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.     

Progesterone receptor mRNA levels during pregnancy, labor, lactation and the estrous cycle in rat uterus

Progesterone plays important roles in the regulation of female reproduction. In this study, progesterone receptor (PR) mRNA levels in rat uterus during pregnancy, labor, lactation and the estrous cycle were examined by competitive RT-PCR. During pregnancy and lactation, PR mRNA levels had decreased on day 20 of pregnancy (P20) and P21 compared with P15 but increased during labor. After a decline on day 1 of lactation (L1), PR mRNA levels had increased again on L3 and L14 compared with P15, P18, P20, P21 and P21pm (at 2200-2300 h on P21). There was no significant change in the PR mRNA level during the estrous cycle. The PR mRNA level did not change during 1 week of progesterone treatment or afterwards. Injection of 17beta-estradiol did not affect PR mRNA levels in rats treated with progesterone or those without any injections. In rats on P18, 17beta-estradiol injection did not change PR mRNA levels after sham-operation but induced an increase in PR mRNA levels of rats ovariectomized 6 h before the treatment. These results suggest that uterine PR mRNA levels are differently regulated during late pregnancy, labor and lactation, and during labor estrogen is one of the essential factors for the increase in PR mRNA levels.     

Molecular mechanisms underlying pig oocyte maturation and fertilization

Since the pig is not only an important farm animal, but also a model animal for biomedical applications, the development of reproductive technologies in this species has been very important. In vitro oocyte maturation and fertilization (IVM-IVF) are basic techniques for a number of oocyte- or embryo-related technologies. The practical aspects for pig oocyte IVM-IVF have been reviewed, while the molecular mechanisms underlying oocyte meiotic maturation and fertilization have not been well summarized, although accumulating data have been obtained in recent one decade. This review will focus on what is known about the molecular mechanisms of porcine oocyte maturation and fertilization such as first meiosis resumption, meiotic spindle assembly, second meiosis metaphase (MII) arrest during oocyte maturation, sperm-egg recognition and fusion, sperm acrosome reaction, second meiosis resumption, sperm chromatin decondensation, and pronucleus formation during fertilization, as well as the establishment of polyspermy block.     

PCR detection of Porcine circovirus type 2 DNA in whole blood,serum, oropharyngeal swab,nasal swab,and feces from experimentally infected pigs and field cases

Porcine circovirus type 2 (PCV2) shedding patterns were investigated by polymerase chain reaction (PCR) for the detection of PCV2 DNA, and the diagnostic suitability of a sample for the PCR was examined by using different types of samples. In the experimental infection, sixteen pigs were inoculated intranasally with PCV2. The samples, including oropharyngeal and nasal swabs, feces, whole blood and serum became positive for PCV2 DNA by PCR immediately after the inoculation, and almost all samples remained positive during the observation period, post-inoculation-day 70. Field samples were collected from 313 pigs in five different age groups. The overall percentages of positive samples in the whole blood, nasal swabs, and feces detected by PCR were 30.4%, 19.2%, and 20.4%, respectively. The frequency of positive samples increased after the nursery stages and reached a peak in the 3 to 4-month-old pigs. These results indicate that PCV2 infection may occur after weaning, that PCV2 DNA may be present in whole blood for a long period after infection, and that whole blood and serum are the most suitable sample types for the PCR analysis of PCV2.     

A comparative pathological study on canine necrotizing meningoencephalitis and granulomatous meningoencephalomyelitis

Canine necrotizing meningoencephalitis (NME) and granulomatous meningoencephalomyelitis (GME) were compared pathologically. Gross observation exhibited lateral ventricular dilation and discoloration, malacia and/or cavitation of the cerebrum in NME. On the contrary, gross changes were milder in GME, except for occasional visible granulomatous mass formation. Histopathologically, the lesions of NME were distributed predominantly in the cerebral cortex and various degrees of inflammatory and necrotic changes were observed according to clinical stages. Besides, microscopic lesions of GME were mainly distributed in the white matter of the cerebrum, cerebellum and brainstem, which are characterized by perivascular cuffing, multiple granulomas and leptomeningeal infiltrates. Although macrophages and lymphocytes were predominant in the inflammatory lesions of both disorders, macrophages in GME transformed into epithelioid cells and exhibited more massive infiltration. Although lectin RCA-1-reactive cells were numerous in both disorders, lysozyme immunoreactive cells in NME were fewer than that in GME. Parenchymal infiltration of MAC387-positive cells was common in GME and limited in NME. The number of CD3-positive lymphocytes in the GME lesions tended to be greater than in NME, though the difference was not statistically significant. Morphological and immunohistochemical differences of the lesions, in particular, the characteristics of infiltrative macrophages may reflect these different pathogeneses of the two disorders.     

