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Background Scaffolds containing two kinds of regular pore size that support differentiation and proliferation of induced pluripotent stem cells (iPSC) are important approaches to artificial pancreas. Methods This study aimed to differentiate iPSC in dual-sized gelatin (Gel)-alginate (Alg)-hyaluronic acid (HA) scaffolds containing inverted colloidal crystal (ICC) topography with 70-µm and 160-µm pore sizes for insulin production. The pore surface of dual-sized Gel-Alg-HA ICC scaffolds was grafted with growth differentiation factor 8 (GDF-8) and Wnt3a, and treated with retinoic acid (RA) and noggin to improve the differentiation of iPSC toward endodermic cells, and then islet cells. Findings An increasing weight percentage of Gel increased the adhesion efficiency of iPSC. Moreover, Alg and HA helped to promote the proliferation and survival rate of iPSC, which resulted in a higher availability of iPSC for differentiation into insulin-production cells. Immunofluorescence staining, flow cytometry and western blot evidenced that dual-sized Gel-Alg-HA ICC scaffolds grafted with GDF-8 and Wnt3a enhanced the differentiation of iPSC into endodermic cells. When subsequent treatment with RA and noggin, endodermic cells differentiation into insulin-production cells in the scaffolds were improved. Precisely controlling the physical and biomedical properties of dual-sized Gel-Alg-HA ICC scaffolds can guide iPSC to differentiate first into endodermic cells, and then into pancreatic β-cells for diabetic treatment.

Recent advances in attempting maturation of pancreatic β cells derived from induced pluripotent stem cells (iPSCs) have initiated a gateway to developing innovated materials and protocols for artificial pancreas implantation in diabetic management. In this study, we proposed a tissue-engineering scheme using iPSCs-laden aldehyde hyaluronic acid (AHA)-polyacrylamide (PAAM) inverted colloidal crystal constructs (ICCCs) associated with controlled release of induction molecules to generate insulin-producing cells (IPCs). Based on physicochemical properties, AHA-PAAM inverted colloidal crystal scaffolds (ICCSs) at a 1:1 weight ratio of AHA to PAAM were used, modified with growth and differentiation factor 8, and grafted with CHIR99021-loaded solid lipid nanoparticles (CHIR99021-SLNs-GDF8-AHA-PAAM ICCSs) to differentiate iPSCs into definitive endoderm (DE) cells. Induction with fibroblast growth factor 7-grafted retinoic acid-loaded SLNs (FGF7-RA-SLNs) further differentiated DE cells into insulin-producing cells (IPCs) in the constructs. An introduction of sustained delivery facilitated bioavailability of CHIR99021 and RA during DE and islet formation, and yielded a high quantity of secreted insulin from IPCs after stimulation of glucose. Loaded CHIR99021 and grafted FGF7 acted as supporters to promote the main function of grafted GDF8 and loaded RA in DE and islet growth. From the highest expression of pancreatic and duodenal homeobox 1, we have reached the appropriate concentration of GDF8, CHIR99021, RA and FGF7 for regenerating IPCs. These data showed that the 2-stage guided differentiation of iPSCs into IPCs in CHIR99021-SLNs-GDF8-AHA-PAAM ICCSs can be promising in ameliorating insulin secretion by β-cell regeneration.

Management of glioblastoma multiforme (GBM), a cerebral glioma with highest-grade malignancy, is a crucial challenge to current anticancer treatment because the lack of GBM-targeting capability of antitumor drug, such as carmustine (BCNU) with a short half-life, along with low blood-brain barrier (BBB) permeability to achieve the clinical efficacy. Hence, polyethyleneimine-poly(lactic-co-glycolic acid) nanoparticles comprising stable core-shell structure with tamoxifen were crosslinked with sialic acid (SA-TAM-PEI-PLGA NPs) as nanocarriers to improve bioavailability of BCNU for pharmacotherapy. The brain-targeted experiments revealed that an increasing SA level improved BBB permeability for BCNU, and was associated with a reduction in transendothelial electrical resistance and an increase in propidium iodide penetration of endothelial monolayer. Immunocytochemical staining images evidenced that surface SA was critical in triggering transcytosis through N-acetylglucosamine on human cerebral microvessel endothelia, and surface crosslinking restrained macrophage phagocytosis. A high TAM level on the carrier surface reduced the survival rate of human U87, and western blots showed that conjugated TAM induced apoptosis of the cancer cells. The use of SA-TAM-PEI-PLGA NPs carrying BCNU as a pharmaceutical preparation is an effective strategy to overcome the BBB restriction and activate apoptosis of tumor cells to prevent the proliferation of GBM.

Quercetin (QU) and rosmarinic acid (RA) were loaded in phosphatidic acid-liposomes (QU/RA-PA-liposomes) with surface apolipoprotein E (ApoE) using a process of thin-film hydration, followed by covalent crosslinking to activate biological pathways for penetrating the blood–brain barrier (BBB) and redeeming the neuronal apoptosis from attack of β-amyloid 1-42 (Aβ1-42) and neurofibrillary tangles. The conjugation of liposomes with PA improved the activity of QU and RA against neurotoxicity of Aβ1-42. The fluorescent images of brain capillaries revealed that surface modification with ApoE improved the permeation ability of QU/RA-PA-ApoE-liposomes across the BBB. In addition, the highest therapeutic efficacy was obtained in the case of QU/RA-PA-ApoE-liposomes, compared to other QU/RA formulations studied using in vivo Aβ1-42-insulted rats mimicking Alzheimer’s disease (AD). The cellular and molecular evidence from AD rats included the decrease in Aβ1-42 plaque formation and interleukin-6 secretion, increase in the neuronal count in Nissl staining, and reduction in the expression of phosphorylated extracellular signal-regulated kinase 1/2, c-Jun N-terminal kinase, p38 kinase and tau protein at serine 202 as well as caspase-3. The use of PA-ApoE-liposomes as a dual targeting formulation enhances the QU and RA ability to infiltrate the BBB, docks Aβ1-42 plaques and can be a potent approach to rescue degenerated neurons from AD.

Internal aqueous phase was incorporated in solid lipid nanoparticles (SLNs) containing 5 lipid components to form dual-phase SLNs (DPSLNs), which were then conjugated with transferrin (Tf) to transport nerve growth factor (NGF), rosmarinic acid (ROA), curcumin (CURC) and quercetin (QU) for enhanced activity against neuronal apoptosis. Particle size, physical stability, loading rate of drugs and releasing rate of drugs were measured to determine the most appropriate lipid composition of DPSLNs. The results showed that DPSLNs not only acted as a drug carrier, but also served to decrease drug-induced cytotoxicity to blood-brain barrier (BBB) cells through sustained release. QU-CURC-ROA-NGF-DPSLNs with modified Tf on the surface facilitated the capacity of drugs to infiltrate the BBB. Interaction between lecithin and Tf assisted in BBB-targeting ability of DPSLNs. Moreover, addition of cardiolipin in DPSLNs improved the therapeutic efficacy via recognizing β-amyloid around neurons. Combining QU, CURC, ROA and NGF in DPSLNs promoted their roles in the reduction of neurotoxicity by mutual collaboration. Immunofluorescence and western-blot analysis revealed suppressed expressions of caspase-3, extracellular signal-regulated kinases 1/2 (ERK1/2), c-Jun N terminal kinase and p38, and enhanced expressions of cAMP-response element-binding protein (CREB) and ERK5 after treatment with QU-CURC-ROA-NGF-DPSLNs. Hence, the drug-loaded DPSLNs with grafted Tf can be promising in rescuing degenerated neurons for Alzheimer's disease therapy.

