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The fungus Xylaria sp. KYJ-15 was isolated from Illigera celebica. Based on the one strain many compounds (OSMAC) strategy, the strain was fermented on potato and rice solid media, respectively. As a result, two novel steroids, xylarsteroids A (1) and B (2), which are the first examples of C28-steroid with an unusual β- and γ-lactone ring, respectively, along with two new dihydroisocoumarin glycosides, xylarglycosides A (3) and B (4), were identified. Their structures were elucidated by spectroscopic methods, X-ray diffraction and electronic circular dichroism (ECD) experiments. All isolated compounds were evaluated for cytotoxicity, DPPH radical scavenging activity, acetylcholinesterase inhibitory and antimicrobial effect. Compound 1 exhibited potent AChE inhibitory activity with an IC50 value of 2.61 ± 0.05 μmol·L-1. The β-lactone ring unit of 1 is critical for its AChE inhibitory activity. The finding was further confirmed through exploring the interaction of 1 with AChE by molecular docking. In addition, both compounds 1 and 2 exhibited obvious antibacterial activity against Bacillus subtilis with a minimum inhibitory concentration (MIC) of 2 μg·mL-1. Compounds 3 and 4 exhibited antibacterial activities against Staphylococcus aureus with MICs of 4 and 2 μg·mL-1, respectively, which also exhibited DPPH radical scavenging activity comparable to the positive control with IC50 values of 9.2 ± 0.03 and 13.3 ± 0.01 μmol·L-1, respectively.
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Humans , Acetylcholinesterase , Molecular Docking Simulation , Anti-Bacterial Agents , Glycosides , Lactones , PainABSTRACT
Objective:To investigate the accumulation of porphyrin metabolites [uroporphyrinogen (UP) Ⅰ and coproporphyrinogen (CP) Ⅲ] induced by 5-aminolevulinic acid (5-ALA) in the urine of rats with colorectal cancer.Methods:The rat model of colorectal cancer was established by dimethylhydrazine (DMH). Urine samples were collected from 30 colorectal cancer rats (colorectal cancer group) and 30 normal rats (normal group). Each animal was given 5-ALA (50 mg/kg) by gavage, and urine was collected after 2, 4, 6 and 8 h. The contents of urinary porphyromogen Ⅰ and porphyromogen faecalis Ⅲ in urine were detected by high performance liquid chromatography (HPLC).Results:There was no significant difference in the contents of UP Ⅰ and CP Ⅲ in urine between colorectal cancer group and normal group before oral administration of 5-ALA ( P>0.05). After oral administration of 5-ALA, the contents of UP Ⅰ and CP Ⅲ in urine of colorectal cancer group were significantly higher than those of normal group ( P<0.05). The contents of UP Ⅰ and CP Ⅲ in urine of colorectal cancer group reached the highest value at 4 hours. According to the receiver operating characteristic (ROC) curve drawn from 4-hour test results, the threshold value of UP Ⅰ for colorectal cancer diagnosis was 50.43 μmol/g, with corresponding sensitivity 96.7%, and the specificity 63.3%, respectively. The threshold value of CP Ⅲ for colorectal cancer diagnosis was 108.85 μmol/g, with corresponding sensitivity 66.7%, and the specificity 86.7%, respectively. Conclusions:The accumulation of porphyrin metabolites induced by 5-ALA in the urine of rats with colorectal cancer is significant. The porphyrin metabolites in urine may be a new tumor marker of colorectal cancer, which provides an experimental basis for the diagnosis of colorectal cancer.
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Objective:To explore the effect and mechanism of 5-Aminolevulinic Acid-Photodynamic Therapy (5-ALA-PDT) on the apoptosis of the human colonic carcinoma HT-29 cells.Methods:HT-29 cells were cultured in vivio and divided into four groups: blank control group, 5-ALA group, PDT group and 5-ALA-PDT group.The control group was not given photosensitizer and light treatment; 5-ALA group was given photosensitizer ; PDT group was given light treatment; 5-ALA-PDT group was given photosensitizer and light treatment at the same time. Flow cytometry was used to observe the apoptosis of HT-29 cells. Reverse transcription polymerase chain reaction (RT-PCR) was used to observe the expression of B-type lymphoma-2 (Bcl-2) and Bcl-2 associated X protein (Bax) in HT-29 cells. Ultraviolet spectrophotometry was used to detect the expression of Caspase-3. Results:The apoptotic rate of 5-ALA-PDT group was significantly higher than that of blank control group, 5-ALA group and PDT group ( P<0.05). Compared with the blank control group, 5-ALA-PDT group and PDT group, the expression of Bcl-2 in the 5-ALA-PDT group was statistically significant ( P<0.05), but there was no significant difference in Bax expression among the four groups ( P>0.05). The expression of Bax/Bcl-2 in 5-ALA-PDT group was significantly higher than that in blank control group, 5-ALA group and PDT group ( P<0.05). The expression of Caspase-3 in 5-ALA-PDT group was significantly higher than that in blank control group, 5-ALA group and PDT group ( P<0.05). Conclusions:5-ALA-PDT can induce apoptosis of HT-29 cells, and its mechanism may be related to the induction of apoptosis through Bax/Bcl-2 pathway.
