Isolation and Characterization of Pepsin-Soluble Collagen from the Skin of Peru Squid (Dosidicus gigas)

Pepsin-soluble collagen (PSC) was isolated from Peru squid (Dosidicus gigas) skin and physicochemical properties of the PSC were determined. The PSC exhibited a maximum absorbance at 220 nm. Sodium dodecyl sulfate polyacrylamide gel electrophoresis suggested the collagen containing α1 and α2 chain was classified as type I collagen. Amino acid composition indicated that the collagen had lower amino acid content than that of mammalian collagen. Denaturation temperature (T_d) of the PSC was 26.8°C. The PSC had relatively high solubility in alkaline condition or NaCl concentrations below 2%. Fourier transform infrared spectroscopy investigations showed the existence of helical arrangements of collagen. The lyophilized collagen had a uniform and regular network structure. These results suggested that Peru squid skin was a potential source of collagen for further research and application.     

益生菌在水产动物肠道内黏附机制的研究进展

双歧杆菌、乳酸杆菌、芽孢杆菌、酵母是益生菌制剂中最常用、最具代表性的菌种。大多数商品化的益生菌制剂是将这些菌以单一的或复合的形式,混合在粉料载体中或添加些蛋白性原料,然后将益生菌制剂直接混合在饲料中添加或制成混合液直接加入水体。各种菌通过肠道并在肠道壁黏附发挥益生作用,能否黏附定植于肠道壁是益生菌作用的前提,也是菌种筛选的重要指标。已有的研究表明,各类微生物的黏附是非特异性的,即与菌体的结构有关,菌体能接近肠道上皮细胞,菌体的特殊配体进一步与宿主细胞相应的受体之间特异性结合,既黏附素一受体学说。     

4种天津近海产贝类牛磺酸和糖胺聚糖的粗提及含量比较

以毛蚶(Arco subcrenat)、紫贻贝(Mytilus edulis)、脉红螺(Rapana venosa)和四角蛤蜊(Mactra veneriformis)的软体部分为原料,经水煮法粗提牛磺酸和酶解法粗提糖胺聚糖,并比较了4种贝类中牛磺酸和糖胺聚糖的含量。结果表明,四角蛤蜊、毛蚶、脉红螺中的牛磺酸平均含量分别为2.708、3.532、3.662g/kg,以脉红螺中牛磺酸含量最为丰富;四角蛤蜊、紫贻贝、毛蚶中糖胺聚糖平均含量为8.27%、4.53%、3.87%,以四角蛤蜊的含量最高。     

Impact of serial activation of embryonic pax2 and WT1 genes on transdifferentiation of renal tubular epithelial cells

AIM: To investigate the mechanisms of transdifferentiation of renal tubular epithelial cells by the observation of serial activation of embryonic pax2 and WT1 genes in chronic renal failure model of 5/6 nephrectomized injury. METHODS: Embryo of mice, cultured renal tubule cells and chronic renal failure rat model of 5/6 nephrectomized injury were established. The expressions of paired Box gene (pax2) and Willm’s tumor gene (WT1), as well as α-smooth muscle actin (α-SMA), a phenotype of mesenchymal cells, were detected by RT-PCR and immunohistochemistry techniques. RESULTS: (1) Expressions of pax2 and WT1 mRNA began at 11.5 d and 14 d of embryo and increased gradually, and expression in trace quantity at 2 weeks after birth. Immunohistochemistry analysis revealed that WT1 only expressed in the glomerular podocytes and expressions of pax2 and WT1 in adult renal tubular cells were negative. (2) Deno-expression of pax2 and WT1 in some tubular cells appeared at week 2 and peaked at week 4 in 5/6 nephrectomized rats, both showing a same trend of expression. However, pax2 showed another peak at week 10 afterwards. (3) Re-expressions of pax2 and WT1 in TEC at 0.5 and 24 h after treated with IL-1α (10 μg/L) or AngII (10-9 mol/L) were observed respectively, followed by upregulation of α-SMA expression, and mesenchymal cells characters were shown. The effects were inhibited by Ang II receptor 2 antagonist and WT1 antibody, respectively. CONCLUSION: Adult renal tubule cells acquire re-activation of embryonic pax2 and WT1 genes and phenotypes of mesenchymes when challenge with certain injuries. Deno-expression of pax2 and WT1 genes closely relates with high concentration of bioactive facors, such as AngII and IL-1, which are involved in the mechanisms of renal tubule cells transdifferentiation.     

