Role of CXCR4 and SDF-1 in mammary tumor metastasis in the cat

It has recently been suggested that the chemokine receptor CXCR4 and its ligand SDF-1 (CXCL12) promote metastasis of various cancers in humans. Since feline mammary tumors also metastasize to distant organs frequently, we used real-time quantitative PCR to examine the expression of feline CXCR4 (fCXCR4) in ten feline mammary tumor cell lines and seven feline mammary tumor tissues, and also the expression of feline SDF-1 (fSDF-1) in various organs. Cell lines derived from metastatic regions expressed more fCXCR4 than those derived from primary tumors. Mammary tumor tissues overexpressed more fCXCR4 than normal mammary tissues. Organs with high levels of fSDF-1 expression represent common sites of metastasis. Migration assays using the feline mammary tumor cell line NAC were also performed to test the activity of TN14003 and TC14012, antagonists of human CXCR4, to antagonize fCXCR4 expressed on NAC cells. TN14003 and TC14012 inhibited migration of NAC cells. We conclude that fCXCR4 may be a therapeutic target for feline mammary tumors.     

不同土壤水分条件下草莓果实在模拟贮运中的损伤易感性

通过模拟运输振动实验研究了种植在不同土壤水分条件下草莓果实在贮运中的损伤易感性。试验结果表明,栽培在较高土壤水分条件下的果实含有较高的水分,贮运中果实损伤易感性与果实含水量、果实鲜重、土壤湿度呈显著或极显著相关。模拟振动实验发现:在高土壤水分条件下栽培的草莓果实在振动前、振动后及贮藏后的呼吸速率分别为生长在低土壤水分果实的124.9%、119.0%和133.2%。振动的果实比未振动的果实降低了28%-36%的蔗糖,增加了4.5%-9.8%的果糖和7.1%-10.1%葡萄糖,说明运输振动促进了蔗糖降解为葡萄糖和果糖的水解过程。     

Investigation of serum concentrations and immunohistochemical localization of α1-acid glycoprotein in tumor dogs

The purpose of this study was to measure the dynamics of serum α1-acid glycoprotein (AGP) concentration in dogs with various tumors, and to investigate the localization of AGP in some tissues using immunohistochemical staining. Sera were obtained from 171 dogs bearing tumors of various types. Serum AGP concentration was measured by single radial immunodiffusion. Tumors occurring in the liver and spleen were also investigated immunohistochemically using anti-canine AGP antibody. Mean serum AGP levels were 749 ± 602 mg/L in dogs with carcinoma (n = 39), 1,014 ± 971 mg/L with sarcoma (n = 18), and 887 ± 935 mg/L with round cell tumors (n = 46), all significantly higher than serum AGP level in healthy dogs (n = 137, 364 ± 106 mg/L). Mean serum AGP levels were significantly higher than in healthy dogs in complex mammary gland carcinoma (n = 5, 876 ± 721 mg/L), malignant melanoma (n = 7, 1,010 ± 821 mg/L), and hepatocellular carcinoma (n = 5, 936 ± 741 mg/L) among carcinomas, hemangiosarcoma (n = 5, 1,740 ± 1,323 mg/L) among sarcomas, and lymphoma (n = 19, 1,072 ± 965 mg/L) and histiocytic tumor (n = 6, 1,800 ± 1,387 mg/L) among round cell tumors. In an immunohistochemical investigation of AGP localization, both weak and strong staining for anti-AGP antibody were seen in hepatic tissue in dogs with primary non-tumorous lesions originating in the spleen (hematoma) and elevated serum AGP, but all tumor tissue in the spleen was negative. Among dogs with primary tumor lesions of the spleen (hemangiosarcoma) and elevated serum AGP levels, both weak, moderate and strong staining for anti-AGP antibody were seen in hepatic tissue, while strong positive staining was apparent in all tumorous tissue from the spleen. In primary tumor lesions in the liver (hepatocellular carcinoma), both moderate and strong staining for anti-AGP antibody were seen in normal hepatic tissue, and both weak, moderate and strong staining were seen in tumor tissues of the liver. AGP levels thus appear to be elevated in dogs with carcinomas, sarcomas, and round cell tumors. With some of these malignant tumors, localization of AGP in tumor tissue was seen.     

Prognosis of canine patients with nasal tumors according to modified clinical stages based on computed tomography: a retrospective study

To evaluate the efficacy of clinical staging based on computed tomography (CT) imaging over the World Health Organization (WHO) staging system based on radiography for nasal tumors in dogs, a retrospective study was conducted. This study used 112 dogs that had nasal tumors; they had undergone radiography and CT and had been histologically confirmed as having nasal tumors. Among 112 dogs, 85 (75.9%) were diagnosed as adenocarcinoma. Then they were analyzed for survival time according to each staging system. More than 70% of the patients with adenocarcinoma were classified as having WHO stage III. The patients classified under WHO stage II tended to survive longer than those classified under WHO stage III. Dogs classified under WHO stage III were further grouped into CT stages III and IV, and CT stage III patients had a significantly longer survival time than CT stage IV patients. In addition, patients treated with a combination of surgery and radiation had a significantly longer survival time than the patients who did not receive any treatment in CT stage III. On the other hand, different treatment modalities did not show a significant difference in the survival time of CT stage IV dogs. The results suggest that WHO stage III dogs may have various levels of tumor progression, indicating that the CT staging system may be more accurate than the WHO staging system.     

