italic>α-Conotoxin ArIB[V11L,V16D] is currently the most optimal selective inhibitor of α7 nicotinic acetylcholine receptor (nAChR) known. In order to explore chemical modification methods and enrich its application in targeting nAChR, this study utilized the linker to covalently connect camptothecin and 7-amino-4-methylcoumarin to the [2,4] disulfide bond of ArIB[V11L,V16D]. Therefore, two peptide-drug conjugates (PDCs), ArIB[V11L,V16D]-5 and ArIB[V11L,V16D]-6, and one fluorescent-labeled peptide, ArIB[V11L,V16D]-7 were constructed. Cytotoxicity evaluation showed that the IC₅₀ values against non-small cell lung cancer cell line A549 of the two PDCs were respectively 1.3 and 4.1 times of camptothecin, indicating slight reduction in activity at the cellular level which was related to the linker structure. Fluorescence spectrum scanning revealed that the excitation and emission wavelength of the fluorescent-labeled peptide were 340 nm and 403 nm respectively, and the fluorescence features of 7-amino-4-methylcoumarin as a marker were retained without fluorescence quenching. This modification strategy laid a solid foundation for the further application of α-conotoxin ArIB[V11L,V16D] in PDCs and fluorescent probes.

OBJECTIVE To evaluate the protective effect of Agaricus bisporus intracellular polysaccharides(IPS) and exopolysaccharides (EPS) on immunological liver injury induced by concanavalin A (Con A). METHODS Mice were pretreated with IPS and EPS (100, 200 and 400 mg kg- 1, ig) daily for 12 d. Immunological liver injury was induced by Con A 25 mg·kg-1 byinjection via the tail vein of mice.Eight hours after injection of Con A, the indexes of the liver, spleen and thymus, serum level of glutamicpyruvic transaminase (GPT), glutamic-oxalacetic transaminase (GOT), tumor necrosis factor-α (TNF-α) and interferon- γ (IFN- γ), splenic lymphocyte percentages of CD4 + and CD8 + , and liver homogenate content of superoxide dismutase (SOD) and malondialdehyde (MDA) were measured. Liver pathological changes were observed by HE staining. RESULTS Compared with normal group, the autoimmune liver injury in mice induced by Con A resulted in an increase in the liver index (P<0.01) , spleen index (P<0.01), the activity of GPT (P<0.01) and GOT (P<0.01), the content of TNF- α (P<0.01) and IFN- γ (P< 0.01), and the level of MDA (P<0.01), but a decrease in the thymus index (P<0.01), the percentage of CD4+ (P<0.01) , the ratio of CD4+/CD8+ (P<0.01), and SOD activity (P<0.01). Compared with model group, treatment with IPS (200 and 400 mg·kg-1) and EPS (200 and 400 mg·kg-1 ) respectively resulted in an increase in the thymus index (P<0.01) but in a decrease in the liver index and spleen index (P<0.01). Similarly, the activity of GOT and GPT was decreased obviously (P<0.01), and the content of TNF-α and IFN-γ in IPS and EPS 200 and 400 mg·kg-1 groups was decreased. Compared with model group, the activity of SOD in IPS and EPS (200 and 400 mg·kg- 1) group was increased (P<0.01) while MDA was decreased (P<0.01). Moreover, the percentage of CD4 + Iymphocytes decreased (P<0.01), whereas no significant difference was found in the ratio of CD4 +/CD8 + .Pathological changes of the liver were observed under a microscope. Pretreatment with IPS and EPS could effectively reduce the liver injury induced by Con A. CONCLUSION IPS and EPS have certain protective effect on immunological liver injury, which may be related to their ability to clean up free radicals, control lipid peroxidation and regulate the balance of the immune system.

