Immune Cell-Mediated Biodegradable Theranostic Nanoparticles for Melanoma Targeting and Drug Delivery.

Although tremendous efforts have been made on targeted drug delivery systems, current therapy outcomes still suffer from low circulating time and limited targeting efficiency. The integration of cell-mediated drug delivery and theranostic nanomedicine can potentially improve cancer management in both therapeutic and diagnostic applications. By taking advantage of innate immune cell's ability to target tumor cells, the authors develop a novel drug delivery system by using macrophages as both nanoparticle (NP) carriers and navigators to achieve cancer-specific drug delivery. Theranostic NPs are fabricated from a unique polymer, biodegradable photoluminescent poly (lactic acid) (BPLP-PLA), which possesses strong fluorescence, biodegradability, and cytocompatibility. In order to minimize the toxicity of cancer drugs to immune cells and other healthy cells, an anti-BRAF V600E mutant melanoma specific drug (PLX4032) is loaded into BPLP-PLA nanoparticles. Muramyl tripeptide is also conjugated onto the nanoparticles to improve the nanoparticle loading efficiency. The resulting nanoparticles are internalized within macrophages, which are tracked via the intrinsic fluorescence of BPLP-PLA. Macrophages carrying nanoparticles deliver drugs to melanoma cells via cell-cell binding. Pharmacological studies also indicate that the PLX4032 loaded nanoparticles effectively kill melanoma cells. The "self-powered" immune cell-mediated drug delivery system demonstrates a potentially significant advancement in targeted theranostic cancer nanotechnologies.

Effects of Y-27632 on the osteogenic and adipogenic potential of human dental pulp stem cells in vitro.

BACKGROUND: Human dental pulp stem cells (hDPSCs) possess mesenchymal stem cell properties, originating from migrating neural crest cells. hDPSCs have received extensive attention in the field of tissue engineering and regenerative medicine due to their accessibility and ability to differentiate in several cell phenotypes. In this study, we cultured hDPSCs with Y-27632 to observe their biological behaviors changes. METHODS: The hDPSCs were separately cultured with Y-27632 (0, 0.156, 0.312, 0.625, 1.25, 2.50, 5, 10, 20, 40 µm) for 24, 48, 72 h to select the suitable concentration and time using CCK-8. Then, the hDPSCs were cultured with 2.50 µm Y-27632 for 48 h to analyzed the biological behaviors changes by 5-Ethynyl-2'-deoxyuridine (EdU), plate cloning, transwell, scratch, and Annexin V FITC/PI assays, separately. Additionally, osteogenic calcium nodules and lipid droplets were analyzed using alizarin red staining and oil red O staining, respectively. qRT-PCR was used to analyze the expression of osteogenesis, adipogenesis, stemness maintenance, and inflammation related genes. RESULTS: The hDPSCs proliferation was significantly enhanced after cultured with 2.50 µm Y-27632 for 48 h, but there was no significant difference in migration and apoptosis. Observation of alkaline phosphatase (ALP) activity, osteogenic and adipogenic differentiation abilities of hDPSCs, Y-27632 treatment clearly decreased the ALP activity and osteogenic differentiation ability, increased the adipogenic differentiation ability. Furthermore, Y-27632 decreased the CD73, CD90, CD105, CD166, TLR4, and NF-κB p65 genes expression, but increased the IL-8 gene expression. CONCLUSIONS: The biological behaviors of hDPSCs could be changed when they cultured with Y-27632.

Isolation and Culture of Primary Human Gingival Epithelial Cells using Y-27632.

