Abnormal expression and role of MicroRNA-214-3p/SLC8A1 in neonatal Hypoxic-Ischaemic encephalopathy.

Neonatal hypoxic-ischaemic encephalopathy (HIE) refers to brain damage caused by intra-uterine distress and asphyxia/hypoxia during the perinatal and neonatal periods. MicroRNA (MiR)-214-3p plays a critical role in cell growth and apoptosis. The aim of this study was to investigate the expression and role of miR-214-3p in neonatal HIE development, and to explore the underlying mechanisms. The expression of miR-214-3p was significantly down-regulated, while that of Slc8a1, a direct target of miR-214-3p, was significantly up-regulated, in the brain tissue of neonatal HIE rats. The over-expression of miR-214-3p promoted the proliferation and inhibited the apoptosis of neurones, while its down-regulation had the opposite effect. Our results indicate that miR-214-3p expression was down-regulated in neonatal HIE rats, and the up-regulation of miR-214-3p expression protected against HIE development by inhibiting neuronal apoptosis.

LncRNA MAFG-AS1-induced acute myeloid leukemia development via modulating miR-147b/HOXA9.

Recent references discovered that lncRNAs acted roles in malignant cancer development. However, the role of MAFG-AS1 in acute myeloid leukemia (AML) development remains unknown. MAFG-AS1 and miR-147b were determined in AML cells and specimens using qRT-PCR assay. Cell proliferation was detected by CCK-8 analysis and flow cytometry was carried out to measure cell cycle. Luciferase reporter analysis was done to determine the mechanism of MAFG-AS1 and miR-147b. We noted that MAFG-AS1 was overexpressed in AML cells and in serum and bone narrow from AML compared with normal controls specimen. Elevated expression of MAFG-AS1 increased cell growth, cycle and EMT in AML cell HL-60 cell. MAFG-AS1 sponged miR-147b expression in HL-60 cell. Moreover, miR-147b was downregulated in AML cells and in serum and bone narrow from AML compared with normal control specimen. miR-147b was negatively correlated with MAFG-AS1 in the serum and bone narrow of AML cases. We illustrated that HOXA9 was one target of miR-147b and ectopic expression of MAFG-AS1 enhanced HOXA9 expression HL-60 cell. Forced expression of MAFG-AS1 induced cell growth, cycle, and EMT via promoting HOXA9. These data illustrated that MAFG-AS1 acted as one oncogenic gene and accelerated AML progression via modulating miR-147b/HOXA9 axis.

Magnet-targeted delivery of bone marrow-derived mesenchymal stem cells improves therapeutic efficacy following hypoxic-ischemic brain injury.

Stem cell transplantation may represent a feasible therapeutic option for the recovery of neurological function in children with hypoxic-ischemic brain injury; however, the therapeutic efficacy of bone marrow-derived mesenchymal stem cells largely depends on the number of cells that are successfully transferred to the target. Magnet-targeted drug delivery systems can use a specific magnetic field to attract the drug to the target site, increasing the drug concentration. In this study, we found that the double-labeling using superparamagnetic iron oxide nanoparticle and poly-L-lysine (SPIO-PLL) of bone marrow-derived mesenchymal stem cells had no effect on cell survival but decreased cell proliferation 48 hours after labeling. Rat models of hypoxic-ischemic brain injury were established by ligating the left common carotid artery. One day after modeling, intraventricular and caudal vein injections of 1 × 105 SPIO-PLL-labeled bone marrow-derived mesenchymal stem cells were performed. Twenty-four hours after the intraventricular injection, magnets were fixed to the left side of the rats' heads for 2 hours. Intravoxel incoherent motion magnetic resonance imaging revealed that the perfusion fraction and the diffusion coefficient of rat brain tissue were significantly increased in rats treated with SPIO-PLL-labeled cells through intraventricular injection combined with magnetic guidance, compared with those treated with SPIO-PLL-labeled cells through intraventricular or tail vein injections without magnetic guidance. Hematoxylin-eosin and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining revealed that in rats treated with SPIO-PLL-labeled cells through intraventricular injection under magnetic guidance, cerebral edema was alleviated, and apoptosis was decreased. These findings suggest that targeted magnetic guidance can be used to improve the therapeutic efficacy of bone marrow-derived mesenchymal stem cell transplantation for hypoxic-ischemic brain injury. This study was approved by the Animal Care and Use Committee of The Second Hospital of Dalian Medical University, China (approval No. 2016-060) on March 2, 2016.

