Immunohistochemical detection of imidazolone, a novel advanced glycation end product, in kidneys and aortas of diabetic patients.

To investigate the role of the Maillard reaction in the pathogenesis of diabetic complications, we produced several clones of monoclonal antibodies against advanced glycation end products (AGEs) by immunizing mice with AGE-modified keyhole limpet hemocyanin, and found that one clone (AG-1) of the anti-AGE antibodies reacted specifically with imidazolones A and B, novel AGEs. Thus, the imidazolones, which are the reaction products of the guanidino group of arginine with 3-deoxyglucosone (3-DG), a reactive intermediate of the Maillard reaction, were found to be common epitopes of AGE-modified proteins produced in vitro. We determined the erythrocyte levels of imidazolone in diabetic patients using ELISA with the monoclonal anti-imidazolone antibody. The imidazolone levels in the erythrocytes of diabetic patients were found to be significantly increased as compared with those of healthy subjects. Then we studied the localization of imidazolone in the kidneys and aortas obtained from diabetic patients by immunohistochemistry using the antibody. Specific imidazolone immunoreactivity was detected in nodular lesions and expanded mesangial matrix of glomeruli, and renal arteries in an advanced stage of diabetic nephropathy, as well as in atherosclerotic lesions of aortas. This study first demonstrates the localization of imidazolone in the characteristic lesions of diabetic nephropathy and atherosclerosis. These results, taken together with a recent demonstration of increased serum 3-DG levels in diabetes, strongly suggest that imidazolone produced by 3-DG may contribute to the progression of long-term diabetic complications such as nephropathy and atherosclerosis.     

Two cases of HTLV-I associated myelopathy (HAM) after cardiac operation

Sixty-one and 51-year-old males had progressive walk disturbance after cardiac operations at the intervals of 1 and 2 years. They had received blood transfusion at their operations. Physical examination revealed spastic paraparesis, sensory disturbance of the lower and rectobladder disorder. High titers of anti-Human T-lymphotrophic Virus type-I (HTLV-I) antibody were found in sera and CNF in both cases. They were diagnosed as HTLV-I associated myelopathy (HAM). Because they lived in Aichi Prefecture where the virus is non-epidemic, they were possibly infected through the blood transfusion at their operations. For prevention of HAM, the anti-HTLV-I antibody of all donor blood should be checked before transfusion.     

Extraosseous metastases of hepatocellular carcinoma detection and therapeutic assessment with Tc-99m PMT SPECT

Owing to recent advances in imaging technology and radiologic intervention, survival rates in patients with hepatocellular carcinoma have improved markedly. However, such prolonged survival has resulted in an increase in extrahepatic metastases. Tc-99m (Sn)-N-pyridoxyl-5-methyltryptophan (Tc-99m PMT), developed for hepatobiliary scintigraphy, has been used to visualize extrahepatic metastases, with most related reports limited to osseous metastases. The authors report two cases of hepatocellular cancer presenting as a hypopharyngeal metastasis and intraperitoneal dissemination along the tract of a fine-needle biopsy. Lesions undetectable on planar imaging could be visualized by Tc-99m PMT SPECT.     

Inhibitory effect of murine kidney extracts on the proliferation of murine mast cells

We examined the effect of murine kidney extract (MKE) on the clonal growth of mast cells from murine peritoneal cells. Adding MKE resulted in a 40% inhibition of colony formation of mast cells in a methylcellulose culture, and a 90% decrease in mast cell numbers and histamine content in mast cells in a liquid culture containing stem cell factor and interleukin-3. The mast cell inhibitory factors in MKE were heat sensitive proteins of approximately 560 and 24 kDa. These results suggest that MKE contains regulators that suppress the growth of murine mast cells and histamine synthesis.     