Establishment of a LacZ marker rescue assay to detect infectious RD114 virus

Cats have an infectious endogenous retrovirus, named RD114 virus, and there is a possibility that RD114 virus has contaminated live attenuated vaccines, for which feline cells are used as a substrate. To monitor infectious RD114 virus in vaccines for cats, we developed a LacZ marker rescue assay to detect infectious RD114 virus. Among four human cell lines examined, TE671 cells (human rhabdomyosarcoma) were most susceptible to RD114 virus and supported RD114 replication efficiently. Infection was enhanced approximately 5 times by the addition of polybrene at concentrations of 2 to 8 microg/ml in the medium during viral adsorption. A 4-hr viral adsorption period was sufficient to obtain the maximum titer. By inoculating samples into TE671 cells transduced with the lacZ marker gene, the limiting diluted sample (i.e., less than 10 infectious units) was detected at 12 days post-inoculation by the LacZ marker rescue assay. Based on the results obtained in this study, we propose a standard protocol of the LacZ marker rescue assay to detect infectious RD114 virus.     

Role of beta1 integrins in adhesion of canine mastocytoma cells to extracellular matrix proteins

The interactions of tumor cells with the extracellular matrix (ECM) are a crucial step in invasion and metastasis. Integrins are adhesive molecules forming heterodimers with alpha and beta subunits that play a definitive role in these interactions. In this study, mastocytoma (mast cell tumor: MCT) cell-ECM interaction was investigated using 3 canine MCT cell lines: CM-MC (originating from cutaneous MCT), VI-MC (originating from intestinal MCT), and CoMS (originating from oral MCT). Flow cytometric analysis showed that all cells highly expressed the integrin beta1 and alpha1 through alpha5 subunits and that they moderately expressed the alpha6 subunit. In adhesion studies, CoMS weakly but spontaneously adhered to fibronectin (FN), which was enhanced by phorbol ester (TPA), while CM-MC and VI-MC required cell activation by TPA to adhere to FN. Anti-beta1 and alpha5 integrin antibodies strongly inhibited cell adhesion to FN in CM-MC and CoMS and moderately inhibited cell adhesion in VI-MC. Only VI-MC adhered to laminin (LN) under activation by TPA. Anti-beta1 integrin antibodies strongly inhibited cell adhesion to LN, but all anti-alpha integrin antibodies failed to inhibit cell adhesion to LN. No cells adhered to collagen types I and IV. Canine MCT cells from different origins expressed similar integrin patterns; however, there were some differences in adhesive behavior in response to various ECM proteins and activating stimuli.     

Identification of tumor-initiating cells in a canine hepatocellular carcinoma cell line

Tumor-initiating cells (TICs) or cancer stem cells (CSCs), a small subset of tumor cells, are involved in tumor initiation, progression, recurrence and metastasis. In human hepatocellular carcinoma (HCC), TICs are enriched with cell surface markers and have the ability to self-renew and differentiate tumors at a high frequency. We established a canine HCC cell line, HCC930599, and analyzed it for stem and progenitor cell marker expression using flow cytometry. HCC930599 showed high CD44 and CD29, moderate CD90, and low CD133, CD34, CD24, CD117, and CD13 expression. CD90⁺CD44⁺ and CD90⁻CD44⁺ cells were characterized using the in vitro sphere assay and an in vivo transplant model. CD90⁺CD44⁺ cells acquired enhanced self-renewal capacity, proliferative activity and tumourigenicity compared with CD90⁻CD44⁺ cells, suggesting that TICs exist in the HCC930599 cell line and that CD90 is a marker for enriched TICs. Understanding TIC characteristics may help elucidate hepatic carcinogenesis and HCC therapy development.     

Erwinia isolates from the bacterial shoot blight of pear in Japan are closely related to Erwinia pyrifoliae based on phylogenetic analyses of gyrB and rpoD genes

The phylogenetic relationships among Erwinia amylovora biovar 4 (the pathogen of bacterial shoot blight of pear in Japan), other biovars of E. amylovora, and Erwinia pyrifoliae were investigated using the sequences of 16S rRNA, gyrB, and rpoD genes. The tested isolates formed two distinct monophyletic groups in the phylogenetic trees constructed based on the gyrB gene, rpoD gene, or a combination of the three genes: group 1 contained E. amylovora biovars 1, 2, and 3; group 2 contained E. amylovora bv. 4 and E. pyrifoliae. This phylogenetic analysis showed that E. amylovora bv. 4 was more closely related to E. pyrifoliae than to other biovars of E. amylovora. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession numbers AB242876 to AB242925.     

Neutralizing antibodies against feline parvoviruses in nondomestic felids inoculated with commercial inactivated polyvalent vaccines

The virus neutralization (VN) antibody titers of serum samples from 18 individuals representing 8 carnivore species vaccinated with commercial polyvalent vaccines optimized for domestic cats containing inactivated feline panleukopenia virus (FPLV) were evaluated against canine parvovirus type 2 (CPV2). In addition, the titers among 5 individuals from 4 carnivore were evaluated against antigenic variants of feline parvoviruses; FPLV, CPV2, CPV2a, CPV2b, CPV2c, mink enteritis virus type 1 (MEV1) and MEV2. The polyvalent vaccines induced cross-reactive VN titers against antigenic variants of feline parvoviruses in nondomestic felids. However, we observed very low cross-reactive VN antibody in lions and Siberian tigers, therefore we should pay attention to CPV infections in these animals even if they were vaccinated with inactivated FPLV vaccines.