Advances in liposomal formulation carrying multiple neuroprotective drugs, such as ceftriaxone (CEF), FK506, and nilotinib, can point toward an approach to obviating the difficulties in Parkinson's disease (PD) treatment. We prepared functionalized liposomes decorated with glutathione (GSH) to penetrate the blood-brain barrier (BBB) and cardiolipin (CL) to link up apoptotic neurons. Further, the effect of CEF-FK506-nilotinib-GSH-CL-liposomes on a PD model established by SH-SY5Y cells with 1-methyl-4-phenylpyridinium-induced neurotoxicity was investigated. An increment of the mole percentage of dihexadecyl phosphate and CL increased the particle size and the absolute value of ζ potential, improved the entrapment efficiency of CEF, FK506, and nilotinib, and reduced the drug-releasing rate. The toxicity studies revealed that CEF, FK506, and nilotinib-encapsulated liposomes could enhance the survival of SH-SY5Y cells. Western blot and immunofluorescence revealed that incorporation of CL in a lipid bilayer ameliorated the docking of CEF-FK506-nilotinib-GSH-CL-liposomes at α-synuclein (α-syn), indicating a better targeting capability of the liposomes to degenerated neurons. Treatment with CEF-FK506-nilotinib-GSH-CL-liposomes reduced the expression of Bax and α-syn and promoted the expression of Bcl-2, tyrosine hydroxylase, and the dopamine transporter. GSH- and CL-conjugated liposomes showed combined activity of targeting the BBB and α-syn and augmented the efficiency of the three drugs in rescuing dopaminergic neurons for neurodegenerative therapy.

A three-dimensional scaffold enables the formation of realistic tissue architecture and promotes physiological responses to activate the differentiation of induced pluripotent cells (iPSCs) into pancreatic islets. Poly(vinyl alcohol) (PVA), alginate (Alg) and gelatin (Gel) were conjugated with methacrylic anhydride (MA) and were photo-crosslinked to prepare PVAMA-AlgMA-GelMA hydrogel scaffolds using a water-in-oil self-assembly method. The swelling ratio, porosity and cell entrapment in the hydrogel were investigated with various molar percentages of polymer matrix. PVAMA and AlgMA assisted in improving the water content of the hydrogel, which created high porosity for the migration of iPSCs, and GelMA enhanced the cell entrapment efficiency. The self-assembled scaffolds with composition of PVAMA:AlgMA:GelMA = 1:1:2 were grafted with activin A and bone morphogenic protein 4 (BMP4) to induce iPSCs to definitive endoderm (DE) lineage, followed by treatment with retinoic acid-loaded solid lipid nanoparticles for controlled release and guided differentiation of DE cells into insulin-producing cells. Flow cytometry analysis and immunochemical staining evidenced that the scaffolds with crosslinked BMP4 boosted the role of surface activin A at activin A:BMP4 = 3:1 in endodermal differentiation of iPSCs to produce the highest level of insulin after pancreatic induction and glucose stimulation. The iPSC-laden PVAMA-AlgMA-GelMA hydrogel constructs composed of repeated units and modified with activin A and BMP4 can be promising in generating pancreas-mimetic structure and supporting the functions of pancreatic islets.

The capillary electrophoresis (CE) technique has been widely used to improve our understanding of biochemistry, and has become one of the best choices to address critical issues in neurobiology. In this study, doubled-emulsified solid lipid nanoparticles (DESLNs) grafted with PPFLMLLKGSTR (PM12) were fabricated to carry retinoic acid (RnA) and nerve growth factor (NGF), and to differentiate induced pluripotent stem cells (iPSCs) into nerve cells. The membrane charge of differentiated neuron-like cells was investigated using a high-performance CE. We found that surface PM12 enhanced the uptake of RnA- and NGF-loaded DESLNs (RnA-NGF-DESLNs) via strong interaction with cellular integrin α3β1 and α5β1, as evidenced by immunofluorescence assay. Further, highly expressed βIII-tubulin, a typical neuronal marker, suggested the improved neurite outgrowth of differentiating iPSCs. The enhanced electrophoretic mobility also demonstrated the reproduction of neurons from iPSCs treated with various kinds of DESLNs. After longer induction, the major CE elution peak appeared in shorter time, evidencing an increase in electrostatic potential (more negative) and effective neuronal regeneration. Induction with PM12-RnA-NGF-DESLNs to differentiate iPSCs resulted in the largest electrophoretic mobility (more negative), zeta potential and membrane charge. Thus, this study showed the ability of CE to analyze the charge variation during guided differentiation of iPSCs with drug carriers, and the CE method can be promising in evaluating the formation of matured neurons from iPSCs.

Liposomes (lip) carrying pharmaceuticals have shown promise in their ability to advance the therapy for neurodegenerative diseases. However, the low nerve-targeting capacity and poor penetration rate of lip through the blood-brain barrier (BBB) are major hurdles to achieving successful treatment. Herein, we developed lip incorporating cardiolipin (CL) and phosphatidic acid (PA) to promote their capability against hyperphosphorylation of tau protein, and a transactivator of transcription (TAT) peptide to permeate the BBB for delivering nerve growth factor (NGF), rosmarinic acid (RA), curcumin (CURC) and quercetin (QU). We derived an optimization method to assess a better composition of phospholipids in the lip loaded with the four medicines. Experimental results revealed that this optimized lip increased the viability of SK-N-MC cells insulted with β-amyloid peptide (Aβ) fibrils and prevented Wistar rat brain from producing hyperphosphorylated tau. CL and PA and the grafted TAT peptide on the carrier surface improved the rescue efficiency by inhibiting Aβ deposition and reducing the expressions of phosphorylated extracellular signal-regulated protein kinase 1/2 (p-ERK1/2), c-Jun N-terminal protein kinase, p38, tau at serine 202 and caspase-3. The lip also enhanced the expressions of p-ERK5 and p-cyclic adenosine monophosphate response element-binding protein. The amalgamated activity of NGF, RA, CURC and QU, and the effect of charged CL/PA on Aβ deposits supported the therapeutic efficacy of lip. The optimized TAT-NGF-RA-CURC-QU-CL/PA-lip can be a capable drug delivery system to cross the BBB and protect Alzheimer's disease brains from tau hyperphosphorylation. STATEMENTS OF SIGNIFICANCE: The therapeutic efficiency of liposomes (lip) against neurodegenerative disorder depends on their nerve-targeting capacity and ability to permeate the blood-brain barrier (BBB). Lip was developed incorporating cardiolipin (CL) and phosphatidic acid (PA) to promote their target specificity against hyperphosphorylation of tau protein, and a transactivator of transcription (TAT) peptide to permeate the BBB. We have successfully derived an optimization method using a new mathematical expression for the first time to assess a better composition of phospholipids in lip loaded with nerve growth factor (NGF), rosmarinic acid (RA), curcumin (CURC) and quercetin (QU). The optimized TAT-NGF-RA-CURC-QU-CL/PA-lip efficaciously down-regulated the expressions of phosphorylated extracellular signal-regulated protein kinase 1/2 (p-ERK1/2), c-Jun N-terminal protein kinase, p38, tau at serine 202 and caspase-3, and up-regulated the expressions of p-ERK5 and p-cyclic adenosine monophosphate response element-binding protein in Alzheimer's disease Wistar rat model.

Significant involvement of oxidative stress in the brain can develop Alzheimer's disease (AD); however, a great number of clinical trials explains the limited success of antioxidant therapy in dealing with this neurodegenerative disease. Here, we established a lipopolymer system of poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) incorporated with phosphatidic acid (PA) and modified with sialic acid (SA) and 5-hydroxytryptamine-moduline (5HTM) to improve quercetin (QU) activity against oxidative stress induced by amyloid-β (Aβ) deposits. Morphological studies revealed a uniform exterior of QU-SA-5HTM-PA-PLGA NPs with a spherical structure and enhanced aggregation with inclusion of PA in the formulation. A better brain-targeted delivery of the lipopolymeric NPs was verified from the high blood-brain barrier (BBB) permeability of QU through strong interactions of surface SA and 5HTM with O-linked N-acetylglucosamine and 5-HT1B receptors, respectively. Immunofluorescence staining images also supported QU-SA-5HTM-PA-PLGA NPs to traverse the microvessels of AD rat brain. Western blot analysis showed that QU-loaded PA-PLGA NPs suppressed caspase-3 expression. The ability of the nanocarriers to recognize Aβ fibrils was demonstrated from the reduced senile plaque formation and the attenuated acetylcholinesterase and malondialdehyde activity in the hippocampus. Hence, the medication of QU-SA-5HTM-PA-PLGA NPs can facilitate the BBB penetration and prevent Aβ accumulation, lipid peroxidation, and neuronal apoptosis for the AD management.

