSC) and bone marrow mesenchymal stem cells (BM-MSC) of healthy people were isolated and cultured by tissue adhesion method, and their differentiation ability was induced by the addition of osteogenic and lipogenic induction, and their surface markers were detected by flow cytometry. The supernatants of BC-MSC, BCN-MSC and BM-MSC of healthy people cultured for 48 h were collected and used for the culture of MCF-7 cells as BC-MSC group, BCN-MSC group and BM-MSC group, respectively, and the control group was the conventional cultured MCF-7 cells. The proliferation ability of MCF-7 cells in each group was detected by methyl thiazol tetrazolium (MTT) assay, the clone formation ability of MCF-7 cells was detected by plate cloning assay, the migration ability of MCF-7 cells was detected by Transwell assay, and the mRNA relative expressions of interleukin (IL)-6 and epithelial mesenchymal transition (EMT)-related genes (E-cadherin, vimentin, snail) were detected by quantitative real-time fluorescence polymerase chain reaction (qRT-PCR) in MCF-7 cells. Western blotting was used to detect expressions of p-STAT3, E-cadherin, vimentin and snail proteins in MCF-7 cells. Luminex liquid microarray technology was used to detect cytokine levels in culture supernatants of different mesenchymal stem cells (MSC). IL-6 neutralizing antibody was added into the supernatant of BC-MSC, MCF-7 cells were cultured with the supernatant (BC-MSC+IL-6 neutralizing antibody group), and then the proliferation and migration abilities of MCF-7 cells were tested, as well as the expression changes of related genes and proteins.Results:BC-MSC, BCN-MSC and BM-MSC were successfully isolated; BC-MSC had positive expressions of CD29, CD44 and CD90 and negative expressions of CD14, CD34 and CD45, which were in line with the characteristics of MSC. MTT assay showed that the absorbance values of MCF-7 cells cultured for 48 h in the control group, BC-MSC group, BCN-MSC group and BM-MSC group were 0.31±0.02, 0.54±0.03, 0.43±0.02 and 0.42±0.02, respectively, and the difference was statistically significant ( F = 56.52, P < 0.05); the results of plate cloning experiments showed that the number of clones in each petri dish of the four groups were 180±9, 439±17, 319±16 and 306±19, respectively, and the difference was statistically significant ( F = 222.70, P < 0.05); Transwell assay showed that the numbers of membrane-penetrating cells in the four groups were 6.5±1.0, 23.2±2.4, 16.0±1.3 and 14.8±2.0, respectively, with the statistically significant difference ( F = 49.44, P < 0.05); qRT-PCR assay showed that the relative expressions of IL-6 mRNA in the control group, BC-MSC group, BCN-MSC group and BM-MSC group were 1.07±0.11, 13.79±3.80, 6.68±1.66 and 6.12±1.52, respectively, and the difference was statistically significant ( F = 107.60, P < 0.05), and the relative expression of E-cadherin mRNA in MCF-7 cells of BC-MSC, BCN-MSC and BM-MSC groups was lower than that of the control group, while the relative expressions of vimentin and snail mRNA were higher than those of the control group, and the differences were statistically significant (all P < 0.05). Western blotting assay showed that the relative expression of E-cadherin mRNA in MCF-7 cells of BC-MSC, BCN-MSC and BM-MSC groups was lower than that of the control group. Western blotting showed that the level of E-cadherin protein in BC-MSC, BCN-MSC and BM-MSC groups was lower than that in the control group, and the levels of vimentin and snail proteins were higher than those in the control group; Luminex liquid microarray technology showed that the content of IL-6 cytokine in the supernatants of BC-MSC, BCN-MSC and BM -MSC cultures were higher, and the relative expressions were 1.75±0.21, 1.00±0.10 and 0.96±0.08, respectively, and the difference was statistically significant ( F = 43.22, P < 0.05). The results of MTT assay showed that the absorbance values of MCF-7 cells in BC-MSC group and BC-MSC+IL-6 neutralizing antibody group were 0.56±0.05 and 0.42±0.04, respectively, and the difference was statistically significant ( t = -3.11, P < 0.05); the results of Transwell assay showed that the numbers of membrane-penetrating cells in the two groups were 30.3±1.5 and 17.3±2.1, respectively, and the difference was statistically significant ( t = -7.12, P < 0.05); qRT-PCR assay showed that the relative expressions of E-cadherin mRNA were 0.44±0.05 and 0.76±0.05 ( t = 6.40, P < 0.01), the relative expressions of vimentin mRNA were 2.90±0.21 and 1.79±0.21 ( t = 5.29, P < 0.01), and the relative expressions of snail mRNA were 3.20±0.20 and 1.91±0.30 ( t = 2.16, P < 0.01); Western blotting assay showed that the degrees to down-regulate the expression of E-cadherin protein and up-regulate the expressions of vimentin and snail proteins in the BC-MSC+IL-6 neutralizing antibody group were weakened compared with the BC-MSC group. Conclusions:BC-MSC can promote the proliferation and migration of breast cancer MCF-7 cells probably through activating IL-6-STAT3 signaling pathway-induced EMT by its secretion of IL-6.

