目的建立免疫富集联合基质辅助激光解吸电离飞行时间质谱法(matrix-assisted laser desorption/ionization time of flight mass spectrometry,MALDI-TOF MS)检测牛奶、鸡蛋中沙门氏菌的方法。方法沙门氏菌经免疫磁珠特异性富集后采用MA...目的建立免疫富集联合基质辅助激光解吸电离飞行时间质谱法(matrix-assisted laser desorption/ionization time of flight mass spectrometry,MALDI-TOF MS)检测牛奶、鸡蛋中沙门氏菌的方法。方法沙门氏菌经免疫磁珠特异性富集后采用MALDI-TOF MS鉴定,并优化富集过程中的抗体、免疫磁珠添加量以及捕获时间,研究其特异性,并用于实际样品中沙门氏菌的检测。结果富集过程最佳反应条件为:每1 mg磁珠中抗体添加量为250μg,1 mL菌样中免疫磁珠添加量为600μL,捕获时间为30 min,免疫磁珠捕获沙门氏菌具有特异性;用于检测牛奶和鸡蛋中的沙门氏菌时,将浓度为3 CFU/mL的沙门氏菌特异性富集后选择性增菌9h即可被MALDI-TOF MS准确鉴定。结论该方法具有选择性好,特异性高,耗时短,结果准确的优点,为食品中沙门氏菌的检测提供相关参考。展开更多
OBJECTIVE To investigate possible protective effect of berberine,an isoquinoline alkaloid,is the major active constituent of Rhizoma coptidis and Cortex phellodendri,on high-fat diet/streptozotocin-induced early diabe...OBJECTIVE To investigate possible protective effect of berberine,an isoquinoline alkaloid,is the major active constituent of Rhizoma coptidis and Cortex phellodendri,on high-fat diet/streptozotocin-induced early diabetic nephropathy in rats and various mechanisms underlie this effect.METHODS The diabetic rat model was generated by a single intraperitoneal injection of streptozotocin(STZ,50 mg·kg-1).Diabetic rats were randomlyassigned into the following five groups:control,DN,losartan(30 mg·kg-1·d-1),berberine(100,200 mg·kg-1·d-1).Berberine and losartan were given intragastricly for nine weeks.At the end of the experiment,urine of each group was collected in a 24 h period.Rats were weighed and then sacrificed.Plasma and kidneys were collected.The levels of blood glucose,creatinine(Cr),triglycerides(TG),total cholesterol(TCH)and malondialdehyde(MDA)in serum were determined using commercial kits according to the manufacturer′s instructions.Transforming growth factor(TGF)-β1and intracellular adhesion molecule-1(IAM-1)mR NA levels were evaluated by RT-PCR.The renal histopathology was observed by light microscopy.Further biochemical analysis of IKKβ1 and p65(nucleus/cytoplasm)was provided using Western blotting techniques.RESULTS Our study has demonstrated that berberine has various pharmacological activities.The DN rats had significantly higher kidney/body weight ratio(17.4±1.4)mg·g-1,and berberine treatment could reduce this ratio change 13.6±0.6 and(11.6±0.8)mg·g-1,respectively;glucose control still remains the only disease-modifying therapy for diabetic complications,FBG was also recorded in the experiment.The findings reveal that the DN group showed a significantly higher glucose level(28.67±2.78)mmol·L-1.Treatment with8 weeks of berberine improved these parameters except blood glucose〔(18.67±2.59)mmol·L-1and 16.45±1.80 vs(28.67±2.78)mmol·L-1:plasma levels of urea nitrogen(15.67±2.48)mmol·L-1and 14.45±2.40 vs(12.26±2.40)mmol·L-1〕;plasma levels of 24 h urine albuminuria〔30.48±1.56 and 25.72±2.24 vs(15.26±0.12)μg·d-1〕;based on these results,berberine supposed to improve renal functions in diabetic rats.Berberine also ameliorated the inflammatory changes of DN in diabetic animals;levels of TG,TCH and MDA in berberine-treatment rats were significantly lower compared with those in the DN group:TG〔2.78±0.24 and 2.45±0.36 vs(5.20±0.60)mmol·L-1〕;TCH〔4.26±0.46 