Thirteen extracting solutions of rare-earth metallofullerenes containing La,Ce,Pr,Nd Sm,Eu,Gd,Tb,Dy,Ho,Er,Tm and Yb respectively have been investigated by means of matrix-assisted laser desorpuon/ ionization time-of-f...Thirteen extracting solutions of rare-earth metallofullerenes containing La,Ce,Pr,Nd Sm,Eu,Gd,Tb,Dy,Ho,Er,Tm and Yb respectively have been investigated by means of matrix-assisted laser desorpuon/ ionization time-of-flight mass spectrometry.The influences of the positive-ion/negative-ion mode,laser intensity,ma trix and mass discrimination to the analytical results are studied,based on which the optimal analytical conditions have been determined.The results show that the extracting solutions contain large quantities of rare-earth metallofullerenes besides empty fullerenes.On the basis of comparing their relative intensities,the different structure stabilities and solubilities of metallofullerenes with different rare-earth metals encapsulated into the fullerene cages,as well as some possible reasons to those differences,are discussed.展开更多
Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone(THAP)and 2,5-dihydroxybenzoic acid(DHB)as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass...Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone(THAP)and 2,5-dihydroxybenzoic acid(DHB)as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer(MALDI-TOF MS).High quality negative-ion spectra of commercial sialylated glycan were obtained with THAP as matrix.Detection limit of the glycan was less than 0.1 pmol.After methyl esterification of sialic acid(SA)residue,sialylated glycans were detected sensitively in the positive-ion mode using DHB as matrix.Neutral and sialylated glycans from the mixture of asialofetuin and fetuin were methylesterified and simultaneously recognized in one manipulation.Methyl esterification of SA residue offers a convenient and sensitive way to identify the structure of N-linked glycans for glycan profiling.展开更多
Matrix-assisted Laser Desorption/Ionization with Time-of-flight Mass Spectrometry (MALDI-TOFMS) was investigated as a method for the rapid identifica-tion of species. Current demand in microbial identi-fication is how...Matrix-assisted Laser Desorption/Ionization with Time-of-flight Mass Spectrometry (MALDI-TOFMS) was investigated as a method for the rapid identifica-tion of species. Current demand in microbial identi-fication is how to compare unknown strains to the known one quickly, semi-automatically and accurately. In this paper, we present a software tool that allows flexibly microbial matching in a user-friendly way, by letting the users to customize comparison parameters including: in vitro transcription enzyme, mass tolerance,minimum fragment length, intensity threshold and corresponding weights. We provide three spectral scoring functions to compute the affin-ity between the species. Therefore, the precision of microbial comparison increases. To test and verify this tool, we employed experimental spectral data based on MALDI-TOFMS and the gene sequences of E.coli and Salmonella. This software is written in Java for cross-platform intention.展开更多
BACKGROUND Burkholderia gladioli(B.gladioli)is regarded as a rare opportunistic pathogen.Only a few patients with abscesses caused by B.gladioli infections have been reported,and these are usually abscesses at the inc...BACKGROUND Burkholderia gladioli(B.gladioli)is regarded as a rare opportunistic pathogen.Only a few patients with abscesses caused by B.gladioli infections have been reported,and these are usually abscesses at the incision caused by traumatic surgery.CASE SUMMARY A 74-year-old male patient with abscesses and pain throughout his body for 1 mo was admitted to our hospital.Some of the abscesses had ruptured with purulent secretions on admission.Color Doppler ultrasound examination of the body surface masses showed mixed masses 75 mm×19 mm,58 mm×17 mm,17 mm×7 mm,and 33 mm×17 mm in size in the muscle tissues of both the right and left forearms,the posterior area of the right knee and the left leg,respectively.Abscess secretions and blood cultures grew B.gladioli.The following 3 methods were used to jointly identify the bacterium:an automatic microbial identification system,matrix-assisted laser desorption/ionization time-of-flight mass spectrometry,and full-length 16 S rDNA sequencing.After 27 d of treatment with meropenem,etimicin,trimethoprim-sulfamethoxazole and other antibiotics,most of his skin abscesses were flat and he was discharged without any symptoms.CONCLUSION This is the first reported case of multiple skin abscesses associated with bacteremia caused by B.gladioli.Our study provides important reference values for the clinical diagnosis and treatment of B.gladioli infections.展开更多
Aim: To identify proteins induced by androgen in Sertoli cells during spermatogenesis. Methods: We analyzed protein profiles in TM4 Sertoli cells treated with dihydrotestosterone (DHT) using surface enhanced laser...Aim: To identify proteins induced by androgen in Sertoli cells during spermatogenesis. Methods: We analyzed protein profiles in TM4 Sertoli cells treated with dihydrotestosterone (DHT) using surface enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Results: We found increases in the expression of a 5.0-kDa protein at 15 min, an 11.3-kDa protein at 24 h and 4.3 kDa, 5.7 kDa, 5.8 kDa, 9.95 kDa and 9.98 kDa proteins at 48 h after the treatment. In contrast, the expression of 6.3 kDa and 8.6 kDa proteins decreased at 30 min, and 4.9 kDa, 5.0 kDa, 12.4 kDa and 19.8 kDa proteins at 48 h after the treatment. The ll.3-kDa protein was identified as macrophage migration inhibitory factor (MIF) known to having various functions. The 9.98-kDa protein was identified as calgizzarin related to calcium channels. The timing of their expression suggests that MIF and calgizzarin are involved in late regulation of spermatogenesis in Sertoli cells by androgen. Conclusion: MIF and calgizzarin are two important androgen-responsive proteins produced by Sertoli cells and they might play a role in regulating spermatogenesis.展开更多
