Establishment of a bi-layered tissue engineered conjunctiva using a 3D-printed melt electrowritten poly-(ε-caprolactone) scaffold.

PURPOSE: To utilize melt electrowriting (MEW) technology using poly-(ε-caprolactone) (PCL) coupled with a 2-step co-culturing strategy for the development of a conjunctival bi-layer synthetic construct. METHODS: Melt electrowritten scaffolds using PCL were fabricated using an in-house-built MEW printer. Human conjunctival stromal cells (CjSCs) and epithelial cells (CjECs) were isolated from donor tissue. A 2-step co-culture method was done by first seeding the CjSCs and culturing for 4 weeks to establish a stromal layer, followed by CjECs and co-culturing for 2 more weeks. Cultured cells were each characterized by morphology and marker expression on immunofluorescence and qPCR. The produced construct was assessed for cellular proliferation using viability assays. The bi-layer morphology was assessed using scanning electron microscopy (SEM), confocal microscopy, and immunofluorescence imaging. The expression of extracellular matrix components and TGF-b was evaluated using qPCR. RESULTS: CjSCs were spindle-shaped and vimentin + while CjECs were polygonal and CK13 + . CjSCs showed consistent proliferation and optimal adherence with the scaffold at the 4-week culture mark. A 2-layered construct consisting of a CjSC-composed stromal layer and a CjEC-composed epithelial layer was appreciated on confocal microscopy, SEM, and immunofluorescence. CjSCs secreted collagens (types I, V, VI) but at differing amounts from natural tissue while TGF-b production was comparable. CONCLUSION: The 3D-printed melt electrowritten PCL scaffold paired with the 2-step co-culturing conditions of the scaffold allowed for the first approximation of a bi-layered stromal and epithelial reconstruction of the conjunctiva that can potentially improve the therapeutic arsenal in ocular surface reconstruction.

A new strategy to sustained release of ocular drugs by one-step drug-loaded microcapsule manufacturing in hydrogel punctal plugs.

PURPOSE: To design an injectable hyaluronate (HA)-based hydrogel system that contains drug-loaded microcapsules as resorbable plugs to deliver ocular drugs. METHODS: In-situ drug-loaded, core-shell-structured chitosan (CS)@HA microcapsules were fabricated via HA hydrosol collecting in electrospun bead-rich CS fibers under continuous stirring. An injectable and cytocompatible hydrogel system with different degrees of chemical crosslinking maintained viscoelastic and sustained drug release for a long-term period of time at body temperature in vitro. RESULTS: With the addition of adipic dihydrazide (ADH) or 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide hydrochloride (EDCI), HA hydrosols transited from liquid to solid state at the gel point, with the G'/G″ ratio varying between 1.43 and 5.32 as a function of crosslinker concentration in the hydrogel phase. Ofloxacin (OFL) release from the mechanically mixed hydrosol system (CS-HA-A0-E0) and the micro-encapsulated hydrosol formulation (CS@HA-A0-E0) were respectively over 80% and 51% of the total drug load leaching out within 24 h. As for the drug-mixed hydrogel systems with low (CS-HA-A0.06-E0.15) and high (CS-HA-A0.06-E0.30) crosslinking density, the OFL release rate reached 38.5 and 46.6% respectively, while the micro-encapsulated hydrogel systems with low (CS@HA-A0.06-E0.15) and high (CS@HA-A0.6-E0.30) showed only (11.9 ± 2.7)% and (17.4 ± 3.5)% drug release respectively. CONCLUSIONS: A one-step in-situ drug-capsulizing method is developed to fabricate a resorbable hydrogel punctal plug with extended drug release. The chemistry of the crosslinking reaction involves the formation of highly biocompatible HA derivatives. Thus, the hydrogel can be used directly in the tear drainage canalicular system.

A biomimetic collagen/heparin multi-layered porous hydroxyapatite orbital implant for in vivo vascularization studies on the chicken chorioallantoic membrane.

