LECT2 mediates antibacterial immune response induced by Nocardia seriolae infection in the northern snakehead.

Leukocyte-derived chemotaxin-2 (LECT2) is a multifunctional immunoregulator that plays several pivotal roles in the host's defense against pathogens. This study aimed to elucidate the specific functions and mechanisms of LECT2 (CaLECT2) in the northern snakehead (Channa argus) during infections with pathogens such as Nocardia seriolae (N. seriolae). We identified CaLECT2 in the northern snakehead, demonstrating its participation in the immune response to N. seriolae infection. CaLECT2 contains an open reading frame (ORF) of 459 bp, encoding a peptide of 152 amino acids featuring a conserved peptidase M23 domain. The CaLECT2 protein shares 62%-84% identities with proteins from various other fish species. Transcriptional expression analysis revealed that CaLECT2 was constitutively expressed in all examined tissues, with the highest expression observed in the liver. Following intraperitoneal infection with N. seriolae, CaLECT2 transcription increased in the spleen, trunk kidney, and liver. In vivo challenge experiments showed that injecting recombinant CaLECT2 (rCaLECT2) could protect the snakehead against N. seriolae infection by reducing bacterial load, enhancing serum antibacterial activity and antioxidant capacity, and minimizing tissue damage. Moreover, in vitro analysis indicated that rCaLECT2 significantly enhanced the migration, respiratory burst, and microbicidal activity of the head kidney-derived phagocytes. These findings provide new insights into the role of LECT2 in the antibacterial immunity of fish.

A comprehensive analysis of the allergenicity and IgE epitopes of myosinogen allergens in Scylla paramamosain.

BACKGROUND: Scylla paramamosain is one of the most common and serious food allergens in Asia. Therefore, research on its prevalence, accurate diagnosis, and IgE-binding pattern of the allergens is crucial. OBJECTIVE: To identify the IgE epitopes of the myosinogen allergens in S. paramamosain using phage peptide library. METHODS: The prevalence of allergy to crabs (AC) and of sensitization was analysed using a questionnaire and a serological assay. BAT was performed by flow cytometry, and its diagnostic performance was evaluated in relation to allergens purified from crab myosinogen. IgE-binding epitopes were identified by phage display using the IgE from patients with AC. Sequence- and structure-based bioinformatics analyses were performed to identify allergenic epitopes. RESULTS: Crab was the most common cause of food allergies in this study. Subjects with AC (n = 30) with clear clinical symptoms were identified by immunoblotting and BAT. All of the myosinogen allergens triggered basophil activation; surface expression of CD63 and CD203c was higher in patients allergic to AK and FLN c than in patients allergic to SCP and TIM. In addition to six conformational epitopes of SCP, six linear epitopes and eight conformational epitopes of AK were identified. Five linear epitopes and three conformational epitopes of TIM, nine linear and ten conformational epitopes of FLN c were also identified, and the sequence VH(I/T) L was appeared in epitopes of both TIM and FLN c. The number of epitopes showed consistency with the value of BAT. CONCLUSIONS AND CLINICAL RELEVANCE: BAT can be used for accurate diagnosis of AC. Identification of particular allergenic motifs could be a valuable tool for prevention, diagnosis, and treatment of food allergies.

[Components of Urinary Nanocrystals and Their Influence on Formation of Calcium Oxalate Stones].

High-resolution transmission electron microscopy, X-ray diffraction, selected area electron diffraction (SAED), and energy dispersive spectroscopy (EDS) were accurately performed to analyze the components of nanocrystals in the urine of patients with calcium oxalate (CaOx) stones. XRD, SAED and FFT detected the presence of calcium oxalate monohydrate (COM), uric acid (UA), and calcium phosphate (CaP). EDS detected the elements of C, O, Ca, with a small amount of N and P. These results showed that the main components of urinary nanocrystals were COM, with a small amount UA and phosphate. HRTEM observation showed that the particle size of urinary nanocrystals was dozens of nanometers. The result was consistent with the calculation by Debye-Scherrer equation. When the urine was filtered through a microporous membrane of 0.45, 1.2, and 3 µm, respectively, the number of diffraction peaks of the obtained urine crystallites increased with the increased pore size, indicating the increase of urinary crystallite species. Crystal nucleation, growth, aggregation, and adhesion of crystals to the renal epithelial cells are important processes for CaOx stone formation. The presence of a large amount of COM crystals in patients' urine is a critical factor for CaOx stones formation. Nano UA and CaP crystallite can induce the CaOx stone formation as central nidus.

Size-dependent toxicity and interactions of calcium oxalate dihydrate crystals on Vero renal epithelial cells.

