A Non-Invasive Method for Monitoring Osteogenesis and Osseointegration Using Near-Infrared Fluorescent Imaging: A Model of Maxilla Implantation in Rats.

Currently, computed tomography and conventional X-ray radiography usually generate a micro-artifact around metal implants. This metal artifact frequently causes false positive or negative diagnoses of bone maturation or pathological peri-implantitis around implants. In an attempt to repair the artifacts, a highly specific nanoprobe, an osteogenic biomarker, and nano-Au-Pamidronate were designed to monitor the osteogenesis. In total, 12 Sprague Dawley rats were included in the study and could be chategorized in 3 groups: 4 rats in the X-ray and CT group, 4 rats in the NIRF group, and 4 rats in the sham group. A titanium alloy screw was implanted in the anterior hard palate. The X-ray, CT, and NIRF images were taken 28 days after implantation. The X-ray showed that the tissue surrounded the implant tightly; however, a gap of metal artifacts was noted around the interface between dental implants and palatal bone. Compared to the CT image, a fluorescence image was noted around the implant site in the NIRF group. Furthermore, the histological implant-bone tissue also exhibited a significant NIRF signal. In conclusion, this novel NIRF molecular imaging system precisely identifies the image loss caused by metal artifacts and can be applied to monitoring bone maturation around orthopedic implants. In addition, by observing the new bone formation, a new principle and timetable for an implant osseointegrated with bone can be established and a new type of implant fixture or surface treatment can be evaluated using this system.

Enzalutamide Induces Apoptotic Insults to Human Drug-Resistant and -Sensitive Glioblastoma Cells via an Intrinsic Bax-Mitochondrion-Cytochrome C Caspase Cascade Activation Pathway.

Glioblastoma multiforme (GBM) is the most common and malignant brain tumor. Temozolomide (TMZ) is the first-line chemotherapeutic drug for treating GBM. However, drug resistance is still a challenging issue in GBM therapy. Our preliminary results showed upregulation of androgen receptor (AR) gene expression in human GBM tissues. This study was designed to evaluate the effects of enzalutamide, a specific inhibitor of the AR, on killing drug-resistant and -sensitive glioblastoma cells and the possible mechanisms. Data mining from The Cancer Genome Atlas (TCGA) database revealed upregulation of AR messenger (m)RNA and protein expressions in human GBM tissues, especially in male patients, compared to normal human brains. In addition, expressions of AR mRNA and protein in human TMZ-sensitive U87 MG and -resistant U87 MG-R glioblastoma cells were elevated compared to normal human astrocytes. Exposure of human U87 MG and U87 MG-R cells to enzalutamide concentration- and time-dependently decreased cell viability. As to the mechanism, enzalutamide killed these two types of glioblastoma cells via an apoptotic mechanism. Specifically, exposure to enzalutamide augmented enzyme activities of caspase-9 rather than those of caspase-8. Moreover, enzalutamide successively triggered an elevation in levels of the proapoptotic Bax protein, a reduction in the mitochondrial membrane potential, release of cytochrome c, cascade activation of caspases-3 and -6, DNA fragmentation, and cell apoptosis in human TMZ-sensitive and -resistant glioblastoma cells. Pretreatment with Z-VEID-FMK, an inhibitor of caspase-6, caused significant attenuations in enzalutamide-induced morphological shrinkage, DNA damage, and apoptotic death. Taken together, this study showed that enzalutamide could significantly induce apoptotic insults to human drug-resistant and -sensitive glioblastoma cells via an intrinsic Bax-mitochondrion-cytochrome c-caspase cascade activation pathway. Enzalutamide has the potential to be a drug candidate for treating GBM by targeting the AR signaling axis.

Cervical Noninvasive Vagus Nerve Stimulation for Migraine and Cluster Headache: A Systematic Review and Meta-Analysis.

