Comparison of arylalkylamine N-acetyltransferase and melatonin receptor type 1B immunoreactivity between young adult and aged canine spinal cord

Melatonin affects diverse physiological functions through its receptor and plays an important role in the central nervous system. In the present study, we compared immunoreactivity patterns of arylalkylamine N-acetyltransferase (AANAT), an enzyme essential for melatonin synthesis, and melatonin receptor type 1B (MT2) in the spinal cord of young adult (2~3 years) and aged (10~12 years) beagle dogs using immunohistochemistry and Western blotting. AANAT-specific immunoreactivity was observed in the nuclei of spinal neurons, and was significantly increased in aged dog spinal neurons compared to young adult spinal neurons. MT2-specific immunoreactivity was found in the cytoplasm of spinal neurons, and was predominantly increased in the margin of the neuron cytoplasm in aged spinal cord compared to that in the young adult dogs. These increased levels of AANAT and MT2 immunoreactivity in aged spinal cord might be a feature of normal aging and associated with a feedback mechanism that compensates for decreased production of melatonin during aging.     

Selective growth-inhibiting effects of compounds identified in Tabebuia impetiginosa inner bark on human intestinal bacteria

The growth-inhibiting activity of anthraquinone-2-carboxylic acid and lapachol identified in the inner bark of taheebo, Tabebuia impetiginosa, toward 10 human intestinal bacteria was evaluated by using a paper disk diffusion bioassay and compared to those of seven lapachol congeners (1,4-naphthoquinone, naphthazarin, menadione, lawsone, plumbagin, juglone, and dichlone) as well as two commercially available antibiotics, chloramphenicol and tetracycline. Anthraquinone-2-carboxylic acid exhibited very strong growth inhibition of Clostridium paraputrificum at 1 microg/disk while 100 microg/disk of lapachol was needed for moderate growth inhibition of the same organism. These two isolates exhibited weak inhibition of Clostridium perfringens and Escherichia coli at 100 microg/disk while no adverse effects were observed on the growth of Bifidobacterium adolescentis, Bifidobacterium bifidum, Bifidobacterium infantis, Lactobacillus acidophilus, and Lactobacillus casei at 1000 microg/disk. Structure-activity relationships indicate that a methyl group in the C-2 position of 1,4-naphthoquinone derivatives might play an important role in antibacterial activity.     

2-fluoroabscisic acid analogues: their synthesis and biological activities

Fluorine was introduced into the 2-position of the side chain of abscisic acid (ABA) analogues by Wittig reaction of alpha-ionone derivatives with ethyl triethylphosphono-2-fluoroacetate. The effects of the fluorinated analogues were evaluated on inhibition of cress seed germination and inhibition of gibberellin-inducible alpha-amylase induction in embryoless barley half-seeds. (2E, 4E)-2-Fluoro-5-(1'-hydroxy-2',6', 6'-trimethyl-2'-cyclohexen-1'-yl)-3-methyl-2,4-pentadienoic acid (5b) showed potent inhibitory activity at the same level as ABA in the cress seed germination test, and 5b also inhibited gibberellin-inducible alpha-amylase induction at 4 x 10(-)(6), 3 times the concentration of ABA (1 x 10(-)(6)) for 50% inhibition of alpha-amylase production. 5b also showed dehydrin induction activity. These results indicate that fluorinated ABA analogues mimic ABA action and can be a lead for a plant growth regulator which regulates plant growth or protects plants from environmental stresses.     

Genetic analysis of ORF5 of recent Korean porcine reproductive and respiratory syndrome viruses (PRRSVs) in viremic sera collected from MLV-vaccinating or non-vaccinating farms

The 23 open reading frame (ORF) 5 sequences of Korean type II porcine reproductive and respiratory syndrome virus (PRRSV) were collected from viremic sera from the (modified live vaccine) MLV-vaccinating and non-vaccinating farms from 2007 to 2008. The samples were phylogenetically analyzed with previous ORF5 sequences, including type I Korean PRRSV, and previously reported or collected sequences from 1997 to 2008. A MN184-like subgroup of type II Korean PRRSV was newly identified in the viremic sera collected from 2007 to 2008. And of the type I PRRSVs, one subgroup had 87.2~88.9% similarity with the Lelystad virus, showing a close relationship with the 27~2003 strain of Spain. The maximum parsimony tree of type II PRRSV from 1997 to 2008 showed that they had evolved to four lineages, subgroups 1, 2, 3 and 4. Most of the recently collected type II PRRSVs belonged to subgroup 4 (48%). The region of three B-cell epitopes and two T-cell epitopes of ORF5 amino acids sequences was considerably different from the MLV in subgroups 3 and 4. In conclusion, the existence of type I PRRSV, which was genetically different from Lelystad virus (Prototype of type I PRRSV), and heterologous type II PRRSVs of viremic pigs detected even in the MLV-vaccinating farms indicated the need for new vaccine approaches for the control of PRRSV in Korea.     

