Baseline Toxicity Model to Identify the Specific and Nonspecific Effects of Per- and Polyfluoroalkyl Substances in Cell-Based Bioassays.

High-throughput screening is a strategy to identify potential adverse outcome pathways (AOP) for thousands of per- and polyfluoroalkyl substances (PFAS) if the specific effects can be distinguished from nonspecific effects. We hypothesize that baseline toxicity may serve as a reference to determine the specificity of the cell responses. Baseline toxicity is the minimum (cyto)toxicity caused by the accumulation of chemicals in cell membranes, which disturbs their structure and function. A mass balance model linking the critical membrane concentration for baseline toxicity to nominal (i.e., dosed) concentrations of PFAS in cell-based bioassays yielded separate baseline toxicity prediction models for anionic and neutral PFAS, which were based on liposome-water distribution ratios as the sole model descriptors. The specificity of cell responses to 30 PFAS on six target effects (activation of peroxisome proliferator-activated receptor (PPAR) gamma, aryl hydrocarbon receptor, oxidative stress response, and neurotoxicity in own experiments, and literature data for activation of several PPARs and the estrogen receptor) were assessed by comparing effective concentrations to predicted baseline toxic concentrations. HFPO-DA, HFPO-DA-AS, and PFMOAA showed high specificity on PPARs, which provides information on key events in AOPs relevant to PFAS. However, PFAS were of low specificity in the other experimentally evaluated assays and others from the literature. Even if PFAS are not highly specific for certain defined targets but disturb many toxicity pathways with low potency, such effects are toxicologically relevant, especially for hydrophobic PFAS and because PFAS are highly persistent and cause chronic effects. This implicates a heightened need for the risk assessment of PFAS mixtures because nonspecific effects behave concentration-additive in mixtures.

Exposure to perfluorooctane sulfonate reduced cell viability and insulin release capacity of ß cells.

Per- and polyfluoroalkyl substances (PFAS) are found to have multiple adverse outcomes on human health. Recently, epidemiological and toxicological studies showed that exposure to PFAS had adverse impacts on pancreas and showed association with insulin abnormalities. To explore how PFAS may contribute to diabetes, we studied impacts of perfluorooctane sulfonate (PFOS) on cell viability and insulin release capacity of pancreatic ß cells by using in vivo and in vitro methods. We found that 28-day administration with PFOS (10 mg/(kg body weight•day)) caused reductions of pancreas weight and islet size in male mice. PFOS administration also led to lower serum insulin level both in fasting state and after glucose infusion among male mice. For cell-based in vitro bioassay, we used mouse ß-TC-6 cancer cells and found 48-hr exposure to PFOS decreased the cell viability at 50 µmol/L. By measuring insulin content in supernatant, 48-hr pretreatment of PFOS (100 µmol/L) decreased the insulin release capacity of ß-TC-6 cells after glucose stimulation. Although these concentrations were higher than the environmental concentration of PFOS, it might be reasonable for high concentration of PFOS to exert observable toxic effects in mice considering mice had a faster removal efficiency of PFOS than human. PFOS exposure (50 µmol/L) to ß-TC-6 cells induced intracellular accumulation of reactive oxidative specie (ROS). Excessive ROS induced the reactive toxicity of cells, which eventually invoke apoptosis and necrosis. Results in this study provide evidence for the possible causal link of exposure to PFOS and diabetes risk.

Environmental Estrogens and Their Biological Effects through GPER Mediated Signal Pathways.

Many environmental chemicals have been found to exert estrogenic effects in cells and experimental animals by activating nuclear receptors such as estrogen receptors and estrogen-related receptors. These compounds include bisphenols, pesticides, polybrominated diphenyl ethers (PBDEs), organophosphate flame retardants, phthalates and metalloestrogens. G protein-coupled estrogen receptor (GPER) exists widely in numerous cells/tissues of human and other vertebrates. A number of studies have demonstrated that GPER plays a vital role in mediating the estrogenic effects of environmental pollutants. Even at very low concentrations, these chemicals may activate GPER pathways, thus affect many aspects of cellular functions including proliferation, metastasis and apoptosis, resulting in cancer progression, cardiovascular disorders, and reproductive dysfunction. This review summarized the environmental occurrence and human exposure levels of these pollutants, and integrated current experimental evidence toward revealing the underlying mechanisms of pollutant-induced cellular dysfunction via GPER. The GPER mediated rapid non-genomic actions play an important role in the process leading to the adverse effects observed in experimental animals and even in human beings.

