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ObjectiveTo explore the pretreatment methods to promote the enzymatic digestion and extraction of active ingredients from Magnoliae Officinalis Cortex dregs（MOCD）， and to provide a reference basis for the utilization of resource components in MOCD. MethodLiquid chromatography-mass spectrometry（LC-MS） was used for qualitative analysis of resource components in MOCD with an Agilent C18 reversed-phase column（3.0 mm×100 mm， 2.7 µm） at the flow rate of 0.4 mL·min-1， the mobile phase was water（A）-acetonitrile（B） for gradient elution（0-3 min， 25%-48%B； 3-6 min， 48%-59%B； 6-10 min， 59%-80%B； 10-20 min， 80%-90%B； 20-25 min， 90%B）， electrospray ionization（ESI） was employed with negative ion mode scanning and scanning range of m/z 50-1 200. A high performance liquid chromatography（HPLC）， which refered to the determination in the 2020 edition of Chinese Pharmacopoeia， was used for quantitative analysis of resource components in MOCD. Four kinds of pretreatment agents were used to separate the resource components from MOCD， and the mechanism of different pretreatment agents was investigated by field emission scanning electron microscopy（FESEM）， X-ray powder diffraction（XRD） and Fourier transform infrared spectroscopy（FT-IR）. ResultMagnolol， honokiol and lignocellulose were identified as the main resource components of MOCD by qualitative and quantitative analysis. Under the conditions of 1% NaOH， reaction temperature at 80 ℃ and reaction time of 60 min， the concentration of reducing sugar produced by the enzymatic hydrolysis was 32.18 g·L-1， which was 79.8% higher than that of the untreated MOCD. After adding tween-80， the enzymatic hydrolysis time was reduced to 1/3 of the original time， the concentration of reducing sugar was increased by 102.0%. And the total recovery of magnolol and honokiol in the pretreatment solution was 69.23%. ConclusionMagnolol， honokiol and lignocellulosic components in MOCD are valuable for development and utilization， the combination of alkaline pretreatment and tween-80 can realize the recovery and utilization of these three resource components， which can provide a new idea for comprehensive utilization of resource components in MOCD.
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Objective:To observe multiple locus variable-number tandem repeat analysis (MLVA) typing of Brucella isolated from Himalayan marmot in Qinghai-Tibet Plateau of Qinghai Province, and to explore the relationship between the strains and strains previous isolated from Qinghai Province. Methods:Blood samples of Himalayan marmot were collected in Qinghai-Tibet Plateau of Qinghai Province from March 2019 to October 2020. Pathogens were isolated and cultured from Brucella antibody positive samples identified by using the rose bengal test (RBT). Conventional biological methods and molecular biological methods (BCSP31-PCR and AMOS-PCR) were used for strain identification. At the same time, MLVA method was used to genotype the isolated strains, and cluster analysis was used to analyze the genetic relationships between the strains based on the genotype of 70 Brucella isolated from different hosts in Qinghai Province. Results:A total of 1 466 blood samples of Himalayan marmot were collected from Qinghai-Tibet Plateau. Two strains of Brucella were isolated and cultured from 64 RBT-positive samples, named QH2013054 and QH2013062, respectively. They were identified as Brucella ovis biotype Ⅲ by conventional and molecular biological methods. The MLVA genotyping results showed that QH2013054 and QH2013062 were different at the Bru16 locus, indicating different MLVA genotypes. Cluster analysis showed that strain QH2013054 had the same MLVA genotype as 7 strains, among which 6 strains were from 3 farmers and 3 sheep from the same family in Gonghe County, and 1 strain was from a farmer in Menyuan Hui Autonomous County. The strain QH2013062 had the same MLVA genotype as 4 strains, including 3 strains from 3 farmers in Menyuan Hui Autonomous County and 1 strain from a farmer in Tu Autonomous County of Huzhu. Conclusions:The strains of Brucella isolated from Himalayan marmot in Qinghai-Tibet Plateau of Qinghai Province have the same MLVA genotype as some strains of Brucella isolated from humans and sheep in Qinghai Province. It is speculated that the host humans, sheep and Himalayan marmot in Qinghai-Tibet Plateau may have a common source of infection.