Protective effect of rAAV2-NTF2 on blood-retinal barrier damage in diabetic rats

AIM: To explore the effect of rAAV2-NTF2 on the blood-retinal barrier damage in early stage diabetic retinopathy. METHODS: Retinal vasculature was observed by Evans blue. NTF2 gene was cloned into adeno-associated virus vector pSNAV. The recombinant pSNAV-NTF2 was transfected into BHK cells. After purification, high-titer rAAV2-NTF2. rAAV2-NTF2 at dose of 4 μL (titer 1.0×1012) were injected into left eyes and rAAV2-EGFP at the same does and titer were injected into the right eyes of 36 rats. After 1 month, diabetes mellitus were induced by STZ. One month after onset of diabetes, EB at a dose of 45 mg/kg was intravenously injected into the rats. Two hours later, both eyes were enucleated immediately after perfusion of 1% PFA-citric acid buffer solution. The retinas were then dissected away and placed in formamide to extract the dye. The concentration of dye was measured by spectrophotometer. RESULTS: Evans blue was contained in normal retinal vessels without leakage, with a very low level of background fluorescence. The vessels staining of diabetic rats’ retina showed increasing fluorescence, indicating the retina-blood barrier damage. Dye concentration, representing the degree of the BRB injury, was higher in retina of 1 month diabetic rats than that in normal rats (4.67 times, P<0.01), indicating retina-blood barrier break-down. Diabetic rats with rAAV2-NTF2 intravitreal injection showed decreased BRB breakdown (P<0.05). CONCLUSION: High expression of NTF-2 decreases the BRB dysfunction in DM rats.     

Prokaryotic expression and identification of a small peptide of osteopontin

AIM: To express osteopontin 13 peptide (OPN 13) in E.coli, and to test the biological activity of the purified products. METHODS: cDNA fragments containing RGD sequences were cloned into prokaryotic expression vector pET-32c(+) including His coding sequence to construct pET-32c-OPN 13 plasmid. E.coli DH5α transformed by pET-32c-OPN 13 plasmid was induced by IPTG at different concentrations for different times to identify the optimal induction condition. Expressed His-OPN 13 fusion protein was purified via Ni-NTA His Bind Resin metal chelation chromatography, and detected by VSMCs adhesion and migration analysis. RESULTS: His-OPN 13 fusion protein was expressed in soluble manner. The fusion proteins were purified via Ni-NTA His Bind Resin affinity chromatography. His-OPN 13 fusion protein specifically inhibited adhesion and migration of VSMCs stimulated by osteopontin in dose-dependent manner. CONCLUSION: The OPN 13 peptide is successfully expressed in E.coli DH5α. The purified His-OPN 13 fusion protein could inhibit the adhesion and migration of VSMCs stimulated by osteopontin.     

Overexpression of SSTR2 inhibits growth of SSTR2-positive tumors via multiple signaling pathways

AIM: To study the anti-proliferation effect of overexpressed SSTR2 in experimental cancer with different profiles of endogenous SSTRs expressions and the possible signaling pathways involved. METHODS: In the first experiment, the growth of the tumor xenografts of the inoculated capan-2 cells and A549 cells overexpressing SSTR2 or LacZ was investigated in nude mice. In the second experiment, the adenoviral vector expressing SSTR2 were introduced into experimental capan-2 xenografts by intratumoral injection. The growth inhibition of these experiment tumors was observed and the potential influences on different signaling pathways were analyzed by immunoassays. RESULTS: Overexpression of SSTR2 inhibited the growth of tumors with different profiles of endogenous SSTRs, including experimental capan-2 xenografts that had endogenous SSTR2 expression. Overexpression significantly affected a number of components in apoptotic pathway, MAPK pathway and angiogenesis. CONCLUSION: SSTR2 is a promising candidate for gene therapy in a wide spectrum of cancers.     

河流故道区梨树-药材立体种植技术研究及分析

河流故道区梨园内间作不同药材,研究立体模式种植技术及环境效益。结果表明：梨树-半夏复合系统中,间作半夏不会造成梨减产,半夏株行距3 cm&#215;15 cm时,产量最高563.2kg/hm^2、收获指数最高4.6、商品指数最高57%。梨树-防风复合系统中,行距为40 cm时,防风产量最高21 610.8 kg/hm^2,收获指数最高9.3、商品指数最高100%。梨树-紫菀复合系统中,提出了氮、磷、钾肥料合理配比的施用量为1 hm^2中N∶P2O5∶K2O为115.8∶292.5∶303.5,紫菀产量为16 049.69 kg/hm^2,紫菀铜含量各处理间无显著差异。对3种复合系统进行土地利用价值分析。结果表明：梨树-半夏的LER为1.8最大,梨树-紫菀的LER为1.4最低。