Effective doses of midazolam oral solution for the prevention of preoperative anxiety in paediatric patients.

BACKGROUND: More than 60% children exhibit anxiety before undergoing an anesthetic-surgical procedure, particularly among pre-school paediatric patients. Oral midazolam can provide procedural sedation for children prior to anesthesia. However, extemporaneous solutions of midazolam are usually prepared from injectable drug solutions, leading to inconsistent efficacy due to variable preparation methods. Xiaoerjing® is the first commercially available oral formulation of midazolam for procedural sedation in children in China. Despite the recommended dosage range of 0.25-0.5 mg/kg, its effective dose is still largely unknown. AIM: To determine the 95% effective dose (ED95) of midazolam oral solution (Xiaoerjing®) for alleviating preoperative anxiety in children prior to mask induction of general anesthesia. DESIGN: The study included 61 children between the ages of 1 and 6 years undergoing elective surgery under general anesthesia. The first patient received a single dose of 0.5 mg/kg midazolam oral solution, which was adjusted for subsequent patients using the biased coin design method based on their response to the previous dose. Doses were increased or decreased at the rate of 0.1 mg/kg. An effective response was defined as having a modified Ramsay sedation score ≥3a, separation anxiety score ≤2, and mask acceptance score ≤2 during inhalational anesthesia induction. RESULTS: Fifty-six children were included in the final analysis. Of those, sedation was successful in 50 patients, with a median separation time of 15 (IQR: 25) min. Midazolam oral solution has an ED95 of 0.8254 mg/kg (95% CI: 0.6915-0.8700 mg/kg) for relieving preoperative anxiety in children. No adverse events occurred following drug administration. CONCLUSION: Midazolam oral solution is a safe and effective medication for relieving preoperative anxiety in children. The ED95 of a single oral dose of midazolam oral solution is 0.8254 mg/kg (95% CI: 0.6915-0.8700 mg/kg).

Combined over-expression of the hypoxia-inducible factor 2α gene and its long non-coding RNA predicts unfavorable prognosis of patients with osteosarcoma.

BACKGROUND: LncRNA hypoxia-inducible factor-2α (HIF-2α) promoter upstream transcript (HIF2PUT) functions as a novel regulatory factor of osteosarcoma stem cells partly by controlling HIF-2α expression. The aim of this study was to investigate the clinical significance of HIF-2α and HIF2PUT in human osteosarcoma. MATERIALS AND METHODS: Quantitative real-time PCR was performed to detect the expression levels of HIF-2α mRNA and HIF2PUT in 82 surgical specimens of primary osteosarcoma and matched non-cancerous bone tissues. Then, the associations of HIF-2α and/or HIF2PUT expression with various clinicopathological features of osteosarcoma patients were statistically analyzed. Moreover, their prognostic value was further evaluated. RESULTS: Compared with non-cancerous bone tissues, HIF-2α mRNA and HIF2PUT expression were both significantly upregulated in osteosarcoma tissues (all P<0.001). Interestingly, the expression levels of HIF-2α mRNA in osteosarcoma tissues were positively correlated with those of HIF2PUT (r=0.28, P=0.009). Additionally, osteosarcoma patients with HIF-2α mRNA and/or HIF2PUT over-expression more frequently had large tumor size (all P<0.05), advanced clinical stage (all P<0.01) and positive distant metastasis (all P<0.01). Moreover, osteosarcoma patients with HIF-2α mRNA and/or HIF2PUT over-expression had a significantly shorter overall and disease-free survival (all P<0.05). Furthermore, Cox multivariate analysis identified that HIF-2α mRNA and/or HIF2PUT expression, clinical stage and distant metastasis were all independent and significant prognostic factors for both overall and disease-free survival (all P<0.05). CONCLUSIONS: HIF-2α and HIF2PUT upregulation may be a common feature in human osteosarcomas with aggressive potency. The over-expression of the two molecules, alone or combined, may predict poor prognosis in osteosarcoma patients.

Uremia-caused changes of ghrelin system in hippocampus may be associated with impaired cognitive function of hippocampus.