BACKGROUND: The vascularization of an orbital implant is a key issue for reducing complications, such as exposure and infection. METHODS: Here, we developed a facile layer-by-layer assembly approach to modify porous hydroxyapatite (pHA) orbital implants with five collagen (COL)/heparin (HEP) multilayers. RESULTS: SEM characterization showed that the average pore size of the pHA/(COL/HEP)5 scaffold was 316.8 ± 77.1 µm. After being coated with five COL/HEP multilayers, the mechanical strength was improved compared with that of the pHA scaffolds. The in vitro assay displayed that the pHA scaffolds covered with COL/HEP multilayers resulted in a larger number of human umbilical vein endothelial cells after being cultured for 14 days. The macroscopic evaluation and semi-quantitative vascular density analysis of the chicken chorioallantoic membrane assay showed that the pHA/(COL/HEP)5 scaffolds resulted in more intense angiogenesis than the pHA scaffolds. CONCLUSIONS: These studies demonstrate that the biomembrane-mimicking coating of COL/HEP multilayers is a simple and effective strategy to endow combined biological performances of pHA orbital implants and to potentially reduce implant-related complications.

An antibacterial and anti-oxidant hydrogel containing hyperbranched poly-l-lysine and tea polyphenols accelerates healing of infected wound.

Both bacteria-infection and excessive inflammation delay the wound healing process and even create non-healing wound, thus it is highly desirable to endow the wound dressing with bactericidal and anti-oxidation properties. Herein an antibacterial and antioxidation hydrogel based on Carbomer 940 (CBM) and hydroxypropyl methyl cellulose (HPMC) loaded with tea polyphenols (TP) and hyperbranched poly-l-lysine (HBPL) was designed and fabricated. The hydrogel killed 99.9 % of methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli (E. coli) at 107 CFU mL-1, and showed strong antioxidation against H2O2 and 2,2-di(4-tert-octylphenyl)-1-picryl-hydrazyl (DPPH) radicals without noticeable cytotoxicity in vitro. The CBM/HPMC/HBPL/TP hydrogel significantly shortened the inflammatory period of the MRSA-infected full-thickness skin wound of rats in vivo, with 2 orders of lower MRSA colonies compared with the blank control, and promoted the wound closure especially at the earlier stage. The inflammation was suppressed and the vascularization was promoted significantly as well, resulting in reduced pro-inflammatory factors including interleukin-6 (IL-6), interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α), and increased anti-inflammatory factors such as interleukin-4 (IL-4) and interleukin-10 (IL-10).

An electrospun scaffold functionalized with a ROS-scavenging hydrogel stimulates ocular wound healing.

Ocular alkali burn is a serious ophthalmic emergency. Highly penetrative alkalis cause strong inflammatory responses leading to persistent epithelial defects, acute corneal perforation and severe scarring, and thereby persistent pain, loss of vision and cicatricial sequelae. Early and effective anti-inflammation management is vital in reducing the severity of injury. In this study, a double network biomaterial was prepared by compounding electrospinning nanofibres of thioketal-containing polyurethane (PUTK) with a reactive oxygen species (ROS)-scavenging hydrogel (RH) fabricated by crosslinking poly(poly(ethylene glycol) methyl ether methacrylate-co-glycidyl methacrylate) with thioketal diamine and 3,3'-dithiobis(propionohydrazide). The developed PUTK/RH patch exhibited good transparency, high tensile strength and increased hydrophilicity. Most importantly, it demonstrated strong antioxidant activity against H2O2 and 2,2-di(4-tert-octylphenyl)-1-picryl-hydrazyl (DPPH). Next, a rat corneal alkali burn model was established, and the PUTK/RH patch was transplanted on the injured cornea. Reduced inflammatory cell infiltration was revealed by confocal microscopy, and lower expression levels of genes relative to inflammation, vascularization and scarring were identified by qRT-PCR and western blot. Fluorescein sodium dyeing, hematoxylin and eosin (H&E) staining and immunohistochemical staining confirmed that the PUTK/RH patch could accelerate corneal wound healing by inhibiting inflammation, promoting epithelial regeneration and decreasing scar formation. STATEMENT OF SIGNIFICANCE: Ocular alkali burn is a serious ophthalmic emergency, characterized with persistent inflammation and irreversible vision loss. Oxidative stress is the main pathological process at the acute inflammatory stage, during which combined use of glucocorticoids and amniotic membrane transplantation is the most widely accepted treatment. In this study, we fabricated a polyurethane electrospun nanofiber membrane functionalized with a ROS-scavenging hydrogel. This composite patch could be a promising amniotic membrane substitute, possessing with a transparent appearance, elasticity and anti-inflammation effect. It could be easily transplanted onto the alkali-burned corneas, resulting in a significant inhibition of stromal inflammation and accelerating the recovery of corneal transparency. The conception of ROS-scavenging wound patch may offer a new way for ocular alkali burn.

