Exploration and advance of key techniques in MALDI mass spectrometry imaging / 药学学报

Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is a new imaging technique with label-free, rapid, and high throughput features. It has bloomed in the analysis on the spatial distribution of biomolecules such as drugs, metabolites, peptides and proteins on the tissue surface in virtue of providing high data throughput from non-targeted full analysis and high accuracy from targeted analysis. The acquisition of MSI signal response with high sensitivity, high spatial resolution, and good stability is directly depended on the appropriate sample preparation approaches, and flexible and various data processing tools will help the non-target data mining to meet the demands of visualization, spatial distribution and multiple index applications so as to reveal the scientific rules beneath the data. This review briefly summarizes the key advances in MALDI-MSI from aspects of sample preparation procedures, data processing and visualization. It also illustrates the characteristics, difficulties and probable solutions derived from these key techniques.

Preparation and identification of rabbit anti-mouse coiled-coil domain containing 189(Ccdc189)polyclonal antibody / 细胞与分子免疫学杂志

Objective To prepare a rabbit anti-mouse coiled-coil domain containing 189 (Ccdc189) polyclonal antibody. Methods The pET-28a-Ccdc189 prokaryotic expression plasmid was constructed and transformed into E.coli BL21. IPTG was used to induce the expression of Ccdc189 prokaryotic protein. Adult male New Zealand rabbits were immunized with purified recombinant protein to obtain rabbit anti-mouse Ccdc189 polyclonal antibody. The specificity of the polyclonal antibody was identified by Western blot analysis, indirect ELISA and immunofluorescence histochemical staining. Results The pET-28a-Ccdc189 recombinant plasmid was successfully constructed and the expression of the Ccdc189 recombinant protein was induced. ELISA revealed that the titer of the polyclonal antibody was 1:1 000 000. Western blot and immunofluorescence staining demonstrated that the Ccdc189 polyclonal antibody could specifically identify the Ccdc189 prokaryotic protein and the Ccdc189 protein in adult wild-type mouse testis. Conclusion A polyclonal antibody with high specificity against mouse Ccdc189 was successfully created.

Survey on cognition of general practice residency training and willingness of teaching among specialists / 中华全科医师杂志

Objective:To survey the cognition of general practice residency training and the willingness of teaching among specialists.Methods:A questionnaire survey was conducted among 221 specialists from 24 departments in Nantong First People's Hospital from May 2021 to June 2021 to investigate the their cognition of general practice residency training program and the teaching willingness.Results:Total 221 questionnaires were distributed and 185 valid ones were retrieved with a response rate of 83.70%. The results showed that 49 specialists (26.49%) well knew the national general practice training policy, 70 (37.84%) knew the most, 52 (28.11%) knew basically, 11 (5.95%) knew little, and 3 (1.62%) did not know at all. Meanwhile, 44 specialists (23.78%) well knew the hospital incentive policies about general practice education, 62 (33.51%) knew the most, 57 (30.81%) knew basically, 18 (9.73%) knew little, and 4 (2.16%) did not know at all. Whether they holding the teaching certificate of general practice was significantly associated with the cognition of national general practice training policy (χ2=14.28, P=0.003) and with their knowledge of residency training program (χ2=16.79, P=0.001), but not associated with knowing the hospital-level incentive policy (χ2=8.18, P=0.075). A total of 170 (91.89%) participants were willing to be clinical teachers of general practice. The reasons for the willingness of teaching were as following: learning more from the teaching in 161 participants (94.71%), expanding sources of patients from rural areas in 102 (60.00%), facilitating promotion in 77 (45.29%), and others in 30 (17.60%). Among 62 specialists holding teaching certificate, 60 (96.77%) were willing to teach general practice residents; while among 123 specialists without teaching certificate, 110 (89.43%) were willing to teach (χ2=4.92, P=0.027). In all hospital incentive policies, promotion of professional titles was most attractive one (82, 44.32%), followed by performance appraisal (63, 34.05%), priority for in-service training (25, 13.51%), and appraisal for excellence award (15, 8.11%). Conclusions:Strengthening trainings for general practice the faculty is helpful to improve their cognition of the general practice residency training programs. And rational hospital incentive policies can enhance the willingness of specialists to teach general practice residency.