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Objective:To investigate the changes of porphyrin metabolites in urine of patients with colorectal cancer before and after operation and their correlation with prognosis.Methods:One hundred patients with colorectal cancer were collected in First Affiliated Hospital of Hebei North University from June 2016 to December 2016, urine was collected before operation, 1 week after operation, 1 year after operation and before recurrence. The contents of urinary porphyrin metabolites of uroporphyrinogenI (UP Ⅰ) and coproporphyrinogen Ⅲ(CP Ⅲ) were detected by high performance liquid chromatography. Toanalyse the changes of UPⅠ and CPⅢ levels before and after operaction of colorectal cancer and their correlation with clinicopathologicalcharacteristics,and the recurrence and metastasis after operation.Results:The levels of UPⅠ and CPⅢ in urine of patients with colorectal cancer after operation were significantly lower than those before operation [(66.80 ± 17.62) μmol/g vs. (35.58 ± 9.32) μmol/g, (20.14 ± 3.14) μmol/g vs. (10.38 ± 0.85) μmol/g] ( P<0.05). The levels of UP Ⅰ and CP Ⅲ in urine of patients with Dukes C/D stage were significantly higher than those with Dukes A/B stage [(45.26 ± 5.26) μmol/g vs. (28.56 ± 3.45) μmol/g, (86.57 ± 6.58) μmol/g vs. (52.48 ± 3.36) μmol/g], the levels of UP Ⅰand CPⅢ in urine of patients with lymph node metastasis were significantly higher than those without lymph node metastasis [(45.44 ± 5.46) μmol/g vs. (30.27 ± 6.07) μmol/g, (86.67 ± 6.87) μmol/g vs. (56.10 ± 11.08) μmol/g], there were significant differences ( P<0.05). Urinary levels of UPⅠ and CPⅢ were independent risk factors for recurrence and metastasis of colorectal cancer after operation ( OR=1.149 and 1.065, P<0.05). Conclusions:Porphyrin metabolites (UPⅠ and CPⅢ) in urine may serve as a new marker for assessing colorectal cancer.
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Objective@#To study the application of urinary 5-aminolevulinic acid (5-ALA)detection in screening and identification of colorectal cancer and adenomatous polyps.@*Methods@#The clinical data of 500 high-risk patients(including 22 cases with colorectal cancer, 134 cases with adenomatous polyps, and 344 cases with other patients) at the First Affiliated Hospital of Hebei North University from January 2018 to October 2018 were collected. High performance liquid chromatography(HPLC) was used to detect urinary 5-ALA and fecal occult blood test was used to detect faeces. Sensitivity and specificity of two methods was compared. At the same time, urine samples of 431 cases(including 22 cases with colorectal cancer, 134 cases with adenomatous polyps and 275 cases with colorectal normal mucosa)were collected, and the difference of the content of urinary 5-ALA among three groups was compared.@*Results@#The sensitivity of urinary 5-ALA for the colorectal cancer screening was74.9%, and the specificity was 72.5%. The sensitivity of urinary 5-ALA for the adenomatous polyps screening was 70.1%, and the specificity was75.0%. The sensitivity of fecal occult blood test for the colorectal cancer screening was 63.6%, and the specificity was 62.1%. The sensitivity of fecal occult blood test for the adenomatous polyps screening was 42.3%, and the specificity was 62.5%. The content of urinary 5-ALA of the colorectal cancer group [(9.35 ± 0.46) μmol/g] was significantly higher than that of the adenomatous polyps group [(7.24 ± 0.64) μmol/g] (P < 0.05) and normal colorectal mucosa group [(3.12 ± 0.24) μmol/g] (P < 0.05), and the content of urinary 5-ALA of the adenomatous polyps group was significantly higher than that of colorectal normal mucosa group (P < 0.05).@*Conclusions@#For screening of colorectal cancer and adenomatous polyps, the content of urinary 5-ALA by HPLC is better than fecal occult blood test, and this approach can do great help to identify colorectal cancer, adenomatous polyps and normal colorectal mucosa.