20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus req...20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus required.Herein,a matrixassisted laser desorption/ionization time-of-f ight tandem mass spectrometry(MALDI-TOF/TOF MS)method is described for rapid and sensitive determination of endogenous 20E in plants.It is based on the use of the(3-(acridin-9-ylamino)phenyl)boronic acid(AYPBA)as the mass tag to assist the MS and tandem MS(MS^(n))analysis of 20E on MALDI-TOF/TOF MS.Good linearity was obtained with a determination coef cient(R^(2))larger than 0.99 in the range of 0.025–2.5μΜ.The limit of detection(LOD)was 2.4 fmol.Acceptable precision and accuracy were gained by intra-and inter-day analysis with relative standard deviations less than 19.5%and relative recoveries ranging from 85.7 to 105.2%.In addition,the AYPBA labeled 20E produced abundant characteristic fragment ions under the high energy collision-induced dissociation,which facilitated the identif cation of 20E by MS^(2)analysis on MALDI-TOF/TOF MS.Using the method,we enabled the identif cation and quantif cation of endogenous 20E in four herbs including Cyanotis arachoidea,Achyranthes bidentata,Spinacia oleracea and Chenopodium quinoa willd.,demonstrating the feasibility of the proposed method for screening of the endogenous 20E in plants.展开更多
BACKGROUND:It has been pointed out that only low-dose arsenic trioxide(ATO)presents therapeutic benefits outweighing the toxic side effects.Low-dose ATO can effectively alleviate acute promyelocytic leukemia(APL). How...BACKGROUND:It has been pointed out that only low-dose arsenic trioxide(ATO)presents therapeutic benefits outweighing the toxic side effects.Low-dose ATO can effectively alleviate acute promyelocytic leukemia(APL). However,it is quite challenging in treating solid tumors. The purpose of this study was to investigate the effect of ATO at low concentrations on the metastatic potential of mouse hepatoma H22 cells and the anti-metastatic mechanism of ATO. METHODS:The metastatic potential of H22 cells was evaluated by adhesion,migration and invasion assays after exposure to a low dose of ATO in vitro.The mouse lung metastatic model induced by injection of H22 cells via the tail vein was adopted for the evaluation of metastatic potential. Different proteins in the lysate of H22 cells exposed to ATO at different concentrations were investigated by surface- enhanced laser desorption and ionization time-of-flight mass spectrometry(SELDI-TOF-MS).Finally,Western blotting analyses were made to detect the expression pattern of MMP-2 and nm23-M1 proteins. RESULTS:Significant cell death started at ATO concentrations above 2μmol/L.The growth and adhesion potential of H22 cells was inhibited in a time-and dose- dependent manner,and the migration and invasion potential of H22 cells was inhibited in a dose-dependent manner while ATO concentration was below 2μmol/L. Mice injected with ATO at a dose of 0.5 mg/kg had fewer lung metastases.However,mice injected with ATO at a dose of 2 mg/kg or 4 mg/kg had a high mortality rate and more liver injuries.A total of 15 different protein peaks were identified between the lysate of H22 cells treated with ATO and controls.Two proteins that peaked atm/z 5302 and 17207 coincided with MMP-2(fragment) and nm23-M1,respectively.Western blotting analyses demonstrated that MMP-2 and MMP-2 fragments were down-regulated and nm23-M1 was up-regulated in H22 cells treated with 2μmol/L ATO for 48 hours. CONCLUSIONS:ATO at a low dose inhibits the metastatic potential of mouse hepatoma H22 cells in vitro and in vivo, and involves down-regulation of MMP-2 and up-regulation of nm23-M1.展开更多
The radial basis function networks were applied to bacterial classification based on the matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOF-MS) data. The classification of bacteri...The radial basis function networks were applied to bacterial classification based on the matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOF-MS) data. The classification of bacteria cultured at different time was discussed and the effect of the network parameters on the classification was investigated. The cross-validation method was used to test the trained networks. The correctness of the classification of different bacteria investigated changes in a wide range from 61.5% to 92.8%. Owing to the complexity of biological effects in bacterial growth, the more rigid control of bacterial culture conditions seems to be a critical factor for improving the rate of correctness for bacterial classification.展开更多
Tsukamurella species are obligate aerobic,gram-positive,weak acid-fast,nonmotile bacilli.They are found in various environments,such as soil,water,sludge,and petroleum reservoir wastewater,and belong to the order Acti...Tsukamurella species are obligate aerobic,gram-positive,weak acid-fast,nonmotile bacilli.They are found in various environments,such as soil,water,sludge,and petroleum reservoir wastewater,and belong to the order Actinomycetales.In 2016,there was a reclassification of species within the genus Tsukamurella,merging the species Tsukamurella tyrosinosolvens(T.tyrosinosolvens)and Tsukamurella carboxydivorans.Tsukamurella species are clinically considered to be a rare opportunistic pathogen,because most reported cases have been related to bacteremia and intravascular prosthetic devices and immunosuppression.To date,it has been isolated only from human specimens,and has always been associated with clinical