枯草杆菌发酵对何首乌抗氧化成分及肝癌细胞抗增殖作用的影响

[目的]探讨何首乌经枯草杆菌发酵后主要抗氧化成分及其对肝癌细胞抗增殖作用效果的变化,为进一步研究何首乌发酵炮制方法及开发何首乌产品提供科学参考.[方法]将生何首乌用枯草杆菌发酵处理后,测定生品和发酵品的游离型蒽醌、结合型蒽醌和总多酚含量等成分的变化,并对比何首乌生品和发酵品对肝癌细胞HepG2抗增殖作用的变化.[结果]何首乌生品和发酵品总游离型蒽醌含量分别为1.56和1.60 mg/g,总结合型蒽醌含量分别为1.24和0.79 mg/g,何首乌经枯草杆菌发酵后结合型蒽醌含量极显著下降(P<0.01,下同).何首乌生品和发酵品水提取液的总多酚含量分别为44.88和74.01 mg/g,乙醇提取液的总多酚含量分别为61.84和93.86 mg/g,何首乌经枯草杆菌发酵后总多酚含量极显著增加.0.025 g/L何首乌生品对肝癌细胞HepG2作用24 h后的细胞抑制率为4.2%,0.25 g/L何首乌生品对肝癌细胞HepG2作用72 h后的细胞抑制率为-11.1%(负值表示促进细胞生长),即随着何首乌生品浓度增加和作用时间延长,其对肝癌细胞生长的作用由抑制转变为促进.经枯草杆菌发酵后,0.025 g/L何首乌发酵品对肝癌细胞HepG2作用24 h后的细胞抑制率为9.9%,0.25 g/L何首乌发酵品对肝癌细胞HepG2作用72 h后的细胞抑制率为20.9%,即随着何首乌发酵品浓度增加和作用时间延长,其对肝癌细胞HepG2有明显的抗增殖效果.[结论]枯草杆菌发酵可降低何首乌的毒性,增强其对肝癌细胞HepG2的抗增殖作用,表明使用枯草杆菌发酵何首乌的炮制方法是一种安全有效的炮制减毒方法,对开发相应何首乌炮制品有重要意义.     

Effects of food enriched with egg yolk hydrolysate (bone peptide) on bone metabolism in orchidectomized dogs

We examined the effects of chicken egg hydrolysate (also known as “bone peptide” or BP) on bone metabolism in 5- to 8-month-old orchidectomized dogs. The bone formation marker serum bone alkaline phosphatase (BAP) and the bone resorption marker urine deoxypyridinoline (DPD) were used as indicators to measure changes in bone metabolism. The following results were observed that Serum BAP was higher in dogs fed BP-enriched food throughout the clinical investigation. Serum BAP was statistically significantly higher in dogs fed BP-enriched food than in dogs fed non-BP-enriched food at 2 months after orchidectomy. This suggests that BP promoted bone formation immediately after orchidectomy.     

Cell-free extract from porcine induced pluripotent stem cells can affect porcine somatic cell nuclear reprogramming

Pretreatment of somatic cells with undifferentiated cell extracts, such as embryonic stem cells and mammalian oocytes, is an attractive alternative method for reprogramming control. The properties of induced pluripotent stem cells (iPSCs) are similar to those of embryonic stem cells; however, no studies have reported somatic cell nuclear reprogramming using iPSC extracts. Therefore, this study aimed to evaluate the effects of porcine iPSC extracts treatment on porcine ear fibroblasts and early development of porcine cloned embryos produced from porcine ear skin fibroblasts pretreated with the porcine iPSC extracts. The Chariot^TM reagent system was used to deliver the iPSC extracts into cultured porcine ear skin fibroblasts. The iPSC extracts-treated cells (iPSC-treated cells) were cultured for 3 days and used for analyzing histone modification and somatic cell nuclear transfer. Compared to the results for nontreated cells, the trimethylation status of histone H3 lysine residue 9 (H3K9) in the iPSC-treated cells significantly decreased. The expression of Jmjd2b, the H3K9 trimethylation-specific demethylase gene, significantly increased in the iPSC-treated cells; conversely, the expression of the proapoptotic genes, Bax and p53, significantly decreased. When the iPSC-treated cells were transferred into enucleated porcine oocytes, no differences were observed in blastocyst development and total cell number in blastocysts compared with the results for control cells. However, H3K9 trimethylation of pronuclear-stage-cloned embryos significantly decreased in the iPSC-treated cells. Additionally, Bax and p53 gene expression in the blastocysts was significantly lower in iPSC-treated cells than in control cells. To our knowledge, this study is the first to show that an extracts of porcine iPSCs can affect histone modification and gene expression in porcine ear skin fibroblasts and cloned embryos.     

