【 Abstract 】 Objective To investigate the effects of 250 ml/m3 carbon monoxide (CO) inhalation or intraperitoneal infusion on lipopolysaccharide (LPS) induced rat intestinal tract injury, and to detect the roles of p38 mitogen-activated protein kinase (MAPK) pathway during CO administration. Methods After received 5 mg/kg LPS or an equal volume of normal saline by intravenous injection, 108 male SD rats were randomly divided into 6 groups: control group, CO inhalation (250 ml/m3) group, CO intraperitoneal infusion (250 ml/m3 at a rate of 2 L/min) group, LPS (5 mg/kg) group, LPS (5 mg/kg)+CO inhalation (250 ml/m3) group and LPS (5 mg/kg)+CO intraperitoneal infusion (250 ml/m3 at a rate of 2 L/min) group. The animals were differently sacrificed at 1, 3 and 6 h for the observation, and the ileum tissues were homogenized for determination the levels of platelet activator factor (PAF), intercellular adhesion molecule-1 (ICAM-1) and interlukin-10 (IL-10) with enzyme-lined immunosorbent assay, the content of maleic dialdehyde (MDA) with thiobarbitric acid, the activity of myeloperoxidase (MPO) with chemical method, the activity of superoxide dismutase (SOD) with hydroxylamine, the activity of phosphorylated p38 MAPK with Western blot, the pathology with light microscope, and the extents of cell apoptosis were showed by the ratio of the apoptotic cells which had less DNA to the total cells of a cell-suspension sample by using the flow cytometry after being stained with propidium iodide. Results Compared with both control, CO inhalation and intraperitoneal infusion group at the same time point, the levels of PAF, ICAM-1, MDA, MPO, cell apoptosis rate and the phosphorylated p38 MAPK protein in LPS group were increased, while IL-10 and SOD were decreased (P ＜ 0.05 or 0.01), and accompanied by severe intestinal tract injury. There were no statistics differences at the different time point in the same group. PAF, ICAM-1, MDA, MPO and cell apoptosis rate in both LPS+CO inhalation group and LPS+CO intraperitoneal infusion group were lower, while IL-10 and SOD were higher than the corresponding value in LPS group at the same time point (all P ＜ 0.05), with ameliorate injury too, but the expression of phosphorylated p38 MAPK was further up-regulated than that of LPS group (all P ＜ 0.05). However, there were no significant differences in these parameters between LPS+CO inhalation group and LPS+CO intraperitoneal infusion group. Conclusion 250 ml/m3 CO inhalation and intraperitoneal infusion exerts the similar protection against LPS induced rat intestinal tract injury via anti-oxidant, anti-inflammation, and anti-apoptosis. This may involve the p38 MAPK pathway.
Objective To explore the regulation of peroxisome proliferator-activated receptor γ coactivator 1α( PGC-1α) and NF-E2-related factor 2( Nrf2) on expression of γ-glutamylcysteine synthetase ( γ-GCS) , and their roles in chronic obstructive pulmonary disease( COPD) . Methods Twenty-four SD rats were randomly divided into a COPD group and a normal control group. COPD model was established by intratracheal instillation of lipopolysaccharide ( LPS) and daily exposure to cigarette smog in the COPD group. The lung function was measured and the pathological changes were observed. The protein and mRNA expressions of PGC-1α, Nrf2, and γ-GCS in lung tissue were measured by immunohistochemistry, Western blot, in site hybridization ( ISH) , and reverse transcription-polymerase chain reaction ( RT-PCR ) ,respectively. Results In the COPD group, the pulmonary function ( FEV0. 3, FEV0. 3 /FVC, PEF) damage and lung pathological changes were conformed as morphological characteristics of COPD. The mRNA of PGC-1α and Nrf2 expressed in lung tissues of two group rats in the region consistent with γ-GCS mRNA. The protein and mRNA expressions of PGC-1αand γ-GCS were markedly increased in the COPD group( all P lt;0. 05) ,and the protein expression of Nrf2 was obviously up-regulated ( P lt; 0. 01) , while Nrf2 mRNA had no significant difference between the two groups( P gt;0. 05 ) . Linear correlation analysis showed that the level ofPGC-1αprotein was positively correlated with the levels of Nrf2 protein and mRNA ( r = 0. 775, 0. 515, all P lt; 0. 01) , and the levels of PGC-1αand Nrf2 protein were positively correlated with the levels of γ-GCS protein ( r = 0. 531, 0. 575, all P lt; 0. 01) and mRNA ( r = 0. 616, 0. 634, all P lt; 0. 01) . Conclusions PGC-1α, which may serve as a co-activator of Nrf2, can up-regulate the expression of γ-GCS gene cooperatively with Nrf2 through a common pathway, which might involve in the oxidative and antioxidative mechanism in the pathogenesis of COPD.
