Objective: To study the chemical constituents from the stems of Acanthopanax gracilistylus. Methods: The chemical constituents of the plant were isolated and purified by column chromatography and their structures were elucidated on the basis of physico-chemical properties and spectral data. Results: A new ent-kaurane glycoside, named kaurane acid glycoside A (16α, 17-dihydroxy-ent-kauran-19-oic 19-[β-D-glucopyranosyl-(1→2)-β-D- glucopyranosyl] ester) (1), was isolated from the n-butanol part. Conclusion: Compound 1 is a new one.

【Objective】 To determine the volume range of suspended erythrocyte and establish its internal control standard. 【Methods】 The theoretical value of suspended erythrocyte volume was calculated according to the screening criteria of healthy blood donors and Quality Requirements for Whole Blood and Blood Components. A total of 2 410 bags of 1 U and 2 U suspended erythrocyte were randomly selected and weighed, and the volume range were formulated by ±2S and ±10% respectively and then compared to determine the volume range in line with the actual situation of our center. 【Results】 The theoretical volume range of 1 U and 2 U suspended erythrocyte were 117-160 mL vs 234-320 mL, and the actual volume range were 142-180 mL vs 276-393 mL. The volume range of 1 U and 2 U suspended erythrocyte formulated by ±2S were 145-181 mL vs 298-358 mL, and by ±10% were 147-179 mL vs 295-361 mL. The hematocrit and hemoglobin content of suspended erythrocyte within the actual volume range met the quality requirements. There were fluctuations in the volume of suspended erythrocyte from different regions. 【Conclusion】 Based on the actual situation of our center and the sampling results of suspended erythrocytes in recent two years, 163 mL±10% and 328 mL±10% were determined as the internal control standards of 1 U and 2 U suspended erythrocyte, respectively. Blood centers should establish accurate and feasible standard of suspended erythrocyte according to the actual situation.

Background Allograft rejection is a main cause of failure of penetrating keratoplasty,especially in the patient with high risk of rejection condition.Previous study on allograft rejection mechanism focused on limbal and corneal neovascularization,but these factors did not explain all the phenomena of allograft rejection.Research found that immune cells appeared in iris and ciliary body when rejection occurred,but the relationship between these immune cells and allograft rejection is unclear Objective This study was to evaluate the relationship between diversity of vascular permeability in the iris and ciliary body and allograft rejection after penetrating keratoplasty.Methods Seventy clean eight-week-old BALB/c mice were divided into allogeneic corneal transplantation group (60 mice) and blank control group (10 mice).Allogeneic corneal transplantation was performed with the same age of C57BL/6 mice as donor and BALB/c mice as the recipients.The grafts were examined under the slit lamp microscope and scored based on the criteria of Hegde.The mice were sacrificed and iris and ciliary tissue were obtained 5,10 days and rejection after surgery.Immunohistochemistry and reverse transcription PCR (RT-PCR) was used respectively to detect the expression diversities of occludin,zonula occludens protein-1 (ZO-1),matrix metalloproteinase-9(MMP-9),major histocompatibility complex-Ⅱ (MHC-Ⅱ),and CCR5,CCR7 and their mRNA in iris and ciliary body.Image-J image analysis software was used to calculate the quantity of positive cells on iris wholemount,and absorbance of target genes (A values).The use and care of the experimental animals complied the ARVO Resolution on the Use of Animals in Research.Results The mean survival time of corneal gratts was (17±3) days after operation.The mean score was 0.6 in 5 days and 0.5 in 10 days,and 3.3 in 18 days after operation.Expression of ZO-1 reduced significantly,and that of MMP-9 increased obviously at the time of rejection.MHC Ⅱ + cells were scattered in iris and ciliary body in normal mice,and the number of the positive cells (cells/field) was increased after operation with a peak value when rejection occurred.A significant difference was seen between normal mice and rejection mice (1559.67±350.29 vs.4021.83±495.18) (P=0.000).The expressions of occludin mRNA and ZO-1 mRNA in the iris and ciliary body decreased obviously in the rejection mice.Compared with normal mice,theA value of ZO-1 and occluding were 36.74±3.13 vs.110.11±11.88 and 57.54±3.41 vs.59.90±3.50respectively,with significant differences between them (all P＜0.05).The expressions of MMP-9 mRNA,CCR5 mRNA and CCR7 mRNA in the iris and ciliary body increased gradually with the time lapse after operation and peaked when the rejection appeared.The A value of MMP-9 mRNA,CCR5 mRNA and CCR7 mRNA were significantly higher than those of normal mice (20.29±1.19 vs.2.77±0.85 for MMP-9 mRNA; 35.43±2.56 vs.9.11±0.29 for CCR5 mRNA,and 60.83±0.87 vs.0.89 ±0.95 for CCR7 mRNA) respectively (all P＜0.05).Conclusions The permeability of vascules in the iris and ciliary body increase during the allograft rejection after penetrating keratoplasty.Increased antigen presenting cells were also detected.

