Silencing a Chitinase Gene, PstChia1, Reduces Virulence of Puccinia striiformis f. sp. tritici.

Chitin is the main component of fungal cell walls, which can be recognized by pattern recognition receptors (PRRs) as pathogen-associated molecular patterns (PAMP). Chitinase in filamentous fungi has been reported to degrade immunogenic chitin oligomers, thereby preventing chitin-induced immune activation. In this study, we identified the chitinase families in 10 fungal genomes. A total of 131 chitinase genes were identified. Among the chitinase families, 16 chitinase genes from Puccinia striiformis f. sp. tritici (Pst) were identified, and the expression of PstChia1 was the highest during Pst infection. Further studies indicated that PstChia1 is highly induced during the early stages of the interaction of wheat and Pst and has chitinase enzyme activity. The silencing of PstChia1 revealed that PstChia1 limited the growth and reduced the virulence of Pst. The expression level of TaPR1 and TaPR2 was induced in PstChia1 knockdown plants, suggesting that PstChia1 is involved in regulating wheat resistance to Pst. Our data suggest that PstChia1 contributes to pathogenicity by interfering with plant immunity and regulating the growth of Pst.

Cassaine diterpenoids from the seeds of Erythrophleum fordii Oliv. and their antiangiogenic activity.

Fourteen undescribed cassaine diterpenoids along with nine known ones were isolated from the seeds of Erythrophleum fordii Oliv. (Leguminosae). In addition, subsequent structural modification yielded ten derivatives. Their chemical structures were established by extensive spectroscopic methods and acid hydrolysis. All the diterpenoids were screened for their antiangiogenic activity using the human umbilical vein endothelial cell (HUVEC) model. Five compounds were active, of which three possessed excellent activity as their effect was better than that of the positive control (SU5416). The structure-activity relationship analysis revealed that the side chain at C-13 was the key part affecting the inhibitory effect. Further study demonstrated that 3ß-hydroxynorerythrosuamine-3-O-ß-D-glucopyranoside and the formate of 3ß-hydroxynorerythrosuamine-3-O-ß-D-glucopyranoside significantly inhibited a series of angiogenic processes including proliferation, migration and capillary-like structure formation of endothelial cells. These findings may provide a new type of antiangiogenic agent for future cancer drug development.

Analysis on Three-Dimensional Strength Influencing Factors and Control Measures of Asphalt Mixtures.

Strength is an important parameter for the design of asphalt pavement materials and structures. To study the influence of various factors on the three-dimensional strength of asphalt mixtures, three aggregate gradations (dense-graded bituminous mixture AC-13, stone mastic asphalt SMA-13 and bituminous stabilization aggregate paving mixture OGFC-13) and two binders (SBS modified bitumen and 70# base bitumen) were used to prepare the asphalt mixture specimens. Among them, SBS+SMA-13 asphalt mixture has the best performance. On this basis, the uniaxial compressive test, uniaxial tensile test and confining triaxial test were conducted on the SBS+SMA-13 asphalt mixture under six oil-stone ratios conditions (5.5%, 5.7%, 5.9%, 6.1%, 6.3%, and 6.5%), six temperatures conditions (5 °C, 10 °C, 15 °C, 20 °C, 25 °C, and 30 °C), and five loading rates conditions (1 mm/min, 2 mm/min, 3 mm/min, 4 mm/min, and 5 mm/min). In addition, a unified three-dimensional strength calculation model considering the influence of temperature, loading rate, and oil-stone ratio was proposed, and the change law of the three-dimensional strength with these above factors was revealed. Furthermore, two sets of three-factor three-level orthogonal tests were carried out on the SMA-13 asphalt mixture. The sensitivity analysis and strength regulation research between three-dimensional strength and each factor were carried out. The results show that the type of asphalt has the greatest influence on the strength of the mixture, the temperature has the second most influence, the loading rate has less influence, and the oil-stone ratio has the least influence. The strength regulations proposed to improve the strength of the asphalt mixture include the use of modified asphalt, high-temperature stability high-quality asphalt, and the lower oil-stone ratio than the Marshall optimal oil-stone ratio. The strength control measures proposed from the perspective of the three-dimensional stress state, the joint failure of each stress components and real stress states are taken into consideration.

