Chinese hamster ovary cell line engineering strategies for modular production of custom extracellular vesicles.

Continuously secreted by all cell types, extracellular vesicles (EVs) are small membrane-bound structures which shuttle bioactive cargo between cells across their external environment. Their central role as natural molecular messengers and ability to cross biological barriers has garnered significant attention in the use of EVs as therapeutic delivery vehicles. Still, harnessing the potential of EVs is faced with many obstacles. A cell line engineering approach can be used to exploit EVs to encapsulate a bespoke cargo of interest. However, full details regarding native EV-loading mechanisms remain under debate, making this a challenge. While Chinese hamster ovary (CHO) cells are well known to be the preferred host for recombinant therapeutic protein production, their application as an EV producer cell host has been largely overlooked. In this study, we engineered CHO DG44 cells to produce custom EVs with bespoke cargo. To this end, genetic constructs employing split green fluorescent protein technology were designed for tagging both CD81 and protein cargoes to enable EV loading via self-assembling activity. To demonstrate this, NanoLuc and mCherry were used as model reporter cargoes to validate engineered loading into EVs. Experimental findings indicated that our custom EV approach produced vesicles with up to 15-fold greater cargo compared with commonly used passive loading strategies. When applied to recipient cells, we observed a dose-dependent increase in cargo activity, suggesting successful delivery of engineered cargo via our custom CHO EVs.

SARS-CoV-2 NSP2 as a Potential Delivery Vehicle for Proteins.

The development of biomolecule delivery systems is essential for the treatment of various diseases such as cancer, immunological diseases, and metabolic disorders. For the first time, we found that SARS-CoV-2-encoded nonstructural protein 2 (NSP2) can be secreted from the cells, where it is synthesized. Brefeldin A and H89, inhibitors of ER/Golgi secretion pathways, did not inhibit NSP2 secretion. NSP2 is likely secreted via an unconventional secretory pathway. Moreover, both secreted and purified NSP2 proteins were able to traverse the plasma membrane barrier and enter both immortalized human umbilical vein endothelial cells and tumor cell lines. After entry, the NSP2 protein was localized in only the cytoplasm. Cytochalasin D, a potent inhibitor of actin polymerization, inhibited the entry of NSP2. NSP2 can carry other molecules into cells. Burkholderia lethal factor 1, a monomeric toxin from the intracellular pathogen Burkholderia pseudomallei, has demonstrated antitumor activity by targeting host eukaryotic initiation translation factor 4A. An NSP2-BLF1 fusion protein was translocated across the cellular membranes of Huh7 cells and mediated cell killing. By using different approaches, including protein purification, chemical inhibition, and cell imaging, we confirm that NSP2 is able to deliver heterologous proteins into cells. NSP2 can act as a potential delivery vehicle for proteins.

Comparison of budesonide vehicles in inducing histologic remission in pediatric eosinophilic esophagitis.

OBJECTIVES: Oral viscous budesonide (OVB) is a common medication used to treat eosinophilic esophagitis (EoE). It is typically mixed with Splenda to produce a slurry, but other delivery vehicles have been used in clinical practice. We aimed to evaluate outcomes of pediatric EoE patients treated with OVB using different drug delivery vehicles. METHODS: We performed a retrospective chart review of pediatric EoE patients treated with OVB. The primary aim was to evaluate rates of histologic remission (defined by <15 eosinophils per high power field in both mid and distal esophagus) after 6-12 weeks of OVB treatment for each delivery vehicle. Secondary aims were to evaluate histologic response and endoscopic response and remission of different delivery vehicles, and to compare the efficacy of different treatment regimens. RESULTS: A total of 111 patients were included in the study. Median treatment duration was 3.4 months. Overall rate of histologic remission with OVB was 52.6%. There was no difference in rates of histologic remission (p = 0.313) or response (p = 0.195 and p = 0.681 in mid and distal esophagus, respectively) among the different vehicle types or treatment regimens. Similarly, there was no difference in endoscopic remission and response among the different vehicle types (p = 0.853 and p = 0.727) or treatment regimens (p = 0.244 and p = 0.157). Patients who achieve histologic remission were more likely to be non-Hispanic Caucasian. CONCLUSION: Our findings suggest there is no difference in histologic and endoscopic outcomes with various delivery vehicles or combination therapy with OVB in the treatment of EoE. More palatable and cost-effective vehicles can be used to treat EoE.