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Objective To study the application of urinary 5-aminolevulinic acid (5-ALA) detection in screening and identification of colorectal cancer and adenomatous polyps. Methods The clinical data of 500 high-risk patients(including 22 cases with colorectal cancer, 134 cases with adenomatous polyps, and 344 cases with other patients) at the First Affiliated Hospital of Hebei North University from January 2018 to October 2018 were collected. High performance liquid chromatography (HPLC) was used to detect urinary 5-ALA and fecal occult blood test was used to detect faeces. Sensitivity and specificity of two methods was compared. At the same time, urine samples of 431 cases (including 22 cases with colorectal cancer, 134 cases with adenomatous polyps and 275 cases with colorectal normal mucosa)were collected, and the difference of the content of urinary 5-ALA among three groups was compared. Results The sensitivity of urinary 5-ALA for the colorectal cancer screening was 74.9% , and the specificity was 72.5% . The sensitivity of urinary 5-ALA for the adenomatous polyps screening was 70.1% , and the specificity was75.0% . The sensitivity of fecal occult blood test for the colorectal cancer screening was 63.6% , and the specificity was 62.1% . The sensitivity of fecal occult blood test for the adenomatous polyps screening was 42.3%, and the specificity was 62.5%. The content of urinary 5-ALA of the colorectal cancer group [(9.35 ± 0.46) μmol/g] was significantly higher than that of the adenomatous polyps group [(7.24 ± 0.64) μmol/g] (P<0.05) and normal colorectal mucosa group [(3.12 ± 0.24) μmol/g] (P<0.05), and the content of urinary 5-ALA of the adenomatous polyps group was significantly higher than that of colorectal normal mucosa group (P<0.05). Conclusions For screening of colorectal cancer and adenomatous polyps, the content of urinary 5-ALA by HPLC is better than fecal occult blood test, and this approach can do great help to identify colorectal cancer, adenomatous polyps and normal colorectal mucosa.
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Objective To explore the differential diagnosis of breast ductal carcinoma in situ (DCIS)and breast ductal carcinoma in situ with microinvasion (DCIS-Mi)by ADCMin ,ADCDR and DCE-MRI,and to analyze the correlation between DCIS-Mi and biological factors. Methods Preoperative breast MRI examinations were performed in 41 patients with DCIS-Mi and 3 7 patients with DCIS.DCIS-Mi and DCIS patients were compared in terms of ADCMin ,ADCMax ,ADCDR ,early enhancement rate (EER)and the morphological characteristics of DCE-MRI.The optimal diagnostic variables were determined by binary Logistic regression,the threshold value of the optimal diagnostic variables was ensured by ROC,and the correlation between DCIS-Mi and biological factors was analyzed by Spearman.Results ADCMin of DCIS-Mi patients was lower than that of DCIS (t＝6.294,P＝0.033),and ADCDR was higher than that of DCIS (t＝9.246,P＝0.020).70.7 3% DCIS-Mi showed non-tumor-like enhancement,inclined to segmental distribution,and internal heterogeneous or cluster ring enhancement;29.27% manifested tumor-like enhancement,internal heterogeneous or ring enhancement,and unclear margin.64.86% DCIS showed non-tumor-like enhancement,inclined to linear distribution,internal homogeneous/heterogeneous enhancement;35.14% expressed tumor-like enhancement,internal homogeneous enhancement,and clear margin.The accuracy,sensitivity and specificity of ADCMin , ADCDR ,tumor or non-tumor internal enhancement features in the diagnosis of DCIS-Mi were higher (84.0%,9 5.3%,9 2.4%;89.3%, 9 5.3%,9 2.4%;85.1%,9 2.5%,9 3.8%;87.4%,9 6.8%,84.7%, respectively).ADCMin and ADCDR threshold value were 1.1 1× 10－3 mm2/s and 0.35×10－3 mm2/s,respectively.ADCMin of patients with DCIS-Mi was positive correlation with ER(－)and PR(－), and negative correlation with HER-2(+)(P＜0.05).ADCDR ,non-tumor distribution,and non-tumor internal enhancement characteristics,the tumor edge and internal enhancement characteristics were negative correlation with ER(－)and PR(－),and positive correlation with HER-2 (+)(P＜0.05).Conclusion ADCMin ,ADCDR and DCE-MRI can be used for the differential diagnosis of DCIS-Mi and DCIS, and provided evidence for clinical treatment plan.
