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Epoetin-beta is extremely useful as a drug for treating anemia in hemodialysis (HD) patients and is widely used for that purpose. The aim of this study was to determine whether once-weekly intravenous administration of epoetin-beta is as effective in maintaining hemoglobin (Hb) concentration as the same weekly dose administered 2 or 3 times per week as maintenance treatment of anemia in HD patients. The subjects were stable HD patients who had been receiving HD for at least 12 months. Using a fixed weekly dose of 3000 or 6000 IU of epoetin-beta, this study evaluated maintenance of improvement of anemia by comparing Hb concentration in the study period (once-weekly) with Hb concentration in the prestudy period (2 or 3 times per week). Of the 112 patients treated with epoetin-beta, 111 patients (full analysis set; 3000 IU, 52 patients; 6000 IU, 59 patients) were evaluated, after excluding one patient whose dose was changed immediately before study initiation. The change in the Hb concentration was maintained within +/-1.5 g/dL in 89.2% of patients (3000 IU, 88.5%; 6000 IU, 89.8%). The mean Hb concentration was 10.42 +/- 0.73 g/dL at study initiation and 10.14 +/- 1.00 g/dL at study completion. Adverse reactions occurred in 9.8% of patients (11 out of 112 patients). The main adverse reactions were malaise and increased blood pressure. Once-weekly intravenous administration of epoetin-beta is useful as maintenance treatment of anemia in HD patients and may be a treatment option.

Rho GTPases regulate several aspects of tissue morphogenesis during animal development. We found that mice lacking the Rho-inhibitory protein, p190-B RhoGAP, are 30% reduced in size and exhibit developmental defects strikingly similar to those seen in mice lacking the CREB transcription factor. In p190-B RhoGAP-deficient mice, CREB phosphorylation is substantially reduced in embryonic tissues. Embryo-derived cells contain abnormally high levels of active Rho protein, are reduced in size, and exhibit defects in CREB activation upon exposure to insulin or IGF-1. The cell size defect is rescued by expression of constitutively activated CREB, and in wild-type cells, expression of activated Rho or dominant-negative CREB results in reduced cell size. Together, these results suggest that activity of the Rho GTPase modulates a signal from insulin/IGFs to CREB that determines cell size and animal size during embryogenesis.

Chronic serum sickness nephritis was induced in rats by sensitization with egg albumin. The glomeruli were then examined by the quick-freezing and deep-etching method with immunohistochemical identification of immune complex deposits on replica membranes. In control rats, the glomerular basement membrane showed a three-layered structure. The middle layer was composed of a compact meshwork of fine fibrils that was connected to adjacent endothelial and epithelial cells by perpendicular fibrils in the inner and outer layers. The mesangial matrix contained a similar but looser meshwork structure. In the nephritic rats, small granular deposits were observed within the meshwork, distorting its fine structure. Larger nodular aggregates extended continuously from the middle layer to the epithelial side. Disruption of the connecting fibrils overlying these larger aggregates was noted. The deposits were stained by antiegg albumin or antirat IgG antibodies. Gold particles immunostaining for the sensitizing antigen were also localized within the deposits. These findings suggest that the deposition of immune complexes occurs in the fibrillary meshworks of the mesangium and lamina densa in this experimental model, with the resultant distortion of their meshwork structures and the formation of nodular aggregates.

The three-dimensional ultrastructure of glomerular sclerosis in the chronic phase of Masugi nephritis was investigated using a quick-freezing and deep-etching method. Newly formed mesangial matrix, which was increased in the axial portions, was composed of fine fibrillar networks similar to those in the lamina densa of the basement membrane. These fibrils were 10-20 nm in diameter and directly attached to the cell membranes of mesangial cells, endothelial cells and podocytes by connecting fibrils. Moreover, thicker fibrils with diameters of 20-30 nm were present in the networks and were connected with cross-bridges. A newly formed matrix of fine fibrillar networks was also seen in the areas of mesangial interposition in the glomerular capillary wall. The border between the matrix and lamina densa was unclear. The fibrils organizing the networks of lamina densa of the glomerular loop were thickened, with some decoration. Connecting fibrils were disrupted in the areas of endothelial detachment. It is suggested that prolonged tissue injury with endothelial detachment might induce mesangial sclerosis composed of fine fibrillar networks. The increase in density of the networks seemingly interfere with the contractile function of mesangial cells, which is followed by alteration of mesangial flow.

An increase in microfilaments in phalloidin-treated hepatocytes of Wistar rats was identified three-dimensionally with myosin subfragment 1 (S1) on replica membranes, using the quick-freezing and deep-etching method. Almost all of the reticular microfilaments around the bile canaliculi and beneath the lateral cell membranes were decorated on their surfaces by S1 attachment. Some showed periodic structures. However, thinner filaments with diameters of 4-7 nm were not decorated by S1. Bundled intermediate filaments around the bile canalicular microfilaments and intermediate filaments localized among cell organelles had smooth surfaces without S1 decoration. The microfilaments decorated by S1 were attached directly to bundled intermediate filaments. The quick-freezing and deep-etching method is useful in analysing cytoskeletal pathology and can be applied to histochemical fields.