Orally administered Streptococcus thermophilus YIT 2001 is a vehicle for the delivery of glutathione, a reactive reduced thiol, to the intestine.

AIMS: We aimed to analyze the behavior of cellular glutathione of Streptococcus thermophilus strain YIT 2001 (ST-1) in the gastrointestinal environment to understand how orally administered glutathione in ST-1 cells is delivered stably to the intestine in a reactive form, which is essential for its systemic bioavailability against lipid peroxidation. METHODS AND RESULTS: Intracellular glutathione was labeled with L-cysteine-containing stable isotopes. ST-1 cells from fresh culture or lyophilized powder were treated with simulated gastric and intestinal juices for 60 min each. The release of intracellular glutathione in digestive juices was quantified via LC-MS/MS. Most of the cellular glutathione was retained in the gastric environment and released in response to exposure to the gastrointestinal environment. During digestion, the membrane permeability of propidium iodide increased significantly, especially when cells were exposed to cholate, without change in the cell wall state. CONCLUSIONS: ST-1 cells act as vehicles to protect intracellular reactive components, such as glutathione, from digestive stress, and release them in the upper intestine owing to the disruption of membrane integrity induced by bile acid.

Extracellular Vesicles in Domestic Animals: Cellular Communication in Health and Disease.

Apoptotic and healthy cells of domestic animals release membrane-enclosed particles from their plasma membrane. These special structures, called extracellular vesicles, play an important role in intercellular communication. In the past, it was believed that their function was mainly to dispose unwanted cell contents and to help maintain cell homeostasis. However, we now know that they have important roles in health and disease and have diagnostic value as well as great potential for therapy in veterinary medicine. Extracellular vesicles facilitate cellular exchanges by delivering functional cargo molecules to nearby or distant tissues. They are produced by various cell types and are found in all body fluids. Their cargo reflects the state of the releasing parent cell, and despite their small size, this cargo is extraordinarily complex. Numerous different types of molecules contained in vesicles make them an extremely promising tool in the field of regenerative veterinary medicine. To further increase research interest and discover their full potential, some of the basic biological mechanisms behind their function need to be better understood. Only then will we be able to maximize the clinical relevance for targeted diagnostic and therapeutic purposes in various domestic animal species.

Exosomes Derived from Mesenchymal Stem Cells: Therapeutic Opportunities for Spinal Cord Injury.

Spinal cord injury (SCI) is a serious neurological condition comprising primary and secondary injury and causing severe neurological impairments. The effect of the conventional treatment is limited, including supportive therapy and emergency surgery. Exosomes derived from mesenchymal stem cells (MSCs-Exos) were previously reported to exert its potential therapeutic effects on SCI. Compared with mesenchymal stem cells (MSCs) transplantation for SCI, MSC-Exos showed several superiorities. In the present review, we summarized the revealed data of mechanisms underlying MSC-Exos repairing of SCI and discussed the issues of MSC-Exos use. Thus, in this review we summarized the latest studies on MSCs-Exos in the therapy of SCI and discussed whether MSCs-Exos can be applied to SCI and the prospects of transformation application.

Mesenchymal stem cells: a trojan horse to treat glioblastoma.

Glioblastoma multiforme (GBM) is the most common and lethal primary tumor of the central nervous system. What makes it so dreadful is the very low survival rate, despite the existence of a standard treatment plan. An innovative and more effective way to treat glioblastoma based on Mesenchymal Stem Cells (MSCs) has been explored recently. MSCs are a group of endogenous multipotent stem cells that could mainly be harvested from adipose tissue, bone marrow, and umbilical cord. Having the ability to migrate toward the tumor using multiple types of binding receptors, they could be used either as a direct treatment (whether they are enhanced or not) or as a delivery vehicle carrying various anti-tumoral agents. Some of these agents are: chemotherapy drugs, prodrug activating therapy, oncolytic viruses, nanoparticles, human artificial chromosome… Promising results have started to surface; however, more evidence is needed to perfect their use as a glioblastoma multiforme treatment option. Alternative treatment, using unloaded or loaded MSCs, leading to a better outcome.