The effect of magnetic guiding BMSCs on hypoxic-ischemic brain damage via magnetic resonance imaging evaluation.

Hypoxic-ischemic brain damage (HIBD) is a critical disease in pediatric neurosurgery with high mortality rate and frequently leads to neurological sequelae. The role of bone marrow mesenchymal stem cells (BMSCs) in neuroprotection has been recognized. However, using the imaging methods to dynamically assess the neuroprotective effects of BMSCs is rarely reported. In this study, BMSCs were isolated, cultured and identified. Flow cytometry assay had shown the specific surface molecular markers of BMSCs, which indicated that the cultivated cells were purified BMSCs. The results demonstrated that CD29 and CD90 were highly expressed, whilst CD45 and CD11b were negatively expressed. Further, BMSCs were transplanted into Sprague Dawley (SD) rats established HIBD via three ways, including lateral ventricle (LV) injection, tail vein (TV) injection, and LV injection with magnetic guiding. Magnetic resonance imaging (MRI) was used to monitor and assess the treatment effect of super paramagnetic iron oxide (SPIO)-labeled BMSCs. The mean kurtosis (MK) values from diffusion kurtosis imaging (DKI) exhibited the significant differences. It was found that the MK value of HIBD group increased compared with that in Sham. At the meantime, the MK values of LV + HIBD, TV + HIBD and Magnetic+LV + HIBD groups decreased compared with that in HIBD group. Among these, the MK value reduced most significantly in Magnetic+LV + HIBD group. MRI illustrated that the treatment effect of Magnetic+LV + HIBD group was best. In addition, HE staining and TUNEL assay measured the pathological changes and apoptosis of brain tissues, which further verified the MRI results. All data suggest that magnetic guiding BMSCs, a targeted delivery way, is a new strategic theory for HIBD treatment. The DKI technology of MRI can dynamically evaluate the neuroprotective effects of transplanted BMSCs in HIBD.

LncRNA SNHG6 Promotes Wilms' Tumor Progression Through Regulating miR-429/FRS2 Axis.

Background: Long noncoding RNA (lncRNA) small nucleolar RNA host gene 6 (SNHG6) has been reported to be an oncogene in a variety of cancers. However, the role of SNHG6 and its associated mechanisms in Wilms' tumor progression remain largely unknown. Methods: The expression of SNHG6, microRNA-429 (miR-429), and FGF receptor substrates 2 (FRS2) messenger RNA (mRNA) was detected by quantitative real-time polymerase chain reaction. Cell proliferation was analyzed through 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and plate colony assay. The apoptosis was assessed by flow cytometry. Cell glycolytic metabolism was analyzed through detecting the lactate dehydrogenase activity, glucose uptake, lactate production, and ATP level. The target relationship between miR-429 and SNHG6 or FRS2 was predicted by miRcode or Starbase and then validated by dual-luciferase reporter assay and RNA pull-down assay. Murine xenograft model was established to validate the function of SNHG6 in vivo. Results: The level of SNHG6 was elevated in Wilms' tumor tissues and cells, and SNHG6 played an oncogenic role to promote the proliferation and glycolysis and restrain the apoptosis of Wilms' tumor cells. MiR-429 was identified as a target of SNHG6, and miR-429 interference partly reversed the inhibitory effects induced by SNHG6 silencing on the malignant behaviors of Wilms' tumor cells. FRS2 mRNA bound to miR-429 in Wilms' tumor cells. SNHG6 upregulated the expression of FRS2 through acting as a sponge of miR-429. MiR-429-induced influences in Wilms' tumor cells were largely counteracted by the overexpression of FRS2. SNHG6 silencing suppressed the Wilms' tumor growth through miR-429/FRS2 axis in vivo. Conclusion: SNHG6 accelerated Wilms' tumor progression through regulating miR-429/FRS2 signaling in vitro and in vivo.

Alteration of Connexin43 expression in a rat model of obesity-related glomerulopathy.

