Objective:To optimize the major parameters of bioinformatics analysis conditions in process of PMF(Peptide Mass Fingerprinting) identify target proteins. Methods:In this process of experiment design and analysis,we made the Bovine Carbonic anhydrase-2 and BSA from 2-DE gel,after proteolysis with porcine trypsin and gathered the peptides,subsequently identified with MALDI-TOF MS and got the peptides mass data. Then,we selected database of Swissprot and MS-Fit search engine,the standard Carbonic anhydrase-2 protein used as model to set up protein search standard conditions. Results:Our study showed that the cysteine was modified by carbamidomethylation more convenient in protein search,at the same time,the better mode of mass tolerance was percentage and 0.1% was suitable tolerance in our search. Then we used BSA mass data to check the accuracy of MS search standard conditions. Conclusions:Results show that the optimized parameters are reliable.

Objective To study if ganglioside (GLS) may promote the recovery the neurological deficits after cerebral ischemia in rats and the me chanism about it.Method Focal cerebral ischemia was made by a 3－ hour occlusion of the right middle cerebral artery(MCA)using nylon monofi1ament in rats and was fo1lowed by reperfusion.GLS was intraperitoneally administered 30 minutes and 3 hours after ischemia and daily during the observation period. Bederson's method was used to evaluate the neurological deficits. The activity of the neurotrophic factors (NTFs) in the periischemic brain tissue extract (BTE) and the effect of GLS on the activity were observed in cultured neuronsfrom the newbom rat cortex and basal ganglia.Result GLS improved the neurologicalfUnction recovery after cerebral ischemia. The BTE of GLS treatment groap supportedneuronal survival and promoted neurite outgrowth about l00 μ m. The number of neuronswas significantly higher in the presence of the BTE from treatment grouP as compared with the BTE from control group (P<0.05).Conclusion it indicated that GLS improved the recovery of the nevrological deficits by potentiating the activety of nevro trophic factors.

BACKGROUND: Flash visual evoked potential(fVEP) can reflect the integrity of visual pathway from retina to pulvinar cortex. The utilization of its characters can accurately evaluate the injury situation of visual conduction pathway.OBJECTIVE: To investigate the feasibility and effects of fVEP on non-traumatic monitoring of intracranial pressure.DESIGN: A repeated measurement design based on patients.PARTICIPANTS: Totally 197 patients with brain injury including 138males and 59 females with an average of(38 ±9) years old were selected from the First Affiliated Hospital of Chongqing Medical University and Xuanwu Hospital Affiliated to Capital University of Medical Sciences. The selected diseases were: epidural hematoma( n=20),subdural hematoma(n = 26),cerebral contusion( n = 4),subdural hematoma companied with cerebral contusion( n = 92),and cerebral contusion companied with intracerebral hematoma( n = 55).METHODS: The level of intracranial pressure of 197 patients with brain trauma was detected with fVEP and epidural manometry. Data of traumatic surveillance were collected after the completion of fVEP. Changes of blood pressure and heart rate in patients were recorded simultaneously.MAIN OUTCOME MEASURES: fVEP results,and levels of intracranial pressure.RESULTS: The average intracranial pressure of 197 patients with brain trauma was(2.75 ± 0.64) kPa,( 1.54 - 4. 02 kPa) . The dispersion of both traumatic and non-traumatic surveillance was relatively big in patients with confirmed visual pathway injury. No typical wave was induced by fVEP in2.79% of patients. Scatter diagram suggested that the consistence between fVEP and epidural manometry was relatively good. Linear regression analysis showed that heart rate was the most sensitive one to the fluctuation of intracranial pressure (β = -0. 369) followed by systolic pressure (β= 0. 316),while diastolic pressure was not so sensitive (β = 0. 147). There was negative correlation between heart rate and intracranial pressure,which could quite sensitively reflect the fluctuation of intracranial pressure,systolic pressure was the next,and diastolic pressure was not easily to be affected by the changes of intracranial pressure.CONCLUSION: Non-traumatic surveillance,fVEP,has favorable consistence with traumatic detection,which can quantitatively evaluate the changes of intracranial pressure non-traumatically.

