Study progress of microRNAs in aplastic anemia / 中华实用儿科临床杂志

Aplastic anemia (AA) refers to a life-threatening bone marrow failure disorder.With respect to the precise pathophysiology of AA at present, it is still unclear.MicroRNA (miRNA), about 22 nucleotides in length, is a kind of small RNAs and it can regulate gene expression post-transcriptionally.According to domestic and foreign reports recently, there are abnormal expressions of several miRNAs in AA patients, suggesting that miRNAs may be involved in the development and progression of AA.This article reviews the study progress of miRNAs in AA.

Analysis of clinical features and genetic variants in a child with creatine deficiency syndrome / 中华医学遗传学杂志

OBJECTIVE@#To explore the clinical features and genetic basis for a patient diagnosed with creatine deficiency syndrome (CDS).@*METHODS@#The patient was subjected to whole exome sequencing (WES). Candidate variant was verified by Sanger sequencing. The level of creatine was determined by using a magnetic resonance spectrum (MRS) method.@*RESULTS@#The patient presented with development delay and poor response to stimuli. No obvious abnormality was found with his muscle tone and strength of his limbs. Borderline EEG was detected. MRI showed abnormal development of the white matter and dysplasia of corpus callosum. Urine organic acid screening has shown increased glycerin-3-phosphate. WES revealed that the patient has carried compound heterozygous variants of the GAMT gene, namely c.412C>T and IVS4-1G>A, which were respectively derived from his mother and father. MRS showed reduced creatine in bilateral basal ganglia. Functional study of the splicing site suggested that the IVS4-1G>A variant has resulted skipping of exon 5 upon splicing.@*CONCLUSION@#The compound variants of the GAMT gene probably underlay the disease in this child. Above finding has enriched the spectrum of GAMT gene variants.

Analysis of 8 274 cases of novel coronavirus nucleic acid detection and co-infection in Wuhan / 中华检验医学杂志

Objective:To investigate the positive rate for 2019-nCoV tests and co-infections in Wuhan district.Methods:A total of 8 274 cases in Wuhan were enrolled in this cross-sectional study during January 20 to February 9 in 2020, and were tested for 2019-nCoV using fluorescence quantitative PCR. Both respiratory tract samples (nasopharynx, oropharynx, sputum and alveolar lavage fluid) and non-respiratory tract samples (urine, feces, anal swabs, blood and conjunctival sac swabs) were collected. If both orf1ab and N genes are positive, they are classified as nucleic acid test positive group; if both orf1ab and N genes are negative, they are classified as negative group; if single gene target is positive, they are classified as suspicious group. Individuals were divided into male group and female group according to sex. At the same time, 316 patients were tested for 13 respiratory pathogens by multiplex PCR.Results:Among the 8 274 subjects, 2 745 (33.17%) were 2019-nCoV infected; 5 277 (63.77%) subjects showed negative results in the 2019-nCoV nucleic acid test; and 252 cases (3.05%) was not definitive (inconclusive result). The age of cases with COVID-19 patients and inconclusive cases was significantly higher than that of cases without 2019-nCoV infection (56>40, t=27.569, P<0.001; 52>40, t=6.774, P<0.001). The positive rate of 13 respiratory pathogens multiple tests was significantly lower in 104 subjects who were positive for 2019-nCoV compared with those in subjects who were negative for 2019-nCoV test (5.77% vs 18.39%, χ 2=24.105, P=0.003). Four types of respiratory tract samples and five types of non-respiratory tract sampleswere found to be positive for 2019-nCoV nucleic acid test. Conclusion:The 2019-nCoV nucleic acid positive rate inmale is higher than infemale. Co-infections should be pay close attention in COVID-19 patients. 2019-nCoV nucleic acid can be detected in non-respiratory tract samples.

