In higher animal cells, duplication of centrosomes is triggered by CDK2/cyclin E-mediated phosphorylation. Nucleophosmin (NPM)/B23, a multifunctional protein, has recently been identified as one of ...the substrates of CDK2/cyclin E in centrosome duplication. Centrosome-bound NPM/B23 dissociates from centrosome upon phosphorylation by CDK2/cyclin E, which in turn triggers initiation of centriole duplication. Duplicated centrosomes remain free of NPM/B23 till mitosis. When the nuclear membrane breaks down during mitosis, NPM/B23 re-localizes to centrosomes. Upon cytokinesis, each daughter cell receives one centrosome bound by NPM/B23, which again dissociates from the centrosome upon exposure to CDK2/cyclin E at mid-late G1 phase of the next cell cycle. Thus, NPM/B23 would constitute one of the licensing systems for centrosome duplication, ensuring the coordination of centrosome and DNA duplication, which limiting duplication once per cell cycle.
Recent research has focused on immunotherapy, particularly with regard to cancer treatment. Programmed death-1 and programmed death ligand 1 (PD-1/PD-L1) pathway blockade is a central topic of the ...promising immunotherapy field. In veterinary medicine, observations of the PD-1/PD-L1 pathway, including the relationship between immune cells and diseases, have increased. In this study, monoclonal antibodies specific to canine PD-1 and PD-L1 were developed, and the antibodies against PD-1 and PD-L1 bind to PD-1 and PD-L1 overexpressing cells, respectively. Additionally, each antibody interfered with the interaction between PD-1 and PD-L1. The expression of PD-1 and PD-L1 was detected on activated T cells from canine peripheral blood mononuclear cells (PBMC), and, remarkably, was the first recorded instance of PD-L1 expression on canine immature dendritic cells. Production of IFN-γ by activated T cells increased significantly when incubated with anti-PD-1 antibody alone and with both anti-PD-1 and anti-PD-L1 antibodies, revealing the functional effects of the antibodies. The antibodies will be useful for research on immune systems and may be the first passive immunotherapy approach in canine cancer patients.
Canine lymphoma/leukemia cell lines with p16 protein expressions: high (17–71 and GL-1) and low (CLBL-1, CLC, Nody-1, and UL-1) were treated in vitro with cyclin-dependent kinase 4 and 6 (CDK4/6) ...inhibitors, palbociclib or abemaciclib. Cell proliferation decreased as a result, with higher IC50 levels observed in the high p16 (17–71 and GL-1) and one low p16 (UL-1) cell lines compared with the low p16 cells (CLBL-1, CLC, and Nody-1). As expected, palbociclib and abemaciclib treatment reduced pRb phosphorylation in a dose-dependent manner, especially in cells with low p16. These results suggest that CDK4/6 inhibitors have potential as new chemotherapeutic agents for canine lymphoma and high p16 protein expression may be used as a biomarker for resistance to CDK4/6 inhibitor therapy.
To investigate the bioactivities of fresh garlic and its processed product, black garlic, we conducted comparative analyses of antioxidant, anti-inflammatory, innate immune activation, and ...anti-cancer activities in addition to the chemical composition (sugar, amino acid, and polyphenol contents) of these materials. Simultaneous assay using neutrophil-like cells showed that fresh garlic exhibited antioxidant and innate immunostimulatory activities, whereas black garlic displayed a potent anti-inflammatory effect. The antioxidant activity index was correlated with phenol and flavonoid contents, while the innate immunostimulatory activity was correlated with fructan content. Furthermore, some black garlics with low fructose content were found to inhibit the proliferation of UM-UC-3 cancer cells, while other black garlics rich in fructose increased UM-UC-3 cell proliferation. It was shown that the processing of fresh garlic could change the composition of sugars, antioxidants, and amino acids, which have different effects on neutrophil-like cells and UM-UC-3 cells, as well as on bioactivities.
Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tickborne infectious disease caused by SFTS virus (SFTSV). We report 7 cases of spontaneous fatal SFTS in felines. Necropsies ...revealed characteristic lesions, including necrotizing lymphadenitis in 5 cases and necrotizing splenitis and SFTSV-positive blastic lymphocytes in all cases. We detected hemorrhagic lesions in the gastrointestinal tract in 6 cases and lungs in 3 cases, suggesting a more severe clinical course of SFTS in felids than in humans. We noted necrotic or ulcerative foci in the gastrointestinal tract in 3 cases, the lung in 2 cases, and the liver in 4 cases. We clarified that blastic lymphocytes are predominant targets of SFTSV and involved in induction of necrotic foci. We also found that thymic epithelial cells were additional targets of SFTSV. These results provide insights for diagnosing feline SFTS during pathological examination and demonstrate the similarity of feline and human SFTS cases.
Abstract
A cat was presented with mast cell tumors (MCTs) of the skin and spleen. During the initial diagnosis, the exon 8 mutation of
c-KIT
was detected in the masses from skin and spleen by a ...commercial laboratory test. Consequently, treatment with toceranib was started. After complete remission, because of recurrence on day 117, the spleen and skin tumors were removed, but the cat eventually died on day 191. The analysis of ten cDNA clones of the
c-KIT
gene cloned from the surgically removed spleen revealed that seven different cDNA patterns were included, indicating the heterogeneity of this gene in the splenic MCT. The seven cDNA nucleotide patterns can be classified into four protein sequence patterns. In addition to the previously known mutations in exon 8, we identified novel mutations in exons 9, 10, and 18; four amino acids deletion in exon 9, and a point mutation in exons 10 and 18. Mouse IL-3-dependent cell line, Ba/F3, was transduced with these mutant clones, and c-KIT phosphorylation and proliferation assays were performed. We found that certain mutations affected the c-KIT phosphorylation status and cell proliferation. This suggests that heterogeneity among the population of tumor cells exists in MCTs, and that the dominant clones of this heterogeneity may contribute to the subsequent tumor cell growth.
