Autophagy inhibitors increase the susceptibility of KRAS-mutant human colorectal cancer cells to a combined treatment of 2-deoxy-D-glucose and lovastatin.

RAS-driven colorectal cancer relies on glucose metabolism to support uncontrolled growth. However, monotherapy with glycolysis inhibitors like 2-deoxy-D-glucose causes limited effectiveness. Recent studies suggest that anti-tumor effects of glycolysis inhibition could be improved by combination treatment with inhibitors of oxidative phosphorylation. In this study we investigated the effect of a combination of 2-deoxy-D-glucose with lovastatin (a known inhibitor of mevalonate pathway and oxidative phosphorylation) on growth of KRAS-mutant human colorectal cancer cell lines HCT116 and LoVo. A combination of lovastatin (>3.75 µM) and 2-deoxy-D-glucose (>1.25 mM) synergistically reduced cell viability, arrested cells in the G2/M phase, and induced apoptosis. The combined treatment also reduced cellular oxygen consumption and extracellular acidification rate, resulting in decreased production of ATP and lower steady-state ATP levels. Energy depletion markedly activated AMPK, inhibited mTOR and RAS signaling pathways, eventually inducing autophagy, the cellular pro-survival process under metabolic stress, whereas inhibition of autophagy by chloroquine (6.25 µM) enhanced the cytotoxic effect of the combination of lovastatin and 2-deoxy-D-glucose. These in vitro experiment results were reproduced in a nude mouse xenograft model of HCT116 cells. Our findings suggest that concurrently targeting glycolysis, oxidative phosphorylation, and autophagy may be a promising regimen for the management of RAS-driven colorectal cancers.

Novel oxazolxanthone derivatives as a new type of α-glucosidase inhibitor: synthesis, activities, inhibitory modes and synergetic effect.

Xanthone derivatives have shown good α-glucosidase inhibitory activity and have drawn increased attention as potential anti-diabetic compounds. In this study, a series of novel oxazolxanthones were designed, synthesized, and investigated as α-glucosidase inhibitors. Inhibition assays indicated that compounds 4-21 bearing oxazole rings exhibited up to 30-fold greater inhibitory activity compared to their corresponding parent compound 1b. Among them, compounds 5-21 (IC50 = 6.3 ±â€¯0.4-38.5 ±â€¯4.6 µM) were more active than 1-deoxynojirimycin (IC50 = 60.2 ±â€¯6.2 µM), a well-known α-glucosidase inhibitor. In addition, the kinetics of enzyme inhibition measured by using Lineweaver-Burk analysis shows that compound 4 is a competitive inhibitor, while compounds 15, 16 and 20 are non-competitive inhibitors. Molecular docking studies showed that compound 4 bound to the active site pocket of the enzyme while compounds 15, 16, and 20 did not. More interestingly, docking simulations reveal that some of the oxazolxanthone derivatives bind to different sites in the enzyme. This prediction was further confirmed by the synergetic inhibition experiment, and the combination of representative compounds 16 and 20 at the optimal ratio of 4:6 led to an IC50 value of 1.9 ±â€¯0.7 µM, better than the IC50 value of 7.1 ±â€¯0.9 µM for compound 16 and 8.6 ±â€¯0.9 µM for compound 20.

Enhanced catalytic activities and modified substrate preferences for taxoid 10ß-O-acetyl transferase mutants by engineering catalytic histidine residues.

