Objective To investigate the expression of integrin avβ3 in both of hepatic stellate cell (HSC)and fibrotic liver tissue,and to demonstrate whether integrin avβ3 is a phenotypical receptor of activated HSC.Methods HSC were isolated from Sprague-Dawley rats and activated by prolonged cultural.The rats were injected with 175 mg of thioacetamide twice a week for 12 weeks to induce liver fibosis.The expressions of avβ3 and a-SMA were identified by immunocytochemical staining.The expression of avβ3 in HSC,normal and fibrotic tissues were examined by real-time PCR and Western blot.Results The expression of avβ3 in activated HSC were up regulated and levels of mRNA and protein were increased to 18 and 5.2 folds on day 14 compared with day 1 and were also higher than control(t=2.39,P＜0.05;t=2.74,P＜0.05).The immunochemistry staining showed that integrin avβ3 was expressed on membrane of activated HSCs.The avβ3 and a-SMA were expressed in portal vein in normal liver,but were also expressed in portal and fibrotic.Conclusions The integrin avβ3 is up-regulated in activated HSC both in vitro and in vivo.It is a phenotypical receptor of activated HSC which involved in liver fibrosis.

OBJECTIVE:To investigate the effects of NOS1AP rs12742393 A/C gene polymorphism on lipid-regulating re-sponse of rosuvastatin calcium. METHODS:Two hundred and tuirty six patients with coronary heart disease(CHD)were selected from cardiology department of our hospital during Jan. 2014-Jun. 2015,and then given rosuvastatin calcium and other symptomatic treatment for 12 weeks. Polymorphism of NOS1AP rs12742393 A/C was detected by PCR-RFLP. The levels of TG,TC,HDL-C and LDL-C were detected by photoelectric colorimetry before treatment and 4,12 weeks after treatment. The serum relationship of genotype with the level of blood lipid was analyzed. RESULTS:Among 236 CHD patients,there were 131 cases of AA genotype (55.5%),98 cases of AC genotype(41.5%) and 7 cases of CC genotype(3.0%);genotype and allele frequencies met the Har-dy-Weinberg balance(P>0.05). There were 132 patients with normal blood lipid and 104 patients with hypercholesterolemia;there was statistical significance in genotype and allele frequencies (P<0.05). Among 104 CHD patients with hypercholesterolemia be-fore treatment,there was no statistical significance in the levels of TG,TC,LDL-C and HDL-C between AA genotype and AC+CC genotype(P>0.05). 4th and 12th week after treatment,the levels of TG,TC and LDL-C in different genotypes were all de-creased significantly;4th week after treatment,the level of LDL-C in AC+CC genotype was significantly lower than AA genotype, and the change compared to before treatment was significantly more than AA genotype,with statistical significance (P<0.05). There was no statistical significance in the level of HDL-C among different genotypes compared to before treatment;there was no statistical significance in the levels of TG,TC and HDL-C 4th,12th week after treatment and their changes compared to before treatment between AA genotype and AC+CC genotype;there was no statistical significance in the level of LDL-C 12th week after treatment and their changes between AA genotype and AC+CC genotype(P>0.05). CONCLUSIONS:NOS1AP rs12742393 A/C gene polymorphism is associated with CHD complicated with hypercholesterolemia;the C allele of NOS1AP rs12742393 may strengthen the response of CHD patients with hy-percholesterolemia to rosuvastatin calcium through influencing the level of LDL-C.

Objective To investigate the correlation between extended high-frequency audiometry and distortion product oto acousticemissions (DPOAE) amplitudes. Methods 60 normal hearing people aged 20~40 years were examined with extended high-frequency audiometry and DPOAE. Results Extended high frequency thresholds negatively correlated to DPOAE. Conclusion There is correlation between these two methods, and they can be used for early creening and diagnosis of hearing loss.

