Photothermal effective CeO2NPs combined in thermosensitive hydrogels with enhanced antibacterial, antioxidant and vascularization performance to accelerate infected diabetic wound healing.

Chronic diabetic wound healing remains a formidable challenge due to susceptibility to bacterial infection, excessive oxidative stress, and poor angiogenesis. To address these issues, a sodium alginate (SA) based photothermal hydrogel dressing with multifunction was fabricated to facilitate wound treatment. Ceria nanoparticles (CeO2NPs) was synthesized, and their antibacterial performance by near-infrared light triggered photothermal effects was first studied and verified in this work. In addition, to release CeO2NPs to achieve antioxidation and pro-vascularization, thermosensitive gelatin (Gel) was utilized to embed the nanoparticles in advance and then composited in SA hydrogel networks. SA network was finally strengthened by acid soaking to form partially crystalline regions to act as natural crosslinkers. Results showed that the Gel/SA/CeO2 hydrogel displayed temperature-responsive release of CeO2NPs, significant antibacterial and antioxidative activity, as well as the ability to remove without injury and promote infected diabetic wound healing with low cytotoxicity, according to antibacterial investigations, cell studies, and in vivo animal studies. This research offers not only a successful method for quickening the healing of diabetic wounds but also a fresh approach to the general use of CeO2NPs.

Sprayable alginate hydrogel dressings with oxygen production and exosome loading for the treatment of diabetic wounds.

Chronic wound unhealing is a common complication in diabetic patients, which is mainly caused by tissue hypoxia, slow vascular recovery, and a long period of inflammation. Here we present a sprayable alginate hydrogel (SA) dressing consisting of oxygen-productive (CP) microspheres and exosomes (EXO) to promote local oxygen generation, accelerate macrophage towards M2 polarization, and improve cell proliferation in diabetic wounds. Results show that the release of oxygen continues for up to 7 days, reducing the expression of hypoxic factors in fibroblasts. In vivo, the diabetic wounds experiment showed that the CP/EXO/SA dressing apparently accelerated full-thickness wound healing characteristics such as the promotion of wound healing efficiency, rapid re-epithelization, favorable collagen deposition, abundant angiogenesis at the wound beds, and shortened inflammation period. EXO synergistic oxygen (CP/EXO/SA) dressing suggests a promising treatment measure for diabetic wounds.

3D light-curing printing to construct versatile octopus-bionic patches.

Reliable, fast and switchable gluing modes are critically important in medical adhesives and intelligent climbing robot applications. The octopus-bionic patch has attracted the attention of many scholars. The suction cup structure of the octopus achieves adhesion through differential pressure, showing strong adhesion in both dry and wet environments. However, the construction of the octopus-bionic patch remains limited in terms of adaptability, personalization and mass production. Herein, a composite hydrogel consisting of gelatin methacryloyl (GelMA), polyethylene glycol diacrylate (PEGDA) and acrylamide (AAM) was developed, and a structure mimicking the octopus sucker was constructed by digital light processing (DLP). The obtained octopus-bionic patch has strong adhesion, good biocompatibility and multi-functionality. Compared with the template method in most studies, the octopus-bionic patch constructed by the DLP printing method has the advantages of customizability and low cost. In addition, the DLP printing method endows the patch surface with an octopus-like groove structure for a better bionic effect.

Fabrication of an Injectable Star-polylactide/Thiolated Hyaluronate Hydrogel as a Double Drug-Delivery System for Cancer Treatment.

Unsatisfactory solid-tumor penetration or rapid metabolism of nanomaterials limits their therapeutic efficacy. Here, we designed an injectable thiolated hyaluronate (HA-SH) hydrogel as a stable drug-releasing platform for in situ tumor treatment. Biodegradable star-shaped polylactide (S-PLLA) was first synthesized and fabricated to porous microspheres to encapsulate hydrophobic curcumin (Cur@S-PLLA), which was then blended with hydrophilic doxorubicin (Dox) and the HA-SH precursor to form composite in situ formable hydrogels [Cur@S-PLLA/(Dox)HA-SH]. The results showed that adding the microspheres improved the performance of the hydrogel, such as decreasing the gelation time from 1080 s to 960 s and also the swelling ratio. The mechanical strength increased from 27 to 45 kPa. In addition, the double drug system guaranteed a sustained release of drugs, releasing Dox at the early stage, with the continuous later release of Cur after gel swelling or S-PLLA degradation to achieve long-lasting tumor suppression, which inhibits the survival of cancer cells. The inhibitory effects of the hydrogels on MCF-7 were studied. The cell activity in the double-loaded hydrogel was significantly lower than that of the control groups, and apparent dead cells appeared in 2 days and fewer living cells with time. Flow cytometry revealed that the Cur@S-PLLA/(Dox)HA-SH group had the highest apoptosis ratio of 86.60% at 12 h, and the drugs caused the cell cycle to be blocked in phase M to reduce cell division. In summary, the innovative release platform is expected to be used in long-lasting tumor suppression and provides more ideas for the design of drug carriers.