Echinacoside Alleviates Cognitive Impairment in Cerebral Ischemia Rats through α 7nAChR-Induced Autophagy / 中国结合医学杂志

OBJECTIVES@#To evaluate the effect of echinacoside (ECH) on cognitive dysfunction in post cerebral stroke model rats.@*METHODS@#The post stroke cognitive impairment rat model was created by occlusion of the transient middle cerebral artery (MCAO). The rats were randomly divided into 3 groups by a random number table: the sham group (sham operation), the MCAO group (received operation for focal cerebral ischemia), and the ECH group (received operation for focal cerebral ischemia and ECH 50 mg/kg per day), with 6 rats in each group. The infarct volume and spatial learning were evaluated by triphenyl tetrazolium chloride staining and Morris water maze. The expression of α7nAChR in the hippocampus was detected by immunohistochemistry. The contents of acetylcholine (ACh), malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD), activities of choline acetyltransferase (ChAT), acetylcholinesterase (AChE), and catalase (CAT) were evaluated by enzyme linked immunosorbent assay. The neural apoptosis and autophagy were determined by TUNEL staining and LC3 staining, respectively.@*RESULTS@#ECH significantly lessened the brain infarct volume and ameliorated neurological deficit in infarct volume and water content (both P<0.01). Compared with MCAO rats, administration of ECH revealed shorter escape latency and long retention time at 7, 14 and 28 days (all P<0.01), increased the α7nAChR protein expression, ACh content, and ChAT activity, and decreased AChE activity in MCAO rats (all P<0.01). ECH significantly decreased MDA content and increased the GSH content, SOD, and CAT activities compared with MCAO rats (all P<0.05). ECH suppressed neuronal apoptosis by reducing TUNEL-positive cells and also enhanced autophagy in MCAO rats (all P<0.01).@*CONCLUSION@#ECH treatment helped improve cognitive impairment by attenuating neurological damage and enhancing autophagy in MCAO rats.

Preparation and anti-tumor activity of a novel antibody-drug conjugate 607-LDM / 药学学报

Antibody-drug conjugates (ADCs) are one of the most important classes of anticancer therapeutics. Human epidermal growth factor receptor-2 (HER2), which is highly expressed in many types of aggressive cancers including breast and ovarian cancer, has been approved as an ideal target for ADCs. Lidamycin (LDM), developed by Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences, is an enediyne-containing antibiotic with potent anti-tumor activity. LDM is a promising payload for ADCs. In the present research, using a special site-directed conjugating technology, we made a novel ADC (607-LDM) with a drug-to-antibody ratio (DAR) of 2 and composed of the anti-HER2 antibody 607 and LDM. The new ADC exhibited potent antitumor activity against human ovarian cancer SKOV3 and breast cancer BT-474 cells. It also induced apoptosis and G2/M arrest. In nude mice with SKOV3 xenografts and a tumor volume of 150-200 mm3, a single intravenous injection 607-LDM at 1 mg·kg-1 induced tumor growth inhibition of 72.4%, which was significant compared to either LDM (50.6%) or antibody (30.2%) treatment alone, or both in combination (50.1%, P < 0.05). All animal experiments were performed in accord with National Regulations and approved by the Animal Experiments Ethical Committee of College of Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences. The novel ADC designed in this study, 607-LDM, is a promising candidate for the treatment of HER2-positive cancers.

