ABSTRACT:Objective To prepare polymeric nanomicelles capable of simultaneously loading doxorubicin (DOX) and Bcl‐2 small interfering RNA (Bcl‐2 siRNA ) , and to explore their in vitro cytotoxicity and cellular uptake in MCF‐7 human breast cancer cells .Methods Copolymer poly (ethylene glycol )‐g‐polyethylenimine‐g‐poly(γ‐benzyl‐L‐glutamate) was synthesized by the combination of reductive amination and carbodiimide methods , and its chemical structure was verified by 1 H NMR .Empty and drug‐loaded copolymeric nanomicelles were prepared by dialysis method and characterized by transmission electron microscope and dynamic light scattering .The ability of the nanomicelles to compress Bcl‐2 siRNA was measured by by agarose gel electrophoresis method . The release profiles of DOX and Bcl‐2 siRNA from the nanomicelles were explored by means of fluorescence spectrometry and dialysis method .The in vitro cytotoxicity and cellular uptake of DOX and Bcl‐2 siRNA co‐loaded nanomicelles in MCF‐7 human breast cancer cells were characterized by MTT assay and confocal laser scanning microscopy , respectively .Results The critical micelle concentration of the copolymer was about 4 mg/L ,and the sizes of self‐assembled empty and drug‐loaded nanomicelles were smaller than 200 nm .The drug‐loading efficiency and drug‐loading content of DOX in the nanomicelles were 88 .7% and 15 .1% ,respectively .The DOX‐loaded nanomicelles could efficiently compress Bcl‐2 siRNA when an N/P ratio was ≥64 .The zeta potential of DOX and Bcl‐2 siRNA co‐loaded nanomicelles was +30 mV .The release behavior of the cargoes from the nanomicells was pH‐sensitive , and the release of Bcl‐2 siRNA was more sensitive to acidic pH than that of DOX . The nanomicelles could simultaneously deliver DOX and Bcl‐2 siRNA into MCF‐7 cells , and the co‐delivered DOX and Bcl‐2 siRNA significantly increased the cytotoxicity of DOX (P<0 .05) .Conclusion The polymeric nanomicelles can co‐load DOX and Bcl‐2 siRNA and deliver them into MCF‐7 cells , and DOX in combination with Bcl‐2 siRNA can synergistically inhibit the growth of MCF‐7 cells and promote cell apoptosis ,suggesting that the nanomicells may be a promising carrier for the co‐delivery for chemotherapeutics and genes .

Objective To observe the improvement of Guangdong Liangcha Granules (GLG) on restraint-stress-induced peroxidation in genital organs of mice. Meth ods Mice models of peroxidation injury in genital organs were induced by 18-hou r restraint stress. Testicular and ovarian malondialdehyde (MDA) level was detec ted by thiobarbituric acid method,glutathione (GSH) content by HPLC,xanthine o xidase (XOD) and GSH-PX activities by colorimetric method,nitric oxide (NO) co ntent by Griess chemical method and oxygen radical absorbance capacity (ORAC) by enzyme-linked fluorescent immunoassay. Results Compared with model group,GLG can markedly reduce MDA level,XOD acitivity and NO contents,in addition,GLG c an effectively increase the ORAC value,GSH content,GSH-PX activity in testis and ovaries. Conclusion Oral treatment of Guangdong Liangcha Granules was found to reduce the status of peroxidation in testis and ovaries,and the improvement may be related to the increase of its free radical scavenging activity and lipid peroxidation.

Objective To investigate the effect of extract ofTangshen-HuazhuoRecipe(TSHZR) on the serum concentrations of transforming growth factorβ1(TGF-β1) and platelet derived growth factor(PDGF) in patients withⅣ stage of diabetic nephropathy(DN).Methods From June 2012 to December 2012, 98 patients ofⅣstage DN in our hospital outpatient were enrolled and randomly divided into treatment group(n=48) and control group(n=50) using random number table. All patients received conventional therapies of controlling blood sugar, lipid, blood pressure and anticoagulant therapy. On such basis, the control group was treated by irbesartan, 150 mg/d, and the treated group treated by TSHZR combined with irbesartan,150 mg/d, for 6 months. Serum TGF-β1 and PDGF were determined with ELISA before and after treatment,and urinary albumin excretion rate,HbA1c,serum creatinine,blood urea nitrogen and lipid profiles were examined as well. ResultsIn the treated group, the TGF-β1 was(172.5±31.3), (123.6±21.2)pg/ml, the PDGF was(860.9± 131.2), (500.6±130.2)pg/ml before the treatment and after the treatment, respectively. The TGF-β1 and PDGF after the treatment were significantly decreased than those before the treatment(P<0.01). After the treatment, TGF-β1 and PDGF in the treated group were statistically significant compared to the control group[TGF-β1 is(157.4±39.6)pg/ml, PDGF is(765.7±161.8)]pg/ml,P<0.01). After the treatment, the treatment group was superior to the control group in TG(1.72±0.25)mmol/L,(2.09±0.27)mmol/L,(P<0.01), TC(4.56± 0.64)mmol/L,(6.11±0.93)mmol/L, (P<0.01), HDL-C(1.56±0.50)mmol/L,(1.36±0.44)mmol/L, (P<0.01), LDL-C(2.46±1.08)mmol/L(3.32±0.87)mmol/L,(P<0.05)and UAER(100.73±204.24)μg/min, (226.24±396.38)μg/min, (P<0.01).Conclusion TSHZR can inhibit the progressive of IV stage of diabetic nephropathy by suppressing TGF-β1 and PDGF expression level.

