Matrix-metalloproteinases-2 and -9 production in bovine endometrial cell culture

In vitro cell culture is a convenient tool for studying cellular mechanisms. In the present study, production of matrix-metalloproteinases (MMPs) in bovine endometrial (containing both epithelial and stromal cells) monolayer cells was examined. Blastocysts attached to the endometrial cells in a monolayer culture were examined for their effects on MMP-2 production. Initial attachment of blastocysts to the monolayer inhibited MMP-2 production by endometrial cells. But once trophoblast cells began to migrate into the endometrial cell layer, MMP-2 production increased, and at the same time MMP-9 production also became evident in the medium. In order to understand how blastocysts affected MMP-2 production, we examined the effect of progesterone, estradiol, insulin-like growth factors (IGFs), tumor necrosis factors (TNFs), and interferon-tau (IFN-tau) supplementation. It was IFN-tau that inhibited the production of MMP-2. In addition, progesterone at a lower dose appeared to inhibit MMP-2 production. Both TNF-alpha and TNF-beta strongly stimulated the production of MMP-2 and MMP-9, whereas IGFs had no effect. Based on these findings, it appears that conceptus has the capacity to inhibit MMP activity.     

Cerebral high-grade astrocytoma (glioblastoma) in a cat

A cerebral tumour was found in the right frontal lobe of a 7-year-old female mongrel cat. The mass showed infiltrative growth and caused deformation of the corpus callosum. Histopathologically, the tumour cells showed anaplasia, pleomorphism and mitotic figures. Necrosis and vascular proliferation were prominent. The neoplastic cells surrounded areas of necrosis, but as an indistinct pseudopalisade formation. Immunohistochemically, low numbers of tumour cells labelled positively for anti-glial fibrillary acidic protein and anti-S100 protein. Electron microscopically, the majority of tumour cells had no filaments and cytoplasmic processes, but the differentiated cells presented cytoplasmic filaments and glycogen granules. Based on these findings, the tumour was diagnosed as cerebral high-grade astrocytoma, glioblastoma.     

Matrix metalloproteinases-2 and -9 activities in bovine follicular fluid of different-sized follicles: relationship to intra-follicular inhibin and steroid concentrations

Matrix metalloproteinases (MMPs) play very important roles in extracellular matrix (ECM) remodeling during ovarian follicular development, ovulation and atresia. The aim of the present study was to determine the content of gelatinases in follicular fluid in various sized bovine follicles. Bovine ovaries were collected from local slaughterhouse and follicular fluid from follicles of 2 to over 25 mm in diameter was collected. Gelatinase activity within the follicular fluid was analyzed by gelatin zymography. The concentration of inhibin in the follicular fluid was also measured by immunoblot analysis. The proMMP-2 and alpha-subunit (alphaN) inhibin was detected in all follicles regardless of their size. The abundance of proMMP-2 varied with follicular size, while alphaN inhibin increased significantly (P<0.01) in follicles of 10-14 and 15-20 mm in size. There was a positive and negative correlation between estradiol (E(2)) and progesterone (P(4)) concentrations with abundance of proMMP-2, respectively. Follicles of diameter over 25 mm had greater proMMP-9 activity than other follicles. These same follicles had significantly (P<0.01) lower inhibin levels than follicles of 10-14 and 15-20 mm in size. In conclusion, these results suggest a significant role of these proteases in growth and development of bovine follicle, particularly proMMP-2 and active MMP-2 activities in the follicular fluid could serve as markers of follicular health while abundance of proMMP-9 may possibly denote a follicular cyst.     

Percutaneous ultrasound-guided over-the-wire catheterization of the portal and hepatic vessels in cattle

A new method for catheterization of the portal and hepatic veins in cattle by means of the over-the-wire system was investigated to maintain more reliable long-term patency of catheters. Four cattle were used to evaluate the success rate, patency and safety of the procedure. The catheters, coated by urokinase were patent as long as they were in situ. In addition, the introducer was useful to prevent the catheter from being broken. No complications developed during the10 days after the procedure. Two cows were then euthanized. Post mortem findings were minimal. The results of the study reported here are promising, the benefits are significant and there is no apparent disadvantage to its use.     

Cell cycle analysis of bovine cultured somatic cells by flow cytometry

This study was undertaken to examine the cell cycle characteristics of bovine fetal and adult somatic cells (fetal fibroblasts, adult skin and muscle cells, and cumulus cells) after culture under a variety of conditions; 1) growth to 60-70% confluency (cycling), 2) serum starvation, 3) culture to confluency. Cell -cycle phases were determined by flow cytometry with propidium iodide staining enabling the calculation of percentages of cells in G0 /G1, S and G2 /M. The majority was in G0/G1 regardless of cell type and treatment. Serum-starved or confluent cultures contained higher percentages of cells in G0/G1 (89.5-95.4%; P < 0.05). Percentages of cells in G0/G1 increased as cell size decreased regardless of the cell type and treatment. In the serum-starved and confluent cultures, about 98% of small cells were in G0/G1 . Serum-starved cultures contained higher percentages of small cells (38.5-66.9%) than cycling and confluent cultures regardless of cell type (P < 0.05) . After trypsinization of fetal fibroblasts and adult skin cells that were serum-starved and cultured to confluency, the percentages of cells in G0/G1 increased (P < 0.05) on incubation for 1.5 (95.7-99.5%) or 3 hr (95.9-98.6%). These results verify that serum starvation and culture to confluency are efficient means of synchronizing bovine somatic cells in G0/G1, and indicate that a more efficient synchronization of the cells in G0/G1 can be established by incubation for a limited time period after trypsinization of serum-starved or confluent cells.     

Hepatic tyrosine aminotransferase activity is affected by chronic heat stress and dietary tyrosine in broiler chickens

1. Two experiments were conducted to investigate the impact of high temperature and dietary tyrosine (Tyr) content on performance and activity of hepatic tyrosine aminotransferase (EC 2.6.1.5.), an enzyme that catalyses the first step in the metabolic degradation of Tyr in broiler chickens. 2. Two-week-old birds were allocated to one of three temperature treatments: 24 degrees C (control), 36 degrees C (heat stress, HS) and 24 degrees C pair-fed (24PF) for 2 weeks and fed on diets containing 100% (Experiment 1) and 50, 100 and 200% (Experiment 2) of the NRC requirement for Tyr. 3. In Experiment 1, exposure of chickens to 36 degrees C for 2 weeks caused significant increase in hepatic tyrosine aminotransferase activity but no significant change in activity of hepatic phenylalanine 4-hydroxylase (EC 1.14.16.1) (an enzyme that catalyses conversion of phenylalanine to Tyr) compared with the 24PF birds. No significant changes attributable to heat stress were detected in hepatic glutamic-oxaloacetic transaminase (EC 2.6.1.1) activity. 4. In Experiment 2, heat stress caused reductions in weight gain and feed intake in chickens on all diets, compared with their control counterparts. Hepatic tyrosine aminotransferase activity was increased by heat stress compared with their 24PF counterparts in chickens fed on the 100 and 200% Tyr diets, while in chickens fed the 50% Tyr diet, it was reduced by heat stress. 5. From these results, it is suggested that hepatic tyrosine aminotransferase activity is affected by heat stress and dietary Tyr content and the increased tyrosine aminotransferase activity with, in part, relatively low phenylalanine hydroxylase activity in hepatic tissues may be involved in the Tyr metabolism characteristic of heat-stressed chickens.