OBJECTIVES: Taxoid 10ß-O-acetyl transferase (DBAT) was redesigned to enhance its catalytic activity and substrate preference for baccatin III and taxol biosynthesis. RESULTS: Residues H162, D166 and R363 were determined as potential sites within the catalytic pocket of DBAT for molecular docking and site-directed mutagenesis to modify the activity of DBAT. Enzymatic activity assays revealed that the kcat/KM values of mutant H162A/R363H, D166H, R363H, D166H/R363H acting on 10-deacetylbaccatin III were about 3, 15, 26 and 60 times higher than that of the wild type of DBAT, respectively. Substrate preference assays indicated that these mutants (H162A/R363H, D166H, R363H, D166H/R363H) could transfer acetyl group from unnatural acetyl donor (e.g. vinyl acetate, sec-butyl acetate, isobutyl acetate, amyl acetate and isoamyl acetate) to 10-deacetylbaccatin III. CONCLUSION: Taxoid 10ß-O-acetyl transferase mutants with redesigned active sites displayed increased catalytic activities and modified substrate preferences, indicating their possible application in the enzymatic synthesis of baccatin III and taxol.

Inhibitory effect of baicalin on allergic response in ovalbumin-induced allergic rhinitis guinea pigs and lipopolysaccharide-stimulated human mast cells.

OBJECTIVE: Baicalin, a flavonoid compound purified from the dry roots of Scutellaria baicalensis Georgi, has generally been used for the treatment of various allergic diseases. However, there is little information about the anti-inflammatory effects of baicalin for allergic rhinitis. This study aims to investigate the anti-allergic effect of baicalin on allergic response in ovalbumin (OVA)-induced allergic rhinitis guinea pigs and lipopolysaccharide (LPS)-stimulated human mast cells. METHODS: Using in vivo models, we evaluated the effect of baicalin on allergic rhinitis symptoms via recording the number of nasal rubs and sneezes. The levels of histamine, OVA-specific immunoglobulin E(IgE), eosinophil cationic protein (ECP) and inflammatory cytokines were measured using enzyme-linked immunosorbent assay (ELISA). The histological changes of nasal mucosa were observed by light microscope after HE staining. In vitro, the release of histamine and ß-hexosaminidase of compound 48/80-induced human mast cells were measured by ELISA and PNP-NAG colorimetry, respectively. The productions of inflammatory cytokines of LPS-stimulated human mast cells were determined using ELISA. Western blot was used to test the protein expression of JAK2, p-JAK2, STAT5, p-STAT5, IKKß, p-IKKß, IκBα, p-IκBα and NF-κB (p65) of LPS-stimulated human mast cells. RESULTS: The oral administration of baicalin at doses of 50 and 200 mg/kg improved allergic rhinitis symptoms and the histological changes of nasal mucosa and decreased the serum levels of histamine, ECP, interleukin (IL)-1ß, IL-6, IL-8, tumor necrosis factor (TNF)-α and OVA-specific IgE in OVA-induced allergic rhinitis guinea pigs. In vitro, baicalin suppressed the release of histamine and ß-hexosaminidase in compound 48/80-induced human mast cells. In addition, baicalin also inhibited the productions of inflammatory cytokines such as IL-1ß, IL-6, IL-8 and TNF-α and suppressed the phosphorylation of JAK2, STAT5, IKKß, IκBα and the nuclear translocation of NF-κB (p65) subunit in LPS-stimulated human mast cells. CONCLUSIONS: These results suggest that baicalin can effectively prevent allergic response in OVA-induced allergic rhinitis guinea pigs and inhibit inflammatory response via blocking JAK2-STAT5 and NF-κB signaling pathways in LPS-stimulated human mast cells. Considered together,the results show that baicalin may be a useful drug in the treatment of allergic rhinitis.

[Inhibitory effect of kaempferol on inflammatory response of lipopolysaccharide-stimulated human mast cells].