S-Benzyl cysteine based cyclic dipeptide super hydrogelator: Enhancing efficacy of an anticancer drug via sustainable release.

Peptide-based low molecular weight supramolecular hydrogels hold promising aspects in various fields of application especially in biomaterial and biomedical sciences such as drug delivery, wound healing, tissue engineering, cell proliferation, and so on due to their extreme biocompatibility. Unlike linear peptides, cyclic peptides have more structural rigidity and tolerance to enzymatic degradation and high environmental stability which make them even better candidates for the above-said applications. Herein, a new small cyclic dipeptide (CDP) cyclo-(Leu-S-Bzl-Cys) (P1) consisting of L-leucine and S-benzyl protected L-cysteine was reported which formed a hydrogel at physiological conditions (at 37°C and pH = 7.46). The hydrogel formed from the cyclic dipeptide P1 showed very good tolerance towards environmental parameters such as pH and temperature and was seen to be stable for more than a year without any deformation. The hydrogel was thermoreversible and stable in the pH range 6-12. Mechanical strength of P1 hydrogel was measured by rheology experiments. Atomic force microscopy (AFM) and field emission scanning electron microscopy (FE-SEM) images revealed that, in aqueous solvents, P1 self-assembled into a highly cross-linked nanofibrillar network which immobilized water molecules inside the cages and formed the hydrogel. The self-assembled cyclic dipeptide acquired the antiparallel ß-sheet secondary structure, which was evident from CD and Fourier transform infrared (FT-IR) studies. The ß-sheet arrangement and formation of amyloid fibrils were further established by ThT binding assay. Furthermore, P1 was able to form a hydrogel in the presence of the anticancer drug 5-fluorouracil (5FU), and sustainable release of the drug from the hydrogel was measured in vitro. The hydrogelator P1 showed almost no cytotoxicity towards the human colorectal cancer cell line HCT116 up to a considerably high concentration and showed potential application in sustainable drug delivery. The co-assembly of 5FU and P1 hydrogel exhibited much better anticancer activity towards the HCT116 cancer cell line than 5FU alone and decreased the IC50 dose of 5FU to a much lower value.

Applications of Synthetic Polymer Discoidal Lipid Nanoparticles to Biomedical Research.

Liposomes are artificially prepared vesicular lipid nanoparticles with a bilayer structure, resembling cell membrane. Their ability to encapsulate various molecules along with excellent biocompatibility makes them ideal delivery vehicles for pharmaceuticals. They can also serve as platforms for membrane proteins to elucidate the structure and function in lipid membranes. Nascent high-density lipoproteins are discoidal lipid nanoparticles with a bilayer structure, which can be reconstituted with their constituents. Such reconstituted nanoparticles, nanodisks, were originally generated in terms of elucidation for mechanisms of lipoprotein metabolisms. At the same time, like liposomes, nanodisks have been developed as delivery vehicles and platforms for membrane proteins in structural biology. From a developmental background, apolipoproteins, their analogs, or fragment peptides were initially used as scaffolding molecules to wrap around the edge of the disk-shaped lipid bilayer. Since the discovery that styrene-maleic acid copolymers produce nanodisks instead of apolipoproteins, variously modified or novel polymers have been synthesized to broaden the applications of polymer nanodisks. This review provides an overview of the types of synthetic polymers used to produce nanodisks, and the biomedical applications of nanodisks to the developments of delivery vehicles and to the structural studies of membrane proteins.

Supramolecular Protein-Polyelectrolyte Assembly at Near Physiological Conditions-Water Proton NMR, ITC, and DLS Study.