Cytoprotective effect of hyaluronic acid and hydroxypropyl methylcellulose against DNA damage induced by thimerosal in Chang conjunctival cells.

BACKGROUND: To investigate genotoxicity of the preservative thimerosal (Thi), and the cytoprotective and antioxidant effects of hyaluronic Acid (HA) and hydroxypropyl methylcellulose (HPMC) on Chang conjunctival cells. METHOD: Cells were divided into three groups. One group was exposed to Thi at various concentrations (0.00001 %∼0.001 %) for 30 min; the other two groups were pretreated with 0.3 % HA or 0.3 % HPMC for 30 min before the Thi exposure. After cell viability was evaluated, alkaline comet assay and detection of the phosphorylated form of the histone variant H2AX (γH2AX) foci were used to determine DNA damage. Reactive oxygen species (ROS) production was assessed by the fluorescent probe, 2', 7'-dichlorodihydrofluorescein diacetate (DCFH-DA). RESULTS: A significant change of cell viability was observed after exposure to 0.001 % Thi for 30 min. DNA single- and double-strand breaks were significantly increased in a dose-dependent manner with Thi exposure. In addition, intracellular ROS induced by Thi was dose-dependent, except at 0.001 % less ROS was induced than at 0.0005 %. However, cells pretreated with 0.3 % HA or 0.3 % HPMC showed significantly increased cell survival, decreased DNA damage, and decreased ROS production compared to cells exposed to Thi alone. Pretreatment with 0.3 % HA was found to be even more protective than 0.3 % HPMC. CONCLUSION: Thi can induce DNA damage in human conjunctival epithelial cells, probably due to oxidative stress. HA and HPMC are protective agents that have antioxidant properties and can decrease DNA damage induced by Thi. Pretreatment of 0.3 % HA may be more protective of the ocular surface than 0.3 % HPMC.

Precisely Tuning the Pore-Wall Surface Composition of Bioceramic Scaffolds Facilitates Angiogenesis and Orbital Bone Defect Repair.