BACKGROUND: Noninvasive vagus nerve stimulation (nVNS) has been proposed as a new neuromodulation therapy to treat primary headache disorders. The purpose of this study was to analyze the effectiveness and safety of peripheral nerve stimulation of the cervical branch of the vagal nerve for primary headache disorders. METHODS: A systematic review and meta-analysis of the literature was carried out on randomized controlled trials of nVNS for treating headaches. We searched the Medline, Embase, and CENTRAL databases until January 29, 2019. A random-effects model was used to report all outcomes. The primary outcomes were a reduction in headache days or attacks and pain-free status within 30 min. Secondary outcomes were: the pain-relief status within 30 min, the pain-relief status at 60 min, abortive medication use, ≥50% responder rate, pain-free status in ≥50% of treated attacks, adverse events, and satisfaction. RESULTS: In total, 983 patients were included from six trials. We found that nVNS was effective in achieving a pain-free status within 30 min (odds ratio [OR], 2.27; 95% confidence interval [CI], 1.16~4.44; p = 0.02), pain-relief status within 30 min (OR, 1.8; 95% CI, 1.17~2.78; p = 0.007), pain-relief status at 60 min (OR, 1.93; 95% CI, 1.2~3.1; p = 0.006), a reduction in abortive medication use (OR, 0.61; 95% CI, 0.41~0.92; p = 0.02), and pain-free status in ≥50% of treated attacks (OR, 2.15; 95% CI, 1.27~3.66; p = 0.005) compared to sham-device treatment. There were no significant differences in decreased headache days (standardized mean difference (SMD), -0.159; 95% CI, -0.357~0.04; p = 0.117), adverse events (OR, 1.084; 95% CI, 0.559~2.104; p = 0.811), or satisfaction (OR, 1.45; 95% CI, 0.97~2.17; p = 0.07) between nVNS and sham-device treatment. The ≥50% responder rate could not be determined (OR, 3.34; 95% CI, 0.83~13.33; p = 0.09; I 2 = 73%). CONCLUSIONS: Cervical nVNS is effective for acute pain relief for migraine and cluster headache. SYSTEMATIC REVIEW REGISTRATION: PROSPERO registration number CRD42019126009.

The Role of Vitamin D in Modulating Mesenchymal Stem Cells and Endothelial Progenitor Cells for Vascular Calcification.

Vascular calcification, which involves the deposition of calcifying particles within the arterial wall, is mediated by atherosclerosis, vascular smooth muscle cell osteoblastic changes, adventitial mesenchymal stem cell osteoblastic differentiation, and insufficiency of the calcification inhibitors. Recent observations implied a role for mesenchymal stem cells and endothelial progenitor cells in vascular calcification. Mesenchymal stem cells reside in the bone marrow and the adventitial layer of arteries. Endothelial progenitor cells that originate from the bone marrow are an important mechanism for repairing injured endothelial cells. Mesenchymal stem cells may differentiate osteogenically by inflammation or by specific stimuli, which can activate calcification. However, the bioactive substances secreted from mesenchymal stem cells have been shown to mitigate vascular calcification by suppressing inflammation, bone morphogenetic protein 2, and the Wingless-INT signal. Vitamin D deficiency may contribute to vascular calcification. Vitamin D supplement has been used to modulate the osteoblastic differentiation of mesenchymal stem cells and to lessen vascular injury by stimulating adhesion and migration of endothelial progenitor cells. This narrative review clarifies the role of mesenchymal stem cells and the possible role of vitamin D in the mechanisms of vascular calcification.

Major Contribution of Caspase-9 to Honokiol-Induced Apoptotic Insults to Human Drug-Resistant Glioblastoma Cells.

Temozolomide (TMZ)-induced chemoresistance to human glioblastomas is a critical challenge now. Our previous studies showed that honokiol, a major bioactive constituent of Magnolia officinalis (Houpo), can kill human glioblastoma cells and suppresses glioblastoma growth. This study was further aimed to evaluate the effects of honokiol on human drug-resistant glioblastoma cells and the possible mechanisms. The results by data mining in the cancer genome atlas (TCGA) database and immunohistochemistry displayed that expression of caspase-9 mRNA and protein in human glioblastomas was induced. Human TMZ-resistant U87-MG-R9 glioblastoma cells were selected and prepared from human drug-sensitive U87-MG cells. Compared to human drug-sensitive U87-MG cells, TMZ did not affect viability of U87-MG-R9 glioblastoma cells. Interestingly, treatment with honokiol suppressed proliferation and survival of human drug-resistant glioblastoma cells in concentration- and time-dependent manners. Compared to caspase-8 activation, honokiol chiefly increased activity of caspase-9 in U87-MG-R9 cells. Successively, levels of cleaved caspase-3 and activities of caspase-3 and caspase-6 in human TMZ-tolerant glioblastoma cells were augmented following honokiol administration. In parallel, honokiol triggered DNA fragmentation of U87-MG-R9 cells. Accordingly, treatment of human TMZ-resistant glioblastoma cells with honokiol induced cell apoptosis but did not affect cell necrosis. Fascinatingly, suppressing caspase-9 activity using its specific inhibitors repressed honokiol-induced caspase-6 activation, DNA fragmentation, and cell apoptosis. Taken together, this study has shown the major roles of caspase-9 in transducing honokiol-induced mitochondria-dependent apoptosis in human drug-resistant glioblastoma cells. Thus, honokiol may be clinically applied as a drug candidate for treatment of glioblastoma patients with chemoresistance.