Polymers with very low surface conductivity can easily accumulate an electric charge through surface friction. If the charged entities are grounded with conductive materials, they may discharge electricity associated with sparks that will interrupt the production process, and more seriously, cause industrial disasters. In order to avoid damage from electrical sparks, an antistatic agent (ASA) is employed with the polymer during production. In this study, conductive iron oxide (Fe3O4) nanoparticles (NPs) with hydrophobic modification by oleic acid (OA) were prepared using a co-precipitation method. Fourier transformed infrared spectra and thermogravimetric analysis confirmed that OA-Fe3O4 was formed through chemisorption interactions between OA and Fe3O4. Fe3O4 NPs were entrapped in solid lipid NPs (SLNs) and poly(lactide-co-glycolide) NPs through hydrophobic interactions to obtain new composite antistatic NPs (CANs). Surfactants and their concentration played a decisive role in the zeta potential and particles size of CANs. Sodium dodecyl sulfate (SDS) created some regular-shaped CANs, and didodecyldimethylammonium bromide (DMAB) and glycerol monostearate (GMS) created irregular-shaped CANs. 10% methanol and 1% Tween 80 yielded an appropriate contact angle with polymer films. The surface electrical resistance value of CANs was found to be in the order of SDS > DMAB > GMS. GMS-based CANs reached a lower electrical resistance value of around 108 Ω/Sq, and can be used as an ASA in engineering practice. Keywords: Composite antistatic nanoparticle, Polymer film, Surface electrical resistance, Sodium dodecyl sulfate, Didodecyldimethylammonium bromide, Glycerol monostearate

Neurodegenerative disorders, such as Alzheimer's disease (AD), present biomedical challenges due to inability of pharmaceuticals to permeate the blood-brain barrier (BBB) and lack of therapeutic specificity in definite sites against multiple pathologies. Phosphatidylcholine (PC)-liposomes carrying curcumin (CURC), quercetin (QU), epigallocatechin gallate (EGCG) and rosmarinic acid (RA) with crosslinked glutathione (GSH) and apolipoprotein E (ApoE) were fabricated to recognize brain microvascular endothelial cells and amyloid beta (Aβ), and reduce tau protein hyperphosphorylation for AD management. Addition of stearic acid to liposomal bilayers ameliorated particle stability, promoted drug entrapment efficiency, and prolonged drug release duration. The triple targeting liposomes boosted the capability of CURC, QU, EGCG and RA for crossing the BBB with the assistance of grafted GSH and ApoE and docking Aβ around SK-N-MC cells using ApoE and PC. Moreover, GSH-ApoE-PC-liposomes benefited the 4 medicines in simultaneously transporting to Aβ1-42-insulted neurons, in functioning against hyperphosphorylated mitogen-activated protein kinases, including p-c-Jun N-terminal protein kinase, p-extracellular signal-regulated protein kinase 1/2 and p-p38, in downregulating tau protein at S202, caspase-3 and interleukin-6, and in upregulating p-cyclic adenosine monophosphate response element-binding protein. GSH-ApoE-PC-liposomes can be promising colloidal carriers in delivering CURC, QU, EGCG and RA to degenerated neural tissue in a controlled manner, targeting pathological factors for neuroprotection, and raising preclinical effectualness for AD treatment.

Background The difficulty of drugs to cross the blood-brain barrier (BBB) to reach specific sites and target multiple factors responsible for neurovegetative diseases necessitates the development of an effective drug delivery system. Methods Rabies virus glycoprotein (RVG)- and transferrin (Tf)-grafted liposomes were constructed to deliver epigallocatechin gallate (EGCG) and FK506 to BBB against neurodegeneration for Parkinson's disease (PD) treatment. Significant findings Surface RVG and Tf revealed substantial improvement in BBB permeability of EGCG and FK506, and incorporation of phosphatidy-l-serine (PS) in RVG-Tf-EGCG-FK506-liposomes ameliorated α-synuclein docking. EGCG and FK506-encapsulated liposomes decreased cytotoxicity to 1-methyl-4-phenylpyridinium-impacted neurons. Immunofluorescence and western blot studies demonstrated better activity of RVG-Tf-EGCG-FK506-PS-liposomes in suppressing Bax, α-synuclein, caspase-3, p-tau protein, p-p38, p-ERK1/2 and p-cJNK expressions, and in increasing Bcl-2, tyrosine hydroxylase, dopamine transporter, p-CREB and p-ERK5 expressions. Multi-targeted RVG-Tf-PS-liposomes advantaged BBB permeation and contributed to neuroprotective effect to promote the efficiency of EGCG and FK506 in PD management.

Pluronic® F127-modified water-dispersible poly(acrylic acid)-bound iron oxide (PF127-PAAIO) nanoparticles have been prepared as diagnostic agents. A blood–brain-barrier penetrating peptide, angiopep-2 (ANG), was further conjugated onto the surface of the PF127-PAAIO (ANG-PF127-PAAIO) for brain targeting. The ANG-PF127-PAAIO shows negligible cell cytotoxicity, better cellular uptake, and higher T2-weighted image enhancement than the PF127-PAAIO in U87 cells. Using an ex vivo blood–brain barrier (BBB) model, we showed that the ANG-PF127-PAAIO shows better permeability to bypass the BBB. This is because the ANG-PF127-PAAIO has a dual-targeting ability, recognition of the low-density lipoprotein receptor-related protein and clathrin-mediated receptor on the U87 surface. Thus, the ANG-PF127-PAAIO is a potential nanotheranostic agent for brain dysfunction.

Effective treatment methods for neurodegenerative disorders in the central nervous system are still facing a great many hurdles due to inability of drugs to permeate the blood–brain barrier (BBB). In this study, a drug carrier system of apolipoprotein E (ApoE)-modified liposomes conjugated with phosphatidic acid (PA) was fabricated to improve BBB penetration and release quercetin (QU) and rosmarinic acid (RA) to inhibit β-amyloid (1–42) (Aβ1-42)-induced Alzheimer's disease (AD). An in vitro BBB model validated the enhanced ability of the dual-functioning ApoE-QU-RA-PA-liposomes to infiltrate the BBB and preserve high viability of endothelia, compared with Tween 80-QU-RA-PA-liposomes. Immunochemical staining images exhibited evidence that ApoE-QU-RA-PA-liposomes were able to penetrate the BBB through the aid of strong attractions between ApoE and low-density lipoprotein receptors in brain microvascular endothelial cells, and also inhibited the apoptosis of Aβ1-42-insulted SK-N-MC cells. In anin vivo AD model, ApoE-QU-RA-PA-liposomes decreased acetylcholinesterase activity and lipid peroxidation level, and lowered Aβ plaque formation. The surface functionalization of liposomes using PA and ApoE can be a promising approach to crossing the BBB and delivering QU and RA to the brain in management of Aβ1-42-induced neurotoxicity.

An in vitro human blood–brain barrier (BBB) model was developed using human brain microvascular endothelial cells (HBMECs) co-cultured with human brain vascular pericytes (HBVPs) and human astrocytes (HAs) at various ratios (HBVPs:HAs = 1:1, 1:2, 1:6), associated with pericyte-conditioned medium (PCM) and astrocyte-conditioned medium (ACM). After 7 days of co-culturing of HBMECs with HBVPs and HAs at a ratio of 1:2, and a 1:1 ratio of PCM and ACM, the transendothelial electrical resistance was enhanced to 319 ± 16.67 Ω × cm2 (225% increase) and the permeability coefficient of propidium iodide was reduced to 2.09 ± 0.5 × 10−6 cm/s (61% decrease). Analysis of three major barrier integrity modulators: transforming growth factor-β1 (TGF-β1), vascular endothelial growth factor (VEGF), and matrix metalloproteinases (MMPs), revealed a higher TGF-β1 expression and lower VEGF and MMP-9 expressions in a co-culture of HBMECs with HBVPs and HAs at a ratio of 1:2. Calcein-AM analysis showed higher P-glycoprotein (P-gp) activity in the model using PCM and ACM with HBVPs:HAs = 1:2 (100%) than that with HBVPs:HAs = 1:1 (55%) and HBVPs:HAs = 1:6 (49%). The effect of TGF-β1 receptor inhibitor SB431542, VEGF inhibitor asterric acid, and MMP-9 inhibitor CTT (H-Cys-Thr-Thr-His-Trp-Gly-Phe-Thr-Leu-Cys-OH) on the BBB model suggested that TGF-β1 up-regulates P-gp activity and VEGF down-regulates P-gp activity. In the presence of PCM and ACM, co-culturing of HBMECs with HBVPs and HAs at a ratio of 1:2 could approach the in vivo BBB in a well representative manner.