disease;human infections are very rare.Reported infections have included pneumonia,brain abscesses,catheter-related bloodstream infections,ocular infections,bacteremia,and sepsis presenting with septic pulmonary emboli in patients who are immunocompromised.To date,there is no commercially available test for identification.On the other hand,sequence-based identification,including matrix-assisted laser desorption ionization time-of-flight mass spectrometry,is an alternative method for identifying clinical isolates that are either slow growers or difficult to identify through biochemical profiling.The golden standards for diagnosis and optimal management still remain to be determined.However,newer molecular biological techniques can provide accurate identification,and contribute to the appropriate selection of definitive therapy for infections caused by this organism.Combinations of several antimicrobial agents have been proposed for treatment,though the length of treatment for infections has yet to be determined,and should be individualized according to clinical response.Immunocompromised patients often experience severe cases due to infection,and life-threatening T.tyrosinosolvens events associated with dissemination and/or failure of source control have occurred.Favorable prognoses can be achieved through earlier identification of the cause of infection,as well as successful management,including appropriate antibiotic therapy together with source control.Further analyses of similar cases are required to establish the most adequate diagnostic methods and treatment regimens for infections.展开更多
Background:We investigated possible biomarkers for endometriosis (EM) using the ClinProt technique and proteomics methods.Methods:We enrolled 50 patients with EM,34 with benign ovarian neoplasms and 40 healthy vol...Background:We investigated possible biomarkers for endometriosis (EM) using the ClinProt technique and proteomics methods.Methods:We enrolled 50 patients with EM,34 with benign ovarian neoplasms and 40 healthy volunteers in this study.Serum proteomic spectra were generated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MS) combined with weak cationic exchange (WCX) magnetic beads.Possible biomarkers were analyzed by a random and repeat pattern model-validation method that we designed,and ClinProtools software,results were refined using online liquid chromatography-tandem MS.Results:We found a cluster of 5 peptides (4210,5264,2660,5635,and 5904 Da),using 3 peptides (4210,5904,2660 Da) to discriminate EM patients from healthy volunteers,with 96.67% sensitivity and 100% specificity.We selected 4210 and 5904 m/z,which differed most between patients with EM and controls,and identified them as fragments of ATP1B4,and the fibrinogen alpha (FGA) isoform 1/2 of the FGA chain precursor,respectively.Conclusions:ClinProt can identify EM biomarkers,which-most notably-distinguish even early-stage or minimal disease.We found 5 stable peaks at 4210,5264,2660,5635,and 5904 Da as potential EM biomarkers,the strongest of which were associated with ATP1B4 (4210 Da) and FGA (5904 Da); this indicates that ATP1B4 and FGA are associated with EM pathogenesis.展开更多
Antarctic ice microalga can survive and thrive in cold channels or pores in the Antarctic ice layer. In order to understand the adaptive mechanisms to low temperature, in the present study we compared two-dimensional ...Antarctic ice microalga can survive and thrive in cold channels or pores in the Antarctic ice layer. In order to understand the adaptive mechanisms to low temperature, in the present study we compared two-dimensional polyacrylamide gel electrophoresis (2-DE) profiles of normal and low temperature-stressed Antarctic ice microalga Chlamydomonas sp. cells. In addition, new protein spots induced by low temperature were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database searching. Well-resolved and reproducible 2-DE patterns of both normal and low temperature-stressed cells were acquired. A total of 626 spots was detected in control cells and 652 spots were detected in the corresponding low temperature-stressed cells. A total of 598 spots was matched between normal and stressed cells. Two newly synthesized proteins (a and b) in low temperature-stressed cells were characterized. Protein spot A (53 kDa, pl 6.0) was similar to isopropylmalate/homocitrate/citramalate synthases, which act in the transport and metabolism of amino acids. Protein spot b (25 kDa, pl 8.0) was related to glutathione S-transferase, which functions as a scavenger of active oxygen, free radicals, and noxious metabolites. The present study is valuable for the application of ice microalgae, establishing an ice microalga Chlamydomonas sp. proteome database, and screening molecular biomarkers for further studies.展开更多
Background Renal biopsy is necessary for diagnosing the pathological changes of primary nephrotic syndrome (NS). However, it is invasive, time-consuming and can not be performed frequent on the same patient. Thus, d...Background Renal biopsy is necessary for diagnosing the pathological changes of primary nephrotic syndrome (NS). However, it is invasive, time-consuming and can not be performed frequent on the same patient. Thus, development of a non-invasive and rapid diagnostic method may improve clinical patient management. Methods Proteomic tool magnetic bead-based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MB-based MALDI TOF MS) was applied to serum to determine peptidome patterns that are characteristic of different pathological changes. Results Serum specimen from 114 patients with NS (62 were minimal change disease (MCD), 30 were membranous nephropathy (MN), and 22 were focal segmental glomerulosclerosis (FSGS)) and 60 normal individuals were analyzed using MB-based MALDI TOF MS. The peptidome pattern was generated by genetic algorithms using a training set of 31 MCD, 15 MN, 11 FSGS and 30 normal individuals and was validated by an independent testing set of the remaining samples. The serum peptidome pattern, based on a panel of 14 peaks, accurately recognized samples from MCD, MN, FSGS and healthy control with sensitivities of 93.5%, 86.7%, 63.6% and 90.0%, and specificities of 98.2%, 94.4%, 100% and 89.5%, respectively. Moreover, one peptide from peptidome pattern was identified by liquid chromatography tandem mass spectrometry (LC MS/MS) as fibrinogen A. Conclusion Detection of the serum peptidome pattern is a rapid, non-invasive, high-throughout, and reproducible method for identifying the pathological patterns of patients with nephrotic syndrome.展开更多