The in vivo potency of polyphosphazene immunoadjuvants is inherently linked to the ability of these ionic macromolecules to assemble with antigenic proteins in aqueous solutions and form physiologically stable supramolecular complexes. Therefore, in-depth knowledge of interactions in this biologically relevant system is a prerequisite for a better understanding of mechanism of immunoadjuvant activity. Present study explores a self-assembly of polyphosphazene immunoadjuvant-PCPP and a model antigen-lysozyme in a physiologically relevant environment-saline solution and neutral pH. Three analytical techniques were employed to characterize reaction thermodynamics, water-solute structural organization, and supramolecular dimensions: isothermal titration calorimetry (ITC), water proton nuclear magnetic resonance (wNMR), and dynamic light scattering (DLS). The formation of lysozyme-PCPP complexes at near physiological conditions was detected by all methods and the avidity was modulated by a physical state and dimensions of the assemblies. Thermodynamic analysis revealed the dissociation constant in micromolar range and the dominance of enthalpy factor in interactions, which is in line with previously suggested model of protein charge anisotropy and small persistence length of the polymer favoring the formation of high affinity complexes. The paper reports advantageous use of wNMR method for studying protein-polymer interactions, especially for low protein-load complexes.

MicroRNA-4731-5p delivered by AD-mesenchymal stem cells induces cell cycle arrest and apoptosis in glioblastoma.

Glioblastoma multiforme (GBM) exhibits the most malignant brain tumor with very poor prognosis. MicroRNAs (miRNAs) are regulatory factors that can downregulate the expression of multiple genes. Several miRNAs acting as tumor-suppressor genes have been identified so far. The delivery of miRNA by mesenchymal stem cell (MSC) due to their ability to specifically target tumors is a new, hopeful therapeutic approach for glioblastoma. The objective of our study is the investigation of the effect of lentivirus-mediated microRNA-4731 (miR-4731) genetic manipulated adipose-derived (AD)-MSC on GBM. The downregulation of miR-4731 in human GBM tumor was detected using the GEO dataset. To evaluate the function of miR-4731, we overexpressed miR-4731 using lentiviral vectors in U-87 and U-251 GBM cell lines. The effects of miR-4731 on cell proliferation and cell cycle of glioma cells were analyzed by wound test and flow-cytometry assay. miR-4731 inhibited the proliferation of GBM cancer cells. Coculturing was used to study the antiproliferative effect of miR-4731-AD-MSCs on GBM cell lines. Direct and indirect coculture of GBM cell lines with miR-4731-AD-MSCs induced cell cycle arrest and apoptosis. Our findings suggest that AD-MSCs expressing miR-4731 have favorable antitumor characteristics and should be further explored in future glioma therapy.

Biological functions and clinical applications of exosomal non-coding RNAs in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide, with a high mortality rate. Its dismal prognosis is attributed to late diagnosis, high risk of recurrence and drug resistance. To improve the survival of patients with HCC, new approaches are required for early diagnosis, real-time monitoring and effective treatment. Exosomes are small membranous vesicles released by most cells that contain biological molecules and play a great role in intercellular communication under physiological or pathological conditions. In cancer, exosomes from tumor cells or non-tumor cells can be taken up by neighboring or distant target cells, and the cargoes in exosomes are functional to modulate the behaviors of tumors or reshape tumor microenvironment (TME). As essential components, non-coding RNAs (ncRNAs) are selectively enriched in exosomes, and exosomal ncRNAs participate in regulating specific aspects of tumor development, including tumorigenesis, tumor metastasis, angiogenesis, immunomodulation and drug resistance. Besides, dysregulated exosomal ncRNAs have emerged as potential biomarkers, and exosomes can serve as natural vehicles to deliver tumor-suppressed ncRNAs for treatment. In this review, we briefly summarize the biology of exosomes, the functions of exosomal ncRNAs in HCC development and their potential clinical applications, including as biomarkers and therapeutic tools.