PURPOSE: Hippocampus plays an important role in spatial learning and memory. Ghrelin, a brain-gut peptide, participates in the mnestic functions of hippocampus through its receptor growth hormone secretagogue receptor (GHS-R) distributed in hippocampus. This study was to investigate whether there was a correlation between the changes of ghrelin system in hippocampus and the spatial cognitive impairment caused by chronic renal failure (CRF). METHODS: Sprague-Dawley rats (male, 180 ± 10 g, 7-8 weeks old) were randomly classified into CRF group and control group (n = 18 per group). The CRF model was constructed by 5/6 nephrectomy and the controls treated with sham operation. By the 8th week after the surgery, the spatial cognitive function of rats was assessed by Morris water-maze test (MWM), the protein expression of ghrelin and GHS-R in the hippocampus by immunohistochemistry, and the mRNA expression by real-time PCR. Statistical analysis was performed using ANOVA, Student-Newman-Keuls-q test and Pearson correlation analysis, and P < 0.05 was considered significant. RESULTS: Compared with the controls, the time spent in "platform" quadrant (TSPQ) of rats with CRF was decreased, but the escape latency (EL) was increased significantly in MWM, and meanwhile the protein and mRNA expression of ghrelin and GHS-R in hippocampus was also increased significantly (P < 0.05 or P < 0.01). Correlation analysis suggested that the TSPQ was negatively but the EL was positively correlated with the mRNA expression of ghrelin and GHS-R (P < 0.01). CONCLUSION: The CRF-caused changes of ghrelin system in hippocampus might be correlated with the CRF-caused cognitive function impairment.

Tea polyphenols may attenuate the neurocognitive impairment caused by global cerebral ischemia/reperfusion injury via anti-apoptosis.

BACKGROUND/AIMS: Global cerebral ischemia/reperfusion (GCIR) may incur neurocognitive impairment. Tea polyphenols (TP) have strong anti-oxidant capacity. This study planned to investigate the protective effect of TP against the neurocognitive impairment caused by GCIR and its mechanism. METHODS: One-stage anterior approach for cerebral four-vessel occlusion (4VO) was used to construct the GCIR model. Sprague Dawley rats were randomly classified into Sham group, GCIR group, and TP group (n = 50 per group). Besides receiving the same 4VO, the rats in TP group were treated with TP (6.4%) injection from the tail vein 30 minutes before cerebral ischemia. Morris water-maze test was used to evaluate the changes in space recognition and memory and open field activity test to assess the activity and motor function of rats. The cell apoptotic study in hippocampal CA1 region at specified time points (12, 24, 48, and 72 hours after surgery) was carried out by the flow cytometry, histology (hematoxylin and eosin staining), and immunohistochemical (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining) examinations. One-way analysis of variance and least significant difference t-test were used and statistical significance considered at P < 0.05. RESULTS: Compared with the GCIR group, the TP group was significantly attenuated in the impairment of space recognition and memory caused by GCIR and so was the neuronal apoptosis in the hippocampal CA1 region (P < 0.05). CONCLUSION: TP may attenuate the impairment of space recognition and memory caused by GCIR via anti-apoptosis.

Identification of the potential molecular targets for human intervertebral disc degeneration based on bioinformatic methods.

The present study aimed to explore potential molecular targets and gain further insights into the mechanism of intervertebral disc degeneration (IDD) progression. Microarray datasets of GSE19943, GSE15227 and GSE34095 were downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) in 3 IDD specimens compared with 3 controls in GSE34095, DEGs in 7 grade III and 3 grade IV samples compared with 5 grade II samples in GSE19943, and differentially expressed miRNAs in 3 degenerated samples compared with 3 controls in GSE15227 were screened. Grade III­ and IV­specific networks were constructed and grade­specific genes were extracted. The network features were analyzed, followed by Gene Ontology (GO) enrichment analysis and pathway enrichment analysis of grade­specific genes and DEGs identified in GSE34095. Furthermore, miRNA­pathway interactions were analyzed using Fisher's exact test. Tumor protein p53 (TP53) was a hub gene in the grade III­specific network and ubiquitin C (UBC) was identified to be a hub gene in the grade IV­specific network. Six significant features were identified by grade­specific network topology analysis. Grade­specific genes and DEGs were involved in different GO terms and pathways. Differentially expressed miRNAs were identified to participate in 35 pathways, among which 6 pathways were significantly enriched by DEGs, including apoptosis. The present study identified that key genes (TP53 and UBC) and miR­129­5p may participate in the mechanism of IDD progression. Thus, they may be potential therapeutic targets for IDD.