In vitro and in vivo antimicrobial activity of antimicrobial peptide Jelleine-I against foodborne pathogen Listeria monocytogenes.

As a human foodborne pathogen, Listeria monocytogenes can cause severe human listeriosis and develop resistance to antibiotics. Antimicrobial peptides (AMPs) are produced from all kingdoms of life and regarded as promising alternatives to conventional antibiotics. Jelleine-I is an AMP identified from honeybees royal jelly. In this study, we explored the activity and action mechanism of Jelleine-I against L. monocytogenes. We found its minimum inhibitory concentration to be 12.5 µg/mL. Membrane permeability analysis revealed that Jelleine-I increased L. monocytogenes cell membrane permeability, causing calcium leakage. Scanning, transmission electron microscopy and fluorescence microscopy revealed that Jelleine-I destroyed membrane integrity, disrupted intracellular structures and interacted with the bacterial DNA. DNA binding analysis demonstrated that Jelleine-I bound to bacterial genomic DNA. Results of reverse transcription-quantitative PCR revealed that Jelleine-I affected bacterial DNA replication gene expression levels. Moreover, Jelleine-I induced cellular reactive oxygen species (ROS) production from fluorescence intensity analysis, and inhibited bacterial biofilm formation. Results of immunomodulation in Galleria mellonella revealed that Jelleine-I increased host hemocyte counts, upregulated host AMP gene (Gloverin and Cecropin D) expression, and inhibited proinfammatory cytokine (tumor necrosis factor α and interleukin 6) production induced by bacterial infection. It efficiently killed bacteria and increased the survival rate of infected insects to 70 %. Furthermore, Jelleine-I increased the G1 to S phase transition in mammalian cells from cells cycle analysis, and cytotoxicity assay results indicated that it promoted cell proliferation without hemolysis or cytotoxicity. Collectively, Jelleine-I possesses antimicrobial, immunomodulatory and cell proliferative activities, and is a promising candidate for preventing L. monocytogenes emergence and dissemination.

Temperature- and pH-responsive injectable chitosan hydrogels loaded with doxorubicin and curcumin as long-lasting release platforms for the treatment of solid tumors.

The efficacy of treating solid tumors with chemotherapy is primarily hindered by dose-limiting toxicity due to off-target effects and the heterogeneous drug distribution caused by the dense extracellular matrix. The enhanced permeability and retention (EPR) effect within tumors restricts the circulation and diffusion of drugs. To overcome these obstacles, hydrogels formed in situ at the tumor site have been proposed to promote drug accumulation, retention, and long-lasting release. We developed a thiolated chitosan (CSSH) hydrogel with a gelation point of 37°C. Due to the pH-sensitive characteristics of disulfides, the prepared hydrogel facilitated drug release in the acidic tumor environment. A drug release system composed of hydrophilic doxorubicin (Dox) and hydrophobic liposome-encapsulated curcumin (Cur-Lip) was designed to enhance the long-lasting therapeutic impacts and reduce adverse side effects. These composite gels possess a suitable gelation time of approximately 8-12 min under physiological conditions. The cumulative release ratio was higher at pH = 5.5 than at pH = 7.4 over the first 24 h, during which approximately 10% of the Dox was released, and Cur was released slowly over the following 24-120 h. Cell assays indicated that the Cur-Lip/Dox/CSSH gels effectively inhibited the growth of cancer cells. These in situ-formed Cur-Lip/Dox gels with long-term drug release capabilities have potential applications for tumor suppression and tissue regeneration after surgical tumor resection.

Liquid Crystal Modified Polylactic Acid Improves Cytocompatibility and M2 Polarization of Macrophages to Promote Osteogenesis.

