Changes in c‐erbB‐2 Immunoexpression in Feline Endometrial Adenocarcinomas

Human epidermal growth factor receptor type 2 (c‐erbB‐2), an oncoprotein with potential prognostic marker and therapeutic use, is overexpressed in several human and animal tumours. But information regarding this molecule in feline tumours is scarce. This study aimed to assess the changes in the immunohistochemical expression of c‐erbB‐2 in feline endometrial adenocarcinomas (FEA) compared to normal endometrium. An immunohistochemistry assay using a specific antibody against c‐erbB‐2 was performed in FEA samples (n = 34) and in normal endometrium in the follicular (FS; n = 12) and luteal (LS; n = 11) stages. In FEA, the c‐erbB‐2 immunoexpression was assessed in neoplastic epithelial cells whilst in normal endometria it was individually evaluated in the surface and the superficial and deep glandular epithelia (SE, SGE and DGE, respectively). In FS and in LS, all the epithelia were positive for c‐erbB‐2; positivity was higher in the SE and the SGE than in DGE. Twenty of the 34 FEA samples (58.8%) were positive for c‐erbB‐2 immunolabelling. Nevertheless, its expression was higher in all the epithelia in the FS compared to FEA (p ≤ 0.0001) or the LS (p = 0.016). The results presented herein suggest that c‐erbB‐2 molecule is differently expressed in the feline endometrium through the oestrous cycle and though it may also be involved in feline endometrial carcinogenesis, a question remains unanswered on the importance of additional pathways of epithelial proliferation in the neoplastic changes in feline endometrium.     

Supplementation with sunflower seeds in beef cattle did not impact on oocyte and in vitro embryo production

Supplementation with compounds rich in linoleic acid, including sunflower seed supplementation, promotes increase in conception rates in cows. We aimed to evaluate whether the sunflower seed (linoleic acid source) supplementation in beef donor females alters the plasma concentrations of cholesterol, triglycerides, HDL and LDL, increases the number and quality of oocytes, increases the cleavage rates and determines an improvement in number and quality of in vitro produced blastocysts. Thus, Nelore females were divided into two groups of 15 animals to receive supplementation with or without sunflower seed for 57 days. Females underwent follicular aspiration and the oocytes were subjected to in vitro embryo production. There was no difference (p > .1) between control group and group supplemented with sunflower seed on the number of displayed follicles; number of aspired oocytes; recovery rate; cleavage rate; number of embryos; number of blastocysts; embryos number of grades I and II; plasma concentrations of total cholesterol, triglycerides; HDL and LDL. Therefore, sunflower seed supplementation in oocyte donors did not increase the number and quality of oocytes, cleavage rates and the number and quality of blastocysts produced in vitro.     

Forest road design with soil sediment evaluation using a high-resolution DEM

In this study, we combined a program to optimize forest road alignments with a method for prediction of surface erosion and related sediment delivered to streams. Combining the forest road design program with the high-resolution digital elevation model made it possible to estimate soil sediment based on a standard methodology, because a relatively accurate road prism could be generated. The combined program properly places and spaces drainage structures based on the Washington State Forest practice board manual. We applied the program to a part of Capitol State Forest in Washington State and optimized horizontal and vertical alignments while estimating movement of soil sediment from roads to streams. We discussed the effects of road surface materials, near-stream culvert distance to stream, and out-sloped forest road template on total road costs and soil sediment delivered to streams. Using lower quality rock surfacing reduced total costs, but the amount of soil sediment from lower quality rock surfacing was 1.5 times more than that on a higher quality rock traction surface. Therefore, lower quality rock surfacing should not be used near the stream. The placement of near-stream culverts 15 m upstream and out-sloped road template were examined using a traction surface on 15-m sections upstream and lower quality rock surfacing on other sections in order to reduce soil sediments to the stream and total costs. As a result, placing near-stream culverts 15 m upstream and using an out-sloped road template significantly reduced total road cost and soil sediment. Finally, we optimized horizontal and vertical alignments while limiting soil sediment to a specified maximum level. The model successfully optimized forest road alignments, which reduced total road cost as well as soil sediment.     

Efficiency of Two Enucleation Methods Connected to Handmade Cloning to Produce Transgenic Porcine Embryos

The purpose of our work was to establish an efficient-oriented enucleation method to produce transgenic embryos with handmade cloning (HMC). After 41–42 h oocytes maturation, the oocytes were further cultured with or without 0.4 μg/ml demecolcine for 45 min [chemically assisted handmade enucleation (CAHE) group vs polar body (PB) oriented handmade enucleation (OHE) group respectively]. After removal of the cumulus cells and partial digestion of the zona pellucida, oocytes with visible extrusion cones and/or polar bodies attached to the surface were subjected to oriented bisection. Putative cytoplasts without extrusion cones or PB were selected as recipients. Two cytoplasts were electrofused with one transgenic fibroblasts expressing green fluorescent protein (GFP), while non-transgenic fibroblasts were used as controls. Reconstructed embryos were cultured in Well of Wells (WOWs) with porcine zygote medium 3 (PZM-3) after activation. Cleavage and blastocyst rates were registered on day 2 and day 7 of in vitro culture respectively. Meanwhile, the total blastocyst cell number was counted on day 7. We found that the difference was only observed between blastocyst rates (38.6 ± 2% vs 48.1 ± 3%) of cloned embryos with GFP transgenic fibroblast cells after CAHE vs OHE. With adjusted time-lapse for zonae-free cloned embryos cultured in WOWs with PZM-3, it was obvious that in vitro developmental competence after CAHE was compromised when compared with the OHE method. OHE enucleation method seems to be a potential superior alternative method used for somatic cell nuclear transfer (SCNT) with transgenic fibroblast cells.     

