Growth Of PC12 Cells Research Articles

Antiproliferative Effects of <i>Ferula assa-foetida’s</i> Extract on PC12 and MCF7 Cancer Cells

Background: Ferula assa-foetida is a herbaceous plant of the Umbelliferae family having a broad spectrum of biological activities such as antiseptic, antibacterial, anti-inflammation, and, anti-tumor activity against a wide range of cancers. Among these features, antitumor activity has become more important in recent years and it still demands more investigations to address the underlying mechanisms. Purpose: This current study was conducted to evaluate the anti-proliferative effect of Ferula assa-foetida on PC12 and MCF7 cells as well as examining its mechanisms. Materials and Methods: Cells were cultured in DMEM medium with 10% fetal bovine serum, 100 units/ml penicillin, and 100 µg/ml streptomycin. Cells incubated with different concentrations of the ethanolic extract of Ferula assa-foetida. Notably, cytotoxicity and apoptosis assay were measured by MTT and PI staining, respectively. Results: The MTT results showed that the ethanolic extract of Ferula assa-foetida in concentrations of 10, 7, 5, and 2.5 μM on both PC12 and MCF7 cells had a significant effect in cell viability and apoptosis induction in comparison to control group. Conclusion: In this study, it was determined that Ferula assa-fetida through the induction of apoptosis prevented the growth of PC12 and MCF7 cells and made a reduction in cell viability with different concentrations in a time- and dose-dependent manner. However, more studies are needed to reveal the mechanisms of Ferula assa-foetida’s extract in apoptosis induction.

The Efficacy of Electrospun PAN/Kefiran Nanofiber and Kefir in Mammalian Cell Culture: Promotion of PC12 Cell Growth, Anti-MCF7 Breast Cancer Cells Activities, and Cytokine Production of PBMC.

BackgroundKefiran is a useful polysaccharide made of branched glucogalactose which is produced by microorganisms. Here the anti-MCF-7 breast cancer cells activity of kefiran and cytokine productions (IL-6) of peripheral blood mononuclear cells (PBMC) treated by kefiran was studied. Also, the effect of using kefiran as a useful and cost-effective scaffold in neural stem cell culture (PC12 cell culture) was investigated.Material and MethodsKefiran was produced from raw milk with 0.5% fat and 10 g of kefir grains. After incubation for 48 hrs at room temperature, the solvent collected (crude kefiran). These samples were kept at 100°C for 1 hr (boiled kefiran) and the supernatant was precipitated by ethanol (pure kefiran). Then, the electrospun nanofibers, pure polyacrylonitrile (PAN), PAN/kefiran 5%, and PAN/kefiran 10% were fabricated and used as scaffolds in the cell culture. The structure of fabricated was studied by SEM and the cytokine production (IL-6) in vitro in the cell culture supernatant of PBMC line after treatment with kefiran (1mg/mL, 5 mg/mL) and kefiran-PAN 5% and 10% were carried out by enzyme-linked immunosorbent assay (ELISA). The attachment of PC12 cells was examined by inverted microscope. Also, cytotoxicity of kefiran for PC12 and MCF7 cells and morphological changes of PC12 cells were evaluated by MTT and Cresyl violet staining (Nissl staining) respectively.ResultsThe mean diameter of fabricated PAN/kefiran 5% and 10% nanofibers were 310.2±43.97 nm. The contact angle measurement results (26.9± 1.9 for the pure PAN scaffold vs 12.3± 1.13 for the PAN/kefiran) revealed enhanced hydrophilicity of scaffolds upon the incorporation of kefiran and PAN. Seeding of PC12 cells on the scaffolds showed that fabrication of kefiran into PAN led to the enhancement of cell attachment, proliferation, and morphological changes. Also, the promotion of PBMC growth and decreasing of MCF7 cell lines viability were shown through MTT assay. No significant changes were measured for the level of IL-6 in PAN/kefiran 5% treated cells compared to the control (p ≥ 0.05).ConclusionThese results suggest superior properties of kefiran/PAN nanofibrous scaffolds for the neural stem cell culture especially for repairing injured spinal cord. Also, the pure kefiran could be used for the enhancement of PBMC growth and reducing the MCF7 cancerous cells growth. So, using biocompatible, anti-bacterial, and anti-tumor kefiran/PAN nanofibers for regenerative medicine seems promising.