Liquid crystalline phases (LC phases) are widely present in an organism. The well-aligned domain and liquidity of the LC phases are necessary for various biological functions. How to stabilize the floating LC phases and maintain their superior biology is still under study. In addition, it is unclear whether the exogenous LC state can regulate the immune process and improve osteogenesis. In this work, a series of composite films (PLLA/LC) were prepared using cholesteryl oleyl carbonate (COC), cholesteryl pelargonate (CP), and polylactic acid (PLLA) via a controlled facile one-pot approach. The results showed that the thermo-responsive PLLA/LC films exhibited stable LC phases at human body temperature and the cytocompatibility of the composites was improved significantly after modification by the LC. In addition, the M2 polarization of macrophages (RAW264.7) was enhanced in PLLA/LC films, and the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) was improved as co-cultured with macrophages. The in vivo bone regeneration of the materials was verified by calvarial repair, in which the amount of new bone in the PLLA-30% LC group was greater than that in the PLLA group. This work revealed that the liquid crystal-modified PLLA could promote osteogenesis through immunomodulation.

Construction of biomimetic artificial intervertebral disc scaffold via 3D printing and electrospinning.

Intervertebral disc (IVD) degeneration is a clinically disease that seriously endangers people's health. Tissue engineering provides a promising method to repair and regenerate the damaged IVD physiological function. Successfully tissue-engineered IVD scaffold should mimic the native IVD histological and macro structures. Here, 3D printing and electrospinning were combined to construct an artificial IVD composite scaffold. Poly lactide (PLA) was used to print the IVD frame structure, the oriented porous poly(l-lactide)/octa-armed polyhedral oligomeric silsesquioxanes (PLLA/POSS-(PLLA)8) fiber bundles simulated the annulus fibrosus (AF), and the gellan gum/poly (ethylene glycol) diacrylate (GG/PEGDA) double network hydrogel loaded with bone marrow mesenchymal stem cells (BMSCs) simulated the nucleus pulposus (NP) structure. Morphological and mechanical tests showed that the structure and mechanical properties of the IVD scaffold were similar to that of the natural IVD. The compression modulus of the scaffold is about 10 MPa, which is comparable to natural IVD and provides good mechanical support for tissue repair and regeneration. At the same time, the porosity and mechanical properties of the scaffold can be regulated through the 3D model design. In the AF structure, the fiber bundles are oriented concentrically with each subsequent layer oriented 60° to the spinal column, and can withstand the tension generated during the deformation of the NP. In the NP structure, BMSCs were evenly distributed in the hydrogel and could maintain high cell viability. Animal experiment results demonstrated that the biomimetic artificial IVD scaffold could maintain the disc space and produce the new extracellular matrix. This engineered biomimetic IVD scaffold is a promising biomaterial for individualized IVD repair and regeneration.

CD47-mediated DTIC-loaded chitosan oligosaccharide-grafted nGO for synergistic chemo-photothermal therapy against malignant melanoma.

Nano-graphene oxide (nGO), an effective drug nanocarrier, is used for simultaneous photothermal therapy (PTT) and near-infrared fluorescence imaging. Dacarbazine (DTIC) is used in the treatment of melanoma with limited clinical efficacy. PTT shows promise in the treatment of skin cancer. Herein, chitosan oligosaccharide (COS)-grafted nGO was further modified with CD47 antibody, and loaded DTIC was prepared using a versatile nanoplatform (nGO-COS-CD47/DTIC) for the treatment of melanoma as a synergistic targeted chemo-photothermal therapy. The in vitro results demonstrated that nGO-COS-CD47/DTIC nanocarriers have excellent biocompatibility, photothermal conversion efficiency, high targeting efficiency, fast drug release under NIR irradiation, and tumor cell killing efficiency. Notably, nGO-COS-CD47/DTIC plus NIR irradiation significantly promoted early cell apoptosis through the mitochondrial apoptosis pathway and exhibited a significant joint function of antitumor efficacy. The demonstrated nGO-COS-CD47/DTIC can provide a highly efficient malignant melanoma therapy using this multifunctional intelligent nanoplatform.

Antibacterial poly (ethylene glycol) diacrylate/chitosan hydrogels enhance mechanical adhesiveness and promote skin regeneration.