Analysis of 8 274 cases of new coronavirus nucleic acid detection and co-infection in Wuhan / 中华检验医学杂志

Objective@#To investigate the positive rate for 2019-nCoV tests and co-infections in Wuhan district.@*Methods@#A total of 8 274 cases in Wuhan were enrolled in this cross-sectional study during January 20 to February 9, 2020, and were tested for 2019-nCoV using fluorescence quantitative PCR. Both respiratory tract samples (nasopharynx, oropharynx, sputum and alveolar lavage fluid) and non-respiratory tract samples (urine, feces, anal swabs, blood and conjunctival sac swabs) were collected. If both orf1ab and N genes are positive, they are classified as nucleic acid test positive group; if both orf1ab and N genes are negative, they are classified as negative group; if single gene target is positive, they are classified as suspicious group. Individuals were divided into male group and female group according to sex. At the same time, 316 patients were tested for 13 respiratory pathogens by multiplex PCR.@*Results@#Among the 8 274 subjects, 2 745 (33.2%) were 2019-nCoV infected; 5 277 (63.8%) subjects showed negative results in the 2019-nCoV nucleic acid test; and 252 cases (3.05%) was not definitive (inconclusive result). The age of cases with COVID-19 patients and inconclusive cases was significantly higher than that of cases without 2019-nCoV infection (40 vs 56, t=27.569, P<0.001; 52 vs 56, t=6.774, P<0.001). The positive rate of 13 respiratory pathogens multiple tests was significantly lower in 104 subjects who were positive for 2019-nCoV compared with those in subjects who were negative for 2019-nCoV test (5.77% vs 18.39%, χ2=24.105, P=0.003). Four types of respiratory tract samples and five types of non-respiratory tract samples were found to be positive for 2019-nCoV nucleic acid test.@*Conclusion@#The 2019-nCoV nucleic acid positive rate in male is higher than in female. Co-infections should be pay close attention in COVID-19 patients. 2019-nCoV nucleic acid can be detected in non-respiratory tract samples.

Protective effect of protein kinase C inhibitor on rat renal vascular endothelial injury induced by lipopolysaccharide / 中华危重病急救医学

OBJECTIVE@#To investigate the protective effect of protein kinase C (PKC) inhibitor rottlerin on rat renal vascular endothelial injury induced by lipopolysaccharide (LPS).@*METHODS@#Rat renal microvascular endothelial cells cultured for 3-6 generations were divided into three groups according to random number table: blank control group in which cells were not challenged, LPS group in which cells were only stimulated by LPS 10 mg/L for 24 hours, and PKC inhibitor group in which cells were treated with PKC inhibitor rottlerin 2 μmol/L 30 minutes before LPS stimulation. The levels of tumor necrosis factor-α (TNF-α) and interleukins (IL-1β, IL-8) were determined by enzyme-linked immunosorbent assay (ELISA). Monolayer permeability was determined by Transwell assay. The expressions of PKC, RhoA and vascular endothelial-cadherin (VE-cadherin) were detected by Western Blot. The morphological characteristic and distribution of F-actin was measured by laser confocal fluorescence microscope.@*RESULTS@#Compared with blank control group, the levels of inflammatory cytokines at 24 hours after 10 mg/L LPS stimulation were significantly increased in LPS group [TNF-α (ng/L): 397.3±25.4 vs. 46.8±8.9, IL-1β (ng/L): 76.7±11.2 vs. 12.6±3.2, IL-8 (ng/L): 574.5±31.4 vs. 73.2±9.6, all P < 0.05], the permeability of endothelial cells was significantly increased (A value: 1.32±0.03 vs. 0.36±0.02, P < 0.05), while the expressions of PKC and RhoA were significantly up-regulated (PKC/β-actin: 0.88±0.02 vs. 0.61±0.03, RhoA/β-actin: 0.96±0.01 vs. 0.49±0.03, both P < 0.05), VE-cadherin expression was significantly down-regulated (VE-cadherin/β-actin: 0.51±0.01 vs. 0.72±0.04, P < 0.05), and the F-actin distribution disorder had obvious stress fiber formation. Compared with LPS group, the levels of inflammatory cytokines were significantly lowered in PKC inhibitor group [TNF-α (ng/L): 127.4±14.6 vs. 397.3±25.4, IL-1β(ng/L): 43.2±7.8 vs. 76.7±11.2, IL-8 (ng/L): 212.7±18.2 vs. 574.5±31.4, all P < 0.05], the permeability of endothelial cells was significantly decreased (A value: 0.81±0.02 vs. 1.32±0.03, P < 0.05), the expressions of PKC and RhoA were significantly down-regulated (PKC/β-actin: 0.44±0.03 vs. 0.88±0.02, RhoA/β-actin: 0.63±0.05 vs. 0.96±0.01, both P < 0.05), the VE-cadherin expression was significantly up-regulated (VE-cadherin/β-actin: 0.69±0.03 vs. 0.51±0.01, P < 0.05), and the F-actin remodeling and stress fiber formation were significantly reduced.@*CONCLUSIONS@#PKC inhibitor could significantly attenuate the damage of vascular endothelial barrier induced by LPS, and plays an important role in endothelial cell barrier.