Treatment principles and optimization of antihypertensive drugs / 中国实用内科杂志

Throughout the guidelines for hypertension published in China and abroad in recent years, the therapeutic principles of hypertension drug treatment mainly include: hypotensive treatment was given according to the overall risk level of patients with hypertension. Intensive antihypertensive therapy should be adopted to achieve maximum cardiovascular benefits, if conditions permit. Comprehensive intervention is needed for factors other than blood pressure,including treatable risk factors, subclinical target organ damage, and a variety of co-existing clinical diseases.The optimized principles of antihypertensive therapy include initiation of combined drug in most patients to raise the rate of blood pressure treatment targets, and the single-pill combination is preferred in combination therapy.

Effect of Periplaneta Aamericana Polypeptide on Angiogenesis / 中国实验方剂学杂志

Objective: To investigate the effect of the Periplaneta Americana polypeptide on the angiogenesis. Method:Methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay and cell scratch assay were used to observe effect of different concentration (6.25,12.5,25,50,100 mg·L-1) of the Periplaneta Americana polypeptide, CⅡ-3 and skimmed cream on the proliferation and migration of human umbilical vein endothelial cells (HUVECs), and a normal group and a thalidomide group were also established in this study. The tubule formation assay was used to detect the effect of different concentration (25, 50, 100 mg·L-1) of the Periplaneta Americana extracts on the formation of tubules in HUVECs cells. The adhesion between HepG2 cells and HUVECs cells was observed by cell adhesion assay. The expression of vascular endothelial growth factor (VEGF) proteins in HUVECs was detected by immunocytochemical staining and enzyme linked immunosorbent assay (ELISA). Result:MTT results showed that the Periplaneta Americana polypeptide could inhibit the proliferation of HUVECs in a dose-dependent manner (PPPPPPPPPConclusion:The Periplaneta Americana polypeptide can inhibit the invasion, metastasis and tube formation of HUVECs, and down-regulate the expression of VEGF in HUVECs. The effect of Periplaneta Americana polypeptide is better than CⅡ-3 and skimmed cream, and the among the polypeptide, the effect of PAP-2 is superior to the other two.

Effect of Leonurine on p38 MAPK Signaling Pathway and miRNA-1 in Cardiomyocyte Hypertrophy Induced by AngⅡ / 中国实验方剂学杂志

Objective: To investigate the inhibitory effect of leonurine on cardiomyocyte hypertrophy induced by angiotensin Ⅱ(Ang Ⅱ) and its effect on p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway and miRNA-1.Method: Cardiomyocyte hypertrophy was induced by Ang Ⅱ (0.1 μmol·L-1) in primary neonatal cardiomyocytes. Experiments were designed in 6 groups as following:normal group, model group, p38 MAPK inhibitor group (SB203580, 10 μmol·L-1), low-dose(5 μmol·L-1), middle-dose(10 μmol·L-1) and high-dose(20 μmol·L-1) group. The cardiomyocyte surface area was measured by image software, and the protein contents were detected by Lowry. The concentrations of ANP in the culture supernatant were measured by enzyme-linked immunosorbent assay (ELISA). The expression level of miRNA-1 was detected by Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR), and the protein expression levels of endothelin-1 (ET-1), p38 MAPK, p-p38 MAPK, myocyte enhancer factor 2 (MEF2), β-myosin heavy chain (β-MHC), α-myosin heavy chain (α-MHC) were detected by Western blot.Result: Compared with normal group, the surface area of cardiomyocyte, the protein contents, the concentrations of ANP, and the protein expression levels of ET-1, p38 MAPK, p-p38 MAPK, MEF2, β-MHC in model group were higher (Pα-MHC and miRNA-1 were lower than those in normal group (Pβ-MHC in high-dose group were lower (Pα-MHC and miRNA-1 were higher than those in model group (PConclusion: Leonurine (20 μmol·L-1) could inhibit cardiomyocyte hypertrophy induced by AngⅡ, and the mechanism is related to the inhibition of activation of p38 MAPK signaling pathway and up-regulation the expression of miRNA-1.

