Advances in the applications of mesenchymal stem cell-conditioned medium in ocular diseases.

Mesenchymal stem cell-conditioned medium (MSC-CM), also known as secretome, is secreted by MSC and contains a variety of bioactive factors with anti-inflammatory, anti-apoptotic, neuroprotection, and proliferation effects. Increasing evidence proved that MSC-CM plays an important role in various diseases, including skin, bone, muscle, and dental diseases. However, the role of MSC-CM in ocular diseases is not quite clear, Therefore, this article reviewed the composition, biological functions, preparation, and characterization of MSC-CM and summarized current research advances in different sources of MSC-CM in corneal and retinal diseases, including dry eye, corneal epithelial damage, chemical corneal injury, retinitis pigmentosa (RP), anterior ischemic optic neuropathy (AION), diabetic retinopathy (DR), and other retinal degenerative changes. For these diseases, MSC-CM can promote cell proliferation, reduce inflammation and vascular leakage, inhibit retinal cell degeneration and apoptosis, protect corneal and retinal structures, and further improves visual function. Hence, we summarize the production, composition and biological functions of MSC-CM and focus on describing its mechanisms in the treatment of ocular diseases. Furthermore, we look at the unexplored mechanisms and further research directions for MSC-CM based therapy in ocular diseases.

Synergetic interaction of capsid proteins for virus-like particles assembly of foot-and-mouth disease virus (serotype O) from the inclusion bodies.

Recombinant virus-like particles (VLP) with single capsid protein have been successfully produced through prokaryotic system, but for those with multiple capsid proteins such as the foot-and-mouth disease virus (FMDV), this approach is more challenging. In this study, in vitro assembly of FMDV VLP was investigated with its capsids VP1, VP2 and VP3 separately expressed as inclusion bodies. After extraction and solubilization, three capsids were purified in denatured state through a flow-through model, increasing its purity to 90%. VLP assembly for FMDV was observed after diluting the mixture of denatured capsids in the ration of 1: 1: 1, while no VLP appeared if the separately diluted and refolded capsids were co-incubated. This result suggests certain synergetic interactions exist among the three capsids, which are crucial for FMDV VLP assembly. Sodium chloride and capsid protein concentration both greatly affect the assembling efficiency. After purification through size exclusion chromatography, VLP with similar diameter and morphology as inactivated FMDV were obtained, which elicited high IgG titers and B cell activation when vaccinated in mouse. It could also induce specific humoral and cellular immune responses in splenocytes proliferative experiments. Our study demonstrated the feasibility of in vitro assembling FMDV VLP from inclusion bodies of VP1, VP2 and VP3 for the first time.

A novel maxillary transverse deficiency diagnostic method based on ideal teeth position.

BACKGROUND: This study proposed a novel maxillary transverse deficiency diagnostic method and evaluated the skeletal Class I and the mild skeletal Class III groups. METHODS: Pre-treatment data from 30 mild skeletal Class III and 30 skeletal Class I patients were collected and uploaded to the Emeiqi Case Management System to design the ideal teeth positions. On these positions, the first bi-molars width was measured at the central fossa and center resistance, the maxillary first bi-premolars width was measured at the central fossa, and the mandibular first bi-premolars width was measured at the distal contact point by Mimics, then width differences of two groups were calculated respectively. RESULTS: At ideal teeth positions, there was no statistically significant difference in the maxillomandibular width in the premolar area between the two groups, but there was in the molar area, and this difference was caused by the difference in mandible width between the two groups. CONCLUSIONS: We proposed a new transverse diagnostic method and found that even the Class I group was not quite up to standard in the molar area on ideal teeth positions, and the Class III group had more severe maxillary transverse deficiency than the Class I group. Meanwhile, the maxillary transverse deficiency in the Class III group was mainly caused by the larger width of the mandible.

The subgenomes show asymmetric expression of alleles in hybrid lineages of Megalobrama amblycephala × Culter alburnus.

