Objective To establish real-time fluorescence quantitative polymerase chain reaction(RFQ-PCR) for measurement of the expression level of a proliferation-inducing ligand(APRIL)and its receptors in children with non-Hodgkin′s lymphoma(NHL).Met-hods Specific primers and TaqMan probes of APRIL and its receptors had been designed,and fluorescence of the PCR products were detected continuously during amplification.According to the standard curves created by plasmid DNA,the expression level of target genes in clinical samples had been determined using software,and these results were presented as the ratios of target genes′ mRNA to ?2 microg-luobulin(?2M)′s mRNA.Results The detection range of RFQ-PCR was between 101-109 ng/L,the coefficient of variation values for both intra-experimental and inter-experimental reproducibility ranged from 1.68% to 5.97% and 6.40% to 10.58%,respectively.The results from 22 samples showed that the expression level of APRIL,B cell maturation antigen(BCMA) and transmembrane activator and calcium modulator and cyclophilin ligand(TACI) in peripheral blood of NHL were significantly higher than those in normal children(P

The study aims to solve the instability problem of methylphenidate (MPH) in plasma, and establish a LC-MS/MS method for simultaneous determining of MPH in human plasma. The stabilities of MPH in different media were studied, and the degradation characteristics of MPH in these media were also investigated by HPLC and LC-MS/MS. To a 200 microL aliquot of freshly collected plasma sample, 10 microL 2% formic acid was added immediately to prevent the hydrolysis of MPH in human plasma samples. Chromatographic separation was performed on a Sapphire C18 column using the mobile phase of methanol - 5 mmol.L-1 ammonium acetate buffer solution containing 0.1% formic acid (46 : 54). MPH was quantified by tandem mass spectrometry operating in positive electrospray ionization mode with multiple reaction monitoring. The detection used the transitions of protonated molecules at m/z 234.2-->84.1 for MPH and m/z 260.3-->183.1 for propranolol (IS), separately. The intra- and inter-assay precisions were all below 5.0%. The accuracies were all in standard ranges. The linear calibration curve was obtained in the concentration range of 0.035-40 ng.mL-1. The methods fulfilled the demand. The method was used to determine the concentration of MPH in human plasma after a single dose of 36 mg MPH tablet to 6 healthy Chinese volunteers. The method is suitable for the precisely determination of MPH and for pharmacokinetic study of MPH in human plasma.
Adult ; Central Nervous System Stimulants ; blood ; pharmacokinetics ; Chromatography, High Pressure Liquid ; Chromatography, Liquid ; Drug Stability ; Humans ; Male ; Methylphenidate ; blood ; pharmacokinetics ; Spectrometry, Mass, Electrospray Ionization ; Tandem Mass Spectrometry ; Young Adult

Objective To observe Meissner's corpuscles in prepuces of different shapes and ages. Methods The Meissner's corpuscles were detected with immunohistochemical stain in 204 prepuce sam- ples of different shapes and ages (3-59 years),and the density of Meissner's corpuscles in every sample was obtained as well.The difference of Meissner's corpuscle densities between phimosis and redundant pre- puce,and correlation between Meissner's corpuscle densities and ages were analyzed with Chi-square test and linear regression,respectively.Results The density of Meissner's corpuscles in redundant prepuce has begun to increase since infancy and reached the peak at the age about 15 years.No significant difference in densities of Meissner's corpuscles between phimosis and redundant prepuce was observed till the age of 20 years,and then there was a trend of disappearance of Meissner's corpuscles in redundant prepuce.A signifi- cantly negative correlation between the densities of Meissner's corpuscles and ages was revealed in redundant prepuce (r=-0.236,P=0.009),whereas an insignificantly positive correlation between the densities of Meissner's corpuscles and ages was shown in phimosis (r=0.193,P=0.084).Conclusions The den- sities of Meissner's corpuscles in redundant prepuce develop synchronically with genital differentiation and accord with the status of sexual function in adult males.The persistent high level of Meissner's corpuscles in adult phimosis might be a mechanism of physiological compensation.

