Human Bone Marrow Neuroblastoma Research Articles

Neuroprotective Effects of the Psychoactive Compound Biatractylolide (BD) in Alzheimer’s Disease

Mitochondria play a central role in the survival or death of neuronal cells, and they are regulators of energy metabolism and cell death pathways. Many studies support the role of mitochondrial dysfunction and oxidative damage in the pathogenesis of Alzheimer's disease. Biatractylolide (BD) is a kind of internal symmetry double sesquiterpene novel ester compound isolated from the Chinese medicinal plant Baizhu, has neuroprotective effects in Alzheimer's disease. We developed a systematic pharmacological model based on chemical pharmacokinetic and pharmacological data to identify potential compounds and targets of Baizhu. The neuroprotective effects of BD in PC12 (rat adrenal pheochromocytoma cells) and SH-SY5Y (human bone marrow neuroblastoma cells) were evaluated by in vitro experiments. Based on the predicted results, we selected 18 active compounds, which were associated with 20 potential targets and 22 signaling pathways. Compound-target, target-disease and target-pathway networks were constructed using Cytoscape 3.2.1. And verified by in vitro experiments that BD could inhibit Aβ by reducing oxidative stress and decreasing CytC release induced mPTP opening. This study provides a theoretical basis for the development of BD as an anti-Alzheimer's disease drug.

Fabrication of Tunable 3D Cellular Structures in High Volume Using Magnetic Levitation Guided Assembly.

Tunable and reproducible size with high circularity is an important limitation to obtain three-dimensional (3D) cellular structures and spheroids in scaffold free tissue engineering approaches. Here, we present a facile methodology based on magnetic levitation (MagLev) to fabricate 3D cellular structures rapidly and easily in high-volume and low magnetic field. In this study, 3D cellular structures were fabricated using magnetic levitation directed assembly where cells are suspended and self-assembled by contactless magnetic manipulation in the presence of a paramagnetic agent. The effect of cell seeding density, culture time, and paramagnetic agent concentration on the formation of 3D cellular structures was evaluated for NIH/3T3 mouse fibroblast cells. In addition, magnetic levitation guided cellular assembly and 3D tumor spheroid formation was examined for five different cancer cell lines: MCF7 (human epithelial breast adenocarcinoma), MDA-MB-231 (human epithelial breast adenocarcinoma), SH-SY5Y (human bone-marrow neuroblastoma), PC-12 (rat adrenal gland pheochromocytoma), and HeLa (human epithelial cervix adenocarcinoma). Moreover, formation of a 3D coculture model was successfully observed by using MDA-MB-231 dsRED and MDA-MB-231 GFP cells. Taken together, these results indicate that the developed MagLev setup provides an easy and efficient way to fabricate 3D cellular structures and may be a feasible alternative to conventional methodologies for cellular/multicellular studies.

Chemical constituents with cytotoxic and anti-inflammatory activity in Hypericum sampsonii and the antitumor potential under the view of cancer-related inflammation

Ethnopharmacological relevanceChronic inflammation has an important role in the development of cancers. Hypericum sampsonii, known as “Yuanbao Cao”, is mainly distributed in the southwest of China. As a folk medicinal plant, “Yuanbao Cao” is traditionally used for treatment of various inflammation-related diseases including swelling, burns, arthritis, and dermatitis, etc. The plant is a promising anticancer herb. However, there is no research on the antitumor potential of this plant from the view of cancer-related inflammation strategy. Aim of the studyTo explore the H. sampsonii in relation to cancer-related chemical constituents with anti-inflammatory and cytotoxic activity in cancer-related inflammation. Materials and methodsThe chemical constituents of H. sampsonii were isolated by repeated chromatography techniques, and their structures were identified mainly by spectroscopic methods and compared to published data. The chemical profile of the herb was analyzed using HPLC. The cytotoxicities of compounds against five cancer cell lines: human melanoma cell (A375), human breast cancer cell (MDA-MB-231), human gastric cancer cell (SGC-7901), human colon cancer cell (SiHa), and human bone marrow neuroblastoma cell (SHSY–5Y), were tested using MTT assay; their anti-inflammatory activities were evaluated by inhibition on NO production in LPS-stimulated RAW 264.7, THP-1 and BV-2 microglial cells. ResultsTwenty-five compounds, including four phenols (1–4), two anthraquinonoids (5 and 6), six xanthones (7–12), one benzophenone (13), one phloroglucinol (14), nine flavonoids (15–23), one sterol (24) and one alkaloid (25), were isolated from the EtOH extract of H. sampsonii. Of them, compounds 3, 4, 6, 7, 10–14, 17, 19, 22 and 23 were reported in H. sampsonii for the first time. HPLC analysis showed that flavonoids were the main constituents in the herb. MTT assay revealed that compounds 1, 2, 5–14, 15, 17, 18, 20, 21, 22 and 25 had selective cytotoxic activities (IC50: 7.52–158.90 μM) against tested cancer cells, in which compound 5, 6, 13 and 14 displayed high activities against A375, MDA-MB-231, SiHa and SHSY-5Y. In the screening experiment of anti-inflammatory activity, most compounds (1–2, 5–23) showed considerable high anti-inflammatory activities (IC50: 10.59–42.75 μM), in which compounds 5, 6, 13, 14, and 15 exhibited high anti-inflammatory activities in LPS-stimulated RAW264.7, THP-1 and BV-2 microglial cells. ConclusionsCompounds 3, 4, 6, 7, 10–14, 17, 19, 22 and 23 were isolated for the first time from H. sampsonii. Compound 5, 6, 13 and 14 displayed high cytotoxic activities against the tested cancer cell lines. Compounds (1–2, 5–23) showed anti-inflammatory activities, of them, compounds 5, 6, 13, 14 and 15 exhibited the high activity. From the view of cancer-related inflammation point, not only the compounds with high cytotoxicity, but those compounds with anti-inflammatory activities, especially the flavonoids, contribute to the antitumor potential of H. sampsonii. The results and viewpoint of present study provide a different insight to better understand the antitumor potential of H. sampsonii, and may also promote the reasonable usage of this folk medical herb.