Heap-up of α-synuclein (α-Syn) and its association with tau protein are esteemed to trigger the onset of Parkinson's disease (PD). The purpose of this study was to develop multi-functional liposomes incorporated with 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), cholesterol, 1,2-dimyristoyl-sn-glycero-3-phosphocholine and phosphatidylserine (PS) to load astragaloside IV (AS-IV) and nestifin-1 (NF-1), followed by grafting with wheat germ agglutinin (WGA) and leptin (Lep) (WGA-Lep-AS-IV-NF-1-PS-liposomes) to protect dopaminergic neurons from apoptosis. Experimental results showed that increasing the mole percentage of DSPC and PS enhanced the particle size, particle stability and entrapment efficiency of AS-IV and NF-1, and reduced the drug releasing rate. Strong affinity of NF-1 to PS was evidenced by nuclear magnetic resonance spectroscopy. WGA-Lep-AS-IV-NF-1-PS-liposomes diminished transendothelial electrical resistance and improved the capacity of propidium iodide, AS-IV and NF-1 to penetrate the blood-brain barrier (BBB). Immunocytochemical staining exhibited the ability of functionalized liposomes to target Lep receptor and α-Syn in MPP+-insulted SH-SY5Y cells. Western blots revealed a substantial reduction of α-Syn and phosphorylated tau protein in the anti-oxidative pathway through interaction with PS. During the course of treatment with WGA-Lep-AS-IV-NF-1-PS-liposomes, the combined activity of AS-IV and NF-1 and recognition capability simultaneously decreased the expression of Bax, and increased the expressions of Bcl-2, tyrosine hydroxylase and dopamine transporter. The liposomes carrying AS-IV and NF-1 can rescue degenerated neurons and are a promising formulation to achieve better PD management.

Rosmarinic acid-loaded polyacrylamide-chitosan-poly(lactide-co-glycolide) nanoparticles (RA-PAAM-CH-PLGA NPs) were grafted with cross-reacting material 197 (CRM197) and apolipoprotein E (ApoE) for targeting of the blood–brain barrier (BBB) and rescuing degenerated neurons. The polymeric nanocarriers were prepared by microemulsion, solvent diffusion, grafting, and surface modification, and CRM197-ApoE-RA-PAAM-CH-PLGA NPs were used to treat human brain-microvascular endothelial cells, RWA264.7 cells, and Aβ-insulted SK-N-MC cells. Experimental results revealed that an increase in the weight percentage of PAAM decreased the particle size, zeta potential, and grafting efficiency of CRM197 and ApoE. In addition, surface DSPE-PEG(2000) could protect CRM197-ApoE-RA-PAAM-CH-PLGA NPs against uptake by RWA264.7 cells. An increase in the concentration of CRM197 and ApoE decreased the transendothelial electrical resistance and increased the ability of propidium iodide and RA to cross the BBB. The order in the viability of apoptotic SK-N-MC cells was CRM197-ApoE-RA-PAAM-CH-PLGA NPs > CRM197-RA-PAAM-CH-PLGA NPs > RA. Thus, CRM197-ApoE-RA-PAAM-CH-PLGA NPs can be a promising formulation to deliver RA to Aβ-insulted neurons in the pharmacotherapy of Alzheimer’s disease.

Nerve growth factor (NGF)-loaded heparinized cationic solid lipid nanoparticles (NGF-loaded HCSLNs) were developed using heparin-stearic acid conjugate, cacao butter, cholesterol, stearylamine (SA), and esterquat 1 (EQ 1). The effect of cationic lipids and lipid matrix composition on the particle size, particle structure, surface molecular composition, chemical structure, electrophoretic mobility, and zeta potential of HCSLNs was investigated. The effect of HCSLNs on the membrane charge of induced pluripotent stem cells (iPSCs) was also studied. The results indicated that the average diameter of HCSLNs was 90–240 nm and the particle size of HCSLNs with EQ 1 was smaller than that with SA. The zeta potential and electrophoresis analysis showed that HCSLNs with SA had a positively charged potential and HCSLNs with EQ 1 had a negatively charged potential at pH 7.4. The high-resolution transmission electron microscope confirmed the loading of NGF on the surface of HCSLNs. Differentiation of iPSCs using NGF-loaded HCSLNs with EQ 1 exhibited higher absolute values of the electrophoretic mobility and zeta potential than differentiation using NGF-loaded HCSLNs with SA. The immunochemical staining of neuronal nuclei revealed that NGF-loaded HCSLNs can be used for differentiation of iPSCs into neurons. NGF-loaded HCSLNs with EQ 1 had higher viability of iPSCs than NGF-loaded HCSLNs with SA. NGF-loaded HCSLNs with EQ 1 may be promising formulation to regulate the membrane charge of iPSCs during neuronal differentiation.

Catanionic solid lipid nanoparticles (CASLNs) with surface wheat germ agglutinin (WGA) and lactoferrin (Lf) were formulated to entrap and release etoposide (ETP), cross the blood–brain barrier (BBB), and inhibit glioblastoma multiforme (GBM) growth. Microemulsified ETP-CASLNs were modified with WGA and Lf to permeate a cultured monolayer of human brain-microvascular endothelial cells (HBMECs) regulated by human astrocytes and to treat malignant U87MG cells. Experimental evidence revealed that an increase in the weight percentage of ETP from 1% to 4% decreased its encapsulation efficiency about 34-44%. In addition, the release rate of ETP from WGA-Lf-ETP-CASLNs decreased with an increase in the concentration of catanionic surfactant from 7.5 µM to 12.5 µM, and WGA-Lf-ETP-CASLNs at 12.5 µM of catanionic surfactant exhibited a feature of sustained release. WGA-Lf-ETP-CASLNs also reduced transendothelial electrical resistance from 245.5 Ω×cm 2 to 191.5 Ω×cm 2 , enhanced the permeability of propidium iodide from 3.62×10 −6 cm/s to 5.61×10 −6 cm/s, induced a minor cytotoxicity to HBMECs, increased the ability of ETP to cross the BBB by about 5.6 times, and improved the antiproliferative efficacy of U87MG cells. The grafting of WGA and Lf is crucial to control the medicinal property of ETP-CASLNs, and WGA-Lf-ETP-CASLNs can be promising colloidal carriers in GBM management.

Self-assembled microgel functionalized with peptides was developed and applied to regenerate neurons from induced pluripotent stem cells (iPSCs). Collagen (COL), hyaluronic acid (HA), and alginate (ALG) were modified with methacrylic anhydride (MA), photocrosslinked for patterned particles, grafted with GRGDSP and Ln5-P4, and self-assembled to integrate the microgel into three-dimensional scaffolds. Physicochemical assessments revealed that the ternary microgel scaffolds had an optimal chemical composition at COLMA:HAMA:ALGMA=1:2:1. In fabricating cell-laden constructs, modified GRGDSP/Ln5-P4 in linear self-assembled scaffolds could significantly improve the entrapment efficiency and viability of iPSCs. In addition, GRGDSP/Ln5-P4 in the microgel constructs triggered the differentiation of iPSCs toward neurons, since the percentage of neurite-like cells could be higher than 98% after induction of nerve growth factor. Self-assembled microgel comprising COLMA, HAMA, ALGMA, and GRGDSP/Ln5-P4 may be promising in producing mature neural lineage from iPSCs, to provide better treatment for damaged nervous tissue.

