四方 賢一

The aim of this study was to clarify the factors influencing the dietary behavior of patients with diabetic nephropathy. One hundred twenty-two patients with type 2 diabetes were recruited from the outpatients of Okayama University Hospital in Okayama, Japan. We performed a cross-sectional study using a questionnaire including 206 items among 18 categories as follows:background factors, coping behavior (coping scale), degree of uncertainty in illness (uncertainty scale), and dietary behavior. The data were analyzed by correlation analysis, t-test, one-way analysis of variance, Pearson correlation analysis, and multiple regression analysis. We found that those patients with microalbuminuria alone tended to recognize more mild about their kidney status than those with macroalbuminuria and chronic renal failure. We also found that common factors influencing the dietary behavior of diabetic patients with and without nephropathy are as follows:1. coping with the problem (beta0.342, p0.01);2. anxiety about prognosis (beta0.344, p0.01);3. sex (beta0.234, p0.05);4. uncertainty regarding treatment (beta0.377, p0.01);5. negative coping (beta0.354, p0.01);and 6. employment status (beta0.367, p0.01). Coping and uncertainty in illness had a significant relation to positive support and lack of support. To maintain appropriate dietary behavior in diabetic patients, medical staff need to determine what the social supports are important for the patient, and also to ensure good communication among healthcare personnel as well as positive support for patients and families.

We developed a "tissue negative staining method" to observe the molecular-level ultrastructure in situ in any portion of the ultrathin sections routinely prepared for electron microscopy. This method was used in electron microscopy of the glomerular basement membranes (GBM). The GBM in patients with nephrotic syndrome was discovered to possess a tunnel structure, designated as "nephrotic tunnel", with lumen large enough to allow free passage of protein molecules. This tunnel seemed to be involved in the etiology of nephrotic syndrome. This new method appears to be applicable to a variety of purposes in biological studies.

In order to clarify the mechanism of proteinuria in diabetic nephropathy, ultrastructural changes of the glomerular basement membrane (GBM) in patients with diabetic nephropathy were examined by electron microscopy using our newly devised "tissue negative staining method". The normal human GBM showed a fine meshwork structure consisting of fibrils forming the small pores. The diameter of these pores was slightly smaller than that of human albumin molecules. The GBM in patients with diabetic nephropathy showed irregular thickening. At higher magnification, hitherto unknown cavities and tunnel structures, which were not seen in normal controls, were observed in the thickened GBM. In some portions, these cavities presented a honeycomb-like appearance. The diameters of the cavities and tunnels were far larger than the dimensions of albumin molecules. These enlarged structures are believed to allow serum protein molecules to pass through the GBM from the capillary lumen to the urinary space. These results suggest that the cause of massive proteinuria in diabetic nephropathy is the disruption of the size barrier of the GBM.

To investigate the role of vitronectin in the progression of diabetic nephropathy, plasma concentrations of vitronectin were measured by enzyme-linked immunosorbent assay in patients with diabetes mellitus and compared with normal control subjects. In diabetic patients with normoalbuminuria and microalbuminuria, plasma concentrations of vitronectin were significantly higher than those of control subjects. Plasma concentrations of vitronectin in diabetic patients with chronic renal failure were significantly lower than those with normal renal function. There was a significant positive correlation between plasma concentration of vitronectin and blood platelet counts. In the early stage of diabetic nephropathy, vitronectin may be increased caused by synthesis from activated platelets. With progression of diabetic nephropathy, plasma vitronectin may be decreased because of accumulation in sclerotic glomeruli and arteriosclerotic lesions. In conclusion, the plasma concentration of vitronectin appears to be an important marker for the progression of diabetic nephropathy.

Sprague-Dawley rats, 6 with aminonucleoside nephrosis and 6 controls, were intravenously injected with human liver ferritin isolated from post mortem liver, and their 24-h urine samples were examined for human ferritin by immunoradiometric assay. In rats with aminonucleoside nephrosis, the amount of excreted ferritin in urine was forty times greater than in control rats. Much more monomeric ferritin was excreted than that of polymeric ferritin. We are the first to have utilized human liver ferritin as a tracer to measure a minor amount of ferritin by a commercially available kit. Our present study seems to indicate a critical role for glomerular basement membrane as a size barrier.

We demonstrated the ultrastructure of rat glomerular basement membrane (GBM) by ultra-high resolution scanning electron microscopy. GBM prepared by sonication methods and conductive-staining could be observed without metal coating at magnifications as high as 400,000 times. The GBM showed an irregular meshwork structure composed of various strands and pores. The width of the strands ranged from 6 to 15 nm, and the diameter of pores ranged from 6 to 50 nm. The present study confirmed our molecular sieve theory of the basement membrane.