Urinary crystals in normal and kidney stone patients often have varying sizes; the interaction between renal epithelial cells and COD crystals generated in the tubular fluid could play an initiating role in the pathophysiology of calcium oxalate nephrolithiasis. This study aims to compare the cytotoxicity of micro/nano-calcium oxalate dihydrate (COD) crystals (50 nm, 100 nm, 600 nm, 3 µm, and 10 µm) toward African green monkey renal epithelial (Vero) cells to reveal the mechanism of kidney stone formation at the molecular and cellular levels. METHODS: Vero cells were exposed to COD crystals of varying sizes at a concentration of 200 µg mL-1 for 6 h. The effects of COD crystals on Vero cell viability, apoptosis rate, and cellular biochemical parameters [lactate dehydrogenase (LDH), superoxide dismutase (SOD), reactive oxygen species (ROS), hyaluronic acid (HA), osteopontin (OPN), and mitochondrial membrane potential (Δψm)] were determined using biochemical and morphological analyses. RESULTS: Vero cell viability and apoptotic rate were closely associated with the size of COD crystals; lower cell viability and higher apoptosis rate were observed in cells exposed to smaller COD crystal size. The expression of SOD, ROS, HA and OPN also changed in a size-dependent manner after exposure to the five different sizes of COD crystals. The area ratio of the (100) face with a high density of Ca2+ ions to the total surface area was also found to influence the severity of cell injury. Cell injury induced by COD crystals was mainly caused by excessive expression of intracellular ROS and reduction of free-radical scavenger SOD. Moreover, binding of large crystals on the cell membrane surface takes more time to cause cell injury than internalized small-sized crystals. The cell death rate was found to be positively correlated with the amount of internalized COD crystals. CONCLUSIONS: although the COD toxicity is often disregarded, the size-dependent cytotoxicity of COD crystals toward Vero cells is demonstrated in this study.

[Characteristics of hydroxyapatite/PMMA nanocomposites for provisional restoration and its biocompatibility with human gingival fibroblasts].

OBJECTIVE: To formulate hydroxyapatite (HA)/polymethyl methacrylate (PMMA) composites with improved cytocompatibility for provisional restoration. METHODS: Nanocomposites with 20 wt%, 30 wt%, 40 wt%, and 50 wt% HA/PMMA (H/P) were developed and examined using X-ray photoelectron spectroscopy (XPS) and scanning electron microscope (SEM). Human gingival fibroblasts were cultured on those HA/PMMA discs and investigated by fluorescent staining on 24 h and MTT assay at 1 d, 3 d, 5 d and 7 d. RESULTS: Chemical integration of HA/PMMA interface was confirmed by XPS. Typical fusiform cells with adhesion spots were detected on 40 wt% and 50 wt% H/P discs. MTT results showed insignificant differences in cell growth between 40 wt% H/P and pure titanium (Ti, P > 0.05), while the other H/P discs showed significantly lower cell growth than pure Ti (P < 0.05). CONCLUSION: 40 wt% H/P might be a promising candidate for provisional dental implant restoration and for esthetic gingival contour.

[Reconstruction of critical sized calvarial defects by porous nano-hydroxyapatite/polyamide 6 composite with bone marrow mesenchymal stem cells in rat].

OBJECTIVE: To evaluate the effects of the nano-hydroxyapatite/polyamide 6 (n-HA/PA6) on the proliferation and osteoblastic differentiation of rat bone marrow mesenchymal stem cells (BMSCs), and the feasibility of using both for constructing tissue engineered bone in the calvarias of rats with critical sized defects. METHODS: The third passage of BMSCs were cultured in osteoblastic medium and seeded on the scaffolds of n-HA/PA6, the proliferation of the BMSCs was tested by MTT (3-{4,5-dimethylthiazol-2yl}-2,5-diphenyl-2H-tetrazolium-bromide) on scheduled dates, and the osteoblastic differentiation of the BMSCs were measured by alkaline phosphatase (ALP) staining. Furthermore, the scaffolds with or without BMSCs in rat calvarial defects, after 4 weeks, 8 weeks, and 16 weeks have been implanted. Histology and scanning electron microscope were used to test the bone healing in the different groups. RESULTS: The BMSCs seeded on the n-HA/PA6 grew well, the proliferation of cells was not affected by the scaffold, and the staining of ALP was also positive. At 4 week and 8 week after implantation, the n-HA/PA6 with BMSCs showed more new bone formation on the surface of scaffolds, with a better osseointegration of implant and host bone when compared with the group of n-HA/PA6 without BMSCs. However, there was no significant difference between these two groups at 16 week. CONCLUSION: The porous n-HA/PA6 has no negative effects on the proliferation and osteoblastic differentiation of rat BMSCs, and using BMSCs as seed cells and n-HA/PA6 as scaffolds is a good choice for constructing tissue engineered bone due to the enhanced new bone formation and osseointegration.

Comparative study on inorganic composition and crystallographic properties of cortical and cancellous bone.

