ABSTRACT
Objective: To investigate the effect and mechanism of Mailuoning Compound for treatment of early steroid-induced osteonecrosis of femoral head (SONFH) in rats by obturator nerve block. Method: 24 rats were injected with endotoxin 10 μg·kg-1 through tail vein. After 24 hours, prednisolone acetate 20 mg·kg-1 was given by intraperitoneal injection, once every 24 hours for 3 consecutive days. After successful modeling, the rats were randomly divided into the model group (n=12), the treatment group (n=12) and the normal control group (n=6). In the treatment group, 2 mL·kg-1 of Mailuoning compound was injected into the obturator nerve from the 4th day, 3 times a week for 8 weeks. The arterial blood was collected from rats on the first day of the 9th week after model building to detect the content of blood lipid; the femoral head was taken to prepare the paraffin section, and the pathological changes of femoral head was observed and the changes of empty bone lacuna rate, bone trabecular area and bone lacuna area were quantitatively analyzed; The changes of bone morphogenetic proteins(BMPs),transforming growth factor-β1(TGF-β1),vascular endothelial cell growth factor(VEGF),and Ⅷ factor related antigen(Ⅷ-R Ag) were quantitatively analyzed by immunohistochemical method. Result: In the model group, the bone trabeculae were sparse, thin, disorganized and broken; some of the bone cells were necrotic and the number of empty bone lacunae was increased. In the treatment group, the number of trabeculae was increased; the structure was clear, most of which was normal bone cells, with a few necrotic bone cells, and the number of empty bone lacunae was decreased obviously. The rate of empty bone lacuna and the area of bone lacuna in the treatment group were significantly lower than those in the model group (Pβ1 and the microvessel density of Ⅷ-R Ag in the treatment group were significantly higher than those in the model group (PPConclusion: Mailuoning compound can improve the microcirculation state of femoral head, promote the formation of new bone and blood vessel in femoral head by regulating the expression of VEGF, BMPs, TGF-β1, Ⅷ-R Ag and down-regulating blood lipid content, thus effectively controlling the development of early SONFH. This can provide a theoretical basis for the treatment of early SONFH.
ABSTRACT
BACKGROUND: Y-27632 is a Rho-associated coil-formed protein kinase inhibitor that can regulate the self-renewal of stem cells, promote clonal formation and cell survival, and regulate and protect neuronal cell growth and development. How to improve the differentiation efficiency of embryonic stem cells into neuron-like cells is highly important for nerve injury repair and nerve regeneration. OBJECTIVE: To investigate the effect of Y-27632 on the differentiation efficiency and function of human embryonic stem cells into neuron-like cells. METHODS: After resuscitation, human embryonic stem cells at passage 16 were subjected to morphological observation and staining identification. The embryoid bodies were prepared by suspension culture, and after 8 days of incubation, the cells were cultured in Sato medium containing different concentrations of Y-27632 (0, 5, 10, 20, 40 μmol/L) for 10 days and identification by staining. After induction for 18 days, the differentiation efficiency and neurite outgrowth were identified by immunofluorescence staining. RESULTS AND CONCLUSION: The human embryonic stem cells co-expressed Oct4 and SSEA-3 stem cell specific markers. After suspension culture for 8 days and further adherent culture for 10 days, the cells could be differentiated into neuron-like cells with neurogenic morphology and expressing Tuj-1. Y-27632, especially at a concentration of 10 μmol/L, not only promoted cell proliferation (a significant increase in adherent cells an Tuj-1 positive cells), but also facilitated cell differentiation into neurons. Immunofluorescence staining findings showed that 10 μmol/L Y-27632 significantly increased the number of Tuj-1 positive cells and neurites and the length of neurites after 18 days of differentiation. These results indicate that Y-27632 not only promotes the differentiation of human embryonic stem cells into neuron-like cells, but also accelerates neurite outgrowth.