Neuroprotective effects of quercetin in a mouse model of brain ischemic/reperfusion injury via anti-apoptotic mechanisms based on the Akt pathway.

The present study provided experimental evidence for the neuroprotective effects of quercetin using a rat model of global brain ischemic/reperfusion (I/R) injury. Pre­treatment with quercetin (5 or 10 mg/kg orally (p.o.); once daily) induced a dose­dependent reduction in I/R­induced hippocampal neuron cell loss, with 10 mg/kg/day being the lowest dose that achieved maximal neuroprotection. Administration of 10 mg/kg quercetin over at least 3 days prior to I/R was required to improve the survival rate of I/R rats. Fluorescence­assisted cell sorting, hematoxylin and eosin staining and terminal deoxynucleotidyl transferase dUTP nick end labeling indicated neuronal cell loss in the CA1 hippocampus. Rats that had undergone transient global cerebral ischemia for 15 min followed by 1 h of reperfusion exhibited a significant increase in reactive oxygen species (ROS) production in the hippocampus. The I/R­induced ROS overproduction in the hippocampus at 1, 12 and 24 h following I/R was significantly decreased by quercetin pre­treatment. Western blot analysis revealed that the neuroprotective effects of quercetin (5 and 10 mg/kg/day, p.o.) were associated with an upregulation of the I/R­induced suppression of B­cell lymphoma­2 (Bcl­2), Bcl extra large and survivin expression as well as phosphorylation of Bcl­2­associated death promoter. Furthermore, the neuroprotective effects of quercetin (5, 10 mg/kg/day) in the brain were associated with an upregulation of Akt signaling. These findings suggested that the inhibition of I/R­induced brain injury by quercetin likely involves a transcriptional mechanism to enhance anti­apoptotic signaling.

Baicalein reduces the invasion of glioma cells via reducing the activity of p38 signaling pathway.

Baicalein, one of the major flavonids in Scutellaria baicalensis, has historically been used in anti-inflammatory and anti-cancer therapies. However, the anti-metastatic effect and related mechanism(s) in glioma are still unclear. In this study, we thus utilized glioma cell lines U87MG and U251MG to explore the effect of baicalein. We found that administration of baicalein significantly inhibited migration and invasion of glioma cells. In addition, after treating with baicalein for 24 h, there was a decrease in the levels of matrix metalloproteinase-2 (MMP-2) and MMP-9 expression as well as proteinase activity in glioma cells. Conversely, the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2 was increased in a dose-dependent manner. Moreover, baicalein treatment significantly decreased the phosphorylated level of p38, but not ERK1/2, JNK1/2 and PI3K/Akt. Combined treatment with a p38 inhibitor (SB203580) and baicalein resulted in the synergistic reduction of MMP-2 and MMP-9 expression and then increase of TIMP-1 and TIMP-2 expression; and the invasive capabilities of U87MG cells were also inhibited. However, p38 chemical activator (anisomycin) could block these effects produced by baicalein, suggesting baicalein directly downregulate the p38 signaling pathway. In conclusion, baicalein inhibits glioma cells invasion and metastasis by reducing cell motility and migration via suppression of p38 signaling pathway, suggesting that baicalein is a potential therapeutic agent for glioma.

Premature senescence and cellular phenotype transformation of mesangial cells induced by TGF-B1.