Salsalate alleviates hyperglycemia by inhibiting endoplasmic reticulum stress in high fat diet induced obese mice / 第二军医大学学报

Objective To explore the effect of salsalate on the glucose metabolism of obese mice induced with high fat diet (HFD). Methods Eight week-old male C57BL/6J mice were fed with HFD in combination with 0.5% of salsalate (SAL group, n＝5) or normal saline (control group, n＝5) for 40 days. The effect of salsalate on serum glucose level was examined by glucose tolerance test (GTT) and insulin tolerance test (ITT). The expressions of endoplasmic reticulumn related proteins, including CCAAT/enhancer-binding protein homologous protein (CHOP), endoplasmic reticulum-localized DnaJ 4 (ERDJ4), glucose regulated protein (GRP)78 and GRP94, were measured by qPCR and Western blotting. Results The random blood glucose level of obese mice were significantly lower in the SAL group than that in the control group (P0.05), and GTT showed that the mice in the SAL group had better glucose tolerance. However, there was no significant difference in fasting insulin level between the two groups. ITT showed there was no difference in the change of blood glucose after insulin stimulation between the two groups. The mRNA expressions of GRP78 and GRP94 and protein expressions of CHOP, ERDJ4, GRP78 and GRP94 were significantly lower in the SAL group than those in the control group (P0.05 or P0.01). Conclusion Salsalate can alleviate the hyperglycemia of obese mice induced with HFD by inhibiting endoplasmic reticulum stress, and the effect is independent of the insulin secretion.

Succession Law of Intestinal Flora after Death in SD Rats / 法医学杂志

OBJECTIVES@#To initially explore the sequential changes in the intestinal flora of corpse for the estimation of postmortem interval （PMI）.@*METHODS@#Rats were sacrificed by cervical dislocation, and samples were taken from their intestines using cotton swab to extract the DNA of intestinal flora. The 16S rRNA V3 universal primers were selected for PCR, and the PCR products were used for denatured gradient gel electrophoresis. The diversity and similarity analysis of intestinal flora were analyzed between groups, and the bands were cut from denaturing gradient gel electrophoresis. After purification, PCR and sequencing, the percentage of major bacteria in each group was obtained.@*RESULTS@#The flora diversity showed a reduced tendency from 1st to 30th day after death （ P<0.05）, while the intra-group similarity showed a downward trend （ P<0.05）. The number of bands and intra-group similarity coefficient （Cs） on the first day was higher than that of other groups （ P<0.05）. The intra-group Cs of the 25th and 30th day had a significant difference compared with the 5th day （ P<0.05）. At the genus level, the intestinal flora was mainly composed of Enterococcus sp. on the 1th and 5th day after death, Bacillus thuringienssis was the dominant species on the 10th, 15th and 20th day, and Enterococcus faecalis became the dominant species on the 25th and 30th day.@*CONCLUSIONS@#The composition and structure of intestinal flora change significantly in rats with the time after death, which indicates that the succession of intestinal flora is related to the postmortem interval.

Determination of Low Level Uranium Isotopes in Vegetation Samples / 分析化学

An analytical method was developed for determination of low-level uranium isotopes in vegetation samples. Dry ashing method was employed to decompose organic matters of vegetation. The sample ash was further digested using multiple acids. Uranium in the prepared sample solution was separated and purified by an extraction chromatography using UTEVA resin. The chemical recovery of uranium in the separation procedure was more than 94% , and more than 99% of Na, K, Ca and other matrix elements and interfering elements were removed. Three natural uranium isotopes were finally measured with high sensitivity ICP-MS/MS. The detection limits of the method for 238U, 235U, 234U were 3. 05, 0. 34 and 0. 04 pg/g, respectively. The detection limits for 238U and 235U were 10 times better than the reported values. Analysis result of U in GBW-10046 standard reference material was in good agreement with reference value, indicating that this method was reliable. The method was successfully applied to determination of uranium isotopes in the vegetation samples collected in Xi′an region, and it was found that the uranium concentrations and isotopic ratios in these vegetation samples fall well into natural level, and there was no significant artificial uranium contamination. This was the first survey of the three natural uranium isotopes in vegetation samples in this region.