and 3.74±0.68 vs(6.26±0.50)mmol·L-1〕;MDA〔4.94±1.19 and 4.28±0.64 vs(4.28±0.64)nu·mL-1〕.Chronic inflammation,as is observed in diabetes,is associated with increased production of TGF-β1and IAM-1.Compared with the renal tissues of DN group,TGF-β1and IAM-1 gene expressions in berberine treated groups were reduced at the dose levels(100 and 200 mg·kg-1).And TGF-β1and IAM-1levels in berberine treated groups were reduced in a dosedependent manner:Relative expression of TGF-β1mR NA level(3.56±0.28 and 3.12±0.14 vs 5.12±0.44);Relative expression of IAM-1 mR NA leve(l1.78±0.56 and 1.42±0.24vs 4.36±0.35).Research finds that the NF-κB signaling pathway is activated in the renal tissue of diabetic mice and berberine inhibits activation of the NF-κB pathway:staining score for IKKβ1(4.34±0.26 and 3.82±0.24 vs 6.23±0.76),staining score for p65(2.34±0.26 and 1.74±0.78 vs 6.23±0.24)in nucleus and staining score for p65(7.21±0.13 and8.15±0.45 vs 4.23±0.54)in cytoplasm.CONCLUSION In this field,berberine suppresses the increased expression of p65 in the nucleus and decreases it in cytoplasm,which leads to the inhibition of the NF-κB pathway.These changes will result in decreasing the transcription and translation of many inflammatory mediators,such as TGF-β1and IAM-1.Additionally,these changes decrease the number of inflammatory cells and mononuclear macrophage infiltration into glomeruli and renal interstitium.These results indicated that berberine can protect the kidney of STZ-diabetic rats by reducing the expression of TGF-β1and IAM-1 in the renal tubulointerstitium.And we propose that berberinemayfunction as an effective therapeutic agent for diabetic nephropathy and attenuate the progression of renal injury.展开更多
文摘目的建立免疫富集联合基质辅助激光解吸电离飞行时间质谱法(matrix-assisted laser desorption/ionization time of flight mass spectrometry,MALDI-TOF MS)检测牛奶、鸡蛋中沙门氏菌的方法。方法沙门氏菌经免疫磁珠特异性富集后采用MALDI-TOF MS鉴定,并优化富集过程中的抗体、免疫磁珠添加量以及捕获时间,研究其特异性,并用于实际样品中沙门氏菌的检测。结果富集过程最佳反应条件为:每1 mg磁珠中抗体添加量为250μg,1 mL菌样中免疫磁珠添加量为600μL,捕获时间为30 min,免疫磁珠捕获沙门氏菌具有特异性;用于检测牛奶和鸡蛋中的沙门氏菌时,将浓度为3 CFU/mL的沙门氏菌特异性富集后选择性增菌9h即可被MALDI-TOF MS准确鉴定。结论该方法具有选择性好,特异性高,耗时短,结果准确的优点,为食品中沙门氏菌的检测提供相关参考。
基金The project supported by Fundamental Research Funds for the Central Universities(21615463)
文摘OBJECTIVE To investigate possible protective effect of berberine,an isoquinoline alkaloid,is the major active constituent of Rhizoma coptidis and Cortex phellodendri,on high-fat diet/streptozotocin-induced early diabetic nephropathy in rats and various mechanisms underlie this effect.METHODS The diabetic rat model was generated by a single intraperitoneal injection of streptozotocin(STZ,50 mg·kg-1).Diabetic rats were randomlyassigned into the following five groups:control,DN,losartan(30 mg·kg-1·d-1),berberine(100,200 mg·kg-1·d-1).Berberine and losartan were given intragastricly for nine weeks.At the end of the experiment,urine of each group was collected in a 24 h period.Rats were weighed and then sacrificed.Plasma and kidneys were collected.The levels of blood glucose,creatinine(Cr),triglycerides(TG),total cholesterol(TCH)and malondialdehyde(MDA)in serum were determined using commercial kits according to the manufacturer′s