Proteomic analysis was applied to generating the map of Arabidopsis mature pollen proteins and analyzing the differentially expressed proteins that are potentially involved in the regulation of Arabidopsis pollen germ...Proteomic analysis was applied to generating the map of Arabidopsis mature pollen proteins and analyzing the differentially expressed proteins that are potentially involved in the regulation of Arabidopsis pollen germination. By applying 2-D electrophoresis and silver staining, we resolved 499 and 494 protein spots from protein samples extracted from pollen grains and pollen tubes, respectively. Using the matrix-assisted laser desorption ionization time-of-flight mass spectrometry method, we identified 189 distinct proteins from 213 protein spots expressed in mature pollen or pollen tubes, and 75 new identified proteins that had not been reported before in research into the Arabidopsis pollen proteome. Comparative analysis revealed that 40 protein spots exhibit reproducible significant changes between mature pollen and pollen tubes. And 21 proteins from 17 downregulated and six upregulated protein spots were identified. Functional category analysis indicated that these differentially expressed proteins mainly involved in signaling, cellular structure, transport, defense/stress responses, transcription, metabolism, and energy production. The patterns of changes at protein level suggested the important roles for energy metabolism-related proteins in pollen tube growth, accompanied by the activation of the stress response pathway and modifications to the cell wall.展开更多
Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and re-versed-phase high performance liquid chromatography. Tw...Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and re-versed-phase high performance liquid chromatography. Two uremic middle molecular fractions (A and B) were obtained from uremic sera and urine and normal urine by gel permeation chromatography, but not from normal sera. The anion exchange chromatographic results of fraction A from different origins demonstrate that subfraction A-3 could be excreted in urine by healthy subject, but accumulated in uremic serum for renal failure of patient with uremia. After desalinization subfraction A-3 was analyzed by MALDI-TOF-MS. The results show that subfraction A-3 consists of six compounds with molecular weight 839, 873, 1007.94, 1106, 1680 and 2015 respectively. Finally, by reversed-phase high performance liquid chromatography, subfraction A-3 was further resolved into six independent fractions. Thus, the isolation and purification of six middle molecular展开更多
The silica-based poly(dimethylsiloxane) (PDMS) microfluidic enzymatic reactor was reported along with its analytical features in coupling with MALDI TOF and ESI MS. Microfluidic chip was fabricated using PDMS cast...The silica-based poly(dimethylsiloxane) (PDMS) microfluidic enzymatic reactor was reported along with its analytical features in coupling with MALDI TOF and ESI MS. Microfluidic chip was fabricated using PDMS casting and O2-plasma techniques, and used for the preparation of enzymatic reactor. Plasma oxidation for PDMS enabled the channel wall of microfluidics to present a layer of silanol (SiOH) groups. These SiOH groups as anchors onto the microchannel wall were linked covalently with the hydroxy groups of trypsin-encapsulated sol matrix. As a result, the leakage of sol-gel matrix from the microchannel was effectively prevented. On-line protein analysis was performed with the microfluidic enzymatic reactor by attachment of stainless steel tubing electrode and replaceable tip, The success of trypsin encapsulation was investigated by capillary electrophoresis (CE) detection, and MALDI TOF and ESI MS analysis. The lab-made device provided excellent extent of digestion even at the fast flow rate of 7.0 μL/min with very short residence time of ca. 2 s. In addition, the encapsulated trypsin exhibits increased stability even after continuous use. These features are the most requisite for high-throughput protein identification.展开更多
基金Project supported by the National Natural Science Foundation of China
文摘Thirteen extracting solutions of rare-earth metallofullerenes containing La,Ce,Pr,Nd Sm,Eu,Gd,Tb,Dy,Ho,Er,Tm and Yb respectively have been investigated by means of matrix-assisted laser desorpuon/ ionization time-of-flight mass spectrometry.The influences of the positive-ion/negative-ion mode,laser intensity,ma trix and mass discrimination to the analytical results are studied,based on which the optimal analytical conditions have been determined.The results show that the extracting solutions contain large quantities of rare-earth metallofullerenes besides empty fullerenes.On the basis of comparing their relative intensities,the different structure stabilities and solubilities of metallofullerenes with different rare-earth metals encapsulated into the fullerene cages,as well as some possible reasons to those differences,are discussed.