The gingival tissue is the first structure that protects periodontal tissues and plays meaningful roles in many oral functions. The gingival epithelium is an important structure of gingival tissue, especially in the repair and regeneration of periodontal tissue. Studying the functions of gingival epithelial cells has crucial scientific value, such as repairing oral defects and detecting the compatibility of biomaterials. As human gingival epithelial cells are highly differentiated keratinized cells, their lifespan is short, and they are difficult to passage. So far, there are only two ways to isolate and culture gingival epithelial cells, a direct explant method and an enzymatic method. However, the time required to obtain epithelial cells using the direct explant method is longer, and the cell survival rate of the enzymatic method is lower. Clinically, the acquisition of gingival tissue is limited, so a stable, efficient, and simple in vitro isolation and culture system is needed. We improved the traditional enzymatic method by adding Y-27632, a Rho-associated kinase (ROCK) inhibitor, which can selectively promote the growth of epithelial cells. Our modified enzymatic method simplifies the steps of the traditional enzymatic method and increases the efficiency of culturing epithelial cells, which has significant advantages over the direct explant method and the enzymatic method.

Synthesis and characterization of citrate-based fluorescent small molecules and biodegradable polymers.

Novel citric acid based photoluminescent dyes and biodegradable polymers are synthesized via a facile "one-pot" reaction. A comprehensive understanding of the fluorescence mechanisms of the resulting citric acid-based fluorophores is reported. Two distinct types of fluorophores are identified: a thiozolopyridine family with high quantum yield, long lifetime, and exceptional photostability, and a dioxopyridine family with relatively lower quantum yield, multiple lifetimes, and solvent-dependent band shifting behavior. Applications in molecular labeling and cell imaging were demonstrated. The above discoveries contribute to the field of fluorescence chemistry and have laid a solid foundation for further development of new fluorophores and materials that show promise in a diversity of fluorescence-based applications. STATEMENT OF SIGNIFICANCE: Photoluminescent materials are pivotal for fluorescence based imaging, labeling and sensing applications. Understanding their fluorescence mechanism is challenging and imperative. We develop a new class of citric acid-derived fluorescent materials in forms of polymers and small molecular dyes by a one-step solvent free reaction. We discovered two different classes of citric acid-derived fluorophores. A two-ring thiozolopyridine structure demonstrates strong fluorescence and exceptional resistance to photo-bleaching. A one-ring dioxopyridine exhibits relative weak fluorescence but with intriguing excitation and solvent-dependent emission wavelength shifting. Our methodology of synthesizing citric acid-derived fluorophores and the understanding on their luminescence are instrumental to the design and production of a large number of new photoluminescent materials for biological and biomedical applications.

Synthesis and characterization of biocompatible antimicrobial N-halamine-functionalized titanium dioxide core-shell nanoparticles.

As one of the most powerful biocides, N-halamine based antimicrobial materials have attracted much interest due to their non-toxicity, rechargeability, and rapid inactivation against a broad range of microorganisms. In this study, novel titanium dioxide-ADMH core-shell nanoparticles [TiO2@poly (ADMH-co-MMA) NPs] were prepared via miniemulsion polymerization using 3-allyl-5,5-dimethylhydantoin (ADMH) and methyl methacrylate (MMA) with nano-TiO2. The produced nanoparticles were characterized by FT-IR, TEM, TGA, and XPS. The UV stability of N-halamine nanoparticles has been improved with the addition of titanium dioxide. After chlorination treatment by sodium hypochlorite, biocidal efficacies of the chlorinated nanoparticles against S. aureus (ATCC 6538) and E. coli O157:H7 (ATCC 43895) were determined. The nanoparticles showed excellent antimicrobial properties against bacteria within brief contact time. In addition, in vitro cell cytocompatibility tests showed that the antibacterial nanoparticles had good biocompatibility.

Fluorescence imaging enabled poly(lactide-co-glycolide).