Background To study diabetic retinopathy (DR) related risk factors is very important in the prevention of DR.Platelet endothelial cell adhesion molecule-1 (PECAM-1) is an important mediator that mediates high blood glucose-induced vascular diseases in diabetic patients.However,its mechanism is still below understood.Objective This clinical study was to investigate the effect of serum level changes of PECAM-1 on DR in type 2 diabetic patients.Methods Fifty-four patients with type 2 diabetes were enrolled from the endocrinology department of the Third People' s Hospital of Nantong City.Fundus examination was performed using the ophthalmoscope and fundus fluorescence angiography (FFA) on all the patients,and these patients were grouped as the non-DR (NDR)group (18 cases),non-proliferative DR(NPDR) group (20 cases) and proliferative DR group (PDR) (16 cases) based on the DR staging criterion of the Chinese Medical Association (1987 version).Eighteen age-and gender-matched normal subjects served as the normal control group.Peripheral blood was collected,and serum PECAM-1 levels were assayed using ELISA.Serum HbA1c levels were detected using the high performance liquid colorimetric(HPLC) method.The correlation of serum PECAM-1 level with serum HbA1c level was analyzed.All results were compared among the groups.Results The serum PECAM-1 levels were (10.907 ± 2.792),(7.024 ±2.377),(5.231 ± 1.816) and (3.817 ± 1.045) μg/L,respectively,in the PDR group,NPDR group,NDR group and normal control group,showing a significant difference among the 4 groups (F =12.630,P =0.02).Serum PECAM-1 content was significantly higher in the PDR group when compared with the NPDR group,NDR group and normal control group (P＜0.05).The serum HbA1c levels were (12.596±3.148)％,(9.118±3.356)％,(5.491±1.017)％ and (4.992 ± 0.725)％ in the PDR group,NPDR group,NDR group and normal control group,respectively,with a significant difference among these 4 groups (F =7.130,P =0.015),and those in the PDR group and NPDR group were significantly elevated in comparison with the NDR group and normal control group (P＜0.05).Significantly positive correlations were seen between serum PECAM-1 level and HbA1 c level in the PDR group,NPDR group and NDR group (r=0.799,P＜0.01 ;r =0.647,P＜0.01 ;r =0.685,P＜0.01).Significantly more patients with a disease course of ≥ 10 years were in the NPDR group in comparison with the PDR group (P =0.023).Conclusions Increase of serum PECAM-1 level is closely associated with blood glucose level,and it is an important factor in the pathogenesis and development of DR.These results imply that control of blood glucose is crucial for the prevention of DR in patient with type 2 diabetes.

Objective To investigate the expression of peroxiredoxin 1 (Prx 1) in hepatocellular carcinoma (HCC) with portal vein tumor thrombosis (PVTT) and to evaluate the relationship between the expressions of Prx 1 and the postoperative recurrence of this disease. Methods Immunohisto chemistry and Western blotting were performed to examine the expression of Prx 1 protein in 40 patients with HCC with PVTT. Experiments on Sprague Dawley (SD) rat hepatoma model were further carried out to observe the pathological changes of Prx 1 by immunohistochemistry. Clinical outcomes were analyzed to find a correlation between the recurrence and positive rate of Prx 1. Results The expression level of Prx 1 was significantly up-regulated in primary tumor tissues than in tumor thrombosis samples (P＜0.01). Immunohistochemistry results showed that the positive rate of Prx 1 in primary tumor tissues were higher than that in tumor thrombosis. Western blotting confirmed a same trend in the level of Prx 1, the average luminosity of the blots were 1534.2 and 735.6, respectively. There was a significant difference in SD rat hepatoma model, the 4, 8, 12, 16, 20 and 24-week positive rates of Prx 1 in liver tumor tissues were 60%, 80%, 75% ,65%, 40% and 25% respectively. Clinical outcomes showed that the time to first postoperative recurrence of Prx 1 in the primary tumor positive group was significantly higher than that in the negative group (6. 3 vs 3. 7 months, P＜0. 01). Conclusions Prx 1 protein was down-regulated in HCC with PVTT. There was a negative correlation between the expression of Prx 1 and recurrence.

The Bacillus subtilis Sac B gene with the vacuolar targeting signal sequence driven by 35S promotor was transferred into Lespedeza thunbergii by Agrobacterium mediated method. Total 62 Kan-resistant plants were obtained, of which 5 plants were proved to be transgenic plants. The transgenic plants were characterized by PCR amplification, PCR-Southern hybridization and RT-PCR. The physiological assay results showed that the transgenic plants were more tolerant to stress than the controls under the condition of 200mmol/L NaCl and 5% PEG, respectively, and that the content of soluble sugar in trnsgenic plants was significantly higher than that of controls in the period of tests (5-15 days) under salt and PEG stress.
Bacillus subtilis ; enzymology ; genetics ; Carbohydrate Metabolism ; Carbohydrates ; chemistry ; Hexosyltransferases ; genetics ; Lespedeza ; drug effects ; genetics ; growth & development ; metabolism ; Plants, Genetically Modified ; Reverse Transcriptase Polymerase Chain Reaction ; Sodium Chloride ; pharmacology ; Solubility ; Stress, Physiological ; drug effects ; Transformation, Genetic ; Transgenes ; genetics

OBJECTIVETo investigate the route and scope of the facial nerve in the temporal region for clinical applications.