基金Supported by the National Natural Science Foundation of China(30800193)Grant from Centre for International Mobility(CIMO),Finland
文摘Native and methyl-esterified sialylated glycans were analyzed with 2,4,6-trihydroxyacetophenone(THAP)and 2,5-dihydroxybenzoic acid(DHB)as matrix by a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer(MALDI-TOF MS).High quality negative-ion spectra of commercial sialylated glycan were obtained with THAP as matrix.Detection limit of the glycan was less than 0.1 pmol.After methyl esterification of sialic acid(SA)residue,sialylated glycans were detected sensitively in the positive-ion mode using DHB as matrix.Neutral and sialylated glycans from the mixture of asialofetuin and fetuin were methylesterified and simultaneously recognized in one manipulation.Methyl esterification of SA residue offers a convenient and sensitive way to identify the structure of N-linked glycans for glycan profiling.
文摘Matrix-assisted Laser Desorption/Ionization with Time-of-flight Mass Spectrometry (MALDI-TOFMS) was investigated as a method for the rapid identifica-tion of species. Current demand in microbial identi-fication is how to compare unknown strains to the known one quickly, semi-automatically and accurately. In this paper, we present a software tool that allows flexibly microbial matching in a user-friendly way, by letting the users to customize comparison parameters including: in vitro transcription enzyme, mass tolerance,minimum fragment length, intensity threshold and corresponding weights. We provide three spectral scoring functions to compute the affin-ity between the species. Therefore, the precision of microbial comparison increases. To test and verify this tool, we employed experimental spectral data based on MALDI-TOFMS and the gene sequences of E.coli and Salmonella. This software is written in Java for cross-platform intention.
基金Supported by Jilin Science and Technology Development Program,No.20190304110YY
文摘BACKGROUND Burkholderia gladioli(B.gladioli)is regarded as a rare opportunistic pathogen.Only a few patients with abscesses caused by B.gladioli infections have been reported,and these are usually abscesses at the incision caused by traumatic surgery.CASE SUMMARY A 74-year-old male patient with abscesses and pain throughout his body for 1 mo was admitted to our hospital.Some of the abscesses had ruptured with purulent secretions on admission.Color Doppler ultrasound examination of the body surface masses showed mixed masses 75 mm×19 mm,58 mm×17 mm,17 mm×7 mm,and 33 mm×17 mm in size in the muscle tissues of both the right and left forearms,the posterior area of the right knee and the left leg,respectively.Abscess secretions and blood cultures grew B.gladioli.The following 3 methods were used to jointly identify the bacterium:an automatic microbial identification system,matrix-assisted laser desorption/ionization time-of-flight mass spectrometry,and full-length 16 S rDNA sequencing.After 27 d of treatment with meropenem,etimicin,trimethoprim-sulfamethoxazole and other antibiotics,most of his skin abscesses were flat and he was discharged without any symptoms.CONCLUSION This is the first reported case of multiple skin abscesses associated with bacteremia caused by B.gladioli.Our study provides important reference values for the clinical diagnosis and treatment of B.gladioli infections.
文摘Aim: To identify proteins induced by androgen in Sertoli cells during spermatogenesis. Methods: We analyzed protein profiles in TM4 Sertoli cells treated with dihydrotestosterone (DHT) using surface enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Results: We found increases in the expression of a 5.0-kDa protein at 15 min, an 11.3-kDa protein at 24 h and 4.3 kDa, 5.7 kDa, 5.8 kDa, 9.95 kDa and 9.98 kDa proteins at 48 h after the treatment. In contrast, the expression of 6.3 kDa and 8.6 kDa proteins decreased at 30 min, and 4.9 kDa, 5.0 kDa, 12.4 kDa and 19.8 kDa proteins at 48 h after the treatment. The ll.3-kDa protein was identified as macrophage migration inhibitory factor (MIF) known to having various functions. The 9.98-kDa protein was identified as calgizzarin related to calcium channels. The timing of their expression suggests that MIF and calgizzarin are involved in late regulation of spermatogenesis in Sertoli cells by androgen. Conclusion: MIF and calgizzarin are two important androgen-responsive proteins produced by Sertoli cells and they might play a role in regulating spermatogenesis.
