Takashi Nishisaka

Narrow Band Imaging Applied to Pleuroscopy for the Assessment of Vascular Patterns of the Pleura

Background: Narrow band imaging (NBI), which enhances blood vessels, is a new endoscopic technology for diagnosing malignancies, but it has not been investigated for pleuroscopy. Objectives: To evaluate the efficacy of NBI applied to pleuroscopy for detecting malignant lesions by assessing vascular patterns of the pleura. Methods: From May 2006 to September 2008, 45 patients with undiagnosed pleural ef-fusion underwent pleuroscopy using a pleura-videoscope with white light (WL) and NBI under local anesthesia. For this prospective study, 73 biopsy specimens were obtained from sites where images under both WL and NBI were recorded and classified regarding vascular patterns. Results: Of the 73 lesions, WL showed blood vessels in 32 lesions, and NBI in 52 lesions (WL vs. NBI; p = 0.0014). The accuracy, sensitivity and specificity in the detection of irregular vascular patterns, e.g. blood vessels with irregular caliber or punctate vessels indicating malignant lesions, were 60.3, 76.5 and 55.4% in WL, and 80.8, 85.3 and 76.9% in NBI, respectively, resulting in a significant increase in NBI (p = 0.0106 for accuracy and p = 0.0494 for specificity). For flat lesions, NBI revealed a higher accuracy rate (90.6%) in the detection of irregular vascular patterns indicating malignant lesions. Conclusion: Our study demonstrated that NBI applied to pleuroscopy displayed blood vessels significantly better than WL. NBI was useful to detect irregular vascular patterns suggesting malignant lesions, especially for flat lesions. Therefore, NBI was considered useful in the selection of optimal biopsy sites by assessing vascular patterns.

Immunohistochemical staining of Reg IV and claudin-18 is useful in the diagnosis of gastrointestinal signet ring cell carcinoma.

Signet-ring cell carcinoma (SRCC) is a unique subtype of adenocarcinoma that is characterized by abundant intracellular mucin accumulation and a crescent-shaped nucleus displaced toward one end of the cell. Identification of an SRCCs primary site is important for better planning of patient management because the treatment and prognosis differs markedly depending on the origin of the SRCC. In the present study, we analyzed the immunohistochemical characteristics of 94 cases of SRCC, including 21 cases of gastric SRCC, 16 of colorectal SRCC, 10 of breast SRCC, and 47 of pulmonary SRCC, with antibodies against Reg IV and claudin-18, which we previously identified as gastric cancer-related genes. We also tested known markers cytokeratin 7, cytokeratin 20, MUC2, MUC5AC, caudal-related homeobox gene 2 (CDX2), thyroid transcription factor-1, mammaglobin, gross cystic disease fluid protein 15, and estrogen receptor. All 21 cases of gastric SRCC and 16 cases of colorectal SRCC were positive for Reg IV, and the remaining SRCCs were negative. Eighteen of 21 (86%) gastric SRCCs and 6 of 16 (38%) colorectal SRCCs were positive for claudin-18, whereas another SRCCs were negative. In conclusion, Reg IV staining and claudin-18 staining can aid in diagnosis of gastrointestinal SRCC.

Role of Point Mutation of the K‐ras Gene in Tumorigenesis of B6C3F1 Mouse Lung Lesions Induced by Urethane

In order to elucidate the role of point mutation of the K‐ras gene in the tumorigenetic process of lung tumors, an experimental model of lung lesions in mice induced by the administration of urethane was used. A total of 135 B6C3F1 male mice, 6 weeks old, were given urethane in the drinking water at 0, 6, 60, 600 or 1200 ppm, and were then killed after varying periods of time. The lung lesions were histologically characterized as hyperplasia, adenoma and adenocarcinoma. Point mutations in codons 12 and 61 of the K‐ras gene were detected by polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) and confirmed by using dideoxy sequencing analysis. K‐ras gene mutation was identified in 9 (23.7%) of 38 lesions classified as hyperplasia, 31 (46.3%) of 67 adenomas, and 3 (50%) of 6 adenocarcinomas. The most frequent mutation was an AT‐to‐TA transversion at the second base of codon 61 and this pattern accounted for 65% of the three mutant forms observed. These results suggest that the point mutation of K‐ras gene is involved in all stages of mouse lung tumorigenesis, i.e., activation of this gene can also influence the later stages of lung lesions.

Heterogeneity of expression of cytokeratin subtypes in squamous cell carcinoma of the lung: With special reference to CK14 overexpression in cancer of high-proliferative and lymphogenous metastatic potential

In surgically resected specimens of squamous cell carcinoma (SCC) of the lung from 45 patients, we immunohistochemically examined the expression of 13 subtypes of cytokeratin (CK), the intermediate filament in cytoplasm of epithelial cells. To investigate heterogeneity of CK, its expression was compared among tumor cell nests with or without keratinization and stratification. Furthermore, the relationship between CK expression and Ki‐67 labeling index or p53 expression was investigated.

Clinical significance of sIL-2R levels in B-cell lymphomas.