instructions.Transforming growth factor(TGF)-β1and intracellular adhesion molecule-1(IAM-1)mR NA levels were evaluated by RT-PCR.The renal histopathology was observed by light microscopy.Further biochemical analysis of IKKβ1 and p65(nucleus/cytoplasm)was provided using Western blotting techniques.RESULTS Our study has demonstrated that berberine has various pharmacological activities.The DN rats had significantly higher kidney/body weight ratio(17.4±1.4)mg·g-1,and berberine treatment could reduce this ratio change 13.6±0.6 and(11.6±0.8)mg·g-1,respectively;glucose control still remains the only disease-modifying therapy for diabetic complications,FBG was also recorded in the experiment.The findings reveal that the DN group showed a significantly higher glucose level(28.67±2.78)mmol·L-1.Treatment with8 weeks of berberine improved these parameters except blood glucose〔(18.67±2.59)mmol·L-1and 16.45±1.80 vs(28.67±2.78)mmol·L-1:plasma levels of urea nitrogen(15.67±2.48)mmol·L-1and 14.45±2.40 vs(12.26±2.40)mmol·L-1〕;plasma levels of 24 h urine albuminuria〔30.48±1.56 and 25.72±2.24 vs(15.26±0.12)μg·d-1〕;based on these results,berberine supposed to improve renal functions in diabetic rats.Berberine also ameliorated the inflammatory changes of DN in diabetic animals;levels of TG,TCH and MDA in berberine-treatment rats were significantly lower compared with those in the DN group:TG〔2.78±0.24 and 2.45±0.36 vs(5.20±0.60)mmol·L-1〕;TCH〔4.26±0.46 and 3.74±0.68 vs(6.26±0.50)mmol·L-1〕;MDA〔4.94±1.19 and 4.28±0.64 vs(4.28±0.64)nu·mL-1〕.Chronic inflammation,as is observed in diabetes,is associated with increased production of TGF-β1and IAM-1.Compared with the renal tissues of DN group,TGF-β1and IAM-1 gene expressions in berberine treated groups were reduced at the dose levels(100 and 200 mg·kg-1).And TGF-β1and IAM-1levels in berberine treated groups were reduced in a dosedependent manner:Relative expression of TGF-β1mR NA level(3.56±0.28 and 3.12±0.14 vs 5.12±0.44);Relative expression of IAM-1 mR NA leve(l1.78±0.56 and 1.42±0.24vs 4.36±0.35).Research finds that the NF-κB signaling pathway is activated in the renal tissue of diabetic mice and berberine inhibits activation of the NF-κB pathway:staining score for IKKβ1(4.34±0.26 and 3.82±0.24 vs 6.23±0.76),staining score for p65(2.34±0.26 and 1.74±0.78 vs 6.23±0.24)in nucleus and staining score for p65(7.21±0.13 and8.15±0.45 vs 4.23±0.54)in cytoplasm.CONCLUSION In this field,berberine suppresses the increased expression of p65 in the nucleus and decreases it in cytoplasm,which leads to the inhibition of the NF-κB pathway.These changes will result in decreasing the transcription and translation of many inflammatory mediators,such as TGF-β1and IAM-1.Additionally,these changes decrease the number of inflammatory cells and mononuclear macrophage infiltration into glomeruli and renal interstitium.These results indicated that berberine can protect the kidney of STZ-diabetic rats by reducing the expression of TGF-β1and IAM-1 in the renal tubulointerstitium.And we propose that berberinemayfunction as an effective therapeutic agent for diabetic nephropathy and attenuate the progression of renal injury.