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The mucosal surfaces of the human body are typically colonized by polymicrobial communities, they exist in the gastrointestinal tract, skin, oral cavity, and genitourinary tract, as well as in anatomical sites previously considered sterile, including the lung. These communities interact with the mucosal immune system to maintain homeostasis in health, emphasizing the important role of " second genome" , which is very important to human health. The human lower respiratory tract also harbor many microbes, the imbalance of microbiome composition maybe critically related with many lung disease. Up to date, researches on microbiome of lung caner is still on the first step. Our review mainly describes the characteristic composition of lung microbiome, and the relationship with diseases, especially with lung cancer. We attempts to provide the basis information for further researches about the relationship between human lung microbiome and lung cancer.
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Objective:The epidemic regularity and trend of human brucellosis in Qinghai Province were analyzed to provide basis for formulating more perfect prevention and control measures.Methods:Using descriptive epidemiological methods, the data of brucellosis reported cases in Qinghai Province from 2005 to 2018 (from the Infectious Disease Report Information Management System of the China Disease Prevention and Control Information System) were collected, and the three distributions (time, region and population distributions) of human brucellosis were statistically described and analyzed.Results:From 2005 to 2018, a total of 418 cases were reported in Qinghai Province, with a incidence of 0.05/100 000, the incidence of different years was statistically different (χ 2 = 11.63, P < 0.05). The incidence of brucellosis remained at a low level from 2005 to 2010, ranged from 0 to 0.05/100 000; and the incidence increased from 2011 to 2018, ranged from 0.27/100 000 to 2.06/100 000. There were reported cases of brucellosis throughout the year, and the onset time was mainly from April to June, accounting for 42.82% (179/418). The 418 cases were distributed in 20 counties (cities, districts), among which the top 5 incidence rates were Menyuan County (15.31%, 64/418), Henan County (13.40%, 56/418), Tianjun County (13.16%, 55/418), Xining City (12.44%, 52/418), and Haiyan County (7.66%, 32/418). In the population distribution, brucellosis cases in the age group from 40 to 49 years old were the most, accounting for 32.78% (137/418), the male to female ratio was 1.58 ∶ 1.00 (256∶162) . And the occupational distribution was dominated by herdsmen, accounting for 57.18% (239/418). Conclusions:The epidemic situation of brucellosis in Qinghai Province is on the rise. It should be strengthened the population monitoring and notification of outbreaks, and health education for all should be implemented.
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Objective:To investigate the role of IL-1β in regulating human placenta-derived mesenchymal stem cells (hPMSCs) to induce the differentiation of activated peripheral blood mononuclear cells (PBMCs) into CD8 + IL-10 + T cell subset in patients with rheumatoid arthritis (RA). Methods:Serum IL-1β levels in RA patients were detected by CBA kit. hPMSCs were isolated from healthy subjects by enzyme digestion. PBMCs that were isolated from RA patients and healthy subjects by Ficoll density gradient centrifugation were activated with PHA and CD3/CD28 monoclonal antibody (McAb) and then co-cultured with hPMSCs. Flow cytometry (FCM) was used to analyze the ability of hPMSCs to induce the differentiation of activated PBMCs into CD8 + IL-10 + T cell subset with the presence of programmed death ligand 1 (PD-L1) and/or PD-L2 McAb and IL-1β. Expression of PD-L1 and PD-L2 on hPMSCs under IL-1β stimulation was analyzed by RT-PCR and FCM. Results:Serum IL-1β levels were significantly higher in RA patients than in healthy subjects. After the activation by PHA and CD3/CD28 McAb, the proportion of CD8 + IL-10 + T cells in PBMCs of RA patients was significantly lower than that of healthy subjects. Moreover, the proportion of CD8 + IL-10 + T cells significantly increased after co-cultured with hPMSCs. The ability of IL-1β-treated hPMSCs to induce activated PBMCs to differentiate into CD8 + IL-10 + T cells was significantly enhanced compared to untreated hPMSCs. IL-1β could up-regulate the expression of PD-L1 and PD-L2 on hPMSCs. Blocking the expression of PD-L1 and/or PD-L2 on hPMSCs significantly reduced the ability of hPMSCs to induce activated PBMCs to differentiate into CD8 + IL-10 + T cells. Conclusions:IL-1β enhanced the capacity of hPMSCs to induce activated PBMCs to differentiate into CD8 + IL-10 + T cells in RA patients by up-regulating the expression of PD-L1 and PD-L2.