The role of DFO in Al (mal) (3)-induced ferroptosis in PC12 cells

Objective: To investigate the mechanism of Al (mal) (3)-induced ferroptosis in rat adrenal pheochromocytoma cells (PC12), to explore the effect of deferoxamine (DFO) . Methods: Taken PC12 cells growing at logarithmic phase and divided into 6 groups: control group, 200 μmol/L Al (mal) (3) group, 0.5% DMSO group, 200 μmol/L DFO group, Al (mal) (3)+DMSO group, Al (mal) (3)+DFO group. DMSO and DFO were added to the DMSO group and the Al (mal) (3)+DMSO group, the DFO group and the Al (mal) (3)+DFO group for 2 h, respectively, Al (mal) (3) was then added to the Al (mal) (3) group, Al (mal) (3)+DMSO group, and the Al (mal) (3)+DFO group to a final concentration of 200 μmol/L. The cell viability was detected by CCK8, the morphology and ROS levels of PC12 cells was observed by inverted microscope, the cell proliferation toxicity and intracellular iron ion content were detected by colorimetry, the GSH content and GSH-PX activity were detected by biochemical method. Results: Al (mal) (3) exposure significantly inhibited the growth of PC12 cells and destroyed the cell morphological structure, resulting in increased LDH activity and intracellular iron ion content in PC12 cells, decreased GSH content and GSH-PX activity, increased ROS levels; the combined treatment of Al (mal) (3)+DFO can significantly improve the cell viability of PC12 cells, improved cell morphology, decreased cell LDH activity and intracellular iron ion content (P>0.05), increased GSH content and GSH-PX activity, decreased ROS levels. Conclusion: Al (mal) (3) can induce ferroptosis in PC12 cells, DFO may inhibit ferroptosis by reducing intracellular iron levels and reducing oxidative damage.

Protective effect of ISO‑1 against advanced glycation end product aggravation of PC12 cell injury induced by Aβ1‑40.

Advanced glycation end products (AGEs) are important pathogenic substances involved in diabetes mellitus (DM) and its complications. AGEs also serve important roles in promoting the development of Alzheimer's disease (AD). Macrophage migration inhibitory factor (MIF), an inflammatory stimulant and a pathogenic factor involved in DM, was previously reported to be present at increased levels in the cerebrospinal fluid of patients with AD and mild cognitive impairment compared with age-matched healthy controls. By investigating the association between AGEs and MIF, and the effects of neuroinflammation on AD, the present study aimed to increase understanding of the specific molecular mechanisms involved in the pathogenesis of DM and AD, and the connection between these diseases. PC12 cells were cultured in vitro; the levels of MIF mRNA and protein were determined using reverse transcription-quantitative (RT-q)PCR and western blot analyses. The optimal concentrations of AGEs and amyloid β 1–40 (Aβ1–40) were also determined in the cell model of AD using Cell Counting Kit-8 and MTT assays. Cell numbers and morphological changes were observed following the treatment of Aβ1–40-stimulated PC12 cells with AGEs and the MIF inhibitor (S,R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1). The mRNA expression levels of interleukin (IL)-1β, IL-6, tumor necrosis factor-α (TNF-α) and MIF were determined via RT-qPCR analysis. The results showed that the levels of MIF mRNA and protein were significantly increased in cells treated with AGEs compared with the control group. In the AD model group, the inhibition of PC12 cell growth was significantly increased, and the mRNA expression levels of IL-1β, IL-6, TNF-α and MIF were also increased. Compared with treatment with AGEs alone, the combination of AGEs treatment with ISO-1 significantly improved the survival rate and resulted in the reduced expression of inflammatory mediators in the AD cell model. Thus, ISO-1 reduced AGEs-mediated damage in the AD cell model. This may be a consequence of AGEs-mediated MIF expression promoting neuritis in the AD cell model, whereas ISO-1 decreased the expression of neuroinflammatory mediators.

