Objective To examine the effects of SMAD4 on the proliferation and apoptosis of ovarian granulosa cells in rats with poly-cystic ovary syndrome(PCOS).Methods A PCOS rat model was established using DHEA,and ovarian granulosa cells were extracted and cultured in vitro.The expression of SMAD4 in ovarian granulosa cells was detected by quantitative real-time PCR and Western blot-ting.SMAD4-siRNA was transfected into ovarian granulosa cells from PCOS rats.The expression of SMAD4 mRNA after transfection was determined by quantitative real-time PCR.Western blotting was performed to detect the expression levels of PCNA,BAX,and BCL-2 proteins after transfection.A CCK-8 assay was performed to evaluate cell growth after siRNA interference.Results The HE staining results revealed that the number of ovarian follicular vacuoles increased and that the number of granulosa cell layers and corpus luteum decreased,thus indicating the establishment of a PCOS model.The FSHR positivity rate exceeded 95％.SMAD4 expression in ovarian granulosa cells was higher in the PCOS group than in the control group(P＜0.05).Furthermore,siRNA effectively reduced SMAD4 expression in ovarian granulosa cells of PCOS rats(P＜0.01),promoted proliferation,and inhibited the apoptosis of granulosa cells.Con-clusion The hindered growth of ovarian granulosa cells in PCOS rats may be linked to the overexpression of SMAD4 mRNA,which sug-gests that targeting SMAD4 could be a promising approach for treating ovulatory abnormalities in patients with PCOS.

Objective To examine the effects of SMAD4 on the proliferation and apoptosis of ovarian granulosa cells in rats with poly-cystic ovary syndrome(PCOS).Methods A PCOS rat model was established using DHEA,and ovarian granulosa cells were extracted and cultured in vitro.The expression of SMAD4 in ovarian granulosa cells was detected by quantitative real-time PCR and Western blot-ting.SMAD4-siRNA was transfected into ovarian granulosa cells from PCOS rats.The expression of SMAD4 mRNA after transfection was determined by quantitative real-time PCR.Western blotting was performed to detect the expression levels of PCNA,BAX,and BCL-2 proteins after transfection.A CCK-8 assay was performed to evaluate cell growth after siRNA interference.Results The HE staining results revealed that the number of ovarian follicular vacuoles increased and that the number of granulosa cell layers and corpus luteum decreased,thus indicating the establishment of a PCOS model.The FSHR positivity rate exceeded 95％.SMAD4 expression in ovarian granulosa cells was higher in the PCOS group than in the control group(P＜0.05).Furthermore,siRNA effectively reduced SMAD4 expression in ovarian granulosa cells of PCOS rats(P＜0.01),promoted proliferation,and inhibited the apoptosis of granulosa cells.Con-clusion The hindered growth of ovarian granulosa cells in PCOS rats may be linked to the overexpression of SMAD4 mRNA,which sug-gests that targeting SMAD4 could be a promising approach for treating ovulatory abnormalities in patients with PCOS.

Objective To analyze the related risk factors affecting the prognosis of elderly patients with severe community acquired pneumonia (SCAP). Methods A retrospective study method was conducted; the elderly (≥ 75 years old) patients with SCAP treated in the First Affiliated Hospital of Hainan Medical College from January 2015 to January 2019 were enrolled. The general data of patients were collected, including sex, age, oxygenation index (PaO2/FiO2), involved organs, presence or absence of following diseases or treatment: damage in multiple lung lobes, septic shock, basic diseases (cardiovascular disease, chronic lung disease, diabetes, hypertension, and cerebrovascular disease), invasive mechanical ventilation, ventilator-associated pneumonia (VAP), misinhalation event, hyponatremia, respiratory acidosis, hypoproteinemia, intubation times, total mechanical ventilation time, etc. According to the prognosis, the patients were divided into a death group and a survival group. The general data were compared between the two groups with different prognoses. Single factor analysis was carried out by selecting variables. The indicators with statistical significant differences in the results of univariate analysis were introduced into the multivariate Logistic regression analysis to analyze the related risk factors affecting the prognosis of elderly patients with SCAP. The receiver operating characteristic (ROC) curve was drawn to analyze the predictive values of risk factors in the patients with SCAP. Results A total of 112 patients were included, 33 died, and the mortality rate was 29.46%. Univariate analysis showed that the following factors were higher in the death group than those in the survival group: organ involvement >2 [69.70% (23/33) vs. 35.44% (28/79)], lung lobe damage ≥ 3 [75.76% (25/33) vs. 51.90% (41/79)], invasive mechanical ventilation [72.73% (24/33) vs. 32.91% (26/79)], diabetes [30.30% (10/33) vs. 12.66% (10/79)], intubation times ≥2 [57.58% (19/33) vs. 48.10% (38/79)], hypoproteinemia [75.76% (25/33) vs. 41.77% (33/79)], hyponatremia [72.73% (24/33) vs. 48.10% (38/79)], respiratory acidosis [66.67% (22/33) vs. 44.30 %(35/79)] and total mechanical ventilation time ≥ 15 days [69.70% (23/33) vs. 40.51 (32/79)]; the factors in the death group lower than those in the survival group were: septic shock [3.03% (1/33) vs. 17.72% (14/79)], chronic lung disease [6.06% (2/33) vs. 25.32% (20/79)] and PaO2/FiO2 [mmHg (1 mmHg = 0.133 kPa): 102.89±14.78 vs. 109.56±14.08],the differences were statistically significant (all P < 0.05); there were no significant differences in gender, age, cardiovascular disease, hypertension, VAP, misinhalation events and cerebrovascular disease between the two groups (all P > 0.05). Multivariate Logistic regression analysis showed that diabetes mellitus [odds ratio (OR) = 1.074, 95% confidence interval (95%CI) = 1.017-1.287, P =0.045], septic shock (OR = 2.765, 95%CI = 1.083-3.411, P = 0.047), hyponatremia (OR = 1.792, 95%CI = 1.128-1.417, P = 0.006), hypoalbuminemia (OR = 2.187, 95%CI = 1.872-5.462, P = 0.046), invasive mechanical ventilation (OR = 5.870, 95%CI = 2.324-23.796, P = 0.001), respiratory acid poisoning (OR = 2.934, 95%CI = 2.454-7.275, P = 0.043), time of mechanical ventilation (OR= 1.986, 95%CI = 2.467-3.483, P = 0.034), number of intubation (OR = 6.760, 95%CI = 2.116-24.696, P = 0.001), PaO2/FiO2 (OR = 1.981, 95%CI = 1.006-1.417, P = 0.007), organ involvement > 2 (OR = 2.924, 95%CI = 2.534-6.285, P = 0.048), chronic lung disease (OR = 2.887, 95%CI = 1.487-3.483, P = 0.039), and lung lobe damage≥3 (OR = 2.754, 95%CI = 1.131-1.798, P = 0.045) were independent risk factors affecting the prognosis of elderly patients with SCAP. ROC analysis showed that hyponatremia, hypoalbuminemia, invasive mechanical ventilation, total mechanical ventilation time, PaO2/FiO2, organ involvement > 2, damage of lung lobes ≥ 3, had predictive values for the prognosis of SCAP [the areas under ROC curve (AUC) were 0.377, 0.267, 0.301, 0.646, 0.650, 0.329, and 0.381, respectively, all P < 0.05]. Conclusions Underlying disease, invasive mechanical ventilation, respiratory acidosis, total mechanical ventilation time, PaO2/FiO2, intubation times ≥ 2, chronic lung disease and lung damage≥ 3 lobes are the independent risk factors for the prognosis of elderly patients with severe community acquired pneumonia. Clinical treatment should focus on the above aspects to minimize the mortality of patients.