Clinical Distribution,Drug Resistance and Drug-resistance Genes of Acinetobacter baumannii in Our Hospital during 2014-2017 / 中国药房

OBJECTIVE:To provide reference for rational drug use in clinic and nosocomial infection control. METHODS:Acinetobacter baumannii(AB)were collected from our hospital during Jan. 2014-Jun. 2017. Drug sensitivity tests were conducted by using K-B method and MIC method. Drug-resistance genes of multidrug-resistant Acinetobacter baumannii(MDR-AB)were amplified by PCR,and compared with GenBank database by using Blast comparison. RESULTS:A total of 1 758 strains of AB were detected,and mainly came from sputum and throat swab(65.24%),followed by urine(18.49%). These infected patients were mainly distributed in the departments of ICU(38.51%)and respiratory medicine(24.00%),respectively. Drug resistance of clinical isolated AB to most commonly used antibiotics were more than 40%,such as compound sulfamethoxazole,piperacillin sodium and tazobactam sodium,gentamicin,cefepime,levofloxacin,minocycline,imipenem,etc.;it had increased year after year. Drug resistance to colistin was lower than 5% and decreased year by year.A total of 673 strains of MDR-AB were detected, and detection rates were 22.77%,29.82%,52.09%,54.33%,respectively.Among 110 strains of MDR-AB,detection rates of TEM, AmpC,IMP,VIM,OXA-23,OXA-24,OXA-51,aac(6′)-Ⅰ,aac(3)-Ⅰ,ant(3″)-Ⅰ,anmA,gyrA,parC gene were 97.27%, 91.82%,49.09%,12.73%、90.91%,12.73%,98.18%,34.55%,60.91%,89.09%,87.27%,77.27%,82.73%,respectively. Results of Blast comparison showed that point mutation occurred in 83rd and 121st base of gyrA gene,144th base of parC gene. CONCLUSIONS:AB mainly come from sputum and throat swab specimens in our hospital,and infected patients are mainly distributed in the departments of ICU and respiratory medicine. Drug resistance is serious,and the detection rate of MDR-AB is increased year by year. Main genes of multidrug-resistant strains mainly include TEM,AmpC,OXA-23,OXA-51,ant(3″)-Ⅰ, anmA,etc.,and mutation of gyrA and parC gene are found. It is necessary to strengthen the management of classification use of antibiotics and strengthen the monitoring of AB drug resistance. According to the results of drug sensitivity test,antibiotics are selected rationally to prevent or delay planting and cross transmission of AB-resistant strain.

Capecitabine combined with docetaxel on tumor marker levels in patients with advanced breast cancer / 国际检验医学杂志

Objective To explore the effects of combination of capecitabine and docetaxel for the treatment on advanced breast cancer and their influence on tumor biomarkers.Methods The 75 cases with advanced breast cancer were randomly recruited from December 2010 to December 2015 in our hospital,and they were divided into the observation group (38 cases) and control group (37 cases) according to the admission time,the patients in control group were treated with docetaxel,while patients in the observation group were treated with combination of capecitabine and docetaxel,the clinical efficacy before and after treatment was observed,and the changes of serum CEA,glucose CA125,CA15-3 and adverse reaction were compared between two groups.Results The effective rate (RR) of the control group was 40.54% (15/37),and the disease control rate (DCR) was 67.57% (25/37).The RR was 63.16% (24/38) and DCR was 86.84% (33/38)in the observation group.There was significant difference between the two groups (P0.05);After treatment,the levels of CEA,CA125 and CA15-3 in both two groups were significantly lower than those before treatment (P<0.05);Moreover,after treatment,the observation group of three tumor biomarkers were significantly lower than those of the control group (P<0.05).Conclusion Combination of capecitabine and docetaxel in the treatment of advanced breast cancer can reduce the serum tumor biomarkers significantly,attenuate the side effects,and the patients are in the good tolerance,it can be widely recommended in clinical use.