Polyaniline Functionalized Graphene Nanoelectrodes for the Regeneration of PC12 Cells via Electrical Stimulation.

The regeneration of neurons is an important goal of neuroscience and clinical medicine. The electrical stimulation of cells is a promising technique to meet this goal. However, its efficiency highly depends on the electrochemical properties of the stimulation electrodes used. This work reports on the preparation and use of a highly electroactive and biocompatible nanoelectrode made from a novel polyaniline functionalized graphene composite. This nanocomposite was prepared using a facile and efficient polymerization-enhanced ball-milling method. It was used to stimulate the growth of PC12 cells under various electrical fields. The enhanced growth of axons and improved wound regeneration of PC12 cells were observed after this treatment, suggesting a promising strategy for neuro traumatology.

Physiochemical and morphological dependent growth of NIH/3T3 and PC-12 on polyaniline-chloride/chitosan bionanocomposites

Biomimetic scaffolds inspired by fields and forces of the natural environment of cells is essential in tissue engineering. This study reports on controlled growth of two model cell lines, NIH/3T3 (promiscuous, fibroblast) and PC-12 (electroresponsive, neural progenitor) cells, given electrical and topographical cues that were delivered from a bionanocomposite of polyaniline-chloride and chitosan (PAn-Cl/CHI). The conductivity and morphology of the scaffold were controlled by varying the wt% of PAn-Cl (0–50 wt%) in CHI and processing methods, air-drying (nanofeatured) versus lyophilization (microporous-reticulated), respectively. Bionanocomposites supported the growth of both cell types independent of the availability of receptor-mediated ligands (laminin). NIH/3T3 cells were less elongated on lyophilized (microporous-reticulated) and more conductive (higher wt% PAn-Cl) composites. PC-12 cells had higher viability and less aggregation when grown on conductive substrates. Air-dried bionanocomposites were more supportive of growth but not attachment of PC-12 cells, suggesting that processing of composites could provide an additional level of engineering control to alter the PC-12 cell attachment and growth. In general, PC-12 cells responded more distinctly and dramatically to the substrate properties than NIH/3T3 cells, supporting a clear role for electrical conductivity on neural cell behavior. Nerve growth factor(NGF)-induced differentiation of PC-12 cells resulted in extensive neurite extension in the presence of adsorbed laminin. In a substrate composition-dependent manner, extension and rate of neurite outgrowth were higher when cultured on the conductive substrates. Overall, this study demonstrates the suitability of conductive PAn-Cl/CHI scaffold to host different cell types and support their responses.

Expression of PDK1 in malignant pheochromocytoma as a new promising potential therapeutic target

Phosphoinositide-dependent kinase 1 (PDK1) is highly expressed in many solid tumors. And several studies have demonstrated that PDK1 has been an emerging and promising target for anti-cancer therapies. However, the role of PDK1 has not been studied so far in malignant pheochromocytoma (PCC). In this study, immunohistochemical staining was performed to investigate the protein level of PDK1 in 63 PCC tissue samples, of which 49 were benign and 14 were malignant. In addition, we evaluated the effect of inhibition of PDK1 with siRNA on cell growth, apoptosis and invasive capacity in PC12 cells and identified the underlying mechanisms. We found that PDK1 was overexpressed in malignant PCC tissues, and knockdown of PDK1 with siRNA significantly inhibited cell proliferation, increased apoptosis induction, and attenuated cell migration and invasive capacity in PC12 cells. We also showed that knockdown of PDK1 significantly reduced the phosphorylation of Akt at threonine 308 (p-Akt T308) but did not alter the serine phosphorylation of Akt on the S473 site (p-Akt S473). Furthermore, we found that the p-Akt expression was noticeably decreased after knockdown of PDK1, but the t-Akt expression did not show a significant decrease. We have demonstrated for the first time that PDK1 is overexpressed in human malignant PCC and plays an important role in the malignant biological behaviors of PC12 cell. Specifically, we have revealed that knockdown of PDK1 could attenuate activation of the Akt signaling. These data suggest that PDK1 could be a new promising potential therapeutic target in human cancer treatment for malignant PCC.