Novel 20% doxycycline-loaded PLGA nanospheres as adjunctive therapy in chronic periodontitis in type-2 diabetics: randomized clinical, immune and microbiological trial.

OBJECTIVE: This study evaluated the clinical, microbiological, and immunological results of poly lactic-co-glycolic acid (PLGA) nanospheres containing 20% doxycycline (DOXY) in the treatment of type-2 diabetic patients (DM-2) with chronic periodontitis (CP). MATERIAL AND METHODS: A parallel, double-blind, randomized, placebo-controlled clinical trial was conducted in DM-2 presenting severe and generalized CP. All patients received one-stage full-mouth ultrasonic debridement (FMUD) and they were randomly divided into two groups: PLAC (n = 20)-local application of placebo PLGA nanospheres, and DOXY (n = 20)-local application of doxycycline-loaded nanospheres; both in six non-contiguous sites. Clinical, metabolic (fasting plasma glucose level-FPG and glycated hemoglobin-HbA1c), cytokine pattern (multiplexed bead immunoassay) and microbiological assessments were performed at baseline, and 1, 3, and 6 months after treatment. RESULTS: Both groups showed clinical improvement in all parameters after treatment (p < 0.05). Deep pockets showed improvements in bleeding on probing-BoP (3 and 6 months), PD (at 3 months), and CAL gain (at 1 and 3 months) favoring DOXY (p < 0.05). The percentage of sites presenting PD reduction and CAL gain ≥ 2 mm was higher in DOXY at 3 months (p < 0.05). DOXY group exhibited a significant increase in the levels of anti-inflammatory interleukin (IL)-10 and a reduction in IL-8, IFN-y, IL-6, and IL-17 (p < 0.05), significant reduction in periodontal pathogens (p < 0.05), and a lower mean percentage of HbA1C at 3 months (p < 0.05). CONCLUSION: DOXY nanospheres may be considered a potential adjunct to mechanical debridement in the therapy of periodontitis in DM-2, offering additional benefits in deep pockets, improving the cytokine profile, and reducing periodontal pathogen levels. CLINICAL RELEVANCE: The use of locally applied doxycycline nanospheres may represent an adjunctive therapeutic approach in the treatment of periodontal disease in type-2 diabetic patients, achieving additional benefits in the local modulation of cytokines, microbial reduction, and clinical parameters, especially in deep pockets.

The Biological Function and Therapeutic Potential of Exosomes in Cancer: Exosomes as Efficient Nanocommunicators for Cancer Therapy.

Cancer therapeutics must be delivered to their targets for improving efficacy and reducing toxicity, though they encounter physiological barriers in the tumor microenvironment. They also face limitations associated with genetic instability and dynamic changes of surface proteins in cancer cells. Nanosized exosomes generated from the endosomal compartment, however, transfer their cargo to the recipient cells and mediate the intercellular communication, which affects malignancy progression, tumor immunity, and chemoresistance. In this review, we give an overview of exosomes' biological aspects and therapeutic potential as diagnostic biomarkers and drug delivery vehicles for oncotherapy. Furthermore, we discuss whether exosomes could contribute to personalized cancer immunotherapy drug design as efficient nanocommunicators.

Generation of microRNA-30e-producing recombinant viral hemorrhagic septicemia virus (VHSV) and its effect on in vitro immune responses.

MicroRNAs (miRNAs) are non-coding small RNAs involved in the regulation of gene expression. In the present study, we firstly reported the use of a fish RNA virus, viral hemorrhagic septicemia virus (VHSV), as a delivery vehicle of a miRNA-30e, and the effect of miR-30e produced by the recombinant VHSV on the immune responses of Epithelioma papulosum cyprini (EPC) cells was investigated. The expression of functional miR-30e using a CMV promoter-driven vector was verified by the significantly lower eGFP expression in cells transfected with a vector containing miR-30e sponge sequence than that in cells transfected with a control vector that had mutated miR-30e sponge sequence. Furthermore, the down-regulation of reporter gene containing 3'-UTR of NF-κb inhibitor α-like protein B (NFκbiαb) by miR-30e was demonstrated, suggesting that miR-30e overexpression can increase immune responses related to NF-κB activation through inhibition of IκB. A miR-30e-expressing recombinant VHSV (rVHSV-A-miR30e) that had primary microRNA-30e sequence between N and P genes was rescued using the reverse genetic method, and the successful expression of miR-30e in the cells infected with rVHSV-A-miR30e was demonstrated using Northern blot and qRT-PCR. Cells infected with rVHSV-A-miR30e showed the increase of NF-κB activation and type I interferon induced genes expression, suggesting that rVHSV-A-miR30e can produce functional miR-30e in fish cells, and VHSV can be used as a vehicle to deliver functional microRNAs in fish.

