加温後のtsAF8細胞の細胞周期

Thermotolerance in tsAF8 cells develops during incubation at 34℃ after heating at 45℃, while it is suppressed by the following incubation at a non-permissive temperature of 39.7℃ after the same heating. The incubation temperature after heating may affect the cell cycle and consequently thermotolerance. In the present study, a relationship between the thermotolerance and the cell cycle of tsAF8 was investigated. The cell cycle fractions and DNA synthesis were measured by flow cytometry using double staining with propidium iodide and bromodeoxyuridine. When the tsAF8 cells were heated at 45℃ for 20 min, and thereafter incubated at 34℃, bromodeoxyuridine uptake in the S phase cells (DNA synthesis) was recovered to 65.1% 6 h after the heating, and the cells showed gradual accumulation in the G(2)/M phase. When the cells were incubated at 39.7℃ after heating at 45℃ for 20 min, then showed inhibition of thermotolerance development, the DNA synthesis was recovered to 15.1% temporarily 6 h after the heating, but it became 0% after 12 h, and the cells did not remarkably accumulate in any phases of the cell cycle. This inhibition of DNA synthesis at 39.7℃ was considered to be the result of cell survival decreasing by a step-down heating. However, the relationship between the thermotolerance and the cell cycle was not found out in tsAF8 cells, because the cells did not accumulate in any phases of the cell cycle under the inhibitory condition of thermotolerance.tsAF8細胞は45℃の加温後34℃で培養すると温熱耐性が速やかに発現するが,加温後,制限温度である39.7℃で培養すると温熱耐性の発現が抑制される｡加温後の培養温度が細胞周期に影響し,その結果として温熱耐性発現に影響を与えている可能性があることから,今回,Propidium Iodide(PI)とbromodeoxyuridine(BrdU)でtsAF8細胞を二重染色し,フローサイトメトリーによって温熱耐性と細胞周期の関係の有無について調べた｡tsAF8細胞を45℃20分の加温後34℃で培養すると,6時間後にはG(1)期の細胞が減少し,12時間後にはG(2)/M期への蓄積が見られた｡しかし,加温後39.7℃で培養した場合には細胞周期の進行がほとんど見られなかった｡BrdU の取込みは,加温せずに39.7℃で培養した場合には活発に行われ,また,45℃20分加温後34℃で培養した場合には,6時間後にはBrdUの取り込みは65.1%まで回復した｡しかし,温熱耐性発現の抑制が観察される45℃20分加温後39.7℃で培養した場合には,BrdUの取込み量は6時間後に一時的に15.1%に回復するが,12時間後には取込み量はゼロとなった｡BrdUの取り込みが阻害されたのはstep-down heatingの現象による細胞生存率の減少が原因だと考えられたが,温熱耐性発現の抑制が観察される条件下では細胞周期の特定の時期への集積がなかったことから,温熱耐性と細胞周期との関係はtsAF8細胞においては見い出されなかった

Relationship between intracellular uptake of adriamycin and membrane potential in ADR resistant Ehrlich ascites tumor cells

We observed adiamycin (ADR) uptake and cellular transmembrane potential [amount of intracellular fluorescence of 3,3'- (Di-n-hexyl)- 2,2'- oxacarbocyanine iodide (NK-2280)] in ADR-resistant cells established from Ehrlich ascites tumor cells (EATC) and wild type EATC. In ADR-resistant cells, ADR uptake and the cellular transmembrane potential decreased as the degree of resistance increased. 4,4'- diisothiocyanatostilbene- 2,2'- disulfonic acid (DIDS) induced markedly decreases of ADR uptake and the cellular transmembrane potential. A good correlation was observed between ADR uptake and transmembrane potential in cultured cells

Anti-EGFR antibody cetuximab is secreted by oral squamous cell carcinoma and alters EGF-driven mesenchymal transition

Genetic amplification, overexpression, and increased signaling from the epidermal growth factor receptor (EGFR) are often found in oral squamous cell carcinoma (OSCC) and thus EGFR is frequently targeted molecularly by the therapeutic antibody cetuximab. We assessed effects of cetuximab in control of EGF-driven malignant traits of OSCC cells. EGF stimulation promoted progression level of mesenchymal traits in OSCC cells, which were attenuated by cetuximab but incompletely. We pursued a potential mechanism underlying such incomplete attenuation of OSCC malignant traits. Cetuximab promoted secretion of EGFR-EVs by OSCC cells and failed to inhibit EGF-driven secretion of EGFR-EVs. Cetuximab was also found to be robustly secreted with the EGFR-EVs by the OSCC cells. Thus, EGF promotes the level of mesenchymal traits of OSCC cells and secretion of EGFR-EVs, which involve cetuximab resistance

