Objective To confirm whether taxotere combined with mild hyperthermia will have synergistic effects,and to explore their joint mechanism of action.Methods Firstly the effective concentration of taxotere was determined by using MTT method to observe the effect of docetaxel on proliferation of human breast cancer cell line MCF-7.Then three samples of in vitro cultured human breast carcinoma MCF-7 cells were prepared,termed the the taxotere group,the taxotere plus mild hyperthermia group and the control group,and treated with effective concentration of taxotere exclusively or in combination with mild hyperthermia,or left without any special treatment.The taxotere plus mild hyperthermia group was subdivided into 5 subgroups according to the temperatures used (39.0 ℃,39.5 ℃,40.0 ℃,40.5 ℃,41.0 ℃) MTT assays were used to measure the proliferation and invasive capacity of the cells and their effective concentrations.Flow cytometry was used to detect cell apoptosis rates and any cell cycle changes in the control group.Western blotting was used to detect any changes in the expression of mitogen-activated protein kinases (MAPKs),Bcl-2/Bax,heat shock protein-70 (HSP-70) and P-gp.Results The taxotere with mild hyperthermia group demonstrated a significantly higher rate of apoptosis than that in the taxotere and control groups.There were also more cells in the G2/M phase observed.Combining taxotere with mild hyperthermia was found after 24 h to have significantly increased p-ERK,p-JNK,p38,HSP-70 and P-gp protein levels and to have significantly decreased Bcl-2 protein expression in contrast with the other two groups.Conclusions Combining taxotere with mild hyperthermia showed synergistic effects in vitro.It seemed to be limiting the accumulation of MCF-7 cells in the G2/M phase and activating the signal pathways of MAPKs while inhibiting the activation of Bcl-2/Bax signal pathways.Combining taxotere and mild hyperthermia can accelerate the expression of HSP70 and P-gp in MCF-7 cells.Hyperthermia might induce chemotherapy resistance.

Objective To investigate the influence of isoquercitrin on the inflammatory factors in LPS-induced RAW264.7 cells.Methods MTT method was used to detect inhibition ratio of RAW264.7 cells induced by isoquercitrin.The level of TNF-α in culture medium was measured by ELISA.Nitric oxide (NO) was detected by Nitrate Assay Kit.Western blotting was used to investigate the influence on the productions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2).Results The half inhibitory concentration (IC50) of isoquercitrin was 65.73 μmol·L-1.LPS had no inhibitory effect on the cells.Compared with LPS group,the level of TNF-α was decreased to 74.80% and 60.57% in isoquercitrin (20,10 μmol·L-1) groups in a dose-dependent manner.The results measured by Nitrate Assay Kit revealed that isoquercitrin (20,10 μmol·L-1) could suppress production of NO,the level of NO decreased to 79.34% and 68.81%(P<0.05).The Western blotting results showed that isoquercitrin (20,15,10 μmol·L-1) inhibited the productions of iNOS and COX-2 (P<0.05).Conclusion Isoquercitrin has anti-inflammatory effects by inhibiting the productions of TNF-α,NO,iNOS and COX-2,and the most effective dose for the inhibition is 10 μmol·L-1.

Objective To monitor the autophagy in osteosarcoma cells by constructing three rLC3B fusion expression vectors,respectively.Methods Rat LC3B gene sequence was amplified by PCR and cloned into pEGFP-C 1 and pmCherry-C1 to construct the fusion expression vector of pEGFP-rLC3B and pmCherry-rLC3B.Subsequently,the EGFP-rLC3B sequence was obtained by PCR with the pEGFP-rLC3B as a template,and cloned into pmCherry-C 1,so the pmCherry-EGFP-rLC3B fusion expression vector was constructed.Three plasmids were transfected into U-2OS cells,and the starvation or Rapamycin was adopted to induce autophagy or the chloroquine or Baf-A1 was used to inhibit autophagy,to verify the above plasmids' function in autophagy detection by laser scanning confocal microscopy.Western blot was used to detect the endogenous LC3B and exogenous EGFPrLC3B,pmCherry-rLC3B and mCherry-EGFP-rLC3B,and to verify the correct expression of exogenous rLC3B and their function of autophagy detection.Finally,cleaved free EGFP was detected by western blot to evaluate the level of autophagic degradation.Results Three fusion expression vectors were constructed successfully through sequencing and restriction enzyme digestion validation.The starvation or Rapamycin was adopted to induce autophagy or the chloroquine or Baf-A 1 was used to inhibit autophagy in transfected U-2OS cells.Clear autophagosomes and autolysosomes were observed by laser scanning confocal microscopy.Endogenous LC3B and exogenous EGFP-rLC3B,pmCherry-rLC3B and mCherry-EGFP-rLC3B were detected through western blot.Finally,western blot verified that the expression of cleaved free EGFP was significantly up-regulated with the increase of starvation time.12 h group increased 1.05 times than the control group and 24 h group increased 1.56 times,showing that the levels of autophagic degradation increased.Conclusion EGFP-rLC3B can be used to detect autophagosome and evaluate the level of autophagic degradation.mCherry-rLC3B can be used to detect autophagosome and autolysosome,but can't distinguish autophagosome from autolysosome.The pmCherry-EGFP-rLC3B has an advantage in the detection of autophagic flux which can distinguish autophagosome from autolysosome.