Tanshinone IIA Attenuates Insulin Like Growth Factor 1 -Induced Cell Proliferation in PC12 Cells through the PI3K/Akt and MEK/ERK Pathways.

The insulin like growth factor 1 (IGF-1) and its receptor (IGF-1R) facilitate tumor proliferation and progression. Tanshinone IIA (TSN) is an active diterpene quinone isolated from the roots of the herbal plant Salvia miltiorrhiza. TSN inhibits the proliferation of various types of cancer cells but its role in the IGF-1R-induced proliferation of pheochromocytoma (PC12) cells and the potential mechanisms are largely unknown. This study aims to investigate the anti-proliferative effect of TSN in PC12 cells and its role on IGF-1R signaling transduction. PC12 cells were treated with IGF-1 with or without TSN, methyl thiazolytetrazolium (MTT) assay, and cell counting kit-8 and flow cytometry were used to evaluate the proliferation of PC12 cells. The role of TSN on the apoptosis of PC12 cells were detected by flow cytometry as well. The effects of TSN and IGF-1 on the phosphorylation of IGF-1R, protein kinase B (Akt), extracellular-signal related kinase 1/2 (ERK1/2) and other downstream targets were analyzed by Western blotting analysis. Our results showed that IGF-1 promoted the growth of PC12 cells in a dose-dependent manner and increased the phosphorylation of IGF-1R, whereas TSN attenuated the effect of IGF-1. Interestingly, TSN did not induce cell apoptosis in PC12 cells. Moreover, TSN attenuated the phosphorylation of Akt and ERK1/2 induced by IGF-1, and the phosphorylation of glycogen synthase kinase-3β, forkhead box O3a (FOXO3a) and c-Raf were also inhibited by TSN. Furthermore, TSN inhibited cell growth induced by IGF-1 and blocked the activation of IGF-1R in SH-SY5Y cells. Taken together, TSN has an inhibitory effect on the proliferation of PC12 cells via down-regulation of the phosphorylated IGF-1R and its downstream signaling.

Flupirtine derivatives as potential treatment for the neuronal ceroid lipofuscinoses.

ObjectiveNeuronal Ceroid Lipofuscinoses (NCL) are fatal inherited neurodegenerative diseases with established neuronal cell death and increased ceramide levels in brain, hence, a need for disease‐modifying drug candidates, with potential to enhance growth, reduce apoptosis and lower ceramide in neuronal precursor PC12 cells and human NCL cell lines using enhanced flupirtine aromatic carbamate derivatives in vitro.MethodsAromatic carbamate derivatives were tested by establishing growth curves under pro‐apoptotic conditions and activity evaluated by trypan blue and JC‐1 staining, as well as a drop in pro‐apoptotic ceramide in neuronal precursor PC12 cells following siRNA knockdown of the CLN3 gene, and CLN1‐/CLN2‐/CLN3‐/CLN6‐/CLN8 patient‐derived lymphoblasts. Ceramide levels were determined in CLN1‐/CLN2‐/CLN3‐/CLN6‐/CLN8 patient‐derived lymphoblasts before and after treatment. Expression of BCL‐2, ceramide synthesis enzymes (CERS2/CERS6/SMPD1/DEGS2) and Caspases 3/8/9 levels were compared in treated versus untreated CLN3‐deficient PC12 cells by qRT‐PCR.ResultsRetigabine, the benzyl‐derivatized carbamate and an allyl carbamate derivative were neuroprotective in CLN3‐defective PC12 cells and rescued CLN1‐/CLN2‐/CLN3‐/CLN6‐/CLN8 patient‐derived lymphoblasts from diminished growth and accelerated apoptosis. All drugs decreased ceramide in CLN1‐/CLN2‐/CLN3‐/CLN6‐/CLN8 patient‐derived lymphoblasts. Increased BCL‐2 and decreased ceramide synthesis enzyme expression were established in CLN3‐derived PC12 cells treated with the benzyl and allyl carbamate derivatives. They down‐regulated Caspase 3/Caspase 8 expression. Caspase 9 expression was reduced by the benzyl‐derivatized carbamate.InterpretationThese findings establish that compounds analogous to flupirtine demonstrate anti‐apoptotic activity with potential for treatment of NCL disease and use of ceramide as a marker for these diseases.