Protective effect of protein kinase C inhibitor on rat renal vascular endothelial injury induced by lipopolysaccharide / 中华危重病急救医学

Objective To investigate the protective effect of protein kinase C (PKC) inhibitor rottlerin on rat renal vascular endothelial injury induced by lipopolysaccharide (LPS). Methods Rat renal microvascular endothelial cells cultured for 3-6 generations were divided into three groups according to random number table: blank control group in which cells were not challenged, LPS group in which cells were only stimulated by LPS 10 mg/L for 24 hours, and PKC inhibitor group in which cells were treated with PKC inhibitor rottlerin 2 μmol/L 30 minutes before LPS stimulation. The levels of tumor necrosis factor-α (TNF-α) and interleukins (IL-1β, IL-8) were determined by enzyme-linked immunosorbent assay (ELISA). Monolayer permeability was determined by Transwell assay. The expressions of PKC, RhoA and vascular endothelial-cadherin (VE-cadherin) were detected by Western Blot. The morphological characteristic and distribution of F-actin was measured by laser confocal fluorescence microscope. Results Compared with blank control group, the levels of inflammatory cytokines at 24 hours after 10 mg/L LPS stimulation were significantly increased in LPS group [TNF-α (ng/L): 397.3±25.4 vs. 46.8±8.9, IL-1β(ng/L): 76.7±11.2 vs. 12.6±3.2, IL-8 (ng/L): 574.5±31.4 vs. 73.2±9.6, all P < 0.05], the permeability of endothelial cells was significantly increased (A value: 1.32±0.03 vs. 0.36±0.02, P < 0.05), while the expressions of PKC and RhoA were significantly up-regulated (PKC/β-actin: 0.88±0.02 vs. 0.61±0.03, RhoA/β-actin: 0.96±0.01 vs. 0.49±0.03, both P < 0.05), VE-cadherin expression was significantly down-regulated (VE-cadherin/β-actin: 0.51±0.01 vs. 0.72±0.04, P < 0.05), and the F-actin distribution disorder had obvious stress fiber formation. Compared with LPS group, the levels of inflammatory cytokines were significantly lowered in PKC inhibitor group [TNF-α (ng/L): 127.4±14.6 vs. 397.3±25.4, IL-1β(ng/L): 43.2±7.8 vs. 76.7±11.2, IL-8 (ng/L): 212.7±18.2 vs. 574.5±31.4, all P < 0.05], the permeability of endothelial cells was significantly decreased (A value: 0.81±0.02 vs. 1.32±0.03, P < 0.05), the expressions of PKC and RhoA were significantly down-regulated (PKC/β-actin: 0.44±0.03 vs. 0.88±0.02, RhoA/β-actin: 0.63±0.05 vs. 0.96±0.01, both P < 0.05), the VE-cadherin expression was significantly up-regulated (VE-cadherin/β-actin: 0.69±0.03 vs. 0.51±0.01, P < 0.05), and the F-actin remodeling and stress fiber formation were significantly reduced. Conclusion PKC inhibitor could significantly attenuate the damage of vascular endothelial barrier induced by LPS, and plays an important role in endothelial cell barrier.

