Shotaro Sato

Strong correlation between the number of CAG repeats in androgen receptor genes and the clinical onset of features of spinal and bulbar muscular atrophy

X-linked spinal and bulbar muscular atrophy (SBMA), a motor neuron disease associated with androgen insensitivity, is caused by androgen receptor gene mutations with an increased number of tandem CAG repeats in exon 1. We investigated the increased number of CAG repeats in androgen receptor genes of 19 SBMA patients and found that this correlated strongly with the age at onset of muscle weakness. Thus, SBMA is the first genetic disease in which a strong correlation between the degree of genetic abnormality (number of CAG tandem repeats) and clinical phenotypic expression is demonstrable. The results further indicate that androgen gene mutation is directly involved in the degeneration of motor neurons.

Immunohistochemical Detection of Intercellular Adhesion Molecule-1 (ICAM-1) and Major Histocompatibility Complex Class I Antigens in Seminoma

Expression of intercellular adhesion molecule-1 (ICAM-1) and major histocompatibility complex (MHC) antigens, and characterization of tumor-infiltrating mononuclear cells (TIM) were examined immunohistologically in 10 specimens of seminoma. ICAM-1 and MHC antigens were not detected on normal spermatogenic cells. ICAM-1 and MHC class I antigens were variably expressed in 7 and 9 seminomas, respectively, whereas class II antigens were not detected. Although the degree of expression of ICAM-1 and MHC antigens was not correlated with any clinical or histopathological factors, neither of the antigens was detected on an anaplastic seminoma. Various numbers of TIM were detected in all of the seminoma, and comprised mainly T cells bearing the lymphocyte function-associated antigen (LFA)-1. No significant correlation was noticed between the degree of lymphocyte infiltration and ICAM-1 or MHC antigen expression. Although ICAM-1 and MHC class I antigens were expressed in seminoma, possibly facilitating an anti-tumor reaction of host, their expression remained low in several cases, despite marked lymphocyte infiltration within the tumor.

Correlation of p53 protein expression in human urothelial transitional cell cancers with malignant potential and patient survival.

The p53 gene product has been detected frequently in various human malignancies. We have studied the expression of p53 protein in urothelial transitional cell cancers (TCCs) and examined its correlation with pathologic grade, stage(pT) and patient survival. Specimens from 69 surgically‐resected TCCs (38 cases of urinary bladder cancer, 17 cases of ureteral cancer and 14 cases of renal pelvic cancer) were examined by immunohistochemical staining, using two anti‐p53 monoclonal antibodies, PAb1801 and PAb240, and a polyclonal antibody, CM‐1. Twenty‐six TCCs (37.6%) were positively stained by at least one of the three antibodies. Statistical analysis showed a significant correlation between p53 expression and high pathologic grade (p less than 0.05, p less than 0.001) or progressive pathologic stage (p less than 0.01). In addition, in 51 of the patients who were available for follow‐up (23 cases of urinary bladder cancer, 13 cases of ureteral cancer, and 15 cases of renal pelvic cancer), the correlation between p53 protein expression and prognosis was examined. The survival of patients exhibiting positive p53 protein expression was significantly worse than those with p53‐negative tumors (p less than 0.05). These results suggest that an immunohistochemical test for p53 protein may be a useful method of evaluating the malignant potential of TCCs. Additionally, expression of p53 protein in TCCs is an indicator of a poor prognosis which should be considered in drawing up treatment strategies.

mRNA and protein expression of p53 mutations in human bladder cancer cell lines

We investigated mRNA and protein expression in p53 gene mutations in four human bladder cancer cell lines using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and Northern blot and Western blot analyses. The following mutations were identified in three of the four cell lines: a missense transversion at codon 110, a missense transition at codon 250 and a non-sense transversion at codon 126. These mutations were located outside previously identified hot spot codons and have rarely been reported in bladder cancer tissues or other neoplasms. Positive intranuclear p53 immunostaining in neoplastic cells in the two missense mutations and the premature stop codon in the non-sense mutation suggested the presence of structural and functional alterations in the p53 protein. Northern and Western blot analyses revealed either an intense or a weak p53 mRNA band together with an intense p53 protein band in the missense mutations, but no p53 mRNA or protein band in the non-sense mutation. A weak p53 mRNA band, but no distinct p53 protein band was observed in the cell line without a mutation and in normal control bladder cells. Our findings suggest that regulation of p53 expression in these cell lines differs at the post-transcriptional and/or post-translational level between the wildtype and the mutant p53 genes and also among different mutant p53 genes. The three cell lines with mutations were derived from high-grade carcinomas; the cell line without mutation was derived from a low-grade carcinoma.

Radioresistance of Intermediate TCR Cells and Their Localization in the Body of Mice Revealed by Irradiation

Extrathymic generation of T cells in the liver and in the intestine was recently demonstrated. We investigated herein whether such T cells, especially those in the liver, are present in other organs of mice. This investigation is possible employing our recently introduced method with which even a minor proportion of extrathymic, intermediate TCR cells in organs other than the liver can be identified. Intermediate TCR cells expressed higher levels of IL‐2Rβ and LFA‐1 than bright TCR cells (i.e., T cells of thymic origin) as revealed by two‐color staining. Although intermediate TCR cells were present at a small proportion in the spleen and thymus, they predominated in these organs after irradiation (9 Gy) and bone marrow reconstitution, or after low dose irradiation (6 Gy). This was due to that intermediate TCR cells were relatively radioresistant, whereas bright TCR cells were radiosensitive. Microscopic observation and immunochemical staining showed that intermediate TCR cells in the spleen localized in the red pulp and those in the thymus localized in the medulla. These intermediate TCR cells displayed a large light scatter, similar to such cells in the liver. The present results suggest that intermediate TCR cells may proliferate at multiple sites in the body.