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OBJECTIVE： To provide reference for promoting the implementation of “two-invoice system” for drug purchase in medical institutions. METHODS： The function of confirmation， enquiry， statistics for “two-invoice system” of drug purchase order were added into relevant module of SPD （supply processing distribution） system to realize the management of “two-invoice system” data by means of information technology. By analyzing the data of the “two-invoice system” of Shanxi dayi hospital （our hospital） from Aug. 2017 to Jun. 2018， the effects of SPD system on “two-invoice system” of drug purchase in our hospital were introduced after the application of SPD system. The effects of SPD system were evaluated in respects of work efficiency， accuracy rate and resource saving， etc. RESULTS： SPD system was used for the management of “two-invoice system” to realize the information management of invoices. The time for checking “two-invoice system” information was shortened from 1.5 min/sheet to 0.1 min/sheet； the accuracy rate was increased from 95％ to 100％， and about 40 000 sheets of paper were saved each year. CONCLUSIONS： The application of SPD system in “two-invoice system” of drug purchase can improve the work efficiency， reduce the labor cost and reduce the waste of resources， it effectively improves the level of management in our hospital.
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Objective To compare the size of the internal target volume (ITV),biological target volume (BTV) and internal biological target volume (IBTV) based on PET-CT and 4DCT for primary nonsmall cell lung cancer (NSCLC),as well as try to apply IBTV in radiotherapy planning.Methods A total of 15 patients with NSCLC were sequentially scanned by an axial enhanced 3DCT,4DCT and 18F-FDG PET-CT in the thoracic region.The gross target volumes (GTVs) of ten phases of 4DCT images were contoured,and ITV was obtained by fusion of ten GTVs.BTV based on PET-CT images was determined by the SUV 2.0.The IBTV was defined by fusion of ITV and BTV.Planning target volumes (PTVs) based on ITV,BTV,and IBTV (PITV,PBTV,PIBTV) were obtained by ITV,BTV and IBTV with a 10-mm expansion respectively.The metrics of PIBTV,PITV and PBTV were compared,and the planning parameters of target volumes and risk organs were evaluated.Results There was no significant difference between ITV and BTV,but there was significant difference between IBTV and ITV and BTV (F=22.533,P ＜ 0.05).To include more than 95％ volume of IBTV,it is necessary to expand the margin of 9.0(6.0,12.0)mm based on BTV or 10.00(7.0,12.0)mm based on ITV.There was no significant difference between the two groups (P ＞ 0.05).Dice's similarity coefficient of BTV and ITV was 0.72(0.54,0.79).The intensity modulated radiotherapy plan based on PBTV can guarantee 85.6％ (80.5％,91.2％) of PITV to reach the prescription dose,compared with 80.2％ (74.4％,87.6％) of PBTV by the plan from PITV.Additionally,the conformity index and homogeneity index were not ideal.The dosimetric parameters of PITV and PBTV in the PIBTV plan were much better than those in PBTV-and PITV plan.Conclusions The radiotherapy plan based on PET-CT or 4DCT could not guarantee a reasonable dose distribution of PTV expanded from ITV or BTV.Thus,using IBTV for radiotherapy is advised.
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The training of academic postgraduate students in critical care medicine emphasizes basic knowledge and underestimates the importance of clinical practice. Research objectives are innovative, but with a lack of practicability, and thus cannot meet the huge needs for clinical research in this major. Under the guidance of the concept of translational medicine , the training of academic postgraduate students should strengthen clinical observation and systematic learning of clinical knowledge, enhance the accumulation of clinical data, improve the quality of data statistics, reinforce the training on clinical research processes, standards, and methods, and emphasize interdisciplinary and multicenter communication and cooperation, so as to break the barrier between basic and clinical research, broaden scientific research thinking , and improve the comprehensive quality of academic postgraduate students in critical care medicine.