Proteomic technology has been widely used in the research of differently-expressed proteins under different physiology and pathologic conditions.In the aspect of the human spermatozoa proteomics,many reports have demonstrated the differently expressed proteins and the roles of sperm under different physiology and pathologic state from proteome level,and these results provided reliable theoretic base for well understanding of sperm molecule mechanism in physiology and pathologic conditions.

Objective To investigate the epidemiology of BDV infection in Yili horses and Yili donkeys and to analyze phylogenetic source of BDV in Yili area, Xinjiang. Methods We established fluo- rescence quantitative nested RT-PCR to detect BDV p24 segment in peripheral blood mononuclear cells (PBMCs) of 518 Yili horses and 206 Yili donkeys in Yili area, Xinjiang. Positive products were validated by detecting BDV p40 segment and plasmid to preclude the contamination, and were sequenced to analyze the homology of gene sequence, amino acid sequence and phylogenetic tree. Results The positive rates of BDV infection in PBMCs of 518 Yili horses and 206 Yili donkeys were 0.97% and 1.94%, respectively. The results of BDV p40 segment verification were positive in all of the samples of BDV p24 positive. All the samples tested were not contaminated by plasmid. There was a homology of the gene sequence of positive PCR samples with strain He/80. And the gene sequence revealed more than 93% identical to H1766 and strain V. Conclusion Our study suggested BDV natural infection in Yili horses and Yili donkeys. The en- demic BDV had a high degree of identity to strain He/80.

Objective To observe the epidemiology characterization of Borna disease virus (BDV) in animal brain in Ili, Xinjiang, and to find out the potential infection of the Borna disease virus to prevent its outbreak. Methods The BDV p24 gene of animal brain tissues in Ili including 200 horses, 75 donkeys and 100 shepherd dogs was detected by fluorescence quantitative nest reverse transcriptase polymer-ase chain reaction(FQ-nRT-PCR). GFP-p24,pMD-19 plasmid contamination was excluded from positive products. Clone sequencing was used to analyze the homology of gene and amino acid sequence. Results BDV p24 gent was found in 3 Ili horses, 4 Ili donkeys and 9 shepherd dogs, and the positive ratio is 1.5%, 5.3% and 9.0%, respectively. The GFP-p24,pMD-19 were not found in BDV p40 gene and plasmid stand-ard. The sequence of BDV p24 amplification production was totally the same as He/80 virus strain. Conclu-sion Natural infection of BDV may exist in the animals(horses, donkeys and dogs)in Ili, and the epidem-ic strain of BDV in this area was homological as He/80 virus strain.