BACKGROUND: Transforming growth factor-ß1 (TGF-ß1) is a polypeptide member of the transforming growth factor ß superfamily of cytokines and performs many cellular functions. Its overexpression may lead to renal fibrosis. AIM: This study planed to investigate the effects of TGF-ß1 on the cell cycle and phenotype of mesangial cells. METHODS: Rat mesangial cells were cultured together with different concentrations (0, 1, 2, 5, and 10 ng/mL) of TGF-ß1 for specified times from 0 min to 72 h. 0 ng/mL TGF-ß1 and 0 min served as controls. Cell cycles were assessed by flow cytometry and α-smooth muscle actin expression (α-SMA) protein expression by western blot analysis. All data were presented as Mean ± SD. Statistical analysis was performed by using one-way analysis of variance and correlation analysis. Results were considered significant at p < 0.05. RESULTS: After 15 min of co-culture with different concentrations of TGF-ß1, the percentage of mesangial cells in G0/G1 phase was significantly elevated compared to the control (p < 0.05). 12 h co-culture induced cell hyperplasia, 24 h co-culture obvious up-regulation of α-SMA (p < 0.01) and one or two cells' myofibroblast phenotype transition, and 36 h co-culture several cells' phenotype transition. Correlation analysis prompted that the TGF-ß1-induced premature aging was time-dependent (p < 0.01). CONCLUSION: TGF-ß1 may induce mesangial cells' premature senescence and myofibroblast-like phenotype transformation time-dependently, which may contribute to the development of early stage of glomerulosclerosis.

An optimal dose of tea polyphenols protects against global cerebral ischemia/reperfusion injury.

Previous studies addressing the protection of tea polyphenols against cerebral ischemia/reperfusion injury often use focal cerebral ischemia models, and the optimal dose is not unified. In this experiment, a cerebral ischemia/reperfusion injury rat model was established using a modified four-vessel occlusion method. Rats were treated with different doses of tea polyphenols (25, 50, 100, 150, 200 mg/kg) via intraperitoneal injection. Results showed that after 2, 6, 12, 24, 48 and 72 hours of reperfusion, peroxide dismutase activity and total antioxidant capacity in brain tissue gradually increased, while malondialdehyde content gradually decreased after tea polyphenol intervention. Tea polyphenols at 200 mg/kg resulted in the most apparent changes. Terminal deoxynucleotidyl transferase-mediated nick end labeling and flow cytometry showed that 200 mg/kg tea polyphenols significantly reduced the number and percentage of apoptotic cells in the hippocampal CA1 region of rats after cerebral ischemia/reperfusion injury. The open field test and elevated plus maze experiments showed that tea polyphenols at 200 mg/kg strengthened exploratory behavior and reduced anxiety of cerebral ischemia/reperfusion injured rats. Experimental findings indicate that tea polyphenols protected rats against cerebral ischemia/reperfusion injury and 200 mg/kg is regarded as the optimal dose.

Uremic anorexia and ghrelin expression in the amygdala.

BACKGROUND/AIMS: Ghrelin can act as a signal for mealtime hunger and meal initiation. Amygdala is indispensable in appetitive behavior motivated by learned emotions. This study was to investigate the alteration of ghrelin in the amygdala of rats with chronic renal failure (CRF) and its relation with uremic anorexia. METHODS: SD rats were randomly classified into CRF group and control group (n=16 per group). The CRF model was constructed using 5/6 nephrectomy. When plasma creatinine (PCr) and blood urea nitrogen (BUN) in the CRF group were twice more than the normal level, food intake (g/24h) was measured and then all rats were killed for detection of ghrelin protein expression in the amygdala using immunohistochemical analysis and mRNA expression using RT-PCT. Statistics was conducted with one-way analysis of variance, Student-Newman-Keuls-q test and correlation analysis. RESULTS: By the 8th week after the surgery, the BUN and PCr of CRF rats exceeded double the normal level, and their food intake was obviously decreased compared with the controls (P<0.05). The protein and mRNA expression of ghrelin in the amygdala of CRF group were significantly reduced, and there was a positive correlation between this reduction and the decrease in food intake (P<0.05). CONCLUSION: The reduction of amygdala's ghrelin in CRF rats may be associated with uremic anorexia.

Palmitic and linoleic acids induce ER stress and apoptosis in hepatoma cells.

