Nobuko Kawaguchi

Endogenous Adipocyte Precursor Cells for Regenerative Soft-Tissue Engineering

Subcutaneous injection of reconstituted basement membrane (Matrigel) in combination with basic fibroblast growth factor induces de novo adipogenesis in which endogenous precursor cells invade the artificially formed Matrigel space, proliferate and differentiate to form adipose tissue. Since this adipogenesis offers us a novel approach for soft-tissue reconstruction without transplanting preadipocytes, the early process was examined by optical and electron microscopy. Formation of multiple layers of fibroblast-like cells at the surface of Matrigel implant was the first response of connective tissue. The cells within four to five layers proximal to Matrigel implant acquired a thick cytoplasm and an enlarged nucleus, and they invaded Matrigel space together with endothelial cells which caused neovascularization. Phagocytotic incorporation and digestion of Matrigel components by well-developed lysosomes appeared to be a stimulus of fibroblast-like cells to mature depending on proximity to Matrigel. The fibroblast-like cells often contacted to the outer surface of capillary over a large area and rapidly accumulated lipid droplets. Electron microscopy of the developing adipocytes showed a well-organized smooth endoplasmic reticulum and mitochondria. This investigation thus revealed the characteristics of adipocyte precursor cells, which can be recruited for regenerative engineering of soft tissues.

Reconstituted basement membrane potentiates in vivo adipogenesis of 3T3-F442A cells

Adipocytes forming fat pad in vivo are surrounded by well developed basement membranes. Synthesis of basement membrane is enhanced during in vitro differentiation of preadipocyte line. In order to know the role of basement membrane in adipogenesis in vivo, we injected 3T3-F442A preadipocytes subcutaneously into nude mice together with or without the reconstituted basement membrane, Matrigel. Histological sections of the fat pads newly formed by injecting the cell alone showed dense population of immature adipocytes and microvessels within 2 weeks and they matured rapidly. In contrast, injection of the cells together with Matrigel showed sparse adipocytes after 2 weeks and they matured slowly over the period of 6 weeks. Quantification of the process by measuring the weight, DNA content, triglyceride content and glycerophosphate dehydrogenase (GPDH) activity of the fat pads showed that injection of the cell alone resulted in early maturation of adipose tissue with fewer adipocytes while the presence of Matrigel decelerated but potentiated the maturation of adipose tissue with 2 fold contents of DNA, triglyceride and GPDH activity. We thus showed that reconstituted basement membrane (Matrigel) supported the survival and maturation of adipocytes.

Hypoxic enhancement of type IV collagen secretion accelerates adipose conversion of 3T3-L1 fibroblasts.

Hypoxic modulation of collagen metabolism appears to be related to pathogenesis of many diseases such as fibrosis of connective tissue after injury and scleroderma. Since most of our understanding of how procollagen assembles within the cell has come from studies on cells cultured under normoxia, it may not be helpful for the etiology of the diseases observed in peripheral tissues under hypoxic conditions. As an experimental model for the hypoxic modulation of collagen metabolism, we cultured 3T3-L1 fibroblasts under low partial oxygen pressure and found that hypoxia enhances secretion of type IV collagen 10-fold and accelerates adipose conversion of the cells. The enhanced secretion of type IV collagen was not accompanied by an appreciable increase of alpha1(IV) and alpha2(IV) mRNAs. Prolyl 4-hydroxylase alpha increased only 3-fold under hypoxia. We suggest that hypoxia creates an environment of prolyl 4-hydroxylase alpha(2)beta(2) tetramers favorable for the folding of type IV procollagen which has many interruptions of the Gly-Xaa-Yaa repeat.

De novo adipogenesis for reconstructive surgery

Autografting of lost soft tissue is an important subject of the plastic and reconstructive surgery and autograft of fat pads has been only technique for this goal. However, the results are disappointing because of absorption of the grafts with time. Adipoblasts or adipocyte precursor cells distribute widely in connective tissues and they can proliferate and mature into adipocytes even in the adult body. In experiments using mice, we found that de novo adipogenesis of endogenous precursor cells can be induced by injecting reconstituted basement membrane, Matrigel, supplemented with more than 1 ng/ml of bFGF. This adipogenesis was reproducibly induced by subcutaneous injection over the chest, lateral abdomen or head. Adipogenesis was induced even in ear cartilage or in muscle. To evaluate the possibility of future application of this de novo adipogenesis to plastic and reconstructive surgery, we have reviewed updated knowledge of the adipogenesis.