Methylpiperidinopyrazole Attenuates Estrogen-Induced Mitochondrial Energy Production and Subsequent Osteoblast Maturation via an Estrogen Receptor Alpha-Dependent Mechanism.

An estrogen deficiency is the main cause of osteoporosis in postmenopausal women. In bone remodeling, estrogen receptors (ERs) can mediate estrogen-transducing signals. Methylpiperidinopyrazole (MPP) is a highly specific antagonist of ER-alpha (ERα). This study was designed to evaluate the effects of MPP on estrogen-induced energy production, subsequent osteoblast maturation, and the possible mechanisms. Exposure of primary osteoblasts isolated from neonatal rat calvarias to MPP did not affect cell morphology or survival. Estradiol can induce translocation of ERα into mitochondria from the cytoplasm. Interestingly, pretreatment of rat calvarial osteoblasts with MPP lowered estrogen-induced ERα translocation. Sequentially, estrogen-triggered expressions of mitochondrial energy production-linked cytochrome c oxidase (COX) I and COX II messenger (m)RNAs were inhibited following pretreatment with MPP. Consequently, MPP caused decreases in estrogen-triggered augmentation of the activities of mitochondrial respiratory complex enzymes and levels of cellular adenosine phosphate (ATP). During progression of osteoblast maturation, estrogen induced bone morphogenetic protein (BMP)-6 and type I collagen mRNA expressions, but MPP treatment inhibited such induction. Consequently, estrogen-induced osteoblast activation and mineralization were attenuated after exposure to MPP. Taken together, MPP suppressed estrogen-induced osteoblast maturation through decreasing chromosomal osteogenesis-related BMP-6 and type I collagen mRNA expressions and mitochondrial ATP synthesis due to inhibiting energy production-linked COX I and II mRNA expressions. MPP can appropriately be applied to evaluate estrogen-involved bioenergetics and osteoblast maturation.

Improved effects of honokiol on temozolomide-induced autophagy and apoptosis of drug-sensitive and -tolerant glioma cells.

BACKGROUND: Temozolomide (TMZ)-induced side effects and drug tolerance to human gliomas are still challenging issues now. Our previous studies showed that honokiol, a major bioactive constituent of Magnolia officinalis (Houpo), is safe for normal brain cells and can kill human glioma cells. This study was further aimed to evaluate the improved effects of honokiol and TMZ on drug-sensitive and -resistant glioma cells and the possible mechanisms. METHODS: TMZ-sensitive human U87-MG and murine GL261 glioma cells and TMZ-resistant human U87-MR-R9 glioma cells were exposed to honokiol and TMZ, and cell viability and LC50 of honokiol were assayed. To determine the death mechanisms, caspase-3 activity, DNA fragmentation, apoptotic cells, necrotic cells, cell cycle, and autophagic cells. The glioma cells were pretreated with 3-methyladenine (3-MA) and chloroquine (CLQ), two inhibitors of autophagy, and then exposed to honokiol or TMZ. RESULTS: Exposure of human U87-MG glioma cells to honokiol caused cell death and significantly enhanced TMZ-induced insults. As to the mechanism, combined treatment of human U87-MG cells with honokiol and TMZ induced greater caspase-3 activation, DNA fragmentation, cell apoptosis, and cell-cycle arrest at the G1 phase but did not affect cell necrosis. The improved effects of honokiol on TMZ-induced cell insults were further verified in mouse GL261 glioma cells. Moreover, exposure of drug-tolerant human U87-MG-R9 cells to honokiol induced autophagy and consequent apoptosis. Pretreatments with 3-MA and CLQ caused significant attenuations in honokiol- and TMZ-induced cell autophagy and apoptosis in human TMZ-sensitive and -tolerant glioma cells. CONCLUSIONS: Taken together, this study demonstrated the improved effects of honokiol with TMZ on autophagy and subsequent apoptosis of drug-sensitive and -tolerant glioma cells. Thus, honokiol has the potential to be a drug candidate for treating human gliomas.

Effects of tebuconazole on cytochrome P450 enzymes, oxidative stress, and endocrine disruption in male rats.