Orbital bone damage (OBD) may result in severe post-traumatic enophthalmos, craniomaxillofacial deformities, vision loss, and intracranial infections. However, it is still a challenge to fabricate advanced biomaterials that can match the individual anatomical structure and enhance OBD repair in situ. Herein, we aimed to develop a selective surface modification strategy on bioceramic scaffolds and evaluated the effects of inorganic or organic functional coating on angiogenesis and osteogenesis, ectopically and orthotopically in OBD models. It was shown that the low thermal bioactive glass (BG) modification or layer-by-layer assembly of a biomimetic hydrogel (Biogel) could readily integrate into the pore wall of the bioceramic scaffolds. The BG and Biogel modification showed appreciable enhancement in the initial compressive strength (∼30-75%) or structural stability in vivo, respectively. BG modification could enhance by nearly 2-fold the vessel ingrowth, and the osteogenic capacity was also accelerated, accompanied with a mild scaffold biodegradation after 3 months. Meanwhile, the Biogel-modified scaffolds showed enhanced osteogenic differentiation and mineralization through calcium and phosphorus retention. The potential mechanism of the enhanced bone repair was elucidated via vascular and osteogenic cell responses in vitro, and the cell tests indicated that the Biogel and BG functional layers were both beneficial for in vitro osteoblastic differentiation and mineralization on bioceramics. Totally, these findings demonstrated that the bioactive ions or biomolecules could significantly improve the angiogenic and osteogenic capabilities of conventional bioceramics, and the integration of inorganic or organic functional coating in the pore wall is a highly flexible material toolbox that can be tailored directly to improve orbital bone defect repair.

Human epidermal growth factor-functionalized cocoon silk with improved cell proliferation activity for the fabrication of wound dressings.

Human epidermal growth factor (hEGF) is a key factor involved in wound healing owing to its powerful ability to stimulate cell proliferation. In this study, we used piggyBac transposon technology to produce transgenic silkworms expressing the hEGF protein fused to truncated heavy chain (FibH-hEGF). The FibH-hEGF fusion protein was successfully expressed and secreted into silkworm cocoons. Compared to wild-type silk, the transgenic silkworm silk had the similar morphology about silks fiber surface and cocoon nets, while the secondary structure between the transgenic silk and wild-type silk was different. Most importantly, transgenic silkworm cocoon silk powder extract significantly increased human fibroblast FIB cell proliferation for a long duration with no apparent cytotoxicity. Our study provides a promising method for obtaining cost-effective and functional biomaterials for the fabrication of wound dressings.

Facile fabrication of elastic, macro-porous, and fast vascularized silicone orbital implant.

Orbital implants with interconnected porous architecture had gained prominence, as they were capable of being colonized by fibrovascular tissue and minimizing complications. However, mechanical properties of orbital implant had received little attention among existing design philosophy. Herein, a compliant porous silicone scaffold was developed by gelatin porogen-leaching method and used as the orbital implant in this study. The silicone scaffolds exhibited desired microstructure and simulated mechanical properties, including high porosity of ~90%, suitable pore size of 280-450 µm, reduced modulus of 50.1 ± 11.7 KPa, and excellent elasticity. in vitro results showed that the porous silicone scaffolds did not exhibit noticeable cytotoxicity and were favorable for both adhesion and proliferation of human vascular ECs. The porous silicone scaffold was easy to be manipulated when implanted into the anophthalmic sockets of rabbits. The implanted scaffolds provided satisfactory volume replacement and induced extensive fibro-vascularization, showing desirable orbital reconstruction effects. Therefore, our novel porous silicone scaffolds may be promising substitutes for current orbital implants.

Tissue engineering of corneal stroma via melt electrowriting.

The cornea serves as the main refractive component of the eye with the corneal stroma constituting the thickest component in a stratified layered system of epithelia, stroma, and endothelium. Current treatment options for patients suffering from corneal diseases are limited to transplantation of a human donor cornea (keratoplasty) or to implantation of an artificial cornea (keratoprosthesis). Nevertheless, donor shortage and failure of artificial corneas to integrate with local tissue constitute important problems that have not been yet circumvented. Recent advances in biofabrication have made great progress toward the manufacture of tailored biomaterial templates with the potential of guiding partially or totally the regeneration process of the native cornea. However, the role of the corneal stroma on current tissue engineering strategies is often neglected. Here, we achieved a tissue-engineered corneal stroma substitute culturing primary keratocytes on scaffolds prepared via melt electrowriting (MEW). Scaffolds were designed to contain highly organized micrometric fibers to ensure transparency and encourage primary human keratocytes to self-orchestrate their own extracellular matrix deposition and remodeling. Results demonstrated reliable cell attachment and growth over a period of 5 weeks and confirmed the formation of a dense and highly organized de novo tissue containing collagen I, V, and VI as well as Keratocan, which resembled very closely the native corneal stoma. In summary, MEW brings us closer to the biofabrication of a viable corneal stroma substitute.