This study is to investigate the inhibitory effect of kaempferol on inflammatory response of lipopolysaccharide(LPS)-stimulated HMC-1 mast cells. The cytotoxicity of kaempferol to HMC-1 mast cells were analyzed by using MTT assay and then the administration concentrations of kaempferol were established. Histamine, IL-6, IL-8, IL-1ß and TNF-α were measured using ELISA assay in activated HMC-1 mast cells after incubation with various concentrations of kaempferol (10, 20 and 40 µmol.L-1). Western blot was used to test the protein expression of p-IKKß, IκBα, p-IκBα and nucleus NF-κB of LPS-induced HMC-1 mast cells after incubation with different concentrations of kaempferol. The optimal concentrations of kaempferol were defined as the range from 5 µmol.L-1 to 40 µmol.L-1. Kaempferol significantly decreased the release of histamine, IL-6, IL-8, IL-1ß and TNF-α of activated HMC-1 mast cells (P<0.01). After incubation with kaempferol, the protein expression of p-IKKß, p-IKBa and nucleus NF-κB (p65) markedly reduced in LPS-stimulated HMC-1 mast cells (P<0.01). Taken together, we concluded that kaempferol markedly inhibit mast cell-mediated inflammatory response. At the same time, kaempferol can inhibit the activation of IKKß, block the phosphorylation of IκBα, prevent NF-KB entering into the nucleus, and then decrease the release of inflammatory mediators.

Analysis of Water Distribution and Muscle Quality of Silver Carp (Hypophthalmichthys molitrix) Chunks Based on Electron-Beam Irradiation.

Electron-beam irradiation (EBI) is an efficient, safe, and nonthermal sterilization technique that is extensively used in food preservation research. Here we report the effects of different EBI doses (0, 4, 8 kGy) and preservation temperatures (room temperature [RT], 4 °C) on the muscle water distribution and muscle quality indices of silver carp chunks (SCCs). The highest entrapped water content was found in the 4-kGy-irradiated/4-°C-stored samples. The expressible moisture content (EMC) of the SCCs increased with increasing irradiation dose and was significantly lower in the RT group than in the 4 °C group. The irradiation dose and preservation temperature had no significant effect on the moisture content, whiteness value and protein content of SCCs (p > 0.05). When the irradiation dose reached 8 kGy, AV value, POV value and TVB value were significantly increased (p < 0.05). The myofibrillar protein content and actomyosin content of the SCCs in the 4 °C group was higher than that of the specimens in the RT group by 0.29−0.98 mg/mL (p < 0.05) and 36.21−296.58 µg/mL (p < 0.05), respectively. Overall, EBI treatment (4 kGy) and low-temperature preservation (4 °C) helped retain the muscle water content of the SCCs and preserve their quality, thereby endorsing the EBI treatment of silver carp products.

Computer-Aided Design of α-L-Rhamnosidase to Increase the Synthesis Efficiency of Icariside I.

Icariside I, the glycosylation product of icaritin, is a novel effective anti-cancer agent with immunological anti-tumor activity. However, very limited natural icariside I content hinders its direct extraction from plants. Therefore, we employed a computer-aided protein design strategy to improve the catalytic efficiency and substrate specificity of the α-L-rhamnosidase from Thermotoga petrophila DSM 13995, to provide a highly-efficient preparation method. Several beneficial mutants were obtained by expanding the active cavity. The catalytic efficiencies of all mutants were improved 16-200-fold compared with the wild-type TpeRha. The double-point mutant DH was the best mutant and showed the highest catalytic efficiency (k cat /K M : 193.52 s-1 M-1) against icariin, which was a 209.76-fold increase compared with the wild-type TpeRha. Besides, the single-point mutant H570A showed higher substrate specificity than that of the wild-type TpeRha in hydrolysis of different substrates. This study provides enzyme design strategies and principles for the hydrolysis of rhamnosyl natural products.

Antimicrobial activities and mechanism of sturgeon spermary protein extracts against Escherichia coli.

This study aimed to evaluate the antimicrobial activities and mechanism of sturgeon spermary protein extracts (SSPE) against Escherichia coli. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined. Cell structural change was analyzed using scanning electron microscopy-energy dispersive X-ray spectrometry and transmission electron microscope. Moreover, pH, zeta potential, membrane potential, intracellular ATP concentrations and the interaction of SSPE with genomic DNA were analyzed. Results showed that molecular weight of SSPE is 13.4 kDa, the content of basic amino acids is the highest, in which arginine accounts for 73.2%. The MIC and MBC of SSPE for E. coli were 0.05 and 5 mg/mL, respectively. After SSPE treatment, cell membrane permeability changes, zeta potential decrease and genomic DNA lysis occurred in E. coli, which indicated it exerted bacteriostatic effects either independently or simultaneously by destroying the cell membrane and genomic DNA. These findings indicated that SSPE has potential to be a natural antiseptic.