The major objective of the present study was to determine the ability of a triazole fungicide tebuconazole to induce cytochrome P450-dependent monooxygenases, oxidative stress, and endocrine-disrupting activity using male rats treated with tebuconazole at 10, 25, and 50 mg/kg p.o. once daily for 28 days. In liver, tebuconazole dose-dependently increased microsomal contents of cytochrome P450 and cytochrome b5 and the activities of NADPH-cytochrome P450 reductase, 7-ethoxyresorufin O-deethylase, methoxyresorufin O-demethylase, pentoxyresorufin O-dealkylase, 7-ethoxycoumarin O-deethylase, aniline hydroxylase, and erythromycin N-demethylase. In kidney, tebuconazole increased 7-ethoxycoumarin O-deethylase activity without affecting other monooxygenase activities. In marked contrast to liver and kidney, tebuconazole decreased testicular 7-ethoxyresorufin O-deethylase, methoxyresorufin O-demethylase, 7-ethoxycoumarin O-deethylase, aniline hydroxylase, and erythromycin N-demethylase activities. The results of immunoblot analysis of liver microsomes of controls and tebuconazole-treated rats revealed that tebuconazole induced CYP1A1/2, CYP2B1/2, CYP2E1, and CYP3A proteins in liver. Additions of tebuconazole to liver microsomes inhibited microsomal 7-ethoxycoumarin O-deethylase activity in vitro (IC50 = 1.50-1.69 µM). Treatment of rats with tebuconazole decreased glutathione content and increased glutathione S-transferase, superoxide dismutase, catalase, and glutathione peroxidase activities in liver; increased superoxide dismutase activities in kidney and testis; but decreased glutathione S-transferase activity in testis. Treatments with tebuconazole decreased serum testosterone concentration and cauda epididymal sperm count. The present study demonstrates that tebuconazole induces a multiplicity of CYPs and oxidative stress in liver; inhibits testicular P450 and glutathione S-transferase activities; and produces anti-androgenic effects in male rats.

Protection of Dexmedetomidine Against Ischemia/Reperfusion-Induced Apoptotic Insults to Neuronal Cells Occurs Via an Intrinsic Mitochondria-Dependent Pathway.

Dexmedetomidine, an agonist of alpha2-adrenergic receptors, is used for critically ill patients to induce and maintain sedation and analgesia. Brain ischemia/reperfusion (I/R) usually causes severe neuronal injuries to intensive care unit patients. This study was aimed to evaluate the effects of dexmedetomidine on I/R-induced insults to neuronal cells and the possible mechanisms. Treatment of neuro-2a cells with dexmedetomidine did not affect cell viability but could protect against I/R-induced cell death. Separately, the I/R-triggered cell shrinkage, DNA fragmentation, and apoptosis in neuro-2a cells were alleviated by dexmedetomidine. As to the mechanisms, exposure of neuro-2a cells to dexmedetomidine substantially attenuated I/R-induced translocation of Bax protein from the cytosol to mitochondria and reduction in the mitochondrial membrane potential (MMP). Successively, dexmedetomidine decreased cytochrome c release from mitochondria to the cytoplasm and consequent cascade activations of caspases-9, -3, and -6 in I/R-treated neuro-2a cells. Interestingly, downregulating caspase-6 activity synergistically improved dexmedetomidine-induced defense against I/R-induced apoptosis of neuro-2a cells. The dexmedetomidine-involved neuroprotection was further confirmed in the other NB41A3 neuronal cells by significantly attenuating I/R-induced changes in the MMP, caspase-3 activation, DNA fragmentation, and cell apoptosis. Taken together, this study has shown the neuroprotective effects of dexmedetomidine against I/R-induced apoptotic insults via an intrinsic Bax-mitochondria-cytochrome c-caspase protease pathway. J. Cell. Biochem. 118: 2635-2644, 2017. © 2016 Wiley Periodicals, Inc.

Honokiol induces autophagic cell death in malignant glioma through reactive oxygen species-mediated regulation of the p53/PI3K/Akt/mTOR signaling pathway.

Honokiol, an active constituent extracted from the bark of Magnolia officinalis, possesses anticancer effects. Apoptosis is classified as type I programmed cell death, while autophagy is type II programmed cell death. We previously proved that honokiol induces cell cycle arrest and apoptosis of U87 MG glioma cells. Subsequently in this study, we evaluated the effect of honokiol on autophagy of glioma cells and examined the molecular mechanisms. Administration of honokiol to mice with an intracranial glioma increased expressions of cleaved caspase 3 and light chain 3 (LC3)-II. Exposure of U87 MG cells to honokiol also induced autophagy in concentration- and time-dependent manners. Results from the addition of 3-methyladenine, an autophagy inhibitor, and rapamycin, an autophagy inducer confirmed that honokiol-induced autophagy contributed to cell death. Honokiol decreased protein levels of PI3K, phosphorylated (p)-Akt, and p-mammalian target of rapamycin (mTOR) in vitro and in vivo. Pretreatment with a p53 inhibitor or transfection with p53 small interfering (si)RNA suppressed honokiol-induced autophagy by reversing downregulation of p-Akt and p-mTOR expressions. In addition, honokiol caused generation of reactive oxygen species (ROS), which was suppressed by the antioxidant, vitamin C. Vitamin C also inhibited honokiol-induced autophagic and apoptotic cell death. Concurrently, honokiol-induced alterations in levels of p-p53, p53, p-Akt, and p-mTOR were attenuated following vitamin C administration. Taken together, our data indicated that honokiol induced ROS-mediated autophagic cell death through regulating the p53/PI3K/Akt/mTOR signaling pathway.

