Plasma matrix metalloproteinase-9 activity in cats with lymphoma.

In this study, plasma MMP-9 activity was evaluated in cats with lymphoma. Plasma samples were obtained from 26 cats with lymphoma before treatment. From 13 of the included 26 cats, plasma samples were obtained 4 weeks after the initiation of treatment. Plasma samples were also obtained from 10 healthy cats as a control. Plasma MMP-9 activity was examined by gelatin zymography and semi-quantitative value (arbitrary unit; a.u.) for each sample was calculated. Relatively high levels of MMP-9 were observed in cats with lymphoma compared with those in healthy control cats. MMP-9 quantification through zymography showed significantly higher activity in cats with lymphoma (median, 0.63 a.u.; range, 0.23-3.24 a.u.) than in healthy controls (0.22 a.u.; 0.12-0.46 a.u.; P < 0.01). MMP-9 activities were significantly different before (0.73 a.u.; 0.30-3.24 a.u.) and after treatment (0.50 a.u.; 0.14-1.32 a.u.; P = 0.017). Measuring plasma MMP-9 activity in cats with lymphoma may become an appropriate monitoring tool for feline lymphoma.

A comparison of surgery and radiation therapy for cT1 esophageal squamous cell carcinoma.

Surgery and radiation therapy have been used to treat esophageal squamous cell carcinoma. However, treatment outcomes have not yet been extensively investigated. The aim of this study was to compare surgery and radiation therapy for clinical T1 esophageal squamous cell carcinoma. A total of 67 clinical T1 esophageal squamous cell carcinoma patients were treated between January 1997 and December 2005; 29 had undergone radical esophagectomy (surgery group) and 38 were treated with definitive radiation therapy (radiation group). The mean patient age was lower in the surgery group than in the radiation group. In surgery group, respiratory complications, anastomotic leaks, recurrent nerve palsies, and anastomotic stenosis occurred in 7, 8, 6, and 5 patients, respectively. In radiation group, leucopenia, esophagitis, pericarditis were observed in 15, 3, and 3 patients, respectively. The 5-year overall survival rate for the surgery group was 68.9%, and 74.3% for the radiation group. There were no significant difference between groups (P= 0.3780). The 5-year relapse-free survival rate in the surgery group was 61.8% and 38.8% in the radiation group. The relapse-free survival rate was significantly higher in the surgery group than in the radiation group (P= 0.0051). The 5-year overall and relapse-free survival rates for tumors invaded into but not through the muscularis mucosa were 83.3% and 75.0%, respectively, in the surgery group and 78.8% and 33.3%, respectively, in the radiation group. There were no significant differences. The 5-year overall survival rates for patients with tumors that invaded the submucosal layer was 64.9% in the surgery group and 66.5% in the radiation group. This difference was not significant (P= 0.8712). The 5-year relapse-free survival rate in the surgery group (56.0%) was significantly higher than that in the radiation group (41.8%; P= 0.0219). In conclusion, surgery may become a standard treatment for cT1 esophageal cancer that can offer longer relapse-free survival, particularly for patients with tumors that invade the submucosa.

p53-dependent hyperthermic enhancement of tumour growth inhibition by X-ray or carbon-ion beam irradiation.

To elucidate p53-dependency on combined treatment with radiation and hyperthermia, growth inhibition and apoptosis were analysed using transplantable human tumour. Human head and neck squamous cell carcinoma (HNSCC) cells carrying different p53 genes were transplanted into the thigh of nude mice. When the mean diameter of tumour reached 5-6 mm, the tumours were exposed to X-rays (2 Gy) or Carbon-ion (C-) beams (1 Gy) followed by heating at 42 degrees C for 20 min. Tumour growth inhibition was evaluated by measuring the diameters of tumour. The induction of apoptosis and accumulation of apoptosis-related proteins were also analysed by immunohistochemical staining. Synergistic enhancement of tumour growth inhibition by hyperthermia was observed in wild-type p53 tumours treated with X-rays or C-beams but not in mutant p53 tumours. The incidence of apoptotic cells and activated-caspase-3-positive cells after combined treatment with them were significantly high in wild-type p53 tumours compared with that in mutant p53 tumours. The hyperthermic enhancement of tumour growth inhibition by X-ray- or C-beam-irradiation was p53-dependent, suggesting that it might be highly correlated with p53-dependent apoptosis.