The clinical characteristics and gene diagnosis of Rubinstein-Taybi syndrome / 临床儿科杂志

Objective To explore the clinical and genetic features of Rubinstein-Taybi syndrome (RSTS). Methods The clinical data of 2 children with RSTS were reviewed and analyzed. Results Two male children (3 years old and 4 months old) were admitted to hospital because of growth retardation. Both of them were characterized by short stature, language and motor retardation, excessive hairiness and cryptorchidism. Case 1 had slightly broad thumbs and toes, and case 2 had distinctive facial features of high arched palate, broad nasal bridge, ptosis, and obviously broad thumbs and toes. Cardiac dysplasia was found in both of them by echocardiography. The c.152T>G (L51X) heterozygous mutation was found in case 1 by high throughput sequencing and genomic chip technology, and this mutation has not been reported. Deletion of 2.5 Mb in chromosome 16p13.3 region was found in case 2. Conclusions The main clinical manifestations of RSTS are excess hair, deformity of thumbs and toes, deformity of the heart development, and growth retardation. Molecular detection can help the clinical diagnosis.

Diterpene ginkgolides meglumine injection inhibits oxidative stress induced by oxygen-glucose deprivation by activating Akt/Nrf2 pathway in SH-SY5Y cells / 中国药理学与毒理学杂志

OBJECTIVE To investigate the protective effects and mechanism of diterpene ginkgolides meglumine injection (DGMI) against oxidative stress induced by oxygen-glucose deprivation (OGD) in SH-SY5Y cells. METHODS SH-SY5Y cells were divided into five groups: normal control, model control (OGD group) and drug(25 mg · L- 1) administration groups including DGMI group, extract of ginkgo biloba leaves injection group (EGBLI) and lactones ginkgo biloba injection group (LGBI). The cells suffered from oxygen-glucose deprivation (OGD) for 4 h, followed by reoxygenation with drugs for 6 h. Then, cell viabilities were detect using CCK-8 assays, reactive oxygen species (ROS) levels using fluorescence probe DCFH-DA and superoxide dismutase (SOD) activities using WST-1 test. Western blotting was used to detected protein levels of hemeoxygenase-1(HO-1), NAD(P)H, quinone oxidore?ductase l (Nqo1), protein kinase B (Akt), phosphorylated Akt (p-Akt), nuclear factor-E2-related factor2 (Nrf2) and phosphorylated Nrf2 (p-Nrf2). The cells were induced by OGD for 4 h, followed by reoxygen?ation and DGMI for 1 h, combined with different concentrations of PI3K inhibitor (LY294002) (at the final concentration of 12.5, 25 and 50 μmol · L-1) before the protein levels of AKT, p-AKT, Nrf2 and p-Nrf2 were detected by Western blotting. RESULTS SH-SY5Y cells induced by OGD for 4 h resulted in an increase in ROS(P<0.01), but a decrease in cell viabilities(P<0.01), SOD activities(P<0.01), and antioxidant protein levels ( Akt, p-Akt, Nrf2, p-Nrf2, HO-1 and Nqo1) (P<0.01). Compared with OGD group, treatment with reoxygenation and drugs (DGMI,EGBLI and LGBI respectively) for 6 h resulted in a decrease in ROS (P<0.01), but an increase in cell viabilities, SOD activities and antioxidant protein levels of p-Nrf2, HO-1, Nqo1 and p-Akt(P<0.05,P<0.01). DGMI group showed the best efficiently. Moreover, after OGD for 4 h, compared with DGMI group, combining reoxygenation and DGMI with LY294002 for 1 h resulted in a concentration-dependent inhibition of the protein levels of p-AKT and p-Nrf2(P<0.01). CONCLUSION DGMI 25 mg · L-1 can inhibit oxidative stress in SH-SY5Y cells induced by OGD by increasing the activity and expression of Nrf2 through PI3K/Akt pathway, which may be one of the mechanisms by which DGMI protects neurons from stroke.