Regeneration of different types of tissues depends on the interplay of stem cells-laden constructs and microenvironments in vivo.

The ability of repair and regeneration of tissues or organs has been significantly improved by using biomaterials-based constructs. Our previous studies found the regeneration of both articular cartilage and subchondral bone by implantation of a poly(lactide-co-glycolide) (PLGA)/fibrin gel/bone marrow stem cells (BMSCs)/(lipofectamine/pDNA-transforming growth factor (TGF)-ß1) construct in vivo, without the step of pre-induced differentiation of the laden stem cells in vitro. To substantiate the ability to regenerate multi-types of tissues by the same constructs, in this study the constructs were implanted into three types of tissues or tissue defects in vivo, including subcutaneous fascia layer, and ear cartilage and eyelid tarsal plate defects. The ear cartilage and eyelid tarsal plate defects were fully regenerated 8 w post-implantation, showing a similar morphology to the corresponding native tissues. In the neo ear cartilage, abundant chondrocytes with obvious lacunas and cartilage-specific extracellular matrices (ECMs) were found. Neo eyelid tarsal plate with mature meibomian gland acinar units was regenerated. Furthermore, expressions of the ECMs-specific genes and proteins, as well as the cell behavior modulatory factors, Sry related HMG box 9 (Sox9) and TGF-ß1 were significantly up-regulated in the regenerated ear cartilages and eyelid tarsal plate than those in the subcutaneously implanted constructs, which were filled with fibrocytes, inflammatory cells, obvious vascularization and slight ECMs deposition. These results confirm firmly the ability to regenerate multi-types of tissues by a stem cells-laden construct via adapting to the microenvironments of corresponding tissues.

Unilateral Levator Aponeurosis Excision for Marcus Gunn Syndrome and Risk Factors of Residual Jaw Winking.

PURPOSE: To investigate the association of ptosis, levator, and jaw winking in Marcus Gunn jaw-winking synkinesis (MGJWS), and the risk factor of preservation and outcomes of the unilateral levator excision and frontalis suspension. METHODS: Clinical features of MGJWS case series from 2011 to 2018 were retrospectively reviewed. Association between jaw winking and ptosis/levator function was statistically analyzed. The patients underwent unilateral levator excision and frontalis suspension using silicone rod or autogenous fascia lata. Clinical outcomes were evaluated in operated patients and the independent risk factors of residual jaw winking were investigated after a long follow-up. RESULTS: There were 42 MGJWS patients in 2011 to 2018, accounting for 2.87% of all congenital blepharoptosis. 80% of mild jaw winking was accompanied with mild ptosis and fair levator function, and moderate-to-severe jaw winking was often accompanied with moderate-to-severe ptosis and poor levator function (P < 0.05). Ptosis showed a strong association with excursion of jaw winking (R = 0.785, P < 0.01). Jaw winking was resolved in all 34 operated patients with good correction of ptosis. Severity of jaw winking is an independent risk factor for the residual synkinesis after surgery. Severe preoperative jaw winking had an 18.05 times increased risk of postoperative residual synkinesis compared with moderate jaw winking (P < 0.05). CONCLUSIONS: In MGJWS eyelid excursion of jaw winking has a direct correlation with ptosis and dysfunction of levator muscle. Unilateral levator aponeurosis excision and frontalis suspension is an efficient approach for MGJWS. Severe jaw winking is a risk factor of residual eyelid synkinesis after surgery.

A biomimetic tarso-conjunctival biphasic scaffold for eyelid reconstruction in vivo.