Electrical stimulation of hindlimb skeletal muscle has beneficial effects on sublesional bone in a rat model of spinal cord injury.

Spinal cord injury (SCI) results in marked atrophy of sublesional skeletal muscle and substantial loss of bone. In this study, the effects of prolonged electrical stimulation (ES) and/or testosterone enanthate (TE) on muscle mass and bone formation in a rat model of SCI were tested. Compared to sham-transected animals, a significant reduction of the mass of soleus, plantaris and extensor digitorum longus (EDL) muscles was observed in animals 6 weeks post-SCI. Notably, ES or ES + TE resulted in the increased mass of the EDL muscles. ES or ES + TE significantly decreased mRNA levels of muscle atrophy markers (e.g., MAFbx and MurF1) in the EDL. Significant decreases in bone mineral density (BMD) (-27%) and trabecular bone volume (-49.3%) at the distal femur were observed in animals 6 weeks post injury. TE, ES and ES + TE treatment significantly increased BMD by +6.4%, +5.4%, +8.5% and bone volume by +22.2%, and +56.2% and+ 60.2%, respectively. Notably, ES alone or ES + TE resulted in almost complete restoration of cortical stiffness estimated by finite element analysis in SCI animals. Osteoblastogenesis was evaluated by colony-forming unit-fibroblastic (CFU-F) staining using bone marrow mesenchymal stem cells obtained from the femur. SCI decreased the CFU-F+ cells by -56.8% compared to sham animals. TE or ES + TE treatment after SCI increased osteoblastogenesis by +74.6% and +67.2%, respectively. An osteoclastogenesis assay revealed significantly increased TRAP+ multinucleated cells (+34.8%) in SCI animals compared to sham animals. TE, ES and TE + ES treatment following SCI markedly decreased TRAP+ cells by -51.3%, -40.3% and -46.9%, respectively. Each intervention greatly reduced the ratio of RANKL to OPG mRNA of sublesional long bone. Collectively, our findings demonstrate that after neurologically complete paralysis, dynamic muscle resistance exercise by ES reduced muscle atrophy, downregulated genes involved in muscle wasting, and restored mechanical loading to sublesional bone to a degree that allowed for the preservation of bone by inhibition of bone resorption and/or by facilitating bone formation.

In vivo dynamic cell tracking with long-wavelength excitable and near-infrared fluorescent polymer dots.

Development of cell-based therapeutic systems has attracted great interest in biomedicine. In vivo cell tracking by fluorescence provides indispensable information for further advancing cell therapy in clinical applications. However, it is still challenging in many cases because of the limited light penetration depth as well as the variations in fluorescent probes, cell lines, and labeling brightness. Here, we designed highly fluorescent polymer dots (Pdots) with far-red-light absorption and near-infrared (NIR) emission for cell tracking. The Pdots consisted of a donor-acceptor polymer blending system where intra-particle energy transfer yielded a narrow-band emission at 800 nm with a high quantum yield of ~0.22. We investigated biocompatibility and cell labeling brightness of the Pdots coated with cell penetrating peptides. Flow cytometry indicated that the cell-labeling brightness of both stem cells and cancer cells increased as much as ~4 orders of magnitude comparing the intensity measurements of labeled cells and controls. Yet, in vivo cell tracking results revealed distinctive fluorescence distribution for the same number of cells that were administered into mice through the tail vein. The stem cells initially accumulated in the lung and remained for seven days, whereas the cancer cells tended to be cleared by the liver in four days. The difference is likely due to the fact that cancer cells are easily attacked by the immune system, whereas stem cells have low immunogenicity. Results obtained herein confirm that NIR-fluorescent Pdots are promising platforms for in vivo cell tracking in small animals.

Rapid bone loss occurs as early as 2 days after complete spinal cord transection in young adult rats.