Objective To observe the short-term,long-term clinical results and complications in refractory juvenile idiopathic arthritis (JIA) treating with TNF-α inhibitors,and to compare 2 evaluation systems.Methods A retrospective review of 52 cases of patients with refractory JIA in Shenzhen Children's Hospital was performed.With reference to International Leagne of Associations for Rheumatology(ILAR)2001 diagnostic criteria,the patients were divided into 4 groups:26 polyarticular JIA patients,14 systemic JIA patients,9 oligoarticular JIA patients and 3 other types of patients.The children with JIA were based on the conventional treatment such as Methotrexate,combination of TNF-α inhibitors treatment.The short-term and long-term clinical outcomes were evaluated and compared with American College of Rheumatology (ACR) and Juvenile Arthritis Disease Activity Score (JADAS).Complications in each group were recorded.Results (1) Short-term outcome assessment:ACR 50 were achieved in 69.2％ of the polyarticular JIA,66.7％ in oligoarticular and 35.7％ in systemic JIA patients on the third month;and by the time of the sixth month it reached to 73.0％ in polyarticular JIA,77.7％ in oligoarticular JIA and 14.3％ in systemic JIA patients on the sixth month.Significant improvement of JADAS after the treatment was observed in each type of JIA,and the differences were statistically significant(all P ＜ 0.05).(2) Long-term outcome assessment:except for the cases missing follow-ups and withdrawal cases,46 patients were able to complete 2 years assessments:according to ACR,effective rate was 84.0％ in polyarticular JIA (21/25 cases),50.0％ in oligoarticular JIA (4/8 cases) and 7.7％ in systemic (1/13 cases) JIA patients;JADAS was significantly decreased in polyarticular JIA patients (76.0％,19/25cases) (P ＜ 0.05),while significant improvement was not observed in oligoarticular JIA and systemic JIA patients(P ＞ 0.05).(3) Complications of upper respiratory tract infection (23.0％,12 cases) and local reaction in injection site (7.6％,4 cases) were noticed.Higher risks of tuberculosis infection and malignancy were not observed.Conclusions (1) TNF-α inhibitors treatment showed a better short-term and long-term outcome in polyarticular and oligoarticular JIA patients,and it may also improve short-term outcome in systemic JIA but with poorer long-term outcome.(2)Two evaluated systems (ACR and JADAS) were well relative,but ACR was capable to compare clinical course between different types of JIA.(3) TNF-α inhibitors treatment was relatively safe with unremarkable adverse reactions.