Conventional 3D porous scaffolds used as tarsal plate substitute may cause corneal irritation and conjunctival mucoid discharge, and even lead to blindness and cicatricial blepharon deformities. In this study, collagen/chitosan (Col/CS) sponges with thickness of 240 µm, 466 µm, and 724 µm were composited onto poly(propylene fumarate)-co-2-hydroxyethyl methacrylate (PPF-HEMA) polymer networks to obtain the corresponding biphasic scaffolds, which simulate the natural anatomy of posterior lamella of eyelid. These three scaffolds exhibited a porous structure with porosity of ∼90%, simulated elastic modulus, appropriate degradation rate and good biocompatibility. Composited with Col/CS sponge of difference thickness, the scaffolds induced different cellular behaviors such as proliferation, distribution and stratification, by regulating the mechanical properties cells sensed as effective modulus. In a rabbit tarso-conjunctival defect model, the grafted biphasic scaffolds promoted re-epithelization with functional regenerated conjunctiva. Hence, the biphasic composite scaffolds may be a promising substitute for tarso-conjunctival repair.

[Effect of porous polyethylene channel implants on volume reconstruction of large orbital wall fractures evaluated by computed tomography measurement].

OBJECTIVE: To assess the clinical effectiveness of porous polyethylene medpor channel implants (MCI) to restore orbital volume in repairing orbital wall fractures, evaluated by computed tomography (CT) and volume measurement program. METHODS: Sixteen patients with unilateral large orbital fractures were included in this study. Computed tomography (CT) scans were used to obtain computer-based orbital volume measurement to assess the change in orbital volume pre- and post-operatively. Pre- and post-operative enophthalmos were measured using a Hertel exophthalmometer. The average time interval between injury and surgery was (17.4+/-10.0) days, and the mean follow-up period was 9 months. RESULTS: Postoperative CT scan showed that most of the MCI to be well positioned. The orbital volume of the injured orbit was significantly increased [mean: (28.16+/-4.32) cm3] as compared with unaffected orbit preoperatively (t=3.044, P<0.01). There was no statistical difference in orbital volume between the injured and normal orbits after orbital reconstruction (t=0.069, P>0.05). The orbital volume change after reconstructive surgery was significantly positively correlated with the decrease of enophthalmos (r=0.715, P<0.01). In order to resolve 2 mm enophthalmos, more than 2.86 cm3 orbital volume augmentation is recommended. CONCLUSIONS: Orbital volume measurement by CT scan is useful in the post-traumatic evaluation of severity of orbital fractures, and it can help predict the degree of late enophthalmos. Orbital reconstruction with the MCI, when indicated, is recommended.

The Natural Diterpenoid Isoforretin A Inhibits Thioredoxin-1 and Triggers Potent ROS-Mediated Antitumor Effects.

Aberrant expression of thioredoxin 1 (Trx1) plays an important role in cancer initiation and progression and has gained attention as an anticancer drug target. Here we report that the recently discovered natural diterpenoid isoforretin A (IsoA) significantly inhibits Trx1 activity and mediates anticancer effects in multiple preclinical settings. The inhibitory effect of IsoA was antagonized by free radical scavengers polyethylene glycol-catalase, polyethylene glycol superoxide dismutase, thiol-based antioxidants N-acetylcysteine and glutathione. Mass spectrometry analysis revealed that the mechanism of action was based on direct conjugation of IsoA to the Cys32/Cys35 residues of Trx1. This conjugation event attenuated reversible thiol reduction of Trx1, leading to ROS accumulation and a broader degradation of thiol redox homeostasis in cancer cells. Extending these in vitro findings, we documented that IsoA administration inhibited the growth of HepG2 tumors in a murine xenograft model of hepatocellular carcinoma. Taken together, our findings highlight IsoA as a potent bioactive inhibitor of Trx1 and a candidate anticancer natural product. Cancer Res; 77(4); 926-36. ©2016 AACR.