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Objective To research the genetic susceptibility of proliferative diabetic retinopathy (PDR) in Han patients with type 2 diabetes from Southern China.Methods A cross-sectional study was performed under the informed consent of the patients.Patients with type 2 diabetes in the Dongguan Eye Study from September 2011 to February 2012 and relative patients treated in Guangdong General Hospital from July 2017 to March 2018 were included in this study,including 100 patients with diabetes mellitus(DM) and 120 patients with PDR.Whole exome sequencing was used to identify DNA mutation in peripheral blood samples from 22 type 2 diabetic patients without retinopathy (DM group) and 23 diabetic patients with PDR (PDR group).Genotype and allele of the nine selected single-nucleotide polymorphisms (SNPs) were tested and analyzed by SnaPshot technology in another 78 DM patients without retinopathy and 97 PDR patients.Results A total of 75 SNPs were associated with PDR (P＜0.01),involving 53 genes.Eleven gene loci were in the exon region and 7 were non-synonymous mutations.Nine exon loci of 8 genes with significant differences were screened out for the verification.SnaPshot SNP genotyping technique found that there were no significant differences in allele and genotype frequency in the nine selected SNPs between PDR group and DM group (all at P＞0.05).However,7 haplotypes distribution frequencies were significantly different between PDR group and DM group (all at P＜0.01).Hapl and Hap4 might reduce the risk of PDR (both at OR＜1,P＜0.05),and Hap2 might increase the risk of PDR (OR＞ 1,P＜0.05).Conclusions The occurrence of PDR probably has a genetic susceptibility in type 2 DM patients of Han nationality in Southern China.
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Objective To investigate the analgesic effect of different concentrations of medetomidine and ropivacaine on knee arthroscopy in elderly patients in the orthopedics department. Methods 72 patients were di-vided into 4 groups,and from February 2016-September 2017 ,different concentrations of dexmedetomidin and ropivacaine were given to the patients after the operation,and compared the analgesic effects and adverse reactions of each group. Results The duration of analgesia in groups C and D were longer than group A and B and there were significant differences in the amount of fentanyl in the 4 groups after 24 h(F = 79.633,P < 0.05 )and there was no significant difference in the incidence of adverse reactions between the 4 groups(P > 0.05);1,3,6, 10,16,24 h after operation,The VAS scores in group C and D were significantly lower than those in group A and B(P < 0.05 ). Conclusion Dexmedetomidine plus ropivacaine can effectively improve the analgesic effect after knee arthroscopy,0.50 g/kg was the best concentration of dexmedetomidin.
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Objective To reconstruct the original three-dimensional conformation of tumor resection tissue through the study of breast-conserving surgery excision specimens for part-mount sub-serial section and pathological three-dimensional (3D) reconstruction,to establish a new margin assessment model,and to guide tumor bed delineation individually for radiotherapy.Methods From February 2016 to February 2017,thirtythree eligible breast cancer patients underwent breast-conserving surgery in Breast Cancer Center of Shandong Cancer Hospital were recruited.The excision specimens were prepared with part-mount sub-serial section,and residual tumors were microscopically outlined,scanned and registered by Photoshop software.The 3D model of residual tumors was reconstructed with 3D-DOCTOR software to evaluate margin status and record pathological type,tumor length and 3D negative margin distance.The gross tumor volume (GTV) was delineated based on clips placed in the lumpectomy cavity.CTV1 and CTV2 were defined by adding uniform 1.00 cm and 1.50 cm margin based on GTV respectively.CTV3 and CTV4 were defined by adding 1.00 cm and 1.50 cm margin based on 3D boundary of excision tumor respectively,and compared the volume differences of CTV1 and CTV3,CTV2 and CTV4.Results Based on the marginal assessment results of 3D pathological reconstruction,the rates of false negatives during the intraoperative rapid pathological examination and postoperative routine pathological margin evaluation were 6.7％ (2/30) and 3.4％ (1/29) respectively.The pathological type of pathological large slice and routine pathological examination was consistent with rate of 93.9％ (31/33).The M(QR) tumor lengths of routine pathological and pathological 3D reconstruction were 1.90 (1.50-2.40) cm and 2.00 (1.60-2.70) cm respectively,with statistical difference between the two groups (Z =-2.438,P =0.015).The M(QR) volumes for CTV1,CTV2,CTV3,CTV4 were 70.76 (49.84-78.07)cm3,110.11 (83.38-126.17) cm3,23.85 (16.46-31.49)cm3 and 38.74 (30.47-50.58) cm3 respectively.There were statistical differences between CTV1 and CTV3,CTV2 and CTV4 (Z =-4.372,P ＜0.001;Z =-4.372,P ＜0.001).Conclusion The application of pathological 3D reconstruction technology can largely compensate for the shortcomings of the traditional margin assessment model,make the decisions of adjuvant treatment after breast-conserving surgery more accurate,and guide the tumor bed delineation individually for radiotherapy.
