This thesis aimed at the Bacillus natto TK-1 screened out from Natto.The lipopeptide was purified using Thin-Layer Chromatography(TLC),and investigated its anti-tumor activity.After acid precipitation and methanol extraction,the lipopeptide was separated on TLC.Then the authors get the monomer surfactin which molecular weight is 1036Da through the High performance liquid chromatography(HPLC),electro spray ionization-mass spectrometry(ESI-MS) and infrared(IR).MTT method was implied to testify the anti-tumor activity of the purified sample from TLC.The results indicated a concentration and time-dependent relationships.After 48h,their IC50 were 40 mg/L.The detection with inverted microscope fluorescence microscope displays that the surfactin will cause a series of Morphological changes to the cells.In TUNEL experiment,the authors noticed that surfactin has the ability to induce apoptosis,besides this inhibition shows an obvious time-dependent relationship.

The study was to develop cis-dichlorodiammineplatinum(DDP)-loaded formulations using a novel type of self-assembled compound composed of block copolymers synthesized by poly(?-glutamic acid)(?-PGA).For the potential of targeting liver cancer cells,D-galactose was conjugated on the prepared ?-PGA.In vitro,DDP can be released from the resulting conjugate in PBS:there was a burst release during the first 8 h,then followed by sustained release.DDP could be easily incorporated into poly(?-glutamic acid)-D-galactose esterifiable derivative through a covalent bond.The yield of DDP incorporation into the esterifiable derivative was 9.4%～10.2%.In vitro experiments conclusively established that the poly(?-glutamic acid)-D-galactose esterifiable derivative-Cisplatin Complex Compound(?-D+-DDP)was much less toxic to normal cell lines than DDP only.The surviving rate of cells treated with ?-D+-DDP compound is higher than those treated with free DDP.Also it has obvious antitumor efficiency on human liver tumor BEL-7402 cells.HE staining indicated that the ?-D+-DDP compound make the BEL-7402 apoptosis.These results indicated that the conjugation of DDP to the esterifiable derivative reduced its cytotoxicity activity,but retains its antitumor activity in vitro.In conclusion,the ?-D+-DDP compound could be used as a potential clinic antitumor drug.The ?-PGA obtained by fermentation can be used as a valuable drug carrier system.

Humans ; Regeneration ; Stem Cell Research ; Stem Cells

Objective To compare biomechanical properties of the helical and straight long PHILOS (proximal humerus internal locking system) plates (Synthes Inc., Switzerland), so as to provide some biomechanical evidence for treating proximal metaphyseal-diaphyseal humeral shaft fractures in clinic. Methods Twelve Synbone artificial bones of right humerus (SYNBONE Inc., Switzerland) were divided into two groups. In control group (n=6), the humerus was fixed with the 10 hole long straight PHILOS plate, while in experimental group (n=6), the humerus was fixed with the same long PHILOS plate which was precontoured for moulding (i.e. helical PHILOS plate). After the proximal metaphyseal-diaphyseal humeral shaft fractures were made in all artificial bones, the biomechanical properties of the specimens in two groups under 6 loading modes (i.e., axial tension and compression, torsion in the same and reverse direction, medial-lateral and anterior-posterior three-point bending) were tested en bloc and compared. ResultsCompare with control group, under 100-500 N tensile and compressive loads, the axial displacement at the fractured end in experimental group increased by about 95% and 58%, respectively. Under 0.6-3 N•m torsional moment in reversed direction, the tensional angle in experimental group was obviously smaller than that in control group, with a decrease of 55%-64%. Under medial-lateral bending moment of 1.5 and 3 N•m, no significant difference was found in deflection of the experiment and control group, while under medial-lateral bending moment of 4.5, 6 and 7.5 N•m, the deflection in experimental group decreased by 20%-30% as compared to control group. Under 0.6-3 N•m torsional moment in the same direction and 1.5-7.5 N•m anterior-posterior bending moment, both the torsional angle and the deflection in experimental group were larger than those in control group, with a significant difference (P＜0.05). Compared with control group, the tensile stiffness and compressive stiffness decreased by 49% and 36%, the torsional stiffness in the same direction decreased by 19% and that in reversed direction increased by 150%, three-point bending stiffness in medial lateral direction increased by 18% and that in anterior posterior direction decreased by 70% in experimental group, all with a significant difference (P＜0.05). ConclusionsCompared with the long straight PHILO plate, the long helical PHILOS plate has better biomechanical properties, which can meet the clinical need of proximal metaphyseal-diaphyseal humeral shaft fracture fixation and postoperative rehabilitation. This surgical technique is expected to be widely applied in clinic, especially with the advantage of minimal invasive surgery.

