小鼠黑色素瘤功能异常CD8~+T细胞基因调控网络构建与分析

疾病是全球范围内危害畜禽健康的主要问题,解析免疫系统调控相关基因成为当前抗病分子育种的研究热点。为研究小鼠黑色素瘤模型功能异常CD8~+T细胞对免疫抗体处理的应答机制,通过基因调控网络方法筛选功能异常CD8~+T细胞在Ig G和PD1抗体处理的特定基因、转录因子和细胞表面受体。结果发现对照组、PD1抗体与Ig G抗体处理组分别有28、17、33个特定表达基因。与对照组相比,Ig G和PD1处理组有2个转录因子Zfx、Zfhx3关闭表达,PD1处理组细胞表面受体Raet1b表达。Raet1b基因的表达可能使小鼠黑色素瘤功能异常CD8~+T细胞恢复功能,结果为抗病育种的相关基因研究提供理论基础。     

Isoprenoid-substituted flavonoids from wood of Artocarpus heterophyllus on B16 melanoma cells: Cytotoxicity and structural criteria

As a result of cytotoxicity-guided fractionation, nine flavonoids, artocarpin (1), cudraflavone C (2), 6-prenylapigenin (3), kuwanon C (4), norartocarpin (5), albanin A (6), cudraflavone B (7), brosimone I (8) and artocarpanone (9) were identified from the methanol extract of the wood of Artocarpus heterophyllus, known commonly as Nangka in Indonesia. A structure–activity investigation of the effect of these isolated compounds (1–9) and structurally related compounds on B16 melanoma cells indicated that isoprenoid moiety substitutions in flavonoids enhance their cytotoxicity, and that the position of attachment and the number of isoprenoid-substituent moieties per molecule influence flavonoid cytotoxicity.     

Primary malignant melanoma in the oesophagus of a foal

A 2‐month‐old filly was evaluated for severe colic. Ultrasound, abdominocentesis and physical examination findings prompted an abdominal exploratory surgery. Perforation of the stomach was discovered during the surgery. The filly was humanely subjected to euthanasia under anaesthesia and post mortem examination was performed. In addition to gastric and duodenal ulceration, a thickened, black area of the proximal oesophagus was discovered. Histopathology of the lesion revealed primary malignant melanoma. Although rare, primary melanoma can occur in noncutaneous locations.     

The Marine Fungal Metabolite,Dicitrinone B,Induces A375 Cell Apoptosis through the ROS-Related Caspase Pathway

Dicitrinone B, a rare carbon-bridged citrinin dimer, was isolated from the marine-derived fungus, Penicillium citrinum. It was reported to have antitumor effects on tumor cells previously; however, the details of the mechanism remain unclear. In this study, we found that dicitrinone B inhibited the proliferation of multiple tumor types. Among them, the human malignant melanoma cell, A375, was confirmed to be the most sensitive. Morphologic evaluation, cell cycle arrest and apoptosis rate analysis results showed that dicitrinone B significantly induced A375 cell apoptosis. Subsequent observation of reactive oxygen species (ROS) accumulation and mitochondrial membrane potential (MMP) reduction revealed that the apoptosis induced by dicitrinone B may be triggered by over-producing ROS. Further studies indicated that the apoptosis was associated with both intrinsic and extrinsic apoptosis pathways under the regulation of Bcl-2 family proteins. Caspase-9, caspase-8 and caspase-3 were activated during the process, leading to PARP cleavage. The pan-caspase inhibitor, Z-VAD-FMK, could reverse dicitrinone B-induced apoptosis, suggesting that it is a caspase-dependent pathway. Our data for the first time showed that dicitrinone B inhibits the proliferation of tumor cells by inducing cell apoptosis. Moreover, compared with the first-line chemotherapy drug, 5-fluorouracil (5-Fu), dicitrinone B showed much more potent anticancer efficacy, suggesting that it might serve as a potential antitumor agent.     

Excision of a feline limbal melanoma and use of nictitans cartilage to repair the resulting corneoscleral defect

A limbal melanoma was surgically removed from a 12-year-old castrated male black domestic shorthair (DSH) cat. The resulting full-thickness eye wall defect was repaired using the autologous third eyelid cartilage. The patient was followed for 85 days postoperatively, during which time there was no recurrence. Other than a small amount of fibrin and blood at the anterior lens capsule, no significant complications were seen. Use of the third eyelid cartilage as graft material following resection of a feline limbal melanoma can be effective for repairing large eye wall defects and preserving ocular function. The third eyelid proved to be a convenient source of graft material. In addition, autologous grafting can reduce the potential for iatrogenic spread of infectious agents such as feline herpes virus.     