METHODSTemporal region dissection was performed on 12 cadavers (24 sides) under light microscope.

RESULTSThere are two branches of the facial nerve in the temporal region from the superior margin of the parotid: the temporal branch and the zygomatic branch. Each of them has two to five branches, which run in the deep layer of the superficial temporal fascia. The temporal branch crosses the zygomatic arch to the temporal region, innervating the frontal muscle, the orbicularis oculi muscle, the corrugator supercilii muscle, and the muscle surrounding the ear, etc. The zygomatic branch goes to the lateral canthus, innervating the orbicularis oculi muscle, the upper and lower eyelid and zygomatic muscles. There are communicating branches among the temporal branches, the zygomatic branches and the supraorbital and lacrimal nerves of the ophthalmic nerve.

CONCLUSIONThe temporal branches and zygomatic branches of the facial nerve run between the deep zone of the superficial temporal fascia and the superficial layer of the profound temporal fascia, where dissection should be avoided during rhytidectomy in order not to damage the facial nerve branches.

Cadaver ; Dissection ; Eyelids ; innervation ; Facial Muscles ; innervation ; Facial Nerve ; anatomy & histology ; Fascia ; innervation ; Humans ; Parotid Gland ; anatomy & histology ; Rhytidoplasty ; Subcutaneous Tissue ; innervation ; Temporal Bone ; Zygoma

OBJECTIVETo explore the feasibility of in vitro chondrogenesis by co-culture of chondrocytes and adipose-derived stromal cells (ADSCs) so as to confirm the hypothesis that chondrocytes can provide chondrogenic microenvironment to induce chondrogenic differentiation of ADSCs.

METHODSHuman ADSCs and porcine auricular chondrocytes were in vitro expanded respectively and then were mixed at the ratio of 7:3 (ADSCs: chondrocytes). 200 microl mixed cells (5.0 x 10(7)/ml) were seeded onto a polyglycolic acid/polylactic acid (PGA/PLA) scaffold, 8 mm in diameter and 2 mm in thickness, as co-culture group. Chondrocytes and ADSCs with the same cell number were seeded respectively onto the scaffold as positive control group and negative control group. 200 microl chondrocytes (1.5 x 10(7)/ml) were seeded as low concentration chondrocyte group. There were 6 specimens in each group. All specimens were harvested after in vitro culture for 8 weeks in DMEM plus 10% FBS. Gross observation, histology, immunohistochemistry, wet weight measurement and glycosaminoglycan (GAG) quantification were used to evaluate the results. Multiple-sample t-test statistics analysis was done to compare the difference of wet weight and glycosaminoglycan(GAG) content between the groups.

RESULTSCells in all groups had fine adhesion to the scaffold and could secrete extracellular matrix. In co-culture group and positive control group, cell-scaffold constructs could maintain the original size and shape during in vitro culture. At 8 weeks, cartilage-like tissue formed in gross appearance and histological features, and abundant type II collagen could be detected by immunohistochemistry. Wet weight and glycosaminoglycan(GAG) content of co-culture group were respectively (174 +/- 12) mg and (7.6 +/- 0.4) mg. There were respectively 75% (P < 0.01) and 79% (P<0.01) of those of positive control group. In negative control group, however, constructs shrunk gradually without mature cartilage lacuna in histology. In low concentration chondrocyte group, constructs also shrunk obviously with small amount of cartilage formation at the edge area of the construct, and wet weight was (85 +/- 5) mg, which was 37% (P<0.01) of that of positive control group.

CONCLUSIONSChondrocytes can provide chondrogenic microenvironment to induce chondrogenic differentiation of ADSCs and thus promote the in vitro chondrogenesis of ADSCs.