Objective:To evaluate the effect of sleep deprivation on cognitive function in septic rats and its relationship with neuronal glycolysis isoenzyme phosphofructokinase-2/fructose-2,?6-diphosphatase 3 (PFKFB3).Methods:Fifty-six healthy male Sprague-Dawley (SD) rats were randomly divided into 4 groups ( n = 14): control group (Con group), sepsis group (LPS group), sepsis+sleep deprivation group (LPS+SD group), sepsis+sleep deprivation+glycolysis inhibitor 3-PO treatment group (LPS+SD+3-PO group). The sepsis model was established by intraperitoneal injection of lipopolysaccharide (LPS) 10 mg/kg. Rats in LPS+SD group were treated with sleep deprivation using a sleep deprivation instrument 24 hours after LPS injection. The LPS+SD+3-PO group was intraperitoneally injected with LPS for 24 hours, and then injected with 3-PO 50 mg/kg, followed by sleep deprivation. Novel object recognition experiments were performed 72 hours after LPS injection. Subsequently, blood and brain tissue samples were collected. The contents of lactate (Lac), reactive oxygen species (ROS) and serum tumor necrosis factor-α(TNF-α), neuron-specific enolase (NSE), pyruvate in brain tissue were detected by enzyme-linked immunosorbent assay (ELISA). Then, the lactate/pyruvate ratio was calculated. Na +-K +-ATPase activity in brain tissue was detected by colorimetry. Morphological changes in hippocampus were detected by hematoxylin-eosin (HE) staining. And the protein expression levels of PFKFB3, ZO-1 and cleaved caspase-3 were measured by Western blotting. Results:Compared with Con group, the novel object recognition index of LPS group was decreased, the levels of NSE, TNF-α, lactate/pyruvate ratio in serum and the levels of Lac, ROS and dry-wet weight ratio in brain tissue were significantly increased, Na +-K +-ATPase activity in brain tissue was decreased, the protein expressions of PFKFB3, caspase-3 were up-regulated, ZO-1 expression was down-regulated, and the neurons in hippocampus were slightly degenerated. Compared with LPS group, the novel object recognition index of LPS+SD group was further decreased [(39.4±5.3)% vs. (54.5±7.6)%)], serum NSE, TNF-α, lactate/pyruvate ratio and brain tissue Lac, ROS, dry-wet weight ratio were further increased [NSE (μg/L): 3.21±0.42 vs. 2.55±0.36, TNF-α (ng/L): 139.4±19.7 vs. 92.2±13.5, lactate/pyruvate ratio: 29.7±5.5 vs. 19.2±4.2, Lac (μmol/g): 19.51±2.33 vs. 11.34±1.52, ROS (kU/g): 117.4±18.7 vs. 78.2±11.8, dry-wet weight ratio: (81.3±9.2)% vs. (64.3±6.6)%], and Na +-K +-ATPase activity was further decreased (mmol·L -1·h -1: 1.88±0.34 vs. 2.91±0.39), the protein expressions of PFKFB3, caspase-3 were further up-regulated and ZO-1 expression was further down-regulated (PFKFB3/β-actin: 0.80±0.11 vs. 0.45±0.07, caspase-3/β-actin: 0.71±0.09 vs. 0.37±0.05, ZO-1/β-actin: 0.31±0.05 vs. 0.61±0.08). The differences were statistically significant (all P < 0.05). HE staining showed that the degeneration of neurons in hippocampus was significantly aggravated. Compared with LPS+SD group, the novel object recognition index of LPS+SD+3-PO group was increased [(50.8±5.9)% vs. (39.4±5.3)%], NSE, TNF-α, lactate/pyruvate ratio of serum and Lac, ROS, dry-wet weight ratio of brain tissue were significantly decreased [NSE (μg/L): 2.60±0.33 vs. 3.21±0.42, TNF-α (ng/L): 103.7±18.3 vs. 139.4±19.7, lactate/pyruvate ratio: 17.4±5.1 vs. 29.7±5.5, Lac (μmol/g): 13.68±2.02 vs. 19.51±2.33, ROS (kU/g): 86.9±14.5 vs. 117.4±18.7, dry-wet weight ratio: (67.7±6.9)% vs. (81.3±9.2)%], and Na +-K +-ATPase activity was increased (mmol·L -1·h -1: 2.82±0.44 vs. 1.88±0.34). The protein expressions of PFKFB3, caspase-3 were down-regulated and ZO-1 expression was up-regulated (PFKFB3/β-actin: 0.50±0.06 vs. 0.80±0.11, caspase-3/β-actin: 0.43±0.06 vs. 0.71±0.09, ZO-1/β-actin: 0.52±0.06 vs. 0.31±0.05). The differences were statistically significant (all P < 0.05). HE staining showed that the degeneration of neurons in hippocampus was significantly improved. Conclusions:Sleep deprivation could aggravate neuroinflammation, neuronal degeneration and apoptosis in septic rats, resulting in destruction of blood-brain barrier and cognitive impairment. 3-PO treatment significantly alleviate the injury and degeneration of hippocampal neurons in septic rats, inhibit neuroinflammation and apoptosis, and improve cognitive dysfunction, which may be related to the inhibition of glycolytic isoenzyme PFKFB3.

Various posttranslational modifications (PTMs) participate in nearly all aspects of biological processes by regulating protein functions, and aberrant states of PTMs are frequently implicated in human diseases. Therefore, an integral resource of PTM-disease associations (PDAs) would be a great help for both academic research and clinical use. In this work, we reported PTMD, a well-curated database containing PTMs that are associated with human diseases. We manually collected 1950 known PDAs in 749 proteins for 23 types of PTMs and 275 types of diseases from the literature. Database analyses show that phosphorylation has the largest number of disease associations, whereas neurologic diseases have the largest number of PTM associations. We classified all known PDAs into six classes according to the PTM status in diseases and demonstrated that the upregulation and presence of PTM events account for a predominant proportion of disease-associated PTM events. By reconstructing a disease-gene network, we observed that breast cancers have the largest number of associated PTMs and AKT1 has the largest number of PTMs connected to diseases. Finally, the PTMD database was developed with detailed annotations and can be a useful resource for further analyzing the relations between PTMs and human diseases. PTMD is freely accessible at http://ptmd.biocuckoo.org.
Databases, Protein ; Disease ; genetics ; Gene Regulatory Networks ; Humans ; Phosphorylation ; Protein Processing, Post-Translational ; Proteins ; metabolism ; Search Engine