Mdivi-1, mitochondrial fission inhibitor,impairs developmental competence and mitochondrial function of embryos and cells in pigs

Mitochondria are highly dynamic organelles that undergo constant fusion/fission as well as activities orchestrated by large dynamin-related GTPases. These dynamic mitochondrial processes influence mitochondrial morphology, size and function. Therefore, this study was conducted to evaluate the effects of mitochondrial fission inhibitor, mdivi-1, on developmental competence and mitochondrial function of porcine embryos and primary cells. Presumptive porcine embryos were cultured in PZM-3 medium supplemented with mdivi-1 (0, 10 and 50 μM) for 6 days. Porcine fibroblast cells were cultured in growth medium with mdivi-1 (0 and 50 μM) for 2 days. Our results showed that the rate of blastocyst production and cell growth in the mdivi-1 (50 μM) treated group was lower than that of the control group (P < 0.05). Moreover, loss of mitochondrial membrane potential in the mdivi-1 (50 μM) treated group was increased relative to the control group (P < 0.05). Subsequent evaluation revealed that the intracellular levels of reactive oxygen species (ROS) and the apoptotic index were increased by mdivi-1 (50 μM) treatment (P < 0.05). Finally, the expression of mitochondrial fission-related protein (Drp 1) was lower in the embryos and cells in the mdivi-1-treated group than the control group. Taken together, these results indicate that mdivi-1 treatment may inhibit developmental competence and mitochondrial function in porcine embryos and primary cells.     

The use of the green fluorescent protein as a biomarker for sapstain fungi

To understand wood colonization by sapstain fungi and their potential biocontrol agents, it is necessary to differentiate these organisms directly on their natural substrates. In the present study the feasibility of transforming with the green fluorescent protein (GFP), the sapstain fungus Ophiostoma piceae and a potential biocontrol agent Cartapip^®, an Ophiostoma piliferum albino strain was assessed. Transformants of the two fungal species were screened by polymerase chain reaction and Southern blot analyses. The GFP was expressed in spores, synnemata and mycelia of the transformants grown in artificial media or wood. The growth, pigmentation and wood colonization of the transformants were similar to that of the non‐transformants, suggesting that the presence of the gfp gene had no negative effect on the biology of the transformants. Using fluorescence and confocal microscopy, the GFP‐expressing fungi were easily differentiated from the wild‐type strains and other fungal species in wood, even 4 months after inoculation. The results show that the use of the GFP system is feasible to monitor Ophiostoma fungi in wood.     

Evaluation of two commercial PRRSV antibody ELISA kits with samples of known status and singleton reactors

Two commercial PRRSV ELISA kits (IDEXX and Bionote) were evaluated for their sensitivity and specificity using 476 PRRS-positive serum samples collected from 7 animal challenge experiments and 1,000 PRRS-negative sera. Both ELISA kits exhibited 100% sensitivity with sera collected 14 to 42 days post-infection, and the results from the kits were highly correlated (R²=0.9207). The specificity of IDEXX or Bionote kit was 99.9% or 99.7%, respectively. In addition, the Bionote ELISA kit was used to examine 100 sera that were determined to be falsely positive either by IDEXX 2XR or 3XR ELISA, and only 7 of these samples were found to be positive. These results indicate that both ELISA kits exhibited similar levels of sensitivity and specificity and would complement one another for the verification of false-positive samples.     

Evaluation of a Collagenase Generated Osteoarthritis Biomarker in Naturally Occurring Canine Cruciate Disease

Objective— To determine the clinical value of a novel osteoarthritis (OA) biomarker in detecting canine cruciate disease.
Study Design— Cross sectional clinical study.
Animals— Dogs (n=22) with cranial cruciate ligament (CCL) rupture and 12 control dogs.
Methods— Concentrations of collagenase-generated cleavage epitope of type II collagen (Col2-3/4C_{long mono}, or C2C) in serum, urine, and joint fluid were compared between a group of dogs with CCL rupture and a control group. Correlation of C2C concentrations to the clinical stage of stifle OA was also evaluated.
Results— There were no significant differences in C2C concentrations in serum, urine, and joint fluid between groups ( P >.05). Subjective scores of lameness, joint effusion, osteophytosis were significantly more severe in the CCL rupture group compared with the control group ( P <.05). There was no significant correlation of C2C concentrations with clinical stage of stifle OA ( P >.05).
Conclusion— This OA biomarker did not detect pathology associated with CCL rupture. Our results suggest that collagenase-specific degradation of type II collagen in articular cartilage may not be involved in the early stage of naturally occurring canine cruciate disease, and that pathology associated with naturally occurring CCL rupture is different from that of experimental OA model.
Clinical Relevance— C2C is not clinically useful in detecting CCL rupture in dogs.     

