Shin Wakui

Localization of Ang-1, -2, Tie-2, and VEGF expression at endothelial-pericyte interdigitation in rat angiogenesis.

Endothelial cells and pericytes play critical role in angiogenesis, which is controlled, in part, by the angiopoietin (Ang)/Tie-2 system and vascular endothelial growth factor (VEGF). Here, we investigated Ang, Tie-2, and VEGF expression within endothelial cells and pericyte interdigitations (EPI), which consist of cytoplasmic projections of pericytes and corresponding endothelial indentations. After subcutaneous implantation of a thermoreversible gelation polymer disc in rats, the capillary density was low on day 5, increased to a peak on day 7, and then decreased on days 10–20. A small number of EPI were observed on day 5, then increased sharply to a peak on day 10, but had decreased on day 20. Light and electron microscopy immunohistochemical and RNA in situ hybridization analyses revealed that Tie-2 localized at endothelial cells, and Ang-2 localized at endothelial cells and pericytes, while Ang-1 and VEGF localized at pericytes, and Ang-1 was most intensely observed at EPI of pericytes. Conventional quantitative RT-PCR and Western blot analyses revealed that the level of Ang-1 was low on days 5–7, then increased on days 10–20, while the level of VEGF was high on days 5–10, but had decreased on day 20. The level of Ang-2 remained high and Tie-2 remained at the level of the control on days 5–20. The present study showed that the angiogenic phase might be initiated by increases in Ang-2 and VEGF, while the microvessel maturation phase might be initiated by a relative increase in Ang-1 and a decrease in VEGF. Moreover, EPI might serve as a pathway for the Ang-1/Tie-2 system, with VEGF promoting pericyte recruitment for microvascular integrity.

Two- and three-dimensional ultrastructural observation of two cell angiogenesis in human granulation tissue

SummaryThe growth of endothelial sprouts from capillary walls in human granulation tissue has been examined by two-and three-dimensional electron microscopy using a serial sectioning method. Within the parent capillary wall endothelial sprouts composed of two layers of relatively immature endothelium was demonstrated. Three-dimensional reconstruction revealed that the two layered endothelial projections extended and/or migrated outward in a bicellular configuration, the slit-like lumen of the endothelial sprout connecting with the parent capillary lumen. These ultrastructural appearances have not been reported previously with sequential composition to the morphological progression of the sprout. In the cytoplasm of the endothelial sprout, abundant intermediate filaments were assumed to play a mechanical role, tension resistance, in the development of the endothelial sprout. The active endothelial sprout in granulation tissue was considered to be at least partially responsible for the growth of the capillary network and subsequent development of granulation tissue.

p53 gene mutations occurring in spontaneous benign and malignant mammary tumors of the dog.

Sixty-three cases of benign and malignant canine mammary tumors were analyzed to define the alteration of exons 5–8 for the p53 tumor suppressor gene using polymerase chain reaction direct sequence analysis with paraffin-embedded tissues. Four missense mutations were found in 38 benign mammary tumors (11%), and five missense (one tumor had two missense mutations) and one nonsense mutations were found in 25 mammary carcinomas (20%). These data suggest that the p53 gene alterations might be initiated at an early stage of canine mammary carcinogenesis and p53 mutations might be associated with malignancy. However, there was no evidence of any relationship between the p53 alterations and the histologic types of tumors or breeds of dogs.

Epidermal growth factor receptor at endothelial cell and pericyte interdigitation in human granulation tissue

Angiogenic immature capillaries in human granulation tissue possess many cytoplasmic interdigitations between endothelial cells and pericytes (CIDEP). Epidermal growth factor (EGF) is a potent mitogenic polypeptide which accelerates angiogenesis in vivo and in vitro. We have recently demonstrated in immature capillaries that EGF is present at the CIDEP and proposed that the CIDEP may be involved in a signaling pathway for EGF (Wakui et al., 1990c. Microvasc. Res. 40, 285-291). This study follows that previous report. In the present study, I have investigated the ultrastructural localization of the EGF receptor (EGF-r) at CIDEP. Immunoreactivity for the EGF-r in immature capillaries was located strictly at the CIDEP in large numbers, at the coated pits and vesicles of endothelial cells, and at some lysosome-like structures of the endothelial cells and the pericytes. On the other hand, immunoreactivity for EGF-r was absent in mature capillaries. At the CIDEP, EGF-r immunoreactivity was present at the cell membrane and a few cytoplasmic elements at the tip of pericyte cytoplasmic projections, but it was completely absent at the corresponding endothelial membrane indentation. The present results support our proposed hypothesis that the CIDEP in immature capillaries act as a pathway for EGF transportation from the endothelial cell to the pericyte by a receptor-mediated process.