It is accepted that alteration of connexin43 (Cx43) expression in glomeruli is a common pathological response in several forms of kidney diseases. To date, however the change of the Cx43 expression in obesity-related glomerulopathy (ORG) has not been reported. In this study, the alteration of Cx43 expression in the glomeruli of rat with ORG was defined. Five-week-old rats were fed with high-fat diet for 18weeks to establish the ORG model, then the histological change of glomeruli, the foot process effacement of podocyte, the markers for podocyte injury (nephrin,podocin and WT1) and Cx43 expression in glomeruli were examined respectively. The results demonstrated metabolic disorder, hyperinsulinemia, systemic inflammation and microalbuminuria in ORG rats. There was significant hypertrophy, glomerular expansion and inflammatory cell infiltration in the kidney of ORG rats compared to the control group. Significant foot process effacement of the podocyte in the glomeruli, nephrin loss and density reduction were shown in the ORG rats, and Cx43 expression was significant upregulated in glomeruli of ORG rats compared to the control group. The results indicate the correlation of overexpressed Cx43 with the obesity related renal inflammation and suggest that Cx43 might be a potential target in the development of obesity related glomerulopathy.

Knockdown of Tlr4 in the Arcuate Nucleus Improves Obesity Related Metabolic Disorders.

High-fat diet-induced hypothalamic metabolic inflammation is emerging as a cause for the development of obesity. It is acknowledged that Toll-like receptor4 (TLR4) signaling plays a crucial role in triggering of the hypothalamic metabolic inflammation during the course of diet-induced obesity. Whether hypothalamic arcuate nucleus (ARC)-restricted TLR4 knockdown improves obesity-related metabolic disorders remains unexplored. In this study, we used TLR4 shRNA lentiviral particles to suppress the TLR4 expression in the hypothalamic ARC of diet-induced obese rat model by stereotaxic injection. Our results demonstrate that ARC-restricted TLR4 knockdown protects obese rats from diet-induced weight gain and energy intake, from diet-induced impaired glucose homeostasis and peripheral insulin resistance, and from high-fat diet-induced hepatic steatosis and adipocyte hypertrophy. Thus, we define ARC-restricted TLR4 knockdown as a potential strategy to combat metabolic disorders associated with obesity.

Obesity increases the risk of renal involvement in children with Henoch-Schönlein purpura.

UNLABELLED: The main aim of this study was to evaluate the relationship between obesity and renal involvement in children with Henoch-Schönlein purpura (HSP). A retrospective study of 141 pediatric patients with HSP was conducted in our hospital. The clinical data of all patients were collected from the electronic medical record management system from January 2010 to June 2014. The possible risk factors of renal involvement, especially obesity, were analyzed using univariate and multivariate analyses. Renal involvement occurred in 45/141 of the patients. A univariate analysis showed that an age more than 7 years at onset, persistent purpura, obesity, time from symptoms onset to diagnosis more than 14 days, and decreased C3 all increased the risk of renal involvement in HSP. The forward stepwise logistic regression analysis indicated obesity (odds ratio (OR) 4.43, 95 % confidence interval (CI) 1.896 to 10.358), age more than 7 years at onset (OR 2.81, 95 % CI 1.142 to 6.907), and persistent purpura (OR 2.57, 95 % CI 1.119 to 5.909) were independent risk factors for renal involvement. CONCLUSIONS: Our results show that obesity can increase the hazard of renal involvement in children with HSP and reconfirm that older age at onset and persistent purpura are the independent risk factors for renal involvement. WHAT IS KNOWN: • There have been some reports that obesity was associated with the development of renal injury. • It is not clear whether obesity can increase the risk of renal involvement in children with HSP. What is New: • The main finding of this study is that obesity can increase the hazard of renal involvement in children with HSP.

Aquaporin-4 knockdown ameliorates hypoxic-ischemic cerebral edema in newborn piglets.

Emerging evidence indicates that the water channel protein aquaporin 4 (AQP4) plays an essential role in water homeostasis and is implicated in the pathogenesis of brain edema. This study aimed to understand the physiological role of AQP4 in hypoxia-ischemia-mediated cytotoxic brain edema. We specifically knocked down AQP4 expression by intracerebral injection of a plasmid containing AQP4 siRNA into a neonatal piglet model. The success of the hypoxia-ischemia-induced piglet model was confirmed by conventional magnetic resonance imaging and diffusion-weighted imaging. AQP4 knockdown led to reduced brain edema accompanied by a higher apparent diffusion coefficient value, compared to the control group injected with a plasmid containing scrambled siRNA. Real-time polymerase chain reaction and immunohistochemical analysis confirmed that AQP4 siRNA significantly reduced AQP4 mRNA and protein expression. Finally, neurological function analysis revealed that AQP4 knockdown significantly improved neurobehavioral manifestation of the piglets after exposure to hypoxia-ischemia. Taken together, these results indicate that AQP4 plays an important role in mediating brain edema in hypoxic-ischemic encephalopathy. Therefore, AQP4 could be a therapeutic target to ameliorate early-stage brain edema.

