Nobuaki Yanagisawa

Regulation of Osteoclast Multinucleation by the Actin Cytoskeleton Signaling Network.

Although it is known that osteoclasts are multinucleated cells that are responsible for bone resorption, the mechanism by which their size is regulated is unclear. We previously reported that an actin‐rich superstructure, termed the zipper‐like structure, specifically appears during the fusion of large osteoclast‐like cells (OCLs). Actin cytoskeleton reorganization in osteoclasts is regulated by a signaling network that includes the macrophage colony‐stimulating factor (M‐CSF) receptor, a proto‐oncogene, Src, and small GTPases. Here, we examined the role of actin reorganization in the multinucleation of OCLs differentiated from RAW 264.7 cells using various pharmacological agents. Jasplakinolide, which stabilizes actin stress fibers, induced the development of small OCLs, and the Src inhibitor SU6656 and the dynamin inhibitor dynasore impaired the maintenance of the podosome belt and the zipper‐like structure. These inhibitors decreased the formation of large OCLs but increased the number of small OCLs. M‐CSF is known to stimulate osteoclast fusion. M‐CSF signaling via Src up‐regulated Rac1 activity but down‐regulated Rho activity. Rac1 and Rho localized to the center of the zipper‐like structure. Rho activator II promoted the formation of small OCLs, whereas the Rho inhibitor Y27632 promoted the generation of large OCLs. These results suggest that the status of the actin cytoskeleton signaling network determines the size of OCLs during cell fusion. J. Cell. Physiol. 230: 395–405, 2015.

Effect of mechanical stretching on expressions of muscle specific transcription factors MyoD, Myf-5, myogenin and MRF4 in proliferated myoblasts.

We examined expression of four important members of myogenic regulatory factors (MRFs) in the myoblasts both at mRNA and protein levels, which were subjected to mechanical stretching in in vitro condition. Our results showed that MyoD expression existed both in the stretch and in the control group at all time periods of the mechanical stimulus. Myf‐5 expressed only at early stage of the stretch group. Although mRNA and protein expressions of myogenin and MRF4 were detected both in the stretch and in the control group at 12 h after the stretching, their expressions were only shown in the stretch group at 24 h after the mechanical stimulus. However, at 36 and 48 h, none of the MRFs examined except MyoD appeared in both groups. Our results suggest that the MRFs are up‐regulated upon mechanical stimulus and each member plays a different major role for either proliferation or differentiation of the myoblasts.

Increased expression of decorin during the regeneration stage of mdx mouse

Satellite cells exist in postnatal muscle tissue and constitute the main source of muscle precursor cells for growth and repair. These cells carry out important roles for skeletal muscle formation postnatally during growth of muscle mass as well as damage-induced regenerative processes. Muscle regeneration supports muscle function in aging and has a role in the functional impairment caused by progressive neuromuscular diseases. Major substances controlling this process are growth factors and extracellular matrix. Myostatin, a member of TGF-β family, was mainly expressed in muscle tissue. Decorin, a member of the small leucine-rich proteoglycan gene family, is composed of a core protein and a dermatan/chondroitin sulfate chain. Recent studies have shown that decorin enhanced the proliferation and differentiation of myogenic cells by suppressing myostatin activity. Thus, decorin appears to be a new molecule in the myostatin signaling pathway and a promising target for treatment of progressive neuromuscular diseases. Therefore, in this study, we examined the localization of decorin as well as myostatin in a muscular dystrophy model in mdx mice and B10 Scott Snells mice as a control to elucidate the differences between decorin and myostatin messages as well as protein distribution. This study revealed increased expression of decorin protein as well as mRNA at the regenerative stage of mdx mice compared to early stages, while only weak expression of decorin was detected in the control mice. Our study contributes to identifying the relationship between decorin and myostatin as well as the development of a therapeutic strategy for progressive neuromuscular diseases.

The expression of embryonic globin mRNA in a severely anemic mouse model induced by treatment with nitrogen-containing bisphosphonate.