Elevated soluble interleukin-2 receptor (sIL-2R) in sera is observed in patients with malignant lymphoma (ML). Therefore, sIL-2R is commonly used as a diagnostic and prognostic marker for ML, but the mechanisms responsible for the increase in sIL-2R levels in patients with B-cell lymphomas have not yet been elucidated. We first hypothesized that lymphoma cells expressing IL-2R and some proteinases such as matrix metalloproteinases (MMPs) in the tumor microenvironment can give rise to increased sIL-2R in sera. However, flow cytometric studies revealed that few lymphoma cells expressed IL-2R α chain (CD25) in diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL), and most CD25-expressing cells in the tumor were T-cells. Distinct correlations between CD25 expression on B-lymphoma cells and sIL-2R levels were not observed. We then confirmed that MMP-9 plays an important role in producing sIL-2R in functional studies. Immunohistochemical (IHC) analysis also revealed that MMP-9 is mainly derived from tumor-associated macrophages (TAMs). We therefore evaluated the number of CD68 and CD163 positive macrophages in the tumor microenvironment using IHC analysis. A positive correlation between the levels of sIL-2R in sera and the numbers of CD68 positive macrophages in the tumor microenvironment was confirmed in FL and extranodal DLBCL. These results may be useful in understanding the pathophysiology of B-cell lymphomas.

p16/CDKN2 Gene and p53 Gene Alterations in Japanese Non‐smoking Female Lung Adenocarcinoma

Primary lung adenocarcinomas in non‐smoking females are increasing in the USA and Japan. Environmental factors such as passive smoking, asbestos, domestic radon, and hormonal effects have been implicated, but the etiology is still uncertain. We therefore analyzed point mutations of p16 gene, a newly characterized tumor suppressor gene, and compared the results with alterations of p53 gene in 28 primary lung adenocarcinomas in non‐smoking Japanese females. There were no cases with somatic point mutation of p16 gene, except for one case with two germline mutations (silent mutations). In contrast, six out of 16 informative cases showed loss of heterozygosity of p53 gene using a TP53 microsatellite marker and 19 out of 28 cases showed expression of oncoprotein using DO‐7 immunohistochemistry. These findings suggest that p16 gene alteration is a rare event in primary lung adenocarcinomas in Japanese non‐smoking females, compared with alterations of the p53 gene.

Characteristic gene expression in stromal cells of gastric cancers among atomic‐bomb survivors

To elucidate the mechanism of radiation‐induced cancers, molecular analysis of cancers in atomic‐bomb survivors is important. In our study, we developed a custom oligonucleotide array of 208 genes. We analyzed gene expression profiles of gastric cancers (GCs) from atomic‐bomb survivors and identified 9 genes with significantly lower expression in GCs from exposed patients than in GCs from nonexposed patients. Among these 9 genes, expression of versican and osteonectin was investigated in greater detail using immunohistochemistry in 116 GCs from 64 exposed and 52 nonexposed patients who developed GC after the bombing. In the Stage I/II GCs, the clinicopathologic, phenotypic and proliferative characteristics of GCs from exposed and nonexposed patients did not differ significantly; however, versican and osteonectin were expressed at much lower levels in the area of tumor‐associated stroma of exposed patients than in nonexposed patients (p = 0.026 and p = 0.024, respectively). These results suggest that the characteristics of tumor‐associated stromal cells differ between GCs from exposed and nonexposed patients.

Primary angiitis of the central nervous system mimicking dissemination from brainstem neoplasm: a case report

BACKGROUND Diagnosis of PACNS is very difficult because of the variety of clinical manifestation, especially neurologic and MRI findings. We report the case of a 68-year-old woman with PACNS. CASE DESCRIPTION Her clinical symptoms mimicked malignant brainstem neoplasm disseminated to third and fourth ventricles and cervical spinal cord. Brain biopsy could not be correctly diagnostic. In spite of a trial of high-dose steroids, repeated MRI showed an increasing number of lesions. Two weeks after surgery, she died of respiratory dysfunction. Postmortem examination suggested PACNS, after consideration of histories, laboratory and radiologic findings, and the lack of systemic disease. CONCLUSION Although brain biopsy represents the gold standard measure, combination with several other examinations might be necessary to arrive at an early and definitive diagnosis. PACNS should be considered the differential diagnosis of intracranial tumors.

IgG4-related pleural disease presenting as a massive bilateral effusion.

A 74-year-old woman with massive bilateral pleural effusion, which was exudative in nature, and with mononuclear cell predominance underwent a pleuroscopy. Parietal pleura were thickened and partly reddish in color. Biopsy specimens taken from the parietal pleura revealed lymphoplasmacytic inflammation with fibrosis. As her performance status rapidly worsened with thoracentesis, we performed bilateral pleurodesis using talc. Pathologic evaluation of the pleural biopsy specimen with immunohistochemical staining revealed 91 IgG4-positive plasma cells per high-power field and an IgG4/IgG ratio of 91%. Thus, the diagnosis of pleuritis from IgG4-related disease was established. Our case suggests that IgG4-related disease is one of the causes of pleural effusion, and it should be included in the differential diagnosis of unexplained pleuritis.

Numerous conglomerate inclusions in slowly progressive familial amyotrophic lateral sclerosis with posterior column involvement

A 59-year-old woman with slow progression of the loss of motor function and predominant lower motor manifestation during a 14-year period showed familial amyotrophic lateral sclerosis (fALS) with posterior column involvement, neuropathologically. Conglomerate inclusions (CIs) were observed in the remaining neurons in various areas, including the spinal anterior horn, posterior horn, Clarks column, accessory cuneate nucleus, tegmental reticular formation, motor nucleus of the trigeminal nerve, nucleus of the facial nerve, hypoglossal nucleus, medial nucleus of the thalamus, dentate nucleus, and motor cortex (Betz cells). Immunohistochemically, it was newly identified that the CIs showed marked immunoreactions with antibodies to phosphorylated and non-phosphorylated neurofilaments and to 64, 120, and 200 kD neurofilaments. The CIs were partially immunoreactive with the anti-ubiquitin antibody, although they reacted only weakly (or not at all) with anti-Cu/Zn superoxide dismutase (SOD1) antibody. Ultrastructurally, the CIs were comprised of neurofilaments. These data suggest that this case might have been different from an example of fALS with Ile 113 Thr mutation in the SOD1 gene.