OBJECTIVETo investigate the effects of aminoguanidine cream on the proliferation of keratinocytes (KC), content of advanced glycosylation end products (AGE) and oxidative stress in skin tissue of rats with diabetes.

METHODSStearic acid, liquid paraffin, vaseline, lanolin, isopropyl myristate fat, glycerol, 50 g/L alcohol paraben, aminoguanidine hydrochloride etc. were mixed in certain proportion to make aminoguanidine cream, and cream without aminoguanidine was used as matrix. The dorsal skin of normal rats were harvested and treated by aminoguanidine cream with dose of 5, 10 g/L, or 5 g/L together with 10 g/L azone. The transdermal effect was respectively measured at post treatment hour 2, 4, 7, 10, 12, 24. Thirty SD rats were divided into normal control (NC, n = 6), diabetes (D, n = 8), aminoguanidine cream-interfered (AI, n = 8), matrix cream-interfered groups (MI, n = 8) according to the random number table. Diabetes was reproduced by intraperitoneal injection of STZ (65 mg/kg) in rats of D, AI, and MI groups, and rats in NC group were injected with 0.05 mmol/L citrate buffer as control. One week later, dorsal skin of rats in AI and MI groups were respectively treated with 10 g/L aminoguanidine cream and matrix cream by external use for 4 weeks. AGE content was determined with fluorescence detection from skin collagen extract. KC cell cycle was detected by flow cytometry. Skin tissue specimens were obtained for determination of levels of superoxide dismutase (SOD), malondialdehyde (MDA), myeloperoxidase (MPO), and total antioxidant capacity. Data were processed with t test.

RESULTSTransdermal effect of aminoguanidine cream with dose of 10 g/L was better than that with 5 g/L or 5 g/L + 10 g/L azone cream. One rat was not induced successfully in MI group. Four weeks after model reproduction, 4 rats died in D group and 1 rat died in AI group. The AGE content in D group was obviously higher than that in NC group [(36.8 +/- 2.6), (24.6 +/- 2.7) U per milligram hydroxyproline, respectively, t = 7.2, P < 0.01], and that in AI group [(28.6 +/- 3.7) U per milligram hydroxyproline] was also lower as compared with that in D group (t = -3.9, P < 0.05). There was no significant difference in AGE content between MI [(32.2 +/- 5.2) U per milligram hydroxyproline] and D groups (t = 1.6, P > 0.05). The percentage of KC in S phase was obviously lower in D group than in NC group [(5.3 +/- 0.6)%, (7.6 +/- 0.9)%, respectively, t = 4.50, P < 0.01], while that in MI group [(9.2 +/- 1.5)%] was higher as compared with that in D group ( t = 4.90, P < 0.01). It was more higher in AI group than in D group on KC percentage in S and G2/M phase (with t value respectively 6.80, 3.17, P values all below 0.01). The oxidative stress indexes of skin tissue in D group were all higher than those in NC group, in which levels of MPO and SOD showed statistical difference (with t value respectively 4.4, 3.7, P values all below 0.05). The oxidative stress indexes were all lower in AI group than in D group, especially in SOD level (t = -1.4, P < 0.05). Levels of MAD, MPO in MI group were significantly lower than those in D group (with t value respectively 2.6, 2.9, P values all below 0.05).