基金supported by the National Key R&D Program of China(2017YFC0906800)the National Natural Science Foundation of China(21635006,31670373 and 21721005)
文摘20-Hydroxyecdysone(20E)derived from plants has a wide range of physiological and pharmacological ef ects on animals and humans,and rapid and sensitive methods for screening of the endogenous 20E in plants are thus required.Herein,a matrixassisted laser desorption/ionization time-of-f ight tandem mass spectrometry(MALDI-TOF/TOF MS)method is described for rapid and sensitive determination of endogenous 20E in plants.It is based on the use of the(3-(acridin-9-ylamino)phenyl)boronic acid(AYPBA)as the mass tag to assist the MS and tandem MS(MS^(n))analysis of 20E on MALDI-TOF/TOF MS.Good linearity was obtained with a determination coef cient(R^(2))larger than 0.99 in the range of 0.025–2.5μΜ.The limit of detection(LOD)was 2.4 fmol.Acceptable precision and accuracy were gained by intra-and inter-day analysis with relative standard deviations less than 19.5%and relative recoveries ranging from 85.7 to 105.2%.In addition,the AYPBA labeled 20E produced abundant characteristic fragment ions under the high energy collision-induced dissociation,which facilitated the identif cation of 20E by MS^(2)analysis on MALDI-TOF/TOF MS.Using the method,we enabled the identif cation and quantif cation of endogenous 20E in four herbs including Cyanotis arachoidea,Achyranthes bidentata,Spinacia oleracea and Chenopodium quinoa willd.,demonstrating the feasibility of the proposed method for screening of the endogenous 20E in plants.
文摘BACKGROUND:It has been pointed out that only low-dose arsenic trioxide(ATO)presents therapeutic benefits outweighing the toxic side effects.Low-dose ATO can effectively alleviate acute promyelocytic leukemia(APL). However,it is quite challenging in treating solid tumors. The purpose of this study was to investigate the effect of ATO at low concentrations on the metastatic potential of mouse hepatoma H22 cells and the anti-metastatic mechanism of ATO. METHODS:The metastatic potential of H22 cells was evaluated by adhesion,migration and invasion assays after exposure to a low dose of ATO in vitro.The mouse lung metastatic model induced by injection of H22 cells via the tail vein was adopted for the evaluation of metastatic potential. Different proteins in the lysate of H22 cells exposed to ATO at different concentrations were investigated by surface- enhanced laser desorption and ionization time-of-flight mass spectrometry(SELDI-TOF-MS).Finally,Western blotting analyses were made to detect the expression pattern of MMP-2 and nm23-M1 proteins. RESULTS:Significant cell death started at ATO concentrations above 2μmol/L.The growth and adhesion potential of H22 cells was inhibited in a time-and dose- dependent manner,and the migration and invasion potential of H22 cells was inhibited in a dose-dependent manner while ATO concentration was below 2μmol/L. Mice injected with ATO at a dose of 0.5 mg/kg had fewer lung metastases.However,mice injected with ATO at a dose of 2 mg/kg or 4 mg/kg had a high mortality rate and more liver injuries.A total of 15 different protein peaks were identified between the lysate of H22 cells treated with ATO and controls.Two proteins that peaked atm/z 5302 and 17207 coincided with MMP-2(fragment) and nm23-M1,respectively.Western blotting analyses demonstrated that MMP-2 and MMP-2 fragments were down-regulated and nm23-M1 was up-regulated in H22 cells treated with 2μmol/L ATO for 48 hours. CONCLUSIONS:ATO at a low dose inhibits the metastatic potential of mouse hepatoma H22 cells in vitro and in vivo, and involves down-regulation of MMP-2 and up-regulation of nm23-M1.
基金Supported by Foundation for Young Mainstay TeachersEducation Ministry of China.
文摘The radial basis function networks were applied to bacterial classification based on the matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOF-MS) data. The classification of bacteria cultured at different time was discussed and the effect of the network parameters on the classification was investigated. The cross-validation method was used to test the trained networks. The correctness of the classification of different bacteria investigated changes in a wide range from 61.5% to 92.8%. Owing to the complexity of biological effects in bacterial growth, the more rigid control of bacterial culture conditions seems to be a critical factor for improving the rate of correctness for bacterial classification.
