BACKGROUND:Silk fibroin, chitosan, and nano hydroxyapatite are natural materials, and they al have good biological activity and physical or chemical properties. As tissue engineering materials, they have been already widely used in clinic or research work, but there are some defects in the application of these three kinds of materials. OBJECTIVE:To discuss the preparation and characteristics of silk fibroin/chitosan/nano hydroxyapatite complicated scaffolds which could be used in bone tissue engineering. METHODS:Silk fibroin, chitosan, and nano hydroxyapatite were separately prepared into 2%solution, and then mixed at the ratio of 1:1:0.5, 1:1:1, 1:1:1.5 respectively. The three-dimensional complicated scaffolds were prepared by those mixed liquids through repeated freeze drying and chemical crosslinking technology. Scanning electron microscope was used to detect the pore size of the scaffolds. Porosity, water absorption rate, and hot-water loss rate were determined. Mechanical tester was used to measure the tensile and compressive modulus of dried three-dimensional scaffolds. RESULTS AND CONCLUSION:The silk fibroin/chitosan/nano hydroxyapatite complicated scaffold in the dry state had no special smel , appeared to be a stabilized solid cylinder, and exhibited clear resiliency and flexibility with a touch. With the increased content of nano hydroxyapatite, the porosity, water absorption rate and average pore size of the scaffolds appeared to be decreased, while the hot-water loss rate and compressive strength were increased. The scaffold prepared at 1:1:1 was better for bone tissue engineering, and the average pore size, water absorption rate and hot-water loss rate were 85.67 μm, (135.65±4.56)%and (22.84±1.06)%, respectively, closer to the needs of the bone tissue engineering. Uniform pores were found within the scaffold at 1:1:1, showing the network structure, developed transport among pores, and the network structure was approximately 10μm.

Objective To investigate the feasibility of restoration of cartilage defect with silk fibrin/chitosan(SF-CS)biological scaffold compound by induced bone marrow mesenchymal stem cells (BMSCs)in the elderly rabbits.Methods BMSCs were extracted,cultured and induced to differentiate,then inoculated into SF-CS three-dimensional scaffold restoration.54 rabbits(aged 16-18months)were divided into scaffold restoration,single scaffold and control groups(n=18 per group).The right knee joint was used for building cartilage defect model and implanted by scaffolds.General observation,tissue staining and modified Wakitani histological scoring were performed at 4,8 and 12weeks after operation.Results SF-CS scaffold was structured by multiple interlinked pores.The average pore size was 151.72 μm.The porosity was(92.72±4.78)％.The imbibition rate was (141.10± 6.87)％.BMSCs was grown well and proliferated dynamically in SF-CS scaffold after induction.At 12 weeks,the cartilage defect was basically repaired,type Ⅱ collagen was positively expressed and the scaffold was almost assimilated in scaffold restoration group.In single scaffold group,the cartilage defect was repaired mainly by fiber tissue,type Ⅱ collagen was less expressed and the scaffold almost degraded while the cartilage defect was repaired badly in control group.The scaffold restoration group was superior to single scaffold and control groups(P＜0.05)in improving the Wakitani score.Conclusions The SF-CS scaffold as BMSCs carrier may restore cartilage defect in knee joint of the elderly rabbits.

BACKGROUND:Poly lactic acid as an excellent delivery has good biocompatibility.
OBJECTIVE:To prepare recombinant human bone morphogenetic protein-2 (rhBMP-2)/poly lactic acid (PLA) sustained release microspheres, and to study its physical and chemical properties.
METHODS:The rhBMP-2/PLA sustained release microspheres were prepared using w/o/w solvent evaporation method. Scanning electron microscopy, laser particle size, zeta potential, and swel ing properties were detected. ELISA kit was utilized for measurement of encapsulation efficiency, drug-loading rate and in vitro drug release rate.
RESULTS AND CONCLUSION:Under the scanning electron microscope, rhBMP-2/PLA sustained release microspheres were approximately circle with excellent dispersion. The uniform spheres were visible with a mean particle size of 839.6 nm. The zeta potential were (-32.93±3.74) mV. The swel ing coefficient was 1.157±0.059. The drug-loading rate and encapsulation efficiency of rhBMP-2/PLA sustained release microspheres were (88.943±2.878)%and (0.026±0.001)%respectively. The drug release rate at 1 day was about 10.199%, then the drug release was relatively constant, and til 19 days, the cumulative drug release rate was 54.643%. These findings indicate that the constructed rhBMP-2/PLA sustained release microspheres meet the requirement of the Chinese Pharmacopoeia (10th edition) that the encapsulation efficiency is not less than 80%and the microspheres have a good slow-release function in vitro.