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Objective To evaluate the role of Toll-like receptor 4 (TLR4) in cardiomyocyte apoptosis induced by hypoxia-reoxygenation (H/R) in newborn rats.Methods Cardiomyocytes obtained from newborn Sprague-Dawley rats,aged 1-3 days,were primarily cultured in DMEM culture medium supplemented with 20％ fetal bovine serum and divided into 4 groups (n=10 each) using a random number table:control group (group C),H/R group,negative control group (con-siRNA group) and TLR4-siRNA group.Cardiomyocytes were routinely cultured for 24 h in group C.Cardiomyocytes were subjected to hypoxia for 2 h followed by 4 h of reoxygenation in group H/R.Cardiomyocytes were transfected with siRNA and TLR4-siRNA at the final concentration of 80 nmol/L in con-siRNA group and TLR4-siRNA group,respectively.After successful establishment of H/R injury model,the cell survival rate was measured by MTT assay,flow cytometry was used to detect apoptosis in cardiomyocytes,the expression of TLR4 mRNA was determined by real-time polymerase chain reaction,and the expression of TLR4,Bcl-2,Bax and caspase-3 was detected by Western blot.The apoptosis rate was calculated.Results Compared with group C,the cell survival rate was significantly decreased,the expression of TLR4 protein and mRNA,caspase-3 and Bax was up-regulated,the apoptosis rate was increased and the expression of Bcl-2 was down-regulated in the other three groups (P＜0.01).Compared with H/R and con-siRNA groups,the cell survival rate was significantly increased,the expression of TLR4 protein and mRNA,caspase-3 and Bax was down-regulated,the apoptosis rate was decreased and the expression of Bcl-2 was up-regulated in group TLR4-siRNA (P＜0.01).Conclusion TLR4 is involved in cardiomyocyte apoptosis induced by H/R in newborn rats.
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Objective To evaluate the effect of oxycodone on acute lung injury (ALI) induced by lipopolysaecharide (LPS) in rats. Methods Thirty-six pathogen-free healthy male Sprague-Dawley rats, weighing 250-300 g, were divided into 3 groups (n= 12 each) using a random number table: sham opera-tion group (group S), LPS-induced ALI group (group A) and oxycodone group (group O). ALI was in-duced by injecting LPS 8 mg∕kg intravenously in A and O groups, while the equal volume of normal saline was given instead in group S. Oxycodone 2 mg∕kg was injected intravenously at 10 min before LPS injection in group O, while the equal volume of normal saline was given instead in S and A groups. Rats were sacri-ficed at 6 h after LPS injection, and the broncho-alveolar lavage fluid (BALF) was collected for detection of tumor necrosis factor-alpha (TNF-α) and interleukin-1beta (IL-1β) concentrations by enzyme-linked im-munosorbent assay. Pulmonary specimens were obtained for microscopic examination of the pathological changes and for determination of wet∕dry weight ratio (W∕D ratio) and expression of Toll-like receptor 4 (TLR4) in lung tissues (using real-time polymerase chain reaction and Western blot). Results Compared with group S, the TNF-α and IL-1β concentrations in BALF, W∕D ratio, pathological scores and expres-sion of TLR4 were significantly increased at 6 h after LPS injection in A and O groups (P<0. 05). Com-pared with group A, the TNF-α and IL-1β concentrations in BALF, W∕D ratio, pathological scores and expression of TLR4 were significantly decreased at 6 h after LPS injection in group O (P<0. 05). Conclu-sion Oxycodone can attenuate LPS-induced ALI in lung tissues, and the mechanism is related to down-regulating the expression of TLR4 and inhibiting inflammatory responses of rats.