A proteomics study to explore the role of adsorbed serum proteins for PC12 cell adhesion and growth on chitosan and collagen/chitosan surfaces.

The aim of this article is to apply proteomics in the comparison of the molecular mechanisms of PC12 cell adhesion and growth mediated by the adsorbed serum proteins on the surfaces of chitosan and collagen/chitosan films. First, the chitosan and the collagen/chitosan films were prepared by spin coating; and their surface morphologies were characterized by scanning electron microscopy, X-ray energy dispersive spectroscopy, contact angle measurement and Fourier transform infrared spectroscopy. Subsequently, cell proliferation experiments on two materials were performed and the dynamic curves of protein adsorption on their surfaces were measured. Then, proteomics and bioinformatics were used to analyze and compare the adsorbed serum proteins on the surfaces of two biomaterials; and their effects on cell adhesion were discussed. The results showed that the optimum concentration of chitosan film was 2% w/v. When compared with chitosan film, collagen/chitosan film promoted the growth and proliferation of PC12 cells more significantly. Although the dynamic curves showed no significant difference in the total amount of the adsorbed proteins on both surfaces, proteomics and bioinformatics analyses revealed a difference in protein types: the chitosan surface adsorbed more vitronectin whereas collagen/chitosan surface adsorbed more fibronectin 1 and contained more cell surface receptor binding sites and more Leu-Asp-Val sequences in its surface structure; the collagen/chitosan surface were more conducive to promoting cell adhesion and growth.

Optical Activation of TrkA Signaling.

Nerve growth factor/tropomyosin receptor kinase A (NGF/TrkA) signaling plays a key role in neuronal development, function, survival, and growth. The pathway is implicated in neurodegenerative disorders including Alzheimer's disease, chronic pain, inflammation, and cancer. NGF binds the extracellular domain of TrkA, leading to the activation of the receptor's intracellular kinase domain. As TrkA signaling is highly dynamic, mechanistic studies would benefit from a tool with high spatial and temporal resolution. Here we present the design and evaluation of four strategies for light-inducible activation of TrkA in the absence of NGF. Our strategies involve the light-sensitive protein Arabidopsis cryptochrome 2 and its binding partner CIB1. We demonstrate successful recapitulation of native NGF/TrkA functions by optical induction of plasma membrane recruitment and homo-interaction of the intracellular domain of TrkA. This approach activates PI3K/AKT and Raf/ERK signaling pathways, promotes neurite growth in PC12 cells, and supports survival of dorsal root ganglion neurons in the absence of NGF. This ability to activate TrkA using light bestows high spatial and temporal resolution for investigating NGF/TrkA signaling.

A Single-Step Surface Modification of Electrospun Silica Nanofibers Using a Silica Binding Protein Fused with an RGD Motif for Enhanced PC12 Cell Growth and Differentiation.

In this study, a previously known high-affinity silica binding protein (SB) was genetically engineered to fuse with an integrin-binding peptide (RGD) to create a recombinant protein (SB-RGD). SB-RGD was successfully expressed in Escherichia coli and purified using silica beads through a simple and fast centrifugation method. A further functionality assay showed that SB-RGD bound to the silica surface with an extremely high affinity that required 2 M MgCl2 for elution. Through a single-step incubation, the purified SB-RGD proteins were noncovalently coated onto an electrospun silica nanofiber (SNF) substrate to fabricate the SNF-SB-RGD substrate. SNF-SB-RGD was characterized by a combination of scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, and immunostaining fluorescence microscopy. As PC12 cells were seeded onto the SNF-SB-RGD surface, significantly higher cell viability and longer neurite extensions were observed when compared to those on the control surfaces. These results indicated that SB-RGD could serve as a noncovalent coating biologic to support and promote neuron growth and differentiation on silica-based substrates for neuronal tissue engineering. It also provides proof of concept for the possibility to genetically engineer protein-based signaling molecules to noncovalently modify silica-based substrates as bioinspired material.