[Research of factors related to invasion and metastasis in choroidal melanoma].

OBJECTIVE: To investigate the expression of factors related to invasion and metastasis in choroidal melanoma and to determine their relationships with malignant features. METHODS: The expression of Connexin43 (Cx43), epithelial cadherin (E-cadherin), phosphatidylinositol 3-kinase (PI3K) and connective tissue growth factor (CTGF) in choroidal melanoma and nevi were detected by immunohistochemistry, and the correlation between these factors and clinicopathological features were analyzed. RESULTS: Positive rates of Cx43, E-cadherin, PI3K and CTGF were 74.07% (20/27), 44.4% (12/27), 74.07% (20/27) and 66.67% (18/27) in choroidal melanoma tissues, respectively; and 33.33% (5/15), 86.67% (13/15), 33.33% (5/15) and 20.00% (3/15) in the nevi tissues, respectively. There were significant differences in the expression of these markers between the two groups (χ(2) = 5.060, P = 0.024; χ(2) = 5.490, P = 0.019; χ(2) = 5.060, P = 0.024; χ(2) = 6.637, P = 0.010). The expression rates of Cx43 protein were 40% (4/10), 88.89% (8/9) and 100% (8/8) in spindle, mixed and epithelioid cell type, respectively. The expression of these data was related to histological type (χ(2) = 9.874, P = 0.007). The expression rates of PI3K protein were 42.86% (3/7), 75% (9/12) and 100% (8/8), in small, medium and large tumors, respectively, and their expression were co-related to the tumor size (χ(2) = 6.357, P = 0.042). Positive rates of Cx43, E-cadherin, PI3K and CTGF were 50% (6/12), 83.33% (10/12), 50% (6/12) and 41.66% (5/12), respectively, in choroidal melanoma tissues without sclera invasion and were 93.33% (14/15), 40% (6/15), 93.33% (14/15) and 86.67% (13/15), respectively, in choroidal melanoma tissues with violation involved the sclera. There were significant differences of the expression of these markers between the two groups (χ(2) = 4.457, P = 0.016; χ(2) = 3.546, P = 0.028; χ(2) = 4.457, P = 0.016; χ(2) = 4.218, P = 0.019). CONCLUSION: Increased expression of Cx43, PI3K and CTGF and decreased expression of E-cadherin are involved in the processes of invasion and metastasis of choroidal melanoma.

Expression and significance of factors related to angiogenesis in choroidal melanoma.

AIM: TO INVESTIGATE EXPRESSION OF FACTORS RELATED TO ANGIOGENESIS: HIF-1α, iNOS, COX-2 and VEGF in choroidal melanoma and its clinical significance. METHODS: Fifty samples of choroidal melanoma and 15 samples of melanocytic nevi of the eyelid identified by pathology were collected. Immunohistochemistry SP method was used to examine the expression of HIF-1α, iNOS, COX-2 and VEGF in these samples. The comparison among groups was done by SPSS 13.0 software. RESULTS: The positive expression rates of HIF-1α, iNOS, COX-2 and VEGF in choroidal melanoma group were significantly higher than those in eyelid nevi group (χ(2)=6.5542, 7.7224, 8.5828, 15.1749). The positive expression rate of VEGF was associated with the tumor size (χ(2)=10.9194), but was not associated with pathological type (χ(2)=2.0712) and the situation of scleral invasion (χ(2)=5.4289). The positive expression rate of HIF-1α was associated with the tumor size (χ(2)=7.1216) and pathological type (χ(2)=9.0889), but was not associated with the situation of scleral invasion (χ(2)=3.3586). The positive expression rate of iNOS was associated with the tumor size (χ(2)=9.5503), but was not associated with pathological type (χ(2)=1.9450) and the situation of scleral invasion (χ(2)=2.3810). The positive expression rate of COX-2 was associated with the tumor size (χ(2)=7.2970), but was not associated with pathological type (χ(2)=1.8421) and the situation of scleral invasion (χ(2)=0.4018). The expression of HIF-1α, iNOS and COX-2 were significantly associated with the expression of VEGF (r=0.9429, 1, 0.9857). The expression of COX-2 was significantly associated with the expression of iNOS (r=0.9857). The expression of HIF-1α was significantly associated with the expression of COX-2 (r=0.9857). The expression of HIF-1α was significantly associated with the expression of iNOS (r=0. 9429). CONCLUSION: The expression of HIF-1α, iNOS and COX-2 protein in choroidal melanoma were higher and may relate to angiogenesis and stimulate tumor growth. Determination of HIF-1α, iNOS and COX-2 may be helpful for the diagnosis and therapy of this tumor.