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Objective To evaluate the effects of dexmedetomidine or dezocine alone or the combination of the two agents on the emergence agitation in patients undergoing thoracic surgery.Methods Eighty ASA Ⅰ or Ⅱ patients,aged 18-64 yr,weighing 48-75 kg,scheduled for elective thoracic surgery under general anesthesia,were randomly divided into 4 groups (n =20 each):control group (group C),dexmedetomidine group (group DEX),dezocine group (group DEZ) and dexmedetomidine + dezocine group (group DEX + DEZ).In group DEX,dexmedetomidine 0.5 μg/kg was injected intravenously at 15 min before induction of anesthesia,followed by continuous infusion of dexmedetomidine at 0.4 μg· kg-1 · h-1 until beginning of chest closure.Dezocine 0.1 mg/kg was injected intravenously after beginning of chest closure in group DEZ.In group DEX + DEZ,dexmedetomidine 0.5 μg/kg was injected intravenously at 15 min before induction of anesthesia,followed by continuous infusion of dexmedetomidine at 0.4 μg·kg-1 ·h-1 until beginning of chest closure and then dezocine 0.1 mg/kg was injected intravenously.While the equal volume of normal saline was given instead in group C.Flurbiprofen axetil 50 mg was injected intravenously at beginning of skin closure in each group.Venous blood samples were collected for determination of plasma C-reactive protein (CRP),TNF-α and IL-10 levels at 10 min before induction of anesthesia (T1),before skin closure (T2),immediately after extubation (T3) and 15 min after extubation (T4).Side effects such as agitation during emergence from anesthesia were recorded.Sedation was assessed using Ramsay score.Results Compared with group C,the levels of plasma CRP and TNF-α at T2-4 and ratio of TNF-α/IL-10 were significantly decreased,the levels of IL-10 were increased at T2-4,the degree and incidence of agitation were decreased,and Ramsay score was increased in the other three groups (P ＜ 0.05).Compared with groups DEX and DEZ,the levels of plasma CRP and TNF-α at T2_4 and ratio of TNF-α/IL-10 were significantly decreased,the levels of IL-10 were increased at T2-4,and the degree and incidence of agitation were decreased in group DEX + DEZ (P ＜0.05).No side effects such as hypotension,bradycardia,respiratory depression,nausea and vomiting were observed in the four groups.Conclusion Dexmedetomidine or dezocine alone or combination of the two agents can decrease the degree and occurrence of emergence agitation and inhibit the inflammatory response simultaneously,and the combination of the two agents provides better efficacy than either alone in patients undergoing thoracic surgery.
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Objective To evaluate the effects of heme oxygenase-1 (HO-1) mediated by cell penetrating peptide PEP-1 on liverinjury induced by intestinal ischemia/reperfusion (I/R) in rats.Methods Twenty-four male Sprague-Dawley rats,aged 7-9 weeks,weighing 210-260 g,were randomly divided into 3 groups (n =8 each):sham operation group (group S),intestinal I/R group (group I/R) and PEP-1/HO-1 group (group HO).To establish a model of intestinal I/R,intestines were exteriorized and the superior mesenteric artery was exposed and occluded for 45 min ischemia,and then the clamp was removed for 120 min reperfusion.The PEP-1/HO-1 fusion protein 0.5 mg was injectedvia ihe left iliac vein 30 min prior to ischemia in group HO.The superior mesenteric artery was only exposed but not occluded in group S.At the end of reperfusion,blood samples were collected from the right common carotid artery for measurement of serum aspartate aminotransferase (AST),alanine aminotransferase (ALT) activities.The rats were then sacrificed and livers were removed for microscopic examination and for determination of malondialdehyde (MDA) content and superoxide dismutase (SOD) activity in livertissues.Results Compared with group S,serum AST and ALT activities and MDA content in liver tissues were significantly increased,while SOD activity in liver tissues was decreased in groups I/R and HO (P ＜ 0.05).Compared with group I/R,serum AST and ALT activities and MDA content in liver tissues were significantly decreased,while SOD activity in liver tissues was increased in group HO (P ＜0.05).Liver injury induced by intestinal I/R was significantly attenuated in group HO compared with group I/R (P ＜ 0.05).Conciusioon HO-1 protein mediated by cell penetrating peptide PEP-1 can attenuate liver injury induced by intestinalI/R in rats.
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Objective To evaluate the effect of dexmedetomidine pretreatment on lipopolysaccharide (LPS)-induced release of inflammatory mediators in primary microglias in neonatal rats.Methods Purified primary microglias were seeded in the plate and randomly divided into 3 groups with 10 holes in each group:control group (group C),LPS group (group L) and dexmedetomidine pretreatment group (group D).In group C,the cells were incubated in serum-free DMEM for 24 h.In group LPS,the cells were incubated with LPS (final concentration 1 μg/ml) for 24 h.In group D,the cells were pretreated with dexmedetomidine (final concentration 1 ng/ml) for 1 h,then LPS (final concentration 1 μg/ml) was added and the cells were incubated for 24 h.The levels of nitric oxide (NO) (by Greiss method) and prostaglandin E2 (PGE2),IL-1β and TNF-α (by ELISA) in the supernatant were measured after 24 h of incubation.The expression of inducible nitric oxide synthase (iNOS) mRNA in the cells was determined by RT-PCR.Results Compared with group C,the levels of NO,PGE2,IL-1β and TNF-α,and expression of iNOS mRNA were significantly increased in groups L and D (P ＜ 0.05).There was no significant difference in the indexes mentioned above between groups D and L (P ＞ 0.05).Conclusion Pretreatment with dexmedetomidine has no significant effect on LPS-induced release of inflammatory mediators in primary microglias in neonatal rats.