The utility of three-dimensional dynamic contrast-enhanced

Aneurysmal bone cysts (ABCs) are classified as bone-related lesions based on the 2005 World Health Organization histological classification of odontogenic tumors. Most ABCs are diagnosed using a combination of conventional radiography, computed tomography, magnetic resonance imaging (MRI), and digital subtraction angiography. ABCs should be differentiated from true cysts or other pseudocysts because their treatment is different. Additionally, unlike other cysts, ABCs pose a hemorrhagic risk in surgery; thus, preoperative evaluation of intralesional blood flow is required. Here we report a case of a mandibular ABC in a 39-year-old woman and focus on its dynamic contrast-enhanced MRI (DCE-MRI) features. On DCE-MRI, the lesion was divided into two areas according to the enhancement pattern: the blood-pooling and blood-flow areas. The series of DCE-MR images of the blood-pooling area showed marked enhancement of the margin, but no enhancement in the inner part of the cavity. Additionally, the time-signal intensity curve (TIC) demonstrated no change in the signal intensity (SI) until approximately 15 min after gadolinium-diethylenetriamine penta-acetic acid (Gd-DTPA) administration. In contrast, the series of DCE-MR images of the blood-flow area exhibited marked enhancement in the cyst cavity in the early phase. The TIC showed a rapid increase in SI in the early phase, followed by a rapid decrease until 150 s, and finally a gradual decrease until approximately 15 min after Gd-DTPA administration. Thus, in the current patient, preoperative DCE-MRI clearly delineated the vessel-rich area within the lesion

Diagnostic value of dynamic contrast-enhanced MRI for unilocular cystic-type ameloblastomas with homogeneously bright high signal intensity on T2-weighted or STIR MR images

Typical MR images of ameloblastomas on T2-weighted image (WI) or short inversion time inversion-recovery (STIR) show multiple bright high-signal-intensity loci on a high-signal-intensity background. Unilocular cystic-type ameloblastomas show homogeneously bright high signal intensity on T2WI or STIR as a water-like signal intensity. Therefore, it is difficult to distinguish unilocular cystic-type ameloblastoma from other cystic lesions such as keratocystic odontogenic tumors, radicular cysts (residual cysts) and dentigerous cysts only on the basis of MRI signal intensity. In the present study, we evaluated whether contrast-enhanced (CE)-T1WI and dynamic CE-MRI (DCE-MRI) could provide additional information for differential diagnosis in unilocular cystic-type ameloblastoma. Images from 12 cases of suspected unilocular cystic-type ameloblastoma were evaluated in the present study. Of them, 5 had areas suspected of indicating a solid component on T1WI and T2WI (or STIR). Ten had undergone additional CE-T1WI and DCE-MRI. On 5 of 10 cases of CE-T1WI, a tiny enhancement area was detected. On 6 of 10 DCE-images, a time-course enhanced area which was suspected to be a solid component was detected. CE-T1WI was helpful in the diagnosis of ameloblastoma because the tiny enhanced areas were taken to indicate possible solid components. Moreover, the rim-enhancement area on CE-T1WI could be divided into small regions of interest, and some of these showed slightly increased enhancement on DCE-MRI, which was taken to indicate a solid component and/or intramural nodule with focal invasion of ameloblastoma tissue. DCE-MRIs of the four remaining cases, which provided no clues to the diagnosis of ameloblastoma in the manner of the above descriptions, showed thicker rim enhancement than odontogenic cysts. Thus, CE-T1WI and DCE-MRI were helpful in the differential diagnosis of unilocular cystic-type ameloblastomas with homogeneously bright high signal intensity on T2WI or STIR

Oyygen uptake of adriamycin resistant cells of Ehrlich ascites tumor

エールリッヒ腹水癌細胞を用いアドリアマイシンに対する耐性細胞（ADR耐性細胞）を樹立した。電子顕微鏡を用い撮影写真から細胞質当たりのミトコンドリア（MT）の割合を面積比で求めた。親株に比較して1μg/ml ADR耐性細胞では1.32倍、10μg/ml ADR耐性細胞では1.47倍であった。これらの細胞の呼吸を測定した。耐性細胞の内発呼吸は親株に比較して増加していた。1μg/ml ADR耐性細胞では1.45倍、10μg/ml ADR耐性細胞では1.49倍であり、MTの増加量とほぼ同じ割合であった。これらのことから、細胞が耐性になるとエネルギー消費が高まるために細胞内MTが増加し、その結果呼吸（酸素消費）が増加することが推察された。Adriamycin-resistant cells of Ehrlich ascites tumor cells were established in our laboratory. Using electron microscope, the area of mitochondria (MT) per cytoplasm of ADR-resistant cells were measured with planimeter. The values of wild-type cells, 1μg/ml ADR-resistant cells and 10μg/ml ADR-resistant cells were 39.3, 51.8 and 57.7 μ(2) per 1,000 μ(2) of cytoplasm, respectively. Oxygen consumption of 1 μg/ml ADR-resistant cells and 10 μg/ml ADR-resistant cells were 1.45-fold and 1.49-fold compared to that of wild-type cells, respectively. These results indicate that ADR-resistant cells require more energy to work efflux pump than wild-type cells