Heat-induced growth inhibition and apoptosis in transplanted human head and neck squamous cell carcinomas with different status of p53.

To examine p53-dependency in hyperthermic cancer therapy, heat-induced growth inhibition and apoptosis in transplanted human head and neck squamous cell carcinoma (HNSCC) tumours were analysed with different status of p53 into nude mice. The tumour tissue from HNSCC cell line (SAS) transfected with mutant p53 gene (SAS/mp53) or control vector containing neo gene (SAS/neo) was transplanted into the subcutaneous tissue of the thigh of nude mice using a trocar. Hyperthermia was performed at 42 degrees C when the mean diameter of tumour was 5-6mm. The diameter of tumours was measured using vernier calipers and tumour weight (TW) and the relative tumour weight (RW) was calculated. Tumour regrowth delay was evaluated when the RW reached 5-fold against the control group. The accumulation of p53 and Bax proteins was examined by an immunohistochemical technique. Apoptotic cells in the sections were detected by staining of DNA ends using an immunohistochemical technique. SAS/mp53 tumours showed more heat-resistance than SAS/neo tumours. The p53-positively staining cells were observed in untreated SAS/mp53 tumours, but not in untreated SAS/neo tumours. After heat treatment, the accumulation of p53 and Bax proteins was observed in SAS/neo tumours, but little in SAS/mp53 tumours. The incidence of apoptotic cells induced by heat treatment was very low in SAS/mp53 tumours compared with SAS/neo tumours. In conclusion, the heat-induced growth inhibition of a transplanted HNSCC may be correlated with the induction of p53-dependent Bax-mediated apoptosis. Thus, p53 status appears to be one of the useful parameters for the predictive assays in hyperthermic cancer therapy.

p53-dependent thermal enhancement of cellular sensitivity in human squamous cell carcinomas in relation to LET.

PURPOSE: To investigate the dependence on p53 gene status of the thermal enhancement of cellular sensitivity against different levels of linear energy transfer (LET) from X-rays or carbon-ion (C-) beams. MATERIALS AND METHODS: Two kinds of human squamous cell carcinoma cell lines were used with an identical genotype except for the p53 gene. SAS/mp53 cells were established by transfection with mutated p53 (mp53) gene to SAS cells having functional wild-type p53 (wtp53). As the control, a neo vector was transfected to the SAS cells (SAS/neo cells). Both cells were exposed to X-rays or accelerated C-beams (30-150 KeV microm(-1)) followed by heating at 44 degrees C. Cellular sensitivity was determined by colony-forming activity. Induction of apoptosis was analysed by Hoechst 33342 staining of apoptotic bodies and agarose-gel electrophoresis for the formation of DNA ladders. RESULTS: It was found that (1) there was no significant difference in cellular sensitivity between SAS/neo and SAS/mp53 cells to LET radiation of >30 KeV microm(-1), although the radiosensitivity of SAS/neo cells to X-rays was higher (1.2-fold) than that of SAS/mp53 cells; (2) there was an interactive thermal enhancement of radiosensitivity below an LET of 70 KeV microm(-1) in SAS/neo cells, although only additive thermal enhancement was observed in SAS/mp53 cells through all LET levels examined; (3) low-LET radiation induced apoptosis only in SAS/neo cells; (4) high-LET radiation at an isosurvival dose-induced apoptosis of SAS/neo cells at a higher frequency compared with that with low-LET radiation; (5) high-LET radiation-induced p53-independent apoptosis in SAS/mp53 cells; and (6) thermal enhancement of cellular sensitivity to X-rays was due to induction of p53-dependent apoptosis. CONCLUSIONS: The findings suggest that thermal enhancement of radiosensitivity may result from p53-dependent apoptosis induced by inhibition of p53-dependent cell survival system(s) through either regulation of the cell cycle or induction of DNA repair. It is also suggested that the analysis of p53 gene status of cancer cells may predict response to combined therapies with low-LET radiation and hyperthermia.

The dependence of p53 on the radiation enhancement of thermosensitivity at different let.