Objective To study the anti-inflammatory,antitussive,and analgesia effects of Jinhua qingre capsules.Methods The anti-inflammatory effect was assessed by the methods include xylene-induced mouse auricular swelling and histamine-induced pigment oozing from skin vessel in rats;The antitussive and analgesic effect were assessed by ammonia water induced cough model and acetic acid-induced twisting method.Results In anti-inflammation experiment,the high dose (12 g·kg-1) and moderate (6 g·kg-1) groups of Jinhua qingre capsules showed significant inhibitory effect on auricular swelling and significant difference compared with control group (P ＜ 0.01,P ＜ 0.05.);the high dose (10 g· kg-1) and moderate dose (5 g·kg-1) groups of Jinhua Qingre capsules play a marked inhibitory role in the increase in mouse peritoneal capillary permeability (P ＜ 0.01,P ＜ 0.05).In antitussive experiment,high dose (12 g· kg-1) and moderate dose (6 g· kg-1) groups had significant inhibitory effect on cough caused by ammonia water (P ＜ 0.01,P ＜ 0.05) compared with control group.In analgesic experiment,the high dose (12 g·kg-1),moderate dose (6 g·kg-1),and low dose (3 g·kg-1) groups effectively reduced the writhing frequency of mice (P ＜ 0.01,P ＜ 0.05).Conclusion Jinhua qingre capsules have potential effects on anti-inflammatory,antitussive,and analgesic.

To verify the reliability of targeted detecting HER2 positive cancer cells and clinical pathological tissue specimens with a recombinant anti HER2 single chain antibody in single chain Fv fragment (scFv) format, we have constructed the fusion variable regions of the ScFv specific for HER2/neu. labeled a green-fluorescent protein(GFP). The humanized recombinant Anti HER2 ScFv-GFP gene was inserted into pFast Bac HT A, and expressed in insect cells sf9. Then the recombinant fusion protein Anti HER2 ScFv-GFP was properly purified with Ni2+-NTA affinity chromatography from the infected sf9 cells used to test the specificity of the fusion antibody for HER2 positive cancer cells. Firstly, the purified antibody incubated with HER2 positive breast cancer cells SKBR3, BT474 and HER2 negative breast cancer cells MCF7 for 12 h/24 h/48 h at 37 degrees C, in order to confirm targeted detecting HER2 positive breast cancer cells by Laser Confocal Microscopy. Furthermore, the same clinical pathological tissue samples were assessed by immunohistochemistry (IHC) and the fusion antibody Anti HER2 ScFv-GFP in the meanwhile. The data obtained indicated that the recombinant eukaryotic expression plasmid pFast Bac HT A/Anti HER2 ScFv-GFP was constructed successfully In addition, obvious green fluorescent was observed in insect cells sf9. When the purified fusion antibody was incubated with different cancer cells, much more green fluorescent was observed on the surface of the HER2 positive cancer cells SKBR3 and BT474. In contrast, no green fluorescent on the surface of the HER2 negative cancer cells MCF7 was detected. The concentration of the purified fusion antibody was 115.5 microg/mL, of which protein relative molecular weight was 60 kDa. The analysis showed the purity was about 97% and the titer was about 1:64. The detection results of IHC and fusion antibody testing indicated the conformity. In summary, the study showed that the new fusion antibody Anti HER2 ScFv-GFP can test HER2 positive cancer cells, indicating a potential candidate method for clinical HER2 positive specimens detection.
Animals ; Breast Neoplasms ; diagnosis ; pathology ; Female ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; Humans ; MCF-7 Cells ; Receptor, ErbB-2 ; analysis ; Recombinant Fusion Proteins ; genetics ; Sf9 Cells ; Single-Chain Antibodies ; genetics