Establishment of a patient-derived xenograft mouse model of pleomorphic leiomyosarcoma

Soft tissue sarcomas are difficult to treat using chemotherapy owing to a current deficiency in candidate drugs for specific targets. Screening candidate compounds and analyzing therapeutic targets in sarcomas is insufficient, given the lack of an appropriate human sarcoma animal model to accurately evaluate their efficacy, as well as the lack of an adequate technical protocol for efficient transplantation and engraftment of sarcoma specimens in patient-derived xenograft (PDX) models. Accordingly, in this study, we sought to identify the optimal type of sarcoma and develop a protocol for generating a PDX model. We characterized a PDX mouse model using histopathological and immunohistochemical analyses to determine whether it would show pathological characteristics similar to those of human sarcomas. We achieved engraftment of one of the 10 transplanted sarcoma specimens, the xenografted tumor of which exhibited massive proliferation. Histologically, the engrafted sarcoma foci resembled a primary tumor of pleomorphic leiomyosarcoma and maintained their histological structure in all passages. Moreover, immunohistochemical analysis revealed the expression of specific markers of differentiation to smooth muscle, which is consistent with the features of leiomyosarcoma. We thus demonstrated that our pleomorphic leiomyosarcoma PDX mouse model mimics at least one aspect of human sarcomas, and we believe that this model will facilitate the development of novel therapies for sarcomas.     

Retrospective study of canine nasal tumor treated with hypofractionated radiotherapy

The object of this study was to evaluate hypofractionated multiportal field and two-portion (rostral and caudal portions divided by the eyelid) radiation therapy for canine nasal tumors. Sixty-three dogs underwent multiportal hypofractionated radiation therapy. The radiation field was divided into rostral and caudal portions by the eyelid. Treatments were performed four times for 57 dogs. The median irradiation dose/fraction was 8 Gy (range, 5-10 Gy); the median total dose was 32 Gy (10-40 Gy). Improvement of clinical symptoms was achieved in 53 (84.1%) of 63 cases. Median survival time was 197 days (range, 2-1,080 days). Median survival times with and without destruction of the cribriform plate before radiotherapy were 163 and 219 days, respectively. There was no significant difference between them. No other factors were related to survival according to a univariate analysis. All radiation side effects, except one, were grade I according to the VRTOG classification. It was not necessary to treat any dogs for skin side effects. One dog (1.6%) developed an oronasal fistula 1 year after completion of radiation therapy. This radiation protocol may be useful in reducing radiation side effects in dogs with cribriform plate destruction.     

Species richness of the understory woody vegetation in Japanese cedar plantations declines with increasing number of rotations

The possibility of restoring natural broadleaf forests may be decreased by the effects of plantation management, particularly in sites that undergo repeated rotation. We investigated the following two working hypotheses about the effects of repeated plantation of conifers on the natural regeneration of woody saplings in cool-temperate Japanese cedar plantations: (1) that repeated plantation of conifers decreases sapling species richness, and (2) that repeated plantation of conifers changes sapling species compositions. Our result supported the first hypothesis, because species richness was significantly lower in second-rotation plantations than in first-rotation plantations. The second hypothesis was not supported, because no significant or substantial differences in species composition were observed between plantations with different numbers of rotations. However, the abundance of tree (nonshrub) and gravity-dispersed species decreased after the second rotation of large saplings, albeit not those of small saplings, suggesting that response to repeated rotation depended on sapling size. Our results suggest that it is important to consider factors affecting the maintenance of a species in the plantations, such as distance from natural forests and seed sources, to minimize the effects of repeated plantation.     