文摘Tsukamurella species are obligate aerobic,gram-positive,weak acid-fast,nonmotile bacilli.They are found in various environments,such as soil,water,sludge,and petroleum reservoir wastewater,and belong to the order Actinomycetales.In 2016,there was a reclassification of species within the genus Tsukamurella,merging the species Tsukamurella tyrosinosolvens(T.tyrosinosolvens)and Tsukamurella carboxydivorans.Tsukamurella species are clinically considered to be a rare opportunistic pathogen,because most reported cases have been related to bacteremia and intravascular prosthetic devices and immunosuppression.To date,it has been isolated only from human specimens,and has always been associated with clinical disease;human infections are very rare.Reported infections have included pneumonia,brain abscesses,catheter-related bloodstream infections,ocular infections,bacteremia,and sepsis presenting with septic pulmonary emboli in patients who are immunocompromised.To date,there is no commercially available test for identification.On the other hand,sequence-based identification,including matrix-assisted laser desorption ionization time-of-flight mass spectrometry,is an alternative method for identifying clinical isolates that are either slow growers or difficult to identify through biochemical profiling.The golden standards for diagnosis and optimal management still remain to be determined.However,newer molecular biological techniques can provide accurate identification,and contribute to the appropriate selection of definitive therapy for infections caused by this organism.Combinations of several antimicrobial agents have been proposed for treatment,though the length of treatment for infections has yet to be determined,and should be individualized according to clinical response.Immunocompromised patients often experience severe cases due to infection,and life-threatening T.tyrosinosolvens events associated with dissemination and/or failure of source control have occurred.Favorable prognoses can be achieved through earlier identification of the cause of infection,as well as successful management,including appropriate antibiotic therapy together with source control.Further analyses of similar cases are required to establish the most adequate diagnostic methods and treatment regimens for infections.
基金grants from the National Natural Science Foundation of China,supported by Peking University People's Hospital Research and Development Funds
文摘Background:We investigated possible biomarkers for endometriosis (EM) using the ClinProt technique and proteomics methods.Methods:We enrolled 50 patients with EM,34 with benign ovarian neoplasms and 40 healthy volunteers in this study.Serum proteomic spectra were generated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MS) combined with weak cationic exchange (WCX) magnetic beads.Possible biomarkers were analyzed by a random and repeat pattern model-validation method that we designed,and ClinProtools software,results were refined using online liquid chromatography-tandem MS.Results:We found a cluster of 5 peptides (4210,5264,2660,5635,and 5904 Da),using 3 peptides (4210,5904,2660 Da) to discriminate EM patients from healthy volunteers,with 96.67% sensitivity and 100% specificity.We selected 4210 and 5904 m/z,which differed most between patients with EM and controls,and identified them as fragments of ATP1B4,and the fibrinogen alpha (FGA) isoform 1/2 of the FGA chain precursor,respectively.Conclusions:ClinProt can identify EM biomarkers,which-most notably-distinguish even early-stage or minimal disease.We found 5 stable peaks at 4210,5264,2660,5635,and 5904 Da as potential EM biomarkers,the strongest of which were associated with ATP1B4 (4210 Da) and FGA (5904 Da); this indicates that ATP1B4 and FGA are associated with EM pathogenesis.
基金Supported by the National Natural Science Foundation of China(40406003)
文摘Antarctic ice microalga can survive and thrive in cold channels or pores in the Antarctic ice layer. In order to understand the adaptive mechanisms to low temperature, in the present study we compared two-dimensional polyacrylamide gel electrophoresis (2-DE) profiles of normal and low temperature-stressed Antarctic ice microalga Chlamydomonas sp. cells. In addition, new protein spots induced by low temperature were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database searching. Well-resolved and reproducible 2-DE patterns of both normal and low temperature-stressed cells were acquired. A total of 626 spots was detected in control cells and 652 spots were detected in the corresponding low temperature-stressed cells. A total of 598 spots was matched between normal and stressed cells. Two newly synthesized proteins (a and b) in low temperature-stressed cells were characterized. Protein spot A (53 kDa, pl 6.0) was similar to isopropylmalate/homocitrate/citramalate synthases, which act in the transport and metabolism of amino acids. Protein spot b (25 kDa, pl 8.0) was related to glutathione S-transferase, which functions as a scavenger of active oxygen, free radicals, and noxious metabolites. The present study is valuable for the application of ice microalgae, establishing an ice microalga Chlamydomonas sp. proteome database, and screening molecular biomarkers for further studies.
基金This work was supported by gran'ts from the National Science Fund for Distinguished Young Scholars of China (No. 30925019), the National Natural Science Foundation of China (No. 30871166), and the Key Clinical Research Program, Ministry of Health, China (2007).