Objective To study the relation of matrix metalloproteinase-9 in serum and tumor necrosis fac- tor-?in serum and amniotic fluid in predicting ehorioamnhionitis in patients with premature rupture of membranes (PROM).Methods The levels of MMP-9 in serum and TNF-?in serum and amniotie fluid were measured by ra- dioimmunoassay and ELISA in 67 cases with premature rupture of membranes as study group and 40 cases normal full-termed pregnant women as controls group.Results(1)The levels of TNF-?in amniotie fluid and MMP-9 in serum in study group were significantly higher than those in controls group(P0.05).(2)In study group,the levels of MMP-9 of serum in0.05).Conclusions The levels of TNF-?in amniotic fluid and MMP-9 in serum were valuable clinical indices for identification of chorioamnionitis in patients with PROM.The levels of MMP-9 in serum also could assess the time of rupture of membranes and the degree of ehorioamnionitis.

The marine biological source of mineral drugs recorded in Chinese Pharmacopoeia (2010 version) mainly including pearl, nacre, clam shell, common oyster shell, ark shell, cuttle bone, and sea-ear shell are widely used in clinical. Calcium carbonate and a small amount of protein are the main components in this type of drugs. In this paper, a systematical and comparable study were carried out by determination of calcium carbonate by EDTA titration method, the crystal of calcium carbonate by X-Ray powder diffraction and the total amino acids (TAAs) of the hydrolyzed samples by ultraviolet spectrophotometry method. As a result, the crystal structure is calcite for common oyster shell, mixture of calcite and aragonite for nacre and sea-ear shell, aragonite for the other drugs. The content of calcium carbonate ranged from 86% to 96%. Cuttle bone has the highest amount of TAAs among the seven drugs which reached 1.7% while clam shell has the lowest content of 0.16% on average. In conclusion, an effective method was developed for the quality control of marine mineral drugs by comprehensive analysis of calcium carbonate and TAAs in the seven marine mineral drugs.
Amino Acids ; analysis ; chemistry ; Animal Shells ; chemistry ; Animals ; Calcium Carbonate ; analysis ; chemistry ; Crystallization ; Edetic Acid ; chemistry ; Mollusca ; chemistry ; classification ; Pharmaceutical Preparations ; analysis ; chemistry ; standards ; Quality Control ; Reproducibility of Results ; Seawater ; Species Specificity ; Spectrophotometry, Ultraviolet ; X-Ray Diffraction

Objective:The recombinant human retinal pigment epithelium-derived factor(PEDF)protein to be obtained and the angiogenesis of the rPEDF to be identified.Methods: PEDF gene gene was amplified by PCR and cloned into pET32a,rPEDF protein was expressed in E.coli BL21 and confirmed by SDS-PAGE and Western blot.The rPEDF was purified by Ni-NTA on denature condition.The concentration of the rPEDF was determined by Bradford method.The angiogenesis of the rPEDF was determined by chick chorioallantoic membrane(CAM) method.Results: The expression plasmid pET32a-PEDF was constructed successfully.The rPEDF was expressed with stable efficiency in E.coli BL21.The results of the CAM experiment showed that the rPEDF had notable angiogenesis effect in the concentration 0.4、0.04 ng/ml,but had no effect in 4 ng/ml.Conclusion:The PEDF gene was cloned and expressed efficiently,the angiogenesis of the rPEDF to be identified and the activity was worked in certain range.The results can facilitate studying its function and spreading its application.