Fluorescent biomaterials have attracted significant research efforts in the past decades. Herein, we report a new series of biodegradable, fluorescence imaging-enabled copolymers, biodegradable photoluminescent poly(lactide-co-glycolide) (BPLP-co-PLGA). Photoluminescence characterization shows that BPLP-co-PLGA solutions, films and nanoparticles all exhibit strong, tunable and stable photoluminescence. By adjusting the molar ratios of L-lactide (LA)/glycolide (GA) and (LA+GA)/BPLP, full degradation of BPLP-co-PLGA can be achieved in 8-16 weeks. The fluorescence decay behavior of BPLP-co-PLGA can be used for non-invasive monitoring of material degradation. In vitro cytotoxicity and in vivo foreign body response evaluations demonstrate that BPLP-co-PLGA exhibits similar biocompatibility to poly(lactide-co-glycolide) (PLGA). The imaging-enabled BPLP-co-PLGA was fabricated into porous scaffolds whose degradation can be monitored through non-invasive imaging and nanoparticles that show theranostic potential demonstrated by fluorescent cellular labeling, imaging and sustained 5-fluorouracil delivery. The development of inherently fluorescent PLGA copolymers is expected to impact the use of already widely accepted PLGA polymers for applications where fluorescent properties are highly desired but limited by the conventional use of cytotoxic quantum dots and photobleaching organic dyes. STATEMENT OF SIGNIFICANCE: This manuscript describes a novel strategy of conferring intrinsic photoluminescence to the widely used biodegradable polymers, poly(lactide-co-glycolide) without introducing any cytotoxic quantum dots or photo-bleaching organic dyes, which may greatly expand the applications of these polymers in where fluorescent properties are highly desired. Given the already significant impact generated by the use of PLGA and alike, this work contributes to fluorescence chemistry and new functional biomaterial design and will potentially generate significant impact on many fields of applications such as tissue engineering, molecular imaging and labeling, and drug delivery.

Design of antimicrobial peptides conjugated biodegradable citric acid derived hydrogels for wound healing.

Wound healing is usually facilitated by the use of a wound dressing that can be easily applied to cover the wound bed, maintain moisture, and avoid bacterial infection. In order to meet all of these requirements, we developed an in situ forming biodegradable hydrogel (iFBH) system composed of a newly developed combination of biodegradable poly(ethylene glycol) maleate citrate (PEGMC) and poly(ethylene glycol) diacrylate (PEGDA). The in situ forming hydrogel systems are able to conform to the wound shape in order to cover the wound completely and prevent bacterial invasion. A 2(k) factorial analysis was performed to examine the effects of polymer composition on specific properties, including the curing time, Young's modulus, swelling ratio, and degradation rate. An optimized iFBH formulation was achieved from the systematic factorial analysis. Further, in vitro biocompatibility studies using adult human dermal fibroblasts (HDFs) confirmed that the hydrogels and degradation products are not cytotoxic. The iFBH wound dressing was conjugated and functionalized with antimicrobial peptides as well. Evaluation against bacteria both in vitro and in vivo in rats demonstrated that the peptide-incorporated iFBH wound dressing offered excellent bacteria inhibition and promoted wound healing. These studies indicated that our in situ forming antimicrobial biodegradable hydrogel system is a promising candidate for wound treatment.

Click chemistry plays a dual role in biodegradable polymer design.

Click chemistry plays a dual role in the design of new citrate-based biodegradable elastomers (CABEs) with greatly improved mechanical strength and easily clickable surfaces for biofunctionalization. This novel chemistry modification strategy is applicable to a number of different types of polymers for improved mechanical properties and biofunctionality.

Development of intrinsically photoluminescent and photostable polylactones.

A method of introducing intrinsically photo luminescent properties to biodegradable polymer is introduced, exemplified by the synthesis of intrinsically photoluminescent polylactones that enable non-invasive monitoring and tracking of material degradation in vivo in realtime, as well as the formation of theranostic nanoparticles for cancer imaging and drug delivery.

Synthesis and characterization of biomimetic citrate-based biodegradable composites.