Integrative analysis of long non-coding RNA and messenger RNA expression in toll-like receptor 4-primed mesenchymal stem cells of ankylosing spondylitis.

BACKGROUND: The precise pathogenesis of ankylosing spondylitis (AS) is still largely unknown at present. Our previous study found that toll-like receptor 4 (TLR4) downregulated and performed immunoregulatory dysfunction in mesenchymal stem cells from AS patients (AS-MSCs). The aim of this study was to explore the expression profiles of long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs) in TLR4-primed AS-MSCs, and to clarify the potential mechanisms. METHODS: The immunoregulatory effects of MSCs were determined after TLR4 activation. Next, the differentially-expressed (DE) lncRNAs and mRNAs between AS-MSCs and TLR4-primed AS-MSCs [stimulated by lipopolysaccharide (LPS)] were identified via high-throughput sequencing followed by quantitative real-time PCR (qRT-PCR) confirmation. Finally, bioinformatics analyses were performed to identify the critical biological functions, signaling pathways, and associated functional networks involved in the TLR4-primed immunoregulatory function of AS-MSCs. RESULTS: A total of 147 DE lncRNAs and 698 DE mRNAs were identified between TLR4-primed AS-MSCs and unstimulated AS-MSCs. Of these, 107 lncRNAs were upregulated and 40 were downregulated (fold change ≥2, P<0.05), while 504 mRNAs were upregulated and 194 were downregulated (fold change ≥2, P<0.05). Five lncRNAs and five mRNAs with the largest fold changes were respectively verified by qRT-PCR. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses demonstrated that the DE mRNAs and lncRNAs were highly associated with the inflammatory response, such as NOD-like receptor (NLR) signaling pathway, the TNF signaling pathway and the NF-κB signaling pathway. Cis-regulation prediction revealed eight novel lncRNAs, while trans-regulation prediction revealed 15 lncRNAs, respectively. Eight core pairs of lncRNA and target mRNA in the lncRNA-transcription factor (TF)-mRNA network were as follows: PACERR-PTGS2, LOC105378085-SOD2, LOC107986655-HIVEP2, MICB-DT-MICB, LOC105373925-SP140L, LOC107984251-IFIT5, LOC112268267-GBP2, and LOC101926887-IFIT3, respectively. CONCLUSIONS: TLR4 activation in AS can enhance the immunoregulatory ability of MSCs. Eight core pairs of lncRNA and target mRNA were observed in TLR4-primed AS-MSCs, which could contribute to understanding the potential mechanism of AS-MSC immunoregulatory dysfunction.

Microbial Cell Factory of Baccatin III Preparation in Escherichia coli by Increasing DBAT Thermostability and in vivo Acetyl-CoA Supply.

Given the rapid development of genome mining in this decade, the substrate channel of paclitaxel might be identified in the near future. A robust microbial cell factory with gene dbat, encoding a key rate-limiting enzyme 10-deacetylbaccatin III-10-O-transferase (DBAT) in paclitaxel biosynthesis to synthesize the precursor baccatin III, will lay out a promising foundation for paclitaxel de novo synthesis. Here, we integrated gene dbat into the wild-type Escherichia coli BW25113 to construct strain BWD01. Yet, it was relatively unstable in baccatin III synthesis. Mutant gene dbat S189V with improved thermostability was screened out from a semi-rational mutation library of DBAT. When it was over-expressed in an engineered strain N05 with improved acetyl-CoA generation, combined with carbon source optimization of fermentation engineering, the production level of baccatin III was significantly increased. Using this combination, integrated strain N05S01 with mutant dbat S189V achieved a 10.50-fold increase in baccatin III production compared with original strain BWD01. Our findings suggest that the combination of protein engineering and metabolic engineering will become a promising strategy for paclitaxel production.