Objective To study the national surveillance results and learn the current situation of iodized salt consumption at household level in 2008, and to find out the remaining problems and to provide scientific basis for developing control strategies against iedine deficiency disorders. Methods In 2008, in accordance with the requirements of the "National Iodine Deficiency Disorders Surveillance Program (Trial)", the surveillance was conducted at county level in 31 provinces and at division level in Xinjiang Production and Construction Corps. In each county 9 townships were randomly selected according to their sub-area positions of east, west, south, north and center;4 villages were randomly sampled in each chosen township;8 households were randomly selected in each chosen village. In every county with 9 or less townships, 1 township was randomly selected respectively in the east, west, south,north and center sub-areas;4 villages were randomly sampled in each chosen township;15 households were randomly selected in each chosen village. Edible salt from these households was collected. Iodized salt coverage rate, proportion of qualified iodized salt and consumption rate of the qualified iodized salt of the households in each province were counted and analyzed. Iodized salt was determined by direct titration;the salt samples from Sichuan and other enhanced salt were detected by arbitration. Results Totally 2817 counties (districts, cities, banners) and 14 divisions of the Xinjiang Production and Construction Corps reported the monitoring results, monitoring coverage reached 99.96%(2831/2832). Mean of iodine content was 31.51 mg/kg.Sixteen provinces had a variation coefficient of iodine content for more than 20%. A total of 826 968 households were tested of their edible salt, in which iodized salt 798 725 copies, non-iodized salt 28 243 copies, and unqualified iodized salt 20 270 copies. Weighted by population,at national level, the coverage rate of iodized salt was 97.48%, qualified rate of iodized salt 97.16%, and consumption rate of qualified iodized salt was 94.79%.Twenty seven provinces (autonomous regions and municipalities) and Xinjiang Production and Construction Corps had a qualified iodized salt coverage rate of above or equal 90.00%. Tibet, Hainan, Xinjiang and Tianjin provinces (regions) had a qualified iodized salt coverage rate lower than 90.00%. Further, 2487 counties had the rate high or equal 90.00% accounting for 87.82% (2487/2831) of complementing monitoring counties. One hundred and four counties and 1 division of the Xinjiang Production and Construction Corps had the coverage rate of iodized salt below 80.00%. Conclusions Sixteen provinces(autonomous regions and municipalities) have relatively a high degree of variation coefficient in salt iodine content. The quality of iodized salt needs to be improved. The coverage rate of iodized salt and the qualified iodized salt at national level are both above or equal 90.00%. However, the non-iodized salt problem is still serious and have a relatively lower coverage of iodized salt in Tibet, Hainan and Xinjiang.

Objective To understand the current level of iodized salt coverage in areas with intensified monitoring measure in China in 2008. Methods In accordance with the "National Iodine Deficiency Disorders Surveillance Program (Trial)" of Ministry of Health issued in 2007, the selected key counties (cities, districts and banner) were divided into 5 sub-areas, 1 non-iodine townships(towns, street offices) was sampled randomly in each sub-area, 4 administrative villages (neighborhood committees) were sampled from each selected township;15households salt samples in each selected village were randomly collected. All salt samples were detected by semiquantitative kit at first. The salt samples that can not be determined by the kit were tested by direct titration and the arbitration act (GB/T 13025.7-1999) detection. Iodized salt determination criteria: reagent color change in semiquantitative test kit or iodine content ≥ 5 mg/kg were identified as iodized salt. Otherwise, the salt samples were identified as non-iodized salt. Results All the provinces(autonomous regions, municipalities) except Tibet in China had conducted a specific survey on iodized salt coverage in non-iodized salt high-risk areas, which revealed that the national coverage rate of iodized salt was 93.01%(130 928/140 770). At the provincial level, twenty provinces and the Xinjiang Production and Construction Corp had a iodized salt coverage over 90%, while the other six provinces (Beijing, Xinjiang, Zhejiang, Fujian, Tianjin and Jiangxi) between 80% - ＜ 90% and the rest four provinces,such as Guangxi, Qinghai, Guangdong and Hainan, lower than 80%. At the county level, 64.57%(277/429) of all the surveillance counties had a iodized salt coverage over 95% while 10.02%(43/429) lower than 80%. Among all the six types of areas where specific survey were conducted, areas with incomplete iodized salt distribution network and areas with crude salt production had a iodized salt coverage lower than 90%, 81.74%(4978/6090) and 86.53%(17 098/19 759), respectively. In raw salt production area, there were 10 out of 21 provinces with iodized salt coverage rate below 90%, it consisted of 47.6%(10/21) of the total monitoring provinces in the same type areas.There were 8 out of 16 provinces with iodized salt coverage rate lower than 90% in the areas with faultiness iodized salt network, it consisted of 50.0%(8/16) of the sampling provinces in the same type area. Conclusions Most provinces(21) in China have a relatively high iodized salt coverage at household level during this specific survey.Areas with incomplete iodized salt distribution network and crude salt production are the most affected areas by noniodized salt. Aiming at the high-risk non-iodized salt areas discovered during this survey, corresponding intervention measures should be implemented with joint efforts from sectors concerned.