Inflammation and oxidative stress are key mediators in AKB48-induced neurotoxicity in vitro

Synthetic cannabinoids were introduced into market in early 2000s; since these “legal highs” are dramatically popular among youth, it becomes a deadly problem. Synthetic cannabinoids have high affinity to cannabinoid receptors; leading to various clinical symptoms. AKB48 (Apinaca) has been classified as a third-generation synthetic cannabinoid for the first time in 2014.The toxicity profile of AKB48 is unclear due to little information that mainly obtained from clinical and forensic cases; however, it is believed to be similar with other psychoactive substances. Thus, we aimed to investigate the possible toxicity mechanisms of AKB48 in SH-SY5Y (human bone marrow neuroblastoma) cell line. IC50 value of AKB48 was calculated as 160.91 μM by MTT assay. AKB48 treatment enhanced (≥1.2-fold) the fluorescence intensity indicating increased reactive oxygen species production; however, glutathione levels did not changed in the range of 25–200 μM exposure concentrations. Cannabinoid type-1 receptor (CB1) expression was increased ≥15-fold in the range of 25–50 μM of AKB48, while cannabinoid type-2 receptor (CB2) did not expressed in SH-SY5Y cells. Interleukin-6 (IL-6) and tumor necrosis factor-α (TNF- α) were up-regulated with a dose-dependent manner, and the profiles were almost identical; however, mitogen-activated protein kinase 8 (MAPK 8) was only upregulated with 25 μM of AKB48 and nuclear factor kappa B (NF-ĸB) did not change. Our results should raise the concerns about the safety associated with synthetic cannabinoids uses.

Zbtb38 is a novel target for spinal cord injury.

Spinal cord injury (SCI) is currently incurable since treatments applied to clinic are limited to minimizing secondary complications and the mechanisms of injury-induced spinal cord damage are poorly understood. Zbtb38, also called CIBZ, is highly expressed in spinal cord and it functions as a negative regulator in SCI-induced apoptosis. We show here that Zbtb38 is downregulated under endoplasmic reticulum (ER) stress, which promotes ER stress-associated apoptosis in human bone marrow neuroblastoma cells. In the traumatic SCI mice, ER stress presented in injured spinal cord induced repression of Zbtb38 expression and triggered Zbtb38-mediated apoptosis. ChIP-QPCR analysis revealed that ATF4, an ER-stress inducible transcription factor, directly activated Zbtb38 transcription by binding to the Zbtb38 promoter. However, this binding was significantly reduced following SCI, leading to a sharp decrease in Zbtb38 expression. Restoring Zbtb38 function in injured spinal cord by injection of lentivirus containing Zbtb38 into SCI mice, significantly alleviated secondary damage of spinal cord with decreased ER stress-associated apoptosis and partially recovered spinal cord functions. These findings demonstrate that restoration of Zbtb38 expression can reduce secondary tissue damage after SCI, and suggest that a therapeutic strategy for targeting Zbtb38 may promote functional recovery of spinal cord for patients with SCI.

Biatractylolide Modulates PI3K-Akt-GSK3β-Dependent Pathways to Protect against Glutamate-Induced Cell Damage in PC12 and SH-SY5Y Cells.

