基于流域尺度的水环境分区识别与评价——以辽河流域(浑太水系)山水林田湖草沙一体化保护和修复工程为例

以“辽河流域(浑太水系)山水林田湖草沙一体化保护和修复工程”项目为依托,针对浑太水系流域上游修复区,建立了主成分分析、层次聚类、水质标识指数与反距离权重插值法相结合的水环境评价体系。利用主成分分析法进行指标降维筛选,对所选指标进行方差检验,通过聚类分析以及差异化将173个监测点分组,利用水质标识指数法对分组后的均值进行评价,再将结果分配到对应监测点。基于流域尺度与行政区划相结合的方法进行水环境评价分区,对研究区进行分区反距离插值,得到浑太水系流域上游的水环境状况。结果表明:173个监测点共分为14组,一组处于Ⅳ类水质,一组处于劣Ⅴ类水质,剩余12组均为Ⅲ类水质。以浑河流域和太子河流域为大界限,浑河流域被划分为5个评价区,太子河流域被划分为4个评价区,其中有6个分区整体处于Ⅲ类水平;浑太流域上游水质状况较好,仅太子河流域中的清河、浑河流域中的大柳河为Ⅳ类,以及浑河流域内的李石河和古城子河流域为劣Ⅴ类,其余均为Ⅲ类;清河和大柳河主要为五日生化需氧量(BOD 5)超标,李石河和古城子河均为金属镉超标。进行流域尺度的水环境分区识别与评价,可打破以往水环境评价中的行政区域界限,从而以中小流域为单位,统筹把握流域水环境状况。研究成果可为浑太流域上游水环境治理提供依据,建立的研究思路和结果也可为其他流域的水环境评价工作提供参考与借鉴。

Isolation, Purification and Cryopreservation of Cells from Neonatal Bovine Testis

The development and application of spermatogonial stem cell technology have an important significance in animal cloning, preservation of endangered species and spermatogenesis research. In this study, the seminiferous epithlium cells were isolated and purified, the ceils were cryopreserved after identification, and the effects of different purification and cyopreservation methods on bovine testicular cells were studied. The results showed that there were spermatogonial stem cells and sertoli cells in the neonatal bovine seminiferous tubules, differential adherent selection methods could effectively separate these two cell types. Spermatogonial stem cells were positive after AKP, C-kit, and OCT-4 identification; sertoli ceils were positive after oil red O and vimentin identification. Frozen stock solution supplemented with 10% DMSO had the best effect in spermatogonial stem cell cryopreservation, while fxozen stock solution supplemented with 10% of ethylene glycol and 0.1 mmol. L^-1 trehalose had the best effect in sertoli cells crvooreservation.

CDC25B Involved in Proliferation of Sertoli Cells of New Born Calves Through FSH and Possibly Being Key Regulating Factor

The effects of FSH on the proliferation of sertoli cells of new born calves were studied in order to provide some data for theoretical research and practical use of spermatogenesis in vitro. Different concentrations of FSH （0, 0.01, 0.02, 0.04, and 0.08 IU· mL^-1） were taken to treat bovine sertoli cells in vitro culture, the number of sertoli cells and the expression of seven genes were determined at 6, 12 and 24 h after FSH treatments. FSH could significantly promote the proliferation of in vitro cultured sertoli cells. FSH had no significant effects on the expression of CDC25A and could significantly improve the expression of CDC25B. 0.04 IU· mL^-1 and 0.08 IU· mL^-1 FSH treatments decreased the expression of CDC25C at 12 h. 0.08 IU· mL^-1 FSH treatment decreased the expression of CDC25C at 24 h. 0.04 IU. mLI FSH could significantly decrease the expression of GSK-3β and improve the expression of β-catenin at 6, 12 and 24 h. 0.02, 0.04 and 0.08 IU· mL^-1 FSH treatments enhanced the expressions of CYCLIND1 and C-MYC. In conclusion, FSH promoted the proliferation of sertoli cells and 0.04 IU· mL^-1 FSH concentration could significantly promote the proliferation of in vitro cultured sertoli cells. FSH promoted the proliferation of sertoli cells by CDC25B and WNT/ β-eatenin and CDC25B might be the key regulator to the proliferating rate of sertoli cells of bovine calf.

