OBJECTIVETo observe the level of intracellular reactive oxygen species (ROS) in rats with severe burn and pulmonary microvascular endothelial cells (PMVECs) treated with serum of rat with burn injury, and to investigate the relationship between ROS and apoptosis of PMVECs.

METHODS(1) Twenty-four SD rats were divided into sham injury group ( n = 3) and burn group (n = 21) according to the random number table (the same grouping method below). Rats in burn group were inflicted with 30% TBSA full-thickness scald on the back, and rats in sham injury group were sham injured. Blood samples were collected from abdominal aorta at post injury hour 6, 12, 24, 36, 48, 60, 72 respectively from 3 rats of burn group. The serum content of ROS was assayed by ELISA. The same determination was performed in rats of sham injury group. (2) Five rats were subjected to scald injury as above, and burn serum was prepared 24 hours after injury. Another 5 rats without receiving any treatment were used to prepare normal serum. (3) Marginal pulmonary tissue was harvested from 20 SD young rats. Cells were cultured with tissue block method and indentified with immunohistochemical staining. The third passage of PMVECs in logarithmic phase were inoculated in 6-well plates and 12-well plates. PMVECs in both plates were divided into 4 groups: normal serum group, burn serum group, normal serum + MnTBAP group, and burn serum + MnTBAP group, with 3 wells in each group. Cells in the former 2 groups were respectively cultured with special nutrient solution of endothelial cells without serum added with 15% healthy rat serum or 15% burn rat serum. Cells in the latter 2 groups were cultured with the same culture conditions as in the former two groups correspondingly with addition of 100 µmol/L MnTBAP in the nutrient solution. After being cultured for 24 h, the content of ROS in PMVECs in 6-well plates was detected with flow cytometry. The apoptosis of PMVECs in 12-well plates was observed with acridine orange-ethidium bromide staining, and the apoptosis rate was calculated. Data were processed with one-way analysis of variance and LSD-t test.

RESULTS(1) The serum contents of ROS in rats of burn group were respectively (187 ± 21), (235 ± 22), (231 ± 25), (291 ± 20), (315 ±23) nmol/mL at post injury hour 24, 36, 48, 60, 72, which were significantly higher than that in sham injury group [(141 ± 19) nmol/mL, with t values respectively 7. 86, 9. 57, 13. 87, 14.98, 18.40, P values below 0.01]. (2) Primary cells grew slowly and showed a cobblestone appearance. After passages, cells grew with orderly distribution. The positive rate of coagulation factor VIII of cells was (96 ± 5)% , and thus they were identified as PMVECs. (3) In normal serum group, burn serum group, normal serum + MnTBAP group, and burn serum + MnTBAP group, the contents of ROS in PMVECs were respectively 798 ± 40, 1 294 ± 84, 763 ± 59, 926 ± 42 ( F =93.01, P <0.01), and the apoptosis rates of PMVECs were respectively (6.2 ± 1.3)%, (57.3 ± 6. 7)%, (3.7 ± 0. 8)%, (28.7 ± 5. 7)% (F = 224.50, P <0.01) after being cultured for 24 h. Compared with those of normal serum group, the content of ROS and apoptosis rate of PMVECs in burn serum group increased significantly (with t values respectively 10.40 and 49.06, P values below 0.01). The content of ROS and apoptosis rate of PMVECs in burn serum + MnTBAP group were significantly lower than those in burn serum group (with t values respectively 7.48 and 23.94, P values below 0.01).

CONCLUSIONSSerum content of ROS was increased in severely burned rats. Burn rat serum stimulation on PMVECs can lead to the increase of the intracellular ROS and induce apoptosis. However application of MnTBAP can scavenge ROS and reduce the apoptosis induced by burn rat serum.

Animals ; Apoptosis ; Burns ; blood ; therapy ; Endothelial Cells ; pathology ; Enzyme-Linked Immunosorbent Assay ; Lung ; blood supply ; Oxygen ; Rats ; Reactive Oxygen Species ; blood ; Serum ; metabolism

Objective:To determine national trends for breast conserving surgery and to explore the factors affecting the scale of breast conserving surgery in China.Methods:A questionnaire survey was mailed to 110 hospitals with an year′s volume of more than 200 breast cancer surgeries in each center in China concerning hospital variations and percentage of breast conserving surgery.Results:The overall proportion of breast conserving surgery is 21.9% for operable breast cancer in China. There is a significant positive correlation between local Gross Domestic Product (GDP) and the rate of breast conserving surgery ( P=0.001). Hospitals with higher annual operation volume have higher breast-conserving ratios( P=0.042). Compared with non-teaching hospitals, more patients with stage I breast cancer underwent breast conserving surgery in teaching hospitals ( P=0.021). After breast-conserving surgery, the proportion of positive margins needing reoperation had a lower percentage and in cancer hospitals it was the lowest ( P=0.023). The method of pathological evaluation and the remedy strategy for positive margin was not related to per capita GDP and hospital category ( P>0.05). Conclusions:This survey demonstrates the current practices of breast conserving surgery in China. Local GDP, hospital category and tumor stage were factors influencing breast conserving surgery. Breast conserving surgery in China is still at a low level compared with developed countries.

Chemicals possessing reactive electrophiles can denature innate proteins leading to undesired toxicity, and the overdose-induced liver injury by drugs containing electrophiles has been one of the major causes of non-approval and withdraw by the US Food and Drug Administration (FDA). Elucidating the associated proteins could guide the future development of therapeutics to circumvent these drugs' toxicities, but was largely limited by the current probing tools due to the steric hindrance of chemical tags including the common "click chemistry" labels. Taking the widely used non-steroidal anti-inflammatory drug acetaminophen (APAP) as an example, we hereby designed and synthesized an APAP analogue using fluorine as a steric-free label. Cell toxicity studies indicated our analogue has similar activity to the parent drug. This analogue was applied to the mouse hepatocellular proteome together with the corresponding desthiobiotin-SH probe for subsequent fluorine-thiol displacement reactions (FTDRs). This set of probes has enabled the labeling and pull-down of hepatocellular target proteins of the APAP metabolite as validated by Western blotting. Our preliminary validation results supported the interaction of APAP with the thioredoxin protein, which is an important redox protein for normal liver function. These results demonstrated that our probes confer minimal steric perturbation and mimic the compounds of interest, allowing for global profiling of interacting proteins. The fluorine-thiol displacement probing system could emerge as a powerful tool to enable the investigation of drug-protein interactions in complex biological environments.