Some problems were settled in making permanent slide for predacious nematode fungi by slide cultivation and cotton blue stain. Method of small-hole culture covered with slides could obtain high quality images, which solved the problem that trapping device couldn't be observed with high-power microscopes and oil-immersion microscopes. Scraping slide technique improved the method of making temporary slides.

BACKGROUND: Prevention of implant from inflammation was an effective method to reduce expulsion rate of stainless steel micro-screw implant, and develop new type of antibiotic material.OBJECTIVE: To evaluate the cytotoxicity of a new type of antibiotic stainless steels.METHODS: Metal test samples (antibiotic stainless steel, medical stainless steel, and medial pure titanium) were made into rectangular solids with length of 15 mm × 10 mm × 3 mm. Samples were cleaned with high temperature and high pressure. Alloy leaching liquor was prepared with DMEM culture media according to the ratio betwean surface area and volume of culture solution (3 cm~2/mL). The leaching liquor was maintained in incubator at 37 ℃ for 96 hours, and then degerming was performed using microporous membrane. 6.4% phenol was added, which was considered as the positive control group, and DMEM culture media was considered as the blank control group. Growth of MG63 cells was observed under inverted phase contrast microscope;absorbanca of cells cultured for 24, 48, 72, 96, and 120 hours was detected using MTT test; cytotoxicity of antibiotic stainless steels was evaluated.RESULTS AND CONCLUSION: ① At 24 hours after culture, calls in the positive control group was abnormal; while, cells in other groups were well adherent-grew. ② After 48 hours of culture, with the culture time increased,cytotoxicity was detected out in the positive control group; cells in other groups and blank control groups were normal and grew well. Afew of cells in stainless steels group showed karyopyknosis. ③ The absorbance was the highest of medical pure titanium, and then of antibiotic stainless steel and of medical stainless steel, while there was no significant difference between the three materials. ④ The level of cytotoxicity was grade 0. The results suggested that the antibiotic stainless steel which had the same cytotoxicity grade as medical stainless steel and pure titanium was in line with the requirement of its clinical application.

Objective To estimate the biological effects of leptin on human HaCaT keratinocytes and explore their molecular mechanisms.Methods Cell counting kit-8 (CCK-8) was used to evaluate the proliferation of cultured HaCaT cells treated with different concentrations of leptin for 24 and 48 hours.Some HaCaT cells were classified into four groups to remain untreated,be treated with leptin (100 μg/L) and piceatannol (a specific inhibitor of STAT3 phosphorylation) alone or in combination for 24 hours,respectively,followed by the evaluation of cell proliferation using CCK-8 kit.Flow cytometry was performed to assess cell cycle of HaCaT cells treated with leptin of 100 μg/L,Western blot to determine the phosphorylation level of Erk1/2 and STAT3 in HaCaT cells treated with leptin of 100 μg/L for different durations.Statistical analysis was done by Student's t-test for unpaired data using GraphPad Prism 5 software.Results The proliferation of HaCaT cells was accelerated to different degrees after treatment with leptin of 50 and 100 μg/L for 24 and 48 hours,and the accelerating effect was in a dose-dependent manner within 24 hours (r =0.9989,P ＜ 0.05).Piceatannol apparently inhibited the promotive effect of leptin on the proliferation of HaCaT cells.There was an obvious elevation in the percentage of cells at S phase ((57.70 ± 5.88)％ vs.(42.50 ± 7.55)％,P ＞ 0.05),but a significant decrease in that at G0/G1 phase ((39.70 ± 1.57)％ vs.(45.20 ± 1.44)％,P ＜ 0.05),with a significant increase in proliferation index (0.603 ±0.0157 vs.0.564 ± 0.0144,P ＜ 0.05) in HaCaT cells treated with leptin of 100 μg/L for 24 hours compared with the untreated controls.Western blot showed that leptin of 100 μg/L markedly enhanced the phosphorylation level of STAT3 in HaCaT cells.Conclusion Leptin may upregulate the proliferation of HaCaT cells through activation of STAT3 pathway.

Limonin existed in citrus fruits has been shown to have anti-bacterial, anti-viral, anti-feedant, anti-nociceptive, anti-inflammatory activities and anti-carcinogenic activities. But the clinical use is limited by its low bioavailability. The aim of this study is to observe the absorption and secretion transport mechanisms of limonin in intestine which can pave the way for the further study and clinical use. The transport characteristics and mechanisms of limonin in rat were studied by in situ intestine perfusion and in vitro Caco-2 cells method. The intestinal absorption of limonin was probably via a facilitated diffusion pathway which was poor and without segment-selection. Verapamil and ketoconazole improved the absorption remarkably according to the result of in vitro Caco-2 cells study; however, probenecid had no significant effect on the absorption. The P-gp efflux and CYP3A4 metabolism were involved in the poor intestinal absorption and low bioavailability of limonin. The exploration of the intestinal absorption mechanism is crucial to the design of dosage form and clinical use of limonin.