OBJECTIVE: To comparatively investigate the inorganic composition and crystallographic properties of cortical and cancellous bone via thermal treatment under 700 °C. METHODS: Thermogravimetric measurement, infrared spectrometer, X-ray diffraction, chemical analysis and X-ray photo-electron spectrometer were used to test the physical and chemical properties of cortical and cancellous bone at room temperature 250 °C, 450 °C, and 650 °C, respectively. RESULTS: The process of heat treatment induced an extension in the a-lattice parameter and changes of the c-lattice parameter, and an increase in the crystallinity reflecting lattice rearrangement after release of lattice carbonate and possible lattice water. The mineral content in cortical and cancellous bone was 73.2wt% and 71.5wt%, respectively. For cortical bone, the weight loss was 6.7% at the temperature from 60 °C to 250 °C, 17.4% from 250 °C to 450 °C, and 2.7% from 450 °C to 700 °C. While the weight loss for the cancellous bone was 5.8%, 19.9%, and 2.8 % at each temperature range, the Ca/P ratio of cortical bone was 1.69 which is higher than the 1.67 of stoichiometric HA due to the B-type CO3²â» substitution in apatite lattice. The Ca/P ratio of cancellous bone was lower than 1.67, suggesting the presence of more calcium deficient apatite. CONCLUSION: The collagen fibers of cortical bone were arrayed more orderly than those of cancellous bone, while their mineralized fibers ollkded similar. The minerals in both cortical and cancellous bone are composed of poorly crystallized nano-size apatite crystals with lattice carbonate and possible lattice water. The process of heat treatment induces a change of the lattice parameter, resulting in lattice rearrangement after the release of lattice carbonate and lattice water and causing an increase in crystal size and crystallinity. This finding is helpful for future biomaterial design, preparation and application.

[Initial immunoreaction study on PVA/n-HA + PA66 biological composite material after being implanted into animal body].

OBJECTIVE: To observe immunoreaction to PVA/n-HA+ PA66 biological composite material after being implanted into animal body. METHODS: PVA/n-HA+PA66 composite materials were implanted into mouse subcutaneous tissue. Histologial examination was performed at 2, 4, 6 weeks, the amount of CD3, CD4+, CD8+ in blood and IL-1, IL-6, TNF-alpha in spleen cells was measured at the same time. RESULTS: The amount of CD3, CD4+, CD8+ in mouse blood and IL-1, IL-6, TNF-alpha in mouse spleen cells were not different with the control groups at 2, 4, 6 weeks (P > 0.05). The fibrous tissue and some blood vessels were found growing into the porous materials, which resulted in composite materials intergration of proliferative structure. The reject reaction was not found. CONCLUSION: PVA/n-HA+PA66 biological composite material does not cause immune rejection after being implanted into animal body, verifying its good biocompatibility.

[Effect of silver-hydroxyapatite/titania nanocomposites on proliferation, vitality and ultrastructure of osteoblasts].

OBJECTIVE: Osteoblasts behaviors were investigated when cultured in the presence of silver-hydroxyapatite/titania nanoparticles (nAg-HA/TiO2) in vitro. METHODS: Rat calvaria osteoblasts were cultured in the presence of 10 mirog/mL-500 mirog/mL nHA or nAg-HA/TiO2 for 2 h, 6 h, 8 h, 24 h, 72 h and 120 h. Cell proliferation, vitality and ultrastructure were tested using cell counting chamber, MTT assay, and transmission electron microscope (TEM). RESULTS: Both cell proliferation and vitality were restricted by addition of nanoparticles, while the respiration rates as an index for unit cell viability were enhanced. nAg-HA/TiO2 at low concentration showed superiority. No remarkable ultrastructural changes were observed in the osteoblasts exposed to nanoparticles. The differences in cell proliferation and cell vitality between nAg-HA/TiO2 group and nHA group at the same concentration were insignificant. CONCLUSION: nAg-HA/TiO2 restricted cells proliferation, but triggered unit cell viability. Cytocompatibility of nAg-HA/TiO2 is almost the same as that of nHA's.

[Characterization and antibacterial effect of Ag-nHA-nTiO2/polyamide 66 nanocomposite membrane on oral bacteria].