PURPOSE: The aim of this study is to investigate the dependence of p53-gene status on the radiation enhancement of thermosensitivity at different levels of linear energy transfer (LET). METHODS AND MATERIALS: We used two kinds of human glioblastoma transfectants of A-172 cells bearing the wild-type p53 gene, A-172/neo cells with control vector containing the neo gene and A-172/mp53 cells with both the dominant negative mutated p53 gene and neo gene. We exposed these cells to X-rays and accelerated carbon-ion (C-) beams (13-200 KeV/microm) followed by heating at 44 degrees C. Cellular sensitivities were determined using clonogenic assay. RESULTS: The radiation enhancement of thermosensitivity was LET-dependent for the A-172/neo cells, but this was not clearly demonstrated in the A-172/mp53 cells. The supraadditive radiation enhancement of thermosensitivity was observed in A-172/neo cells at the LET range of 13 to 70 KeV/microm, though only an additive effect was observed at higher LET. In A-172/mp53 cells, only an additive effect was observed through all the LET examined. CONCLUSION: These results indicate that the radiation enhancement of thermosensitivity is p53- and LET-dependent. Our results suggest that the combined use of high-LET radiation and hyperthermia brings useful application for cancer therapeutic purposes.

WAF1 accumulation by carbon-ion beam and alpha-particle irradiation in human glioblastoma cultured cells.

PURPOSE: There have been no reports about the effects of heavy-ion beams on the expression of the WAF1 gene, although ionizing radiation such as y-rays and X-rays is well known to induce WAF1 (p21/CIP1/sdi1) gene expression in a p53-dependent manner. In the present study, it was examined whether WAF1 accumulation was induced after carbon-ion (C-) beam or alpha-particle irradiation in four glioblastoma cell lines. MATERIALS AND METHODS: A colony assay for radiosensitivity and Western blot analysis of WAF1 were applied to two human glioblastoma cell lines, A-172 bearing wild-type p53 (wtp53) and T98G bearing mutated p53 (mp53). A-172/neo and A-172/mp53 were transfected with a control vector (containing only a neo selection marker) and a mp53 expression vector respectively. RESULTS: The amount of WAF1 increased markedly after X-ray irradiation in A-172 and A-172/neo cells but not in T98G and A-172/mp53 cells. The level of WAF1 reached a plateau at 3-10 h after X-ray irradiation at 5 Gy in A-172 and A-172/neo cells. Likewise, the levels of WAF1 in A-172 and A-172/neo cells reached a plateau at 3-10 h and 6-24 h after C-beam (3.0 Gy) and alpha-particle (4.5 Gy) irradiation respectively. The amount of WAF1 increased markedly in a dose-dependent manner 10 h after X-ray, C-beam or alpha-particle irradiation in A-172 and A-172/neo cells but not in T98G or A-172/mp53 cells. In addition, cell survival assay showed that these cell lines were most sensitive to C-beams, less sensitive to alpha-particles and least sensitive to X-rays at 10% survival. There was no difference in sensitivity among these cell lines against C-beam and alpha-particle irradiation whereas wtp53 cells (A-172 and A-172/neo) were more sensitive to X-rays than mp53 cells (A-172/mp53 and T98G). CONCLUSIONS: These results indicate that C-beams and alpha-particles induce p53-dependent WAF1 accumulation as well as is the case with X-rays, suggesting that WAF1 protein accumulation may not contribute to cell killing.

Restoration by glycerol of p53-dependent apoptosis in cells bearing the mutant p53 gene.