Proline-rich cell wall proteins are widely spread in plants and are believed to function by modeling the architecture of the cell wall surrounding specific cell types. Five genes encoding proline-rich proteins were isolated from cotton cDNA libraries. The most common characteristic of these proteins is the abundant proline residues that occur in repeating motifs of at least two consecutive Pros. Based on amino acid composition, repetitive motifs and domain organization, the five members can be divided into two subgroups: one group similar to common PRPs including GhPRP3, GhPRP6, GhPRP5 and GhPRP4 was composed of two domains, an N-terminal hydrophobic domain (or signal peptide) followed by a proline-rich domain containing different proline-rich repetitive motifs; the other group different from common PRPs including GhPRPL lies in it contains an N-terminal hydrophilic domain, eight repetitive copies of pentapeptide (similar to PPKKE) lies in the C-terminal domain. Expression studies of the six GhPRPs have been carried out by quantitative realtime RT-PCR. The results showed that GhPRP3 and GhPRP5 were preferentially expressed in 10 dpa fiber, little transcripts was detected in other tissues examined. GhPRPL highly expressed in cotyledons, whereas smaller or negligible amounts of its transcripts were detected in other tissues. The remaining two genes, GhPRP4 and GhPRP6, were expressed in all the tissues analysed, but their transcript level is different. GhPRP4 mRNA is most abundant in hypocotyls, and then in anther, while GhPRP6 expressed highly in fiber, and then in 10 dpa ovule. Furthermore, the results showed that the fiber-specific GhPRP3 and GhPRP5 were also developmentally regulated, suggesting that the genes may play important roles during cotton fiber development.

Objective To provide the basis for clinical diagnosis and treatments of colorectal cancer by studying the clinical characteristics and incidence tendency of colorectal cancer in recent 12 years in Kunming. Methods The data of 1628 colorectal cancer patients from 2001 to 2012 in the First Affiliated Hospital of Kunming Medical University were collected. The clinic pathological data of all patients was divided into 2001-2006 (Agroup) and 2007-2012 (B group) two consecutive time periods for statistical analysis according to its time of onset. Results The incidence of colorectal cancer in the city rose from 49.2%to 56.0%, and the incidence of colorectal cancer in rural areas decreased from 50.8%to 44%( <0.05) .The location ratio of colorectal cancer in the rectum decreased from 74.8%to 73.9%, in the left half colon ratio increased from 14.8%to 19%, while the right colon ratio decreased from 10.4% to 7.1% ( < 0.05) . The change in the three parts had statistically significant difference between two consecutive time periods. The percentage of colorectal cancer patients in Dukes A stage rose from 13.4% to 18.8%, while the percentage of patients in Dukes D stage decreased from 22.3% to 14.5% ( <0.01) . Conclusions The incidence of colorectal cancer in Kunming gradually increases.The morbidity in the city is significantly higher than that in the country. The ratio of left half colon cancer is rising, but the ratio of rectal cancer and right half colon cancer is falling. The early diagnosis rate of colorectal cancer is on the rise.