【Abstract】ObjectiveTo study the application of ultrasonically activated scalpel in laparoscopic intestinal adhesion release.MethodsIntestinal adhesion release with ultrasonically activated scalpel under laparoscope was performed in 29 patients suffered from intestinal adhesive obstruction after gynecological operation. ResultsAll operations were successfully performed, and none of them converted into open surgery. Intestinal disruption occurred durring operation in 2 patients with extensive intestinal denseadhesion which were mended successfully under laparoscope. The operative duration was 30-150 min (mean 45 min). Postoperative complications such as bowel leakage, bleeding, abdominal infection were not experienced. Postoperative hospital stay was 3-7 days (mean 4 days). No case had relapse symptom such as abdominal distention or pain after 1-24 months of followup. ConclusionCompared with electric scalpel, ultrasonically activated scalpel can improve the operative safety, lessen tissue damage, shorten operative time, and reduce the chance of relapse in laparoscopic operation in gynecology.
Objective To study the effects of peroxisome proliferators-activated receptor (PPAR) γ on the growth of human hepatocellular carcinoma cells and explore the roles of phosphatase and tensin homologue deleted on chromosome ten (PTEN) and phospho-Akt in this process. Methods SMMC-7721 cells were treated with 15-d-PGJ2 or pioglitazone, which were two kinds of PPARγ ligands, at different concentrations. The viability of SMMC-7721 cells was evaluated by MTT assay. The cell cycle was analyzed by flow cytometry. PTEN mRNA level was determined by RT-PCR. The protein expressions of PTEN and pAkt were measured by Western blot analysis. Results It was demonstrated through MTT assay that both 15-d-PGJ2 and pioglitazone had an inhibitory effect on the growth of SMMC-7721 cells in a time- and dose- dependent manner. According to flow cytometry detection, more cells were arrested in G0/G1 phase. Increased expression of PTEN mRNA was detected in 15-d-PGJ2 or pioglitazone-treated cells through RT-PCR. Increased expression of PTEN protein and decreased expression of pAkt were confirmed by Western blot analysis. Conclusion The ligands of PPARγ could inhibit SMMC-7721 cells proliferation in a time- and dose- dependent manner. The upregulation of PTEN may be involved in the underlying mechanism.
ObjectiveTo investigate the expression of extracellular signalregulated kinase (ERK) and p38 mitogenactivated protein kinase （p38 MAPK） in autogenous vein grafts during vascular remodeling.MethodsAn autogenous vein graft model was established by transplanting the right jugular vein to infrarenal abdominal aorta in 80 Wistar rats. Vein graft samples were harvested 6 hours, 24 hours, 3 days, 7 days, 2 weeks, 4 weeks, 6 weeks and 8 weeks after surgery. Gene expression of ERK and p38 MAPK was measured by reverse transcriptionPCR. Western blot was used to detect the expression of protein products and phosphorylation protein products of ERK and p38 MAPK. Apoptosis of vascular smooth muscle cells (VSMCs) was determined by TUNEL. Proliferating cell nuclear antigen(PCNA) of VSMCs also was studied.ResultsThe expression of ERK1 mRNA and p38 MAPK mRNA increased considerably after surgery. ERK1 mRNA reached the peak on the 7th day 〔（33.2±14.2）%, P＜0.01〕, but p38 MAPK mRNA reached the peak on the second week after surgery 〔(58.8±26.2）%, P＜0.01〕. The expression of ERK1/2 detected by western blot reached the peak during 1 to 2 weeks and decreased gradually to normal level 6 weeks after surgery. The expression of p38 MAPK reached the peak during 2 to 4 weeks and decreased to 1/4 to 1/2fold 8 weeks after surgery. There was a positive relationship between ERK1 and PCNA(r=0.759 6，P＜0.01) and a positive relationship between p38 MAPK and apoptosis(r=0.892 2，P＜0.01). ConclusionActivation of MAPK system exists in autogenous vein grafts and it may become a new target for the therapy of stenosis after vein grafts.