Various functional active hydrogels have been widely applied in tissue regeneration, especially in fields of wound repair as they are similar to the natural extracellular matrix (ECM) and can maintain moist at the wound site. However, preparing a hydrogel with multifunctional properties including high mechanical properties, excellent biocompatibility and long-term antibacterial activity is still a challenge. Herein, we developed a series of double network hydrogels based on poly (ethylene glycol) diacrylate (PEGDA) and chitosan (CS) or thiolated chitosan (TCS). The hydrogels presented in situ forming properties, good mechanical strength, adhesiveness, antibacterial activity and biocompatibility. The sample with the optimal formula of 15 wt% of PEGDA and 2 wt% of CS or TCS showed excellent mechanical adhesiveness, sustained release of antibacterial peptide and plasmid DNA, and it significantly accelerated in vivo wound healing process in a full-thickness skin defect model by reducing the inflammation and promoting the angiogenesis, meaning that the prepared hydrogels are excellent candidates for wound dressing.

Fabrication, antibacterial activity and cytocompatibility of quaternary ammonium chitooligosaccharide functionalized polyurethane membrane via polydopamine adhesive layer.

In this study, to enhance the antibacterial activity and cytocompatibility of the electrospinning polyurethane (PU) fibrous membrane, quaternary ammonium chitooligosaccharide (G-COS) was immobilized on the fibrous membrane surface via an intermediate layer of polydopamine (PDOPA) to obtain the G-COS functionalized PU (G-C-D-PU), as a control, chitooligosaccharide (COS) functionalized PU fibrous membrane (C-D-PU) was prepared, too. Surface composition, morphology, hydrophilicity and surface energy of the original and modified PU fibrous membranes were characterized, which revealed that the surface roughness and hydrophilicity of the PU fibrous membrane were obviously increased by modified with COS and G-COS, respectively. Antibacterial experiment against E. coli and S. aureus indicated that antibacterial activity of the G-C-D-PU fibrous membrane was markedly superior to that of pure PU and C-D-PU fibrous membranes. In vitro cells culture experiments revealed that the adhesion and proliferation of NIH-3T3 cells on the PU fibrous membrane were improved by successively immobilized with PDOPA and COS as well as G-COS with the concentration of 2 g/L and 6 g/L. Moreover, the G-C-D-PU fibrous membranes with relative high G-COS content were more beneficial to the enhancement of antibacterial activity, but on the contrary, those with relative low G-COS content were more in favor of cell attachment and proliferation.

Dual drug loaded coaxial electrospun PLGA/PVP fiber for guided tissue regeneration under control of infection.

Electrospinning promisingly fabricate mats for Guided Tissue Regeneration (GTR). Due to a chronic inflammatory pathology in periodontal, it is highly desirable to develop a novel GTR mats to realize tissue regeneration under control of infection. In the study, coaxial electrospinning was firstly conducted to fabricate dual drug loaded fiber mats with core/shell structure. Naringin-loaded polyvinylpyrrolidone was designed as core fiber to enrich tissue regeneration and metronidazole-loaded poly(lactic-co-glycolic acid) as shell fiber to inhibit bacterial. TEM revealed that the fibers with distinct core/shell structure were in an outer diameter of 1.5-1.7 µm with an inner diameter of <1.0 µm. The loading of dual drug decreased the tensile strength and elongation of the coaxial fiber mats. On in vitro assessment, metronidazole had a short-term release while naringin had a long-term release behavior in all the coaxial mats. The colonization of anaerobic bacteria on the mats effectively were inhibited over 21 days. Furthermore, the dual drug loaded coaxial fiber mats were observed to positively supported the adhesion and proliferation of MC3T3-E1 and was conductive to high alkaline phosphatase express. Thus, a simple and effective coaxial electrospinning approach was demonstrated for the fabrication of anti-infective GTR mats with promoting tissue regeneration.

A novel biomimetic scaffold with hUCMSCs for lumbar fusion.

Discectomy and lumbar fusion are common clinical approaches to treating intervertebral disc (IVD) degeneration with the aid of autologous bone and/or biomaterials. Biomaterials are considered suitable if they are biodegradable and can guide tissue regeneration. These features are necessary for total IVD removal, when it degenerates, as lumbar fusion treatment, avoiding secondary damage caused by the use of autogenous bone. In this work, a novel biomimetic porous chitosan/poly(l-lactic acid) scaffold with human umbilical cord mesenchymal stem cells (hUCMSCs) was applied in lumbar fusion. Hierarchically porous chitosan scaffolds were prepared using molds, porogens and freeze drying, and then poly(l-lactic acid) networks were distributed throughout the interior scaffold to enhance mechanical strength. We conducted cell culture in vitro and animal experiments in vivo. The scaffolds were incubated in hUCMSCs medium and co-cultured for 8 days, and then implanted into half-destroyed IVDs in New Zealand rabbits. The results show that scaffolds with hUCMSCs had greater ability to guide disc regeneration than the blank control and autologous bone, as determined using X-rays, computed tomography, and histologic analyses. Findings confirmed the role of hUCMSCs stimulation in bone-tissue healing and IVD regeneration. This hMUMSCs-based approach, together with the strategy proposed for incorporating osteoblastic cells into scaffolds that promote endogenous or synthetic repair mechanisms, may then be used to develop strategies for the stem-cell-based healing of other acute injuries and chronic diseases.