LC-MS based measurement of androgens and its clinical significance in diagnosis of Sj(o)gren's syndrome / 中国基层医药

Objective To evaluate the clinical value of 6 androgens in the serum of Sj?gren's syndrome (SS) measured by liquid chromatography tandem mass spectrometry (LC-MS).Methods 32 SS patients (SS group) and 25 healthy people (healthy group) were included in this study.6 androgens in the serum were analyzed by LC-MS after prepared.PCA,PLS-DA models and t-test were used to class differentiation of androgens between two groups.Results The results of PLS-DA showed that SS group and healthy group could be well classed by 6 androgens.The levels of testerone (T),dihydrotestosterone (DHT),dehydroepiandrosterone (DHEA),androsterone and DHEAS in SS group were significantly lower than those in healthy group (t=8.536,2.438,3.172,4.158,4.489,all P<0.05).The samples before or after menopause could be distinguished between the two groups by PLS-DA.Conclusion The significant difference of androgens was discovered between SS patients and healthy people via the measurement of 6 androgens in serum.It may arise a new idea for diagnosis and treatment of SS.

Comparision of the Efficiency of Different Promoters Driving the Expression of CD19-specife Chimeric Antigen Receptor (CAR)-redirected T Lymphocytes Mediated by Lentiviral Vector / 医学研究杂志

Objective Three kinds of different internal promoters were inserted into the lentiviral vector to drive the expression of CD19-specifc chimeric antigen receptor (CAR)-redirected T lymphocytes with the green fluorescent protein(GFP) and the GFP expression efficiency of transduced cells driven by lentivirus were compared.Methods Three GFP reporter lentivirus vectors carrying different promoters and anti-CD19-CAR,including CMV,EF1α,and CMV-EF1α were selected.Three different vectors' names abbreviate to pHAGE-CMV-EFlα-EGFP-CD19,pHAGE-CMV-EGFP-CD19,pHAGE-EF1α-EGFP-CD19.Human embroic kideny 293T cells were cotransfected with the three plasmids by calcium phosphate DNA precipation.And the expression of GFP was observed under fluorescent microscope after transfecting 293T cells,and virus supernatant was collected after 72 h and centrifuged.Nucleic acid copy number in RNA and the expression of EGFP and CD3 zeta were identified through fluorescence quantitative PCR,flow cytometry and western blotting,respectively.The titers of the lentiviral vectors were determinde by scoring GFP expression following swerial dilutions of the viral supernatant on 293T cells.T lymphocytes was infected with lentivims produced from these reporter vectors.Then,fluorescence microscope,wesyern blotting were used to detect the GFP expression strength.Results Results of fluorescence microscope maintained that different internal promotors driving the expressing effect is different.293T as targeted cell,the transfected 293T cells were found containing strong expression of GFP.The amount of the 293T cells expressing GFP driven by CMV-EF1α promoter was the largest,while the EF1α promoter was the smallest.Flow cytometry results show that the rate of transduction is 72.4％,20.6％ and 14.5％,respectively.T lymphocytes as targeted cell,Western blot showed that CMV-EF1α was the largest among all promoters,while the CMV was the smallest.Conclusion In the study of gene expression mediated by lenuvirus,proper cell lines and promoters should be selected to obtain high efficiency.