Cyclin-dependent kinase inhibitor p16 (CDKN2A) primarily functions as a negative regulator of the retinoblastoma protein (pRb) pathway to prevent pRb phosphorylation, thus playing a critical role in ...cell cycle arrest. In canine lymphoma cells, methylation due to inactivation of the p16 gene has been reported. However, its protein expression has not been examined in previous studies. In our in vitro study, the gene and protein expression of p16 and phosphorylated pRb were examined simultaneously in eight canine lymphoma and leukemia cell lines (17-71, CLBL-1, GL-1, CLC, CLGL-90, Ema, Nody-1, and UL-1). Methylation of the p16 gene was also explored using the demethylation drug 5-Aza-2′-deoxycytidine (5-Aza). After 5-Aza treatment, p16 gene and protein expression increased and pRb phosphorylation decreased, suggesting that both hypermethylation of the p16 gene and pRb hyperphosphorylation occurred in four out of eight cell lines (CLBL-1, CLC, Nody-1, and UL-1). Moreover, the estimation of p16’s protein expression was better than that of p16’s mRNA expression because the expression of the protein was more stable than those of the gene, and highly related to the phosphorylation of pRb. These results revealed that p16’s protein expression could be a promising biomarker for canine lymphoma cells.
Glucocorticoids, among the most widely utilized drugs in veterinary medicine, are employed to treat a wide variety of diseases; however, their use often induces adverse events in dogs. The efficacy ...of glucocorticoids usually depends on dosage, although differences in sensitivity to glucocorticoids in individual animals have been reported. Glucocorticoids bind to the cytoplasmic glucocorticoid receptor (GR), which is expressed in almost all cells. These receptors are key factors in determining individual sensitivity to glucocorticoids. This study examined individual differences in glucocorticoid sensitivity in dogs, focusing on reactivity of the GR to prednisolone.
We first molecularly cloned the GR gene from a healthy dog. We discovered a mutant GR in a dog suspected to have iatrogenic Cushing syndrome. The mutant GR had extra nucleotides between exons 6 and 7, resulting in a truncated form of GR that was 98 amino acids shorter than the wild-type dog GR. The truncated GR exhibited very low reactivity to prednisolone, irrespective of concentration.
We have identified the truncated form of canine GR in a dog with iatrogenic Cushing syndrome. This truncated form showed the very less sensitivity to glucocorticoid in vitro, unfortunately, we could not elucidate its clinical significance. However, our data is a first report about the function of canine GR, and will facilitate the analysis of canine glucocorticoid sensitivity.
A 1-year-old spayed female Miniature Schnauzer had chronic hyponatremia, accompanied by polyuria and polydipsia. Blood tests and urinalysis revealed severe hyponatremia, low plasma osmolality with ...euvolemia, and increased sodium excretion in urine. Hypothyroidism and hypoadrenocorticism were ruled out as causes. These findings led to the diagnosis of syndrome of inappropriate secretion of antidiuretic hormone (SIADH). Magnetic resonance imaging (MRI) showed dilation of the lateral ventricles, indicating severe hydrocephalus. Tolvaptan, a vasopressin V2 receptor antagonist commonly used in human SIADH, was administered along with water restriction. This treatment resulted in a consistent increase in plasma sodium levels without any adverse effects. This case report represents the first documented evidence of the therapeutic efficacy of tolvaptan in treating SIADH in a dog.
Recombinant canine interferon-γ (rc-IFNγ; InterdogⓇ) was exclusively approved as a therapeutic for canine atopic dermatitis. However, it has been used off-label for the treatment of canine cancer. We ...examined the inhibitory effect of rc-IFNγ on the growth of canine tumor cell lines and analyzed its mechanism of action. Three (CTB-p, CTB-m, and CNM-m) out of seven mammary gland tumor cell lines and two (VIMC and CoMS) out of four mast cell tumor cell lines showed remarkable growth inhibition after treatment with rc-IFNγ. However, one (CLBL-1) out of nine lymphoma cell lines showed a significant amount of cell death. Using CTB-p and CTB-m cell lines, we showed that STAT1 was essential for inducing the growth inhibitory effect of rc-IFNγ. Although rc-IFNγ induced G1 growth arrest in CTB-p cell line, treatment with rc-IFNγ did not alter the expression of cell cycle regulatory proteins. In this study, we observed direct cytotoxicity or cytostatic effects of rc-IFNγ in canine tumor cell lines. However, the detailed mechanisms responsible for these effects need to be elucidated in the future.
•Recombinant canine interferon-γ (rc-IFNγ) showed the inhibitory effect on the growth of several canine tumor cell lines.•rc-IFNγ induced G1 growth arrest in one of the canine mammary gland tumor cell lines.•STAT1 is essential for inducing the growth inhibitory effect of rc-IFNγ.
PDF