Bifidobacterium spp: the promising Trojan Horse in the era of precision oncology.

Selective delivery of therapeutic agents into solid tumors has been a major challenge impeding the achievement of long-term disease remission and cure. The need to develop alternative drug delivery routes to achieve higher drug concentration in tumor tissue, reduce unwanted off-target side effects and thus achieve greater therapeutic efficacy, has resulted in an explosive body of research. Bifidobacterium spp. are anaerobic, nonpathogenic, Gram-positive bacteria, commensal to the human gut that are a possible anticancer drug-delivery vehicle. In this review, we describe Bifidobacterium's microbiology, current clinical applications, overview of the preclinical work investigating Bifidobacterium's potential to deliver anticancer therapy, and review the different strategies used up to date. Finally, we discuss both current challenges and future prospects.

Preparation of Lipid Nanodisks Containing Apolipoprotein E-Derived Synthetic Peptides for Biocompatible Delivery Vehicles Targeting Low-Density Lipoprotein Receptor.

High-density lipoprotein (HDL) particles that are formed in vivo adopt a disk-shaped structure, in which the periphery of the discoidal phospholipid bilayer is surrounded by apolipoprotein. Such discoidal nanoparticles can be reconstituted with certain apolipoproteins and phospholipids and are commonly called lipid nanodisks. Apolipoprotein E (apoE), one of the HDL constituent proteins, serves as a ligand for the low-density lipoprotein (LDL) receptor. Thus, it is considered that biocompatible delivery vehicles targeting LDL receptors could be prepared by incorporating apoE as the protein component of lipid nanodisks. To enhance targeting efficiency, we designed lipid nanodisks with a large number of ligands using a peptide with the LDL receptor-binding region of apoE combined with a high lipid affinity sequence (LpA peptide). In our study, the LpA peptide spontaneously formed discoidal complexes (LpA nanodisks) of approximately 10 nm in size, equivalent to native HDL. LpA peptides on nanodisks adopted highly α-helical structures, a competent conformation capable of interacting with LDL receptors. As anticipated, the uptake of LpA nanodisks into LDL receptor-expressing cells (HepG2) was higher than that of apoE nanodisks, suggesting an enhanced targeting efficiency via the enrichment of LDL receptor-binding regions on the particle. Biodistribution studies using 111In-labeled LpA nanodisks showed little splenic accumulation and prolonged retention in blood circulation, reflecting the biocompatibility of LpA nanodisks. High accumulation of 111In-labeled LpA nanodisks was observed in the liver as well as in implanted tumors, which abundantly express LDL receptors. Thus, LpA nanodisks are potential biocompatible delivery vehicles targeting LDL receptors.

Alendronate-Anionic Clay Nanohybrid for Enhanced Osteogenic Proliferation and Differentiation.