The Pathoenesis of HuangQin Tang / 浙江中医药大学学报

Objective] The pathogenesis of HuangQin Tang is discussed little in six meridians syndrome differentiation, and which meridian syndrome does it belong to isn’t clear. So this article tries to discuss the belonging and pathogenesis of it. And the key point of using it in clinic will also be involved.[Method] Combining the observation of efficacy of XiaoChaiHu Tang and HuangQin Tang and the elaboration articles are related to that two prescriptions to analyze the difference of that two prescriptions in pathogenesis.[Result]It’s found that HuangQin Tang and XiaoChaiHu Tang all belong to ShaoYin in six meridian syndrome.But their pathogenesis is different. HuangQin Tang should be used in heat syndrome of ShaoYang disease which caused by heat pathogen of ShaoYang meridian and diarrhea will appear because belly is shocked. However, XiaoChaiHu Tang is used to drive cold pathogen of ShaoYang meridian. [Conclusion]Both of HuangQin Tang and XiaoChaiHu Tang belong to ShaoYang Meridian, but it’s known that HuangQin Tang is used less. And less discussion of HuangQin Tang should be blamed. The more we know on pathogenesis of HuangQin Tang, the better we use it.

Involvement of p38-p53 signal pathway in neuroprotective effects of DGMI on SH-SY5 Y cells damaged by oxygen-glucose deprivation / 中国药理学通报

Aim To investigate the protective effects of Diterpene Ginkgolides Meglumine Injection(DGMI)on SY5 Y cells damaged by oxygen-glucose deprivation and its functional mechanisms.Methods After 4 h of OGD,the cells were treated with 25 mg·L-1 drugs for 1 h.Subsequently,cell viabilities were measured by cell counting kit-8(CCK-8 kit)and cell apoptosis was measured by flow cytometric analysis.Furthermore, the mitochondrial membrane potential was detected by rhodamine123 staining.The levels of phospho-p38, phospho-p53,Bcl-2,Bax and cleaved caspase-9/3 were evaluated by western blot.Results DGMI signif-icantly increased the cell viabilities of SY5 Y cells dam-aged by OGD,and reduced OGD-elicited dissipation of mitochondrial membrane potential and cell apoptosis. Furthermore,DGMI also reduced p-p38,p-p53,Bax/Bcl-2 ratio,cleaved caspase-9 and cleaved caspase-3. Conclusion DGMI shows good neuroprotective effects on SY5 Y cells after oxygen-glucose deprivation.The underlying mechanisms may be associated with the sup-pression of p38/p53/Bcl-2 /caspase-9/caspase-3 sig-naling pathway.

Diterpene ginkgolides meglumine injection attenuates oxygen-glucose deprivation-induced apoptosis of nerve cells via inhibition of calpain signaling pathway / 中国药科大学学报

@#To investigate the anti-apoptotic effect of diterpene ginkgolides meglumine injection(DGMI)on SH-SY5Y cells induced by oxygen-glucose deprivation/reoxygenation(OGD/R), and to explore its mechanisms. After 4 h of OGD, the SH-SY5Y cells were treated with 25 mg/L DGMI for 1 h. The release of lactic dehydrogenase(LDH)was measured by cytotoxicity detection kitplus. Cell apoptosis was detected by caspase-3/7 assays. Cell death was detected by ELISA. The concentration of ［Ca2+］i in cytoplasm was measured by Fluo-3 AM and the levels of calpain and cleaved capaease-12 were evaluated by western blot. As we expected, DGMI significantly decreased the release of LDH, the concentration of ［Ca2+］i, the protein levels of calpain and cleaved caspase-12. Furthermore, DGMI injection also attenuated the activities of caspase-3/7 and the contents of cytoplasmic histone-associated- DNA-fragments. These data demonstrated that the DGMI injection showed good anti-apoptotic effect in SH-SY5Y cells induced by OGD/R. The mechanisms may be associated with the inhibition of Ca2+/calpain/caspase-12/caspase-3 signaling pathway.