Biatractylolide, isolated from the ethyl acetate extract of Atractylodes macrocephala, has shown various pharmacological activities such as antitumor and antioxidant activities. In this work, we aim to study the protective effect of biatractylolide on glutamate-induced rat adrenal pheochromocytoma cell (PC12) and human bone marrow neuroblastoma cell line (SH-SY5Y) injury and preliminarily explore its mechanism. The results showed that glutamate was cytotoxic with an inhibitory concentration 50% (IC50) of 8.5 mM in PC12 and 10 mM in SH-SY5Y cells. In this work, the preincubation with biatractylolide (10, 15, and 20 μM) observably improved cell viability, inhibited the apoptosis of cells induced by glutamate, and reduced the activity of LDH. AO staining revealed that apoptosis of cells was decreased. Additionally, the results of western blotting manifested that pretreatment with biatractylolide could downregulate GSK3β protein expression and upregulate p-Akt protein expression, thereby protecting PC12 and SH-SY5Y cells from injury. All these findings indicate that biatractylolide has a neuroprotective effect on glutamate-induced injury in PC12 and SH-SY5Y cells through a mechanism of the PI3K-Akt-GSK3β-dependent pathways.

Human umbilical cord mesenchymal stem cells infected with adenovirus expressing HGF promote regeneration of damaged neuron cells in a Parkinson's disease model.

Parkinson's disease (PD) is a neurodegenerative movement disorder that is characterized by the progressive degeneration of the dopaminergic (DA) pathway. Mesenchymal stem cells derived from human umbilical cord (hUC-MSCs) have great potential for developing a therapeutic agent as such. HGF is a multifunctional mediator originally identified in hepatocytes and has recently been reported to possess various neuroprotective properties. This study was designed to investigate the protective effect of hUC-MSCs infected by an adenovirus carrying the HGF gene on the PD cell model induced by MPP+ on human bone marrow neuroblastoma cells. Our results provide evidence that the cultural supernatant from hUC-MSCs expressing HGF could promote regeneration of damaged PD cells at higher efficacy than the supernatant from hUC-MSCs alone. And intracellular free Ca2+ obviously decreased after treatment with cultural supernatant from hUC-MSCs expressing HGF, while the expression of CaBP-D28k, an intracellular calcium binding protein, increased. Therefore our study clearly demonstrated that cultural supernatant of MSC overexpressing HGF was capable of eliciting regeneration of damaged PD model cells. This effect was probably achieved through the regulation of intracellular Ca2+ levels by modulating of CaBP-D28k expression.

Cellular prion protein imaging analysis with aptamer-labeled Ru(bpy) 3 2+ -doped silica nanoparticles

A new type of highly selective aptamer-labeled fluorescent silica nanoparticles [Apt-tris(2,2′-bipyridyl)ruthenium(II)@SiO2 NPs] were prepared through the reverse microemulsion method by using prolonged fluorescence lifetime ruthenium complexes of tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy) 3 2+ ) as the source of the fluorescence for cellular prion protein imaging. Investigations showed that the newly prepared Ru(bpy) 3 2+ @SiO2 NPs possessed superior advantages of strong fluorescence, low toxicity, and easy surface modification for bioconjugation. Cell imaging experiments indicated that Apt-Ru(bpy) 3 2+ @SiO2 NPs had great tendency to human bone marrow neuroblastoma cells (SK-N-SH cells), since they can express large amount of prion protein on the surface of the cell, while in HeLa cells this phenomenon disappeared for the reason that HeLa cells cannot express prion proteins.

Reactive oxygen species(ROS) involving in inhibiting proliferation of human bone marrow neuroblastoma cells (SH-SY5Y) by giant typhonium rhizome water extract

Objective To investigate whether the reactive oxygen species involved in inhibiting proliferation of human bone marrow ceils of neuroblastoma cells （SH-SY5Y） process by giant typhonium rhizome water extract, and to investigate the initial mechanism of anti-tumor effect of giant typhonium rhizome in vitro. Methods After giant typhonium rhizome water extract processing of human bone marrow neuroblastoma cells （SH-SY5Y）at 24 h and 48 h, the cells were collected, and western blotting reaction methods such as flow cytometry was used to study the levels of ROS and SH-SY5Y apoptosis. Results The levels of ROS elevated fxom before dosing （4.9± 1.6）% to （39.7±11.8）% in the process of giant typhonium rhizome water extract induced apoptosis in SH-SY5Y; after ROS inhibition was added, SH-SY5Y apoptosis level induced by giant typhonium rhizome water extract had decreased. Conclusion ROS involved in the process of giant typhonium rhizome water extract inhibiting cell proliferation in SH-SY5Y. Key words: Giant typhonium rhizome; Reactive oxygen species; Human bone marrow neuroblastoma cells： Flow cytometry

Toxicity of graphene oxide and multi-walled carbon nanotubes against human cells and zebrafish