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Objective To evaluate the regulatory role of acetylcholine receptor in muramyl dipeptide (MDP)-induced activation of Nod-like receptor 2/receptor-interacting protein 2 (2NLR2/RIP2) pathway in macrophages of mice.Methods RAW264.7 cells at the logarithmic growth phase were seeded in 12-well plates (density 1 × 106 cells/ml,2 ml/well),a total of 108 wells.The cells were randomly divided into 3 groups (n =36 each) using a random number table:control group (group C),MDP group (group M),and GTS-21 (a7nAChR specific agonist) group (group G).The cells were routinely cultured in group C.MDP with the final concentration of 10 μg/ml was added to the culture medium in group M.MDP with the final concentration of 10μg/ml and GTS21 with the final concentration of 50 μg/ml were added to the culture medium in group G.The cells were incubated for 24 h.At 1,6 and 24 h of incubation with MDP,12 wells were chosen and the cell suspension was obtained for measurement of NLR2 mRNA expression (by real-time fluorescent quantitative PCR),RIP2 expression (by Western blot),and concentrations of tumor necrosis factor-alpha (TNF-α) and high mobility group box-1 (HMGB1) in the culture media (by ELISA).Results Compared with group C,the levels of NLR2 mRNA,RIP2,TNFα and HMGB1 were significantly increased at each time point in group M (P ＜ 0.05).Compared with group M,the levels of NLR2 mRNA,RIP2,TNF-α and HMGB1 were significantly decreased at each time point in group G (P ＜ 0.05).Conclusion Acetylcholine receptor can suppress MDP-induced transduction of NLR2/RIP2 pathway in macrophages of mice.
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Objective To evaluate the effects of postconditioning with propofol on lipopolysaccharide (LPS)-induced inflammatory responses of microglial cells in rat brain tissues.Methods The primary cultured microglial cells in brain tissues of Sprague-Dawley rats were seeded in 24 multi-well plates at a density of 1 × 105 cells/ml,and the microglial cells of 100 wells were randomly divided into 5 groups (n =20 each) using a random number table:control group (group C),LPS group (group L),and propofol 25,50 and 100 μmol/L groups (P25,P50,P100 groups).The cells were cultured routinely in group C.LPS 1 μg/ml was added and the cells were incubated for 24 h in group L.In P25,P20,and P100 groups,when the cells were incubated for 24 h with LPS 1 μg/ml,propofol with the final concentrations of 25,50 and 100 μmol/L was added,respectively.The cells were collected at 1 h of incubation with propofol for determination of the expression of inducible nitric oxide synthase (iNOS) mRNA,cyclooxygenase-2 (COX-2) mRNA,tumor necrosis factor-α (TNF-α) mRNA and interleukin-1β (IL-1β) mRNA (by RT-PCR).The supematant was separated for determination of the concentrations of nitric oxide (NO) (by Griess method) and pmstaglandin E2 (PGE2),TNF-α and IL-1β (by ELISA).Results Compared with group C,the expression of iNOS mRNA,COX-2 mRNA,TNF-α mRNA and IL-1β mRNA was significantly up-regulated and the concentrations of NO,PGE2,TNF-α and IL-1β in the supematant were increased in group L (P ＜ 0.01).Compared with group L,the expression of iNOS mRNA,COX-2 mRNA,TNF-α mRNA and IL-1β mRNA was significantly down-regulated and the concentrations of NO,PGE2,TNF-α and IL-1β in the supernatant were decreased in P50 and P100 groups (P ＜ 0.05 or 0.01),while no significant change in the indexes mentioned above was found in P25 group (P ＞ 0.05).Conclusion Postconditioning with propofol 50 and 100 μmol/L can inhibit LPS-induced inflammatory responses of microglial cells in rat brain tissues.