Development of a model for inducing transient insulin resistance in the mare: preliminary implications regarding the estrous cycle

Peripheral insulin resistance is the failure of proper cellular glucose uptake in response to insulin. Insulin resistance and hyperinsulinemia are associated with several disease states in the horse and reproductive function disturbances in humans, including polycystic ovarian syndrome. To test the hypothesis that insulin resistance (IR) and hyperinsulinemia disrupt the estrous cycle in mares, two experiments were conducted to first develop a model to induce IR and to then examine the effect of this model on the duration of the estrous cycle. In Exp. 1, a hyperinsulinemic-euglycemic clamp (HEC) procedure was performed on seven mares to determine insulin sensitivity before and immediately following infusion of a heparinized lipid solution. The HEC procedure was repeated 1 wk after lipid infusion. Mares developed IR following the lipid infusion (P < 0.05), and some individuals maintained IR for up to 1 wk. Mares also exhibited increased blood insulin both immediately following treatment and 1 wk later (P < 0.05). In Exp. 2, induction of insulin resistance by lipid solution was not accompanied by changes in circulating concentrations of luteinizing hormone, and duration of the luteal phase, compared with the duration of untreated luteal phases. Nonetheless, lipid infusion and the resultant insulin resistance were associated with an increased interovulatory period (P < 0.05), and peak concentrations of progesterone (P < 0.05) were higher during the treated vs. untreated luteal phases of the estrous cycle. The results from the preliminary study suggest that infusion of a lipid solution may induce transient insulin resistance and hyperinsulinemia. The resulting insulin resistance and hyperinsulinemia may modify characteristics of the estrous cycle, perhaps at the level of the ovary.     

Oviductal Fluid Proteins Associated with the Bovine Zona Pellucida and the Effect on In Vitro Sperm–Egg Binding,Fertilization and Embryo Development

Studies have demonstrated that oviductal fluid (ODF) proteins associate with eggs of numerous species including the bovine. In this study, the association of three ODF proteins, the bovine oestrus‐associated protein, osteopontin (OPN), lipocalin‐type prostaglandin D synthase (L‐PGDS), with the bovine zona pellucida (ZP) was demonstrated by immunohistochemistry and western blot. The biological function of ODF derived egg‐associated OPN and L‐PGDS in sperm binding, fertilization and embryonic development was also explored. In vitro matured bovine oocytes were pre‐incubated with ODF collected by cannula from cows in oestrus, or ODF with antibodies to OPN, L‐PGDS and bovine serum albumin (BSA). Following incubation, oocytes were inseminated with 1 × 10⁵ frozen‐thawed spermatozoa, and they were evaluated for sperm binding, fertilization and embryonic development in vitro. Pre‐treatment of ODF with antibodies to all of proteins reduced sperm binding to the ZP and fertilization in vitro. Cleavage rates were not significantly different among incubations, but rates of embryo development were significantly decreased. We conclude that antibodies to OPN, L‐PGDS and BSA react with oocytes incubated with ODF and inhibit sperm binding, fertilization and embryonic development in vitro, suggesting a potential role of these proteins in these events.     

鹿蹄草等中草药对油桃采后褐腐病抑制效果研究

在离体和活体条件下,测定鹿蹄草、厚朴等中草药单剂及鹿蹄草为主的各中草药复配组合对油桃采后褐腐病菌的抑制效果。结果表明:11号(鹿蹄草+厚朴+大黄),14号(鹿蹄草+高良姜+虎杖)复配组合在离体试验中抑菌效果最好,10 mg/mL浓度的抑制率分别达到89.77%,85.50%;两种组合的最小抑菌浓度(MIC)分别是1.25,2.5 mg/mL;最小杀菌浓度(MBC)分别是2.5,5.0 mg/mL。活体抑菌测定结果表明,11号和14号复配组合对油桃褐腐病菌有不同程度延缓侵染和抑制作用,但不能完全防治;0.05 mg/mL扑海因与11号组合20 mg/mL,0.10 mg/mL扑海因与11号组合10 mg/mL均能完全抑制褐腐病菌在油桃果实上的发生和蔓延。     

Biomarker evidence for photosynthesis during neoproterozoic glaciation

Laterally extensive black shales were deposited on the S?o Francisco craton in southeastern Brazil during low-latitude Neoproterozoic glaciation approximately 740 to 700 million years ago. These rocks contain up to 3.0 weight % organic carbon, which we interpret as representing the preserved record of abundant marine primary productivity from glacial times. Extractable biomarkers reflect a complex and productive microbial ecosystem, including both phototrophic bacteria and eukaryotes, living in a stratified ocean with thin or absent sea ice, oxic surface waters, and euxinic conditions within the photic zone. Such an environment provides important constraints for parts of the "Snowball Earth" hypothesis.