Polyacrylamide (PAAM) and chitosan were fabricated by inverted colloidal crystal (ICC) method for scaffolds comprising regular pores. The hybrid PAAM-chitosan ICC scaffolds were grafted with poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) for a rougher pore surface and grafted with transactivator of transcription von Hippel-Lindau (TATVHL) peptide for a better differentiation of induced pluripotent stem (iPS) cells toward neural lineage. By scanning electron microscopy, we found that iPS cells cultured in PAAM-chitosan ICC scaffolds with PLGA NPs at 1.0 mg/mL and TATVHL peptide at 15 μg/mL elongated the axonal length to 15 μm. A combination of PLGA NPs and TATVHL peptide favored the adhesion of iPS cells, reduced the embryonic phenotype after cultivation, and guided the production of βIII tubulin-positive cells in PAAM-chitosan ICC scaffolds. In addition to the differentiation toward neurite-like cells, an increase in the content of TATVHL peptide in PAAM-chitosan ICC scaffolds inhibited the differentiation of iPS cells toward astrocytes. ICC scaffolds composed of PAAM, chitosan, PLGA NPs, and TATVHL peptide can be an efficacious matrix to differentiate iPS cells toward neurons and retard the glial formation for nerve regeneration.

Yung-Chih Kuo,1 Che-Yu Lin,1 Jay-Shake Li,2 Yung-I Lou3 1Department of Chemical Engineering, 2Department of Psychology, National Chung Cheng University, Chia-Yi, 3Department of Accounting, Providence University, Taichung, Taiwan, Republic of China Abstract: Curcumin (CRM) and nerve growth factor (NGF) were entrapped in liposomes (LIP) with surface wheat germ agglutinin (WGA) to downregulate the phosphorylation of kinases in Alzheimer’s disease (AD) therapy. Cardiolipin (CL)-conjugated LIP carrying CRM (CRM-CL/LIP) and also carrying NGF (NGF-CL/LIP) were used with AD models of SK-N-MC cells and Wistar rats after an insult with β-amyloid peptide (Aβ). We found that CRM-CL/LIP inhibited the expression of phosphorylated p38 (p-p38), phosphorylated c-Jun N-terminal kinase (p-JNK), and p-tau protein at serine202 and prevented neurodegeneration of SK-N-MC cells. In addition, NGF-CL/LIP could enhance the quantities of p-neurotrophic tyrosine kinase receptor type 1 and p-extracellular signal-regulated kinase 5 for neuronal rescue. Moreover, WGA-grafted CRM-CL/LIP and WGA-grafted NGF-CL/LIP significantly improved the permeation of CRM and NGF across the blood–brain barrier, reduced Aβ plaque deposition and the malondialdehyde level, and increased the percentage of normal neurons and cholinergic activity in the hippocampus of AD rats. Based on the marker expressions and in vivo evidence, current LIP carriers can be promising drug delivery systems to protect nervous tissue against Aβ-induced apoptosis in the brain during the clinical management of AD. Keywords: liposome, Alzheimer’s disease, β-amyloid, neurodegeneration, blood–brain barrier, wheat germ agglutinin

Of late, cancer has become a terrible disease affecting people throughout the world. Keeping this in mind, we tried to design drugs that are more lipophilic, target-specific, water-soluble, cytoselective and fluorescent. In this regard, we reported novel ruthenium(II)-p-cymene imidazophenanthroline scaffolds as effective DNA targeting agents. The planarity of imidazophenanthroline ligands caused the Ru(II) complex to be a good intercalator. An extended π-electronic conjugation was introduced in the imidazophenanthroline moieties through the Suzuki and Sonogashira coupling reactions. Here, we synthesized nine Ru(II) complexes (16a–b, 17a–d, and 19a–c). Among these, [(η6-p-cymene)RuCl(K2-N,N-2-(4′-methyl-[1,1′-BIphenyl]-4-yl)-1H-imidazo[4,5-f][1,10]phenanthroline)]·PF6 (16b) exhibited the best potency and selectivity with excellent cellular uptake; [(η6-p-cymene)RuCl(K2-N,N-2-(4-(phenylethynyl)phenyl)-1H-imidazo[4,5-f][1,10]phenanthroline)]·PF6 (17a) acted as a cytoselective probe for live cell imaging.

Amphiphilic solid lipid nanoparticles (ASLNs) with surface PPFLMLLKGSTR peptide (Ln5-P4) (Ln5-P4/ASLNs) were prepared to load nerve growth factor (NGF) and retinoic acid (RA) and to guide the differentiation of induced pluripotent stem cells (iPSCs) toward neurons. Beeswax (BW) and lecithin played predominant roles in microemulsion and in the average diameter, zeta potential, encapsulation efficiency of NGF and RA and release kinetics of NGF- and RA-loaded Ln5-P4/ASLNs (Ln5-P4/NGF-RA-ASLNs). An increasing BW weight percentage from 0% to 75% decreased the particle size and zeta potential along with improved encapsulation efficiency of RA and NGF with enhanced positive expression of β-tubulin III to 93.72% in cultured cells. Strong affinity of Ln5-P4/NGF-RA-ASLNs to α3β1 integrin expressed on iPSCs facilitated internalization of Ln5-P4/NGF-RA-ASLNs. The capability of Ln5-P4/NGF-RA-ASLNs to induce neuronal differentiation was much higher than that of free NGF-ASLNs and RA-ASLNs, as visualized using immunochemical staining. Flow cytometry analysis indicated that Ln5-P4 on NGF-RA-ASLNs promoted the uptake of NGF and RA by iPSCs and accelerated neuronal production. Ln5-P4/NGF-RA-ASLNs are a promising colloidal delivery system to generate mature neurons from iPSCs and can be potential for treating neurodegenerative disease and nerve injury in regeneration medicine.

Solid lipid nanoparticles (SLNs) conjugated with aprotinin (Apr) and melanotransferrin antibody (Anti-MTf) were used to carry doxorubicin (Dox) across the blood–brain barrier (BBB) for glioblastoma multiforme (GBM) chemotherapy. Dox-entrapped SLNs with grafted Apr and Anti-MTf (Apr-Anti-MTf-Dox-SLNs) were applied to a cellular monolayer comprising human brain-microvascular endothelial cells (HBMECs) with a regulation of human astrocyte (HAs) and to a proliferated colony of U87MG cells. Based on the average particle diameter, zeta potential, entrapping efficiency of Dox, and grafting efficiency of Apr and Anti-MTf, we found that 1,2-dipalmitoyl-sn-glycero-3-phosphocholine of 40% (w/w) in lipids was appropriate for fabricating Apr-Anti-MTf-Dox-SLNs. In addition, Apr-Anti-MTf-Dox-SLNs could prevent Dox from fast dissolution and did not induce a serious cytotoxicity to HBMECs and HAs when compared with free Dox. Moreover, the treatment with Apr-Anti-MTf-Dox-SLNs enhanced the ability of Dox to infuse the BBB and to inhibit the growth of GBM. The current Apr-Anti-MTf-Dox-SLNs can be a promising pharmacotherapeutic preparation to penetrate the BBB for malignant brain tumor management.

Solid lipid nanoparticles (SLNs) conjugated with tamoxifen (TX) and lactoferrin (Lf) were applied to carry anticancer carmustine (BCNU) across the blood-brain barrier (BBB) for enhanced antiproliferation against glioblastoma multiforme (GBM). BCNU-loaded SLNs with modified TX and Lf (TX-Lf-BCNU-SLNs) were used to penetrate a monolayer of human brain-microvascular endothelial cells (HBMECs) and human astrocytes and to target malignant U87MG cells. The surface TX and Lf on TX-Lf-BCNU-SLNs improved the characteristics of sustained release for BCNU. When compared with BCNU-loaded SLNs, TX-Lf-BCNU-SLNs increased the BBB permeability coefficient for BCNU about ten times. In addition, TX-BCNU-SLNs considerably promoted the fluorescent intensity of intracellular acetomethoxy derivative of calcein (calcein-AM) in HBMECs via endocytosis. However, the conjugated Lf could only slightly increase the fluorescence of calcein-AM. Moreover, the order of formulation in the inhibition to U87MG cells was TX-Lf-BCNU-SLNs>TX-BCNU-SLNs>Lf-BCNU-SLNs>BCNU-SLNs. TX-Lf-BCNU-SLNs can be effective in infiltrating the BBB and delivering BCNU to GBM for future chemotherapy application.