Antibacterial activity and cytocompatibility of chitooligosaccharide-modified polyurethane membrane via polydopamine adhesive layer.

The aim of this study was to provide a convenient surface modification method for polyurethane (PU) membrane and evaluate its influence on hydrophilicity, antibacterial activity and cell functions, which are the most important factors for wound dressings. For this purpose, chitooligosaccharide (COS) was modified onto the surface of PU membrane based on the self-polymerization of dopamine (DOPA). Surface composition, morphology, hydrophilicity and surface energy of the original and modified PU membranes were characterized. Surface roughness and hydrophilicity of the PU membrane were obviously increased by modified with polydopamine (PDOPA) and COS. Antibacterial experiment against Escherichia coli and Staphylococcus aureus indicated that antibacterial activity of PU membrane increased only slightly by modified with PDOPA, but increased significantly by further modified with COS. Cells culture results revealed that COS-functionalized PU membrane is more beneficial to the adhesion and proliferation of NIH-3T3 cells compared to the original and PDOPA-modified PU membranes.

Rapid biomimetic mineralization of collagen fibrils and combining with human umbilical cord mesenchymal stem cells for bone defects healing.

Collagen biomineralization is regulated by complicated interactions between the collagen matrix and non-collagenous extracellular proteins. Here, the use of sodium tripolyphosphate to simulate the templating functional motif of the C-terminal fragment of non-collagenous proteins is reported, and a low molecular weight polyacrylic acid served as a sequestration agent to stabilize amorphous calcium phosphate into nanoprecursors. Self-assembled collagen fibrils served as a fixed template for achieving rapid biomimetic mineralization in vitro. Results demonstrated that, during the mineralization process, intrafibrillar and extrafibrillar hydroxyapatite mineral with collagen fibrils formed and did so via bottom-up nanoparticle assembly based on the non-classical crystallization approach in the presence of these dual biomimetic functional analogues. In vitro human umbilical cord mesenchymal stem cell (hUCMSC) culture found that the mineralized scaffolds have a better cytocompatibility in terms of cell viability, adhesion, proliferation, and differentiation into osteoblasts. A rabbit femoral condyle defect model was established to confirm the ability of the n-HA/collagen scaffolds to facilitate bone regeneration and repair. The images of gross anatomy, MRI, CT and histomorphology taken 6 and 12weeks after surgery showed that the biomimetic mineralized collagen scaffolds with hUCMSCs can promote the healing speed of bone defects in vivo, and both of the scaffolds groups performing better than the bone defect control group. As new bone tissue formed, the scaffolds degraded and were gradually absorbed. All these results demonstrated that both of the scaffolds and cells have better histocompatibility.

Investigation of the antimicrobial activity and biocompatibility of magnesium alloy coated with HA and antimicrobial peptide.

Implant-associated infection is one of the biggest problems in orthopedic surgery. Antimicrobial peptides (AMPs) are well-known components of the innate immunity and less susceptible to the development of pathogen resistance compared to conventional antibiotics. Magnesium alloys as potential biodegradable bone implants have been received much attention in biomaterials field. This study investigated the deposition of calcium phosphate (CaP) coatings and loading of AMPs on the magnesium alloy surface by a biomimetic method. Scanning electron microscope (SEM) results presented that a microporous and plate-like CaP coating was processed on the magnesium alloy surface. X-ray diffractometry (XRD) and Fourier transform infrared spectroscopy (FTIR) analysis showed the main component of coating was hydroxyapatite (HA). Degradation assay in vitro showed that the HA coating deposited onto the magnesium alloy was corroded more slowly than the bare one. The amount of AMP loaded in the HA coating was 11.16±1.99 µg/cm2. The AMP loaded onto HA coatings had slow release for 7 days. The AMP-loaded coating showed antimicrobial activity against Staphylococcus aureus. Its bacterial inhibition rate exceeded 50% after 4 days and the antibacterial effect was sustained for 7 days. The coated magnesium alloys loaded with AMP could improve rat bone marrow mesenchymal stem cells (rBMMSCs) proliferation. Furthermore, it could also promote alkaline phosphatase (ALP) activity of rBMMSCs. Both radiographic evaluation and histopathology analysis demonstrated that implantation of the coated magnesium alloy into the rabbit femoral condyle had promoted bone repair and showed anti-inflammatory effect. The results showed that the AMP loaded onto HA coatings on the magnesium alloy surface could be considered an ideal orthopedic implant against S. aureus infection.