含植物提取物的天然沸石日粮对稻鸭生长性能和肠组织的影响

试验旨在研究饲粮中添加沸石制品、植物提物、蛭石（即ZEM）对稻鸭生长性能和肠组织学的影响,试验将14日龄稻鸭分成4组,每组3个重复,每个重复6只鸭（雌雄各半）。稻鸭分别饲喂含ZEM0、0．1、0．5、1．0g·kg^-1的商品鸭碎麦芽日粮63d,结果发现,9周后含ZEM0．1和0．5g·k^-1组稻鸭体增重高于对照组。在饲喂ZEM组稻鸭任意肠段内用低倍显微镜测得的肠绒毛高度、绒毛面积、细胞面积和细胞有丝分裂数目等观测值均高于对照组;饲喂ZEM 0．5g·kg^-1组十二指肠绒毛高度、细胞面积和细胞有丝分裂数目增加显著（P〈0．05）,饲喂ZEM0．1g·kg^-1组空肠细胞分裂也显著增多（P〈0．05）。在扫描电子显微镜下观测到的结果为,所有饲喂ZEM组稻鸭在十二指肠和回肠的绒毛顶部都有突出的上皮细胞和细胞群集。在饲喂ZEM0．1g·kg^-1组稻鸭的空肠中,可数次观测到绒毛回。这些肠组织学上的改变表明,小肠绒毛和上皮细胞功能有可能已经被激活。从目前的结果可以看出,饲喂ZEM组稻鸭十二指肠和回肠的小肠绒毛和上皮细胞都表现出强大的功能,且饲喂ZEM可使稻鸭体增重增加。以上试验结果都表明了饲粮中ZEM的添加水平可至1．0g·kg^-1。     

Molecular identification of Erysipelothrix isolates from the tonsils of healthy cattle by PCR

For 79 isolates from the tonsils of healthy cattle identified as Erysipelothrix by cultivation, biochemical and serological tests, genotypic identification was performed by polymerase chain reaction (PCR) using four species-specific sets of oligonucleotide primers (ER1F-ER1R, ER2F-ER2R, ER3F-ER3R and ER4F-ER4R). The results of PCR for 79 bovine isolates were compared with those of serological typing. For 19 isolates, serotyping and genotyping results were the same. PCR allowed for the identification of 36 untypable isolates as Erysipelothrix species, strain 1. Serotyping and genotyping results of the remaining 24 isolates were different. Supplemental tests are frequently needed for Erysipelothrix identification.     

Notes on beech (Fagus crenata Blume) seed and seedling mortality due to rodent herbivory in a northernmost beech forest, Utasai, Hokkaido

We investigated the mortality of post-dispersal seeds and current-year seedlings of a beech species (Fagus crenata Blume) at the northernmost extent of its geographical range at Utasai (Kuromatsunai), Hokkaido, Japan. Introduced seeds that were protected from rodent predation by wire cages had a higher survival rate from winter to the following spring (mean ± SD: 84 ± 16.7%,n = 10) than controls without cages (mean ± SD: 12 ± 17.9%,n = 10). Unprotected transplanted current-year seedlings with almost fully opened primary leaves never survived more than three days (0%,n = 30), while more than 80% (n = 30) of seedlings within cages survived to the end of the growing season. These results indicate that rodent herbivory is a major mortality factor that strongly prevents the establishment of beech seedlings. In a natural population, we observed that 90% of seedlings (n = 197) disappeared within 10 days after their emergence and rodent herbivory caused this heavy mortality. A herbivorous rodent, the gray-backed vole,Clethrionomys rufocanus bedfordiae, was remarkably abundant in late June (101.5 voles/ha), suggesting that this species strongly affects the formation of the seedling bank. The presence ofC. rufocanus bedfordiae may be one of the reasons for the scarcity of beech seedlings and saplings and the rarity of recruitment in this northernmost beech forest.     