Objective To explore the change of gene expression of stress activated protein kinase (SAPK) and its upstream signalregulated molecule ——mitogen activated protein kinases(MAPKs) (MKK4 and MKK7) in hypertrophic scar and autocontrol normal skin. Methods The total RNA was isolated from 8 hypertrophic scars and 8 auto-control skin, and then mRNA was purified. The gene expressions of MKK4, MKK7 and SAPK were examined with reverse transcriptionpolymerase chain reaction(RT-PCR) method. Results In hypertrophic scar, both MKK7 and SAPK genes weakly expressed. In auto-control skin, the expression of these 2 genes was significantly elevated in comparison with hypertrophic scar (Plt;0.01). The expression levelsof these 2 genes were 1.5 times and 2.6 times as long as those of hypertrophic scar, respectively. Gene expression of MKK4 had no significant difference between autocontrol skin and hypertrophic scar (Pgt;0.05). Conclusion Decreased gene expression of MKK7 and SAPK which results in reducing cell apoptosis might be one of the mechanisms for controlling the formation of hypertrophic scar.
ObjectiveTo investigate the effects of overexpression of alpha/beta hydrolase domain-containing protein 5 (ABHD5) on the invasion and migration of human colon cancer cell line HCT116 and the pathway of adenosine monophosphate-activated protein kinase (AMPK)/mechanistic target of rapamycin (mTOR).MethodsThe expression of ABHD5 in colon cancer tissues and its relationship with clinicopathological features was analyzed by UALCAN database. HCT116 cells were cultured in vitro and transfected with ABHD5 recombinant plasmid, then they were divided into control group, negative transfection group and ABHD5 transfection group. Real time quantitative PCR (qRT-PCR) was used to detect the expression of ABHD5 mRNA in HCT116 cells. The proliferation of HCT116 cells was detected by CCK-8 method. Transwell assay was used to detect the invasion and migration of HCT116 cells. The expression of matrix metalloprotein 9 (MMP-9), E-cadherin, Snail, and AMPK/mTOR pathway proteins p-AMPK, AMPK, p-mTOR and mTOR were detected by Western blot.ResultsThe results of the UALCAN showed that compared with normal colon tissues, the expression of ABHD5 mRNA in colon cancer tissues was decreased (P<0.05), and which in the adenocarcinoma and the N1 stage was lower than that of the mucinous adenocarcinoma (P<0.05) and N0 stage (P<0.05), respectively. Compared with the control group and the negative transfection group, the expression of ABHD5 mRNA in the ABHD5 transfection group was increased (P<0.05), the proliferation inhibition rate of HCT116 cells in the ABHD5 transfection group was increased (P<0.05), the numbers of migration and invasion cells in the ABHD5 transfection group were decreased (P<0.05), the expressions of MMP-9, Snail, p-mTOR and mTOR were reduced, and the expressions of E-cadherin, p-AMPK and AMPK were increased (P<0.05).ConclusionsThe overexpression of ABHD5 can inhibit the invasion and migration of colon cancer HCT116 cells, activate AMPK, and inhibit the expression of mTOR. It suggests that ABHD5 may play a role in inhibiting colon cancer by affecting AMPK/mTOR pathway.
ObjectiveTo investigate the expression changes and the repair effect of mitogen and stress- activated protein kinase 1 (MSK1) on spinal cord injury (SCI) in rats.MethodsOne hundred and twenty male Sprague Dawley (SD) rats (weighing 220-250 g) were used for the study, 70 of them were randomly divided into sham-operation group and SCI group (n=35), the rats in SCI group were given SCI according to Allen’s method, and the sham-operation group only opened the lamina without injuring the spinal cord; spinal cord tissue was collected at 8 hours, 12 hours, 1 day, 2 days, 3 days, 5 days, and 7 days after invasive treatment, each group of 5 rats was used to detect the expression of MSK1 and proliferating cell nuclear antigen (PCNA) by Western blot assay. Another 20 SD rats were grouped by the same method as above (n=10). In these rats, a negative control lentiviral LV3NC dilution was injected at a depth of approximately 0.8 mm at the spinal cord T10 level. The results of transfection at 1, 3, 5, 7, and 14 days after injection were observed under an inverted fluorescence microscope to determine the optimal transfection time of the virus. The other 30 SD rats were randomly divided into group A with only SCI, group B with a negative control lentiviral LV3NC injected after SCI, and group C with MSK1 small interfering RNA (siRNA) lentivirus injected after SCI, with 10 rats each group. The Basso, Beatlie, Bresnahan (BBB) score of hind limbs was measured at 1, 3, 5, 7, and 14 days after treatment; spinal cord tissue collected at the optimal time point for lentivirus transfection was detected the expression changes of MSK1 and PCNA by Western blot and the localization by immunofluorescence staining of MSK1 and PCNA proteins.ResultsWestern blot assay showed that there was no significant changes in the expression of MSK1 and PCNA at each time points in the sham-operation group. In the SCI group, the expression of MSK1 protein was gradually decreased from 8 hours after injury to the lowest level at 3 days after injury, and then gradually increased; the expression change of PCNA protein