Graphene possesses unique physical and chemical properties, which have inspired a wide range of potential biomedical applications. However, little is known about the adverse effects of graphene on the human body and ecological environment. The purpose of our work is to make assessment on the toxicity of graphene oxide (GO) against human cell line (human bone marrow neuroblastoma cell line and human epithelial carcinoma cell line) and zebrafish (Danio rerio) by comparing the toxic effects of GO with its sister, multi-walled carbon nanotubes (MWNTs). The results show that GO has a moderate toxicity to organisms since it can induce minor (about 20%) cell growth inhibition and slight hatching delay of zebrafish embryos at a dosage of 50 mg/L, but did not result in significant increase of apoptosis in embryo, while MWNTs exhibit acute toxicity leading to a strong inhibition of cell proliferation and serious morphological defects in developing embryos even at relatively low concentration of 25 mg/L. The distinctive toxicity of GO and MWNTs should be ascribed to the different models of interaction between nanomaterials and organisms, which arises from the different geometric structures of nanomaterials. Collectively, our work suggests that GO does actual toxicity to organisms posing potential environmental risks and the result is also shedding light on the geometrical structure-dependent toxicity of graphitic nanomaterials.

Highly selective light scattering imaging of chromium (III) in living cells with silver nanoparticles

In this contribution, we present a highly selective chromium ion (Cr(3+))-induced aggregation of citrate-capped silver nanoparticles, which could be applied for the imaging of the distribution of Cr(3+) in cells. It was found that selective aggregation of citrate-capped silver nanoparticles occurs at room temperature in the presence of Cr(3+) in aqueous medium of pH 6.8, resulting in color change from yellow to pink in 10 min and enhanced localized surface plasmon resonance (LSPR) scattering signals. Tenfold of other metal ions including Al(3+), Ca(2+), Co(2+), Cu(2+), Fe(2+), Fe(3+), Hg(2+), La(3+), Mg(2+), Ni(2+), Pb(2+), Tb(3+) and Zn(2+) had no response. Mechanism analysis showed that the aggregation is mainly dependent on the chelation of Cr(3+) ion with the citrate ion capped on silver nanoparticles, forming crosslinking aggregates of silver nanoparticles. With the Cr(3+)-induced enhancement of LSPR scattering signals, Cr(3+) in cytoplasm of human bone marrow neuroblastoma cells could be imaged with dark-field light scattering imaging technique.

Aptamer-Based Silver Nanoparticles Used for Intracellular Protein Imaging and Single Nanoparticle Spectral Analysis

Aptamer-adapted silver nanoparticles (Apt-AgNPs) were developed as a novel optical probe for simultaneous intracellular protein imaging and single nanoparticle spectral analysis, wherein AgNPs act as an illuminophore and the aptamer as a biomolecule specific recognition unit, respectively. It was found that streptavidin-conjugated and aptamer-functionalized AgNPs show satisfactory biocompatibility and stability in cell culture medium, and thus not only can act as a high contrast imaging agent for both dark-field light scattering microscope and TEM imaging but also can inspire supersensitive single nanoparticle spectra for potential intercellular microenvironment analysis. Further investigations showed that caveolae-related endocytosis is likely a necessary pathway for Apt-AgNPs labeled PrP(c) internalization in human bone marrow neuroblastoma cells (SK-N-SH cells). The integrated capability of Apt-AgNPs to be used as light scattering and TEM imaging agents, along with their potential use for single nanoparticle spectral analysis, makes them a great promise for future biomedical imaging and disease diagnosis.

Cuprous oxide nanospheres as probes for light scattering imaging analysis of live cells and for conformation identification of proteins

A facile solution-phase synthesis route of highly uniform Cu 2O nanospheres (Cu 2O NPs) with the size of 57.7 ± 4.7 nm was developed, and then the nanoparticles were applied to live cell imaging under a common dark-field microscope. Starting from copper(II) salts, the synthesis of Cu 2O NPs was made in the presence of cetyltrimethylammonium bromide (CTAB) by reducing the copper(II) with sodium borohydride (NaBH 4) in aqueous medium and by aging process in the air. Monitoring of morphology evolution process of Cu 2O NPs with scanning electron microscopy (SEM) and measuring of the UV–visible spectra showed that the synthesis of Cu 2O NPs follows the reduction–oxidation coupled process of Cu 2+ into Cu 0 species at first and then the resulted Cu 0 species into Cu 2O NPs in the air. Light scattering (LS) features have been measured with a common spectrofluorometer and a common dark-field microscope, and it was found that the as-prepared Cu 2O NPs display strong blue scattering light and can be applied for cell imaging. If incubated with human bone marrow neuroblastoma, transferrin-conjugated Cu 2O NPs can get into the cells and show strong pure blue light in cytoplasm. Further investigations showed that the Cu 2O NPs could be applied for probes for conformation of proteins.