A Retrospective Analysis of Human Parvovirus B19 Infection in Childhood Immune Thrombocytopenia / 医学研究杂志

Objective To confirm the role of human parvovirus B 19 ( B19) infection in childhood immune thrombocytopenia ( ITP) patients.Methods A total of 416 cases of newly diagnosed childhood ITP patients from Jan .2011 to Dec.2013 had been summarized to be cases group , Then a total of 130 childhood patients with common respiratory tract infection were selected randomly as the control group . All patients had been divided in grouped by age as 0.05) was found.All the ITP patients had not been given anti -B19 treatment.The PLT remission rate,respectively, after treated with the same pro-tocol including glucocorticoid and/or immunoglobulin had a declining trend as the ages increasing .The B19 infection groups of all ages al-so had no significant difference PLT remission rate had been confirmed in non -B19 infection patients in each age groups (all P>0.05). Conclusions B19 infection may not be a major cause in childhood newly ITP , and treatment protocol with no anti -B19 treatment had no influence on the clinical curative efficacy .

Neuroprotective effects of YXETNZ injection on SH-SY5 Y cells against injury induced by oxygen-glucose deprivation / 中国药理学通报

Aim To investigate the protective effects of YXETNZ injection on SH-SY5 Y cells damaged by oxygen-glucose deprivation ( OGD ) , and explore its functional mechanisms. Methods After 4 h of OGD, the cells were treated with 25 mg·L-1 drugs for 1 h. Subsequently, cell viabilities were measured by cell counting kit-8 ( CCK-8 kit ) and cell apoptosis was measured by caspase-3/7 assay kit according to manu-facturer’ s instructions. Furthermore, cell death was also detected by ELISA. The levels of phospho-Akt, phospho-PKA,phospho-Bad were evaluated by western blot. Results Oxygen-glucose deprivation significant-ly decreased the cell viabilities of SH-SY5Y cells, while YXETNZ injection significantly increased cell vi-abilities, phospho-Akt, phospho-PKA and phospho-Bad. Furthermore, YXETNZ injection also reduced the activities of caspase-3/7 and cytoplasmic histone-asso-ciated-DNA-fragments contents. Conclusion Our re-searches demonstrat that YXETNZ injection shows good neuroprotective effects on SH-SY5 Y cells after oxygen-glucose deprivation. The underlying mechanisms may be associated with activation of PI3 K/Akt/Bad/caspase-3/7 , cAMP/PKA/Bad/caspase-3/7 signaling pathway.

Molecular analysis on chronic hepatitis B patients with low-level HBsAg / 中华检验医学杂志

Objective To investigate the molecular characteristics and epidemiological signification of patients with low-level HBsAg. Methods PCR and gene sequencing were used to detect HBV DNA and Tyr-Met-Asp-Asp(YMDD) mutant in 136 serum samples with low-level HBsAg and 44 sernm samples with high-level HBsAg. Genotyping was performed in 47 cases with HBV DNA 10~5 copies/L by concentration method and 37 cases with high-level HBsAg. S gene sequences and serotypes were analyzed in 14 cases with HBV DNA 105 copies/L and 29 cases with high-level HBsAg. S gene sequences were compared with the consensus sequence of Chinese strain by BioEdit software. Results The HBV DNA-positive rate, YMDD mutation rate and HBV DNA load (logarithm) in low-level and high-level HBsAg group were 34.6% (47/136), 0% (0/136), 6.5±1.4 and 84.1% (37/44), 9.1% (4/44), 8.9±1.8, respectively. There was statistically significant differences between two groups (for concentration method,χ~2 = 30.8, P < 0.05; for direct method, χ~2 = 53.5, P < 0.05; for YMDD mutation ratio, P = 0.003, For HBV DNA (log), t = 6.5, P < 0.05). The genotypes in low-level HBsAg group included type B (16/47), type C (5/47) and non-classified ones(26/47). There were significant differences between two groups (χ~2=21.8, P <0.01). The serotypss included adw (7/14), ayw (4/14), adr (2/14) and ayr (1/14). There were significant differences in genotypes (χ~2 = 13.5, P < 0.05) but not in serotypes between two groups (χ~2 = 4.7, P >0.05). S gene sequencing results showed no S gnne variation was detected, but there were 6 single nucleotide polymorphisms in 16 cases, which would not result in the alternation of amino acid. Conclusions Low-replication phenomenon of HBV DNA was present in patients with low-level HBsAg. The major genotyps and serotype was type B and adw/ayw, respectively. Polymorphic variants have been found in the S gene. The existence of low-level HBsAg might be related with its own molecular characteristics resulting in low expression of HBsAg or immune tolerance induced by low-level HBsAg after HBV infection.