AIM: To construct prokaryotic expression vector of human angiogenesis inhibitor arresten gene and express recombinant arresten in Escherichia coli. METHODS: Human arresten gene was amplified from recombinant plasmid pGEM-Arr with polymerase chain reaction (PCR), and then cloned into prokaryotic expression vector pRSET by means of recombinant gene technology. The recombinant plasmid pRSET-Arr was transformed into E.coli BL21(DE3), and recombinant arresten was expressed in the bacteria under induction of IPTG. The expressed products were detected by SDS-PAGE analysis. RESULTS: Restriction analysis indicated that the arresten gene was successfully inserted into the expression vector, and DNA sequencing verified that the reading frame of the recombinant vector was correct. Recombinant arresten was successfully expressed in Escherichia coli; its molecular weight was about 26 kD and its amount was approximately 30% of total bacterial proteins.CONCLUSION: The successful construction of prokaryotic expression vector containing human arresten gene and the effective expression of recombinant arresten in Escherichia coli laid the foundation for further study on its biological functions.

The eukaryotic expression of human arresten gene and its effect on the proliferation of in vitro cultured vascular smooth cells (VSMCs) in vitro were investigated. COS-7 cells were transfected with recombinant eukaryotic expression plasmid pSecTag2-AT or control plasmid pSecTag2 mediated by liposome. Forty-eight h after transfection, reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of arresten mRNA in the cells, while Western blot assay was applied to detect the expression of arresten protein in concentrated supernatant. Primary VSMCs from thoracic aorta of male Sprague-Dawley rats were cultured using the tissue explant method, and identified by immunohistochemical staining with a smooth muscle-specific anti-alpha-actin monoclonal antibody before serial subcultivation. VSMCs were then co-cultured with the concentrated supernatant and their proliferation was detected using Cell Counting Kit-8 (CCK-8) in vitro. The results showed that RT-PCR revealed that the genome of arresten-transfected cells contained a 449 bp specific fragment of arresten gene, suggesting the successful transfection. Successful protein expression in supernatants was confirmed by Western blot. CCK-8 assay showed that the proliferation of VSMCs were inhibited significantly by arresten protein as compared with control cells (F=40.154, P<0.01). It was concluded that arresten protein expressed in eukaryotic cells can inhibit proliferation of VSMCs effectively in vitro, which would provide possibility to the animal experiments.

Objective:To investigate the effect of short single reverse-α fixation of nasobiliary tube after endoscopic nasobiliary drainage.Methods:From January 2019 to October 2019, the patients who performed with endoscopic nasolbiliary drainage in Tongji University Affiliated Shanghai East Hospital were randomly divided into experimental group (short single reverse-α fixation, 155 cases) and control group (routine reverse-α fixation, 137 cases). The incidences of the fixation time, prolapse rate, nursing time, scores of nasal comfort and bile flow rate of nasobiliary duct were evaluated between the two groups.Results:The average time of nasobiliary duct fixation was (18.31±1.67) s, the prolapse rate was 5.2% (8/155) and the time of nursing was (35.03±2.68) s, which were lower than those of the control group (46.50±5.50) s, 13.9% (19/137) and (72.07±7.63) s. The difference was statistically significant ( t or χ 2 values were 60.795, 6.570, 56.629, P<0.01 or 0.05). The comfort score of the experimental group was (4.61±1.06) points, the bile flow rate was (241.52±53.95) ml/days, which were higher than (5.76±0.76) points and (174.09±47.55) ml/days of the control group, the difference was statistically significant ( t values were 10.448, 11.265, P<0.01). Conclusions:Shortening the length of nasobiliary duct combined with single reaction "α" method has the advantages of simper operation and nursing, low prolapse rate, high comfort and good drainage effect. It is worthy of clinical application.