BACKGROUND:Silk fibroin/chitosan/nano hydroxyapatite (SF/CS/nHA) composite scaffold constructed in preliminary experiments has good physical and chemical properties.
OBJECTIVE:To study the capacity and mechanism of SF/CS/nHA composite scaffold for repair of rabbit radial bone defects.
METHODS:Thirty-six New Zealand white rabbits were selected to make animal models of right radial bone defects, and then randomly divided into SF/CS/nHA group, SF/CS group and blank control group. Blank control group had no treatment after modeling. X-ray radiography, gross observation and histopathological observation were performed at 4, 8, 12, 16 weeks postoperatively.
RESULTS AND CONCLUSION:Sixteen weeks after surgery, bone defects in the SF/CS/nHA group were completed replaced by normal bone tissue on X-ray images, and the bone marrow cavity showed complete recanalization with new bone formation;hematoxylin-eosin staining showed bone trabecula and many fusiform bone cells. In the SF/CS group, the bone mineral density in the defect area was slightly lower than that of the normal bone tissues, the bone marrow cavity was partly rehabilitated, and many chondrocytes were seen around bone cells that arranged irregularly with no bone trabecula or bone lamel a. In the blank control group, the images of bone calcification were consistent with normal bone tissues, and a closed bone ununion was formed at each end;hematoxylin-eosin staining showed that the blank control group was fil ed by fibrous connective tissue and a smal amount of bone-like tissues. SF/CS/nHA composite scaffold is better for repair of rabbit radial bone defects.

Objective To investigate the influence of recombinant human bone morphogenetic protein‐2(rhBMP‐2)of poly lactic acid(PLA) release microspheres for compatibility of rabbit bone marrow mesenchymal stem cells(BMSCs) .Methods The rh‐BMP‐2‐PLA release microspheres were prepared by w/o/w multiple emulsion volatilizing method and then cocultured BMSCs .The effects of rhBM P‐2‐PLA release microspheres on the cytotoxicity and relative proliferation rate by MTTassay .Evaluation of mate‐rials biocompatibility by inverted microscope and scanning electron microscope(SEM) .Results The rhBMP‐2‐PLA release micro‐spheres in various concentration of leaching solution and BMSCs training of uninfected cells .Experimental group and control group in 4 different time cell proliferation OD values by analysis of repeated measurement variance between time OD values were statisti‐cally significant(P=0 .000) ,the experimental group and control group OD values are statistically significant(P=0 .025) ,the exper‐imental group higher than the control group ,experimental group OD value time there was a significant interaction effect and the group number ,the change trend are obviously different(P=0 .006) .Inverted microscope to observe materials normal cell prolifera‐tion ,SEM found that vaccination cells surrounding rhBMP‐2‐PLA release microspheres of 7 days later ,the cells grew well and split proliferation activity .Conclusion rhBMP‐2‐PLA release microspheres of BMSCs has non‐toxic and has compatibility .

BACKGROUND:Bone morphogenetic protein-2 (BMP-2) is a key to bone formation and repair.However,it has some disadvantages such as easy to lose and degrade and difficult to sustain continuous effect.OBJECTIVE:To study the preparation and properties of silk fibroin/chitosan/nano-hydroxyapatite (SF/CS/nHA) scaffold loading BMP-2.METHODS:After silk degumming,dissolution and purification,2％ SF solution was obtained.BMP-2 was dissolved in 2％ CS solution,and then fully mixed with equal volume of SF solution and proper amount of nHA.At last,the SF/CS/nHA scaffold loading BMP-2 was prepared using freeze-drying method as experimental group.The SF/CS/nHA scaffold was soaked in the BMP-2 solution as control group.The scaffold porosity was measured by Archimedes method,the surface morphology of the scaffold was observed by scanning electron microscope,the compressive strength was measured by universal testing machine.Scaffolds in the two groups were soaked in PBS,and the release of BMP-2 was measured by ELISA method at different time points.RESULTS AND CONCLUSION:(1) The scaffolds in the two groups had irregular porous structure,interconnected pores and uneven pore wall.There was no significant difference between the two groups in mean pore diameter,porosity and maximum compressive strength.(2) On the 1st day,the release rate of BMP-2 was 4.63％ in the experimental group,and the release curve increased slowly.After 28 days,the release curve of BMP-2 was transferred to the plateau stage.But in the control group,the release rate of BMP-2 on the 1stday was 58.84％,and it was a significant initial burst release.The release curve increased rapidly,and was transferred to the platform stage on the 10th day.The release rate of BMP-2 release was significantly different between the two groups at days 1,2,4,10 (P ＜ 0.05).These results show that the SF/CS/nHA scaffold loading BMP-2 could sustainably and slowly release BMP-2.