Honokiol inhibits sphere formation and xenograft growth of oral cancer side population cells accompanied with JAK/STAT signaling pathway suppression and apoptosis induction.

BACKGROUND: Eliminating cancer stem cells (CSCs) has been suggested for prevention of tumor recurrence and metastasis. Honokiol, an active compound of Magnolia officinalis, had been proposed to be a potential candidate drug for cancer treatment. We explored its effects on the elimination of oral CSCs both in vitro and in vivo. METHODS: By using the Hoechst side population (SP) technique, CSCs-like SP cells were isolated from human oral squamous cell carcinoma (OSCC) cell lines, SAS and OECM-1. Effects of honokiol on the apoptosis and signaling pathways of SP-derived spheres were examined by Annexin V/Propidium iodide staining and Western blotting, respectively. The in vivo effectiveness was examined by xenograft mouse model and immunohistochemical tissue staining. RESULTS: The SP cells possessed higher stemness marker expression (ABCG2, Ep-CAM, Oct-4 and Nestin), clonogenicity, sphere formation capacity as well as tumorigenicity when compared to the parental cells. Treatment of these SP-derived spheres with honokiol resulted in apoptosis induction via Bax/Bcl-2 and caspase-3-dependent pathway. This apoptosis induction was associated with marked suppression of JAK2/STAT3, Akt and Erk signaling pathways in honokiol-treated SAS spheres. Consistent with its effect on JAK2/STAT3 suppression, honokiol also markedly inhibited IL-6-mediated migration of SAS cells. Accordingly, honokiol dose-dependently inhibited the growth of SAS SP xenograft and markedly reduced the immunohistochemical staining of PCNA and endothelial marker CD31 in the xenograft tumor. CONCLUSIONS: Honokiol suppressed the sphere formation and xenograft growth of oral CSC-like cells in association with apoptosis induction and inhibition of survival/proliferation signaling pathways as well as angiogenesis. These results suggest its potential as an integrative medicine for combating oral cancer through targeting on CSCs.

Oxidative stress-induced apoptotic insults to rat osteoblasts are attenuated by nitric oxide pretreatment via GATA-5-involved regulation of Bcl-X L gene expression and protein translocation.

Nitric oxide (NO) has biphasic effects on regulating osteoblast survival and death. This study was aimed to evaluate the effects of NO pretreatment on hydrogen peroxide (HP)-induced insults of rat osteoblasts and the possible mechanisms. Exposure of osteoblasts prepared from rat calvarias to HP significantly increased intracellular reactive oxygen species levels, decreased alkaline phosphatase activity and cell survival, and ultimately induced cell apoptosis. However, NO pretreatment lowered HP-induced oxidative stress and apoptotic insults. In parallel, HP increased Bax levels and its translocation from the cytoplasm to mitochondria. NO pretreatment caused significant attenuations in HP-induced modulations in Bax synthesis and translocation. In contrast, pretreatment with NO enhanced levels and translocation of antiapoptotic Bcl-XL protein in rat osteoblasts. RNA analyses further revealed that HP inhibited Bcl-XL mRNA expression without affecting Bax mRNA levels. In comparison, NO induced Bcl-XL mRNA production and alleviated HP-caused inhibition of this mRNA expression. As to the mechanism, HP suppressed RNA and protein levels of transcription factor GATA-5 in rat osteoblasts. Pretreatment with NO induced GATA-5 mRNA and protein expressions and simultaneously attenuated HP-induced inhibition of this gene's expression. Consequently, GATA-5 knockdown using RNA interference inhibited Bcl-XL mRNA expression and concurrently lowered NO's protection against HP-induced apoptotic insults. Therefore, this study showed that NO can protect rat osteoblasts from HP-induced apoptotic insults. The protective mechanisms are mediated by GATA-5-mediated transcriptional induction of Bcl-X L gene, and translocational modulation of Bcl-XL and Bax proteins.

Roles of microRNA-1 in hypoxia-induced apoptotic insults to neuronal cells.