Objective To study the changes of gastrointestinal movement function in rats with chronic unpredictable mild stress(CUMS) and explore the mechanisms underlying it.Methods The rats were divided into stress model group and control group.The stress model rats were induced by 21-day chronic unpredictable mild stress as well as social-isolated fed.The rate of ink propulsion of gastrointestinal tract and the contraction of intestinal canal in rats were observed.Immunohistochemistry was adopted to detect the expression of OT in rats.Results (1) After the models were induced,weight-gain and sucrose preference of model group ((69.97 ± 9.81) g,(49.05± 5.98) g) were lower than those in control group ((116.27 ± 13.60) g,(83.51 ± 3.08) g) (P ＜ 0.001),and both the crossing-score and rearing-score ((24.00 ± 13.52),(3.90 ± 2.51)) were lower than those in control group ((53.60 ± 27.98),(11.50 ± 8.85)) in the open-field test.(2) The rate of ink propulsion of model group ((67.33 ± 6.24) ％) was decreased when compared to the control group ((76.83 ± 10.00) ％) (P ＜ 0.05),and the intestinal canal contraction amplitude and contraction frequency ((1.37 ± 0.18) g,(0.58 ± 0.02) S-1) were lower than those in control group ((1.88 ± 0.13) g),(0.62 ± 0.04) S-1) (P ＜ 0.05).(3) Compared with the control group (6.07 ± 3.71),OT immunoreactive substance was increased in model group (59.17 ± 16.08) of rats (P＜0.001).Conclusion Chronic stress can cause the decrease in gastrointestinal movement function of rats.These changes may be related to the increased expression of OT in paraventricular nucleus.

Objective:To investigate the relationship between the serum alkaline phosphatase (ALP), prostate specific antigen (PSA) and bone metastasis in initially diagnosed prostate cancer (PCa) patients in different ISUP(International Society of Urological Pathology)groups.Methods:The 368 initial diagnosed prostate cancer patients recruited from January 2013 to December 2018 were retrospectively analyzed, including 247 cases in the Third Affiliated Hospital of Sun Yat-sen University, 111 cases in the Yuebei People's Hospital Affiliated to Medical College of Shantou University and 10 cases in Shenzhen Hospital of Southern Medical University. According to whether there was bone metastasis at the initial diagnosis, it was divided into 230 cases in the bone metastasis group and 138 cases in the non bone metastasis group. There was no significant difference between the two groups in age [(71.9±9.4) years and (71.2±8.7) years], body mass index (BMI) [(23.1±3.7) kg/m 2 and (23.7±2.6) kg/m 2]. There were significant differences in PSA [(307.3±847.0) ng/ml and (84.5±257.3) ng/ml] and ALP [(174.5±270.8) U/L and (71.0±23.2) U/L] between the two groups. In different PSA subgroups, there were 45 cases in PSA <10 ng/ml, 35 cases in PSA 10-20 ng/ml and 288 cases in PSA >20 ng/ml. The differences of ALP and PSA between bone metastasis group and non-bone metastasis group based on different ISUP stratification were analyzed, ROC curves were used to predict their risks of bone metastasis. Results:There were 3(1.3%), 22(9.6%), 34(14.8%), 85(37.0%) and 86 (37.4%) prostate cancer patients with bone metastasis from ISUP group 1 to 5, and 14(10.1%), 19(13.8%), 29(21.0%), 32(23.2%) and 44(31.9%) without bone metastasis, respectively. There was significant difference in the serum ALP levels between the bone metastasis group and the boneless metastasis group in the ISUP group 4(157.6±207.7 vs. 66.5±17.0) and 5(189.4±257.5 vs. 69.2±18.4)( P<0.001) and PSA levels had difference in the ISUP group 3(240.3±313.0 vs. 42.4±42.1), 4(152.3±184.5 vs. 44.7±33.3) and 5(435.2±1006.3 vs. 60.8±84.8)( P<0.001). There was statistically significant between the bone metastasis group and the without(336.1±882.2 vs. 139.3±328.1) when PSA>20 ng/ml( P=0.006). ROC curve analysis: the cut-off values of ALP were 115.5, 109.0, 75.5 and 86.0 U/L from ISUP group 2 to 5 respectively, the sensitivity was 23.8%, 56.5%, 66.4% and 50.6% respectively, the specificity was 99.7%, 93.4%, 78.3% and 89.2% respectively, and the accuracy were 59.4%, 73.1%, 69.7% and 63.3%, respectively. The cut-off values of PSA were 39.5, 93.1, 54.2 and 28.9 ng/ml from ISUP group 2 to 5 respectively, the sensitivity was 64.4%, 68.4%, 87.4% and 88.3% respectively, and the specificity was 79.5%, 90.6%, 63.7% and 61.6% respectively, and the accuracy were 71.6%, 78.1%, 80.1% and 79.2%, respectively. Conclusion:ALP increased significantly in ISUP group ≥4 and PSA in ISUP group ≥3, which related to bone metastasis in patients with initial diagnosed prostate cancer.