文摘Background Renal biopsy is necessary for diagnosing the pathological changes of primary nephrotic syndrome (NS). However, it is invasive, time-consuming and can not be performed frequent on the same patient. Thus, development of a non-invasive and rapid diagnostic method may improve clinical patient management. Methods Proteomic tool magnetic bead-based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MB-based MALDI TOF MS) was applied to serum to determine peptidome patterns that are characteristic of different pathological changes. Results Serum specimen from 114 patients with NS (62 were minimal change disease (MCD), 30 were membranous nephropathy (MN), and 22 were focal segmental glomerulosclerosis (FSGS)) and 60 normal individuals were analyzed using MB-based MALDI TOF MS. The peptidome pattern was generated by genetic algorithms using a training set of 31 MCD, 15 MN, 11 FSGS and 30 normal individuals and was validated by an independent testing set of the remaining samples. The serum peptidome pattern, based on a panel of 14 peaks, accurately recognized samples from MCD, MN, FSGS and healthy control with sensitivities of 93.5%, 86.7%, 63.6% and 90.0%, and specificities of 98.2%, 94.4%, 100% and 89.5%, respectively. Moreover, one peptide from peptidome pattern was identified by liquid chromatography tandem mass spectrometry (LC MS/MS) as fibrinogen A. Conclusion Detection of the serum peptidome pattern is a rapid, non-invasive, high-throughout, and reproducible method for identifying the pathological patterns of patients with nephrotic syndrome.
基金Supported by a competitive research grant (30421002) for Creative Research Groups sponsored by the National Natural Science Foundation of China
文摘Proteomic analysis was applied to generating the map of Arabidopsis mature pollen proteins and analyzing the differentially expressed proteins that are potentially involved in the regulation of Arabidopsis pollen germination. By applying 2-D electrophoresis and silver staining, we resolved 499 and 494 protein spots from protein samples extracted from pollen grains and pollen tubes, respectively. Using the matrix-assisted laser desorption ionization time-of-flight mass spectrometry method, we identified 189 distinct proteins from 213 protein spots expressed in mature pollen or pollen tubes, and 75 new identified proteins that had not been reported before in research into the Arabidopsis pollen proteome. Comparative analysis revealed that 40 protein spots exhibit reproducible significant changes between mature pollen and pollen tubes. And 21 proteins from 17 downregulated and six upregulated protein spots were identified. Functional category analysis indicated that these differentially expressed proteins mainly involved in signaling, cellular structure, transport, defense/stress responses, transcription, metabolism, and energy production. The patterns of changes at protein level suggested the important roles for energy metabolism-related proteins in pollen tube growth, accompanied by the activation of the stress response pathway and modifications to the cell wall.
基金This work was supported by the State Key Fundamental R & D Project (Grant No. G1999064707)the National Natural Science Foundation of China (Grant No. 59873011) Trans-Century Training Program Foundation for the Talents by the Ministry of Education,
文摘Isolation and comparison of uremic sera and urine and normal sera and urine were performed by gel permeation chromatography, anion exchange chromatography and re-versed-phase high performance liquid chromatography. Two uremic middle molecular fractions (A and B) were obtained from uremic sera and urine and normal urine by gel permeation chromatography, but not from normal sera. The anion exchange chromatographic results of fraction A from different origins demonstrate that subfraction A-3 could be excreted in urine by healthy subject, but accumulated in uremic serum for renal failure of patient with uremia. After desalinization subfraction A-3 was analyzed by MALDI-TOF-MS. The results show that subfraction A-3 consists of six compounds with molecular weight 839, 873, 1007.94, 1106, 1680 and 2015 respectively. Finally, by reversed-phase high performance liquid chromatography, subfraction A-3 was further resolved into six independent fractions. Thus, the isolation and purification of six middle molecular
基金Project supported by the National Natural Science Foundation of China (Nos. 30572227, 20299030), Shanghai Science Research Foundation (No 03JC14005), National Basic Research Priority Program (No. 001 CB510202) and National High Technology Key Project (No. 2002BAC11A11).
文摘The silica-based poly(dimethylsiloxane) (PDMS) microfluidic enzymatic reactor was reported along with its analytical features in coupling with MALDI TOF and ESI MS. Microfluidic chip was fabricated using PDMS casting and O2-plasma techniques, and used for the preparation of enzymatic reactor. Plasma oxidation for PDMS enabled the channel wall of microfluidics to present a layer of silanol (SiOH) groups. These SiOH groups as anchors onto the microchannel wall were linked covalently with the hydroxy groups of trypsin-encapsulated sol matrix. As a result, the leakage of sol-gel matrix from the microchannel was effectively prevented. On-line protein analysis was performed with the microfluidic enzymatic reactor by attachment of stainless steel tubing electrode and replaceable tip, The success of trypsin encapsulation was investigated by capillary electrophoresis (CE) detection, and MALDI TOF and ESI MS analysis. The lab-made device provided excellent extent of digestion even at the fast flow rate of 7.0 μL/min with very short residence time of ca. 2 s. In addition, the encapsulated trypsin exhibits increased stability even after continuous use. These features are the most requisite for high-throughput protein identification.