STUDY DESIGN: Animal study. OBJECTIVE: This study examined how soon after spinal cord injury (SCI) bone loss occurs, and investigated the underlying molecular mechanism. METHODS: Eight-week-old male Wistar rats underwent complete transection of the thoracic spinal cord at T3-4 or sham operation (n = 10-12 per group). Blood, hindlimb bone samples, and bone marrows were collected at 2 and 7 days after SCI. RESULTS: The neurologically motor-complete SCI causes loss of bone mass and deterioration of trabecular bone microstructure as early as 2 days after injury; these skeletal defects become more evident at 7 days. These changes are associated with a dramatic increase in levels of bone resorption maker CTX in blood. Alternations of gene expression in hindlimb bone tissues and bone marrow cells at the first week after SCI were examined. Gene expressions responsible for both bone resorption and formation are increased at 2 days post-SCI, and the associated bone loss and bone deterioration are likely the result of higher levels of osteoclastic resorption over osteoblastic formation, as may be extrapolated from findings at molecular levels. CONCLUSIONS: Rapid bone loss occurs as early as 2 days after motor-complete SCI and interventions for inhibiting bone resorption and prompting bone formation should start as soon as possible after the injury to prevent bone loss.

Binding and activity of polybrominated diphenyl ether sulfates to thyroid hormone transport proteins and nuclear receptors.

Polybrominated diphenyl ethers (PBDEs) can be metabolized to hydroxylated PBDEs (OH-PBDEs), which play important roles in their disruption effects on the thyroid hormone (TH) system. Recently, multiple in vitro studies suggested that OH-PBDEs might be further metabolically transformed to PBDE sulfates. However, information about the bioactivity of PBDE sulfate metabolites is limited. In the present study, we explored the possible disruption effects of PBDE sulfates to the TH system by studying their binding and activity towards TH transport proteins and nuclear receptors. We found PBDE sulfates could bind to two major TH transport proteins (thyroxine-binding globulin and transthyretin). Besides, PBDE sulfates could also bind to two subtypes of TH nuclear receptors (TRα and TRß) and showed agonistic activity towards the subsequent signaling pathway. Moreover, the PBDE sulfates showed higher binding potency to TH transport proteins and TRs compared with their corresponding OH-PBDE precursors. Molecular docking results showed that replacement of hydroxyl groups with sulfate groups might lead to more hydrogen bond interactions with these proteins. Overall, our study suggested that PBDE sulfates might disturb the TH system by binding to TH transport proteins or TRs. Our finding indicated a possible mechanism for the TH system disruption effects of PBDEs through their sulfate metabolites.

Brightness Enhancement of Near-Infrared Semiconducting Polymer Dots for in Vivo Whole-Body Cell Tracking in Deep Organs.

In vivo visualization of cell migration and engraftment in small animals provide crucial information in biomedical studies. Semiconducting polymer dots (Pdots) are emerging as superior probes for biological imaging. However, in vivo whole-body fluorescence imaging is largely constrained by the limited brightness of Pdots in near-infrared (NIR) region. Here, we describe the brightness enhancement of NIR fluorescent Pdots for in vivo whole-body cell tracking in deep organs. We first synthesize semiconducting polymers with strong absorption in orange and far-red regions. By molecular doping, the weak broad-band fluorescence of the Pdots was significantly narrowed and enhanced by 1 order of magnitude enhancement, yielding bright narrow-band NIR emission with a quantum yield of ∼0.21. Under an excitation of far-red light (676 nm), a trace amount of Pdots (∼2 µg) in the stomach can be clearly detected in whole-body fluorescence imaging of live mice. The Pdots coated with a cell-penetrating peptide are able to brightly label cancer cells with minimal cytotoxicity. In vivo cell tracking in live mice indicated that the entrapment and migration of the tail-vein-administered cells (∼400 000) were clearly visualized in real time. These Pdots with deep-red excitation and bright NIR emission are promising for in vivo whole-body fluorescence imaging.

Sclerostin Antibody Reverses the Severe Sublesional Bone Loss in Rats After Chronic Spinal Cord Injury.