PURPOSE: Effective heat-induced cell death in cultured cells bearing a mutant p53 (mp53) gene was sought by glycerol treatment which led to conformational change from mp53 to wild-type p53 (wtp53) in p53-null murine fibroblasts transfected with mp53. MATERIALS AND METHODS: Heat sensitivity was measured using a colony-forming assay. For heat-induced apoptosis, gel electrophoresis was applied to detect DNA fragmentation and Hoechst33342 staining for apoptotic bodies. Glycerol (0.6 M) was applied to the cultured cells 48 h before heating at 44 degrees C in a water bath. RESULTS: Wtp53 transfectants (MT158/wtp53-1) were sensitive to heat stress compared with mp53 transfectants (MT158/mp53-2 cells), and the combined treatment with glycerol enhanced cell killing only in the MT158/mp53-2 cells. After glycerol pretreatment for 48 h, the subsequent heat treatment enhanced DNA fragmentation and apoptosis in MT158/mp53-2 cells, while DNA fragmentation and apoptotic bodies were not enhanced with heat treatment alone in these cells. In contrast, DNA fragmentation or apoptotic bodies were clearly observed in MT158/wtp53-1 cells 3-24h after heat treatment. Treatment with glycerol alone did not induce apoptosis in the transfectants. CONCLUSIONS: Glycerol appears to function as a chemical chaperone that restores mp53 to wtp53 function.

Correlation between gamma-ray-induced G2 arrest and radioresistance in two human cancer cells.

PURPOSE: The correlation between radioresistance and gamma-ray-induced G2 arrest was examined in two human cancer cell lines, HeLa (cervical carcinoma) and MeWo (melanoma). METHODS AND MATERIALS: Cellular radioresistance was examined by a colony formation assay and Hoechst 33342 staining. G2 arrest induced by gamma-rays was examined by flow cytometry, and the accumulation of cyclin B1 and cdc2 proteins was analyzed using Western blotting. RESULTS: HeLa was more resistant (10% survival dose[D10] = 10 Gy) than MeWo (D10 = 4 Gy) to gamma-rays. In HeLa, cell cycle analysis showed that G2 arrest was induced 10 or 24 h after irradiation of 10 or 4 Gy, respectively. In contrast, no clear G2 arrest in MeWo was observed after irradiation. Western blot analysis showed that cell cycle regulators, cyclin B1 and cdc2, were accumulated in HeLa but not in MeWo. The accumulation of cyclin B1 and cdc2 reached peak levels 24-34 h after irradiation of 10 Gy, and 24 h after irradiation of 4 Gy. In addition, Hoechst staining revealed similar increase in apoptotic bodies with time after irradiation in HeLa and MeWo at isosurvival doses. CONCLUSION: Radioresistance of these human cancer cells is closely correlated with gamma-ray-induced G2 arrest, and cyclin B1 and cdc2 are possible regulators of G2 arrest.

Analyses of microsatellite instability and the transforming growth factor-beta receptor type II gene mutation in sporadic breast cancer and their correlation with clinicopathological features.

To determine the incidence of microsatellite instability (MSI) and its relationship with both clinicopathologic parameters and patient survival, 101 cases of breast cancer were investigated. In addition, transforming growth factor-beta (TGF-beta) receptor type II (RII) gene mutation was also examined to clarify the relation to MSI in breast cancer development. MSI and RII gene mutation were screened by single strand conformation polymorphism (SSCP). The mutations of the RII gene were confirmed by a direct sequence. An association between the MSI status and the clinicopathological features was examined to assess the potential of the MSI status as a prognostic indicator in sporadic breast cancer cases. MSI was detected in 12 of 101 (11.9%) breast cancer cases. The positive MSI breast cancer cases showed relatively more advanced disease than negative MSI cases, and also exhibited relatively poorer prognoses. No RII gene mutations were observed in any of the breast cancer cases. Our data suggest that the MSI status may thus be a useful indicator for the prognosis of sporadic breast cancer cases. However, the breast seems to be an infrequent target organ for cancer development through RII gene mutations. As a result, tumor progression through this pathway appears to be related to organ specificity. For positive MSI breast cancers, other target genes therefore still need to be identified.

Pigmented neuroectodermal tumor of infancy in the epididymis. A case report.

BACKGROUND: Pigmented neuroectodermal tumor of infancy (PNTI) is a rare pigmentary tumor. Cytologic study of this tumor had not been performed before. In the present study, cytology of such a tumor was done in addition to histology, electron microscopy and immunohistochemistry. CASE: A 5-month-old infant had a scrotal tumor. After tumorectomy, touch smear for cytology revealed small and large tumor cells. The small cells showed a high nuclear/cytoplasmic ratio, and the large cells contained pigment in their cytoplasm. Histologically, the small cells and large, pigmented cells showed a typical alveolar and tubular pattern, respectively. The small cells neuroblastlike and the large cells showed melanocytelike features ultrastructurally and immunohistochemically. CONCLUSION: Cytology has not been applied to the diagnoses of PNTI. In our opinion, fine needle aspiration cytology could be helpful in the preoperative diagnosis of this tumor.

