Yoshitake Ito

Age-related decrease in the number of hemopoietic stem cells and progenitors in senescence accelerated mice

Senescence accelerated mice (SAM-P) were used for the study of the possible aging of hemopoiesis. The number of peripheral leukocytes decreased significantly with age, whereas hematocrit showed only a slight decrease. Although the number of total nucleated cells in the bone marrow increased, the number of hemopoietic stem cells (CFU-S) as well as that of granulocyte-macrophage colony forming cells (GM-CFC) showed a decrease in old mice. A significant decrease in the number of GM-CFC was observed in the spleen of old SAM-P mice, whereas no decrease was found in the number of CFU-S. Such a profound reduction of the recruitment of GM-CFC from CFU-S in the spleen together with a reduction of bone marrow hemopoiesis may be responsible for the decrease in the number of peripheral leukocytes in the old mice. SAM-P mice could provide a good model for the study of the aging of hemopoietic system.

Age-related attenuation of HSP47 heat response in fibroblasts

The collagen-binding heat shock protein of molecular weight 47,000 (HSP47), resident in the endoplasmic reticulum (ER), is assumed to play a specific role as a molecular chaperon in the processing of procollagen molecules. The present investigation of age-related alteration in the HSP47 heat response in cultured murine and human fibroblasts revealed expression in cells with a low population doubling level (PDL) derived from young mice and people more inducible by heat treatment than those from older mice and people. On the other hand, cells with a high PDL showed a very low heat response in terms of HSP47 expression regardless of the donor age. Northern blot analysis of HSP47 m-RNA indicated that the age related attenuation of HSP47 expression was regulated by transcriptional mechanisms. Furthermore, immunofluorescent analysis using a monoclonal antibody against the carboxylterminal propeptide of type I procollagen revealed far greater retention of procollagen molecules in the ER lumen of cells from old persons than in those from young persons. This was particularly prominent in heat-treated cells from old persons, indicating the possibility that the observed decrease in HSP47 heat response might cause blockage of procollagen transport to the Golgi and therefore secretion.

Age-related change in brush borders of rat kidney cortex

Age-related change of rat renal brush borders was examined with electron microscopy and biochemical procedures. Total activity of renal brush border enzymes, such as alkaline phosphatase and leucine aminopeptidase in the homogenate, was significantly decreased with age. Acid phosphatase activity and protein content were not significantly changed with age. Specific activity of leucine aminopeptidase in brush border fraction was significantly decreased at a later stage of age. Protein content of brush border fraction was decreased significantly with age. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, some of the proteins disappeared during aging. Electron microscopic observations of kidney cortex showed that microvilli of renal brush borders from old rats were observed to be fewer than those from the young; epithelial cells in young rats have more densely packed brush border projections than those in old rats. From these results, it is suggested that during aging renal brush borders are degraded, and that protein components of the brush borders were different between old and young.

Morphological aspects of aging liver: half a century of progress in Japan

The most important morphological aspects of the physiological changes of the liver with age are decreases in the numbers of hepatocytes and their mitochondria, coupled with increases in their volumes. The latter is considered to partially compensate for the decrease in numbers of hepatocytes and mitochondria, but leads to a more limited reserve capacity and thus to a condition in which liver function can be more readily exhausted. The binuclearity and polyploidization of the hepatocytes increase with age and are considered to be due to inhibition of their cell division process. Our studies on the parabiosis between young and old rats and mice, on the implantation of isolated hepatocytes in the spleens of rats of different ages, and on the regenerative process after partial hepatectomy in rats of different ages, suggested the existence of humoral inhibitory factor(s) for the division of the cells and their mitochondria. Morphological studies on the native and Hawaiian Japanese, as well as on the USA Caucasians and the Costa Rican Latinos revealed that a difference in the process of the age-related decrease in hepatocyte number among different ethnicities is mainly due to differences in nutritional conditions rather than to genetic factors. Experimental studies are now being carried out on the effect(s) of various dietary conditions on age-related changes of the liver, and the results obtained support the findings of the geographical studies.

Occlusal tooth wear in male F344/N rats with aging

With the aim of clarifying the aging properties of an animal model, the progress of occlusal tooth wear (OTW) of molars in male F344/N rats was monitored. Dried maxilla and mandible specimens from 61 male F344/N rats, aged 7 to >30 months, were used. The levels of OTW of all molars were monitored with aging. The cuspis dentis of molar teeth were worn out by 7 months (M) of age, and the occlusal surface became flat. As for each molar tooth (M(1-3), numbered in accordance with its position), OTW of M(1) was more severe in the lower than in the upper jaw, whereas M(3) was more severe in the upper than the lower jaw. OTW of M(2) in both the upper and the lower jaws progressed rapidly after 27M. OTW in male F344/N rats progressed faster than in females. However, when compensated for life span, both genders had similar profiles in OTW progress with aging. This study suggested that male rats were more convenient than females as a model for gerodontological research because of the earlier course of OTW progress.

Roles of naofen, a novel WD-repeat-2 protein, in the CCl4-treated livers--a possible relationship to cell proliferation.