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At present,there are still many problems in critical care medicine education and teaching,such as imperfect theoretical system,inconsistent teaching materials,uneven teaching staff,lack of practice opportunities and places.The development of intensive care medical teaching should be combined with "organ-system-centered" teaching reform,strengthen the construction of the theoretical system,drive the development of teaching staff,build high-quality practice bases,promote the curriculum to be more specialized,optimized and standardized,so as to promote the development of the subject.
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Objective In order to explore the association between complement C3 and primary open angle glaucoma (POAG),the serum complement C3 level was detected and the polymorphism of the C3 exon gene was analyzed in patients with POAG.Methods A prospective case-control study was designed.A total of 45 patients with POAG visiting Eye &ENT hospital of Fudan University were collected from December 2014 to December 2015,and 45 age-/gender-matched healthy subjects from yearly health screening were collected as normal controls.Serum C3 concentration was detected by immunoturbidimetric assay.Meanwhile,genomic DNA was extracted from peripheral blood leukocytes,and sequencing for C3 exons was followed with PCR to analyze the gene polymorphism.Statistical analysis was performed by use of SPSS 20.0 software.Student's t test was used to evaluate the difference of serum C3 level between two groups.Chisquare test was used to analyze the difference in the distribution of C3 exon gene polymorphism between two groups.The one-way ANOVA was utilized to analyze the difference of serum C3 level among different genotypes of C3 exon.Results The serum level of C3 in POAG (104.81 ± 29.15)mg/dl was significantly lower (t =-3.162,P =0.002) compared to controls (121.06 ± 18.39) mg/dl.There were a total of seven single nucleotide polymorphisms (SNPs) (rs2230201,rs2230204,rs2230205,rs428453,rs423490rs7951,rs539822147) within the C3 exon gene region,but no mutation was detected.There was no difference in the frequency and distribution of these SNPs between POAG and controls,as well as,there was also no significant difference in serum C3 level among these SNPs genotypes.Conclusions Decreased serum C3 level in POAG patients indicated that complement C3 might be involved in the pathomechanism of glaucoma.No significant abnormality of C3 exon gene was detected,so the gene polymorphism having an impact on serum C3 level could be excluded as a reason probably.
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Objective To compare the geometric differences of gross tumor volumes(GTV)and displacements of selected clips propagated by rigid image registration(RIR)and deformable image registration (DIR)at end-inhale phase(CT0)and end-exhale phase(CT50)based on four-dimensional computed tomography(4DCT)of the whole breast after breast-conserving surgery(BCS). Methods Forty-four patients who underwent 4DCT simulation scans after BCS were selected. The GTV and displacements of selected metal clips at CT0and CT50were manually delineated by the same radiotherapy physician. Subsequently,the GTV and displacements of selected clips from CT0images were transformed and propagated to CT50images using RIR and DIR.The geometric differences of GTV and displacements of surgical clips from DIR were compared with those from RIR based on the dice similarity coefficient(DSC)and the displacements of the center of mass(COM)in the three-dimensional(3D)directions. Results The mean DSC was 0.86± 0.04 for RIR and 0.87± 0.04 for DIR(P=0.000).The displacements of COM in 3D directions from RIR were significantly greater than those from DIR(1.22 mm vs. 1.10 mm,P=0.000).In the anterior-posterior direction,the displacements from RIR were significantly greater than those from DIR for both GTV and selected clips(P=0.000).However,in the left-right and superior-inferior directions,there were no significant differences in displacements between RIR and DIR for both GTV and the selected clips(all P>0.05). Conclusions DIR can improve the overlap for GTV registration from 4DCT scans at CT0and CT50.Furthermore,DIR is superior to RIR in reflecting GTV and the displacements of selected clips in anterior-posterior direction induced by respiratory movement.