To date, no efficacious therapy exists that will prevent or treat the severe osteoporosis in individuals with neurologically motor-complete spinal cord injury (SCI). Recent preclinical studies have demonstrated that sclerostin antibody (Scl-Ab) can prevent sublesional bone loss after acute SCI in rats. However, it remains unknown whether sclerostin inhibition reverses substantial bone loss in the vast majority of the SCI population who have been injured for several years. This preclinical study tested the efficacy of Scl-Ab to reverse the bone loss that has occurred in a rodent model after chronic motor-complete SCI. Male Wistar rats underwent either complete spinal cord transection or only laminectomy. Twelve weeks after SCI, the rats were treated with Scl-Ab at 25 mg/kg/week or vehicle for 8 weeks. In the SCI group that did not receive Scl-Ab, 20 weeks of SCI resulted in a significant reduction of bone mineral density (BMD) and estimated bone strength, and deterioration of bone structure at the distal femoral metaphysis. Treatment with Scl-Ab largely restored BMD, bone structure, and bone mechanical strength. Histomorphometric analysis showed that Scl-Ab increased bone formation in animals with chronic SCI. In ex vivo cultures of bone marrow cells, Scl-Ab inhibited osteoclastogenesis, and promoted osteoblastogenesis accompanied by increased Tcf7, ENC1, and the OPG/RANKL ratio expression, and decreased SOST expression. Our findings demonstrate for the first time that Scl-Ab reverses the sublesional bone loss when therapy is begun after relatively prolonged spinal cord transection. The study suggests that, in addition to being a treatment option to prevent bone loss after acute SCI, sclerostin antagonism may be a valid clinical approach to reverse the severe bone loss that invariably occurs in patients with chronic SCI.

Mesoporous Carbon Nanospheres as a Multifunctional Carrier for Cancer Theranostics.

Optical nanomaterials with intense absorption in near-infrared (NIR) region hold great promise for biomedical applications such as photothermal therapy (PTT) and photoacoustic imaging (PAI). In this work, we report mesoporous carbon nanospheres (Meso-CNs) with broadband and intense absorption in the UV-Vis-NIR region (300-1400 nm) and explore their potential as a multifunctional platform for photoacoustic imaging and chemo-photothermal therapy. Methods: Meso-CNs were prepared by a "silica-assisted" synthesis strategy and characterized by transmission electron microscope and optical spectroscopy. We investigated the photothermal conversion and photoacoustic imaging of Meso-CNs in comparison with single-walled carbon nanotubes (SWCNTs), graphene and gold nanorods (GNRs). In vitro cellular assays and in vivo chemo-photothermal combination therapy were performed. Results: The absorption coefficients of Meso-CNs are 1.5-2 times higher than those of SWCNTs and graphene and are comparable to those of GNRs in both the first and the second near-infrared optical windows (NIR-I and NIR-II) of tissues. When exposed to an NIR laser, the photothermal and photoacoustic signal generation of Meso-CNs are also stronger than those of SWCNTs, graphene, and GNRs. DOX was loaded into Meso-CNs with a high efficiency (35 wt%) owing to the unique mesoporous structure. Particularly, the drug release from Meso-CNs is sensitive to both pH and NIR light stimulation. In vivo chemo-photothermal combination therapy demonstrates a remarkable inhibition effect on tumor growth under NIR laser treatment. Conclusions: We have developed Meso-CNs for photothermal conversion and photoacoustic imaging. The porous structure also serves as a drug carrier and the drug release can be controlled by pH and external light. The high drug loading capacity, superior photothermal and photoacoustic generation, together with the apparent chemo-photothermal therapeutic effect, make Meso-CNs a promising platform for cancer theranostics.

Bisphenol AF and Bisphenol B Exert Higher Estrogenic Effects than Bisphenol A via G Protein-Coupled Estrogen Receptor Pathway.

Numerous studies have indicated estrogenic disruption effects of bisphenol A (BPA) analogues. Previous mechanistic studies were mainly focused on their genomic activities on nuclear estrogen receptor pathway. However, their nongenomic effects through G protein-coupled estrogen receptor (GPER) pathway remain poorly understood. Here, using a SKBR3 cell-based fluorescence competitive binding assay, we found six BPA analogues bound to GPER directly, with bisphenol AF (BPAF) and bisphenol B (BPB) displaying much higher (∼9-fold) binding affinity than BPA. Molecular docking also demonstrated the binding of these BPA analogues to GPER. By measuring calcium mobilization and cAMP production in SKBR3 cells, we found the binding of these BPA analogues to GPER lead to the activation of subsequent signaling pathways. Consistent with the binding results, BPAF and BPB presented higher agonistic activity than BPA with the lowest effective concentration (LOEC) of 10 nM. Moreover, based on the results of Boyden chamber and wound-healing assays, BPAF and BPB displayed higher activity in promoting GPER mediated SKBR3 cell migration than BPA with the LOEC of 100 nM. Overall, we found two BPA analogues BPAF and BPB could exert higher estrogenic effects than BPA via GPER pathway at nanomolar concentrations.