Changes of receptor mRNAs for oxytocin and estrogen during the estrous cycle in rat uterus

Oxytocin receptor (OTR) mRNA levels increase dramatically near term and is potently stimulated by estrogen because increased OTR mRNA levels result from estrogen treatment in ovariectomized rat uterus. In this study, OTR, estrogen receptor (ER) alpha and ERbeta mRNA levels in the rat uterus during the estrous cycle were examined by quantitative RT-PCR. OTR mRNA levels during the estrous cycle began to increase on diestrus (P<0.05, vs value on estrus), reached maximal increase both in the morning (1000-1130 hr) and afternoon (1600-1630 hr) on proestrus (P<0.01, vs metestrus, diestrus and estrus) and then declined on estrus. In contrast ER alpha mRNA levels began to decrease on diestrus, reached statistical significance both in the morning and the afternoon on proestrus (P<0.01, vs metestrus, diestrus and estrus) and returned to the value of metestrus on estrus. ERbeta mRNA levels were low in the morning and the afternoon on proestrus (P<0.01, vs metestrus and estrus) and also returned to metestrus values on estrus. Treatments with estrogen for 3 days significantly decreased both ERalpha and ERbeta mRNA levels. It can be concluded from these results that during the estrous cycle, OTR mRNA levels in rat uterus predominantly increase at proestrus with a decrease in ERalpha and ERbeta mRNA levels, which is probably due to the increased estrogen levels in circulation before ovulation.     

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.     

Langerhans cells within the follicular epithelium and the intradermal sweat duct in equine insect hypersensitivity "Kasen"

Histopathologic and electron microscopic observations were given on Langerhans cells (LCs) within the follicular epithelium (FE) and intradermal sweat duct (ISD) of equine "Kasen". By light microscopy, LCs were present in the greatest numbers within the FE and ISD than within the epidermal layer and the normal skin, with an occasional formation of several aggregated foci. By electron microscopy, LCs within the FE and ISD widely extended their dendritic processes between the keratinocytes and contained Birbeck granules (Bgs), mitochondria, rough endoplasmic reticula and ribosomes in the cytoplasm. Numerous Type 2 LCs, with a number of Bgs and endocytosis, and Type 3 LCs, with multivesicular bodies and endosomes of various sizes, were recognized within the FE and ISD, although inactive Type 1 LCs, with a narrow and lucid cytoplasm, were rarely seen. LCs observed within the FE and ISD in the "Kasen" skin lesions might express the particular stage corresponded to recognize, intake and process the antigens which permeate them.     

Thyroid hormone augments GLUT4 expression and insulin-sensitive glucose transport system in differentiating rat brown adipocytes in culture

The effects of triiodothyronine (T3) on differentiation-dependent expression of GLUT and responses of glucose transport to insulin and norepinephrine (NE) were investigated. Precursor cells of brown adipocytes isolated from the interscapular brown adipose tissue of newborn rats were cultured in the absence or presence of various concentrations of T3. Western bolt analysis revealed that treatment with T3 resulted in an increased expression of GLUT4, in a dose-dependent manner, whereas GLUT1 contents were unchanged. In parallel with the increase in GLUT4 expression, T3 improved insulin sensitivity for glucose transport, being accompanied by an increase in maximal transport rate and a reduction of ED(50). In contrast, T3-treatment of the brown adipocytes during the differentiation process had little effect on NE-regulatable glucose transport system. These results suggest that T3 plays a predominant role in the development of insulin-sensitive glucose transport during differentiation of brown adipocytes.