Pantoprazole (PPZ) Inhibits RANKL-Induced Osteoclast Formation and Function In Vitro and Prevents Lipopolysaccharide- (LPS-) Induced Inflammatory Calvarial Bone Loss In Vivo.

Bone remodeling is a process delicately balanced between osteoclastic bone resorption and osteoblastic bone formation. Osteoclasts (OCs) are multinucleated giant cells formed through the fusion of monocytic precursors of the hematopoietic stem cells lineage. OCs are the exclusive cells responsible for the resorption and degradation of the mineralized bone matrix. Pantoprazole (PPZ), a proton pump inhibitor (PPI), is commonly prescribed to reduce excess gastric acid production for conditions such as gastroesophageal reflux disease and peptic ulcer disease. Studies have found contradictory effects of PPI therapy on bone metabolism due to the lack of understanding of the exact underlying mechanism. In this study, we found that PPZ inhibits receptor activator of nuclear factor-κB (NF-κB) ligand- (RANKL-) induced osteoclastogenesis from bone marrow monocytic/macrophage (BMMs) precursors and the bone-resorbing activity of mature OCs. Correspondingly, the expression of OC marker genes was also attenuated. At the molecular level, PPZ treatment was associated with reduced activation of the ERK MAPK signaling pathways crucial to OC differentiation. Additionally, the in vivo administration of PPZ protected mice against lipopolysaccharide- (LPS-) induced inflammatory calvarial bone erosion, as a result of the reduced number and activity of OCs on the calvarial bone surface. Although PPI use is associated with increased risk of osteoporosis and bone fractures, our study provides evidence for the direct inhibitory effect of PPZ on OC formation and bone resorption in vitro and in vivo, suggesting a potential therapeutic use of PPZ in the treatment of osteolytic disease with localized bone destruction.

Enhanced catalytic activity of Au core Pd shell Pt cluster trimetallic nanorods for CO2 reduction.

Herein, Au core Pd shell Pt cluster nanorods (Au@Pd@Pt NRs) with enhanced catalytic activity were rationally designed for carbon dioxide (CO2) reduction. The surface composition and Pd-Pt ratios significantly influenced the catalytic activity, and the optimized structure had only a half-monolayer equivalent of Pt (θ Pt = 0.5) with 2 monolayers of Pd, which could enhance the catalytic activity for CO2 reduction by 6 fold as compared to the Pt surface at -1.5 V vs. SCE. A further increase in the loading of Pt actually reduced the catalytic activity; this inferred that a synergistic effect existed among the three different nanostructure components. Furthermore, these Au NRs could be employed to improve the photoelectrocatalytic activity by 30% at -1.5 V due to the surface plasmon resonance. An in situ SERS investigation inferred that the Au@Pd@Pt NRs (θ Pt = 0.5) were less likely to be poisoned by CO because of the Pd-Pt bimetal edge sites; due to this reason, the proposed structure exhibited highest catalytic activity. These results play an important role in the mechanistic studies of CO2 reduction and offer a new way to design new materials for the conversion of CO2 to liquid fuels.

[Surgical treatment for distal humerus type C fractures].

Distal humerus type C fracture is a rare and complicated intra-articular injury. Non-surgical treatment is difficult to achieve accurately reduction and reconstruction of articular surface, while open reduction and internal fixation is the best treatment option. Olecranon osteotomy could provide adequate surgical exposure, and is the most commonly used surgical method, but complications such as non-union osteotomy, internal fixation failure occurred. To avoid sacrificing integrity of olecranon, paratricipital approach, triceps reflecting approach, triceps reflectin ganconeus pedicle approach, triceps splitting and triceps tongue-shaped flap approach have been applied to fracture revealed. However, there is a certain contradiction of choice between surgical exposure and extension function of elbow due to limitations of different approaches. With the promotion of "double-column" theory, double plates has significant mechanical advantages over single plates. Even if parallel double-plate has more advantageous than vertical double-plate in vitro biomechanical experiments, it is not clear whether there is any difference between two methods in clinical application. Elbow arthroplasty may be the final choice for C-type fractures that could not be reconstructed on articular surface, but its long-term efficacy remains to be observed due to technical limitations.