Bright Polymer Dots Tracking Stem Cell Engraftment and Migration to Injured Mouse Liver.

Stem cell therapy holds promise for treatment of intractable diseases and injured organs. For clinical translation, it is pivotal to understand the homing, engraftment, and differentiation processes of stem cells in a living body. Here we report near-infrared (NIR) fluorescent semiconductor polymer dots (Pdots) for bright labeling and tracking of human mesenchymal stem cells (MSCs). The Pdots exhibit narrow-band emission at 775 nm with a quantum yield of 22%, among the highest value for various NIR probes. The Pdots together with a cell penetrating peptide are able to track stem cells over two weeks without disturbing their multipotent properties, as confirmed by the analyses on cell proliferation, differentiation, stem-cell markers, and immunophenotyping. The in vivo cell tracking was demonstrated in a liver-resection mouse model, which indicated that the Pdot-labeled MSCs after tail-vein transplantation were initially trapped in lung, gradually migrated to the injured liver, and then proliferated into cell clusters. Liver-function analysis and histological examination revealed that the inflammation induced by liver resection was apparently decreased after stem cell transplantation. With the bright labeling, superior biocompatibility, and long-term tracking performance, the Pdot probes are promising for stem cell research and regenerative medicine.

Myostatin inhibits osteoblastic differentiation by suppressing osteocyte-derived exosomal microRNA-218: A novel mechanism in muscle-bone communication.

Muscle and bone are closely associated in both anatomy and function, but the mechanisms that coordinate their synergistic action remain poorly defined. Myostatin, a myokine secreted by muscles, has been shown to inhibit muscle growth, and the disruption of the myostatin gene has been reported to cause muscle hypertrophy and increase bone mass. Extracellular vesicle-exosomes that carry microRNA (miRNA), mRNA, and proteins are known to perform an important role in cell-cell communication. We hypothesized that myostatin may play a crucial role in muscle-bone interactions and may promote direct effects on osteocytes and on osteocyte-derived exosomal miRNAs, thereby indirectly influencing the function of other bone cells. We report herein that myostatin promotes expression of several bone regulators such as sclerostin (SOST), DKK1, and RANKL in cultured osteocytic (Ocy454) cells, concomitant with the suppression of miR-218 in both parent Ocy454 cells and derived exosomes. Exosomes produced by Ocy454 cells that had been pretreated with myostatin could be taken up by osteoblastic MC3T3 cells, resulting in a marked reduction of Runx2, a key regulator of osteoblastic differentiation, and in decreased osteoblastic differentiation via the down-regulation of the Wnt signaling pathway. Importantly, the inhibitory effect of myostatin-modified osteocytic exosomes on osteoblast differentiation is completely reversed by expression of exogenous miR-218, through a mechanism involving miR-218-mediated inhibition of SOST. Together, our findings indicate that myostatin directly influences osteocyte function and thereby inhibits osteoblastic differentiation, at least in part, through the suppression of osteocyte-derived exosomal miR-218, suggesting a novel mechanism in muscle-bone communication.

Multilayered upconversion nanocomposites with dual photosensitizing functions for enhanced photodynamic therapy.

The energy transfer from upconversion nanoparticles (UCNPs) to photosensitizers has been widely used for generating reactive oxygen species (ROS) in photodynamic therapy (PDT) by near infrared (NIR) excitation. However, the poor spectral overlap of lanthanide ions and conventional photosensitizers leads to low PDT efficiency. In this study, we construct a multilayered upconversion nanoplatform with dual photosensitizers to efficiently use the UV and visible upconversion emissions in NIR-responsive PDT. The nanoplatform consists of three functional layers, which are the upconversion nanoparticle as a core, and light-sensitive conjugated polymer and apo-transferrin-titanocene (Tf@Tc) as shells. Under NIR irradiation, apparent energy transfer occurs from the core to the polymer and Tc components in the shell, producing reactive oxygen species and free radicals for cancer cell killing. In vitro cellular assays show the synergistic therapeutic effect of the conjugated polymer and Tc as photosensitizers. In vivo animal studies show that tumor growth is significantly inhibited in the mice receiving the theranostic platform and NIR irradiation. Based on these observations, the multilayered upconversion nanocomposites can find potential applications in NIR-mediated anti-tumor therapy.