Hyperthermic enhancement of tumour growth inhibition by accelerated carbon-ions in transplantable human esophageal cancer.

The study examined the effects of combination of hyperthermia (42 degrees C) and 290 MeV/u carbon-ion (C-) beams or 200 kVp X-rays on tumour regrowth delay of transplantable human esophageal cancer as an in vivo model for radiotherapy of cancer. The C-beams were more effective in the tumour growth inhibition than X-rays. The relative biological effectiveness (RBE) of C-beams against X-rays was 2.00. It was observed that the interactive hyperthermic (42 degrees C, for 30 min) enhancement of tumour regrowth delay by high-linear energy transfer (LET) C-beams was similar to that of combination of low-LET X-rays with hyperthermia. The thermal enhancement ratios (TER) were 6.10 and 5.57 for X-rays and C-beams, respectively. These results suggest that hyperthermic treatment is effective in radiotherapy not only by low-LET radiation but also by high-LET radiation such as C-beams. In conclusion, the depression of the tumour growth by the combined treatment of hyperthermia (42 degrees C) and the C-beams strongly suggests the available possible application of interdisciplinary cancer therapy for refractory tumours.

Adhesive property of toxin-coregulated pilus of Vibrio cholerae O1.

The adhesive property of toxin-coregulated pilus (TCP) to the human intestine jejunum), and whether or not TCP mediates the adhesion of Vibrio cholerae 395 organisms to the intestinal epithelium were investigated using visually proving methods. The purified TCP did not agglutinate human erythrocytes nor adhere to the surface of human intestinal epithelium. V. cholerae 395 adhered to the epithelium, but the adhesion was not inhibited by blocking the pili with the Fab fraction of anti-TCP IgG. The organisms adhered to the intestine treated with purified TCP in advance, as well as to the intact intestine. These findings suggest that TCP is not involved in the adhesion of these organisms to the intestinal epithelium.

Effects of accelerated carbon-ions on growth inhibition of transplantable human esophageal cancer in nude mice.

We have studied the effectiveness of 290 MeV/u carbon-ion (C-) beams (linear energy transfer (LET) of 70 keV/mm) and 200 kVp X-rays on tumor growth inhibition as an in vivo model for radiotherapy of cancer. We measured the size of tumor growth of transplantable human esophageal cancer in nude mice after radiation with C-beams and compared this with X-rays as the control. A significant inhibition of tumor growth was observed by C-beams as compared with X-rays. The relative biological effectiveness (RBE) of C-beams against X-rays was 2.02. Histopathological studies showed that C-beams at 20 Gy induced prominent necrosis in the central region and multinucleate giant cells and inflammatory cells in peripheral regions of the tumor, whereas X-rays at 20 Gy induced only mild necrosis. The high RBE of C-beams obtained in this study provides in vivo evidence that C-beams are more effective than conventional X-rays for radiotherapy of cancer.

Cardiac rhabdomyoma of a neonate: an autopsy case report.

A primigravida delivered a cyanosed female infant with a very low Apgar score. Cardiac anomaly of the fetus was detected at 32 weeks of gestation by ultrasonography. The baby died on the day of delivery. Autopsy revealed multiple tumor masses in the interventricular septum and ventricular walls. The tumor originating from the interventricular septum was the largest and measured 3.7 x 3 cm. Histologically, the tumor was composed of large polygonal glycogen-laden cells and 'spider-cells'. Eosinophilic giant histiocytic cells were also observed in the spleen. Ultrastructural features of the tumor cells correlated with those of typical cardiac muscle cells.

An autopsy case of malignant lymphoma with Lyell's syndrome.