Natural bone apatite crystals, which mediate the development and regulate the load-bearing function of bone, have recently been associated with strongly bound citrate molecules. However, such understanding has not been translated into bone biomaterial design and osteoblast cell culture. In this work, we have developed a new class of biodegradable, mechanically strong, and biocompatible citrate-based polymer blends (CBPBs), which offer enhanced hydroxyapatite binding to produce more biomimetic composites (CBPBHAs) for orthopedic applications. CBPBHAs consist of the newly developed osteoconductive citrate-presenting biodegradable polymers, crosslinked urethane-doped polyester and poly (octanediol citrate), which can be composited with up to 65 wt % hydroxyapatite. CBPBHA networks produced materials with a compressive strength of 116.23 ± 5.37 MPa comparable to human cortical bone (100-230 MPa), and increased C2C12 osterix gene and alkaline phosphatase gene expression in vitro. The promising results above prompted an investigation on the role of citrate supplementation in culture medium for osteoblast culture, which showed that exogenous citrate supplemented into media accelerated the in vitro phenotype progression of MG-63 osteoblasts. After 6 weeks of implantation in a rabbit lateral femoral condyle defect model, CBPBHA composites elicited minimal fibrous tissue encapsulation and were well integrated with the surrounding bone tissues. The development of citrate-presenting CBPBHA biomaterials and preliminary studies revealing the effects of free exogenous citrate on osteoblast culture shows the potential of citrate biomaterials to bridge the gap in orthopedic biomaterial design and osteoblast cell culture in that the role of citrate molecules has previously been overlooked.

[Analysis of the factors for predicting the outcomes of interferon-α and entecavir treatments for chronic hepatitis B with positive HBeAg].

OBJECTIVE: To analyze the predictive factors of the therapeutic effects of interferons (IFNs) and entecavir (ETV) treatments for 48 weeks in patients with chronic hepatitis B (CHB) positive for HBeAg. METHODS: This retrospective analysis compared the treatment efficacy of IFNs and ETV in 129 CHB patients positive for HBeAg. Twenty-seven of the patients were treated with PEG-IFNα-2a (180 µg once a week, PEG-IFN group), 51 patients with conventional IFNα (5 MIU three times a week, IFN group), and 51 with ETV (0.5 mg once daily, ETV group) for 48 weeks. RESULTS: After completion of the treatment cycles, the patients in ETV group showed a significantly higher HBV DNA undetectable rate and a significantly lower HBeAg seroconversion rate than those in PEG-IFN and IFN groups (P<0.05); HBeAg seroconversion rates were similar between PEG-IFN group and IFN group (Χ(2)=0.709, P=0.400). In PEG-IFN and ETV groups, HBeAg seroconversion rates were not associated with age, gender, baseline HBeAg, baseline HBV DNA and baseline ALT. In IFN group, HBeAg seroconversion rates were associated with baseline HBeAg (P=0.048) but not with age, gender, baseline HBV DNA and baseline ALT. In PEG-IFNα-2a group, ROC analysis showed that the sensitivity and specificity of HBeAg seroconversion at 48 weeks were 0.778 and 0.889, respectively, when the decline rate of HBeAg between baseline and week 24 exceeded 97.81%, with the corresponding positive and negative predictive values (PPV and NPV) of 0.778 and 0.889, respectively; the sensitivity and specificity of HBeAg seroconversion at 48 weeks were 0.889 and 0.722, respectively, when the decline rate of HBeAg between week 12 and week 24 was over 42.75%, with the corresponding PPV and NPV of 0.615 and 0.929, respectively. CONCLUSION: Treatments with PEG-IFNα-2a and conventional IFNα for 48 weeks can achieve a higher HBeAg seroconversion rate than ETV, but the latter produces a higher HBV DNA undetectable rate. For PEG-IFNα-2a treatment, the decline rate of HBeAg between baseline and week 24 over 97.81% is the best predicting factor for HBeAg seroconversion at week 48 in CHB patients positive for HBeAg.

Dual growth factor releasing multi-functional nanofibers for wound healing.

The objective of this research is to develop a dual growth factor-releasing nanoparticle-in-nanofiber system for wound healing applications. In order to mimic and promote the natural healing procedure, chitosan and poly(ethylene oxide) were electrospun into nanofibrous meshes as mimics of extracellular matrix. Vascular endothelial growth factor (VEGF) was loaded within nanofibers to promote angiogenesis in the short term. In addition, platelet-derived growth factor-BB (PDGF-BB) encapsulated poly(lactic-co-glycolic acid) nanoparticles were embedded inside nanofibers to generate a sustained release of PDGF-BB for accelerated tissue regeneration and remodeling. In vitro studies revealed that our nanofibrous composites delivered VEGF quickly and PDGF-BB in a relayed manner, supported fibroblast growth and exhibited anti-bacterial activities. A preliminary in vivo study performed on normal full thickness rat skin wound models demonstrated that nanofiber/nanoparticle scaffolds significantly accelerated the wound healing process by promoting angiogenesis, increasing re-epithelialization and controlling granulation tissue formation. For later stages of healing, evidence also showed quicker collagen deposition and earlier remodeling of the injured site to achieve a faster full regeneration of skin compared to the commercial Hydrofera Blue® wound dressing. These results suggest that our nanoparticle-in-nanofiber system could provide a promising treatment for normal and chronic wound healing.