Osteodifferentiation of mesenchymal stem cells on chitosan/hydroxyapatite composite films.

Chitosan (Ch) is one of the most commonly used natural biomaterials. Osteodifferentiation of mesenchymal stem cells (MSCs) on Ch has drawn extensive interest. Hydroxyapatite (HA) is a component of skeleton and teeth with good biocompatibility. Combination with HA may be a good method to modify Ch to facilitate cellular behaviors and functions on it. In this study, Ch/HA film was prepared and characterized. Its potential to benefit cellular behaviors and osteodifferentiation of MSCs was evaluated. Resultantly, physical properties of composite Ch/HA, including water-in-air contact angle, tensile strength, elastic modulus, and breaking elongation were favorably modified. In cellular culture medium, Ch/HA films absorbed more Ca(2+) than Ch films, and more HA crystalline growths on Ch/HA films. 3-(4,5-Dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay and morphological features showed better proliferation and adhesion of MSCs on Ch/HA films. Osteodifferentiation of MSCs on Ch/HA was promoted, indicated by modified transcription level of osteocalcin, osteopontin, collagen I, alkaline phosphatase (ALP), and induced ALP activity. These data suggest biocompatibility of Ch is modified after being blended with HA, which promotes osteodifferentiation of MSCs. This can be a promising approach to modify Ch for its applications in bone tissue engineering.

[Preparation of chitosan/hydroxyapatite membrane and its effect on cell culture].

Compound membranes of chitosan/hydroxyapatite were prepared by blending. The physical performance showed that the air-water contact angles decreased from chitosan's 103 degrees to chitosan/hydroxyapatite's 57 and the water adsorption rate increased slightly. When immersed into culture medium, the materials adsorbed Ca2+, and low crystalline hydroxyapatite deposited on the surface of the membranes. Chitosan/hydroxyapatite compound membranes could enhance the attachment and proliferation of mescenchymal stem cells (MSCs). After 12 days' induction on the materials, the alkaline phosphatase (ALP) activity value of MSCs on the compound membrane was 10.1, being much higher than 1.6 on chitosan membrane (P<0.01). All these results indicate that chitosan does not have very good affinity for MSCs, but the biocompatibility of chitosan can be apparently enhanced after mixing with hydroxyapatite. The compound membrane stimulates MSCs to differentiate into osteoblasts and it may be a good potential material for bone substitution.

Molecular and functional characterization of a c-type lysozyme from the Asian corn borer, Ostrinia furnacalis.

Some lepidopteran lysozymes have been reported to display activity against Gram-positive and Gram-negative bacteria, in contrast to most lysozymes that are active only against Gram-positive bacteria. OstrinLysC, a c-type lysozyme, was purified from the Asian corn borer, Ostrinia furnacalis Guenée (Lepidoptera: Pyralidae), and shows activity against Gram-positive and Gram-negative bacteria. The NH2-terminal amino acid sequence was determined by Edman degradation and used in a homology cloning strategy. The gene coding for OstrinLysC contains three exons and two introns. The expression profile of the OstrinlysC gene was examined by quantitative real-time PCR. Following injection of the larvae with bacteria, the OstrinlysC gene is strongly up-regulated in immune tissues. Transcripts were also detected in gut tissue. After feeding the larvae with bacteria, OstrinlysC transcripts increased in immune tissues. A very low level of transcript abundance was also detected in gut tissue. These results suggested that the OstrinlysC gene is involved in immune responses. The three dimensional structure of OstrinLysC was predicted. Based on comparison of the 3-D structure of OstrinLysC with that of silkworm lysozyme and chicken lysozyme, we hypothesize that the positive charge-rich surface and the short loop-2, which is close to the cluster of hydrophobic residues, may play important roles in the interaction with the outer membrane of Gram-negative bacterial cell walls.