BACKGROUND: Alendronate (AL), a drug for inhibiting osteoclast-mediated bone-resorption, was intercalated into an inorganic drug delivery nanovehicle, layered double hydroxide (LDH), to form a new nanohybrid, AL-LDH, with 1:1 heterostructure along the crystallographic C-axis. Based on the intercalation reaction strategy, the present AL-LDH drug delivery system (DDS) was realized with an enhanced drug efficacy of AL, which was confirmed by the improved proliferation and osteogenic differentiation of osteoblast-like cells (MG63). METHODS: The AL-LDH nanohybrid was synthesized by conventional ion-exchange reaction and characterized by powder X-ray diffraction (PXRD), high-resolution transmission electron microscopy (HR-TEM), and Fourier transform infrared (FT-IR) spectroscopy. Additionally, in vitro efficacy tests, such as cell proliferation and alkaline phosphatase (ALP) activity, were analyzed. RESULTS: The AL was successfully intercalated into LDH via ion-exchange reaction, and thus prepared AL-LDH DDS was X-ray single phasic and chemically well defined. The accumulated AL content in MG63 cells treated with the AL-LDH DDS nanoparticles was determined to be 10.6-fold higher than that within those treated with the intact AL after incubation for 1 hour, suggesting that intercellular permeation of AL was facilitated thanks to the hybridization with drug delivery vehicle, LDH. Furthermore, both in vitro proliferation level and ALP activity of MG63 treated with the present hybrid drug, AL-LDH, were found to be much more enhanced than those treated with the intact AL. This is surely due to the fact that LDH could deliver AL drug very efficiently, although LDH itself does not show any effect on proliferation and osteogenic differentiation of MG63 cells. CONCLUSION: The present AL-LDH could be considered as a promising DDS for improving efficacy of AL.

Injectable Systems for Intra-Articular Delivery of Mesenchymal Stromal Cells for Cartilage Treatment: A Systematic Review of Preclinical and Clinical Evidence.

Stem cell-based therapy is a promising approach to treat cartilage lesions and clinical benefits have been reported in a number of studies. However, the efficacy of cell injection procedures may be impaired by cell manipulation and damage as well as by cell dissemination to non-target tissues. To overcome such issues, mesenchymal stromal cell (MSC) delivery may be performed using injectable vehicles as containment systems that further provide a favorable cell microenvironment. The aim of this systematic review was to analyze the preclinical and clinical literature on platelet-rich plasma (PRP), hyaluronic acid (HA), and hydrogels for the delivery of MSCs. The systematic literature search was performed using the PubMed and Web of science databases with the following string: "(stem cells injection) AND (platelet rich plasma OR PRP OR platelet concentrate OR biomaterials OR hyaluronic acid OR hydrogels)": 40 studies (19 preclinical and 21 clinical) met the inclusion criteria. This review revealed an increasing interest on the use of injectable agents for MSC delivery. However, while negligible adverse events and promising clinical outcomes were generally reported, the prevalence of low quality studies hinders the possibility to demonstrate the real benefits of using such injectable systems. Specific studies must be designed to clearly demonstrate the added benefits of these systems to deliver MSCs for the treatment of cartilage lesions and osteoarthritis.

Drug Delivery Nanoparticles with Locally Tunable Toxicity Made Entirely from a Light-Activatable Prodrug of Doxorubicin.

PURPOSE: A major challenge facing nanoparticle-based delivery of chemotherapy agents is the natural and unavoidable accumulation of these particles in healthy tissue resulting in local toxicity and dose-limiting side effects. To address this issue, we have designed and characterized a new prodrug nanoparticle with controllable toxicity allowing a locally-delivered light trigger to convert the payload of the particle from a low to a high toxicity state. METHODS: The nanoparticles are created entirely from light-activatable prodrug molecules using a nanoprecipitation process. The prodrug is a conjugate of doxorubicin and photocleavable biotin (DOX-PCB). RESULTS: These DOX-PCB nanoparticles are 30 times less toxic to cells than doxorubicin, but can be activated to release pure therapeutic doxorubicin when exposed to 365 nm light. These nanoparticles have an average diameter of around 100 nm and achieve the maximum possible prodrug loading capacity since no support structure or coating is required to prevent loss of prodrug from the nanoparticle. CONCLUSIONS: These light activatable nanoparticles demonstrate tunable toxicity and can be used to facilitate future therapy development whereby light delivered specifically to the tumor tissue would locally convert the nanoparticles to doxorubicin while leaving nanoparticles accumulated in healthy tissue in the less toxic prodrug form.