Identifying and comparing the chemical constituents of wild silkworm cocoon and silkworm cocoon is of great significance for understanding the domestication of silkworm. In this study, we used high temperature and high pressure and methanol-water system to extract cocoon chemical constituents. We used UHPLC-MS to identify and compare cocoon chemical constituents of wild silkworm and domestic silkworm Dazao and Haoyue strains. The cocoon metabolic fingerprints of wild silkworm and domestic silkworm Dazao and Haoyue strains were obtained by using the UHPLC-MS in the positive ion mode and negative ion mode. By annotation, we found that cocoon chemical compounds with high abundances contained amino acids, flavonoids, alkaloids, terpenes, organic acids, and lignans. PLS-DA showed that the cocoon components were significantly different among the wild silkworm and two domestic silkworm strains Dazao and Haoyue. Proline, leucine/isoleucine and phenylalanine showed significantly higher abundances in the cocoon of domestic silkworm Dazao strain than in those of wild silkworm and domestic silkworm Haoyue strain. The flavonoid secondary metabolites are abundant in the Dazao cocoon, including quercetin, isoquercetin, quercetin 3-O-sophoroside, quercetin-3-O-α-L-rhamnoside, quercetin-3-O- rutinoside, and kaempferol. The other secondary metabolites, alkaloids, terpenes and lignans, showed higher abundances in the wild silkworm cocoon than in the domestic silkworm cocoon, including neurine, candicine, pilocarpidine, artemisiifolin, eupassopin, and eudesobovatol. By exposing cocoons to UV light and observing the green fluorescence of flavonoids, we found that Dazao cocoon had the most flavonoids, and Haoyue cocoon had least flavonoids and wild silkworm cocoon had mediate flavonoids. Alkaloids and organic acids are good anti-insect and antimicrobial agents, which have high abundance in the wild silkworm cocoon and could enhance the defense ability of wild silkworm cocoon. Flavonoids are abundant in the cocoon of domestic silkworm Dazao strain, which the main factors are leading to the yellow-green cocoon of Dazao.
Animals ; Bombyx ; Chromatography, High Pressure Liquid ; Flavonoids ; Mass Spectrometry

Ankylosing spondylitis (AS) combined with lower cervical fracture is often categorized into unstable fracture, with a high incidence of neurological injury and a high rate of disability and morbidity. As factors such as shoulder occlusion may affect the accuracy of X-ray imaging diagnosis, it is often easily misdiagnosed at the primary diagnosis. Non-operative treatment has complications such as bone nonunion and the possibility of secondary neurological damage, while the timing, access and choice of surgical treatment are still controversial. Currently, there are no clinical practice guidelines for the treatment of AS combined with lower cervical fracture with or without dislocation. To this end, the Spinal Trauma Group of Orthopedics Branch of Chinese Medical Doctor Association organized experts to formulate Clinical guidelines for the treatment of ankylosing spondylitis combined with lower cervical fracture in adults ( version 2024) in accordance with the principles of evidence-based medicine, scientificity and practicality, in which 11 recommendations were put forward in terms of the diagnosis, imaging evaluation, typing and treatment, etc, to provide guidance for the diagnosis and treatment of AS combined with lower cervical fracture.