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Objective To understand the relationship between habitat choice of Himalaya marmot and vegetation elements such as vegetation coverage,grassland type,vegetation type and so on by using geographic information technology in Qinghai.Methods Himalayan marmot field survey data from Qinghai Provincial Institute for Endemic Disease Control and Prevention and vegetation coverage,grassland type and other remote sensing and geographic information data was processed for spatial overlay using ArcGIS tool.The vegetation characteristic information of the location of Himalayan marmot was searched and analyzed statistically,and then the initial quantitative analysis of Himalayan marmot on the selection of habitat vegetation conditions was made.Results After screening,84 samples of Himalayan marmot were retained.The mean of vegetation coverage was 0.708 5,the range was 0.313 3-1.000 0.Totally 54.76％ (46/84) of the Himalayan marmot samples was in the vegetation coverage of 0.70 to 1.00.Among the nine grassland types,the alpine meadows accounted for 39.78％ of total area of Qinghai,with 59 Himalayan marmot sample points distributed,which accounting for 70.24％ (59/84).Himalayan marmot mainly feed on some of the roots of grassland plants,including Poa pratensis,Alpine Kobresia,Potentilla,Polygonum viviparum,Kobresia cordata,Oxytropis,Aster and other miscellaneous grass.Conclusions Vegetation conditions is one of the important factors of Himalayan marmot in habitat selection.Quantitative analysis of the relationship between Himalayan marmot spatial distribution and vegetation factors by using geographic information technology,can deepen the understanding of marmot habitat selection,and provide a basis for grassland plague surveillance and control.
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Objective To analyze the main biochemical indexes in the patients with type 2 diabetes mellitus (T2DM) and complications.Methods A total of 112 cases of patients with T2DM in the Zigong Municipal Third People's Hospital were selected from March 2015 to March 2016 and divided into the T2DM without complications group (61 cases) and T2DM with complications group (51 cases) according to whether having complications.Contemporaneous 55 volunteers undergoing physical examination were selected as the control group.The levels of insulin,fasting blood glucose (FBG),D-dimer (D-D) and serum lipids (HDL,LDL,TC and TG) were compared among three groups.Results The levels of FBG,D-D,TC,TG and LDL in the T2DM without complications group and T2DM with complications group were higher than those in the control group,while the levels of insulin and HDL were lower than those in the control group,the differences were statistically significant (P<0.05);moreover,except HDL,the levels of insulin,FBG,D-D,LDL,TC and TG had statistical difference between the T2DM with complications group and T2DM without complications group (P<0.05).Conclusion Regularly detecting the biochemical indexes of blood glucose and lipids in diabetic patients is an important mean to preventing and treating diabetes mellitus and its complications.
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Objective To improve the effectiveness of internship nursing students in infectious diseases by implementing peer teaching method. Methods The nursing students were divided into two groups according to the order of student number. The control group was given the teaching method of the general outline. On the basis of the outline teaching, the experimental group gave counseling and counseling to the students in their professional and psychological aspects. Results There were significant differences between the two groups in terms of the results of the three tests and the theoretical evaluation, and the results of the test group were better than those of the control group (P<0.05). Conclusion Peer- to- peer learning method can improve the practical effect of surgical nursing students in infectious diseases, and can be popularized in a certain extent.