Evaluation of computer-based volume measurement and porous polyethylene channel implants in reconstruction of large orbital wall fractures.

PURPOSE: To describe the use of computer-based orbital volume measurement as a predictor of late enophthalmos, and to assess the effectiveness of the MedPor (Porex Surgical Products Group, Newnan, GA) porous polyethylene channel implant to restore orbital volume in repairing large orbital wall fractures. METHODS: Sixteen patients with unilateral large orbital fractures were included. Computed tomographic (CT) scans were used to obtain computer-based orbital volume measurement to predict the likelihood of late enophthalmos and to assess the change in orbital volume before and after surgery. The effectiveness of a channel implant was evaluated by the orbital volume and postoperative exophthalmetric measurement. RESULTS: The average time interval between injury and surgery was 17.4 +/- 10 days, and the mean follow-up was 9 months. The orbital volume of the injured orbit was significantly increased (mean, 4.22 +/- 2.61 cm2) compared with the unaffected orbit before surgery (t = 3.046, P = 0.005). There was not a significant difference in orbital volume between the two orbits after orbital reconstruction (t = 0.069, P = 0.945). The orbital volume change after reconstructive surgery was significantly positively correlated with the decrease of enophthalmos (r = 0.715, P = 0.001; enophthalmos [E] = 0.72; volume increment [V] = 0.06). To resolve 2 mm enophthalmos, more than 2.9 cm3 orbital volume augmentation is recommended for early reconstructive surgery. Postoperative CT scan showed most of the channel implants to be well positioned. CONCLUSIONS: Computer-based orbital volume measurement from a CT scan is useful in the posttraumatic evaluation of orbital fractures, and it can help predict the degree of late enophthalmos that can be expected. Orbital reconstruction with the MedPor channel implant (Porex Surgical Products Group), when indicated, is recommended, especially for large orbital wall fractures.

Liposomal C6 Ceramide Activates Protein Phosphatase 1 to Inhibit Melanoma Cells.

Melanoma is one common skin cancer. In the present study, the potential anti-melanoma activity by a liposomal C6 ceramide was tested in vitro. We showed that the liposomal C6 (ceramide) was cytotoxic and anti-proliferative against a panel of human melanoma cell lines (SK-Mel2, WM-266.4 and A-375 and WM-115). In addition, liposomal C6 induced caspase-dependent apoptotic death in the melanoma cells. Reversely, its cytotoxicity was attenuated by several caspase inhibitors. Intriguingly, liposomal C6 was non-cytotoxic to B10BR mouse melanocytes and primary human melanocytes. Molecularly, liposomal C6 activated protein phosphatase 1 (PP1) to inactivate Akt-mammalian target of rapamycin (mTOR) signaling in melanoma cells. On the other hand, PP1 shRNA knockdown or exogenous expression of constitutively activate Akt1 (CA-Akt1) restored Akt-mTOR activation and significantly attenuated liposomal C6-mediated cytotoxicity and apoptosis in melanoma cells. Our results suggest that liposomal C6 activates PP1 to inhibit melanoma cells.

Exposure rate of unwrapped hydroxyapatite orbital implants in enucleation surgery.

OBJECTIVE: To document the long-term exposure rate of unwrapped coralline hydroxyapatite (HA) orbital implants and explore possible risk factors. DESIGN: This retrospective case series (May 2008-April 2013) reviewed the 234 patients with anophthalmia who underwent insertion of an unwrapped HA orbital implant by one of two different surgical closing techniques. RESULTS: Of the 234 cases, 151 underwent a rectus end-to-end suturing closure technique and 83 underwent a rectus orthotopic suturing closure technique. The time of follow-up ranged from 25 months to 69 months (mean 41.9 months). Implant exposure developed in 11 cases. Three in the rectus end-to-end suturing closure group (2.0%) and eight in the rectus orthotopic suturing closure group (9.6%). In the rectus end-to-end suturing technique, a crosswise fixation of vascularised rectus muscle tissue is formed across the front of the implant; in this group the incidence of implant exposure was reduced (OR=8.11, p=0.013). Prior ocular surgery was found to be a factor increasing the incidence of HA exposure (OR=2.73, p=0.032). CONCLUSIONS: The placement of an unwrapped HA orbital implant with rectus end-to-end suturing in enucleation surgery was associated with a low rate of exposure in most cases. The end-to-end suturing creates a joint-like structure over the HA sphere, protecting the Tenon's capsule and conjunctiva from its rough surface and reducing the risk of implant exposure. Prior ocular surgery may be another risk factor for HA exposure.