ABSTRACT
<p><b>OBJECTIVE</b>To observe the impact of adenosine A1 receptor stimulation on extracellular signal-regulated kinase 1/2 (ERK1/2) signal pathways on angiotensin II (AngII) stimulated cardiomyocytes of neonatal rats in vitro.</p><p><b>METHODS</b>Cardiomyocytes of neonatal rats were cultured in vitro. Cardiomyocytes hypertrophy was induced by AngII (0.1 µmol/L). The antihypertrophic effect of adenosine A1 receptor stimulation via adenosine A1 receptor agonist R-PIA (1 µmol/L) was observed in the presence or absence of ERK1/2 inhibitor 1, 4-Diamino-2, 3-dicyano-1, 4-bis(o-aminophenylmercapto) butadiene (U0126) 1 µmol/L, PKC inhibitor Ro-31-8220 (50 nmol/L), and pertussis toxin (PTX, 5 mg/L). The total protein content was assayed by the method of Lowry. The expression of mRNA of atrial natriuretic peptide (ANP) was determined by RT-PCR. [Ca(2+)]i was measured by confocal microscope using Fluo-3/AM as fluorescent indicator. The relative expression of ERK1/2 was determined by Western blot.</p><p><b>RESULTS</b>Compared with normal control group, AngII induced significant cardiomyocyte hypertrophy. Compared with AngII group, R-PIA significantly inhibited AngII-induced increase of the protein content, cardiomyocytes volume and expression of ERK1/2, calcium ion fluorescence intensity, similar as U0126 and Ro-31-8220. The inhibitory effects on AngII induced cardiomyocytes hypertrophy of R-PIA were lost when preincubated with PTX.</p><p><b>CONCLUSION</b>Adenosine A1 receptor can inhibit AngII induced cardiomyocyte hypertrophy through downregulating ERK signal pathways and reducing intracellular Ca(2+).</p>
Subject(s)
Animals , Female , Male , Rats , Adenosine A1 Receptor Agonists , Pharmacology , Angiotensin II , Pharmacology , Calcium , Metabolism , Cardiomegaly , Cells, Cultured , MAP Kinase Signaling System , Myocytes, Cardiac , Metabolism , Pathology , Rats, Sprague-Dawley , Receptor, Adenosine A1 , MetabolismABSTRACT
Objective To demonstrate the inhibitory effect of adenosine A_1 receptor agonist R(-)-N6-(2-phe- nylisopropyl) adenosine (R-HA) and cross-talk between adenosine A_1 receptor and CaMKII on Isoproterenol (Iso)- induced hypertrophy in cultured myocardial cells in neonatal rats.Methods The protein synthesis was determined by incorporation of [~3H]-leucine into myocyte protein.The expression of CaMK Ⅱ ? B was determined by Western- blot.The [Ca~(2+)]i transient was measured in myocytes loaded with fura-2 by the spectrofluorometric method. Results R-PIA (1?mol/L) inhibited Iso(10 ?mol/L)-induced increase of [~3 H-]-leucine incorporation [(R-PIA: 974.8?58.6) vs (Iso:1220.8?240.5) count per min per well,P
ABSTRACT
<p><b>AIM</b>In order to study the effects of kappa-opioid receptor activation, we used cultured cardiomyocytes to study the inhibitory effects of U50,488H on cellular proliferation, protein content in the presence or absence of nor-binaltorphimine (nor-BNI).</p><p><b>METHODS</b>The cellular proliferation was determined with crystal violet staining and the protein content was assayed with Lowry's method.</p><p><b>RESULTS</b>A kappa-opioid receptor agonist U50,488H at 0.1 micromol/L-10 micromol/L inhibited the cellular proliferation and protein content of cultured myocardial cells in a dose-dependent manner. The inhibitory effects of U50,488H were completely blocked by pretreatment with nor-BNI, a specific kappa-opioid receptor antagonist at 1 micromol/L.</p><p><b>CONCLUSION</b>U50,488H inhibited the cultured myocardial cells' growth. The inhibitory effects of U50,488H are involved in mediating the action of kappa-opioid receptor stimulation.</p>