BackgroundMammalian erythropoiesis can be divided into two distinct types, primitive and definitive, in which new cells are derived from the yolk sac and hematopoietic stem cells, respectively. Primitive erythropoiesis occurs within a restricted period during embryogenesis. Primitive erythrocytes remain nucleated, and their hemoglobins are different from those in definitive erythrocytes. Embryonic type hemoglobin is expressed in adult animals under genetically abnormal condition, but its later expression has not been reported in genetically normal adult animals, even under anemic conditions. We previously reported that injecting animals with nitrogen-containing bisphosphonate (NBP) decreased erythropoiesis in bone marrow (BM). Here, we induced severe anemia in a mouse model by injecting NBP injection in combination with phenylhydrazine (PHZ), and then we analyzed erythropoiesis and the levels of different types of hemoglobin.MethodsSplenectomized mice were treated with NBP to inhibit erythropoiesis in BM, and with PHZ to induce hemolytic anemia. We analyzed hematopoietic sites and peripheral blood using morphological and molecular biological methods.ResultsCombined treatment of splenectomized mice with NBP and PHZ induced critical anemia compared to treatment with PHZ alone, and numerous nucleated erythrocytes appeared in the peripheral blood. In the BM, immature CD71-positive erythroblasts were increased, and extramedullary erythropoiesis occurred in the liver. Furthermore, embryonic type globin mRNA was detected in both the BM and the liver. In peripheral blood, spots that did not correspond to control hemoglobin were observed in 2D electrophoresis. ChIP analyses showed that KLF1 and KLF2 bind to the promoter regions of β-like globin. Wine-colored capsuled structures were unexpectedly observed in the abdominal cavity, and active erythropoiesis was also observed in these structures.ConclusionThese results indicate that primitive erythropoiesis occurs in adult mice to rescue critical anemia because primitive erythropoiesis does not require macrophages as stroma whereas macrophages play a pivotal role in definitive erythropoiesis even outside the medulla. The cells expressing embryonic hemoglobin in this study were similar to primitive erythrocytes, indicating the possibility that yolk sac-derived primitive erythroid cells may persist into adulthood in mice.

Root resorption of maxillary primary incisors in relation to position of successive permanent incisors by Micro-CT

The aim of this study is to elucidate the root resorption of primary incisors in relation to the development of permanent incisors. We observed the maxillas of the dry skulls of Indian children, using Micro-CT, and measured distance between the root surface of the primary incisor and the bony crypt of the permanent incisor. The bony crypt of the maxillary permanent incisor, which was situated at the lingual side of the primary incisor, grew upward towards the mouth as the tooth eruption stage proceeded. Root resorption was clearly seen at the primary dentition stage, and it proceeded to from root surface towards root canal at the first molar eruption. With the advance of the stage, the distance between the root surface of the primary incisor and the bony crypt of the permanent incisor became shorter, especially the distance between the root of the primary lateral incisor and the bony crypt of the permanent central incisor did. By using Micro-CT, we could confirm more clearly that the growth of the bony crypt of the central permanent incisor deeply influence the progress of root resorption of the primary lateral incisor.

Morphology and relationships of the biceps brachii and brachialis with the musculocutaneous nerve

IntroductionMajor anatomical textbooks generally state that the biceps brachii muscle (BB) is composed of long and short heads, whereas the brachialis muscle (BR) consists of a single head. However, the numbers of heads comprising the BB and the BR are very variable. The purpose of this study was to investigate how the branching patterns of the musculocutaneous nerve (MC) influence the number of heads of the BB and the BR.Materials and methodsMorphological examinations of the BB and MC were conducted using cadavers of 22 Japanese individuals, and morphological examinations of the BR and the MC were conducted in 9 of those 22 individuals.ResultsA three-headed BB was observed in 7 of the 22 specimens (31.8%). Most of these specimens showed a Type III branch pattern (after penetrating the long head or the short head, the MC innervated the supernumerary head or communicated with the main root again). The number of BR heads was categorized into three types: Type A, two heads (superficial and deep heads, 22.2%); Type B, three or four heads (two or three superficial heads and one deep head, 44.4%); and Type C, multiple heads (33.3%). Among these categories, branches of the MC in Type A specimens were most simple.ConclusionA supernumerary head of the BB seemed to be present if the MC penetrates it. The BR basically consists of superficial and deep heads, and the number of superficial heads is affected by branches of the MC.