Expression of Inflammatory Factors in Bovine Reproductive Tract During Follicular and Luteal Phases

In order to improve reproduvtive efficieny and understand reproduvtive defense mechanism, the oviduct, uterine horn and uterine body of bovine were used to detect the changes of inflammatory factors and the relationship between estrogen and progesterone receptor protein during estrous cycle by real-time PCR and Elisa method. The results showed that interleukin-4（IL-4）, interleukin-6（IL-6）, interleukin-10（IL-10）, interleukin-1α（IL-1α） and interleukin-1β（IL-1β） were expressed in cow oviduct, uterine horn and uterine body. In the follicular phase and the luteal phase, m RNA expression of five inflammatory factors in the uterine horn and uterine body was higher than that in the oviduct. In the follicular phase, IL-10 was highly expressed in the uterine horn and uterine body, IL-4 was highly expressed in the uterine horn, uterine body and oviduct. Additionally, in the luteal phase, IL-6 and IL-1β were highly expressed in the uterine horn, uterine body and oviduct, and the highest expression of IL-1β was observed in the uterine horn. The levels of Estrogen Receptor（ERα） protein in the oviduct, uterine horn and uterine body significantly increased in the follicular phase. The levels of Progesterone Receptor（PR） protein in the same portions of the reproductive tract in the luteal phase were significantly higher than those in the follicular phase. IL-4 and IL-10 in the cow reproductive tract might play a major role in the follicular phase, while IL-6 and IL-1β might play a major role in the luteal phase. The expression of five inflammatory factors was not directly regulated by ERα and PR.

Study on Semen Freezing Preservation of German Shepherd Dogs

In order to prolong semen preservation and improve pregnancy rate, semen freezing was studied with German shepherd dogs for experimental animals. The semen of four dogs was collected 40 times in four dilution frozen into two formulations, according to the sperm motility to compare the advantages and disadvantages. The result indicated that the sperm motility of the pellet frozen semen in dilute 2 was significantly higher than that in dilution 1, 3, and 4 （P0.01）. The sperm motility of dogs semen with fried smoked method was notablely higher than that of frozen semen of program method （P0.01）. The dilution which contained yolk-Tris mainly was the best; the pellet semen frozen with fried smoked method was superior to tuble semen frozen with program freezing method; sperm motility of pellet semen was higher than that of tuble semen in the same dilution. The conception rate and litter size of the natural matting were higher than AI.

富血小板血浆促进成纤维细胞的生长和迁移

目的探讨富血小板血浆对成纤维细胞生长和迁移的作用。方法获取大鼠成纤维细胞,提取大鼠富血小板血浆。分别以大鼠血清(对照组)、血小板浓度为4×1011/L、8×1011/L和12×1011/L的富血小板血浆处理细胞,采用MTT试验检测不同浓度的富血小板血浆对成纤维细胞增殖的作用;分别以大鼠血清(对照组)、血小板浓度为4×1011/L和8×1011/L的富血小板血浆处理细胞,通过Annexin V-FITC/PI染色法观察富血小板血浆对成纤维细胞凋亡的影响;Transwell实验检测富血小板血浆对成纤维细胞迁移的作用;通过Western blot实验检测富血小板血浆对成纤维细胞中TGF-β1、VEGF和bFGF蛋白的影响。结果与对照组相比,当血小板浓度为4×1011/L、8×1011/L和12×1011/L的富血小板血浆作用细胞24 h时,对细胞增殖的促进比例分别为(23.66±5.83)%、(65.45±9.76)%和(90.59±7.69)%;不同血小板浓度的富血小板血浆均可以抑制成纤维细胞的凋亡,并促进其迁移;与对照组相比其差异具有统计学意义(P<0.05)。富血小板血浆能够促进成纤维细胞中TGF-β1、VEGF和b FGF的表达。结论富血小板血浆能够通过增加成纤维细胞中的多种生长因子实现促进成纤维细胞生长和迁移的作用。

富血小板血浆促进骨髓间充质干细胞对大鼠骨缺损的修复作用

目的探讨富血小板血浆(platelet-rich plasma, PRP)对大鼠骨缺损的修复作用。方法提取大鼠骨髓间充质干细胞(bone marrow stromal cells, BMSCs),获得血小板浓度为2×10~5/L、2×10~6/L和2×10~7/L的PRP,分别将不同浓度的PRP与BMSCs共培养,通过MTT实验观察PRP对BMSCs细胞增殖的影响,ELISA实验检测PRP对BMSCs中碱性磷酸酶(alkaline phosphatase,ALP)的作用。构建大鼠股骨节段缺损模型,分别使用BMSCs和PRP(血小板2×10~5/L)混合物;BMSCs和PRP(血小板2×10~6/L)混合物;BMSCs和PRP(血小板2×10~6/L)来填补模型中的缺陷部位。8周后处死大鼠,评估缺损修复情况。通过Western Blot和Real-Time PCR检测断端骨组织内BMP-2和Collagen I含量,免疫组织化学染色检测组织内BMP-2含量。结果不同浓度的PRP对BMSCs的增殖和ALP的表达情况均有促进作用,且随血小板浓度的增加而加剧。PRP的添加能够促进大鼠骨缺损模型中的骨再生作用,促进BMP-2和Collagen I的表达,其促进作用具有剂量依赖性。BMSCs单独处理组能够在一定程度上促进骨缺损的修复,但作用较慢。PRP单独应用对骨缺损的修复作用较小。结论 PRP具有促进BMSCs对大鼠骨缺损的修复作用。