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Objective To study the false negative results of Gelle test .Methods Hearing test data between August 2013 to July 2014 were retrieved from outpatient records with hearing complaints of tinnitus or hearing loss . Recruiting criteria:no history of otitis media ,normal tympanic membrane ,no radiologic manifestations of lesions in middle ear and mastoid and abiltly of clear expression of changes of hearing .A total of 60 patients(120 ears ) were tested with conventional pure tone audiometry (PTA) and Gelle test .Acoustic immittance and distortion product otoacoustic emission (DPOAE) were tested in 110 and 113 ears ,respectively .Results Among 120 ears ,7 were di‐agnosed as conductive hearing loss ,23 as mixed hearing loss ,52 as sensorineural hearing loss ,and 38 as normal hearing by pure tone audiometry .The negative Gelle test results were detected in 5 ,11 ,16 and 3 ears ,respective‐ly .110 ears were tested with tympanometry ,types A ,B and C tympanograms were found in 102 ears ,2 ears and 6 ears ,respectively ,and the negative Gelle test results were 22 ,2 and 4 ,respectively .DPOAEs were recordable in 56 ears and negative in 57 ears ,and negative Gelle tests were recorded in 12 and 17 ears respectively .Conclusion False-negative results were found in all types of PTA ,tympanometry and DPOAE .It indicates that Gelle test might not be sensitive and accurate enough to evaluate the integrity and mobility of ossicular chain .Gelle test com‐bined with PTA and DPOAE may be better for assessment of hearing loss and entity of ossicular chain .
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<p><b>OBJECTIVE</b>To develop an allele-specific PCR (AS-PCR)/restriction fragment length polymorphism (RFLP) assay for CYP1B1 gene haplotypes predisposing to primary congenital glaucoma (PCG).</p><p><b>METHODS</b>Twenty Chinese PCG patients and 20 healthy controls were recruited. Peripheral blood sample was subjected to direct sequencing for common single nucleotide polymorphisms (SNPs) of the CYP1B1 gene. Based on the results, CYP1B1 gene haplotypes were constructed by PCR-RFLP and AS-PCR combined with RFLP.</p><p><b>RESULTS</b>Four SNPs loci were identified by sequencing, which included rs10012 G>C (S1 in exon 2), rs1056827 T/G (S2 in exon 2), rs1056836 C/G (S3 in exon 3) and rs1056837T>C (S4 in exon 3). The distribution of such loci showed different characteristics between the two groups. 50% of the PCG patients had rs10012 G>C and rs1056827 T>G, while 25% of PCG patients had rs1056836 C>G and rs1056837T>C. As for the controls, 25% had rs10012 G>C and rs1056827 T>G, 10% had rs1056836 C>G and rs1056837T>C. None of the SNP loci has presented alone. PCR-RFLP was carried out to confirm the results of SNPs typing, but could not confirm the linkage between the SNP loci. By contrast, AS-PCR combined with RFLP has achieved specific amplification for rs10012 G>C and thorough differentiation of 1056827 T>G polymorphism. Similar results have been obtained by the same method for rs1056836 C>G and rs1056837T>C typing and linkage disequilibrium analysis.</p><p><b>CONCLUSION</b>The AS-PCR/RFLP assay has successfully constructed the haplotypes of the CYP1B1 gene. For its accuracy, efficiency and specificity, the method may be used for constructing haplotypes for hereditary disease studies.</p>
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Alleles , Base Sequence , Cytochrome P-450 CYP1B1 , Genetics , Gene Frequency , Genetic Predisposition to Disease , Genetics , Genotype , Glaucoma , Genetics , Haplotypes , Linkage Disequilibrium , Polymerase Chain Reaction , Methods , Polymorphism, Restriction Fragment Length , Polymorphism, Single Nucleotide , Reproducibility of Results , Sequence Analysis, DNA , MethodsABSTRACT
Objective To investigate the effect of parathyroid hormone (PTH) on the epithelial to mesenchymal transition (EMT) in human renal proximal tubular epithelial cells (HK-2 cells),and determine the role of β-catenin signaling pathway.Method The expression of α-smooth muscle actin (α-SMA),E-cadherin and β-catenin in HK-2 cells was measured by real-time PCR,Western blotting and immunofluorescence technique.The signaling pathway by which PTH activated EMT in HK -2 cells was identified by using synthetic β-catenin siRNA.Results Parathyroid hormone (10-10mol/ L) increased α-SMA expression and decreased E-cadherin expression in HK-2 cells (P＜ 0.01,respectively).Untreated cells showed the expression of E-cadherin,whereas α-SMA staining was noticeably increased in cells treated with PTH.β-catenin activity was significantly increased after exposed to PTH.Theα-SMA expression was decreased strongly and E-cadherin expression was increased after β-catenin siRNA transfection (all P ＜ 0.05).Conclusion PTH significantly induces epithelial to mesenchymal transition in HK-2 cells throughβ-catenin signaling pathway.