Vibration acceleration enhances proliferation, migration, and maturation of C2C12 cells and promotes regeneration of muscle injury in male rats.

Vibration acceleration (VA) using a whole-body vibration device is beneficial for skeletal muscles. However, its effect at the cellular level remains unclear. We aimed to investigate the effects of VA on muscles in vitro and in vivo using the C2C12 mouse myoblast cell line and cardiotoxin-induced injury in male rat soleus muscles. Cell proliferation was evaluated using the WST/CCK-8 assay and proportion of Ki-67 positive cells. Cell migration was assessed using wound-healing assay. Cell differentiation was examined by the maturation index in immunostained cultured myotubes and real-time polymerase chain reaction. Regeneration of soleus muscle in rats was assessed by recruitment of satellite cells, cross-sectional area of regenerated muscle fibers, number of centrally nucleated fibers, and conversion of regenerated muscle from fast- to slow-twitch. VA at 30 Hz with low amplitude for 10 min promoted C2C12 cell proliferation, migration, and myotube maturation, without promoting expression of genes related to differentiation. VA significantly increased Pax7-stained satellite cells and centrally nucleated fibers in injured soleus muscles on Day 7 and promoted conversion of fast- to slow-twitch muscle fibers with an increase in the mean cross-sectional area of regenerated muscle fibers on Day 14. VA enhanced the proliferation, migration, and maturation of C2C12 myoblasts and regeneration of injured rat muscles.

Variability in Physical Inactivity Responses of University Students during COVID-19 Pandemic: A Monitoring of Daily Step Counts Using a Smartphone Application.

This study investigated the changes in physical inactivity of university students during the COVID-19 pandemic, with reference to their academic calendar. We used the daily step counts recorded by a smartphone application (iPhone Health App) from April 2020 to January 2021 (287 days) for 603 participants. The data for 287 days were divided into five periods based on their academic calendar. The median value of daily step counts across each period was calculated. A k-means clustering analysis was performed to classify the 603 participants into subgroups to demonstrate the variability in the physical inactivity responses. The median daily step counts, with a 7-day moving average, dramatically decreased from 5000 to 2000 steps/day in early April. It remained at a lower level (less than 2000 steps/day) during the first semester, then increased to more than 5000 steps/day at the start of summer vacation. The clustering analysis demonstrated the variability in physical inactivity responses. The inactive students did not recover daily step counts throughout the year. Consequently, promoting physical activity is recommended for inactive university students over the course of the whole semester.

Vibration acceleration promotes endochondral formation during fracture healing through cellular chondrogenic differentiation.

Vibration acceleration through whole body vibration has been reported to promote fracture healing. However, the mechanism responsible for this effect remains unclear. Purpose of this study was to determine whether vibration acceleration directly affects cells around the fracture site and promotes endochondral ossification. Four-week-old female Wistar Hannover rats were divided into two groups (vibration [V group] and control [C group]). The eighth ribs on both sides were cut vertically using scissors. From postoperative day 3 to 11, vibration acceleration using Power Plate® (30 Hz, low amplitude [30-Low], 10 min/day) was applied in the V group. Mature calluses appeared earlier in the V group than in the C group by histological analysis. The GAG content in the fracture callus on day 6 was significantly higher in the V group than in the C group. The mRNA expressions of SOX-9, aggrecan, and Col-II in the fracture callus on day 6 and Col-X on day 9 were significantly higher in the V group than in the C group. For in vitro analysis, four different conditions of vibration acceleration (30 or 50 Hz with low or high amplitude [30-Low, 30-High, 50-Low, and 50-High], 10 min/day) were applied to a prechondrogenic cell (ATDC5) and an undifferentiated cell (C3H10T1/2). There was no significant difference in cell proliferation between the control and any of the four vibration conditions for both cell lines. For both cell lines, alcian blue staining was greater under 30-Low and 50-Low conditions than under control as well as 30-High and 50-High conditions on days 7 and 14. Vibration acceleration under 30-L condition upregulated chondrogenic gene expressions of SOX-9, aggrecan, Col-II, and Col-X. Low-amplitude vibration acceleration can promote endochondral ossification in the fracture healing in vivo and chondrogenic differentiation in vitro.

Expression Analysis of Serotonin Receptors, Serotonin Transporter and l-Amino Acid Decarboxylase in the Mouse Sphenopalatine Ganglion by RT-PCR, Northern Blot Analysis and In Situ Hybridization.