Case Studies in Flatland Farmland Neighborhood Landscape Complex Units: Asami District

This chapter discusses the case study of Asami District, located in the eastern part of Matsusaka City in Mie Prefecture between the Kushida River in the east and the Kongo River in the west, as a flatland farmland neighborhood landscape complex unit. Asami District was developed during the 8th century, and in some parts still uses Jori-sei, a traditional land allotting system, that encourages and maintains rich biodiversity in the area. However, as traditional paddy field allotting based on Jori-sei imposes a stronger level of labor to aged farmers, the modern agricultural land improvement project is progressing rapidly in this area. The loss of these undeveloped fields with ecological, historical, and cultural value could negatively impact the area; as such, a well-considered improvement method for sustainable land management of the area is required. As a consequence, the relationship between biodiversity and landscape management labor accounts is clarified and future scenarios that are balancing them are considered.

Labor Forces and Landscape Management

Many mature countries, including Japan, are confronting population decline and aging societies. Such a radical decrease of the population could adversely affect the sustainability of landscape management, as the issue is exacerbated by the lack of skilled local labor forces in landscape management. It is important, therefore, to develop methods for providing or re-providing appropriate labor for the sustainable management of landscapes, and how to sustainably reduce or shrink landscape management areas according to the available labor. In this chapter, an original concept of landscape management labor accounts is proposed, and the background, fundamentals, and perspective is explained.

Managing Urban and Rural Agricultural Landscape Processes in Japan

In Japan, the goal of land use policy, which was established in the City Planning Act in 1968 and the Act on Establishment of Agricultural Promotion Regions in 1969, was to divide the urban and rural area by zoning in the expanding periods, but many researchers have pointed out that it has failed and urban sprawl has expanded in the urban periphery areas due to loose land use regulations. However, today in the shrinking periods, the need of diversion of agricultural land to residential land is declining. On the other hands, urban residents have recently shown more interest in the food safety and the urban agriculture, and the expectation to the various ecological services of agricultural fields in urban area such as disaster prevention, environmental improvement, and landscape preservation has been increasing. Under these circumstances, Urban Agriculture Promotion Basic Plan was established in 2015 and Urban Green Space Conservation Law was revised in 2017. This chapter introduces the laws and some recent attempts of urban agriculture from the following four points; (1) To promote further entry of various motivated entities into agriculture such as urban residents, and companies which promote new agricultural businesses, (2) To evaluate the mixture of urban and rural areas positively by regarding urban agricultural fields not as “the place to be developed” but as “the place to be preserved”, (3) To improve the self-sufficiency rate by the high technology collection, and (4) To balance the productivity and the biological diversity by branding of the environmentally-friendly agriculture.

Sustainable Landscape Management and Landscape Management Labor Accounts

There exist many kinds of landscape elements such as garden trees, roadside trees, urban gardens, parks, paddy fields, other farmlands, bamboo bushes, forest and so on. Human labor is required to maintain these green features. In this article, the concept of ‘landscape management labor accounts’ is proposed and investigated through some case studies. An annual personal labor account is a personal labor account required to maintain a targeted landscape element for 1 year. A personal labor account per area is the annual personal labor density needed to maintain one area for 1 year. Annual personal labor accounts range widely from 3 to 2340 man-hours per year. Also personal labor densities also show wide variation from 0.01 to 85.8. Five working style types can be distinguished according to the relationship between annual personal labor accounts and personal labor densities.

Integration of Landscape Management Labor Accounts

This chapter summarizes the results of the case studies in previous chapters and then classifies the working style types by analyzing the working time and working area of all neighborhood landscape complex units. Then, the total labor accounts of neighborhood landscape complex units are compared to one another. In addition, the integration of landscape labor accounts from small scale to large scale is considered. On a city scale, the targeted area is Nagoya City, the total labor accounts of which are calculated and future prospects for sustainable landscape management are proposed. On a regional scale, the targeted area is Chubu Metropolitan District, the total labor accounts of which are calculated in 1 km mesh and the future situations, particularly the shortage of landscape management plans, are examined through comparing the current and forecasted populations.

Conclusion: Towards Sustainable Labor Force Management of Landscapes

This chapter summarizes the key concepts of this book. In the shrinking and aging societies throughout Japan, the issues of landscape management are serious. In this book, the concept of landscape management labor accounts is proposed toward a comprehensive and integrated understanding of labor force management of the ecological elements and units in different landscapes from a small to large scales. The framework of landscape management labor accounts will provide a good viewpoint for analyzing current problems and determining future solutions that involve the cooperation of various stakeholders. In the future, collaborative development of functional studies on ecological services, structural studies on green infrastructure, and affordability studies on landscape management labor accounts might provide an integrated planning and design process with not only the aspects of ecological values considered but also economic and social values in a sustainable and resilient manner as well.