Synthesis of mesoporous-silica-coated Gd2O3:Eu@silica particles as cell imaging and drug delivery agents.

Mesoporous-silica-coated Gd2O3:Eu/silica nanoparticles were synthesized by a multistep chemical process and characterized by XRD, TEM and N2 adsorption/desorption isotherms in terms of size, morphology and porosity. The core Gd2O3:Eu obtained by this method was highly luminescent upon excitation, giving the function of cell imaging upon incubation with the human cervical carcinoma (HeLa) cells. The outer porous silica shell is able to load the anticancer drug with a relatively high loading efficiency and release the loaded drugs at a sustained rate. The HeLa cells can be killed effectively on incubation with the core-shell porous particles loaded with the anticancer drug DOX. Meanwhile, the accumulation of mesoporous nanoparticles loaded with drugs in the target location could be monitored via fluorescence imaging. Therefore, the core-shell hybrid nanoparticles presented in this work are potential multifunctional biomaterials for smart detection or diagnosis and therapy in future biomedical engineering.

A Soluble Activin Receptor IIB Fails to Prevent Muscle Atrophy in a Mouse Model of Spinal Cord Injury.

Myostatin (MST) is a potent regulator of muscle growth and size. Spinal cord injury (SCI) results in marked atrophy of muscle below the level of injury. Currently, there is no effective pharmaceutical treatment available to prevent sublesional muscle atrophy post-SCI. To determine whether inhibition of MST with a soluble activin IIB receptor (RAP-031) prevents sublesional SCI-induced muscle atrophy, mice were randomly assigned to the following groups: Sham-SCI; SCI+Vehicle group (SCI-VEH); and SCI+RAP-031 (SCI-RAP-031). SCI was induced by complete transection at thoracic level 10. Animals were euthanized at 56 days post-surgery. RAP-031 reduced, but did not prevent, body weight loss post-SCI. RAP-031 increased total lean tissue mass compared to SCI-VEH (14.8%). RAP-031 increased forelimb muscle mass post-SCI by 38% and 19% for biceps and triceps, respectively (p < 0.001). There were no differences in hindlimb muscle weights between the RAP-031 and SCI-VEH groups. In the gastrocnemius, messenger RNA (mRNA) expression was elevated for interleukin (IL)-6 (8-fold), IL-1ß (3-fold), and tumor necrosis factor alpha (8-fold) in the SCI-VEH, compared to the Sham group. Muscle RING finger protein 1 mRNA was 2-fold greater in the RAP-031 group, compared to Sham-SCI. RAP-031 did not influence cytokine expression. Bone mineral density of the distal femur and proximal tibia were decreased post-SCI (-26% and -28%, respectively) and were not altered by RAP-031. In conclusion, MST inhibition increased supralesional muscle mass, but did not prevent sublesional muscle or bone loss, or the inflammation in paralyzed muscle.

Amplified Singlet Oxygen Generation in Semiconductor Polymer Dots for Photodynamic Cancer Therapy.

This paper described the energy-transfer amplified singlet oxygen generation in semiconductor polymer dots (Pdots) for in vitro and in vivo photodynamic therapy. Hydrophobic photosensitizer tetraphenylporphyrin was facilely doped in the nanoparticles consisting of densely packed semiconductor polymers. Optical characterizations indicated that the fluorescence of Pdots was completely quenched by the photosensitizer, yielding an energy transfer efficiency of nearly 100% and singlet-oxygen generation quantum yield of ∼50%. We evaluated the cellular uptake, dark toxicity, and photodynamic therapy of the Pdot photosensizer in human gastric adenocarcinoma cells. The in vitro studies indicated that cancer cells were efficiently destroyed at very low dose of the Pdots such as 1 µg/mL by using the light dose of 90 J/cm(2), which is considerably less than that in clinical practice. The antitumor effect of the Pdots was further evaluated in vivo with human gastric adenocarcinoma xenografts in Balb/c nude mice, which show that the xenograft tumors were significantly inhibited and eradicated in some cases. Our results indicate the energy transfer amplified Pdot platforms have great therapeutic potential for treating malignant cancers.

Sclerostin antibody preserves the morphology and structure of osteocytes and blocks the severe skeletal deterioration after motor-complete spinal cord injury in rats.