Molecular mechanism of panaxydol on promoting axonal growth in PC12 cells.

Nerve growth factor (NGF) promotes axonal growth in PC12 cells primarily by regulating the RTK-RAS-MEK-ERK pathway. Panaxydol, a polyacetylene isolated from Panax notoginseng, can mimic the effects of NGF. Panaxydol promotes neurite outgrowth in PC12 cells, but its molecular mechanism remains unclear. Indeed, although alkynol compounds such as panaxydol can increase intracellular cyclic adenosine 3',5'-monophosphate (cAMP) levels and the ERK inhibitor U0126 inhibits alkynol-induced axonal growth, how pathways downstream of cAMP activate ERK have not been investigated. This study observed the molecular mechanism of panaxydol-, NGF- and forskolin-induced PC12 cell axon growth using specific signaling pathway inhibitors. The results demonstrated that although the RTK inhibitor SU5416 obviously inhibited the growth-promoting effect of NGF, it could not inhibit the promoting effect of panaxydol on axonal growth of PC12 cells. The adenylate cyclase inhibitor SQ22536 and cAMP-dependent protein kinase inhibitor RpcAMPS could suppress the promoting effect of forskolin and panaxydol on axonal growth. The ERK inhibitor U0126 inhibited axonal growth induced by all three factors. However, the PKA inhibitor H89 inhibited the promoting effect of forskolin on axonal growth but could not suppress the promoting effect of panaxydol. A western blot assay was used to determine the effects of stimulating factors and inhibitors on ERK phosphorylation levels. The results revealed that NGF activates the ERK pathway through tyrosine receptors to induce axonal growth of PC12 cells. In contrast, panaxydol and forskolin increased cellular cAMP levels and were inhibited by adenylyl cyclase inhibitors. The protein kinase A inhibitor H89 completely inhibited forskolin-induced axonal outgrowth and ERK phosphorylation, but could not inhibit panaxydol-induced axonal growth and ERK phosphorylation. These results indicated that panaxydol promoted axonal growth of PC12 cells through different pathways downstream of cAMP. Considering that exchange protein directly activated by cAMP 1 (Epac1) plays an important role in mediating cAMP signaling pathways, RNA interference experiments targeting the Epac1 gene were employed. The results verified that Epac1 could mediate the axonal growth signaling pathway induced by panaxydol. These findings suggest that compared with NGF and forskolin, panaxydol elicits axonal growth through the cAMP-Epac1-Rap1-MEK-ERK-CREB pathway, which is independent of PKA.

Electrically conductive nanocomposite hydrogels embedded with functionalized carbon nanotubes for spinal cord injury

Electrically conductive hydrogels incorporated with CNTs support PC12 cell growth and differentiation and hold promise for nerve regeneration.

CAMP-induced activation of protein kinase A and p190B RhoGAP mediates down-regulation of TC10 activity at the plasma membrane and neurite outgrowth.

