Comparison of porcine reproductive and respiratory syndrome virus growth in media supplemented with fetal bovine serum or a serum replacement.

Commercially available serum replacements are often used in cell culture as a cheaper and less variable substitute for fetal bovine serum (FBS). The growth of porcine reproductive and respiratory syndrome virus (PRRSV) isolates in CRL11171 cells maintained in a medium supplemented with FBS was compared with virus propagation in the same cell line maintained in the same medium with a serum replacement. The PRRSV replicated significantly better when the cell culture medium was supplemented with FBS. The results of this study have implications for the use of serum replacement-supplemented medium for PRRSV diagnosis by virus isolation.     

Beta-hydroxybutyrate levels in peripheral blood and ketone bodies supplemented in culture media affect the in vitro chemotaxis of bovine leukocytes.

The role of ketone bodies on chemotactic capacities of leukocytes was characterized in two experiments. Experiment I was performed to investigate the association between serum beta-hydroxybutyrate concentrations (BHB) and in vitro chemotaxis of leukocytes. Cows were divided into low-BHB, medium-BHB, and high-BHB ones and classified according to their BHB. Leukocytes from high-BHB cows had a significantly lower chemotactic differential than leukocytes from low-BHB cows (p < 0.01). The effect of adding ketone bodies into in vitro chemotaxis cultures on leukocytes chemotaxis was studied in Experiment II. Either individual or a combination of commercial ketone bodies - sodium salts of BHB (BHBA), lithium salt of acetoacetate (ACAC), and acetone (Acetone) - were diluted in culture media and divided into eight concentrations corresponding to concentrations of bovine subclinical and clinical ketosis. For leukocytes from medium- and high-BHB cow, the chemotactic indexes of leukocytes were reduced by ACAC and Acetone. Chemotactic differentials of cultures with ACAC and acetone supplementation from both sources of leukocytes were significantly lower than that of the control culture (p < 0.05). For leukocytes from high-BHB cows, chemotactic indexes were suppressed in a ketone-body environment. In conclusion, leukocytes from naturally-occurring ketotic cows have lower chemotactic differentials than those from non-ketotic cows, and a chemotactic capacity indicated by a chemotactic differential is impaired when leukocytes migrate in an environment with ketone bodies in vitro.     

Results of endocrinologic studies in culture media after coculture of bovine oocytes with granulosa cells

The steroid hormones progesterone (P.), and testosterone (T.) were radio-immunologically determined in 108 medium samples, following co-culturing of bovine oocytes with granulosa cells. P. and T. values recorded from a control group were lower with significance than those recorded from co-culturing groups, that is 72 +/- 21 ng/ml and 264 +/- 84 pg/ml as compared to 208 +/- 138 ng/ml and 2,168 +/- 1,595 pg/ml in the oocyte plus fresh granulosa cell co-culturing group as well as 364 +/- 215 ng/ml and 825 +/- 233 pg/ml in the oocyte plus pre-incubated granulosa cell co-culturing group. These rises were accompanied by decline in maturation rate, increase in oocyte degeneration, and rises in the rates of fertilisation and segmentation.     

Development of a cell culture line of bovine fetal endometrial cells.

Monolayers of bovine fetal endometrial cells were established as primary culture cells within 1 to 2 weeks. After the 2nd passage, these cells were inoculated with bovine viral diarrhea virus. Effects of the virus were observed each day with a light microscope. Specific cytopathic effects consisting of degeneration and sloughing of the cells and a well-defined pattern of cytoplasmic vacuolation were observed at 5 days after inoculation.     

The use of the direct immunoperoxidase test to detect the multiplication of rinderpest virus in bovine kidney cell culture

The direct immunoperoxidase test has been used to detect rinderpest virus antigens in infected bovine kidney cell cultures to study the multiplication of the virus. Infected bovine kidney coverslip cultures were sequentially tested with peroxidase-labelled, anti-rinderpest globulins at 3, 6, 24, 48, 72, 96, 120 and 144 h post-infection. The progressive virus-specific cytopathic changes compared well with the increase in the number of cells showing the presence of viral antigens when tested by the direct immunoperoxidase test. The specificity of the reaction was confirmed by using negative and antiserum-blocked BK cell cultures on coverslips.     

Growth and metabolism of murine and bovine embryos in bovine uterine flushing-supplemented culture media.

The aim of this study was to compare the development and metabolic activity of cultured murine and bovine embryos in 2 standard media (HAM F-10 and RPMI) in the presence or absence of bovine uterine flushings. Murine morulae (n = 653) and day 7 bovine embryos (n = 273) were cultured for 18 h or 36 h in either HAM F-10 or RPMI in the presence or absence of bovine uterine flushings. After culture, the development, quality, and metabolic activity (glucose utilization or methionine uptake and incorporation) of embryos was assessed. It was found that HAM F-10 (without uterine flushings) was a more suitable medium than RPMI for optimal development and metabolism of murine and bovine embryos. Poor quality and development, as well as decreased metabolism, were evident after culture of murine embryos in RPMI; in contrast, this medium had no adverse effects on bovine embryos in culture. Supplementation of HAM F-10 with bovine uterine flushings improved the growth of murine embryos and the protein synthesis (as measured by an increased methionine incorporation) for both murine and bovine embryos. However, supplementation with bovine uterine flushings could not overcome deficiencies of an inappropriate medium (RPMI) for murine embryos. Supplementation of a well-defined culture medium with uterine flushings increased metabolism of embryos in culture, and thus might help to increase pregnancy rates after transfer of such embryos to recipient cows.     