Evaluation of adverse reactions in dogs following intravenous mesenchymal stem cell transplantation

Background

Recent studies have assessed the therapeutic potential and drawbacks of mesenchymal stem cells (MSCs). The adverse reactions of intravenous transplantation of bone marrow (BM)-derived MSCs were examined at varying doses and frequencies of administration.Nine healthy beagle dogs were purchased from a commercial laboratory. The dogs were distributed equally (n = 3 per group) and randomly into three groups. All dogs received allogeneic BM-derived MSCs: 2 × 10⁶ once (group A), 2 × 10⁷ once (group B), and 2 × 10⁶ for three consecutive days (group C). Various laboratory examinations, multi-detector computed tomography features and histopathology were evaluated to clarify the clinical and diagnostic features of adverse reactions of MSCs administration, prior to receiving MSCs (pre procedure) and on days 1, 3, and 7 post transplantation.

Results

Only one dog had clinical signs during and after MSCs transplantation. Dogs receiving 2 × 10⁶ MSCs showed increased numbers of lymphocytes but the total white blood cell counts were not elevated (P < 0.01). Multi-detector computed tomography (MDCT) revealed pulmonary parenchymal changes in one dog and histopathologic examination revealed pulmonary parenchymal edema and hemorrhage in four dogs. The presence of pulmonary thromboembolism was not detected in either examination.

Conclusions

We considered the presence of pulmonary edema and hemorrhage as possible adverse reactions after intravenous MSCs transplantation; however these results should be cautiously interpreted.     

The effects of enrofloxacin on canine tendon cells and chondrocytes proliferation <Emphasis Type="Italic">in vitro</Emphasis>

Enrofloxacin, a fluoroquinolone antibiotic has been used widely in humans and domestic animals, including dogs, because of its broad-spectrum activity and relative safety. The side effects of fluoroquinolone, induced tendinopathy, tendonitis, spontaneous tendon rupture and cartilage damage, remain incompletely understood. In the present study, we investigated the in vitro effects of enrofloxacin on cell proliferation and induction of apoptosis in canine Achilles tendon cells and chondrocytes. Cell growth and proliferation after treating with enrofloxacin for 2–6 days was quantified by a colorimetric 2,3-bis{2-methoxy-4-nitro-5-sulfophenyl}-2H-tetrazolium-5-carboxyanilide inner salt (XTT) assay. The results showed that enrofloxacin could inhibit the proliferation of canine tendon cells and chondrocytes at increasing concentrations (10–200 μg/ml). The inhibition of proliferation of canine tendon cells and chondrocytes after exposure to enrofloxacin were associated with induction of apoptosis, as evidenced by the typical nuclear apoptotic condensed nuclei found using Hoechst 33258 staining. It was demonstrated that canine tendon cells and chondrocytes treated with 200 μg/ml enrofloxacin for 4 days exhibited apoptotic features and fragmentation of DNA. Enrofloxacin also increased the apoptosis of canine tendon cells and chondrocytes in a dose and time-dependent manner. The results indicate that enrofloxacin inhibits cell proliferation, induces apoptosis and DNA fragmentation, which might explain enrofloxacin-induced tendinopathy and cartilage damage.     

RNA干扰技术和家蚕杆状病毒表达系统在纤维素酶研究中的应用前景

植物通过光合作用产生的纤维素是地球上最丰富、最廉价的可再生资源。纤维素酶是糖苷水解酶的一种,能有效地将农作物秸杆等富含纤维素的物质水解为葡萄糖,进而发酵产生生物乙醇,从而解决农业、再生能源以及环境污染等问题。随着生物化学、分子生物学以及基因工程等多种交叉学科的快速发展,获得适合工业化的高活力的纤维素酶指日可待。RNA干扰是利用双链RNA特异性地降解相应序列的mRNA,从而特异性地阻断相应基因的表达,是后基因组时代的一种强有力的调控目的基因表达的实验工具。家蚕杆状病毒表达系统是一种快速、高效表达外源基因的技术手段,目前该系统的发展和应用已经非常成熟。本文综述了纤维素酶的特性以及近年来国内外对纤维素酶、RNA干扰和家蚕杆状病毒表达系统的研究进展,并对该两种实验技术手段在今后纤维素酶研究中的应用前景作了预测和展望。     

Isolation and characterization of antimicrobial-resistant Escherichia coli from national horse racetracks and private horse-riding courses in Korea