The application of nasal synchronized intermittent mandatory ventilation in primary apnea of prematurity.

We aimed to evaluate the efficacy of nasal synchronized intermittent mandatory ventilation (nSIMV) in preterm infants with primary apnea of prematurity (AOP). Forty-four preterm infants with AOP were divided into the nSIMV group or nasal continuous positive airway pressure (nCPAP) group. Clinical symptoms, signs and blood gas results following nSIMV or nCPAP were compared between the two groups. Infants receiving nSIMV had a greater reduction in apneic spells and a greater decrease in bradycardia than those receiving nCPAP. Compared with the nCPAP group, the nSIMV group had a lower incidence of respiratory support failure (9.1% vs. 27.3%; p<0.05), a lower incidence of hypercarbia (4.5% vs. 18.2%; p<0.05) and a lower incidence of gastrointestinal complications (4.5% vs. 13.6%; p<0.05). This study showed that nSIMV was more effective in respiratory support in preterm infants with AOP.

[Risk factors for recurrent pneumonia in children: a case-control study].

OBJECTIVE: To study the risk factors for recurrent pneumonia in children without underlying diseases. METHODS: A case-control study was conducted in 106 children with recurrent pneumonia (case group) and 106 age, gender- and weight-matched children with pneumonia but no recurrence (control group). The children in both groups had no underlying disease. The risk factors for recurrent pneumonia were investigated by the Chi-Square analysis and the multivariate logistic regression model. RESULTS: The Chi-Square analysis showed that the percentages with the history of wheezing, allergy (food or medicine) and eczema and the percentage of transient neutropenia in the case group were significantly higher than those in the control group. The multivariate logistic regression analysis showed that the wheezing history (OR=13.387, 95% CI: 5.541-32.343), allergic history (food or medicine) (OR=4.267, 95% CI: 2.081-8.751) and transient neutropenia (OR=3.606, 95% CI: 1.806-7.202) were the independent risk factors of recurrent pneumonia. CONCLUSIONS The wheezing history, allergic history and transient neutropenia may increase the risk of recurrence of pneumonia in pneumonic children without underlying diseases.

Effects of 11beta-hydroxysteroid dehydrogenase inhibitor on body weight and glucose tolerance in Sprague-Dawley rats fed with a high-fat diet.

OBJECTIVE: Many studies have shown that glucocorticoids play a crucial role in the development of obesity and insulin resistance. This study investigated the therapeutic effects of long-term inhibition of glucocorticoid activity on obesity and insulin resistance. METHODS: Four-week-old male Sprague-Dawley (SD) rats were randomly fed with a high-fat diet (fat content accounting for 20% of total calorie) (control group, n=8) or with a high-fat diet along with glycyrrhetic acid (GE, 800 mg/L), an inhibitor of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) for 24 weeks (GE-treated group, n=9). The body weights and the amount of food intake were monitored weekly and daily, respectively. After 24 weeks of GE treatment, oral glucose tolerance tests were performed. Blood glucose was measured by glucose oxidase method. The levels of plasma glucocorticoids, insulin and leptin were measured with radioimmunoassay. The levels of serum cholesterol and triglyceride were determined with an automatic measuring analyzer. RESULTS: The food intake amount decreased significantly in the GE-treated group from 6 weeks and body weight gain was markedly less from 8 weeks after GE administration compared with the control group. After 24 weeks of treatment, the plasma levels of leptin and insulin in GE-treated rats were significantly reduced compared with the control group. The serum levels of cholesterol and triglyceride decreased markedly compared with the control group and the levels of blood glucose were significantly lower 15, 30, 60 and 120 minutes after oral glucose load in the GE-treated group compared with the control group. CONCLUSIONS: Long-term GE treatment may contribute to resisting diet-induced obesity and insulin resistance.

[Effects of D-limonene on leukemia cells HL-60 and K562 in vitro].