was opposite to MSK1. The expression of MSK1 in SCI group was significantly lower than that in the sham-operation group at 1, 2, 3, and 5 days after injury (P<0.05), and the expression of PCNA protein of SCI group was significantly higher than that of the sham-operation group at 8 hours and 1, 2, 3, 5, and 7 days after injury (P<0.05). The fluorescence expression of both the SCI group and the sham-operation group has be found and peaked at 7 days. There was a positive correlation between fluorescence intensity and time in 7 days after transfection. With the prolongation of postoperative time, the BBB scores of groups A, B, and C showed a gradually increasing trend. The BBB score of group C was significantly lower than those of groups A and B at 5, 7, and 14 days after treatment (P<0.05). After transfection for 7 days, Western blot results showed that the relative expression of MSK1 protein in group C was significantly lower than that in groups A and B (P<0.05); and the relative expression of PCNA protein was significantly higher than that in groups A and B (P<0.05). Immunofluorescence staining showed that MSK1 was expressed in the nuclei of the spinal cord and colocalized with green fluorescent protein, neuronal nuclei, and glial fibrillary acidic protein (GFAP). The relative expression area of MSK1 positive cells in group C was significantly higher than that in group B (P<0.05), and the relative expression areas of PCNA and GFAP positive cells were significantly lower than those in group B (P<0.05).ConclusionLentivirus-mediated MSK1 siRNA can effectively silence the expression of MSK1 in rat spinal cord tissue. MSK1 may play a critical role in the repair of SCI in rats by regulating the proliferation of glial cells.
OBJECTIVE: To observe the protein expression of phosphorylated form of P38 mitogen-activated protein kinase(P38MAPK) and c-Jun in hypertrophic scar skin and to explore their influences on the formation and maturation of hypertrophic scar. METHODS: The expression intensity and distribution of phosphorylated form of P38MAPK and c-Jun were examined with immunohistochemistry and pathological methods in 16 cases of hypertrophic scar skin and 8 cases of normal skin. RESULTS: In normal skin, the positive signals of phosphorylated form of P38MAPK mostly distributed in basal lamina cells of epidermis, while c-Jun was mainly located in epidermal cells and endothelial cells. The positive cellular rates of two proteins were 21.3% +/- 3.6% and 33.4% +/- 3.5% respectively. In proliferative hypertrophic scar skin, the particles of phosphorylated P38MAPK and c-Jun were mainly located in epidermal cells and some fibroblasts. The positive cellular rates of two proteins were significantly elevated to 69.5% +/- 3.3% and 59.6% +/- 4.3% respectively (P lt; 0.01). In mature hypertrophic scar, the expression of these proteins decreased but was still higher than that of normal skin. CONCLUSION: The formation and maturation of hypertrophic scar might be associated with the alteration of phosphorylated P38MAPK and c-Jun protein expression in hypertrophic scar.
ObjectiveTo investigate the antibody concentration and immune status of intensive care medical staff after vaccination against COVID-19. Methods From October 1, 2021 to February 28, 2022, the serial numbers of 47 hospitals were randomly selected by cluster stratified random sampling method. Blood samples were collected from 192 medical staff in intensive care department who had received inactivated novel coronavirus vaccine in 7 hospitals. The antibody concentration was determined by chemiluminescence method to find the antibody rule. Logistic regression analysis was used to determine the related factors affecting the production of antibodies. ResultsTotal antibody concentration of 192 blood samples was 23.25 (5.09, 270.22), IgG concentration was 0.94 (0.15, 4.48), IgM concentration was 0.05 (0.03, 0.12). Logistic regression analysis showed that the total antibody concentration might be related to gender and age, and the IgG concentration was significantly related to whether the third injection was administered. One hundred and twenty-seven people received 2 doses of inactivated vaccine, and the positive rate of IgG was the highest within 1 to 2 months, and decreased significantly after 3 months. The positive rate of IgG antibody was 95.4% within 60 days after receiving 3 doses of vaccine, 70% within 1 month after receiving the third dose of vaccine, and 100.0% within 1 to 2 months (P<0.05). The total antibody positive rate was 96.3% in people aged 17 to 35 years and 73.3% in people aged 36 to 58 years, showing statistical difference (P<0.05). The total antibody production rate of those who received the third dose of vaccine was 100.0%, and no severe case of COVID-19 occurred during the sampling period. Conclusions After the first, second, and third doses of COVID-19 vaccine, the total antibody concentration of the virus gradually increases to 100.0%, indicating initial immunity. However, the antibody concentration decreased gradually after 3 months of inoculation. The concentration of IgG in women is higher than that in men, and the concentration of antibody in young people is higher than that in middle-aged and elderly people during the same period.