Objective To investigate the preventive and adverse effects of postnatal inhalation of Budesonide in early stage on bronchopulmonary dysplasia (BPD) in very low birth weight infants.Methods A total of 105 cases of high risk premature infants with BPD,who were born in the Neonatal Intensive Care Unit (NICU) from Shenzhen Maternity and Child Healthcare Hospital from July 15,2015 to December 25,2016,and their gestational age ≥ 27 weeks and ＜ 32 weeks or birth weight ≥ 1 000 g and ＜ 1 500 g were collected for a prospective randomized controlled trial,and were randomly divided into 3 groups:early inhalation group(34 cases),late inhalation group(34 cases) and non-inhalation group(37 cases).The oxygen time,and the incidence of BPD,periventricular-intraventricular hemorrhage (IVH),retinopathy of prematurity (ROP),necrotizing enterocolitis of the newborns (NEC),patent ductus arteriosus in preterm infants (PDA),sepsis and hyperglycemia of infants in 3 groups were compared.Results The average oxygen time in early inhalation group was 9 days,while in late inhalation group and the non-inhalation group was 15 days and 18 days,respectively.The average oxygen time in early inhalation group was significantly lower than that in the late inhalation group and the non-inhalation group,with the difference being statistically significant (H =6.09,P ＜ 0.05).The noninvasive ventilation time in early inhalation group was 3 days,while both the late inhalation group and non-inhalation group were 6 days.The noninvasive ventilation time in early inhalation group was significantly lower than that in the late inhalation group and non-inhalation group,with the difference being statistically significant (H =6.17,P ＜0.05).The incidence of BPD in the early inhalation group,late inhalation group and non-inhalation group were 14.7％ (5/34 cases),20.6％ (7/34 cases) and 37.8％ (14/37 cases),respectively.The incidence of BPD in non-inhalation group was significantly higher than that in the early inhalation group and late inhalation group,with the difference being statistically significant (x2 =12.017,P ＜ 0.05).There were no significant differences in IVH,ROP,NEC,PDA,sepsis and hyperglycemia among the 3 groups (all P ＞ 0.05).Conclusions Postnatal inhalation of Budesonide in early stage in high risk very low birth weight infants can reduce the incidence of BPD and the oxygen time,and the adverse reactions are not obvious.

OBJECTIVE: To investigate the relationship of polycyclic aromatic hydrocarbons( PAH) metabolites,DNA oxidative damage and ring finger protein 2( RING2) expression in coke oven workers. METHODS: A judgment sampling method was used to select 497 coke oven workers in a steel plant as exposure group and 175 water treatment workers in the same plant as control group. The levels of urinary 1-hydroxypyrene, 2-hydroxynathalene, 2-hydroxyfluorene,9-hydroxyphenanthrene and 8-hydroxy deoxyguanosine(8-OHd G) were detected by high performance liquid chromatography.The RING2 expression in whole blood was measured by reverse transcription-polymerase chain reaction. RESULTS: The relative expression of urinary 1-hydroxypyrene,2-hydroxynathalene,2-hydroxyfluorene,9-hydroxyphenanthrene and RING2 in exposure group were higher than that in control group( P < 0. 01). The logistic regression analysis indicated that the higher the level of 1-hydroxypyrene,the higher the risk of high-RING2 expression( P < 0. 05) after adjusting for factors such as sex,age,smoking status,alcohol drinking,2-hydroxynathalene,2-hydroxyfluorene and 9-hydroxyphenanthrene.In 1-hydroxypyrene middle and high level groups,the 8-OHd G concentration of high-RING2 expression workers was significantly higher than those of low-RING2 expression workers( P < 0. 05). CONCLUSION: With the increase of urinary1-hydroxypyrene,the risk of high-RING2 expression was elevated,the degree of DNA oxidative damage was gradually increased.

Objective To evaluate the value of monitoring non-invasive cardiac output parameters in medical treatment of patent ductus arteriosus (PDA) in premature infants.Method Premature infants with PDA diagnosed three days after birth (gestational age:28 ～ 31 weeks or birth weight of 1 000 ～ 1 799 g) admitted to the neonatal intensive care unit (NICU) of our Hospital from February 2016 to August 2016 were enrolled in the study.These premature infants were assigned into treated PDA group (the treatment group) and untreated PDA group (the observation group) based on results of non-invasive cardiac output parameters CI and MD,with aorta CI ≥2.95 L/(min · m2),MD ≥21.50 m/min and pulmonary artery CI ≥4.55 L/(min · m2),MD ≥26.50 m/min as cut-off values.Statistical analysis was carried out using t test,x2 test.The closure rate of arterial duct of two groups and changes in non-invasive cardiac output parameters before and after the closure of arterial duct in the treatment group were compared.Result The overall closure rate of arterial duct was 85.1％ (57/67).The closure rate of arterial duct of the treatment group was 70.8％ (17/24),that of the observation group was 93.0％ (40/43),and the difference had statistical significance (P ＜ 0.05);Comparing the following parameters before and after ductal closure in the treatment group,the difference of pulmonary artery flow time (FT),aorta stroke volume index (SVI) and the integral of the flow profile (Vti) had statistical significance (P ＜ 0.05) [(217.6±19.3) ms vs.(235.8 ±21.4) ms,(22.4±6.0)ml/m2 vs.(25.2 ±7.7)ml/m2,(15.1 ± 4.1) cm vs.(17.2 ±5.3) cm].In the treatment group,after arterial duct was closed,aorta and pulmonary artery CI,MD decreased to some degree,but the difference had no statistical significance (P ＞ 0.05).Conclusion Non-invasive cardiac output parameters including aorta and pulmonary artery CI,MD have certain guiding significance for PDA drug treatment among premature infants;after PDA drug treatment,arterial duct closure condition cannot be judged simply by the changes of aorta and pulmonary artery CI,MD,ultrasonic cardiogram examination results should also be considered.