Effect of dihydromyricetin on proliferation and apoptosis of breast cancer MCF-7 cells / 国际检验医学杂志

Objective To investigate the effect of dihydromyricetin on proliferation and apoptosis of breast cancer MCF-7 cells.Methods From March 2014 to February 2015,breast cancer MCF-7 cells were treated with 99% pure DMY as an inhibitor.MTT assay,flow cytometry and immunocytochemistry were used to analyze the proliferation,apoptosis and protein expression of breast cancer cell MCF-7.Results When the DMY concentration was higher than 20 μg/mL,the inhibitory effect appeared,but not good.When 40 and 80 μg/mL DMY were used,the proliferation of MCF-6 cells were significantly inhibited,and have different degrees of sensitivity to it.When DMY was 80 μg/mL,the IC50 was 226.9 μg/mL.The inhibition rate and IC50 were compared with 0 μg/mL DMY,there was significant difference(P 50%,especially in DMY with 80 μg/mL,the positive rate was 10.00%.Compared with 0 μg/mL DMY,the difference was significant(P<0.05).Conclusion The use of dihydromyricetin in breast cancer patients can effectively inhibit the rapid increase of cancer cells,accelerate apoptosis,slow down the patient′s condition,the effect is outstanding.

Prethrombotic status and long-time thromboembolic events in primary hypertensive patients with or without elevated homocysteine level / 中华心血管病杂志

<p><b>OBJECTIVE</b>To evaluate the association between homocysteine level and prethrombotic status and long-term thromboembolic events in patients with primary hypertension.</p><p><b>METHODS</b>Results between 110 hypertensive patients with elevated homocysteine (HCY) level were compared with 110 hypertensive patients with normal HCY level which were enrolled from October 2003 to November 2009. Fibrinogen (FIB), viscosity, thrombomodulin (TM), granule membrane protein (GMP-140), prethrombin F1+2 fragment (F1+2), D-dimer fragment (D-Dimer) and antithrombin III (AT-III) were measured and correlated to HCY and prethrombotic state. The endpoints of the study were arterial and venous thromboembolic events. The variables linked with arterial and venous thromboembolic events were included in Cox proportional hazard models. The event-free survival was illustrated with Kaplan-Meier survival curves and compared by the Log-rank test.</p><p><b>RESULTS</b>The patients were followed up for 8-122 months (median follow-up time was 85 months). Compared with hypertensive patients with normal HCY, the plasma level of TM ((4.8±1.2) µg/L vs. (4.5±1.0) µg/L, P = 0.045), GMP-140 ((18.8±3.2) µg/L vs. (17.1±4.3) µg/L, P = 0.001), F1+2 ((1.2±0.4) nmol/L vs. (1.0±0.6) nmol/L, P = 0.004) were significantly higher while the plasma level of AT-III ((95.3±10.4) % vs. (98.6±10.6)%, P = 0.021) was significantly lower in hypertensive patients with elevated HCY level. FIB, viscosity of plasma and D-dimer were similar between the two groups. Multiple regression analyses indicated that HCY level was negatively correlated with AT-III (β = -0.199, P = 0.011) and positively correlated with age (β = 0.217, P = 0.04), female gender (β = 5.667, P = 0.001) and TM (β = 2.341, P = 0.003). Cox multivariate analysis revealed that age and HCY level were independent prognostic risk factors of thromboembolic events (OR 1.046, 95% CI 1.013-1.082, OR 1.052, 95% CI 1.027-1.078, respectively) (all P < 0.05). Kaplan-Meier curves showed that there was a significant difference in the event-free survival between the two groups (Log-rank test, P = 0.027).</p><p><b>CONCLUSIONS</b>Compared with normal HCY hypertensive patients, the levels of plasma prothrombin activators such as TM, GMP-140 and F1+2 were significant increased and anti-thrombin factor such as AT-III was significant decreased in hypertensive patients with elevated HCY. Old age and high HCY level were independent prognostic risk factors of thromboembolic events. The event-free survival in hypertensive patients with elevated HCY is lower than in hypertensive patients with normal HCY level.</p>