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Objective To investigate the changes in expression of hippocampal neuronal gap function protein connexin 36 (Cx36) induced by postoperative cognitive dysfunction (POCD) in aged rats.Methods Ninety male SD rats aged 20 months weighing 500-600 g were randomly divided into 3 groups ( n =30 each):group normal control (group C); group sham operation (group S) and group POCD.POCD was induced by splenectomy.Cognitive function was assessed by using open field test and maze test on the 1st,3rd and 7th day after operation (T1,T2,T3 ).The ultrastructure of gap junction in the CA1 region of hippocampas was examined with thin-section transmission electron microscope.The expression of Cx36 was detected by immuno-histochemical method.Results POCD significantly decreased the number of grid cross,the rearing and correct responses and increased the time the animab spent in the central square and total reaction time at T1 in group POCD as compared with group C.Cx36 expression was significantly decreased at T1 in group POCD as compared with group C.The ultrastructure of gap junctions underwent significant change at T1 in group POCD.Conclusion Hippocampal neuronal Cx36 may be involved in the cognitive dysfunction after splenectomy in aged rats.
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Objective To investigate the effects of repeated intraperitoneal dexmedetomidine on the cognitive function in rats with chronic cerebral ischemia.Methods Forty-eight male Sprague-Dawley rats,aged 3-4months,weighing 250-300 g,were randomly divided into 4 groups (n =12 each) ∶ sham operation group (group S),chronic cerebral ischemia group (group IS),dexmedetomidine treatment 1 group (group DXM1) and dexmedetomidine treatment 2 group (group DXM2).Dexmedetomidine 5 μg/kg was injected intraperitoneally at 30 min before occlusion of bilateral common carotid arteries and 3,12,24 and 48 h after occlusion in group DXM1,and at 3,12,24 and 48 h after occlusion in group DXM2.The cognitive function was assessed by Morris water maze 2 weeks after occlusion.The apoptosis was examined by TUNEL.The expression of Bcl-2 protein in hippocampus was detected by Western blot.Results Compared with group S,the escape latency was significantly prolonged from 2nd day to 5th day after the place navigation test in group IS and on 2nd day after Morris water maze test in groups DXM1 and DXM2,and the time of staying in 1 st quadrant was significantly shortened,the apoptotic rate was increased,and the expression of Bcl-2 was up-regulated in groups IS,DXM1 and DXM2 (P ＜ 0.05).Compared with group IS,the escape latency was significantly shortened from 3rd day to 5th day after the place navigation
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Objective To investigate the effects of penehyclidine hydrochloride ( PHC) on lipopolysaccharide (LPS) -induced endothelial cells injury and its mechanism. Methods ECV-304 was cultured in RPMI1640 in a 5% humidified CO2 atmosphere at 37 ℃. Then cultured cells were used to assess the following treatments: control group, LPS group (1 μg/mL) and PHC group(2 μg/mL). At the end of the experiments, supernatant was collected for determination of lactate dehydrogenase ( LDH), and cells were collected for determination of malondialdehyde (MDA), superoxide dismutase (SOD), and nitric oxide (NO) levels. And extracellular regulated kinasel/2( ERKl/2)and JNK MAPK (mitogen-activated protein kinases, MAPK) protein expressions were determined using Western blot technique. Analysis of variance (ANOVA) was used for statistical analysis to compare values among all groups. A significant difference was presumed for a probability value < 0.05. Results Compared with control group, LDH leakage [(1642 ± 367) U/L vs (169±33)U/L], the contents of MDA[(13. 2 ± 1. 2) nmol/L vs (7. 2 ±0. 8)nmol/mL] and NO levels [(143.2 ± 10.3) μmol/L vs(85.5 ±4.1) μmol/L], expressions of ERK1/2 and JNK were remarkably increased and SOD activities[(41.2 ±2.7) U/mL vs (61. 1 ±2.8) U/mL] were obviously decreased in LPS group. PHC markedly decreased LDH leakage [(392 ±90) U/L], MDA contents [(8. 6 ± 1. 3) nmol/ mL] and NO levels [(92.1 ±6.6) μmol/L], ERK1/2 activation and enhanced SOD activities [(58.0± 3.0) U/mL]. Conclusions PHC could protect endothelial cells against LPS-induced cell injury. The effect of PHC is likely mediated through inhibition of ERK1/2 MAPK activation.