CONCLUSIONSAminoguanidine cream can promote KC proliferation and appropriately reduce oxidative stress through inhibiting AGE formation to a certain extent in skin tissue of rats with diabetes. Signal use of matrix cream can also reduce oxidative stress in skin tissue of rats with diabetes.

Administration, Cutaneous ; Animals ; Cell Proliferation ; Diabetes Mellitus, Experimental ; metabolism ; pathology ; Glycation End Products, Advanced ; metabolism ; Guanidines ; administration & dosage ; pharmacology ; Keratinocytes ; drug effects ; Male ; Ointments ; administration & dosage ; pharmacology ; Oxidative Stress ; drug effects ; Rats ; Rats, Sprague-Dawley ; Skin ; drug effects ; metabolism ; pathology

Objective To investigate the effects of advanced glycation end-products(AGEPs)on the function of normal keratinocytes in vitro so as to explore the role of AGEPs in impaired wound healing. Methods Normal rat keratinocytes were incubated with different concentrations of AGEPs.After 48 hours of culturing,the cell proliferation rates were measured by MTT colorimetric determination.The cell cycle distributions and apoptosis were analyzed with flow cytometry,and the migration was investigated by 24-well fluorimetric cell migration assay kit by exposing to 100?g/ml AGEPs.Nuclear extracts from these cells were examined for binding of nucleotides containing NF-?B consensus by immunocytochemistry and EMSA in vitro.Results The proliferations of normal keratinocytes were significantly arrested and many cells were induced to early apoptosis compared with control ones(P＜0.05)by exposing to AGEPs for 48 hours. Meanwhile AGEPs also irritated keratinocytes migration compared with control ones(P＜0.05).Inhibiting the activation of NF-?B could partly recover the proliferation of keratinocytes,reverse apoptosis and attenu- ate migration.Conclusion AGEPs are correlated with the migration,proliferation and apoptosis of kera- tinocytes by NF-?B.

AIMTo observe the effect of vitamin E (VE) on ovarian apoptosis-related protein Bcl-2 and Bax and its impact on antioxidant capacity in aged female rats and to study the senility-delaying effect and mechanism of VE on ovary.

METHODSNatural aging female rats were given different doses of exogenous VE. Then apoptosis regulatory protein Bcl-2, Bax expression in ovarian grandlose cells were detected by using immunohistochemical methods and Western blot. The contents of serum total superoxide dismutase (SOD) activity and malondialdehyde (MDA) were detected by using biochemical methods.

RESULTSContrasted with adult control group, the level of Bcl-2 expression in Senile control group was lower and the level of Bax expression was higher (P < 0.01), Serum SOD activity decreased and the level of MDA significantly increased (P < 0.01). Contrasted with senile control group, the level of Bcl-2 expression increased in VE group, the level of Bax expression decreased (P < 0.05), the level of MDA expression significantly decreased (P < 0.01).

CONCLUSIONVE can regulate apoptosis-related protein Bcl-2, Bax expression and confront free radical damage which contribute to a protective effect for ovarian grandiose cells.

Aging ; Animals ; Antioxidants ; pharmacology ; Apoptosis ; drug effects ; Female ; Granulosa Cells ; cytology ; Ovary ; cytology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rats ; Rats, Wistar ; Vitamin E ; pharmacology ; bcl-2-Associated X Protein ; metabolism

Objective To establish an HPLC method for the determination of anti-influenza drug GZ830 and its related substances, and investigate the degradation of GZ830. Methods The degradation of GZ830 under the conditions with different pH and temperatures was investigated with HPLC method to explore the degradation principle. Results The HPLC method for the determination of GZ830 was established. The linear relationship between the drug and the peak area was good in the concentration range of 160-1200 μg/ml (r=0.9998). GZ830 was easily degraded under alkaline conditions and could not tolerate high temperatures. The degradation of GZ80 did not occur when its aqueous solution was kept at 40℃ for 6 h. However, after heating at 80℃ for 6 h, about 2.4％GZ80 was degraded and the degradation rate reached about 7％ after kept at 115℃ for 30 min. Conclusion Under the optimized chromatographic conditions, GZ830 was well separated from the degraded products, and the HPLC method possessed high sensitivity and good specificity for the determination of GZ80, and the degradation of GZ80 was different under different conditions.