In order to study the effects of Ca2+ in the biosynthesis of salvianolic acid B (Sal B) induced by salicylic acid (SA) in the young seedlings of Salvia miltiorrhiza, we used confocal laser scanning microscopy and high performance liquid chromatography to measure the change of relative fluorescence intensity of Ca2+ and the contents of Sal B induced by SA before and after the application of extracellular calcium channel inhibitors (VP and LaCl3), intracellular calcium channel inhibitor (LiCl), as well as intracellular calmodulin antagonist (TFP). SA could induce the calcium burst, and the Ca2+ peak could last to 2-3 min in the guard cells of S. miltiorrhiza, which prompted the biosynthesis of Sal B after the Ca2+ burst. Both Vp or LaCl3, and LiCl or TFP could inhibit the burst of Ca2+ and the biosynthesis of Sal B. The above results demonstrated that Ca2+ from the extracellular and the intracellular calcium store regulate the biosynthesis of Sal B elicited by salicylic acid in S. miltiorrhiz young seedlings.
Benzofurans ; metabolism ; Calcium ; metabolism ; Plant Leaves ; metabolism ; Salicylic Acid ; pharmacology ; Salvia miltiorrhiza ; metabolism ; Seedlings ; metabolism ; Signal Transduction

Halohydrin dehalogenase is of great significance for biodegradation of the chlorinated pollutants, and also serves as an important biocatalyst in the synthesis of chiral pharmaceutical intermediates. A putative halohydrin dehalogenase (HheTM) gene from Tistrella mobilis KA081020-065 was cloned and over-expressed in Escherichia coli BL21 (DE3). The recombinant enzyme was purified by Ni-NTA column and characterized. Gel filtration and SDS-PAGE analysis showed that the native form of HheTM was a tetramer. It exhibited the highest activity at 50 degrees C. The nature and pH of the buffer had a great effect on its activity. The enzyme maintained high stability under the alkaline conditions and below 30 degrees C. HheTM catalyzed the transformation of ethyl(S)-4-chloro-3-hydroxybutyrate in the presence of cyanide, to give ethyl (R)-4-cyano-3-hydroxybutyrate, a key intermediate for the synthesis of atorvastatin.
3-Hydroxybutyric Acid ; chemistry ; Bacterial Proteins ; genetics ; metabolism ; Cloning, Molecular ; Escherichia coli ; Hydrolases ; genetics ; metabolism ; Hydroxybutyrates ; chemistry ; Recombinant Proteins ; genetics ; metabolism ; Rhodospirillaceae ; enzymology ; genetics

Objective To investigate the effect of allgeneic hematopoietic stem cell transplantation (allo-HSCT) for β-thalassemia major. Methods Twenty-four β-thalassemia major patients with median age of 4 years (range: 2～15 years), 18 boys and 6 girls, received allo-HSCT.They were classified into class Ⅱ-Ⅲ according to Pesaro thalassemia classification. Twenty-three transplantations were from sibling donor and 1 was from mother, either HLA-identical (n = 23) or HLA-mismatched (5/6) (n = 1). Fifteen patients received bone marrow transplantation (BMT) plus peripheral blood stem cell transplantation (PBSCT), and 9 were subjected to umbilical cord blood transplantation (UCBT). The conditioning regimen consisted of busalphan, cyclophosphamide,fludarabine, plus hydroxyurea before transplantation. Graft-versus-host disease (GVHD) prophylaxis included CsA, methotrexate, antilymphpcute globulin, and mycophenolate mofetil. The median follow-up period was 13 months (range: 3～69). Results Of 24 patients, there were 21 cases (87. 5 %) of disease-free survival, 1 (4. 2 %) transplantation-related death, and 2 cases (8. 3 %) of rejection. Three-year overall survival and disease-free survival rate was 91.7 % and 87. 5 %respectively. The cumulative incidence of grade Ⅱ -Ⅳ acute GVHD and chronic GVHD was 16. 7 %and 20. 3 %, particularly cumulative extensive chronic GVHD was 5. 0 %. Conclusion The sibling donor BMT plus PBSCT is an effective and safe way to treat β-thalassemia major. Cord blood is an important source of hematopoietic stem cells for HSCT. The protocol GVHD prophylaxis of CsA,MTX, ATG with a low-dose and short course of MMF can effectively reduce the incidence of severe acute GVHD, improve the outcome of thalassemia transplantation.