Objective To evaluate the effect of leptin on K17 expression in HaCaT human keratinocytes.Methods Some cultured HaCaT cells were treated with leptin (100 ng/ml) or remained untreated for 24 hours followed by the quantification of K17 mRNA expression by real-time PCR and detection of K17 protein expression by Western blot and immunofluorescence staining.To investigate the action mechanism of leptin,some cultured HaCaT cells were divided into several groups to be treated with leptin (100 ng/ml) alone,Piceatannol (an inhibitor of the STAT3 pathway) + leptin (100 ng/ml),PD-98059 (an inhibitor of the Erk1/2 pathway) + leptin (100 ng/ml),respectively for 24 hours,with the cells receiving no treatment as the negative control.Subsequently,the mRNA and protein expressions of K17 were measured by the above methods.Statistical analysis was done by the two-sample ttest.Results The mRNA expression of K17 was significantly higher in HaCaT cells treated with leptin alone than in those remaining untreated (3.086 7 ± 0.186 1 vs.1.000 0 ± 0.000 0,P ＜ 0.01),but significantly downregulated in HaCaT cells treated with Piceatannol + leptin and those with PD-98059 + leptin compared with those treated leptin alone (0.674 1 ± 0.060 0 and 0.855 0 ± 0.390 3 vs.2.242 7 ± 0.188 7,both P ＜ 0.01).The results of Western blot and immunofluorescence staining were in agreement with those of real-time PCR.Conclusions Leptin can induce K17 expression in HaCaT cells,likely by activating the STAT3 and Erk1/2 signaling pathways.

Objective:To compare the efficacy and safety between direct anterior approach (DAA) and posterior approach (PA) in total hip arthroplasty.Methods: This study evaluated postoperative results of 92 consecutive total hip arthroplasties performed by a single surgeon;44 from the DAA,and 48 from PA.The age,body mass index,operation time,blood loss,hospital stay,positioning of the artificial hip,postoperative Harris score and postoperative complications were recorded and analyzed.Results: Both the average age of the patients separately (58.0±11.9) years in DAA group and (61.0±10.4) years in PA group and the body mass index (25.1±3.7) in DAA group and (24.7±3.3) in PA group,showed no significant difference between the two groups.The DAA group had significantly reduced the hospital stay (3.8±1.7) days vs.(4.9±2.3) days for the PA group (P<0.05) and operation time was (76.0±17.4) min in DAA group,and (71.0±14.3) min in PA group (P>0.05).The amount of blood loss: in group DAA (238.0±55.3) mL,and in group PA (387.0±61.2) mL (P<0.05).There was no statistical difference in the positioning of the artificial hip: the cup anteversion in DAA group and PA group was 17.3°±5.3° vs.18.6°±5.1°,the cup inclination was 38.5°±5.7° vs.37.7°±5.2°.In DAA group,there was significantly less use of assistive devices [(24.6±7.8) d vs.(31.7±10.2) d,P<0.05],and the pain was significantly lower.Harris score at the end of 6 weeks of the follow-up: in DAA group 85.7±5.4,and in PA group 81.3±6.1 (P<0.05);at the end of the last follow-up: in DAA group 93.4±4.7,and in PA group 92.3±5.3 (P>0.05).Complications were encountered in the two groups.There were two intraoperative complications (4.4％),1 great trochanter fracture and 1 lateral cutaneous nerve injury in DAA group.No dislocation was observed in DAA group.One dislocations and 1 groin pain were recorded in PA group.No prosthesis loosening,deep vein thrombosis,sciatic nerve injury and other complications occurred in the two groups.Conclusion: Total hip arthroplasty using the anterior approach allows for superior recovery and better stability.

Objective To explore the expressions of matrix metalloproteinases-1(MMP-1) and tissue inhibitor of metalloproteinase-1(TIMP-1) in sternomastoid muscle and explore the pathogenesis of sternomastoid muscle fibrosis in congenital fibra muscular torticollis in children.Methods The hyperplastic state of collagen fiber were determine by Masson collagen stainning method and muscular torticollis and fibra torticollis was differed,obtained 22 cases of the fibra torticollis group.Immunohistochemical method was used to determine the expression of MMP-1 and TIMP-1 in sternomastoid muscle of fibra torticollis and compared them with 6 cases of control group.Results By the immunohistochemical method,the expression of MMP-1 in the experiment group significantly decreased than that in control group(P0.05).Conclusion In congenital fibra torticollis,the sternomastoid muscle fibrosis is related to the decrease of MMP-1.