Objective To study the influence of serum containing Fuzheng Toudu Qudu Recipe, a Chinese formula with the actions of supporting healthy qi to expel and remove toxicity, on serum levels of interleukin 2 (IL-2) and soluble interleukin 2 receptor (sIL-2R) at different stages of CD34+derived dendritic cells (DC) of patients with minimal residual disease of myelogenous leukemia ( MRD-L) , and to explore the biological mechanism of Fuzheng Toudu Qudu Recipe in promoting CD34+ to transform into DC in MRD-L patients. Methods Bone marrow mononuclear cells ( BMMC) were separated from the bone marrow of acute myeloid leukemia patients at complete remission stage by using Ficoll centrifugation. CD34+ cells were isolated by using immuno-magnetic mircobeads method, and then were cultured with various concentrations of Chinese medicine medicated serum and cytokines in vitro for the induction of DC. The morphologic characteristics of DC were observed with the inverted phase contrast microscope, and the expression levels of DC surface molecules such as CD83, CD80, CD86, CD1a and HLA-DR were detected by using flow cytometry. On culturing day 0, 6 and 9, serum levels of IL-2 and sIL-2R of each group were measured by enzyme-linked immunosorbent assay ( ELISA). Results ( 1) Chinese medicine medicated serum combined with cytokines was effective on promoting CD34+ to differentiate into DC with typical morphology, and inducing DC to have high expression of CD80, CD83, CD86 and HLA-DR, which differed from those in fetal calf serum (FCS) group and blank rabbit serum group (P<0.01). Middle- and low-dose combination groups increased expression of CD1a, which differed from high-dose combination group and cytokines group ( P<0.01). ( 2) Content of IL-2 in combination groups was higher than that in blank rabbit serum group on culturing day 0. In the combination groups, IL-2 was higher on culturing day 6 and 9 than that on culturing day 0. Middle and high-dose combination groups had higher IL-2 content on culturing day 9 than on culturing day 6 ( P<0.05 or P<0.01). At the same time point, combination groups had higher IL-2 content than the blank rabbit serum group (P<0.05 or P<0.01). (3) Content of sIL-2R in the combination groups was lower than that in blank rabbit serum group on culturing day 0. In the combination groups, sIL-2R was lower on culturing day 9 than that on culturing day 0 and 6 ( P<0.01) . High-dose combination group had lower sIL-2R content on culturing day 6 than that on culturing day 0 (P<0.05), and the difference of sIL-2R in other groups was insignicant between on culturing day 6 and on culturing day 0 ( P>0.05) . At the same time point, combination groups had lower IL-2 content than the blank rabbit serum group (P<0.05 or P<0.01). Conclusion Fuzheng Toudu Qudu Recipe is effective on increasing serum content of IL-2 and reducing sIL-2R content, and the changes of cytokine contents are more obvious along with the maturity of DC, which indicates that the recipe plays positive effect in the process of promoting CD34+cells to differentiate into DC.

Norepinephrine (NE) can raise blood pressure and speed up heart rate. However, because its effect of raising heart rate is less than that of reflex reduction of heart rate caused by the increase of blood pressure, NE causes more heart rate decrease in patients. A case of tachyarrhythmia caused by low dose NE was admitted to department of intensive care unit (ICU) of Shijiazhuang Third Hospital. The heart rate of the patient increased with the elevation of NE application dose. A variety of antiarrhythmic drugs was invalid. The related examination was prescribed to eliminate the cause of arrhythmia caused by the disorder of electrolysis and thyroid function, and found that heart rate decreased as the dose of NE tapered. After NE was stopped, the patient recovered sinus rhythm. During one month of follow-up, the patient's heart rhythm was normal. Therefore, the occurrence of tachyarrhythmia is related to NE.

The eukaryotic expression of human arresten geneand its effect on the proliferation of in vitro cultured vascular smooth cells (VSMCs) in vitro were investigated. COS-7 cells were transfected with recombinant eukaryotic expression plasmid pSecTag2-AT or control plasmid pSecTag2 mediated by liposome. Forty-eight h after transfection, reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of arresten mRNA in the cells,while Western blot assay was applied to detect the expression of arresten protein in concentrated supernatant. Primary VSMCs from thoracic aorta of male Sprague-Dawley rats were cultured using the tissue explant method, and identified by immunohistochemical staining with a smooth muscle-specific anti-αactin monoclonal antibody before serial subcultivation. VSMCs were then co-cultured with the concentrated supernatant and their proliferation was detected using Cell Counting Kit-8 (CCK-8) in vitro. The results showed that RT-PCR revealed that the genome of arresten-transfected cells contained a 449 bp specific fragment of arresten gene, suggesting the successful transfection. Successful protein expression in supernatants was confirmed by Western blot. CCK-8 assay showed that the proliferation of VSMCs were inhibited significantly by arresten protein as compared with control cells (F=40.154, P＜0.01). It was concluded that arresten protein expressed in eukaryotic cells can inhibit proliferation of VSMCs effectively in vitro, which would provide possibility to the animal experiments.