Novel Ultrasonographic Scoring System of Sinusoidal Obstruction Syndrome after Hematopoietic Stem Cell Transplantation

Sinusoidal obstruction syndrome (SOS)/hepatic veno-occlusive disease (VOD) is a well-documented complication after hematopoietic stem cell transplantation (HSCT). Transabdominal ultrasonography (US) enables the visualization of blood flow abnormalities and is therefore useful for the diagnosis of SOS/VOD. We herein prospectively evaluated accuracy of a novel US diagnostic scoring system of SOS/VOD based on US findings. We carried out US in 106 patients on day 14 and when SOS/VOD was suspected after allogeneic HSCT. Among 106 patients, 10 patients (9.4%) were diagnosed as SOS/VOD by Baltimore or Seattle criteria. According to univariate analysis of 17 US findings (US-17 screening), we established a novel scoring system (HokUS-10) consisting of 10 parameters, such as gallbladder wall thickening, ascites, and blood flow signal in the paraumbilical vein. The sensitivity and specificity were 100% and 95.8%, respectively. Diagnostic performance of the HokUS-10 was significantly better than US-17 screening. In 4 of 10 patients US detection of SOS/VOD preceded to clinical diagnosis. The HokUS-10 scoring system is useful in the diagnosis of SOS/VOD; however, our results should be validated in other cohorts.     

Effects of amifostine on the proliferation and differentiation of megakaryocytic progenitor cells

This study investigated the effects of amifostine, a clinically usable radioprotector or chemoprotector, on the proliferation and differentiation of normal and X-irradiated cluster of differentiation 34 positive (CD34+) megakaryocytic progenitor cells (colony-forming unit in megakaryocytes, CFU-Meg) from human placental and umbilical cord blood (CB) in vitro. Amifostine significantly accelerated megakaryocyte colony formation in a plasma clot culture supplemented with recombinant human thrombopoietin because of an increase in immature CFU-Meg-derived large megakaryocyte colony formation. An analysis of the cells that were harvested from the culture showed that amifostine induced a 70- and an 83-fold increase in the total cell and CFU-Meg numbers, respectively, and produced hyperploid megakaryocytes of more than 8 N ploidy. The radioprotective effect of amifostine on the clonal growth of X-irradiated CD34+ CFU-Meg was observed by treatment before or after irradiation. These findings suggest that the action of amifostine extends from immature CFU-Meg to the terminal differentiation of megakaryopoiesis, and its radioprotective effect is shown in megakaryopoiesis and thrombopoiesis.     

Induction of vasculogenesis in breast cancer models

Recently, there have been reports of postnatal vasculogenesis in cases of ischaemia models. The aim of the present study is to provide evidence of postnatal vasculogenesis in breast-cancer-bearing mice. Based on cell surface antigen expression, we isolated endothelial precursor cells from bone marrow, peripheral blood and tumour-infiltrating cells from mice that had received six human breast cancer xenografts. In all three areas (bone marrow, peripheral blood and tumour-infiltrating cells), endothelial precursor cell population was elevated in all transplanted mice. Differentiation and migration activities of endothelial precursor cells were measured by comparing levels of the endothelial precursor cell maturation markers Flk-1, Flt-1, Tie2, VE-cadherin and CD31 among these three areas. The endothelial precursor cell population was 14% or greater in the gated lymphocyte-size fraction of the inflammatory breast cancer xenograft named WIBC-9, which exhibits a hypervascular structure and de novo formation of vascular channels, namely vasculogenic mimicry (Shirakawa et al, 2001). In vitro, bone marrow-derived endothelial precursor cells from four human breast cancer xenografts proliferated and formed multiple clusters of spindle-shaped attaching cells on a vitronectin-coated dish. The attaching cells, which incorporated DiI-labelled acetylated low-density lipoprotein (DiI-acLDL) and were negative for Mac-1. The putative bone marrow derived endothelial precursor cell subset, which was double positive of CD34 and Flk-1, and comparative bone marrow derived CD34 positive with Flk-1 negative subset were cultured. The former subset incorporated DiI-acLDL and were integrated with HUVECs. Furthermore, they demonstrated significantly higher levels of murine vascular endothelial growth factor and interleukin-8 in culture supernatant on time course by enzyme-linked immunosorbent assay. These findings constitute direct evidence that breast cancer induces postnatal vasculogenesis in vivo.