Hypoxia is a common occurrence in brain tumors and traumatic brain injury. microRNA (miR)-1 participates in the regulation of brain development and neuronal function. Interestingly, miR-1 can mediate ischemia-induced injury to cardiomyocytes. This study was designed to evaluate the roles of miR-1 in hypoxia-induced insults to neurons and the possible mechanisms. Exposure of neuro-2a cells to oxygen/glucose deprivation (OGD) or cobalt chloride decreased cell viability and induced cell apoptosis in time-dependent manners. In parallel, OGD caused augmentation of cellular Bax and cytochrome c levels, a reduction in the mitochondrial membrane potential (MMP), activation of caspase-3, and fragmentation of DNA. miR-1 was induced in neuro-2a cells by OGD. Knocking down miR-1 expression using specific antisense inhibitors significantly alleviated OGD-induced neuronal death. Administration of OGD to neuro-2a cells induced heat-shock protein (HSP)-70 messenger (m)RNA and protein expressions. A bioinformatic search revealed that miR-1-specific binding elements exist in the 3'-untranslated region of HSP-70 mRNA. Overexpression of miR-1 simultaneously attenuated OGD-induced HSP-70 mRNA and protein expressions. In comparison, knocking down miR-1 expression synergistically enhanced OGD-induced HSP-70 mRNA. As to the mechanism, reducing miR-1 expression lowered OGD-induced alterations in the MMP, caspase-3 activation, DNA fragmentation, and cell apoptosis. Taken together, this study shows that miR-1 can target HSP-70 expression and consequently mediate hypoxia-induced apoptotic insults to neuro-2a cells via an intrinsic Bax-mitochondrion-caspase protease pathway.

Genistein induces oestrogen receptor-α gene expression in osteoblasts through the activation of mitogen-activated protein kinases/NF-κB/activator protein-1 and promotes cell mineralisation.

Oestrogen and oestrogen receptors (ER) play critical roles in the maintenance of bone remodelling. Genistein, structurally similar to 17ß-oestradiol, is a phyto-oestrogen that may be beneficial for treating osteoporosis. In the present study, we evaluated the effects of genistein on the regulation of ERα gene expression and osteoblast mineralisation using MC3T3-E1 cells and primary rat calvarial osteoblasts as our experimental models. Exposure of MC3T3-E1 cells and primary rat osteoblasts to genistein at ≤ 10 µm for 24 h did not affect the cell morphology or viability. However, treatment of MC3T3-E1 cells with 10 µm-genistein enhanced the phosphorylation of extracellular signal-regulated kinase 1/2, p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase 1/2 in a time-dependent manner. Sequentially, genistein increased the translocation of NF-κB and c-Jun from the cytoplasm to the nucleus. Consequently, exposure of MC3T3-E1 cells to genistein induced ERα mRNA expression in concentration- and time-dependent manners. In parallel, the amounts of cytosolic and nuclear ERα in MC3T3-E1 cells were increased following genistein administration. Additionally, genistein also increased the levels of ERα mRNA and nuclear ERα protein in rat calvarial osteoblasts. A bioinformatic search revealed that there are several ERα-specific DNA-binding elements in the 5'-promoter regions of the bone morphogenetic protein-6, collagen type I and osteocalcin genes. As a result, genistein could induce the expressions of these osteoblast differentiation-related genes in primary rat osteoblasts. Co-treatment with genistein and traditional differentiation reagents synergistically increased osteoblast mineralisation. Therefore, the present study showed that genistein can induce ERα gene expression via the activation of MAPK/NF-κB/activator protein-1 and accordingly stimulates differentiation-related gene expressions and osteoblast mineralisation.

The Ratio of Plasma Amyloid-ß 1-42 over Serum Albumin Can Be a Novel Biomarker Signature for Diagnosing End-Stage Renal Disease-Associated Cognitive Impairment.

BACKGROUND: Cognitive impairment (CI) is one of the major complications in chronic kidney disease patients, especially those with end-stage renal disease (ESRD). Limited biomarkers have been found that can significantly predict ESRD-associated cognitive decline. OBJECTIVE: This cohort study aimed to investigate de novo biomarkers for diagnosis of the ESRD-associated CI. METHODS: In this cohort study, qualified samples were divided into control (with an estimated glomerular filtration rate (eGFR) of≥60 mL/min and a Mini-Mental State Examination (MMSE) score of > 27), ESRD without CI (eGFR < 15 and MMSE > 27), and ESRD with CI (eGFR < 15 and MMSE < 27) groups. Levels of plasma amyloid-ß (Aß)1 - 42, serum indoxyl sulfate, and hematologic and biochemical parameters were measured. RESULTS: Compared to the control group, levels of blood urea nitrogen, creatinine, and indoxyl sulfate were elevated in ESRD patients both without and with CI. Interestingly, ESRD patients with CI had the lowest levels of serum albumin. In contrast, levels of plasma Aß1 - 42 were significantly higher in the ESRD with CI group than in the control and ESRD without CI groups. In addition, the ratio of plasma Aß1 - 42 over serum albumin was significantly higher in the ESRD with CI group than in the control or ESRD without CI groups. Importantly, the area under the receiver operating characteristic curve (AUROC) for CI in the total population by the ratio of Aß1 - 42 over albumin was 0.785 and significant (p < 0.05). CONCLUSIONS: This cohort study has shown that the ratio of plasma Aß1 - 42 over serum albumin can be a de novo biomarker for the diagnosis and prognosis of ESRD-associated cognitive decline.