Cyclic AMP plays a pivotal role in neurite growth. During outgrowth, a trafficking system supplies membrane at growth cones. However, the cAMP-induced signaling leading to the regulation of membrane trafficking remains unknown. TC10 is a Rho family GTPase that is essential for specific types of vesicular trafficking. Recent studies have shown a role of TC10 in neurite growth in NGF-treated PC12 cells. Here, we investigated a mechanical linkage between cAMP and TC10 in neuritogenesis. Plasmalemmal TC10 activity decreased abruptly after cAMP addition in neuronal cells. TC10 was locally inactivated at extending neurite tips in cAMP-treated PC12 cells. TC10 depletion led to a decrease in cAMP-induced neurite outgrowth. Constitutively active TC10 could not rescue this growth reduction, supporting our model for a role of GTP hydrolysis of TC10 in neuritogenesis by accelerating vesicle fusion. The cAMP-induced TC10 inactivation was mediated by PKA. Considering cAMP-induced RhoA inactivation, we found that p190B, but not p190A, mediated inactivation of TC10 and RhoA. Upon cAMP treatment, p190B was recruited to the plasma membrane. STEF depletion and Rac1-N17 expression reduced cAMP-induced TC10 inactivation. Together, the PKA-STEF-Rac1-p190B pathway leading to inactivation of TC10 and RhoA at the plasma membrane plays an important role in cAMP-induced neurite outgrowth.

Investigation of surface topography and stiffness on adhesion and neurites extension of PC12 cells on crosslinked silica aerogel substrates.

Fundamental understanding and characterization of neural response to substrate topography is essential in the development of next generation biomaterials for nerve repair. Aerogels are a new class of materials with great potential as a biomaterial. In this work, we examine the extension of neurites by PC12 cells plated on matrigel-coated and collagen-coated mesoporous aerogel surfaces. We have successfully established the methodology for adhesion and growth of PC12 cells on polyurea crosslinked silica aerogels. Additionally, we have quantified neurite behaviors and compared their response on aerogel substrates with their behavior on tissue culture (TC) plastic, and polydimethylsiloxane (PDMS). We found that, on average, PC12 cells extend longer neurites on crosslinked silica aerogels than on tissue culture plastic, and, that the average number of neurites per cluster is lower on aerogels than on tissue culture plastic. Aerogels are an attractive candidate for future development of smart neural implants and the work presented here creates a platform for future work with this class of materials as a substrate for bioelectronic interfacing.

MiR-181d-5p promotes neurite outgrowth in PC12Cells via PI3K/Akt pathway.

miRNAs dysregulate in spinal cord injury (SCI) and have been demonstrated to play a crucial role in neurite outgrowth. However, the underlying mechanism remains elusive. In this study, we constructed a mouse model of SCI, extracted RNA from injured spinal cord tissue for the use of microarray assay. miR-181d-5p which is one of the most significantly expressed miRNAs in miRNA-mRNA network, abundantly expressed in center system and highly conserved across different spices, was chosen for our further study. To demonstrate whether miR-181d-5p can promote neurite outgrowth in PC12 cells via PI3K/Akt signaling pathway, we performed function analysis of miR-181d-5p with LV-miR-181d-5p and LV-sh-GFP to infect PC12 cells. Through microarray assay analysis, we totally found 262 significantly expressed miRNAs and 2973 target genes in SCI and observed that their expression dynamically changed postinjury. Here, we provided enough evidences that the overexpression of miR181d-5p significantly decreased the expression of PTEN, upregulated p-Akt expression, increased neurite outgrowth-related proteins (GAP-43 and NF-200) and synaptic vesicle-related proteins (Synapsin and PSD95), and then promoted neurite outgrowth in PC12 cells. Furthermore, we confirmed that miR-181d-5p could directly target to the 3'-UTR of PTEN mRNA through dual-luciferase report assay. Our study supports that aberrant expression of miRNAs is involved in the pathogenesis of SCI, miR-181d-5p plays an important role in neurite growth in PC12 cells via PI3K/Akt signaling pathway and may be a candidate target for the treatment of SCI in the future.

Isoliquiritigenin Induces Cytotoxicity in PC-12 Cells In Vitro.