Apoptosis and Expression of Caspase in Peripheral-blood Lymphocyte(PBLC) of Systemic Lupus Erythematosus(SLE) Patients / 医学研究杂志

Objective To study the apoptosis of PBLC in SLE patients and the roles of caspase in the pathogeneses of SLE.Methods PBLC apoptosis was evaluated by vitro cell culture in 25 patients with SLE(11 active stage,14 catabasis),and 18 controls.The expression of caspase of PBLC was also detected by Western blot.Results In SLE,compared with controls,the apoptosis of PBLC were reduced before cell culture while those were increased after culture 24 and 48hrs.Expressions of caspase-2,3 in active stage patients were lower than those of catabasis group and controls.Conclusions Abnormal PBLC apoptosis results in decreasing Caspase2,3 in SLE patients.

Attenuation of GVHD for allo-bone marrow transplantation recipient by FasL-Fas pathway in an H-2 haplotype disparate mouse combination / 华中科技大学学报（医学）（英德文版）

In order to explore a new special and effective way to prevent graft versus host disease (GVHD) after allogenic bone marrow transplantation (allo-BMT), the stem cell antigen-1 (Sca-1) + early hematopoietic cells (EHC) from BALB/c mouse (H-2d) were introduced with exogenous mouse Fas ligand (mFasL) cDNA gene by the retrovirus-mediated gene transfer and expanded for one week, and then they were co-cultured with the spleen mononuclear cells (SMNC) from BAC mouse (H-2dxb) as one way mixed lymphocyte reaction (OWMLR). The cytotoxicity of treated BAC mouse SMNC against Na2 51CrO4 labeling SMNC from BALB/c mouse was observed. The bone marrow mononuclear cells (BMMNC) from BAC mouse treated by the above methods were transplanted into lethally-irradiated congenic BALB/c mice to observe the occurrence of GVHD. The results showed that the SMNC from BAC mouse after OWMLR with exogenous mFasL cDNA gene-transduced hematopoietic cells (HC) from BALB/c mouse in a ratio of 1 to 5 exhibited an obvious inhibition of the cytotoxicity against the BALB/c mouse spleen cells at different effector/target ratios as compared to the control group (P<0.01). The grade I GVHD or no GVHD and the 80% survival rate at day 60 post-BMT were observed in the BALB/c mouse receiving BAC mouse BMMNC treated with similar way, while the grade II - III GVHD and the 20% survival rate were noted in the control group (P<0.01). It is suggested that the attenuation of GVHD in allo-BMT recipient could be successfully achieved through FasL-Fas pathway in an H-2 haplotype disparate mouse combination.

Functional assay and high throughput screening for G-protein coupled receptors / 中国药理学通报

G-protein coupled receptor (GPCR) family is the large st group of therapeutic targets. High throughput screening (HTS) plays critical ro les in early stage of drug discovery. Based on signaling pathway stimulated by l igand binding on the GPCRs, many functional assay technologies have been develop ed for HTS. These technologies include Fluorometric microvolume assay technology (FMAT), Fluorescence polarization (FP), Competitive enzyme-linked immunosorben t assay (ELISA), Scintillation proximity assay (SPA), Melanophore assay, Reporte r gene assay and Calcium assay. The principles and applic- ations of these technologies were summarized in this review. We particularly emphasize those techniques of nonradioactive, su bstrate and cofactor free assays, which will be the major approaches for HTS, su ch as reporter gene and calcium assays.