The use of chemotherapy against brain tumors faces various limitations to achieving its therapeutic effect, due to both the inability of anticancer agents to cross the blood–brain barrier (BBB) and the formation of brain cancer stem cells (BCSCs). Without adequate exposure, these chemotherapeutic drugs cannot have an antiproliferative effect on the tumors. Here, we developed curcumin (CCM)-loaded chitosan-poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) modified with sialic acid (SA) to permeate the BBB and with anti-aldehyde dehydrogenase (anti-ALDH) to target BCSCs. An increased chitosan concentration plays a pivotal role in maintaining a steady release of CCM from NPs. The viability of BBB cells and transendothelial electrical resistance were maintained after treatment with NPs for 4 h. Immunochemical staining of human brain microvascular endothelial cells confirmed that modification of SA on the surface of NPs greatly helped in permeation of the BBB through the use of N-acetylglucosamine. In addition, immunofluorescence images evidenced the assistance of anti-ALDH in inhibiting U87MG cells and BCSCs through targeting ALDH. ALDH was colocalized with CD44 in U87MG cells and BCSCs. The cell viability assay of U87MG cells and BCSCs supported the high level of inhibition after treatment with anti-ALDH-modified NPs. The drug delivery system in this study was designed in such a way to deliver CCM into the brain and subsequently inhibit the proliferation of glioblastoma cells and BCSCs.

Solid lipid nanoparticles (SLNs) stabilized with catanionic surfactants and grafted with serotonergic 1B receptor subtype antagonist (S1BRSA) were employed to encapsulate carmustine (BCNU) (denoted as S1BRSA/BCNU–CASLNs) for brain-targeted delivery. BCNU–CASLNs were fabricated by microemulsion method and the monolayer of human brain-microvascular endothelial cells (HBMECs) regulated by human astrocytes were treated with S1BRSA/BCNU–CASLNs to evaluate the BCNU permeability across the blood–brain barrier. An increase in the catanionic surfactant concentration promoted the surface charge and toxicity to HBMECs and U87MG cells of glioblastoma multiforme (GBM). The catanionic surfactant concentration at 1 mM yielded the smallest particle size and highest entrapment efficiency of BCNU. An increase in the weight percentage of Compritol 888 ATO (CA) reduced the particle size and surface charge and enhanced the grafting efficiency of S1BRSA, viability of HBMECs, and BCNU permeability. A maximal entrapment efficiency of BCNU emerged at 50% (w/w) CA. An increase in the S1BRSA concentration reduced the grafting efficiency of S1BRSA and improved the BCNU permeability. The developed S1BRSA/BCNU–CASLNs, showing an anticancer efficacy against the growth of malignant GBM U87MG cells, were efficacious carriers in delivering BCNU to HBMECs and could be administered by intravenous injection in clinical application.

Polymeric nanoparticles (NPs) combined with lipids can have profound effects on treatment efficacy in patients with neurological disorders such as Alzheimer's disease (AD). We developed polyacrylamide (PAAM)-cardiolipin (CL)-poly(lactide-co-glycolide) (PLGA) NPs grafted with surface 83-14 monoclonal antibody (MAb) to carry rosmarinic acid (RA) and curcumin (CUR). This drug delivery system was used to cross the blood-brain barrier (BBB) and enhance the viability of SK-N-MC cells insulted with β-amyloid (Aβ) deposits. Experimental evidence revealed that an increase in the concentration of 83-14 MAb enhanced the permeability coefficient of RA and CUR using the nanocarriers. The levels of phosphorylated p38 and phosphorylated tau protein at serine 202 in degenerated SK-N-MC cells were in the order: Aβ > (Aβ + RA-CUR) > (Aβ + 83-14 MAb-RA-CUR-PAAM-PLGA NPs) > (Aβ + 83-14 MAb-RA-CUR-PAAM-CL-PLGA NPs) ≈ control. The viability of SK-N-MC cells reduced with time and CL in 83-14 MAb-RA-CUR-PAAM-CL-PLGA NPs advantaged Aβ-targeted delivery of RA-CUR. These results evidenced that the current 83-14 MAb-RA-CUR-PAAM-CL-PLGA NPs can be a promising pharmacotherapy to permeate the BBB and reduce the fibrillar Aβ-induced neurotoxicity.

The differentiation of induced pluripotent stem cells (iPSCs) in biomaterial scaffolds is an emerging area for biomedical applications. This study proposes, for the first time, the production of pancreatic cells from iPSCs in gelatin-poly(lactide-co-glycolide) nanoparticle (PLGA NP) scaffolds. The porosity and swelling ratio of the scaffolds decreased with increases in gelatin and PLGA NP concentrations. The adhesion efficiency of iPSCs in gelatin-PLGA NP scaffolds was found to be higher at 6.7% (w/w) PLGA NP. A 3-step induction of iPSCs was used to differentiate into pancreatic islet cells using activin A, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), and retinoic acid (RA). The ability of iPSCs to differentiate into pancreatic islet cells in a scaffold was demonstrated by immunofluorescence staining and flow-cytometry analysis. The results indicate that the concentration of activin A, LY294002, and RA plays a decisive role in the differentiation of iPSCs into pancreatic cells. Activin A and LY294002 induce the iPSCs into endoderm and RA induces endoderm into islet cells. A maximum insulin secretion by glucose stimulation was obtained with a higher concentration (2μM) of RA. The use of activin A-grafted gelatin-PLGA NP scaffolds induced by LY294002 and RA can be a promising approach to developing pancreatic islet cells from iPSCs.

Regenerative medicine has witnessed a paradigm shift from synthetic implants and tissue grafts to a bone tissue engineering (BTE) approach that incorporates biodegradable bioceramic composite scaffolds with biological cells. Polysaccharides, a biodegradable naturally occurring polymer can be useful either as carriers systems for active biomolecules or as cell carriers. It plays a crucial role to mimic the natural extra cellular matrix (ECM) for the development of 3D scaffold material in BTE and regenerative medicine. Alginate, a natural polysaccharide acts a biodegradable gel when crosslinked with calcium ions. Further, it encapsulates and immobilizes a variety of cells for immunoisolatory and biochemical processing applications. The bone-forming osteoblast cells can easily be attached to the alginate surface and therefore can proliferate well. Hence, alginate is widely utilized in BTE for its bioavailability, biodegradability, and ease of access.

Solid lipid nanoparticles (SLNs) with surface monoclonal antibody (MAb), poloxamer 407 (P407), and polysorbate 80 (Tween 80) (defined as P-T-MAb/SQV-SLNs) were used to enhance the transport efficacy of antiviral saquinavir (SQV) across the blood–brain barrier (BBB). Internalizations of P-T-MAb/SQV-SLNs by RAW264.7 cells and by human brain-microvascular endothelial cells were quantified by chemical assay and examined by immunochemical staining. The results revealed that a decrease in the weight percentage of Dynasan 114 (DYN) in internal lipids, comprising DYN, palmitic acid, and cacao butter, increased the particle size and zeta potential of P-T-MAb/SQV-SLNs. In addition, the external P407 and Tween 80 could stabilize P-T-MAb/SQV-SLNs and reduce phagocytosis by RAW264.7 cells. Tween 80 on P-T-MAb/SQV-SLNs also benefited the delivery of SQV across the BBB. Moreover, MAb grafted on P-T-MAb/SQV-SLNs promoted the permeability of SQV across the blood–brain barrier. The grafting of MAb and coating of P407 and Tween 80 on the surface of SLNs demonstrate an effective strategy to prevent phagocytosis by RAW264.7 cells and to promote the targeting delivery of SQV across the BBB for inhibiting retroviral growth.