Potential role of hCG in apoptosis of human luteinized granulosa cells

The corpus luteum (CL) forms after ovulation and acts as a temporary endocrine gland that produces progesterone (P4), a hormone that is essential for implantation and maintenance of pregnancy in mammals. In pregnant women, human chorionic gonadotropin (hCG) secreted by the conceptus prevents luteolysis. hCG also increases the survival of cultured human luteinized granulosa cells (hLGCs). To clarify the maintenance mechanism of the human CL, we investigated the effects of hCG and P4 receptor antagonists, onapristone (OP) and RU486, on the viability of hLGCs. With the patients’ consent, hLGCs were isolated from follicular aspirates for in vitro fertilization. The cells were cultured with hCG (0.1, 1, 10, 100 IU/ml), OP (10, 25, 50, 100 μM), RU486 (100 μM), P4 (1, 10, 25, 50 μM) or some combination of the four for 24 h. Cell viability was significantly increased by hCG (100 IU/ml) and significantly decreased by OP (100 μM) compared with the control. Cells treated with hCG and OP together were significantly less viable than the control and OP-treated cells. The combined treatment also significantly increased CASP3 activity and cleaved CASP3 protein expression. Furthermore, P4 addition reversed the reduction in cell viability caused by the combination of hCG and OP treatment. The overall findings suggest that hCG cooperates with P4 to increase survival of hLGCs and to induce apoptosis when P4 action supported by hCG is attenuated in the human CL.     

Expressions of estrogen receptors in the bovine corpus luteum: cyclic changes and effects of prostaglandin F2alpha and cytokines

Estrogen (E) exerts its function by binding to two intracellular estrogen receptors, ERalpha and ERbeta. Although ERs have been reported to be expressed in the bovine corpus luteum (CL), the mechanisms that control ER expression in the bovine CL are not fully understood. To determine the possible regulatory mechanisms of ERalpha and ERbeta that meditate distinct E functions, we examined 1) the changes in the protein expressions of ERs in the CL throughout the luteal phase and 2) the effects of prostaglandin (PG) F2alpha, tumor necrosis factor-alpha (TNFalpha) and interferon-gamma (IFNgamma) on the expressions of ERs in cultured bovine luteal cells. Western blot analyses revealed that ERalpha and ERbeta proteins were expressed throughout the luteal phase. The ERalpha protein level was high at the early luteal (Days 2-3 after ovulation) and mid-luteal stages (Days 8-12) and was extremely low at the regressed luteal stage (Days 19-21). The ERbeta protein level increased from the early to developing luteal stage, remained at the same level at the mid-luteal stage and decreased thereafter. The ratio of ERbeta to ERalpha was higher in the regressed stage than in the other stages. Luteal cells obtained from mid-stage CLs (Days 8-12) were incubated with PGF2alpha (0.01-1 microM), TNFalpha (0.0145-1.45 nM) or IFNgamma (0.0125-1.25 nM) for 24 h. PGF2alpha and TNFalpha inhibited ERa and ERbeta mRNA expressions. IFNgamma suppressed ERbeta mRNA expression but did not affect the expression of ERalpha mRNA. However, the ERalpha and ERbeta protein levels were not affected by any of the above treatments. These data indicate that PGF2alpha, TNFalpha and IFNgamma regulate ERalpha and ERbeta mRNA expressions in bovine luteal cells. Moreover, the changes in the ERbeta/ERalpha ratio throughout the luteal phase suggest that ERalpha is associated with luteal maintenance. Therefore, a dramatic decrease in ERalpha at the regressed luteal stage could result in progression of structural luteolysis in the bovine CL.     

Analysis of the carbohydrates in an East African traditional beverage, togwa

In sub-Saharan Africa, togwa is a widely consumed beverage containing saccharified starch as a basis, being made from cereals and their malt flours. Togwa was prepared from maize flour and finger millet malt in a laboratory, and trace element and amino acid contents in the togwa were analyzed. The major constituents of togwa are carbohydrates, which are starch and starch hydrolysates that seem to be smoothly digested after ingestion. Carbohydrates in field and laboratory togwa were analyzed in detail by two different HPLC systems: normal phase chromatography and a gel permeation chromatography-multi angle laser light scattering system. Togwa was found to contain a very wide range of size distributions of molecules from glucose to starch that have molecular masses in the millions. It was concluded that togwa is an easily digestible and nutritious food.