Expression of connexin 43 and E-cadherin in choroidal melanoma.

AIM: To investigate the expression of connexin 43 and epithelial cadherin (E-cadherin) in choroidal melanoma, to explore the clinical and pathological implications of expression of these proteins, and to determine their relations with malignant features. METHODS: The expression of connexin 43 and E-cadherin in choroidal melanoma were detected by immunohistochemistry and correlated with clinicopathological features. RESULTS: Positive rates of connexin 43 in choroidal melanomas and benign pigmented nevus tissues were 75% and 40% respectively with significant differences between the two groups (χ(2)=5.607, P=0.009). Positive rates of E-cadherin in choroidal melanomas and benign pigmented nevus tissues were 40% and 75% respectively with significant differences between the two groups (χ(2)=5.214, P=0.010). Significant overexpression of connexin 43 and reduction of E-cadherin expression was associated with the invasion to the sclera, and there were respectively significant differences between without and with scleral invasion groups (χ(2)=2.880, P=0.040; χ(2)=2.778, P=0.046). Overexpression of connexin 43 were correlated with tumor cell types and the expression of connexin 43 and E-cadherin may be correlated with each other. CONCLUSION: The increased expression of connexin 43 and the decreased expression of E-cadherin may be involved in the process of invasion of choroidal melanoma. The overepression of connexin 43 and reduction of E-cadherin may contribute to the development of choroidal melanoma.

Decrease of genital organ weights and plasma testosterone levels in rats following oral administration of leuprolide microemulsion.

Studies were conducted to develop oral leuprolide microemulsions using oleic acid as an absorption enhancer and to evaluate its absorption and pharmacological responses in rats. Oral administration of leuprolide microemulsion at a dose of 3 mg/kg showed a greater in vivo exposure level (C(max) and AUC) than its saline solution. When male rats were orally given a microemulsion formulation of leuprolide acetate at 0.25, 0.5, and 1mg/day for 14 consecutive days, a significant decrease in testis, prostate and seminal vesicle weights was observed. In a 35-day study, the reduction of the male genital organ weights by once a day treatment (2 mg/rat, qd) was similar to that by twice a day treatment (1 mg/rat, bid) at the same dose level. From both 14- and 35-day studies, plasma testosterone levels were sharply increased at the beginning of the treatment, and then significantly decreased to below normal control level which was also maintained during the treatment. In female rats, similar reduction of uterus and ovary weights was obtained following oral administration of leuprolide microemulsion for 35 days. These antagonistic activities from oral leuprolide microemulsion were similar to a single subcutaneous injection of Lupron depot (3.75 mg/rat), a commercial leuprolide product. The results indicated that leuprolide absorbed into systemic blood circulation from the oral microemulsion containing oleic acid reached the plasma level which can exert its pharmacological effects. Increasing oral absorption of leuprolide observed in this study could be mediated by improved membrane permeation from oleic acid and reduced enzymatic degradation from microemulsions. These findings suggest that systemic absorption of highly water-soluble protein or peptide drugs could be enhanced by oral microemulsions containing oleic acid.