Objective: Combination therapy with irinotecan (CPT-11), 5-fluorouracil (5-FU) and leucovorin is widely used for the treatment of metastatic colorectal cancer. However, little is known about the safety of chemotherapy of malignancies in hemodialysis (HD) patients. We encountered a patient with colorectal carcinoma on HD. We decided to administer combination chemotherapy while monitoring the pharmacodynamics of the patient. Case Report: A 74-year-old male received regular HD 3 times a week because of type 1 diabetes mellitus. He was diagnosed with stage IV colorectal cancer in November 2004. After sigmoidectomy, the patient received chemotherapy: a weekly schedule of CPT-11 (50 mg/m2) was administered followed by l-leucovorin (10 mg/m2) and 5-FU (400 mg/m2) just after HD. During the course, the plasma concentrations of both SN-38, an active metabolite of CPT-11, and 5-FU were not increased compared with those of patients with normal renal function. Our patient presented with grade III hematological toxicity that was easily recovered by granulocyte colony-stimulating factor. Conclusion: These data suggest that the dose-reduced Saltz regimen can be feasible for colorectal cancer patients receiving dialysis as postoperative adjuvant chemotherapy.

In Japan, calcimimetics and other phosphate binders such as lantanum carbonate are not available for patients on long-term hemodialysis (HD), so we prospectively evaluated the clinical efficacy of the combination of sevelamer hydrochloride and calcium carbonate (CaCO3) for hyperphosphatemia. The study group comprised 65 HD patients who had been administered CaCO3 (>or=1500 mg/day) for hyperphosphatemia [>or=6.0 mg/dL (>or=1.94 mmol/L)]. At the beginning of the study the dose of CaCO3 was reduced by 1500 mg/day and the patients divided into two groups according to the dose of additional sevelamer hydrochloride: group A 2250 mg/day; group B 3000 mg/day. Oral active vitamin D therapy was unchanged. Fourteen patients (21.5%) dropped out because of adverse effects and of the 51 remaining patients 35 (53.8%) suffered from gastrointestinal problems. Serum phosphate concentration decreased significantly [from 7.5+/-0.8 mg/dL (2.42+/-0.26 mmol/L) to 6.6+/-1.3 mg/dL (2.13+/-0.42 mmol/L), P

Perlecan, a large, multi-domain, heparan sulfate proteoglycan originally identified in basement membrane, interacts with extracellular matrix proteins, growth factors and receptors, and influences cellular signalling. Perlecan is present in a variety of basement membranes and in other extracellular matrix structures. We have disrupted the gene encoding perlecan (Hspg2) in mice. Approximately 40% of Hspg2-/- mice died at embryonic day (E) 10.5 with defective cephalic development. The remaining Hspg2-/- mice died just after birth with skeletal dysplasia characterized by micromelia with broad and bowed long bones, narrow thorax and craniofacial abnormalities. Only 6% of Hspg2-/- mice developed both exencephaly and chondrodysplasia. Hspg2-/- cartilage showed severe disorganization of the columnar structures of chondrocytes and defective endochondral ossification. Hspg2-/- cartilage matrix contained reduced and disorganized collagen fibrils and glycosaminoglycans, suggesting that perlecan has an important role in matrix structure. In Hspg2-/- cartilage, proliferation of chondrocytes was reduced and the prehypertrophic zone was diminished. The abnormal phenotypes of the Hspg2-/- skeleton are similar to those of thanatophoric dysplasia (TD) type I, which is caused by activating mutations in FGFR3 (refs 7, 8, 9), and to those of Fgfr3 gain-of-function mice. Our findings suggest that these molecules affect similar signalling pathways.

The mesangium is the central connective core of the individual lobules of glomerular tufts. The mesangial cells are surrounded by matriceal materials. This mesangial matrix (MM) is similar in appearance, but not identical to the peripheral glomerular basement membrane (GBM). The MM is composed of more coarsely fibrillar and slightly less electron-dense materials than that of the GBM observed on conventional ultrathin sections by electron microscopy. Moreover, the MM is a matter of interest to many investigators, because it is believed to play a role in the mediation of molecules passing through the glomerulus [1,2], and is involved in a variety of kidney diseases [3]. However, the three-dimensional organization of the MM is still poorly defined. The purpose of this study is to clarify the three-dimensional ultrastructure of the MM in situ by quick-freeze and deep-etch replica methods at high resolution [4, 5].

Ultrastructure of glomerular basement membrane by quick-freeze and deep-etch methods. The glomerular basement membrane of rat kidneys were three-dimensionally observed by quick-freeze and deep-etch replica methods at high resolution. The middle layer (lamina densa) was composed of 6 to 10 nm fibrils which formed a meshwork structure. The space between the fibrils had polygonal shape. The average long dimension of the space between fibrils was 17 nm and the short one was 13 nm. At the outer layer (lamina rara externa), fibrils connected podocytes perpendicularly with the meshwork of the middle layer. At the inner layer (lamina rara interna), similar perpendicular fibrils also connected endothelial cells with the meshwork of the middle layer. This is the first report to visualize the three-dimensional meshwork structure of the middle layer (the lamina densa) in situ. The function of anchoring podocytes to the lamina densa was suggested in the perpendicularly arranged fibrils of the outer layer. The quick-freeze and deep-etch method is useful in analyzing filamentous ultrastructure in glomeruli, and will be applied to clarifying pathological ultrastructure in kidney diseases.