Saquinavir (SQV)-loaded catanionic solid lipid nanoparticles (CASLNs) were grafted with surface transferrin (Tf) (Tf/SQV-CASLNs) for targeting the blood–brain barrier (BBB). Tf/SQV-CASLNs were cultured with human brain-microvascular endothelial cells (HBMECs) to demonstrate the particulate uptake via Tf receptors for the BBB transport. The results revealed that an increase in the concentration of catanionic surfactants from 1 mM to 2 mM enlarged the average diameter of Tf/SQV-CASLNs from 150 nm to 500 nm. An increasing weight percentage of Compritol 888 ATO (CA) in lipids enhanced the viability of HBMECs. However, an increasing concentration of catanionic surfactants reduced the viability. The order of the CA weight percentage in the permeability of SQV using Tf/SQV-CASLNs to cross the BBB is 33% > 100% > 0%. In addition, an increase in the concentration of Tf on Tf/SQV-CASLNs enhanced the BBB permeability of SQV. Tf/SQV-CASLNs fabricated with 33% (w/w) CA in the lipid phase, catanionic surfactants of 1 mM, and Tf of 30 μg/mL can be efficient colloidal carriers for delivering SQV into the brain.

Yung-Chih Kuo, Yin-Jung Lee Department of Chemical Engineering, National Chung Cheng University, Chia-Yi, Taiwan, Republic of China Abstract: β-Amyloid (Aβ)-targeting liposomes (LIP) with surface serotonin modulator (SM) and apolipoprotein E (ApoE) were utilized to facilitate the delivery of nerve growth factor (NGF) across the blood–brain barrier (BBB) for neuroprotection in the hippocampus. The therapeutic efficacy of SM- and ApoE-grafted LIP carrying NGF (NGF-SM-ApoE-LIP) was assessed by an in vitro Alzheimer’s disease (AD) model of degenerated SK-N-MC cells and an in vivo AD model of Aβ-insulted Wistar rats. The experimental evidences revealed that the modified SM and ApoE on the surface of LIP increased the permeation of NGF across the BBB without serious damage to structural integrity of tight junction. When compared with free NGF, NGF-SM-ApoE-LIP upregulated the expression of phosphorylated neurotrophic tyrosine kinase receptor type 1 on cholinergic neurons and significantly improved their survival. In addition, NGF-SM-ApoE-LIP could reduce the secretion of acetylcholinesterase and malondialdehyde and rescue hippocampal neurons from apoptosis in rat brains. The synergistic effect of SM and ApoE is promising in the induction of NGF to inhibit the neurotoxicity of Aβ and NGF-SM-ApoE-LIP can be a potent antiapoptotic pharmacotherapy for clinical care of patients with AD. Keywords: Alzheimer’s disease, blood–brain barrier, serotonin modulator, apolipoprotein E, nerve growth factor, liposome

Cholesterol-mediated cationic solid lipid nanoparticles (CSLNs) were formulated with esterquat 1 (EQ 1) and stearylamine as positively charged external layers on hydrophobic internal cores of cacao butter. These CSLNs were employed to deliver saquinavir (SQV) to the brain. The permeability of SQV across the blood-brain barrier (BBB) using SQV-loaded CSLNs (SQV-CSLNs) was estimated with an in vitro model of a monolayer of human brain-microvascular endothelial cells (HBMECs) regulated by human astrocytes. The results revealed that the average diameter of SQV-CSLNs diminished when the weight percentage of cholesterol and EQ 1 increased. The morphological images indicated a uniform size of SQV-CSLNs with compact lipid structure. In addition, an increasing weight percentage of cholesterol and EQ 1 enhanced the zeta potential of SQV-CSLNs. The fluorescent staining demonstrated that HBMECs could internalize SQV-CSLNs. An increase in the weight percentage of cholesterol and EQ 1 also promoted the uptake of SQV-CSLNs by HBMECs. Moreover, a high content of cholesterol and EQ 1 in SQV-CSLNs increased the BBB permeability of SQV. The cholesterol-mediated SQV-CSLNs can be an efficacious drug delivery system for brain-targeting delivery of antiviral agents.

The understanding of differentiating induced pluripotent stem (iPS) cells in porous biomaterials is a critical challenge in recent biotechnological development. This study presents the investigation on the differentiation of iPS cells toward neurons in biomedical scaffolds containing alginate, chitosan, and gelatin with grafted neuron growth factor (NGF). Alginate-chitosan-gelatin scaffolds were prepared by particulate leaching method using polystyrene (PS) microspheres as porogen. In addition, the neuronal differentiation of iPS cells in the constructs was identified by immunochemical staining. The morphological studies demonstrated that an increase in the concentration of PS microspheres from 0.5 to 0.75 g/mL improved the pore regularity in alginate-chitosan-gelatin hydrogels. The effect of composition on the differentiation of iPS cells into neuronal lineage was in the order where alginate:chitosan:gelatin=1:1:3>2:1:2>1:1:1. Moreover, an increase in the concentration of NGF promoted the neuronal production from iPS cells in cultivated constructs. Alginate-chitosan-gelatin scaffolds with surface NGF can guide the differentiation of iPS cells for regenerating neurons.

The ability of cryopreserved chondrocytes to revitalize and propagate is a key biotechnology in cartilage regeneration. This study shows the formation of neocartilage from cryopreserved chondrocytes in scaffolds grafted with elastin and poly-L-lysine. Cryopreserved chondrocytes in elastin- and poly-L-lysine-grafted constructs were cultured in a dynamic bioreactor and assessed by biochemical assay and staining. Elastin demonstrated a better efficacy for recruiting cryopreserved chondrocytes onto the pore surface of constructs than poly-L-lysine. However, surface elastin and poly-L-lysine did not significantly enhance the biocompatibility to cryopreserved chondrocytes. Chondrocytes multiplied from cryopreserved chondrocytes in elastin-grafted constructs is faster than that in poly-L-lysine-grafted constructs. In addition, elastin could stimulate cryopreserved chondrocytes to synthesize more glycosaminoglycans and collagen than poly-L-lysine. Porous biomaterials with surface elastin and poly-L-lysine can maintain active chondrocytic proliferation and extracellular matrix secretion from chondrocytes with appropriate cryopreservation.

Solid lipid nanoparticles (SLNs) comprising complex internal lipids were conjugated with melanotransferrin antibody (MA) to carry anticancer etoposide across the blood-brain barrier (BBB) for managing glioblastoma multiforme (GBM). MA was crosslinked on the surface of etoposide-loaded SLNs (ETP-SLNs) to target human brain-microvascular endothelial cells (HBMECs) and U87MG cells. The experimental evidences showed that an increase in the tripalmitin weight percentage in lipids enhanced the particle size and viability of U87MG cells, however decreased the etoposide loading efficiency, MA conjugation efficiency, and permeability coefficient for etoposide across the BBB. A high level of MA on the particle surface increased the atomic ratio of nitrogen to phosphorus and permeability coefficient for propidium iodide and etoposide across the BBB, however reduced the MA conjugation efficiency, transendothelial electrical resistance, and viability of U87MG cells. Based on immunochemical staining, we found that MA on ETP- SLNs triggered the melanotransferrin-mediated transcytosis and promoted the growth-inhibitory efficacy to U87MG cells. MA- conjugated ETP-SLNs can be a promising colloidal delivery system for malignant GBM pharmacotherapy.

Understanding of charged groups on nanostructured lipid particles is an important issue in nanomedicine development. This study presents the relation between noncovalent interactions and the composition of ionogenic groups in the assembled lipid layer of cationic solid lipid nanoparticles (CSLNs). Innovated CSLNs containing cacao butter, cholesterol, stearylamine, and esterquat 1 (EQ 1) were fabricated by modified solvent diffusion method. The results revealed that the average diameter of CSLNs decreased when the weight percentage of cholesterol and EQ 1 increased. X-ray photoelectron spectroscopy and Fourier-transform infrared spectroscopy evidenced strong interactions between cholesterol and EQ 1. These noncovalent interactions exhibited hydrophobic and cation–aromatic characteristics and mediated the distribution of lipids in the external layer of CSLNs. In addition, an increase in the weight percentage of cholesterol and EQ 1 enhanced the zeta potential, electrophoretic mobility, and fixed charge dens...