Naofen (GenBank accession no. EF613262), a newly found intracellular protein in the WD-repeat-2 protein family, has been cloned as an anti-verotoxin II antibody immunoreactive substance, and the nucleotide- and amino acid-sequences have been clarified. The present study was undertaken to evaluate the roles of naofen especially in carbon tetrachloride (CCl4-induced cirrhosis model of rats, also in partial hepatectomy. Naofen mRNA expressions were observed from the early phases of cirrhosis development and during regenerative phases after partial hepatectomy, more remarkable in the former. Naofen immunoreactive fragments located in the vascular endothelial cells and peri-vascular spaces in normal livers especially in Glissons areas, being strongly stained in the connective tissues 8 weeks after starting CCl4-injections, besides in the cytoplasm of hepatocytes in pseudo-lobules. In contrast, partial hepatectomy caused a small increase of naofen expressions in the whole hepatocytes, and significantly in the endothelial cells of portal veins and hepatic arterioles. Furthermore, in parallel to the degree of naofen mRNA and protein expressions, the rates of double-nuclei cells to total hepatocytes in the Glissons areas increased in both cirrhosis and partial hepatectomy, suggesting a relationship between naofen expression and mitosis. In in-vitro studies with cell lines, vascular endothelial growth factor, a cell proliferation stimulant, increased the naofen mRNA expressions in HepG(2) cell lines, whereas paclitaxel, a cytotoxic anti-cancer drug, diminished them in NRK52E, both concentration-dependently. These results indicated that naofen immunoreactive fragments play an important role in the cell proliferation, relevant for analyzing the regenerative phases during cirrhosis developments and after partial hepatectomy.

A role of anti-verotoxin antibody immunoreactive peptide, Virp5, from rat spinal cord

An anti-verotoxin 2 (VT2) antibody immunoreactive 5-kDa polypeptide (Virp5), has been obtained through screening of the rat spinal cord cDNA library with the aid of anti-VT2 antibody. Virp5 was mainly expressed in the central nervous system, liver and kidney, and localized at glia-like cells and nerve fibers in the central nervous system, vascular endothelial cells and hepatic cells in the liver, as well as epithelial cells of distal tubules in the kidney. Intravenous administration of purified Virp5 elicited a dose-dependent increase in blood pressure. These results suggest that Virp5 commonly exists in the body, being partly playing a role in regulating the blood pressure.

Parvalbumin expression in the diaphragm of aged rats.

Parvalbumin is a calcium-binding protein involved in the mechanism of Ca(++) exchange between the sarcoplasmic reticulum and the myofibrils. There is a direct correlation between the amount of parvalbumin and the velocity of contraction-relaxation of skeletal muscle fibers. Male Donryu rats (6, 24 and 30 months of age) were housed in a specific pathogen free room and fed with 60% of the mean daily quantity of rodent chow consumed by rats fed ad libitum. To assess the expression of parvalbumin in the rat diaphragm, transversal sections were processed immunohistochemically using polyclonal antiparvalbumin antibody. The stained sections were inspected on an image analysis device to evaluate the intensity of the immunoreaction and to take morphological measurements of the muscle fibers. The optical density data allowed the distinction of fibers in fast- and slow-twitch types. The results show that the number of fibers did not change in young and old rats, but the percentage of muscle fibers containing parvalbumin decreased with age. Moreover, the fast-twitch fibers showed a remarkable decrease in their diameter at 30 months of age, but the slow-twitch ones did not. The results suggest that, as the organism gets old, the fiber type composition of the diaphragm changes to adapt to different physiological and/or metabolic exigencies.

Morphological age changes in the diaphragm of rats under different protein diets.

Male Donryu rats were housed in a specific pathogen free room and fed with 60% of the mean daily quantity of diet consumed by rats fed ad libitum The animals were divided into three groups of 50 and each group fed different amounts and types of dietary protein. The group P40 was fed with a diet containing 40% of total volume as vegetable protein; the agroup P10 with 10% of total volume as vegetable protein; and the group CE7 with 18% of total volume as mixed vegetable and animal proteins. Subsets of the animals were sacrificed at the ages of 6, 12, 18, 24 and 30 months. The P40 group gained body weight faster than other two groups during the first months. However, after 12 months, the body weight began to decrease, reaching the lowest mean body weight among the three groups by the end of the study. The three groups did not show any significative difference in the muscle fiber number index and in the number of capillaries per muscle fiber. The calculated diaphragm volume index increased up to a peak at 12 months in all three groups decreasing gradually thereafter, but did not show any significant difference due to dietary protein intake. The data suggest that the morphological age changes occurring in the diaphragm muscle are likely unrelated to dietary conditions and are more likely due to other factors such as changes in the neural activity during aging.

Morphometrical analysis of autofluorescent granules in bronchial epithelia of human lung cancer cases and suncus fed BHA.

ヒト肺癌症例の肺とBHA投与スンクス肺を用いて気管支上皮内の自家蛍光色素顆粒の量を微計測した.ヒト肺腺癌例では気管支上皮内の顆粒が, 非癌例, 扁平上皮癌例に比して有意に多く, 特に男性の喫煙者に多かった.BHA投与スンクスの気管支上皮においては14か月後には色素顆粒が増加し同時に腺腫様増殖が生じた.以上のことから気管支上皮の発癌過程について脂質代謝の面から考察した.