OBJECTIVETo study the protective effect of Sarcandra glabra extract (SGE) on immune system in restrained mice.

METHODThe male C57BL/6 mice were randomly divided into normal control group, stress control group, 125, 500 mg x kg(-1) SGE group. The spleen lymphocyte suspensions of each group were prepared. The parameters of spleen T cells subsets, NK cell and NKT cell proportion and number was detected by Flow cytometry.

RESULTSGE regulated the balance of T cell subsets, increased the percent of NK cells and NKT cell proportion and number in restrained mice.

CONCLUSIONSGE has immunologic protective effect in restrained mice probably via the amelioration of immune cells proportion and number.

Animals ; Drugs, Chinese Herbal ; pharmacology ; Killer Cells, Natural ; drug effects ; immunology ; Magnoliopsida ; chemistry ; Male ; Mice ; Mice, Inbred C57BL ; Natural Killer T-Cells ; drug effects ; immunology ; Restraint, Physical ; Stress, Psychological ; immunology ; T-Lymphocyte Subsets ; drug effects ; immunology

Objective: To investigate the possible mechanisms of tumor-associated macrophages (TAMs) in regulating epithelia-mesenchymal transition (EMT) of Hep3B hepatoma cells, since EMT is closely associated with the malignancy of hepatoma cells and tumor microenvironment plays an important role in regulating EMT of hepatoma cells, and to provide new regimens for the clinical studies and treatment of liver cancer. Methods: Human monocytic leukemia THP-1 cells were successfully induced to TAMs. With TAMs as target cells, they were co-cultured with the supernatant of Hep3B hepatoma cells or Hep3B hepatoma cells, and Western blot and RT-PCR were used to measure the change in the expression of Toll-like receptor 4 (TLR4) in TAMs. The expression of TLR4 in TAMs was downregulated by transient plasmid transfection with shRNA. With Hep3B hepatoma cells as target cells, the supernatants of TAMs and TAMs transfected with shRNA TLR4 plasmid were used for intervention, and Western blot was used to measure the protein expression of E-cadherin, N-cadherin, and vimentin. The two-sided t-test was used for comparison of the means of two independent samples. Results: THP-1 cells were successfully induced to TAMs. According to the results of Western blot, compared with the control-CM group, the TAM-CM group had a significant reduction in the protein expression of E-cadherin and significant increases in the protein expression of N-cadherin and vimentin in Hep3B cells. After the expression of TLR4 in TAMs was downregulated, the culture solution of TAMs was used for the intervention of Hep3B cells (shRNA group), and compared with the TAM-CM group, the shRNA group had a significant increase in the expression of E-cadherin and significant reductions in the protein expression of N-cadherin and vimentin in Hep3B cells. Western blot and RT-PCR showed that the expression of TLR4 in TAMs was influenced by Hep3B cells. Conclusion TAMs can promote EMT of Hep3B hepatoma cells, and downregulation of the expression of TLR4 in TAMs may reduce EMT of Hep3B hepatoma cells, suggesting that TLR4 on the surface of TAMs may be a key molecule involved in the interaction between TAMs and Hep3B hepatoma cells.

Objective To analyze the influencing factors of hematoma complicated from ultrasound-guided minimally invasive surgery for benign breast masses.Methods Retrospective analysis was performed in 412 patients with a total of 516 masses underwent the ultrasound-guided minimally invasive surgery for benign breast masses from January 2011 to December 2015 in Xiaoshan Hospital. Theχ2 test was used to univariately analyze risk factors of hematoma formation after ultrasound-guided minimally invasive surgery for benign breast masses. Logistic regression analysis was used to multivariately analyze risk factors of hematoma formation.Results All masses were resected completely, however, hematomas with long diameter≥1.0 cm were formed in 43 masses one week after surgery, and all hematomas were completely absorbed after six months. There were significant differences in the incidence of hematoma between the groups of different needle sizes, numbers of needle cutting, masses sizes, blood flow grades, depth, resection numbers and effective compression time of postoperative bandages (χ2=16.917, 14.548, 39.971, 23.333, 29.137, 36.819 and 39.864, respectively, allP<0.001). The needle sizes, the numbers of needle cutting, the masses sizes, blood flow grades, depth, resection numbers and the effective compression time constituted the risk factors of hematoma formation after the minimally invasive surgery.Conclusions The risk factors of the hematoma formation after ultrasound-guided minimally invasive surgery for benign breast masses included the different size of the needle, the number of different cutting needles, different size of the masses, the grade of blood flow, the number of resection and the different effective compression time of postoperative bandages. We could prevent the occurrence of hematoma in advance by screening patients and take corresponding measures.