Objective To investigate the correlation of the expressions of advanced glycation end products(AGE) and the receptor for advanced glycation end products(RAGE) in serum and placenta with the pathogenesis of preeclampsia. Methods From December 2013 to June 2014, 32 women with severe preeclampsia who received cesarean section in the Affiliated Hospital of Qingdao University were recruited in the study, defined as the severe preeclampsia group. 30 healthy pregnant women who received cesarean section in the same hospital were recruited as the control group. ELISA was used to measure the maternal serum AGE, soluble receptor for advanced glycation end products (sRAGE) and tumor necrosis factor-α(TNF-α) in these women. Furthermore, ELISA was also used to measure AGE and TNF-α in the placenta. The localizations of AGE and RAGE protein in placentas were detected by immunohistochemical SP method. RAGE and TNF-α mRNA expression in placentas were measured by real-time quantitative PCR. AGE, RAGE and TNF-αprotein expression in placentas were measured by western blot, respectively. Results (1) The serum levels of AGE,sRAGE and TNF-αin the severe preeclampsia group were (538 ± 75),(367 ± 86) and (322 ± 40) ng/L,respectively. They were significantly higher than those in the control group[(454 ± 50), (286 ± 35) and (270 ± 35) ng/L, respectively](P<0.05). The levels of AGE showed positive correlation with the levels of TNF-α(r=0.588,P<0.05),while the levels of sRAGE showed no correlation with TNF-α(r=-0.041, P>0.05). (2) In the severe preeclampsia group, the levels of AGE and TNF-αin placentas were (500 ± 82) and (334 ± 57) ng/L, which were higher than those in the control group [(431 ± 74) and (263 ± 46) ng/L, respectively](P<0.05). The levels of AGE showed positive correlation with the levels of TNF-ɑ(r=0.406,P<0.05). (3)AGE and RAGE protein mainly located in the syncytiotrophoblasts, macrophages and vascular endothelial cells in the placentas of the two groups. AGE expressed mainly in the cytoplasm, and RAGE expressed in the cytoplasm and cell membranes.(4)RAGE and TNF-αmRNA expression in the placentas of the severe preeclampsia group were 12.6 ± 4.6 and 10.4 ± 2.4, which were significantly higher than those in the control group (0.9 ± 0.4 and 3.5 ± 0.9,P<0.01). (5) The expressions of AGE、RAGE and TNF-αprotein in placentas of the severe preeclampsia group were 0.68 ± 0.06, 0.82 ± 0.08 and 0.76 ± 0.08. All were significantly higher than those of the control group (0.46 ± 0.05,0.42 ± 0.09 and 0.52 ± 0.07;P<0.01). Conclusions The levels of AGE and RAGE in serum and placentas elevated in the severe preeclampsia group, and the expression of TNF-αalso elevated. These indicated that AGE and RAGE might be involved in the systemic inflammatory response and local inflammatory response in placentas, and then caused the preeclampsia.

To improve ethanol production in Saccharomyces cerevisiae,an integration plasmid pUPGKAT with PGK promoter(phosphoglycerate kinase promoter),adh1 gene(the coding sequences of alcohol dehydrogenaseⅠ) and CYC1 terminator(Cytochrome c transcription terminator) was constructed.Firstly,a fusion fragment composed of PGK promoter and adh1 gene was generated by over lap extension PCR and ligated into pUG6 resulting in plasmid pUPGKA.Subsequently,CYC1 termi nator was amplified from pSH65 by PCR and ligated to the SpeⅠand SacⅡrestriction site of pUPGKA.To integrate PGK-adh1-CYC1 into S.cerevisiae genome,pUPGKAT was digested by TthⅢⅠand the lin-earized plasmid was used to transform S.cerevisiae YS2-△adh2(adh2 disrupted strain) by lithium acetate method.The yeast mutant YS2-△adh2-adh1 which had the adh1 gene placed under the PGK promoter and harbored the adh2 deletion was constructed.Anaerobic fermentation showed overexpression of adh1 by PGK promoter resulted in a 8.84% higher ethanol production compared to YS2-△adh2.

OBJECTIVETo discuss the effect of Taohong Siwu Decoction (TSD) in regulating functions of endothelial cells and treating arteriosclerosis obliterans (ASO).

METHODSThe ASO model was prepared by using high-fat diet plus intimal injury. They were randomly divided into the model group (n = 10), the normal control group (n = 9), the low dose TSD group (group A, n = 12), the middle dose TSD group (group B, n = 10), and the high dose TSD group (group C, n = 9). Eight weeks after modeling, the limb blood perfusion was observed using laser Doppler flowmetry. The arterial morphology was observed using light microscope and transmission electron microscope. The number of circulating endothelial cells (CECs) was determined using Percoll density gradient centrifugation method. Serum levels of TNF-alpha, IL-1, ET-1, and NO were detected using double antibody sandwich assay of enzyme linked immunosorbent assay (ELISA).

RESULTSThe ASO rat model was successfully established. Blood lipids levels significantly increased, the blood perfusion of left hind limbs significantly decreased, the number of CECs in the peripheral blood significantly increased, the arterial lumen was irregularly narrowed, the ultra-structure of vessel walls was damaged, serum levels of TNF-alpha, IL-1, and ET-1 significantly increased, and the serum level of NO significantly decreased in the model group, showing statistical difference when compared with the normal control group (P < 0.01). Compared with the model group, significant improvement in the aforesaid indices was shown in group B and C (P < 0.05, P < 0.01).

CONCLUSIONSThe injury and abnormal functions of endothelial cells is an important pathological process of ASO. As an effective recipe for treating ASO, TSD could protect vascular endothelial cells and improve the secretion function of vascular endothelial cells.

Animals ; Arteriosclerosis Obliterans ; blood ; drug therapy ; Diet, High-Fat ; adverse effects ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Endothelial Cells ; metabolism ; Endothelin-1 ; blood ; Endothelium, Vascular ; cytology ; Interleukin-1 ; blood ; Male ; Nitric Oxide ; blood ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha ; blood

0.05 indicating no statistical difference.However,the noise measurements for the L and C groups were 30.05 and 27.80,respectively,with P