To investigate the effects of D-limonene (D-L) on the cell growth and apoptosis in HL-60, K562 cells and to elucidate its mechanism, the influence of D-L on proliferation of HL-60 and K562 cells was determined by propidium iodide assay, the expression levels of mutant p53, bcl-2, bax gene were detected by cell morphological analysis, flow cytometry and immunohistochemistry staining, the D-L-inducing HL-60 and K562 cell apoptosis in vitro was observed systematically. The results showed that D-L inhibited HL-60 and K562 cell growth in a dose- and time-dependent manner with the IC50 of 0.75 mmol/L similarly, D-L induced apoptosis of HL-60 and K562 cells, and expression of bcl-2 gene was down regulated by D-L in a concentration-dependent manner in HL-60 cells. The bcl-2, mutant type of p53 genes were down regulated while bax gene was up regulated by D-L in a concentration-dependent manner in K562 cells. It is concluded that D-L can inhibit proliferation and induce apoptosis of HL-60 and K562 cells. The bcl-2, mutant type of p53 and bax may be involved in the gene regulation of D-L-induced apoptosis.

[Effects of leptin and insulin on leptin receptors mRNA expression in human hepatocellular HepG2 cells].

OBJECTIVE: To study the leptin receptor isoforms regulation by leptin and insulin. METHODS: Human hepatocellular carcinoma cells of the line HepG2 were cultured in DMEM containing 10% FBS in six-well plate and were incubated for 24 hours in serum-free medium containing 0, 10(-9), 10(-8), 10(-7), and 10(-6) mol/L of human leptin or insulin. Using the semi-quantitative RT-PCR technique, the mRNA expressions of long (OB-Rb) and short (OB-Ra: OB-R219.3) leptin receptor isoforms were measured. RESULTS: OB-Rb and OB-R219.3 mRNAs were expressed in this cell line. Leptin of the concentrations of 10(-7) approximately 10(-6) mol/L significantly inhibited the OB-Rb mRNA expression, with the maximum decrease (by 43%) at the concentration of 10(-6) mol/L. Similarly the mRNA expression of OB-R219.3 was also markedly reduced in cells treated with leptin of the concentrations of 10(-8) approximately 10(-6) (mol/L), with the maximum inhibition (by 49%) at the concentrations of 10(-6) mol/L. Insulin showed no effect on OB-Rb and OB-R219.3 mRNAs expression in HepG2 cell. CONCLUSION: In HepG2 cells, leptin down-regulates the expressions of OB-Rb and OB-R219.3 mRNAs, and insulin has no effect on OB-Rb and OB-R219.3 mRNAs, which contributes at least partly to an understanding of the mechanism of leptin resistance in vivo and suggests that leptin-induced receptor down-regulation may be relevant to leptin resistance at sites of peripheral action.

Effects of leptin and dexamethasone on long and short leptin receptor mRNA.

BACKGROUND: Leptin exerts its effects by using both long (OB-Rb) and short (OB-Ra) receptors. Although leptin resistance or insensitivity is reported to be a cause of human obesity, there have not been enough studies to clarify regulation of leptin receptors. METHODS: The authors studied leptin receptor regulation by leptin and dexamethasone in the human hepatocellular carcinoma cell (HepG2). Using a quantitative RT-PCR technique, the authors demonstrate expression of OB-Ra and OB-Rb mRNA after the incubation for 24 h with leptin or dexamethasone at various concentrations (10(-9)-10(-6) M). RESULTS: Leptin (10(-7)-10(-6) M) significantly inhibited expression of OB-Rb mRNA, with maximum inhibition (43% of control) at 10(-6) M. Expression of OB-R219.1 and OB-R219.3, two short isoforms of leptin receptor, were also reduced in cells treated with leptin most remarkably at 10(-7) M for OB-R219.1 (44% of the control) and at 10(-6) M for Ob-R219.3 (49% of the control). In contrast, dexamethasone (10(-8)-10(-6) M) significantly increased OB-Rb mRNA levels, with a maximum increase (204% of the control) at 10(-7) M, and OB-R219.1 and OB-R219.3 mRNA expression was also markedly increased at 10(-9)-10(-6) M. The peak values were 254% of the control for OB-R219.1 and 246% of the control for OB-R219.3 at 10(-7) M. CONCLUSIONS: In HepG2 cells, leptin inhibits and dexamethasone increases OB-Ra and OB-Rb mRNA expression. It is suggested that glucocorticoids as well as leptin itself contribute to regulatory effects of leptin through changes in gene expression of leptin receptors.