In vivo vascularization of MSC-loaded porous hydroxyapatite constructs coated with VEGF-functionalized collagen/heparin multilayers.

Rapid and adequate vascularization is vital to the long-term success of porous orbital enucleation implants. In this study, porous hydroxyapatite (HA) scaffolds coated with vascular endothelial growth factor (VEGF)-functionalized collagen (COL)/heparin (HEP) multilayers (porosity 75%, pore size 316.8 ± 77.1 µm, VEGF dose 3.39 ng/mm(3)) were fabricated to enhance vascularization by inducing the differentiation of mesenchymal stem cells (MSCs) to endothelial cells. The in vitro immunofluorescence staining, quantitative real-time polymerase chain reaction (qRT-PCR), and western blotting results demonstrated that the expression of the endothelial differentiation markers CD31, Flk-1, and von Willebrand factor (vWF) was significantly increased in the HA/(COL/HEP)5/VEGF/MSCs group compared with the HA/VEGF/MSCs group. Moreover, the HA/(COL/HEP)5 scaffolds showed a better entrapment of the MSCs and accelerated cell proliferation. The in vivo assays showed that the number of newly formed vessels within the constructs after 28 d was significantly higher in the HA/(COL/HEP)5/VEGF/MSCs group (51.9 ± 6.3/mm(2)) than in the HA (26.7 ± 2.3/mm(2)) and HA/VEGF/MSCs (38.2 ± 2.4/mm(2)) groups. The qRT-PCR and west rn blotting results demonstrated that the HA/(COL/HEP)5/VEGF/MSCs group also had the highest expression of CD31, Flk-1, and vWF at both the mRNA and protein levels.

Scanning electron microscopic studies of Supramid Extra from the patients displaying recurrent ptosis after frontalis suspension.

PURPOSE: To investigate the morphologic changes in Supramid Extra suture harvested from patients displaying recurrent ptosis who had undergone a frontalis suspension for congenital ptosis. DESIGN: Observational case series. METHODS: Using scanning electron microscopy, we examined the morphologic changes in Supramid Extra (S. Jackson Inc, Alexandria, Virginia, USA) harvested from 20 consecutive patients (23 eyelids) with recurrent ptosis who had undergone frontalis suspension. Disintegration was graded as mild, moderate, or severe. RESULTS: Examination of unused Supramid showed the smooth surface of the sheath and round, well-demarcated polyfilaments on cross section. However, implanted Supramid showed different degrees of disintegration. The surface of the sheath had a rough texture with cracks and peeling. On cross section, an accumulation of debris among the polyfilaments and irregular architectures with decreased diameters of the polyfilaments were noted in some cases. On the surface of polyfilaments, different degrees of debris were noted in all cases. The degree of the changes was not exactly correlated with either the time of the recurrence or the duration of implantation. However, as the duration of implanted Supramid increased, greater changes on the surface of the sheath were noted. With implantation of less than 37 months, mild changes (44%) were predominant; between 37 and 48 months, moderate changes (60%) were noted, and with more than 48 months of implantation, severe changes (44%) were predominant. CONCLUSIONS: Supramid undergoes morphologic degradation by hydrolysis after implantation. This change might be considered one of the possible factors that facilitate the recurrence of ptosis.