Ser217Leu polymorphism of the HPC2/ELAC2 gene associated with prostatic cancer risk in Japanese men

The HPC2/ELAC2 gene may be associated with hereditary/familial prostate cancer (PCa). Two common missense variants (Ser217Leu and Ala541Thr) have been reported in the gene. We performed mutational, allelotyping and expression analyses and a molecular epidemiological study to clarify the relations between this gene and prostatic diseases, including PCa and benign prostatic hyperplasia (BPH) in Japanese men. We screened for mutations in 109 patients with PCa including 11 patients from 1 hereditary and 9 familial PCa. Loss of heterozygosity and expression were analyzed. An epidemiological study was done in sporadic PCa (n=98) and BPH (n=143) using 1 novel (Ser627Leu) and 2 previously described polymorphisms of the HPC2/ELAC2 gene. Somatic or germline mutations were not confirmed in any cases of PCa. Loss of heterozygosity at 2 microsatellites, D17S1289 and D17S520, was detected in 1 of 38 and 1 of 35 cases, respectively. Expression analysis revealed decreased or absent mRNA expression in 6 of 38 tumors. Epidemiologic analysis showed that a Leu allele at codon 217 was significantly more frequent in patients with PCa than in controls (10.2% vs. 3.5%, odds ratio = 3.11; 95% confidence interval, 1.22–7.90). At codon 541, all patients with PCa or BPH and all control subjects had the Ala/Ala genotype. At codon 627, the incidence of the Leu variant was slightly, but not significantly, higher in patients with BPH than in controls (7.0% vs. 2.8%, odds ratio = 2.59, 95% confidence interval, 0.94–7.13, not statistically significant). We concluded that germline/somatic mutations of HPC2/ELAC2 are uncommon in PCa. Similarly, allelic imbalances at the gene locus and changes in expression are rare. Although no difference in allele frequency at Ser217Leu between patients with PCa and controls has been reported in a Western population, this polymorphism is a potential indicator of PCa risk in Japanese men and it should be examined in other ethnic groups.

Differences in prostate cancer grade, stage, and location in radical prostatectomy specimens from United States and Japan

To compared prostate cancer at radical prostatectomy between men in the United States (US) and Japan in the modern era.

p16 (CDKN2/MTS1) Gene Deletions are Rare in Prostatic Carcinomas in the United States and Japan

PURPOSE The incidence of clinically apparent prostatic carcinoma is much higher in the United States than in Japan. Alterations in the p16 tumor suppressor gene have been identified in various tumor types, including cultured prostatic carcinoma cell lines. We studied the possible deletions of either exon 2 or 3 of this gene in primary clinical prostatic carcinomas from Japan and the United States. MATERIALS AND METHODS Genomic DNA was extracted from 36 formalin-fixed, paraffin-embedded clinical prostatic carcinomas from Japan and 27 carcinomas from the United States. Exons 2 and 3 of the p16 gene were amplified using comparative multiplex polymerase chain reactions (PCR) and then analyzed for possible deletions of either exon. RESULTS Two out of 36 (5.6%) carcinomas from Japan clearly demonstrated deletion of p16 exon 2, but this deletion was not detected in any of the 27 carcinomas from the United States. CONCLUSIONS Although slightly higher in Japan than in the United States, the frequency of p16 exon deletions in clinical prostatic carcinomas is very low, and probably is not important in the development of this neoplasm.

Development of the Bovine Ileal Mucosa

The development of the bovine ileal mucosa was studied with particular reference to maturation during the fetal and neonatal period. In this region, by 4-5 months of fetal development, vacuolation of the epithelial cells had occurred on the villi, and the goblet and absorptive cells in the crypts were present. By 6-9 months, the villi were longer and more numerous than in the previous stages. At the same time, the vacuolated cells could be seen predominantly on the upper half of each villus. The absorptive cells and goblet cells were more distinct in the crypt and lower half of each villus. Moreover, the goblet cells showed differences in mucin, while in the submucosa the lymphoid follicles were seen to have enlarged to become a prominent feature of the Peyers patches at this stage. At birth, in suckled animals, the ileal cells on the lower area of each villus and in the crypt appeared more like mature cells. In contrast, there were numerous inclusion bodies in epithelial cells on the upper half of each villus. They appeared in the apical portion of the cytoplasm as vacuoles with stainable or dense contents. By 1 week, however, epithelial cells no longer contained inclusion bodies, and absorptive and goblet cell populations had begun to emerge from the crypts. These histological results suggest that the bovine ileal mucosa has two distinct turning points during its development in the fetus and the neonate. Initially all the mucosal structures are present in fetuses at 6-7 months of gestation, and then the vacuolated cells covering the ileal villi are replaced by mature, nonpinocytosing epithelium which emerges from the crypts on or before the 7th day after birth (ileal closure).

Atypical Leydig Cell Hyperplasia in Adult Rats with Low T and High LH Induced by Prenatal Di(n-butyl) Phthalate Exposure

The present study describes atypical Leydig cell (LC) hyperplasia in 20-week-old Sprague-Dawley rats with low testosterone and high luteinizing hormone levels after prenatal administration of 100 mg/kg/day di(n-butyl) phthalate on days 12 to 21 postconception. Light microscopy revealed LC hyperplasia surrounded by severely degenerated seminiferous tubules. Aggregated LCs had large ovoid nuclei with nucleoli and abundant eosinophilic cytoplasm. Immunohistochemical analysis showed expression of proliferating cell nuclear antigen and vimentin in many hyperplastic LCs. Electron microscopy revealed atypical nuclei, abundant free ribosomes, stripped rough endoplasmic reticulum, intermediate-size filaments, elongated cytoplasmic filopodia, atypical tight junctions, and cilia formations, but smooth endoplasmic reticulum was scarcely observed.

Testicular Epidermoid Cyst and Penile Squamous Cell Carcinoma in a Dog

report is also of interest because of lymphatic and blood vessel invasion and pulmonary metastasis. The squamous cell carcinoma component of these two malignant thymomas arose from neoplastic thymic epithelial cells that had formed tubules and cysts. The malignant cells arose from thymic epithelium that was part of the tumor and not from epithelia of branchial cysts of the mediastinum or from the commonly acquired nonneoplastic cysts of thymic involution.