To prepare virus-like particles containing FluA/B mRNA as RNA standard and control in Influenza RNA detection, the genes coding the coat protein and maturase of E. coli bacteriophage MS2 were amplified and cloned into D-pET32a vector. Then we inserted 6 histidines to MS2 coat protein by QuikChange Site-Directed Mutagenesis Kit to construct the universal expressing vector D-pET32a-CP-His. In addition, the partial gene fragments of FluA and FluB were cloned to the down-stream of expressing vector. The recombinant plasmid D-pET32a-CP-His-FluA/B was transformed to BL21 with induction by IPTG. The virus-like particles were purified by Ni+ chromatography. The virus-like particles can be detected by RT-PCR, but not PCR. They can be conserved stably for at least 3 months at both 4 degrees C and -20 degrees C. His-tagged virus-like particles are more stable and easier to purification. It can be used as RNA standard and control in Influenza virus RNA detection.
Escherichia coli ; genetics ; metabolism ; Influenza A virus ; genetics ; metabolism ; Influenza B virus ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; RNA, Viral ; genetics ; metabolism ; Recombinant Fusion Proteins ; genetics ; metabolism ; Ribonucleases ; chemistry ; Virion ; genetics ; metabolism

A total of 152 strains of endophytic fungi were isolated from the barks of Eucommia ulmoides in three regions (Lueyang country, Zunyi country, Cili country). Based on morphological characteristics and analysis of ITS sequences, these strains were identified into 8 genera. Thereinto Phomopsis, Diaporthe and Alternaria were common genera to Eucommia barks from different sites. But the dominant genus was different: Alternaria was the dominant genus in the barks from Cili country, and Phomopsis was the dominant genus from Zunyi country, then Diaporthe was the one from Lueyang country. According to the similarity coefficient, the composition of the endophytic fungi was distinctly different between the barks from three sites. The diversity and species richness in Lueyang country and Cili country were found higher than those in Zunyi country. The evenness of endophytic fungi was 0.936 5 in Lueyang county, which was higher than 0.737 1 or 0.641 0 in Cili county or Zunyi county, respectively. After phylogenic analysis and calculating the genetic distances of typical strains belong to Phomopsis and its perfect stage--Diaporthe, there was very high genetic diversity in the two genera from our study. In conclusion, the community structure and diversity of endophytic fungi were significant different in Eucommia barks from the three habitats.
DNA, Fungal ; genetics ; DNA, Intergenic ; genetics ; Ecosystem ; Endophytes ; classification ; physiology ; Eucommiaceae ; microbiology ; Fungi ; classification ; genetics ; physiology ; Phylogeny ; Plant Bark ; microbiology

BACKGROUND:316L-Cu antibacterial stainless steel is made by adding a certain amount of copper into the stainless steel fol owed by a special heat treatment to uniformly disperse copper-rich precipitates in stainless steel substrate, thereby harvesting the antibacterial properties. OBJECTIVE:To evaluate the antibacterial activity of the Cu ions released from 316L type Cu-bearing antibacterial stainless steels against Porphyromonas gingivalis, thereby providing biomedical evidence for its clinical application. METHODS:The medical 316L stainless steel samples at a surface area to volume ratio of 0.1 cm2/L were soaked in simulated body fluids at 37 ℃ for 1-10 days. A graphite furnace atomic absorption spectrophotometer was employed to detect the amount of Cu release in the simulated body fluids each day and then the rate of Cu release per day could be determined. The antibacterial activities of the steel samples were evaluated by a standard film-covered method under a scanning electron microscope. RESULTS AND CONCLUSION:The daily Cu releasing amount from the 316L-Cu stainless steel within 10 days was significantly higher than that of 316L stainless steel, and al the values remained nearly constant. With time, the sterilizing rate of 316L-Cu stainless steel was gradual y increased, and reached 100%until the 10th hour. Porphyromonas gingivalis showed some morphological changes at 3 hours after treated with 316L-Cu stainless steel, appeared with cleavage at 6 hours, and mostly disintegrated into pieces at 9 hours. The results indicated that the 316L-Cu antibacterial stainless steel showed excel ent antibacterial property against Porphyromonas gingivalis, slowly release Cu irons, and alter the surrounding microenvironment, which is a highly promising biomaterial and has good clinical value.