Objective To investigate the influence of simvastatin treatment on Toll-like receptor 4 (TLR4) in monocytes of peripheral blood in patients with sepsis and severe sepsis and its significance. Methods A prospective randomized controlled trial was conducted. 106 patients with sepsis and 92 patients with severe sepsis admitted to Department of Critical Care Medicine of Henan Provincial People's Hospital from August 2013 to June 2015 were enrolled. These two groups of patients were randomized into conventional treatment group and simvastatin group. All patients received treatment according to the 2012 International Sepsis Treatment Guidelines, including anti-infection drugs, nutritional support, and palliative treatment, and the patients with severe sepsis were given early goal-directed therapy (EGDT). The patients in simvastatin group received simvastatin 40 mg daily orally for at least 15 days. The peripheral blood was collected and the monocytes were isolated at 1, 5, 10, 15 days after intensive care unit (ICU) admission. TLR4 expression on the surface of TLR4/CD14+ double positive monocytes was determined by flow cytometry, and adverse reaction was observed during treatment. Results TLR4 expression on the surface of monocytes showed a tendency of decreasing with prolongation of simvastatin treatment in the simvastatin group in patients with sepsis (n = 59) or severe sepsis (n = 54). However, in patients with sepsis, TLR4 level was significantly decreased from 10 days in simvastatin group as compared with that of conventional therapy group (n = 47), and it was decreased up to 15 days [mean fluorescence intensity (MFI): 21 (19, 28) vs. 27 (25, 33) at 10 days, Z = 2.198, P = 0.021; 16 (15, 21) vs. 26 (23, 34) at 15 days, Z = 4.611, P = 0.002]. In patients with severe sepsis, there was no significant difference in TLR4 level at different time points between simvastatin group and conventional treatment group (n = 38) [MFI: 55 (52, 63) vs. 56 (48, 65) at 1 day, Z = 0.313, P = 0.692; 47 (42, 56) vs. 49 (41, 58) at 5 days, Z = 0.827, P = 0.533; 40 (35, 42) vs. 42 (37, 45) at 10 days, Z = 1.012, P = 0.301; 33 (30, 38) vs. 38 (35, 41) at 15 days, Z = 0.539, P = 0.571]. No adverse reaction related with simvastatin was found during treatment in patients with sepsis or severe sepsis. Conclusions Statins could significantly down-regulate the TLR4 expression on peripheral blood monocytes in septic patients, while it showed no significant influence on TLR4 expression in patients with severe sepsis. A different effect of statins on TLR4 expression and the downstream inflammation process in sepsis and severe sepsis patients might partially explain the discrepancy in previous reports about the therapeutic effect of statins therapy in sepsis and severe sepsis patients.

Objective To observe the action ofFu Liu Zhen (acupuncture at the six abdominal points) plus moxa-box moxibustion at umbilicus on a community population of qi and yang deficiency.Method Two hundred subjects diagnosed with qi deficiency and/or yang deficiency were randomized into a treatment group and a control group, 100 cases in each group. The two groups both received constitution-associated health education, and the treatment group was additionally intervened byFu Liu Zhen plus moxa-box moxibustion at umbilicus, while the control group didn’t receive any other intervention. The Score of Constitution in Traditional Chinese Medicine (TCM) and the number of people of each constitution were observed prior to the intervention, right after the intervention, 6 months after the intervention and 12 months after the intervention.Result In the treatment group, the scores of qi deficiency and yang deficiency after the intervention, 6 months after the intervention and 12 months after the intervention were significantly different from that prior to the intervention (P<0.01). The scores of the corresponding constitution types (qi deficiency, yang deficiency) after the intervention, 6 months after the intervention and 12 months after the intervention were significantly different from that before the intervention (P<0.01). The proportions of subjects with harmonious constitution, qi deficiency, and yang deficiency in the treatment group after the intervention, 6 months and 12 months after the intervention were significantly different from that prior to the intervention (P<0.01). The proportions of subjects with harmonious constitution, qi deficiency, and yang deficiency in the treatment group after the intervention, 6 months and 12 months after the intervention were significantly different from that in the control group (P<0.01).ConclusionFu Liu Zhen plus moxa-box moxibustion at umbilicus can improve the qi-deficiency and yang-deficiency constitutions in the community population.