Canine limbal melanoma: 30 cases (1992-2004). Part 2. Treatment with lamellar resection and adjunctive strontium-90beta plesiotherapy--efficacy and morbidity

OBJECTIVES: To evaluate the efficacy of lamellar resection and adjunctive 90Sr-beta plesiotherapy in the management of canine limbal melanoma and the frequency and morbidity of early and late-onset side effects following treatment. DESIGN: Retrospective study. ANIMALS STUDIED: Thirty dogs treated for limbal melanoma at the Animal Health Trust (AHT), between 1992 and 2004. METHODS: Medical records were reviewed and information collected on the radiation dose administered, duration of follow-up, and the frequency of early- and late-onset ocular side effects. Follow-up information was obtained by re-examination of patients or completion of a telephone interview with the referring veterinary surgeons or the owners. RESULTS: Recurrence of limbal melanoma occurred in 1 of 30 cases (3%; 95% CI: 0.2-14%). One or more side effects occurred in 16 of 30 cases (53%; 95% CI: 28.1-74.9). The occurrence of side effects was not associated with the number of sites treated or the total surface dose of radiation. Short-term side effects were reported in 16/30 cases (53%) and included corneal scarring (12/30), corneal neovascularization (6/30), conjunctivitis with or without granulation tissue formation (5/30), lipid keratopathy (1/30), and localized bullous keratopathy (2/30). Long-term side effects were reported in 6 of 30 cases (20%) cases and included deep scleral thinning (5/30), focal scleromalacia (3/30), globe perforation (2/30), lipid keratopathy (3/30), localized bullous keratopathy (1/30), and sectoral cortical cataract (1/30). CONCLUSIONS: Lamellar resection and adjunctive 90Sr-beta plesiotherapy is relatively easy to perform, is minimally invasive, and extremely well tolerated by patients. The treatment is highly efficacious in the management of canine limbal melanoma. Modification of the radiation dosimetry, in particular reducing the total dose of radiation and the dose per radiation fraction, is necessary to reduce the incidence of late onset side effects.     

PDGFRs expression in dogs affected by malignant oral melanomas: correlation with prognosis

Canine malignant melanoma (CMM) is the most common canine oral tumour, and up to 70–75% of dogs in stage II–III die within 1 year after surgery. The purpose of this study was to evaluate the expression of platelet‐derived growth factors receptors (PDGFR)‐α and ‐β in stage II and III CMMs and to correlate it with prognosis. PDGFRs expression was evaluated by immunohistochemistry on 48 cases of formalin‐fixed CMM samples and correlated with clinical–pathological findings and outcome after surgery. PDGFRs co‐expression was observed in 37.5% of cases. Positivity for PDGFR‐α and ‐β receptor was present in 54.2 and 47.9% of cases, respectively. Ki67 values >19.5% were ascertained in 66.7% of cases. Statistical analysis showed that PDGFRs co‐expression and Ki67 values > 19.5% were both associated with worse prognosis. PDGFRs expression suggests a role in the pathogenesis and progression of CMM, and α and β co‐expression appears to be associated to worse prognosis.     

Prolongation of survival of dogs with oral malignant melanoma treated by en bloc surgical resection and adjuvant CSPG4‐antigen electrovaccination

Reported post‐surgery 1‐year survival rate for oral canine malignant melanoma (cMM) is around 30%; novel treatments are needed as the role of adjuvant chemotherapy is unclear. This prospective study regards adjuvant electrovaccination with human chondroitin sulfate proteoglycan‐4 (hCSPG4)‐encoded plasmid in 23 dogs with resected II/III‐staged CSPG4‐positive oral cMM compared with 19 dogs with resected only II/III‐staged CSPG4‐positive oral cMM. Vaccination resulted in 6‐, 12‐, 18‐ and 24‐month survival rate of 95.6, 73.9, 47.8 and 30.4%, respectively [median survival time (MST) 684 days, range 78–1694, 8 of 23 dogs alive] and 6‐, 12‐, 18‐ and 24‐month disease‐free interval (DFI) rate of 82.6, 47.8, 26.1 and 17.4%, respectively (DFI 477 days, range 50–1694). Non‐vaccinated dogs showed 6‐, 12‐, 18‐ and 24‐month survival rate of 63.2, 26.3, 15.8 and 5.3%, respectively (MST 200 days, range 75–1507, 1 of 19 dogs alive) and 6‐, 12‐, 18‐ and 24‐month DFI rate of 52.6, 26.3, 10.5 and 5.3%, respectively (DFI 180 days, range 38–1250). Overall survival and DFI of vaccinated dogs was longer in those <20 kg. In vaccinated and non‐vaccinated dogs local recurrence rate was 34.8 and 42%, respectively while lung metastatic rate was 39 and 79%, respectively.     

Mutation analysis and gene expression profiling of ocular melanomas in cats

Feline ocular melanomas show a high malignant behaviour, but adjunctive therapies are non‐existent. The aim of this pilot study was to determine, whether feline ocular melanomas harbour mutations comparable to mutations in human melanomas and to evaluate the gene expression status of genes known to be involved in initiation and progression of human melanomas. Mutation hotspot regions of several genes of feline ocular melanomas were analysed by DNA sequencing and RNA expression levels of the respective genes and others were evaluated by quantitative real‐time polymerase chain reaction (RT‐qPCR). Common mutations found in human melanomas are not present in feline tumours. Gene expression analysis revealed a significant upregulation of KIT and LTA4H, as well as a downregulation of GNAQ, GNA11, BRAF and RASSF1 in feline ocular melanomas. As KIT seems to harbour a potential as target gene in human uveal melanomas, future studies should further investigate the potential of KIT as target for adjunctive therapy in feline ocular melanomas.