Intraductal papilloma of parotid gland in 1 case / 实用口腔医学杂志

1 case of intraductal papilloma of parotid gland was analyzed by means of clinicopathologic data,hematoxylin-eosin and immuno-histochemical staining.Combined with the relevant literature,clinical pathological features,diagnosis and treatment of the salivary gland intra-ductal papilloma were discussed.

Comparison of direct immune-fluorescent assay and real-time quantitative PCR in detecting the Hantavirus / 中华预防医学杂志

<p><b>OBJECTIVE</b>To compare the differences between the direct immuno-fluorescent assay (DFA) and real-time quantitative PCR in detecting the Hantavirus (HV) in rat lungs.</p><p><b>METHODS</b>From April to October in 2012, a total of 479 rats were caught by mouse-trap in residential or wild areas in Huxian, Jingyang, and Meixian of Shaanxi province, where haemorrhagic fever with renal syndrome (HFRS) was highly prevalent. The rats were dissected to take the two lungs, one was frozen and applied immuno-fluorescent assay to detect HV antigen while the other one was extracted its RNA and detected HV nucleic acid by real-time quantitative PCR. Then we compared the positive rate of the two methods.</p><p><b>RESULTS</b>Out of the 479 rats, 105 were caught from residential areas and the other 374 were caught from wild areas. Among the 105 rats caught from residential areas, no HV were detected out neither by DFA nor by real-time quantitative PCR. Among the 374 wild rats, 13.1% (49/374) were detected HV positive by DFA and 14.7% (55/374) were detected HV positive by real-time quantitative PCR. The difference showed no statistical significance (χ(2) = 0.402, P = 0.526). When detecting each lung sample, the HV positive rate was 10.2% (49/479) under the detection by DFA while the HV positive rate was 11.5% (55/479) under the detection by real-time quantitative PCR. The difference had no statistical significance (χ(2) = 1.286, P = 0.257) and the consistency coefficient was 68.2% under the paired chi-square test analysis, which showed high consistency (u = 11.759, P < 0.05). The sensitivity of real-time quantitative PCR to detect HV was 77.6% (38/49) comparing with DFA as standard, and the specificity was 96.1% (413/430). Out of the 9 suspected HV positive sample detected by DFA, 6 were confirmed positive by real-time quantitative PCR and 3 were denied.</p><p><b>CONCLUSION</b>Compared with the DFA, real-time quantitative PCR could also be used to detect the infection of HV in rats, and the result might be more stable.</p>

SNP in differentially methylated region upstream of H19 gene in Chinese Korean nationality / 法医学杂志

OBJECTIVE@#To investigate SNP and distribution of haplotypes in differentially methylated region (DMR) upstream of H19 gene in Chinese Korean nationality in order to provide basic data for forensic application and population genetics research.@*METHODS@#One hundred and one blood samples from unrelated Chinese Korean individuals and 14 blood samples from 5 Chinese Korean intergenerational families which known genetic relationship were collected. The SNP in DMR upstream of H19 gene were investigated by PCR-cycle sequencing and McrBC digestion followed by PCR. The haplotypes detected by parentally imprinted allele (PIA) method and relevant genetic parameters were calculated.@*RESULTS@#Thirteen SNPs (rs10840167, rs2525883, rs12417375, rs4930101, rs2525882, rs2735970, rs2735971, rs11042170, rs2735972, rs10732516, rs2071094, rs2107425, and rs4930098) and five haplotypes were detected in 1 174 bp target product in DMR upstream of H19 gene, with 9 SNPs having high discrimination power as good genetic markers. The average gene diversity (GD) of haplotypes was 0.714. The maternal haplotype was confirmed correctly by PIA method from McrBC-digested products of genomic DNA.@*CONCLUSION@#High polymorphisms exist in DMR upstream of H19 gene in Chinese Korean nationality. And determination of the maternal haplotype could furthermore enhance the forensic identification efficiency of imprinted gene.