Unloading, neural lesions, and hormonal disorders after acute motor-complete spinal cord injury (SCI) cause one of the most severe forms of bone loss, a condition that has been refractory to available interventions tested to date. Thus, these features related to acute SCI provide a unique opportunity to study complex bone problems, potential efficacious interventions, and mechanisms of action that are associated with these dramatic pathological changes. This study was designed to explore the therapeutic potential of sclerostin antibody (Scl-Ab) in a rat model of bone loss after motor-complete SCI, and to investigate mechanisms underlying bone loss and Scl-Ab action. SCI rats were administered Scl-Ab (25 mg/kg/week) or vehicle beginning 7 days after injury then weekly for 7 weeks. SCI resulted in significant decreases in bone mineral density (-25%) and trabecular bone volume (-67%) at the distal femur; Scl-Ab completely prevented these deteriorations of bone in SCI rats, concurrent with markedly increased bone formation. Scanning electron microscopy revealed that SCI reduced numbers of osteocytes and dendrites concomitant with a morphology change from a spindle to round shape; Scl-Ab corrected these abnormalities in osteocytes. In ex vivo cultures of bone marrow cells, Scl-Ab inhibited osteoclastogenesis, and promoted osteoblastogenesis accompanied by increases in mRNA levels of LRP5, osteoprotegerin (OPG), and the OPG/RANKL ratio, and a decrease in DKK1 mRNA. Our findings provide the first evidence that robust bone loss after acute motor-complete SCI can be blocked by Scl-Ab, at least in part, through the preservation of osteocyte morphology and structure and related bone remodeling. Our findings support the inhibition of sclerostin as a promising approach to mitigate the striking bone loss that ensues after acute motor-complete SCI, and perhaps other conditions associated with disuse osteoporosis as a consequence of neurological disorders.

Semiconductor polymer dots induce proliferation in human gastric mucosal and adenocarcinoma cells.

We investigated the cellular uptake behavior and cell viability of semiconducting polymer dots (Pdots) on human gastric adenocarcinoma (SGC-7901) cells and human gastric mucosal (GES-1) cells. MTT studies indicate the Pdot treatment induces obvious cell proliferation in both types of cell lines. We performed further investigations such as reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP) change, which indicate that the cell proliferation is in good agreement with the increase in the ROS and MMP levels. Moreover, expression of protein kinase B (Akt) decreased as the Pdot concentration increases, but the expression of protein dual-phosphorylated Erk (p-Erk) and phosphorylated c-Jun N-terminal kinases (p-JNK) were increased. These effects indicated that the Pdots could promote the growth of SGC-7901 cells and GES-1 cells by appropriately regulating the expressions of protein Akt, p-Erk, and p-JNK.

Identification of functional glucocorticoid response elements in the mouse FoxO1 promoter.

Glucocorticoids stimulate muscle atrophy through a cascade of signals that includes activation of FoxO transcription factors which then upregulate multiple genes to promote degradation of myofibrillar and other muscle proteins and inhibit protein synthesis. Our previous finding that glucocorticoids upregulate mRNA levels for FoxO1 in skeletal muscle led us to hypothesize that the FoxO1 gene contains one or more glucocorticoid response elements (GREs). Here we show that upregulation of FoxO1 expression by glucocorticoids requires the glucocorticoid receptor (GR) and binding of hormones to it. In cultured C2C12 myoblasts dexamethasone did not alter FoxO1 mRNA stability. Computational analysis predicted that the proximal promoter of the FoxO1 gene contained a cluster of eight GRE half sites and one highly conserved near-consensus SRE; the cluster is found between -800 and -2000bp upstream of the first codon of the FoxO1 gene. A reporter gene constructed using the first 2kb of the FoxO1 promoter was stimulated by dexamethasone. Removal of a 5' domain containing half of the GREs reduced reporter gene activity and removal of all GREs in this region ablated activation by dexamethasone. Restriction fragments of the cluster of 8 upstream GREs bound recombinant GR in gel shift assays. Collectively, the data demonstrate that the proximal promoter of the FoxO1 gene contains multiple functional GREs, indicating that upregulation of FoxO1 expression by glucocorticoids through GREs represents an additional mechanism by which the GR drives glucocorticoid-mediated muscle atrophy. These findings are also relevant to other physiological roles of FoxO1 such as regulation of hepatic metabolism.