OBJECTIVE: Undried silver-hydroxyapatite-titania (Ag-nHA-nTiO2) nanoparticles slurry was used to make membrane with polyamide 66 (PA66) by co-polymerization method. The purpose of this study is to test the physical and chemical characteristics and antibacterial ability. METHODS: The morphology, chemical components and structures of the membrane were characterized by atomic absorption spectrometer (AAS), X-ray diffraction (XRD), scanning electron microscope (SEM) and energy-dispersive X-ray analysis (EDX). Staphylococcus aureus (S. aureus), Escherichia coli (E. coli), Porphyromonas gingivalis (P. gingivalis), Fusobacterium nucleatum (F. nucleatum) and Streptococcus mutans (S. mutans) were utilized to test the antibacterial effect. RESULTS: XRD results demonstrated that the membrane have characteristic diffraction peaks of pure hydroxyapatite (HA). A homogeneous distribution of the Ca, P, Ti and Ag element in the membrane was confirmed by EDX. Both surface and section showed porous structure which was confirmed by SEM and the average hole size was 20-30 microm. The bacteria assay reflected to the antibacterial effect, 50.10% of S. aureus and 56.31% of E. coli were killed. However, 91.84% of P. gingivalis, 90.64% of F. nucleatum and 90.49% of S. mutans were killed and pictures of SEM showed obviously fewer cells on the surface. CONCLUSION: The nanocomposite membrane could be one of the bioactive materials with antibacterial properties for oral guided bone regeneration technique.

[Porous polyvinyl alcohol hydrogel composite prepared and studied initially for biocompatibility].

OBJECTIVE: To investigate the feasibility of porous polyvinyl alcohol hydrogel (PVA-H) composite, for tissue engineering scaffold. METHODS: The approach of emulsifier-foaming, freeze-drying and surfactant-cleaning method was developed to gain three kinds of porous scaffolds: porous polyvinyl alcohol (PVA), porous polyvinyl alcohol/nano-hydroxyapatite (PVA/n-HA) and porous polyvinyl alcohol/chitosan (PVA/Cs). We checked the physical features of the above three kinds of materials and further investigated their biocompatibility by SEM image analysis, mechanics test, MTT and muscle implanting. RESULTS: Three kinds of materials had the similar character of high porosity (80%), but the PVA/n-HA had the smallest mean pore size (154.5 microm) and the PVA had the biggest tensile strength (0.60 Mpa). MTT assay demonstrated three kinds of materials to have no toxicity, furthermore, PVA had better absorbance, compared with cell control group (P < 0.05). After muscle implantation, there were many muscular tissues growing into pores of PVA/Cs at the 4th week, although PVA/Cs had stronger inflammatory reaction. Other two kinds of materials had very small inflammatory reaction and there were many fibrous tissues growing into pores at the 1st, 4th, 12th week. CONCLUSION: The features of materials are changed after adding n-HA or Cs, moreover, porous PVA/Cs seems to have the activity of inducing muscle growth. Porous PVA and its composites may be applied to tissue engineering as a long-term or permanent scaffold due to their good biocompatibility, elasticity and hydrophilicity.

[Guided reconstruction of soft tissue defects of the palate with n-HA/PA66 plates: an experimental study in goats.].

PURPOSE: To evaluate the effect of n-HA/PA66 plate as a guided implant for the regeneration of the soft tissue defects in palatal region of goats. METHODS: 12 goats were divided into experimental group and control group at random. 18.0 mm x 14.0 mm soft tissue defect and 15.0 mm x 13.0 mm hard tissue defect were formed artificially in hard palate. n-HA/PA66 plates were implanted to the defect in the experimental group. The tissue regeneration in the defects was observed and the structure of regenerating tissue was analyzed by histology. RESULTS: The mucoperiosteal tissue in the experimental group regenerated along the n-HA/PA66 plates and the mucoperiosteal defects were closed in 6 weeks. The soft and hard tissues were not closed up in control group in 6 weeks. CONCLUSION: n-HA/PA66 plates can guide the regeneration of mucoperiosteal tissue to close the palatal defects.

[Experimental study on the reconstruction of mandibular defects with a new bioactive artificial bone nano-hydroxyapatite/polyamide-66 in dogs].

OBJECTIVE: This study was designed to investigate reconstruction of segmental defect in the mandible using a new bionic materials of nano-hydroxyapatite -polyainides-66 (n-HA/PA66). METHODS: Two defects (15 mm x 10 mm x 5 mm) were created in the mandibular bodies of dogs. One of defects was reconstructed with n-HA/PA66, another not repaired as a blank control. At 2, 4, 8, 12, 16 weeks after operation. Evaluation of effects of n-HA/PA66 on reconstruction of the mandibular defects was carried out by means of radiography and histology. RESULTS: From 2 to 8 weeks after operation, some fiber tissue grew into the space between n-HA/PA66 and mandibular bone. The ossification was observed at 12 weeks post-operation. At 16 weeks, the n-HA/PA66 was connected directly to the mandibular bone by the newborn bone. CONCLUSIONS: The new artificial bone of n-HA/PA66 has the effects of osteoconduction and osteoinduction, with a good biocompatibility and is an ideal bone substitute material for reconstruction of mandibular defect.

[Nano biomedical material and its application].

Nano biomedical material is a new area that shows promising prospect. In this paper, the researches and applications of nano inorganic biomaterial, nano polymer biomaterial and nano composite biomaterial are reviewed. The developmental tendency of nano biomedical material is also forecasted.