Objective To explore the characteristic of the genetic frequency distribution of human leukocyte antigen(HLA)-DR alleles in children with bronchial asthma in Guangxi area.Methods Eighty-four unrelated asthmatic individuals and 168 healthy people without asthma and atopy living in Nanning region of Guangxi as control group were involved in the study.All asthmatics had their serum total IgE levels measured with Pharmacia UniCAP system,and skin-prick test with 10 kinds of inhalant allergens were taken,and pulmonary function were measured among the asthmatic.HLA oligonucleotide array was used to 21 gene frequencies of HLA-DR.Results The frequencies of HLA-DR B1*070X allele and HLA-DR B1*11XX allele among the asthmatic(2.98% and 13.69%)were significantly higher than those in control group(0.3% and 5.95%)(?2 =6.915,9.478 P

Objective: To analyze the influencing factors of iron metabolism assessment in patients with myelodysplastic syndrome. Methods: MRI and/or DECT were used to detect liver and cardiac iron content in 181 patients with MDS, among whom, 41 received regular iron chelation therapy during two examinations. The adjusted ferritin (ASF) , erythropoietin (EPO) , cardiac function, liver transaminase, hepatitis antibody, and peripheral blood T cell polarization were detected and the results of myelofibrosis, splenomegaly, and cyclosporine were collected and comparative analyzed in patients. Results: We observed a positive correlation between liver iron concentration and ASF both in the MRI group and DECT groups (r=0.512 and 0.606, respectively, P<0.001) , only a weak correlation between the heart iron concentration and ASF in the MRI group (r=0.303, P<0.001) , and no significant correlation between cardiac iron concentration and ASF in the DECT group (r=0.231, P=0.053) . Moreover, transfusion dependence in liver and cardiac [MRI group was significantly associated with the concentration of iron in: LIC: (28.370±10.706) mg/g vs (7.593±3.508) mg/g, t=24.30, P<0.001; MIC: 1.81 vs 0.95, z=2.625, P<0.05; DECT group: liver VIC: (4.269±1.258) g/L vs (1.078±0.383) g/L, t=23.14, P<0.001: cardiac VIC: 1.69 vs 0.68, z=3.142, P<0.05]. The concentration of EPO in the severe iron overload group was significantly higher than that in the mild to moderate iron overload group and normal group (P<0.001) . Compared to the low-risk MDS group, the liver iron concentration in patients with MDS with cyclic sideroblasts (MDS-RS) was significantly elevated [DECT group: 3.80 (1.97, 5.51) g/L vs 1.66 (0.67, 2.94) g/L, P=0.004; MRI group: 13.7 (8.1,29.1) mg/g vs 11.6 (7.1,21.1) mg/g, P=0.032]. Factors including age, bone marrow fibrosis, splenomegaly, T cell polarization, use of cyclosporine A, liver aminotransferase, and hepatitis antibody positive had no obvious effect on iron metabolism. Conclusion: There was a positive correlation between liver iron concentration and ASF in patients with MDS, whereas there was no significant correlation between cardiac iron concentration and ASF. Iron metabolism was affected by transfusion dependence, EPO concentration, and RS.
Ferritins ; Humans ; Iron ; Iron Overload ; Liver/metabolism* ; Myelodysplastic Syndromes/therapy* ; Primary Myelofibrosis ; Retrospective Studies ; Splenomegaly