The sphenopalatine ganglion (SPG) is a gathering of the cell bodies of parasympathetic fibers that dominate the nasal gland, lacrimal gland and cerebral blood vessels. The SPG controls nasal secretions, tears, and the dilation of cerebral blood vessels. However, it is unclear how serotonin regulates SPG functions. In this study, we investigated the expression of genes involved in the serotonergic system in the mouse SPG. We examined the mRNA expression levels of 5-HT1A, 5-HT1B, 5-HT1D, 5-HT1F, 5-HT2A, 5-HT2B, 5-HT2C, 5-HT3A, 5-HT3B, 5-HT4, 5-HT5A, 5-HT5B, 5-HT6 and 5-HT7 receptors, as well as serotonin transporter, tryptophan hydroxylases 1 and 2, and L-amino acid decarboxylase (AADC) by RT-PCR. It revealed that the 5-HT3A and 5-HT3B ionotropic receptors and AADC were likely to be highly expressed in the SPG, as measured by RT-PCR. We next performed in situ hybridization on the SPG to examine the expression of these three genes at the cellular level after validating the specificity of each cRNA probe by northern blotting. The 5-HT3A receptor, 5-HT3B receptor, and AADC were expressed in 96.5% ±â€¯1.0%, 29.7% ±â€¯10.7%, and 57.4% ±â€¯2.9% of neuronal cell bodies in the SPG, respectively, indicating that the 5-HT3A receptor was virtually expressed in all SPG neurons. Our results on the expression of these critical serotonin system genes in the parasympathetic SPG provide insight into the pathogenetics of rhinitis, conjunctivitis and headache. Furthermore, our findings suggest that targeting the 5-HT3A receptor might have therapeutic potential in the treatment of these ailments.

(+)-Catechin protects dermal fibroblasts against oxidative stress-induced apoptosis.

BACKGROUND: Oxidative stress has been suggested as a mechanism underlying skin aging, as it triggers apoptosis in various cell types, including fibroblasts, which play important roles in the preservation of healthy, youthful skin. Catechins, which are antioxidants contained in green tea, exert various actions such as anti-inflammatory, anti-bacterial, and anti-cancer actions. In this study, we investigated the effect of (+)-catechin on apoptosis induced by oxidative stress in fibroblasts. METHODS: Fibroblasts (NIH3T3) under oxidative stress induced by hydrogen peroxide (0.1 mM) were treated with either vehicle or (+)-catechin (0-100 µM). The effect of (+)-catechin on cell viability, apoptosis, phosphorylation of c-Jun terminal kinases (JNK) and p38, and activation of caspase-3 in fibroblasts under oxidative stress were evaluated. RESULTS: Hydrogen peroxide induced apoptotic cell death in fibroblasts, accompanied by induction of phosphorylation of JNK and p38 and activation of caspase-3. Pretreatment of the fibroblasts with (+)-catechin inhibited hydrogen peroxide-induced apoptosis and reduced phosphorylation of JNK and p38 and activation of caspase-3. CONCLUSION: (+)-Catechin protects against oxidative stress-induced cell death in fibroblasts, possibly by inhibiting phosphorylation of p38 and JNK. These results suggest that (+)-catechin has potential as a therapeutic agent for the prevention of skin aging.

L-arginine stimulates fibroblast proliferation through the GPRC6A-ERK1/2 and PI3K/Akt pathway.

L-arginine is considered a conditionally essential amino acid and has been shown to enhance wound healing. However, the molecular mechanisms through which arginine stimulates cutaneous wound repair remain unknown. Here, we evaluated the effects of arginine supplementation on fibroblast proliferation, which is a key process required for new tissue formation. We also sought to elucidate the signaling pathways involved in mediating the effects of arginine on fibroblasts by evaluation of extracellular signal-related kinase (ERK) 1/2 activation, which is important for cell growth, survival, and differentiation. Our data demonstrated that addition of 6 mM arginine significantly enhanced fibroblast proliferation, while arginine deprivation increased apoptosis, as observed by enhanced DNA fragmentation. In vitro kinase assays demonstrated that arginine supplementation activated ERK1/2, Akt, PKA and its downstream target, cAMP response element binding protein (CREB). Moreover, knockdown of GPRC6A using siRNA blocked fibroblast proliferation and decreased phosphorylation of ERK1/2, Akt and CREB. The present experiments demonstrated a critical role for the GPRC6A-ERK1/2 and PI3K/Akt signaling pathway in arginine-mediated fibroblast survival. Our findings provide novel mechanistic insights into the positive effects of arginine on wound healing.

XLMR protein related to neurite extension (Xpn/KIAA2022) regulates cell-cell and cell-matrix adhesion and migration.

X-linked mental retardation (XLMR) is a common cause of moderate to severe intellectual disability in males. XLMR protein related to neurite extension (Xpn, also known as KIAA2022) has been implicated as a gene responsible for XLMR in humans. Although Xpn is highly expressed in the developing brain and is involved in neurite outgrowth in PC12 cells and neurons, little is known about the functional role of Xpn. Here, we show that Xpn regulates cell-cell and cell-matrix adhesion and migration in PC12 cells. Xpn knockdown enhanced cell-cell and cell-matrix adhesion mediated by N-cadherin and ß1-integrin, respectively. N-Cadherin and ß1-integrin expression at the mRNA and protein levels was significantly increased in Xpn knockdown PC12 cells. Furthermore, overexpressed Xpn protein was strongly expressed in the nuclei of PC12 and 293T cells. Finally, depletion of Xpn perturbed cellular migration by enhancing N-cadherin and ß1-integrin expression in a PC12 cell wound healing assay. We conclude that Xpn regulates cell-cell and cell-matrix adhesion and cellular migration by regulating the expression of adhesion molecules.