Psychrotrophs were isolated by using four media from low-temperature sewage of sewage treatment plant in Urumqi, Xinjiang. Totally, 154 strains were obtained including 12 filamentous fungi, 46 yeasts, 6 actinomycetes and 90 bacteria. The results of tolerance tests of the isolates to salt, phenol and SDS, and enzyme producing characters of amylase, proteinase and esterase were shown. Then 60 bacterial strains were chosen for 16S rRNA gene sequencing and analysis. The blasting results showed that the strains were assigned to 13 recognized genera , and the Strain 39 exhibited 96.6% similarity to Acinetobacter lwoffii(DSM2403), indicating that it might be a novel species. These results suggested that there were a lot of psychrotrophs and rich bacterial diversity in low-temperature sewage. In addition, which maybe an important and potential library of microbial resources.

OBJECTIVETo evaluate the expression of dental matrix protein-l (DMP1) in porcine oral mucosa fibroblasts (POMF) transfected by DMP1 and the influences of the transfection.

METHODSThe full length of porcine DMP1 cDNA was linked into an eukaryotic expression vector pEGFP-C1. POMF and mesenchymal stem cells (MSC) were transfected with the pEGFP-DMP1. The expression of DMP1, dental sialoprotein (DSP), amelin and ameloblastin (Ambn) gene of transfected POMF and MSC were detected by RT-PCR. The expression of DMP1 and DSP protein was examined by immunocytochemical staining. The formation ratio of mineralized nodules of transfected cells was compared with un-transfected ones after mineralized induction. The formation of mineralized nodules of three-dimensional pellet transfected cells was compared with un-transfected ones after hematoxylin and eosin staining.

RESULTSThe constructed pEGFP-DMP1 could produce 4.7 kb and 1.5 kb fragments. DMP1 gene, DSP gene and Ambn gene were expressed by POMF after transfection. Immunohistochemical staining and the quantitative analysis of protein showed that DMP1 and DSP protein was positive in transfected POMF and MSC. The formation ratio of mineralized nodules of transfected POMF and MSC was higher than that of un-transfected ones (P < 0.05).

CONCLUSIONSThe expression of porcine DMP1 in POME after gene transfection can induce the expression of tooth-development-associated gene Ambn and DSP and enhance the formation of mineralized nodules.

Animals ; Calcification, Physiologic ; Cell Differentiation ; Cells, Cultured ; Extracellular Matrix Proteins ; genetics ; metabolism ; Fibroblasts ; cytology ; metabolism ; Genetic Vectors ; Mouth Mucosa ; cytology ; Phosphoproteins ; genetics ; metabolism ; Swine ; Transfection

OBJECTIVETo investigate the growth and development of brain derived neurophic factor(BDNF)-positive neurons in the frontal lobe of human fetus.

METHODSThe expression of the BDNF-positive neurons in the frontal lobe of human fetus in the 2(nd),3(rd),and 4(th) month of gestation were observed with the streptavidin-biotin-complex/immunoperoxidase(SABC)method.

RESULTSBy the second month of gestation,BDNF-positive neurons were seen in the subventricular layer of the frontal lobe of cerebellum.By the third month of gestation,BDNF-positive neurons in the central layer were in various shapes,with big nucleus,less cytoplasm,and small processes.By the fourth month of gestation,BDNF-positive neurons in the central layer grew larger in size,cytoplasm increased,the BDNF-positive expression was enhanced with deeper dyeing,and the nerve fibers and particles were distributed between neurons;also,the BDNF-positive neurons were seen in the marginal layer of the frontal lobe of cerebrum.