MicroRNA-210 targets antiapoptotic Bcl-2 expression and mediates hypoxia-induced apoptosis of neuroblastoma cells.

MicroRNAs (miRNAs) can regulate cell survival and death by targeting apoptosis-related gene expression. miR-210 is one of the most hypoxia-sensitive miRNAs. In this study, we evaluated the roles of miR-210 in hypoxia-induced insults to neural cells. Treatment of neuro-2a cells with oxygen/glucose deprivation (OGD) induced cell apoptosis in a time-dependent manner. In parallel, OGD time-dependently increased cellular miR-210 levels. Knocking down miR-210 expression using specific antisenses significantly attenuated OGD-induced neural apoptosis. Concurrently, OGD increased hypoxia-inducible factor (HIF)-1α mRNA and protein syntheses. Pretreatment with YC-1, an inhibitor of HIF-1α, reduced OGD-caused cell death. Sequentially, OGD specifically decreased antiapoptotic Bcl-2 mRNA and protein levels in neuro-2a cells. A search by a bioinformatic approach revealed that miR-210-specific binding elements exist in the 3'-untranslated region of Bcl-2 mRNA. Application of miR-210 antisenses simultaneously alleviated OGD-involved inhibition of Bcl-2 mRNA expression. In comparison, overexpression of miR-210 synergistically diminished OGD-caused inhibition of Bcl-2 mRNA expression and consequently induced greater cellular insults. Taken together, this study shows that OGD can induce miR-210 expression through activating HIF-1α. And miR-210 can mediate hypoxia-induced neural apoptosis by targeting Bcl-2.

Pharmacokinetics of desflurane elimination from respiratory gas and blood during the 20 minutes after cardiac surgery.

BACKGROUND/PURPOSE: Desflurane, with a low blood-gas partition coefficient, is an ideal anesthetic to achieve rapid offset and recovery from general anesthesia. Investigation of desflurane elimination from blood and respiratory gas should provide useful information with respect to a patient's recovery from anesthesia. Therefore, this study is designed to characterize the pharmacokinetics of desflurane elimination after cardiac surgery. METHODS: Sixteen patients undergoing coronary artery bypass graft surgery were enrolled. At the end of surgery, multiple gas and blood samples were taken in the 20 minutes before and after stopping desflurane administration, with prior maintenance of a fixed 7% inspired desflurane in 6 L/minute oxygen flow for 60 minutes before the cessation. The blood desflurane concentrations, including internal jugular-bulb blood (Jdes), arterial blood (Ades) and pulmonary arterial blood (PAdes) were analyzed using gas chromatography. The inspiratory desflurane concentration (CIdes) and end-tidal desflurane (CEdes) were measured with an infrared analyzer, and cardiac output was measured using an Opti-Q pulmonary artery catheter. RESULTS: Before cessation of desflurane administration, the inspiratory desflurane concentration (CIdes) was relatively higher than end-tidal (CEdes), arterial (Ades), internal jugular-bulb blood (Jdes), and pulmonary (PAdes) concentrations in sequence (CIdes > CEdes > Ades≈ Jdes > PAdes). During the elimination phase, rapid decay occurred in CEdes, followed by Jdes, Ades and PAdes. Twenty minutes after stopping desflurane administration, the desflurane concentrations were: PAdes > Ades≈ Jdes > CEdes. The decay curves of desflurane concentrations demonstrated two distinct elimination components: an initial, fast 5-minute component followed by a slow 15-minute component. CONCLUSION: Desflurane is eliminated fastest from the lungs, as indicated by CEdes, compared to elimination from circulating blood. The initial, rapid 5-minute desflurane washout reflected the diluting effect of functional residual capacity of the lungs.

Hyperventilation accelerates rise in arterial blood concentrations of sevoflurane in gynecologic patients.