Effects of demethylating agent 5-aza-2 '-deoxycytidine and histone deacetylase inhibitor FR901228 on maspin gene expression in oral cancer cell lines

Maspin, which belongs to the serine protease inhibitor (serpin) superfamily, has been proposed as a potent tumor suppressor that inhibits cell motility, invasion, angiogenesis, and metastasis. In the present study, we examined the effects of 5-aza-2'-deoxycytidine (5-aza-dC), a demethylating agent, and FR901228, a histone deacetylase (HDAC) inhibitor, on maspin expression in oral cancer cell tines. The expression levels of maspin mRNA were divided into two groups, which was the maspin tow-expressed and high-expressed cell lines in the 12 oral cancer cell lines. The maspin promoter contained only a few methylated CpG sites in the maspin low-expressed cell lines. Moreover, the methylation status was not altered after 5-aza-dC treatment. However, the transcription of the maspin gene was clearly increased following 5-aza-dC treatment in a number of oral cancer cell tines. These results imply that an action of 5-aza-dC is separate from induction of promoter demethylation. Treatment with FR901228 resulted in a time-dependent stimulation of the re-expression of maspin mRNA as early as 4 h after treatment in the maspin downregulated cells. The re-expression of the maspin gene may contribute to the recuperation of biological functions linked to FR901228 such as an inhibitory effect on tumor angiogenesis and cell invasion. These results indicate that maspin and its target genes may be excellent leads for future studies on the potential benefits of FR901228, a HDAC inhibitor, in cancer therapy.</p

Depletion of Lipid Efflux Pump ABCG1 Triggers the Intracellular Accumulation of Extracellular Vesicles and Reduces Aggregation and Tumorigenesis of Metastatic Cancer Cells

The ATP-binding cassette transporter G1 (ABCG1) is a cholesterol lipid efflux pump whose role in tumor growth has been largely unknown. Our transcriptomics revealed that ABCG1 was powerfully expressed in rapidly metastatic, aggregative colon cancer cells, in all the ABC transporter family members. Coincidently, genetic amplification of ABCG1 is found in 10–35% of clinical samples of metastatic cancer cases. Expression of ABCG1 was further elevated in three-dimensional tumoroids (tumor organoids) within stemness-enhancing tumor milieu, whereas depletion of ABCG1 lowered cellular aggregation and tumoroid growth in vitro as well as hypoxia-inducible factor 1α in cancer cells around the central necrotic areas in tumors in vivo. Notably, depletion of ABCG1 triggered the intracellular accumulation of extracellular vesicles (EVs) and regression of tumoroids. Collectively, these data suggest that ABCG1 plays a crucial role in tumorigenesis in metastatic cancer and that depletion of ABCG1 triggers tumor regression with the accumulation of EVs and their derivatives and cargos, implicating a novel ABCG1-targeting therapeutic strategy by which redundant and toxic substances may be accumulated in tumors leading to their regression

Assessment of MRI and dynamic contrast-enhanced MRI in the differential diagnosis of adenomatoid odontogenic tumor

The radiographical differentiation of adenomatoid odontogenic tumor (AOT) from dentigerous cysts, calcifying odontogenic cysts, calcifying epithelial odontogenic tumors, odontogenic keratocysts, and amelobastomas is sometimes difficult. We attempted to differentiate AOT from other lesions similar to AOT in radiographic findings using MRI. The MRI features of AOT in our 3 cases included homogeneous low SI in the cystic portion and homogeneous intermediate SI in the solid portion on T1WI, homogeneous high SI in the cystic portion and intermediate to slightly high SI in the solid portion on T2WI, and enhancement of only the solid portion on CE-T1WI although non of the sequences included SI of calcifications. The contrast index curves in the 3 cases of AOT showed a gradual increase to 300 s, which signified a benign tumor. These MRI features were characteristic features of AOT and might be a basis for differentiating AOT from the above possible lesions in radiographic examinations.</p