Effects of sulindac sulfide on proliferation and apoptosis of human breast cancer cell.

The present study aimed to observe the effects of sulindac sulfide on the proliferation and apoptosis of human breast cancer cells MCF-7, and to explore the potential underlying molecular mechanism. The inhibitory ratio was detected using a cell counting kit-8 assay. The changes in cell cycle distribution were assessed using flow cytometry (FCM). Furthermore, the changes in cell apoptosis rates were detected by Hoechst 33258 staining and FCM coupled with Annexin V-FITC/propidium iodide (PI) staining. In addition, the protein expression was detected using western blotting. Sulindac sulfide was able to inhibit the proliferation of breast cancer in a dose- and time-dependent manner. In addition, sulindac sulfide altered the cell cycle of breast cancer cells. The results of Hoechst 33258 staining and FCM coupled with Annexin V-FITC/PI staining demonstrated that sulindac sulfide could significantly induce the apoptosis of MCF-7 cells in a dose-dependent, and time-dependent manner. The western blot analysis demonstrated the protein expression of Bcl-2 was downregulated, and Bax and cleaved caspase-3 were upregulated. The results of the present study suggest that sulindac sulfide can inhibit the proliferation and induce the apoptosis of MCF-7 cells.

Bio-production of Baccatin III, an Important Precursor of Paclitaxel by a Cost-Effective Approach.

Natural production of anti-cancer drug taxol from Taxus has proved to be environmentally unsustainable and economically unfeasible. Currently, bioengineering the biosynthetic pathway of taxol is an attractive alternative production approach. 10-deacetylbaccatin III-10-O-acetyl transferase (DBAT) was previously characterized as an acyltransferase, using 10-deacetylbaccatin III (10-DAB) and acetyl CoA as natural substrates, to form baccatin III in the taxol biosynthesis. Here, we report that other than the natural acetyl CoA (Ac-CoA) substrate, DBAT can also utilize vinyl acetate (VA), which is commercially available at very low cost, acylate quickly and irreversibly, as acetyl donor in the acyl transfer reaction to produce baccatin III. Furthermore, mutants were prepared via a semi-rational design in this work. A double mutant, I43S/D390R was constructed to combine the positive effects of the different single mutations on catalytic activity, and its catalytic efficiency towards 10-DAB and VA was successfully improved by 3.30-fold, compared to that of wild-type DBAT, while 2.99-fold higher than the catalytic efficiency of WT DBAT towards 10-DAB and Ac-CoA. These findings can provide a promising economically and environmentally friendly method for exploring novel acyl donors to engineer natural product pathways.

Activity Essential Residue Analysis of Taxoid 10ß-O-Acetyl Transferase for Enzymatic Synthesis of Baccatin.

Taxoid 10ß-O-acetyl transferase (DBAT) is a key enzyme in the biosynthesis of the famous anticancer drug paclitaxel, which catalyses the formation of baccatin III from 10-deacetylbaccatin III (10-DAB). However, the activity essential residues of the enzyme are still unknown, and the acylation mechanism from its natural substrate 10-deacetylbaccatin III and acetyl CoA to baccatin III remains unclear. In this study, the homology modelling, molecular docking, site-directed mutagenesis, and kinetic parameter determination of the enzyme were carried out. The results showed that the enzyme mutant DBATH162A resulted in complete loss of enzymatic activity, suggesting that the residue histidine at 162 was essential to DBAT activity. Residues D166 and R363 which were located in the pocket of the enzyme by homology modelling and molecular docking were also important for DBAT activity through the site-directed mutations. Furthermore, four amino acid residues including S31 and D34 from motif SXXD, D372 and G376 from motif DFGWG also played important roles on acylation. This was the first report of the elucidation of the activity essential residues of DBAT, making it possible for the further structural-based re-design of the enzyme for efficient biotransformation of baccatin III and paclitaxel.