The potential of constructs comprising induced pluripotent stem (iPS) cells and biopolymers can be high for neurological surgery practice, if the systematic activity of neuronal regeneration is clarified. This study shows a guided differentiation of iPS cells toward neurons in neuron growth factor (NGF)-grafted poly(ε-caprolactone) (PCL)-poly(β-hydroxybutyrate) (PHB) scaffolds. The porosity of PCL-PHB scaffolds enhanced with increasing the concentration of salt particles (porogen) and the weight percentage of PCL. An increase in the graft concentration of NGF elevated the atomic ratios of N/C and O/C on the surface of NGF-grafted PCL-PHB scaffolds. In addition, incorporating heparin and NGF promoted the adhesion and viability of iPS cells in constructs. When the weight percentage of PCL increased, the viability of iPS cells reduced; however, more PCL in constructs benefited the adhesion of iPS cells. Under the influence of heparin and NGF, a high weight percentage of PCL and a long inductive period improved iPS cells to differentiate into neuron-like cells carrying βIII tubulin and inhibited other differentiation(s). The material-driven differentiation in NGF-grafted PCL-PHB constructs can be promising in guiding iPS cells to produce neurons for nerve tissue engineering.

Formation of neocartilage is a critical issue in contemporary regenerative medicine. This study presents the generation of tissue engineering cartilage in TATVHL peptide-grafted scaffolds. Bovine knee chondrocytes were seeded in TATVHL peptide-grafted scaffolds and cultured in a spinner bioreactor. The results revealed that surface TATVHL peptide enhanced the adhesion of bovine knee chondrocytes in scaffolds. However, an increase in the concentration of TATVHL peptide in scaffolds (up to 20 μg/mL) did not cause an evident variation in the cell viability. Surface TATVHL peptide was effective in promoting the quantity of cartilaginous components in constructs after dynamic cultivation. Biochemical assay, scanning electron microscope images, and histological staining demonstrated that surface TATVHL peptide accelerated the proliferation of bovine knee chondrocytes in constructs. In addition, the secretion of glycosaminoglycans and production of collagen in TATVHL peptide-grafted constructs were faster than those in TATVHL peptide-free constructs. TATVHL peptide can be a promising bioactive molecule to improve chondrogenesis in porous biomaterials.

An exposure to an electromagnetic field (EMF) has been shown to enhance the membrane permeability of endothelial cells. This study investigates the effect of EMF on the modulation of P-glycoprotein (P-gp) and multidrug resistance-associated protein (MRP) for delivering saquinavir (SQV) and azidothymidine (AZT). To assess the transport of SQV and AZT, an EMF and P-gp/MRP (transport protein) inhibitors were employed to stimulate confluent human brain-microvascular endothelial cells (HBMECs) with the regulation of human astrocytes. SQV at 40 μM and AZT at 300 μM were acceptable dosages for the viability of HBMECs. Under an EMF, verapamil showed a higher ability to elevate the permeation across the blood-brain barrier (BBB) than probenecid during treating with SQV. The reverse was true during treating with AZT. After an exposure to an EMF and administration of SQV and AZT, P-gp and MRP1 localized on the luminal side of HBMECs, exhibiting polar characteristics of the cell membranes. The synergetic effect of an EMF exposure and efflux protein inhibition can be promising in delivering antiretroviral drugs across the BBB.

This study investigates the capacity of albumin-grafted biomaterials as tissue engineering scaffolds to regenerate cartilaginous components. Porcine knee chondrocytes were seeded and cultivated in porous ternary matrix consisting of polyethylene oxide, chitin, and chitosan with surface albumin. The results revealed that the quantity of albumin did not affect the viability of porcine knee chondrocytes in the constructs. However, a high grafting concentration of albumin favored the adhesion of porcine knee chondrocytes on the scaffolding pore surface. After cultivation over 4 weeks, an increase in the concentration of albumin enhanced the quantities of porcine knee chondrocytes, glycosaminoglycans, and collagen in the constructs. The histological staining of porcine knee chondrocytes showed an active chondrocytic growth in the albumin-grafted constructs. In addition, the safranin-O staining indicated that the surface albumin could stabilize the secretion of glycosaminoglycans. Moreover, the immunochemical staining against type II collagen exhibited a regular production of collagen by phenotypic porcine knee chondrocytes in the constructs. Albumin-grafted polyethylene oxide/chitin/chitosan scaffolds can be a promising biomimetic substrate in neocartilage formation.

In this study, through the extension of an one-dimensional, dissimilarly charged protrusions surface model set up in our previous work, a novel dissimilarly charged protrusion array (DCPA) model immersed in an electrolyte solution, which could simulate realistically both the surface morphology and the surface charged condition profoundly concerned on a biological cell membrane, or on the surface of a micro-scale, modified particle used in biomedical engineering and water treatment, is proposed. Considering the condition of small protrusions, the electrical potential field due to the electrical double layer (EDL) on DCPA model is solved semi-analytically using both the double Fourier series and the perturbation method. The analysis from the numerical result reveals that, a small, dissimilarly charged protrusion can lead to a steep variation in the local EDL configuration, especially compared with that in the condition when the charged surface is taken roughly as a flat surface using a lumped, mean surface charge density.

Aims: Poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs) were grafted with transferrin (Tf) to enhance the transport of nevirapine (NVP) across human brain microvascular endothelial cells (HBMECs). Methods: NVP-loaded PLGA NPs with surface-grafting Tf (Tf/NVP–PLGA NPs) were incubated with HBMECs and immunochemical staining characterized Tf receptors (TfRs). Results: The polydispersity index of Tf/NVP–PLGA NPs was lower than 0.008. The entrapment efficiency of NVP and loading efficiency of Tf was 20–75% and 15–80%, respectively. Tf slightly retarded the release of NVP from PLGA. Dioctadecyldimethylammonium bromide (DODAB)-stabilized Tf/NVP–PLGA NPs reduced the viability of HBMECs to 70–75%. The secretion of TNF-α was inhibited by Tf and stimulated by DODAB. The permeability of NVP across HBMECs reached maxima at 67% DODAB and 0.1–0.2% Tf. An increase in the concentration of Tf enhanced the uptake of Tf/NVP–PLGA NPs via a TfR-mediated mechanism. Conclusion: Tf/NVP–PLGA NPs are efficacious carriers in targeting delivery across HBMECs for viral therapy.

The present study presents the regeneration of cartilage in hybrid scaffolds comprising polyethylene oxide (PEO) and chitosan with surface CDPGYIGSR. This surface peptide was grafted via crosslinking onto the scaffolds. The pores in the scaffolds were interconnected and uniformly distributed with an average diameter about 200-250 μm. A high weight percentage of PEO in the matrix yielded a rugged topography of the pore surfaces. The adhesion of bovine knee chondrocytes (BKCs) in the peptide-grafted scaffolds was more efficient than that in the peptide-free scaffolds. In addition, the constructs with surface peptide could stimulate chondrogenesis with enhanced quantities of BKCs, glycosaminoglycans (GAGs), and collagen over cultivation. The histological staining of the proliferated BKCs and secreted GAGs indicated that the surface peptide favored the production of neocartilage in the constructs. Moreover, the immunochemical staining against type II collagen demonstrated the maintenance of phenotypic chondeocytes on the peptide-grafted surfaces. The peptide-grafted PEO/chitosan scaffolds can be applied to the treatment for injured cartilage in preclinical trials.

Inverted colloidal crystal (ICC) scaffolds grafted with two defined peptides, CSRARKQAASIKVAVSADR and CDPGYIGSR, and bone marrow stromal cells (BMSCs) were applied to the treatment for spinal cord injury (SCI) of model rats. The results revealed that the neuronal survival, characterized by the Nissl staining, in the injured thoracic spine at T10 decreased drastically after SCI without treatment over 4 weeks. The neuronal survival was enhanced by the present treatments. The order in the neuronal survival was BMSCs in peptide-modified ICC construct>BMSCs in peptide-free ICC construct>direct injection of BMSCs. Based on the nestin and neurofilament-H staining, the effect of therapeutic method on the quantity of neuronal precursor cells and axonal growth followed the same order as the neuronal survival. When treated with peptide-modified ICC construct, the expression of glial fibrillary acidic protein and tumor necrosis factor-α significantly reduced, indicating the inhibition of the formation of glial scar tissue and inflammatory cytokine. The ICC topography with the surface peptides can guide the differentiation of BMSCs toward neurons for nerve regeneration and enhance the SCI treatment.