Susceptibility of cell line cultures of bovine kidney origin to infectious bovine rhinotracheitis and parainfluenza-3 viruses

A comparative study was carried out on the susceptibility of primary bovine embryo kidney (PBEK) cell cultures, and that of AUBEK and MDBK cell lines to infectious bovine rhinotracheitis (IBR) and Parainfluenza-3 (PI-3) viruses.

The cytopathic effects induced by the two viruses were rather inconsistent, based on observations of unstained preparations. On the other hand, there was no significant difference between the susceptibility of the PBEK cultures and the cell line cultures to infection with either virus on the basis of the lesions detected in stained preparations, and of the growth curve patterns.

It is concluded that PBEK cell cultures are more sensitive for isolating IBR or PI-3 viruses than are the AUBEK and MDBK cell lines. However, the latter appear to be satisfactory for studies of these two viruses.     

牛乳腺上皮细胞的分离培养

为了对牛乳腺上皮细胞(MECs)进行分离、培养和鉴定,并研究细胞分泌功能,试验通过胶原酶消化法分离得到了牛乳腺上皮细胞,采用传代法对细胞进行纯化,对细胞标志蛋白进行免疫荧光染色鉴定,通过体外诱导和RT-PCR分析鉴定细胞的分泌功能。结果表明:分离到的牛乳腺上皮细胞具有典型乳腺上皮细胞的形态特征,表达广谱角蛋白,经诱导后可分泌β-酪蛋白。     

Effect of bovine seminal plasma on bovine endometrial epithelial cells in culture

The purpose of this study was to investigate the effect of seminal plasma (SP) from bulls of known fertility on bovine endometrial epithelial cells (bEEC) in culture. The bEEC from passage 5, approximately 5.0–13 × 10⁵ cells per flask, were challenged with SP from bulls of high or low fertility (n = 3 and 2, respectively) or PBS (control), at 1% (75 μl) or 4% (300 μl) and were incubated for 72 hr (n = 13 per challenge). Total cell number and viability of bEEC after challenge with 1% SP from either high‐ or low‐fertility bulls (75H or 75L, respectively) did not differ from controls. In contrast, challenge with 4% of SP from high‐ or low‐fertility bulls (300H or 300L) negatively affected bEEC cell number and viability. Challenge with 300 L had a greater adverse effect than 300H. These results suggest that the negative effect of bovine SP on bEEC is both dose‐dependent and fertility‐dependent.     

产蛋鸡日粮中添加酵母细胞壁对鸡蛋品质及血清蛋白指标的影响

试验选用181日龄海兰灰产蛋鸡576只,随机分为6个处理,设对照基础日粮组,抗生素组(基础日粮+500mg/kg10%杆菌肽锌组),4个酵母细胞壁组,在基础日粮的基础上分别添加500、1000、1500、2000mg/kg酵母细胞壁,每个处理6个重复,试验期56d,以研究酵母细胞壁对鸡蛋品质、血清蛋白指标的影响。结果显示:蛋壳强度、蛋壳厚度、血清中的球蛋白、总蛋白的含量,6个处理组均没有显著性差异。哈夫单位、血清蛋白指标出现差异但没表现出规律性。     

Isolation and monolayer culture of bovine oviduct epithelial cells

Oviduct epithelia obtained from 32 cows were cultured. The oviducts were classified into follicular and luteal phases and divided into ampulla and isthmus regions. The epithelial cells were dissociated by enzyme digestion and cultured in plastic dishes with Dulbecco's modified Eagle's medium/Ham's F12 (1:1) containing 10% calf serum. After enzyme treatment, the epithelial suspension showed free ciliated and non-ciliated cells, and cell mass. The non-ciliated cells contained secretory granules in the cytoplasm. The cell mass was composed of ciliated and secretory cells. The cell mass adhered to the dish within 12-24 hr, while the free ciliated cells attached on Day 2 of the culture. The cells grew into confluent monolayers on Day 4. The cell monolayers contained ciliated and non-ciliated cells. The monolayered non-ciliated cells showed a few secretory granules. When the cells were further cultured without subculturing, ciliary activity diminished on Day 5 and was rarely detected on Day 9. When the cells were subcultured on Day 3, ciliary movement was detected on the monolayers for only 2 days. Cell mass that did not adhere to the dish and remained floating in the medium formed ball-like structures on Day 2. Active ciliary beating was observed on the cells that were cultured in the medium supplemented with 10(-5) and 10(-9) M estradiol-17 beta, however, the ciliary activity diminished on Day 5. No difference in the cell growth was observed between the follicular and luteal phases or between the ampulla and isthmus regions.