OBJECTIVES: Hepatic inflammation and degeneration induced by lipid depositions may be the major cause of nonalcoholic fatty liver disease. In this study, we tried to investigate the effects of saturated and unsaturated fatty acids on hepatoma cell apoptosis. METHODS: H4IIE liver cells were treated with palmitic acid, linoleic acid, or both with or without the calcium-specific chelator BAPTA-AM after which the expression of proteins associated with endoplasmic reticulum (ER) stress, apoptosis, caspase-3 levels, and calcium flux were measured. RESULTS: Palmitic or linoleic acid (250 µM) induced H4IIE cell apoptosis, which required calcium flux but not caspase-3. Apoptosis was not observed when cells were co-treated with linoleic acid (125 µM) and palmitic acid (250 µM). Importantly, the release of cytochrome C from mitochondria into cytoplasm during cell apoptosis was specifically detected only when linoleic acid (125 µM), but not palmitic acid (250 µM), was added to the cells. Depletion of intracellular calcium flux by the calcium-specific chelator, BAPTA-AM, abolished linoleic acid-induced apoptosis. Moreover, in the presence of BAPTA-AM, expression of the unfolded protein response (UPR)-associated genes, CHOP, GRP78, and GRP94, was induced by linoleic acid, but not palmitic acid. CONCLUSIONS: The results suggest that linoleic acid promotes cell apoptosis through the release of cytochrome C, only if the intracellular calcium flux is unperturbed and intact. These results confirm that ER stress contributes to fatty acid-induced liver cell apoptosis.

Tea polyphenols increase X-ray repair cross-complementing protein 1 and apurinic/apyrimidinic endonuclease/redox factor-1 expression in the hippocampus of rats during cerebral ischemia/reperfusion injury.

Recent studies have shown that tea polyphenols can cross the blood-brain barrier, inhibit apoptosis and play a neuroprotective role against cerebral ischemia. Furthermore, tea polyphenols can decrease DNA damage caused by free radicals. We hypothesized that tea polyphenols repair DNA damage and inhibit neuronal apoptosis during global cerebral ischemia/reperfusion. To test this hypothesis, we employed a rat model of global cerebral ischemia/reperfusion. We demonstrated that intraperitoneal injection of tea polyphenols immediately after reperfusion significantly reduced apoptosis in the hippocampal CA1 region; this effect started 6 hours following reperfusion. Immunohistochemical staining showed that tea polyphenols could reverse the ischemia/reperfusion-induced reduction in the expression of DNA repair proteins, X-ray repair cross-complementing protein 1 and apurinic/apyrimidinic endonuclease/redox factor-1 starting at 2 hours. Both effects lasted at least 72 hours. These experimental findings suggest that tea polyphenols promote DNA damage repair and protect against apoptosis in the brain.

[Fas and TNFR1 expressions after cerebral ischemia and reperfusion in rats: association with cell apoptosis and the effects of Bcl-2 overexpression].

OBJECTIVE: To investigate the effect of Bcl-2 overexpression on Fas and TNFR1-mediated apoptosis and its possible mechanism in rat hippocampus following global ischemia/reperfusion (IR). METHODS: Ninety healthy male SD rats were randomly divided into sham operated group, IR group and Bcl-2 overexpression group (BT group). Rat model of global IR was established by the 4-V0 method. The expressions of Bcl-2, Fas and TNFR1 and the cell apoptosis in the CA1 and CA3 regions were examined by HE staining, immunohistochemistry and TUNEL method. RESULTS: In IR group, the neurons in the CA1 region showed an obvious reduction in number with disordered arrangement and interstitial edema 48 h after global IR. Such changes were not obvious in BT group. Immunohistochemistry showed that Fas expression in the CA1 region reached the peak level at 6 h in IR group with a greater expression intensity than that in BT group (P<0.05). TNFR1 was expressed at a higher level in IR group than in BT group (P<0.05), reaching the peak level at 24 h. In the sham group, the expression of Fas and TNFR1 was not detected the in CA1 and CA3 regions. Global IR caused increased cell apoptosis in the CA1 and CA3 regions, starting at 6 h and reached peak at 24 to 48 h. The cell apoptosis was less obvious in BT group (P<0.05). CONCLUSION: Fas and TNFR1 are expressed in the CA1 and CA3 regions after global IR in rats, suggesting the involvement of death receptor in cerebral IR injury. Bcl-2 overexpression decreases the expression of Fas and TNFR1 and cell apoptosis after global IR, thus offering protective effect against cerebral IR injury.