Isoliquiritigenin (ISL) has been reported to have a wide range of biological activities. This study evaluated the cytotoxic effect of ISL on norvegicus pheochromocytoma cell line (PC-12 cells) and its possible molecular mechanism. The cytotoxicity in vitro of ISL against PC-12 cells was investigated by MTT assay. The migration and invasion of PC-12 cells were performed by scratch test and transwell assay. Apoptosis was evaluated by microscopy and flow cytometry. The reactive oxygen species (ROS) and mitochondrial membrane potential were studied by fluorescent microscopy. DNA damage of PC-12 cells was analyzed by comet assay. The protein expression of caspase, Bcl-2 family member, autophagy-associated protein Beclin-1, and LC3 was detected by western blot. The autophagy of PC-12 cells was investigated by acridine orange (AO) and monodansylcadaverine (MDC) staining. The IC50 value of ISL against PC-12 cell is 17.8±1.8μM. ISL could suppress PC-12 cell migration and invasion. AO/ethidium bromide staining and flow cytometry suggested that ISL caused apoptosis of PC-12 cells. Significant DNA damages of PC-12 cells treated with ISL were observed in a comet assay. ISL inhibited the cell growth of PC-12 cells at S phase. Exposure of PC-12 cells to ISL increased the levels of cellular reactive oxygen species (ROS) and decreased the mitochondrial membrane potential. Additionally, ISL trigged the release of cytochrome c from the mitochondria to the cytoplasm. The expression levels of caspase-9, caspase-3, caspase-7, Bax, and Bim were upregulated, whereas the expression levels of Bcl-2 and Bcl-x were downregulated. AO and monodansylcadaverine (MDC) staining assay showed that ISL caused autophagy of PC-12 cells. The upregulation of protein Beclin-1 and LC3 was observed in PC-12 cells. Therefore, the results show that ISL induces apoptosis of PC-12 cells through ROS-mediated activation of the intrinsic mitochondria-cytochrome c-caspase protease mechanism and causes the autophagy of PC-12 cells. Graphical Abstract The in vitro cytotoxicity, apoptosis, comet assay, ROS, mitochondrial membrane potential, cell cycle arrest, autophagy, and western blot induced by ISL were investigated.

Nanostructured Multilayer Films Assembled from Poly(dopamine)‐Coated Carbon Nanotubes for Controlling Cell Behavior

AbstractNano‐topographic surfaces have been used as an effective tool to control cell behavior such as adhesion and proliferation. In this study, multilayer films with nano‐topographic features were fabricated by alternatively assembling poly(l‐lysine) (PLL) and poly(dopamine)‐coated carbon nanotubes (CNTs@PDA) layers. The growth of PLL/CNTs@PDA film presented a perfect linear relationship with the number of bilayers. A nanostructured morphology with interpenetrating CNT networks was observed by scanning electron microscopy (SEM). Adhesion and proliferation of endothelial cells (ECs) and smooth muscle cells (SMCs) on the PLL/CNTs@PDA multilayer films have been evaluated. The films support initial adhesion of both ECs and SMCs. Interestingly, the PLL/CNTs@PDA multilayer films were found to promote proliferation of SMCs and inhibited proliferation of ECs. Further, pheochromocytoma (PC12) cells were employed to evaluate the influence of PLL/CNTs@PDA multilayer films on the outgrowth of synapses. We found that the nanostructured surface significantly promoted the synapses of PC12 cell growth and formation. Our findings suggest that cytophilic surfaces with the nanostructured morphology have diverse effects on different cells, which sheds light on new design of biomaterial surfaces in cell‐based applications.

Cdk5 is required for the neuroprotective effect of transforming growth factor-β1 against cerebral ischemia-reperfusion

Transforming Growth Factor β1 (TGF-β1), a well-known neuroprotective and neurotrophic factor in the central nervous system, is also involved in the repair process responses after ischemia-reperfusion injury. Herein, we found that TGF-β1 enhanced Cdk5 expression while decreased Tunel-positive cells compared with the ischemia group, and roscovitine(Cdk5 inhibitor) treatment could blunt these effects. In vitro study, TGF-β1 facilitated Cdk5/p35 complex, the proliferation, neurite growth and differentiation of PC12 cells, effects of which could be blunted by roscovitine and Cdk5 silencing. Moreover, ERK1/2 inhibitor SCH772984 abrogated the effects of TGF- β1 on Cdk5 and Bax levels. Taken together, we conclude that Cdk5 contributes to the neuroprotective function of TGF- β1 via ERK1/2 signaling.