Calcitonin gene-related peptide (CGRP)-immunoreactive nerve plexuses in the renal pelvis and ureter of rats

SummaryThe distribution of calcitonin gene-related peptide-immunoreactive nerve fibers in the renal pelvis and ureter was examined by immunohistochemistry using whole-mount preparations and cryostat sections. The patterns of innervation were contrasted between the pelvis and ureter; the immunoreactive nerve fibers in the pelvis ran parallel to the long axis of each of the circular and longitudinal muscle layers, causing a lattice-like appearance of the nerve fibers. In the ureter, the immunoreactive fibers were accumulated in the subepithelial region and the longitudinal muscle. In both the pelvis and ureter, a portion of the nerve fibers of smaller caliber showed a swollen or beaded structure; they were located in the musculature and beneath the epithelium extending for considerable distances. Ligation of the ureter caused a marked decrease in the immunoreactive nerves in the pelvis and the proximal portion of the ureter, suggesting that the axonal flow in the calcitonin gene-related peptide-containing neurons of the ureter runs towards the pelvis.

MHC class II antigen-associated invariant chain on renal cell cancer may contribute to the anti-tumor immune response of the host

To investigate the association between renal cell cancer (RCC) and the host immune system, we examined the expression of invariant chain (Ii) and HLA-DR on renal cancer. Immunohistochemically, Ii was detected in 53 of the 60 cases of RCC. Significant correlation was found between the expression of Ii and the degree of lymphocyte infiltration. Flow cytometric analysis for HLA-DR and Ii on RCC cell line (ACHN) showed no positive cells, whereas interferon (IFN)-gamma treatment induced HLA-DR. Immunoprecipitation showed the presence of cytoplasmic Ii in ACHN cells. In addition, IFN-gamma-treated ACHN cells showed more intense signals than untreated cells. These results suggest that Ii associated with class II antigens on RCC may contribute to the anti-tumor immune response of the host and that IFN-gamma, which is administered for the treatment of cancer, may increase the immunogenicity of RCC.

Value of dimercaptosuccinic acid single photon emission computed tomography and magnetic resonance imaging in detecting renal injury in pediatric patients with vesicoureteral reflux : comparison with dimercaptosuccinic acid planar scintigraphy and intravenous pyelography

The value of 99mTc-dimercaptosuccinic acid (DMSA) planar renal scintigraphy, DMSA single photon emission computed tomography (SPECT), and magnetic resonance imaging (MRI) in the assessment of renal injury related to vesicoureteral reflux (VUR) was examined in 60 kidneys of 32 pediatric patients (28 bilateral, 4 unilateral) with primary VUR. The results were: (1) detection of minor renal lesions was best with MRI, then DMSA-planar and DMSA-SPECT, and (2) in comparing the positive rate, DMSA-SPECT (85%) and MRI (83.3%) were superior to intravenous pyelography (55%) and DMSA-planar scintigraphy (65%). These results suggest that DMSA-SPECT or MRI may be more sensitive than DMSA-planar scintigraphy and intravenous pyelography in detecting renal injury related to VUR in pediatric patients.

Interferon γ but not tumor necrosis factor α decreases susceptibility of human renal cell cancer cell lines to lymphokine-activated killer cells

SummaryHuman renal cell cancer (RCC) cell lines, ACHN and KRC/Y, with or without exposure to cytokines, were examined for their susceptibility to lymphokine-activated killer (LAK) cells. Flow-cytometric analysis demonstrated constitutional expression of class I antigen on both cell lines, which was enhanced by interferon α (IFNα), IFNγ and tumor necrosis factor α (TNFα). A 4-h51Cr-release cytotoxicity assay demonstrated that pretreatment of both cell lines with IFNγ or IFNα, but not with TNFα, decreased their susceptibility to LAK cells. IFNγ also decreased susceptibility to natural killer cells in a 16-h51Cr-release cytotoxicity assay. IFNγ treatment decreased the susceptibility of ACHN cells in a dose-dependent manner. “Cold”-target competition assay clearly showed that IFNγ- but not TNFα-pretreated cells compete less effectively than do untreated target cells. Pretreatment with IFNγ, however, increased expression of intercellular adhesion molecule-1 (ICAM-1) to a degree comparable to that with TNFα. Northern blot analyses using a 520-base-pair ICAM-1 cDNA as a probe demonstrated that more 3.3-kb mRNA is expressed in IFNγ- and TNFα-pretreated cells. These results suggest that IFNγ-treated RCC cell lines may reduce their ability to be recognized by LAK cells, and that IFN-induced protection of RCC cell lines against LAK cells may depend upon a mechanism independent of the expression of class I antigens or ICAM-1 on tumor cells.

Primary Localized Amyloidosis of the Bladder: A Case of AL (λ) Amyloid Protein and Combination Therapy Using Dimethyl Sulfoxide and Cepharanthin

We report a case of primary localized amyloidosis of the bladder with amyloid deposits which was characterized as being of immunoglobulin light chain origin (AL) including lambda type (A lambda) and P component (AP) using the KMnO4 pretreatment method and immunohistochemical procedures. The patient was treated successfully with intravesical dimethyl sulfoxide instillation and oral administration of high-dose cepharanthin after transurethral resection. Combination therapy with dimethyl sulfoxide and cepharanthin was shown to be useful for primary localized amyloidosis of the bladder.