[Anthropometric analysis of the mandible morphology in young females with different vertical skeletal pattern].

OBJECTIVE: The purpose of this study is to assess the height and width of mandibular cross-sections and the thickness of cortical bone for young females in different vertical skeletal pattern by cone beam CT (CBCT) and determine the characteristics and differences of mandible among different vertical skeletal patterns. METHODS: A total of 64 females, aged from 19 to 40 years old, were enrolled from Department of Orthodontics of School of Stomatology of Shandong University. After examination with CBCT scan, they were divided into 3 groups according to the vertical skeletal patterns (14 in low-angle group, 31 in average-angle group and 19 in high-angle group). The mandibular morphology was evaluated according to the height and width of the mandibular cross-sections and the thickness of cortices. SPSS 16.0 was used to analyze the characteristics and differences of mandible among different vertical skeletal patterns. RESULTS: The height of symphysis in the high-angle group was higher than that in the low-angle group, but the height of molars in the low-angle group was higher than that in the high-angle group (P<0.05). In the regions of anterior teeth and premolars, the width of upper one third of the mandibular cross-sections in the low-angle group was significantly larger than those in the high-angle group, while in the symphysis and lateral incisors regions, the width of lower one third of the mandibular cross-sections in the low-angle group was larger than those in the high-angle group (P<0.05). Except of the region of premolars, the cortical thickness of base in the low-angle group was significant thicker than those in the high-angle group (P<0.05). In the regions of canine, premolar and molar, the cortical thickness of buccal upper one third in the low-group was thicker than those in the high-angle group, but in the regions of canine and the second molar, the cortical thickness of buccal lower one third in the low-angle group was thi- cker than those in the high-angle group (P<0.05). In the regions of the symphysis, the cortical thickness of lingual lower one third in the low-angle group was thicker than those in the high-angle group (P<0.05), but in the regions of premolar and molar of lingual upper one third in the low-angle group was thicker than those in the high-angle group (P<0.05). CONCLUSION: The height and width of the cross-sections of the body of mandible differed more than thickness of cortical bone among the three different vertical skeletal patterns. The height of cross-sections decreases gradually from the anterior region to the posterior region in three groups. The width of cross-sections in the low-group is thicker than those in other two groups. The thickness of cortical bones decreases successively from low, average to high angle group.

Functionalized poly(L-lactide) nanoparticles from electrospun nanofibers.

Crystalline poly(L-lactide) (PLLA) nanoparticles were prepared by aminolysis of electrospun PLLA nanofibers and subsequently labeled by a fluorescent colorant. The size of the nanoparticles could be controlled by either the conditions of the aminolysis reaction or the diameter of the original electrospun fiber. The latter method resulted in higher yield. Although the as-spun nanofibers were generally amorphous, the nanoparticles showed high crystallinity in the typical α-form of PLLA crystals, as revealed by X-ray diffraction and differential scanning calorimetry. Gel-permeation chromatography was used to determine the molecular weight of the formed particles in relation to aminolysis time. After aminolysis, PLLA nanoparticles spontaneously generated amine groups on the surface, which are available for further modifications. In this study the amine groups were reacted with isothiocyanate groups, and fluorescein-5'-isothiocyanate was successfully attached to the PLLA nanoparticles. Smaller particles showed significantly higher fluorescein binding density. Through this simple 'top-down' routine, it was possible to create nanoparticles with tailorable size and specific surface functions. Such materials could serve in bioimaging or nanomedicine applications.