PURPOSE: We investigated whether ventilation volumes affected arterial blood sevoflurane concentration (A (sev)) and its uptake into the body during general anesthesia. METHODS: Thirty female patients undergoing elective gynecologic surgery were randomly allocated into three groups: hyperventilation, normal ventilation, and hypoventilation. Inspiratory (CI(sev)) and end-tidal ((sev)) sevoflurane concentrations were routinely measured by infrared analysis, and A (sev) were analyzed by gas chromatography for 40 min after intubation. Cardiac index and total peripheral vascular resistance were measured with a Finometer. RESULTS: During the first 10 min after sevoflurane administration, A (sev) in the hyperventilation group was the highest and differed significantly from those in the normal ventilation group, followed by those in the hypoventilation group. In addition, hyperventilation significantly increased the slope of A (sev) over time in the first 5 min, but there were no differences in slopes in the 5-10, 10-20, and 20-40 min periods, which indicates no difference in sevoflurane bodily uptake among the three groups after 5 min. CONCLUSION: Hyperventilation accelerated the rate of A (sev) increase immediately after sevoflurane administration, which was time dependent with respect to different alveolar ventilation levels.

Resveratrol triggers the ER stress-mediated intrinsic apoptosis of neuroblastoma cells coupled with suppression of Rho-dependent migration and consequently prolongs mouse survival.

Neuroblastoma, the most common childhood tumor, are highly malignant and fatal because neuroblastoma cells extremely defend against apoptotic targeting. Traditional treatments for neuroblastomas are usually ineffective and lead to serious side effects and poor prognoses. In this study, we investigated the molecular mechanisms of resveratrol-induced insults to neuroblastoma cells and survival extension of nude mice with neuroblastomas, especially in the endoplasmic reticular (ER) stress-intracellular reactive oxygen species (iROS) axis-mediated signals. Resveratrol specifically killed neuroblastoma cells mainly via apoptosis and autophagy rather than necrosis. As to the mechanisms, resveratrol time-dependently triggered productions of Grp78 protein and iROS in neuroblastoma cells. Attenuating the ER stress-iROS signaling axis significantly suppressed resveratrol-induced autophagy, DNA damage, and cell apoptosis. Successively, resveratrol decreased phosphorylation of retinoblastoma protein and induced cell cycle arrest at the S phase, translocation of Bak protein to mitochondria, a reduction in the mitochondrial membrane potential, cascade activation of caspases-9, -3, and -6, and DNA fragmentation. Moreover, weakening the ER stress-iROS axis concomitantly overcome resveratrol-induced decreases in translocation of Rho protein to membranes and succeeding cell migration. Interestingly, administration of resveratrol did not cause significant side effects but could protect the neuroblastoma-bearing nude mice from body weight loss and consequently extended the animal survival. In parallel, resveratrol elevated levels of Grp78 and then induced cell apoptosis in neuroblastoma tissues. This study has shown that resveratrol could kill neuroblastoma cells and extend survival of animals with neuroblastomas by triggering the ER stress-iROS-involved intrinsic apoptosis and suppression of Rho-dependent cell migration. Our results imply the potential of resveratrol as a drug candidate for chemotherapy of neuroblastoma patients.

Lipoteichoic acid induces surfactant protein-A biosynthesis in human alveolar type II epithelial cells through activating the MEK1/2-ERK1/2-NF-κB pathway.

BACKGROUND: Lipoteichoic acid (LTA), a gram-positive bacterial outer membrane component, can cause septic shock. Our previous studies showed that the gram-negative endotoxin, lipopolysaccharide (LPS), could induce surfactant protein-A (SP-A) production in human alveolar epithelial (A549) cells. OBJECTIVES: In this study, we further evaluated the effect of LTA on SP-A biosynthesis and its possible signal-transducing mechanisms. METHODS: A549 cells were exposed to LTA. Levels of SP-A, nuclear factor (NF)-κB, extracellular signal-regulated kinase 1/2 (ERK1/2), and mitogen-activated/extracellular signal-regulated kinase kinase (MEK)1 were determined. RESULTS: Exposure of A549 cells to 10, 30, and 50 µg/ml LTA for 24 h did not affect cell viability. Meanwhile, when exposed to 30 µg/ml LTA for 1, 6, and 24 h, the biosynthesis of SP-A mRNA and protein in A549 cells significantly increased. As to the mechanism, LTA enhanced cytosolic and nuclear NF-κB levels in time-dependent manners. Pretreatment with BAY 11-7082, an inhibitor of NF-κB activation, significantly inhibited LTA-induced SP-A mRNA expression. Sequentially, LTA time-dependently augmented phosphorylation of ERK1/2. In addition, levels of phosphorylated MEK1 were augmented following treatment with LTA. CONCLUSIONS: Therefore, this study showed that LTA can increase SP-A synthesis in human alveolar type II epithelial cells through sequentially activating the MEK1-ERK1/2-NF-κB-dependent pathway.