There is growing evidence that ubiquitin-proteasome system plays an important role for the generation of circadian rhythms in mice as in Drosophila. Here we examined the expression of ubiquitin-related enzymes (Ubce5, UbcM4, Ube2v, Ube2d2, UchL1, UchL3, Ubp41, Ufd1L, β-TrCP) in the suprachiasmatic nucleus (SCN). At mRNA level, the expression of these enzymes were faint to moderate except ubiquitin carboxy-terminal hydrolase L1 (UchL1), a dominant deubiquitinating salvaging enzyme. Although strongly expressed in the SCN, UchL1 mRNA did not show the rhythm in the SCN in both light-dark and constant dark conditions.
The purpose of the current study was to evaluate the bone formation when β-tricalcium phosphate (TCP) was implanted in bone defects of rat femurs. β-TCP granules were applied to defects created in the femurs of 65 male rats who were sacrificed 3, 7, 10, 14 or 30 days later. Bone tissues were embedded in paraffin, serial sections were cut and then stained with hematoxylin-eosin. Histomorphometric analyses were also conducted. Furthermore, total mRNAs were extracted, homogenized, and reverse transcribed, after which quantitative PCR assays were conducted with a LightCycler™ using the double-stranded DNA dye Syber Green I with primers for either rat osteopontin or osteocalcin. Tissues in defects without β-TCP were used as controls. The amount of newly formed bone tissue in the β-TCP implanted group was significantly greater in both the side areas and the central area of defects than in the control group. Expressions of osteopontin and osteocalcin mRNAs of cells in the defects of the experimental group were up-regulated compared with the control group at all time periods. Taken together, these results prove that β-TCP is an appropriate material for osteoconduction and promotes bone formation in bone defects.
Several studies have reported that growth and differentiation of cultured myoblasts can be facilitated by applying appropriate mechanical stimulus. However, the effects of mechanical stimulus on the characteristics of muscle fibers have not yet been fully elucidated. In this study, we gave mechanical stress to C2C12 cells, which were myoblasts derived from mice skeletal muscle. The following myosin heavy chain (MHC) isoforms were investigated in order to clarify muscle characteristics: MHC-2b, 2d and 2a, all of which are fast-twitch fibers. After inoculating cells on a silicone chamber, the chamber was mechanically stretched, and a LightCycler™ was used to measure the mRNA expression of each MHC isoform at several times. The results showed that, with mechanical stretching, the expression of MHC-2b was initially high. On the other hand, without stretching, the expression of MHC-2d increased over time, but with stretching, it was hardly seen. Furthermore, the expression of MHC-2a was significantly high in the stretching group. These results of this study suggest that, when intermittently stimulated, myoblasts express increased levels of MHC-2a isoform. Therefore, it is indicated that myocytes respond to environmental changes not only to facilitate growth and differentiation, but also to alter muscle function actively at the MHC isoform level.
The aim of this study was to evaluate the effects of enamel matrix proteins (EMP) at the early stage of wound healing in the periodontal tissues by a combination treatment with guided tissue regeneration (GTR). Intrabony defects were produced surgically at the distal aspects of both mandibles in six beagle dogs. At 12 weeks following the surgery, the defects were exposed using a full thickness mucoperiosteal flap procedure. Subsequently, the defects were treated by the following treatments: a control group treated with GTR alone, and an experimental group treated with a combination of GTR and EMP. After one, two, four and eight weeks of the treatment, the animals were sacrificed, and sections of the tissue were stained and evaluated microscopically. After one and two weeks, the proliferating cell nuclear antigen (PCNA)-positive cell ratios of the experimental group were significantly greater than that of the control group. After 2 and 4 weeks, new bone and new cementum formation in the experimental group were significantly greater than those in the control group (P < 0.05). However, after 8 weeks, no statistical difference was found between the two groups in new bone or cementum formation. The study results suggest that a maturation of periodontal ligament cells might contribute, during the early stage of periodontal healing, to stimulate a proliferation of periodontal ligament cells.
In this study we tried to identify new genes or proteins in skeletal muscle induced by exercise. We analyzed alterations of protein expression in mouse gastrocnemius muscles induced by swimexercise using two dimensional gel electrophoresis and mass spectrometry. Nine spots were significantly altered between control and swim groups. One of the four protein spots whose expression was decreased was identified as functionally unknown “expressed sequence AI854635” gene. The AI854635 gene has C2H2 type zinc finger motif, and is considered to be a transcription factor. The mRNA of AI854635 gene was expressed in skeletal muscle, brain, kidney, and thymus. To elucidate the function of the AI854635 gene we analyzed mRNA expression levels during C2C12 myoblast differentiation. C2C12 myoblast began to form myotube around 20 h after the initiation of differentiation. The mRNA expression levels of AI854635 gene was significantly induced around 6 h and increased till 48 h, indicating a pivotal role in myoblast differentiation. Although the significance of decreased expression of AI854635 gene induced by swim-exercise is not clear, we found that this gene is involved in myoblast differentiation.
Proteins of the Bcl-2 family are key regulators of apoptosis. Bax can be regarded as pro-apoptotic, whereas Bcl-2 is perceived as anti-apoptotic. It has been proposed that an increased ratio of proapoptotic Bax to anti-apoptotic Bcl-2 can be associated with apoptosis. Since prostaglandin A2 (PGA2) and 2-methoxyestradiol (2-ME) play an active role in the induction of apoptosis, the influence of 20 μg/ml PGA2 and 1 μM 2-ME was investigated on Bax and Bcl-2 expression levels in cervical carcinoma cells. Both PGA2 and 2-ME exposure led to statistically significant increases in Bax expression levels. Cells were shown to be more susceptible to the effects of 2-ME than to the effects caused by PGA2. In contrast, no statistically significant effects were observed on Bcl-2 expression levels after exposure to PGA2 and 2-ME. The Bax/Bcl-2 ratios for PGA2- and 2-ME-exposed cells were 2.06 and 1.87 respectively, normalised against Bcl-2 levels. Further investigation of the function and regulation of the Bcl-2 family will allow researchers to consider potential pathways of apoptosis signaling mechanisms for diseases where apoptosis can potentially be controlled.