CONCLUSIONBDNF-positive neurons may participate in the early development of the frontal lobe of cerebrum of human fetus.

Brain-Derived Neurotrophic Factor ; metabolism ; Fetus ; metabolism ; Frontal Lobe ; embryology ; Humans ; Neurons ; cytology ; metabolism

BACKGROUNDWe investigated the pathogenesis of atherosclerosis in diabetes, and detected the expression of scavenger receptor CD36 in monocytes in patients with type 2 diabetes.

METHODSAccording to the criteria by WHO, diabetic patients were classified into two groups: well controlled diabetic patients (WCP) and poorly controlled diabetic patients (PCP). The expression of CD36 protein and mRNA were evaluated by flow cytometry and reversal transcription polymerase chain reaction (RT-PCR). Plasma levels of accumulation of oxidized LDL (oxLDL) were directly measured by sandwich enzyme-linked immunosorbent assay (ELISA) method.

RESULTSFlow cytometry and RT-PCR showed that the mean fluorescence intensity (MFI) of CD36 in monocyte and CD36 mRNA were significantly higher in the PCP and WCP in comparison with healthy controls (P<0.01). CD36 MFI and mRNA in the PCP were increased by 78% and 36% compared to the WCP. In both groups, CD36 MFI and mRNA were significantly higher in patients with diabetic atherosclerosis in comparison with those without diabetic atherosclerosis (P<0.05). No significant difference was found in CD14 expression between the groups (P>0.05). The concentrations of plasma oxLDL were higher in the PCP group compared to WCP and control group (P<0.05), whereas oxLDL average values did not differ significantly between WCP and control groups (P>0.05). In the WCP and PCP groups, oxLDL levels were higher in patients with diabetic atherosclerosis than those without diabetic atherosclerosis (P<0.05).

CONCLUSIONSThe increased expression of scavenger receptor CD36 may be one of the mechanism of accelerated atherosclerosis in diabetic. The poorly controlled diabetes patients are at higher risk for the vascular complications than the well controlled diabetic patients.

Adult ; Aged ; Atherosclerosis ; etiology ; CD36 Antigens ; genetics ; Diabetes Mellitus, Type 2 ; complications ; Female ; Fluorescent Antibody Technique ; Humans ; Lipoproteins, LDL ; analysis ; Male ; Middle Aged ; Monocytes ; metabolism ; RNA, Messenger ; analysis ; Regression Analysis

OBJECTIVETo study the effect of SOD2 and its C47T mutation on oxidative injury in cochlea hair cells.

METHODSHEI-OC1 cells were transfected with the SOD2 of Ala16 and Vla16. Cells' proliferation ability was determined by MTT assay. The intracellular SOD2 activities were detected by xanthine oxidase method. Intracellular ROS were determined by DCFH-DA after exposure to 100 µmol/L t-BHP and the early apoptotic and necrotic rate or late apoptotic rate were quantified by flow cytometry (FCM) using Annexin V/PI double staining.

RESULTSMTT method showed the transfection of SOD2 gene and empty plasmid did not affect the proliferation capacity. SOD2 vitality in Ala(16) and Val(16) SOD2 transfected cells increased 2.51 and 2.71 times respectively (P < 0.01), but the difference between the two transfection groups was not statistically significant (P > 0.05). After exposed to t-BHP, the majority of the untransfected and empty plasmid transfected cells sent '++' class bright fluorescence, while in Ala(16) and Val(16) SOD2 transfected groups, only about half cells sent '±' ∼ '+' level fuzzy fluorescence. determination of FCM suggested the early apoptotic and necrotic rate or late apoptotic rate decreased after SOD2 transfection (P < 0.01), but the difference between the two genotypes of SOD2 was not statistically significant (P > 0.05).

CONCLUSIONHigh expression of SOD2 below 3.71 times can reduce intracellular ROS level in HEI-OC1 cells, while SOD2 C47T mutation had no effect on them. SOD2 can be considered as NIHL susceptibility gene and its rs4880 SNP may be not directly related to NIHL genetic susceptibility.

Cell Line ; Cochlea ; metabolism ; Hair Cells, Auditory ; metabolism ; Humans ; Mutation ; Oxidative Stress ; Polymorphism, Single Nucleotide ; Reactive Oxygen Species ; metabolism ; Superoxide Dismutase ; genetics