Effect of Tibial Plateau Leveling Osteotomy on Femorotibial Contact Mechanics and Stifle Kinematics

Objective— To evaluate the effects of tibial plateau leveling osteotomy (TPLO) on femorotibial contact mechanics and 3-dimensional (3D) kinematics in cranial cruciate ligament (CrCL)-deficient stifles of dogs.
Study Design— In vitro biomechanical study.
Animals— Unpaired pelvic limbs from 8 dogs, weighing 28–35 kg.
Methods— Digital pressure sensors placed subjacent to the menisci were used to measure femorotibial contact force, contact area, peak and mean contact pressure, and peak pressure location with the limb under an axial load of 30% body weight and a stifle angle of 135°. Three-dimensional static poses of the stifle were obtained using a Microscribe digitizing arm. Each specimen was tested under normal, CrCL-deficient, and TPLO-treated conditions. Repeated measures analysis of variance with a Tukey post hoc test ( P <.05) was used for statistical comparison.
Results— Significant disturbances to all measured contact mechanical variables were evident after CrCL transection, which corresponded to marked cranial tibial subluxation and increased internal tibial rotation in the CrCL-deficient stifle. No significant differences in 3D femorotibial alignment were observed between normal and TPLO-treated stifles; however, femorotibial contact area remained significantly smaller and peak contact pressures in both medial and lateral stifle compartments were positioned more caudally on the tibial plateau, when compared with normal.
Conclusion— Whereas TPLO eliminates craniocaudal stifle instability during simulated weight bearing, the procedure fails to concurrently restore femorotibial contact mechanics to normal.
Clinical Relevance— Progression of stifle osteoarthritis in dogs treated with TPLO may be partly the result of abnormal stifle contact mechanics induced by altering the orientation of the proximal tibial articulating surface.     

The effect of fermented apple diet supplementation on the growth performance and meat quality in finishing pigs

The objective of the present study was to investigate the effect of fermented apple diet (FAD) supplementation on the growth performance and meat quality in finishing Berkshires. The FAD was made from dropped apple mixed with rice bran and barley bran. Until 81 ± 1 kg live weight at 133 ± 1 days, the animals were fed a growing diet, after which experimental samples were fixed at 0, 2, 4 and 6% FAD as C, T1, T2 and T3 in the finishing diets. Growth performance, ADG, ADFI and feed efficiency were improved in T1 than other groups. In carcass parameters, carcass weight was higher ( P  < 0.05) in T1 than in other groups. In meat quality, moisture and crude protein contents decreased ( P  < 0.05) by addition of FAD. pH₂₄ and WHC were higher ( P  < 0.05) in T1 than other groups. In sensory evaluation, marbling of fresh meat and tenderness, juiciness, flavor and overall acceptability of cooked meat were improved by the addition of FAD. According to the results of our experiment, FAD can be used for improvement of meat quality parameters.     

MAGNETIC RESONANCE IMAGING OF THE CANINE BRAIN AT 7 T

The purpose of this study was to describe relevant canine brain structures as seen on T2-weighted images following magnetic resonance (MR) imaging at 7 T and to compare the results with imaging at 1.5 T. Imaging was performed on five healthy laboratory beagle dogs using 1.5 and 7 T clinical scanners. At 1.5 T, spin echo images were acquired, while gradient echo images were acquired at 3 T. Image quality and conspicuity of anatomic structures were evaluated qualitatively by direct comparison of the images obtained from the two different magnetic fields. The signal-to-nose ratio (SNR) and contrast-to-noise ratio (CNR) were calculated and compared between 1.5 and 7 T. The T2-weighted images at 7 T provided good spatial and contrast resolution for the identification of clinically relevant brain anatomy; these images provided better delineation and conspicuity of the brain stem and cerebellar structures, which were difficult to unequivocally identify at 1.5 T. However, frontal and parietal lobe and the trigeminal nerve were difficult to identify at 7 T due to susceptibility artifact. The SNR and CNR of the images at 7 T were significantly increased up to 318% and 715% compared with the 1.5 T images. If some disadvantages of 7 T imaging, such as susceptibility artifacts, technical difficulties, and high cost, can be improved, 7 T clinical MR imaging could provide a good experimental and diagnostic tool for the evaluation of canine brain disorders.