Expression of cytokines in mouse hepatitis B virus X gene-transfected model / 华中科技大学学报（医学）（英德文版）

The expression profile in the mouse hepatitis B virus X (HBx)-transfected model was investigated in order to lay a foundation for further study on the implication of cytokines expression in hepatitis B virus (HBV) infection. Hydrodynamic injection method via the tail vein was used to establish the animal HBx-transfected model. By using microassay, the differential expression of gene in each group was analyzed, which was further confirmed by using real-time PCR and semi-quantitative PCR. Most of chemokine genes such as Ccl2, Ccl5, Ccl9, MIG and IP-10 were up-regulated in the HBx-transfected mouse model versus the control mice, which was coincided with the microarray results. Western blotting and immunohistochemistry were applied to detect the expression of MIG and IP-10 in the liver tissues. Simultaneously, ELISA was adopted to measure the content of IFN-γ in the liver tissues. DNA microassay revealed that the expression of 611 genes changed in HBx-transfected mice as compared with that in pCMV-tag2B-transfected mice, and most of the screened chemokines were up-regulated (including MIG and IP-10). Additionally, IFN-γ protein levels were increased by 20.7% (P<0.05) in pCMV-tag2B-HBx-transfected mice as compared with the untreated mice. IFN-γ protein levels were reduced by 53.9% (P<0.05) in pCMV-tag2B-transfected mice as compared with the untreated mice, which was consistent with the up-regulation of MIG and IP-10. It was suggested HBx transfection could induce the expression of MIG and IP-10 in the liver tissues, which might play the roles in HBV-related liver immunity and cytokines-mediated antiviral effect.

Expression of cytokines in mouse hepatitis B virus X gene-transfected model / 华中科技大学学报（医学）（英德文版）

The expression profile in the mouse hepatitis B virus X (HBx)-transfected model was investigated in order to lay a foundation for further study on the implication of cytokines expression in hepatitis B virus (HBV) infection. Hydrodynamic injection method via the tail vein was used to establish the animal HBx-transfected model. By using microassay, the differential expression of gene in each group was analyzed, which was further confirmed by using real-time PCR and semi-quantitative PCR. Most of chemokine genes such as Ccl2, Ccl5, Ccl9, MIG and IP-10 were up-regulated in the HBx-transfected mouse model versus the control mice, which was coincided with the microarray results. Western blotting and immunohistochemistry were applied to detect the expression of MIG and IP-10 in the liver tissues. Simultaneously, ELISA was adopted to measure the content of IFN-γ in the liver tissues. DNA microassay revealed that the expression of 611 genes changed in HBx-transfected mice as compared with that in pCMV-tag2B-transfected mice, and most of the screened chemokines were up-regulated (including MIG and IP-10). Additionally, IFN-γ protein levels were increased by 20.7% (P<0.05) in pCMV-tag2B-HBx-transfected mice as compared with the untreated mice. IFN-γ protein levels were reduced by 53.9% (P<0.05) in pCMV-tag2B-transfected mice as compared with the untreated mice, which was consistent with the up-regulation of MIG and IP-10. It was suggested HBx transfection could induce the expression of MIG and IP-10 in the liver tissues, which might play the roles in HBV-related liver immunity and cytokines-mediated antiviral effect.