Limited information is available regarding horse-associated antimicrobial resistant (AR) Escherichia (E.) coli. This study was designed to evaluate the frequency and characterize the pattern of AR E. coli from healthy horse-associated samples. A total of 143 E. coli (4.6%) were isolated from 3,078 samples collected from three national racetracks and 14 private horse-riding courses in Korea. Thirty of the E. coli isolates (21%) showed antimicrobial resistance to at least one antimicrobial agent, and four of the AR E. coli (13.3%) were defined as multi-drug resistance. Most of the AR E. coli harbored AR genes corresponding to their antimicrobial resistance phenotypes. Four of the AR E. coli carried class 1 integrase gene (intI1), a gene associated with multi-drug resistance. Pulsed-field gel electrophoretic analysis showed no genetic relatedness among AR E. coli isolated from different facilities; however, cross-transmissions between horses or horses and environments were detected in two facilities. Although cross-transmission of AR E. coli in horses and their environments was generally low, our study suggests a risk of transmission of AR bacteria between horses and humans. Further studies are needed to evaluate the risk of possible transmission of horse-associated AR bacteria to human communities through horse riders and horse-care workers.     

Antioxidative Effects of Astaxanthin against Nitric Oxide‐Induced Oxidative Stress on Cell Viability and Gene Expression in Bovine Oviduct Epithelial Cell and the Developmental Competence of Bovine IVM/IVF Embryos

The aim of the present study was to elucidate the fundamental mechanism of bovine oviduct epithelial cell (BOEC) co‐culture on developmental capacity of bovine in vitro oocyte maturation/in vitro fertilization (IVM/IVF) embryos. We examined the effects of astaxanthin against nitric oxide‐induced oxidative stress on cell viability by MTT assay, lipid peroxidation (LPO) by using thiobarbituric acid (TBA) reaction for malondialdehyde (MDA) and the expression of antioxidant genes (CuZnSOD, MnSOD and Catalase) or apoptosis genes (Bcl‐2, Caspase‐3 and Bax) by RT‐PCR in BOEC. We also evaluated the developmental rates of bovine IVM/IVF embryos co‐cultured with BOEC pre‐treated with astaxanthin (500 μm ) in the presence or absence of sodium nitroprusside (SNP, 1000 μm ) for 24 h. Cell viability in BOEC treated with SNP (50–2000 μm ) lowered, while astaxanthin addition (50–500 μm ) increased it in a dose‐dependent manner. Cell viability in astaxanthin plus SNP (1000 μm ) gradually recovered according to the increase in astaxanthin additions (100–500 mm ). The LPO in astaxanthin group (50–500 μM) gradually decreased in a dose dependent manner and among SNP or astaxanthin plus SNP group, SNP alone and astaxanthin (50 μM) plus SNP shown a significant increase than other groups (p < 0.05). Expression of apoptosis or antioxidant genes was detected by RT‐PCR. Bcl‐2 and antioxidant genes were detected in astaxanthin or astaxanthin plus SNP group, and Caspase‐3 and Bax genes were only found in SNP group. When bovine IVM/IVF embryos were cultured for 6–7 days under co‐culture system such as BOEC treated with astaxanthin in the presence or absence of SNP, the developmental ability to blastocysts in 500 μm astaxanthin group was the highest of all groups. These results suggest that astaxanthin has a antioxidative effect on cell viability and LPO of BOEC, and development of bovine IVM/IVF embryos due to the induction of antioxidant genes and suppression of apoptosis genes.     

Ultrastructural study on scab resistance expressed in epidermal pectin layers of pear leaves

Proliferation and collapse of subcuticular hyphae of Venturia nashicola race 1 were studied ultrastructurally, after inoculation of susceptible Japanese pear cv. Kousui, resistant Japanese pear cv. Kinchaku, resistant Asian pear strain Mamenashi 12 and nonhost European pear cv. Flemish Beauty leaves, to understand the nature of the resistance mechanism. After cuticle penetration by the pathogen, the hyphae were observed at lower frequency in epidermal pectin layers and middle lamellae of leaves of the three resistant plants than in those of susceptible ones. This result suggested that fungal growth was suppressed in the incompatible interaction between pear and V. nashicola race 1. In the pectin layers of all inoculated plants, some hyphae had modifications such as breaks in the plasmalemma with plasmolysis, necrotic cytoplasm and degraded cell walls. More hyphae had collapsed in the leaves of the three resistant plants than in those of the susceptible cv. Kousui. In collapsed hyphae, the polymerized cell walls broke into numerous fibrous and amorphous pieces, showing that the scab resistance might be associated with cell wall-degrading enzymes from pear plants.