Increased resveratrol production in wines using engineered wine strains Saccharomyces cerevisiae EC1118 and relaxed antibiotic or auxotrophic selection.

Resveratrol is a polyphenolic compound with diverse beneficial effects on human health. Red wine is the major dietary source of resveratrol but the amount that people can obtain from wines is limited. To increase the resveratrol production in wines, two expression vectors carrying 4-coumarate: coenzyme A ligase gene (4CL) from Arabidopsis thaliana and resveratrol synthase gene (RS) from Vitis vinifera were transformed into industrial wine strain Saccharomyces cerevisiae EC1118. When cultured with 1 mM p-coumaric acid, the engineered strains grown with and without the addition of antibiotics produced 8.249 and 3.317 mg/L of trans-resveratrol in the culture broth, respectively. Resveratrol content of the wine fermented with engineered strains was twice higher than that of the control, indicating that our engineered strains could increase the production of resveratrol during wine fermentation.

[Case-control studies on complex tibial plateau and posterior condylar fractures treated through combined anterior-posterior (small incision or micro-incision) approach].

OBJECTIVE: To study the therapeutic effects of combined anterior-posterior (small incision or micro-incision) approach for complex tibial plateau and posterior condylar fractures. METHODS: From 2000 to 2008, 79 patients (81 limbs) with complex tibial plateau and posterior condylar fractures were reviewed. There were 45 males and 34 females, ranging in age from 19 to 66 years, with an average of 40.6 years. Thirty-nine limbs were treated using small incision through combined anterior-posterior approach, in which 13 limbs were Schatzker type IV, 15 limbs were type V ,and 11 limbs were type VI. Other 42 limbs were treated using micro-incision through combined anterior-posterior approach, in which 18 limbs were Schatzker type IV, 16 limbs were type V, and 8 limbs were type VI. The Rasmussen scores for knee joint and radio scores were used to evaluate therapeutic effects after the treatment. The complications such as cutaneous necrosis and incision infection were observed. RESULTS: All the patients were followed up. According to Rasmussen criterion, in small incision group, 16 limbs got an excellent result, 13 good, 7 fair and 3 bad; in micro-incision group,above data were 19, 11, 8 and 4 respectively. Comparison between the two groups, P = 0.924. Comparison of complications such as cutaneous necrosis and incision infection: in small incision group,10 limbs had the complications, and in micro-incision group were 4 limbs; the occurrence rate of small incision group were higher than that of micro-incision group (P = 0.047). CONCLUSION: There are no significant differences between the two groups in the knee joint function rehabilitation; however, there is smaller rate for cutaneous necrosis and incision infection in micro-incision group.

Immuno-stimulating properties of diosgenyl saponins isolated from Paris polyphylla.

The effects of three diosgenyl saponins isolated from Paris polyphylla on the immuno-stimulating activity in relation to phagocytosis, respiratory burst, and nitric oxide production in mouse macrophage cells RAW 264.7 have been investigated. Our results showed that all three diosgenyl saponins significantly enhanced phagocytic activity that increased with the concentration of saponins to reach a maximum, and then tended to decrease with higher concentrations. Saponins with sugar moiety directly induced respiratory burst response in RAW 264.7 cells that increased with the concentrations and reached a maximum, then decreased with higher concentrations after 2-h incubations, however, diosgenin had no PMA-triggered respiratory burst response. Treatment of RAW 264.7 cells with saponins with sugar moiety for 24-h caused a significant increase in the production of nitric oxide, while diosgenin had no effect at all. Consequently, relationship between molecular structures of three diosgenyl saponins and their immunomodulatory activities was discussed, and a possible mechanism of immuno-stimulating function of diosgenyl saponins was accordingly explored.