Adipocytes ; cytology ; Animals ; Cell Differentiation ; Cells, Cultured ; Chondrocytes ; cytology ; Coculture Techniques ; Humans ; Swine ; Tissue Engineering ; methods ; Tissue Scaffolds

The paper is aimed to establish a methods for identication of pearl powder and conch powder from different origins. Hermetic aluminum pan was used to encapsulate samples. The optimal testing conditions were: heating rate 10 degrees C x min(-1), sample weight 3 mg and nitrogen gas flow rate 40 mL x min(-1). The enthalpy values of pearl powder and conch powder was obvious different. Identication of pearl powder and conch powder by DSC is a practical method for its accuracy, convenience and practificality.
Animal Shells ; chemistry ; Animals ; Calorimetry, Differential Scanning ; methods ; China ; Discriminant Analysis ; Pinctada ; chemistry ; classification ; Powders ; chemistry

OBJECTIVESTo investigate mechanism for the increasing level of serum vascular endothelial growth factor(VEGF) in tumour patients during radiotherapy and the inhibitory action of the antisense oligodeoxynucleotide (AS-ODN) to the expression of VEGF protein by radiotherapy in the prostate cancer cell line (PC3M).

METHODSTo observe the changes of serum VEGF in the prostate cancer patients during radiotherapy dynamically and the inhibitory action of the antisense oligodeoxynucleotide to the expression of VEGF by radiotherapy in PC3M.

RESULTSThe changes of serum VEGF in three patients receiving radiotherapy had been observed continuously. The levels of serum VEGF began to increase when the patients received radiotherapy and rised up to peak value after fifteen days, then declined to the range of pre-radiotherapy. Irradiating the PC3M cells with X-rays significantly increased the VEGF expression and secretion. The expression of VEGF protein in the group treated by VEGF AS-ODNs and X-ray irradiation decreased significantly than the group treated only by X-ray irradiation.

CONCLUSIONSThe induction of VEGF protein expression by X-ray irradiation in tumor cells may result in the increasing of the VEGF in the prostate cancer patients during radiotherapy and the induction can be blocked by VEGF AS-ODNs.

DNA, Antisense ; pharmacology ; Endothelial Growth Factors ; antagonists & inhibitors ; blood ; genetics ; Gene Expression ; drug effects ; radiation effects ; Humans ; Intercellular Signaling Peptides and Proteins ; blood ; genetics ; Lymphokines ; antagonists & inhibitors ; blood ; genetics ; Male ; Prostatic Neoplasms ; blood ; pathology ; Radiotherapy ; adverse effects ; Tumor Cells, Cultured ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors

OBJECTIVETo identify a naturally presented HLA-A2-restricted epitope of MAGE-A3 antigen, FLWGPRALV (MAGE-A3(271 - 279)), on the surface of a human hepatocellular carcinoma (HCC) cell line HLE.

METHODSSynthetic peptide FLWGPRALV, served as positive control target, was analyzed by HPLC and HPLC-ESI-TOF-MSMS, in order to determine its HPLC elution time, mass-spectrometric characteristics and the lowest detection limitation by the two approaches. 3 x 10(9) HLE cells were collected, peptides naturally presented by major histocompatibility complex (MHC) molecules on the cell surface were isolated by mild acid elution, and concentrated by lyophilization, then the mixtures of peptides were fractioned by HPLC. The ingredient ranged from 2 min before the elution time determined by the synthetic peptide to 2 min after that was collected, concentrated by lyophilization, and analyzed by HPLC-ESI-TOF-MSMS, to identify the existence of the MAGE-A3(271 - 279) peptide.

RESULTSThe HPLC-ESI-TOF-MSMS detection provided an evidence for the existence of a doubly charged ion of (m/z)(2) 529.9, which was further analyzed by collision induced dissociation. The doubly charged ion was ultimately identified as the MAGE-A3(271 - 279) peptide, its amino sequence was FLWGPRALV and its molecular weight was 1058.4 Da.

CONCLUSIONSMAGE-A3(271 - 279) epitope could be naturally presented by HLA-A2 molecules to the surface of HCC cell line and MAGE-A3(271 - 279) peptide may have potential immunotherapeutic value in HCC patients.

Amino Acid Sequence ; Antigen Presentation ; Antigens, Neoplasm ; analysis ; isolation & purification ; Carcinoma, Hepatocellular ; immunology ; pathology ; Cell Line, Tumor ; Chromatography, High Pressure Liquid ; Epitopes, T-Lymphocyte ; analysis ; isolation & purification ; HLA-A2 Antigen ; immunology ; Humans ; Liver Neoplasms ; immunology ; pathology ; Mass Spectrometry ; Neoplasm Proteins ; analysis ; isolation & purification