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Objective To investigate the effect of penehyclidine hydrochloride (PHCD) on the reinstatement of conditioned place preference (CPP) in morphine dependent rats. Methods Forty male adult SD rats weighing 180-220 g were randomly divided into 5 groups (n = 8 each): group control (group C); group morphine (group M) and 3 PHCD groups (group P1-3 ). Morphine 10 mg/kg was injected subcutaneously once a day for 8 days to induce morphine CPP. The rats were then subjected to extinction of CPP for 10 days with normal saline (NS) instead of morphine. After the extinction, the rats were put into the drug-paired side of the box. A single priming dose of morphine 4 mg/kg was injected to reinstate the morphine CPP. In group P1-3 the rats received PHCD 0.5, 1.0 and 1.5 mg/kg intraperitoneally 30 min prior to priming dose of morphine, whereas in group C and M the rats received NS. The second day the rats underwent CPP test. Results Compared with group M, the time spent in the drug-paired side (grey area) was significantly shortened in group P1-3 (P ＜ 0.05 or 0.01 ).Compared with group P1 ,no significant change in the time spent in the drug-paired side (grey area) was found in group P2(P ＞ 0.05), but the time spent in the drug-paired side (grey area) was significantly shortened in group P3 ( P ＜ 0.05). Conclusion PHCD could significantly inhibit the reinstatement of CPP induced by priming dose of morphine in morphine dependent rats and it is related to the dose.
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Objective To investigate the effect of electro-acupuncture at zusanli on Severe thermal injury-induced acute lung injury in rats.Methods Forty male SD rats weighing 200-250 g were used in this study.Thirty percent of the total body surface (TBS) was shaved chemically with 20% sodium sulfate and then exposed to 99-100℃ water for 12 s.The animals with third degree thermal injury involving 30% TBS were randomly divided into 5 groups(n=8 each):group Ⅰ control(group C);groupⅡ thermal injury;group Ⅲ electro-acupuncture at zusanli;group Ⅳ electric stimulation of non-acupoint and group Ⅴ electro-acupuncture at zusanli+α-bungarotoxin α-BGT).In group Ⅲ,Ⅳ,and Ⅴ electro-stimulation(3 v,2 ms,3 Hz) of zusanli or non-acupoint was performed for 12 min immediately after thermal injury model was established and every 8 h.hung specimens were obtained at 48 h after thermal injury for microscopic examination.The pulmonary HMGB-l protein level was measured by ELISA.The expression of HMGB-1 mRNA and protein in the lung was determined by RT-PCR and immuno-histochemistry respectively.Results Thermal injury induced leucocytosis in the interstitial capillaries,interstitial edema,intra-alveolar fibrin deposit,blebbing of type Ⅱ alveolar lining cells and decrease in lamellar body.Both expression of HMGB-1 mRNA and protein in the lung was significantly enhanced at 48 h after thermal injury.Electrical stimulation of zusanli significantly down-regalated the expression of HMGB-1 mRNA and protein in the lung.However,α-BGT pretreatment reversed the effects of electrical stimulation of zusanli.Conclusion Electrical stimulation of zusanli could significantly ameliorate severe thermal injury-induced acute lung injury through inhibition of HMGB-1 mRNA and protein expression and activation of cholinergic anti-inflammatory pathway mediated by nicotinic acetylcholine receptor α7 subunit.
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Objective To investigate the effect of PEP-1-heme oxygenase-1 (PEP-1-HO-1) fusion protein transduction on hypoxia-reoxygenation (H/R) injury in rat H9c2 cells. Methods After construction of the prokaryotic expression plasmid pET15b-PEP-1-hHO-1 containing the human heme oxygenase-1 gene, it was then transformed to make PEP-1-HO-1 fusion protein express. The H9c2 cells were cultured in high-glucose Dulbecco's modified Eagle's medium (DMEM) supplemented with 15% fetal bovine serum and randomly divided into 4 groups (n = 4 each): control group (group C), H/R group, low-concentration fusion protein group (group L-HO), and high-concentration fusion protein group (group H-HO). The cells were exposed to 22 h of hypoxia followed by 8 h of reoxygenation. PEP-1-HO-1 fusion protein was added to the culture medium with a final concentration of 1.0 μ mol/L (group L-HO) or 2.0 μmol/L (group H-HO) before hypoxia. The cells and supernatant of the culture medium were collected after reoxygenation to determine the activity of lactate dehydrogenase (LDH) in the supernatant and the content of malondialdehyde (MDA) and activity of superoxide dismutase (SOD) in the cells. Results The SOD activity was significantly lower, while the MDA content and LDH activity were significantly higher in group H/R, L-HO and H-HO than in group C (P ＜0.05). The SOD activity was significantly higher, while MDA content and LDH activity were significantly lower in group L-HO and H-HO than in group H/R, and in group H-HO than in group L-HO ( P ＜ 0.05). Conclusion PEP-1-HO-1 fusion protein transdution can protect H9c2 cells against H/R injury in rats.