Objective: To investigate the effect of mesenchymal stem cells (MSCs) on apoptosis of breast cancer cell line MCF-7 induced by cisplatin (DDP), MSCs derived from breast cancer (BC-MSCs) or adjacent non-cancerous tissues (BN-MSCs) were isolated, cultured and identified. Methods: BC-MSCs and BN-MSCs were isolated and cultured by tissue adherent method. The differentiation potential of BC-MSCs was detected by osteogenic and adipogenic induction, and cell surface markers of BC-MSCs and BN-MSCs were evaluated by flow cytometry. MCF-7 cells were co-treated with DDP and conditioned medium (CM) collected from BC-MSCs and BN-MSCs after being cultured for 48 hours, respectively. Inhibition rate of cell proliferation was evaluated by MTT. Cell apoptosis and viability were detected by MUSE cell analyzer. Cytokines in MSC-CM were detected by Luminex liquid chip. Interleukin 6 (IL-6) mRNA expressions in MCF-7 cells with different treatment were detected by RT-PCR. Results: The morphology of BC-MSCs and BN-MSCs successfully isolated and cultured was uniform fibroblast-like clusters under the microscope. These cells expressed high levels of CD29 and CD44, but neither CD14 nor CD34 were detected. MSCs could also differentiate into osteoblasts and adipocytes after specific induction. After treatment with 2.5, 5, 10, 20, 40 and 80 μmol/L DDP, the inhibitory rates of proliferation of MCF-7 cells in DDP group were (17.33±2.00)%, (22.37±0.73)%, (30.77±1.23)%, (44.93±1.27)%, (62.03 ±1.97)% and (73.93±1.10)%, respectively. While the inhibitory rates of DDP+ BC-MSCs group were (8.27±0.63)%, (11.50±1.30)%, (20.57±0.93)%, (32.60 ±1.90)%, (52.27±0.73)% and (62.13±2.17)%, respectively. The inhibitory rates of DDP+ BN-MSCs group were (12.90±1.60)%, (16.53±2.87)%, (25.90±1.50)%, (39.40±2.40)%, (57.40±0.70)% and (69.03±1.07)%, respectively. The inhibitory rates of DDP+ BC-MSCs group were significantly lower than those of DDP group (P<0.05). The apoptotic rates of MCF-7 cells in DDP group, DDP+ BC-MSCs group and DDP+ BN-MSCs group were (47.77±1.98)%, (29.20±2.12)% and (37.92±2.21)%, respectively. The apoptotic rates of DDP group was significantly higher than that of DDP+ BC-MSCs group (P<0.05). The cell viabilities of MCF-7 in DDP group, DDP+ BC-MSCs group and DDP+ BN-MSCs group were 0.52±0.02, 0.72±0.02 and 0.64±0.02, respectively. The cell viability of DDP group was significantly lower than that of DDP+ BC-MSCs group (P<0.05). The result of Luminex liquid chip analysis showed that, the level of IL-6 in BC-MSCs group increased 2.50±0.68 fold when compared with BN-MSCs group (P<0.05). The relative expressions of IL-6 mRNA in DDP group and DDP+ BC-MSCs group were 1.02±0.10 and 7.58±0.55, respectively, with a statistically significant difference (P<0.01). The apoptotic rates of MCF-7 cells in DDP+ BC-MSCs group with or without IL-6 neutralizing antibody were (27.41±1.95)% and (42.45±2.87)%, respectively, with a statistically significant difference (P<0.05). The cell viabilities of MCF-7 cells in DDP+ BC-MSCs group with or without IL-6 neutralizing antibody were (72.40±2.60)% and (59.76±3.89)%, respectively, with a statistically significant difference (P<0.05). Conclusions BC-MSCs and BN-MSCs have been isolated and cultured successfully. Compared with BN-MSCs, BC-MSCs could attenuate the effect of DDP on MCF-7 cells, evidently decrease the apoptosis and increase the proliferation and vitality in an IL-6 dependent manner.