We present a case of fatal Lyell's syndrome which developed following a CT examination using omnipaque 3000 contrast medium. A 59-year-old man was suffering from malignant lymphoma. He was readmitted to this hospital due to relapse of fever and lymph node swelling. On the day of readmission, generalized erythema, purpura, and mucosal erosions developed after a CT examination. Steroids and chemotherapy were ineffective, and he expired approximately two weeks after admission. Drug-induced dermatopathy or leukemic cell infiltration in the skin was clinically suspected. Histological findings disclosed toxic epidermal necrolysis.

Characterization of a 50 kDa surface membrane protein on thymic stromal cells as an important factor for early T cell development.

We previously reported that the nude mouse-derived splenic T cell clone, N-9F, exhibits a proliferative response to the SL10.3 thymic epithelial cell clone. In the present study we generated an Armenian hamster mAb, HS9, specific for SL10.3, which inhibited the N-9F's proliferative response to SL10.3. We performed thymocyte repopulation experiments using fetal liver cells and 2'-deoxyguanosine-treated thymic rudiments. After 14 days of culture, donor fetal liver cells proliferated and differentiated to CD4+CD8+ and CD4-CD8+ with some CD4+CD8- cells in the host thymic rudiments. However, most of the thymocytes remained at a CD4-CD8- immature stage in the presence of HS9 and the cell recovery was reduced to 30% of the control. Immunohistostaining and flow cytometry studies revealed that HS9 reacted with stromal cells of fetal thymus at the earliest from day 14 gestation. Neither thymocytes nor lymph node T cells were stained with HS9. HS9 antigen was distributed not only on thymic subcapsular and cortical stromal cells, but also on peripheral B cells in adult mice. The antigen that HS9 detected was found to be a 50 kDa surface membrane protein on thymic stromal cells. On the other hand, the 50 kDa molecule is associated with two other molecules of 80 and 100 kDa on the B cells. These data indicate that the HS9 antigen may have an important role for early T cell development, especially at a stage from CD4-CD8- to CD4+CD8+, and may have some unknown function on B cells.

Effect of 20-methylcholanthrene on the development of atherosclerosis in LAP quails.

We studied the effect of 20-methylcholanthrene, a carcinogen, on atherosclerosis in the ascending aorta and brachiocephalic arteries of hyperlipidemic and atherosclerosis-prone (LAP) quail. A total of 66 quails were divided into 6 groups and fed the following diets: Group I, basal; Group II, basal+low dose of carcinogen; Group III, basal + high dose of carcinogen; Group IV, basal + 0.2% cholesterol; Group V, basal + 0.2% cholesterol + low dose of carcinogen; and Group VI, basal + 0.2% of cholesterol + high dose of carcinogen. The carcinogen was dissolved in corn oil at 2 mg/ml and 4 mg/ml as low and high doses respectively, and was given orally twice weekly. Marked elevation of the serum cholesterol level and significant lipid-rich aortic lesions were observed in all the cholesterol-fed groups after 12 weeks. Although the serum cholesterol level in Group VI was lower than that in Group IV, the severity of the atherosclerotic lesion was greater in the former than in the latter. An immunohistochemical study showed a positive reaction of DBA, PHA and OKM-1 with the lipid-containing cells of aortic intimal lesions.

[Expression of PDGF-A and -B in human coronary atherosclerotic lesion: immunohistochemical and in situ hybridization study].

We studied the role of platelet-derived growth factor (PDGF) on the development of atherosclerosis of human coronary arteries of 63 autopsied cases. Smooth muscle cells in fibrocellular intimal thickening lesion showed no significant immunohistochemical reaction of antibodies for PDGF-A or PDGF-B or PDGF-receptor. In atherosclerotic lesions, foam cells derived from macrophages and smooth muscle cells showed intense immunohistochemical reaction with antibodies of PDGF-B and PDGF-receptor, but not with that of PDGF-A. By in situ hybridization, no significant signals of PDGF-A or PDGF-B or PDGF-receptor were demonstrated in proliferating intimal smooth muscle cells in fibrocellular intimal thickening lesions. Uncomplicated atherosclerotic lesions expressed significant amount of m-RNA of c-sis protooncogene and PDGF-B receptor in foam cells. However, no significant signals of PDGF-A were observed in uncomplicated atherosclerotic lesions. These results suggest that foam cells-producing PDGF-B play an important role for the progression of atherosclerotic lesions in human coronary arteries.