Hybridization drives rapid speciation by shaping novel genotypic and phenotypic profiles. Genomic incompatibility and transcriptome shock have been observed in hybrids, although this is rarer in animals than in plants. Using the newly sequenced genomes of the blunt snout bream (Megalobrama amblycephala [BSB]) and the topmouth culter (Culter alburnus [TC]), we focused on the sequence variation and gene expression changes in the reciprocal intergeneric hybrid lineages (F1-F3) of BSB × TC. A genome-wide transcriptional analysis identified 145-974 expressed recombinant genes in the successive generations of hybrid fish, suggesting the rapid emergence of allelic variation following hybridization. Some gradual changes of gene expression with additive and dominance effects and various cis and trans regulations were observed from F1 to F3 in the two hybrid lineages. These asymmetric patterns of gene expression represent the alternative strategies for counteracting deleterious effects of the subgenomes and improving adaptability of novel hybrids. Furthermore, we identified positive selection and additive expression patterns in transforming growth factor, beta 1b (tgfb1b), which may account for the morphological variations of the pharyngeal jaw in the two hybrid lineages. Our current findings provide insights into the evolution of vertebrate genomes immediately following hybridization.

Progress in Utilizing Dynamic Bonds to Fabricate Structurally Adaptive Self-Healing, Shape Memory, and Liquid Crystal Polymers.

Stimuli-responsive structurally dynamic polymers are capable of mimicking the biological systems to adapt themselves to the surrounding environmental changes and subsequently exhibiting a wide range of responses ranging from self-healing to complex shape-morphing. Dynamic self-healing polymers (SHPs), shape-memory polymers (SMPs), and liquid crystal elastomers (LCEs), which are three representative examples of stimuli-responsive structurally dynamic polymers, have been attracting broad and growing interest in recent years because of their potential applications in the fields of electronic skin, sensors, soft robots, artificial muscles, and so on. Recent advances and challenges in the developments toward dynamic SHPs, SMPs, and LCEs are reviewed, focusing on the chemistry strategies and the dynamic reaction mechanisms that enhance the performances of the materials including self-healing, reprocessing, and reprogramming. The different dynamic chemistries and their mechanisms on the enhanced functions of the materials are compared and discussed, where three summary tables are presented: A library of dynamic bonds and the resulting characteristics of the materials. Finally, a critical outline of the unresolved issues and future perspectives on the emerging developments is provided.

SILICONE OIL-FILLED FOLDABLE CAPSULAR VITREOUS BODY VERSUS SILICONE OIL ENDOTAMPONADE FOR TREATMENT OF NO LIGHT PERCEPTION AFTER SEVERE OCULAR TRAUMA.

PURPOSE: To compare the anatomical and functional outcomes of silicone oil (SO)-filled foldable capsular vitreous body (FCVB) and SO endotamponade in vitrectomy for patients with no light perception after ocular trauma. METHODS: A total of 64 patients (64 eyes) with no light perception caused by severe ocular trauma were divided into FCVB and SO groups based on the surgical treatment. The main outcome measurements were retinal reattachment rate, intraocular pressure, best-corrected visual acuity, and number of operations. RESULTS: Both the FCVB group (29 eyes) and the SO group (35 eyes) showed significant improvement in postoperative best-corrected visual acuity and intraocular pressure. The two groups showed no significant differences in final intraocular pressure and the retinal reattachment rate. The postoperative vision (≥LP) in the FCVB group was significantly worse than in the SO group (FCVB [4/29] vs. SO [18/35], P = 0.003). However, the number of surgeries in the FCVB group was significantly lower than in the SO group (FCVB [1.10] vs. SO [2.23], P < 0.001). CONCLUSION: Vitrectomy combined with SO endotamponade shows better short-term improvement in the treatment of no light perception caused by severe ocular trauma. However, SO-filled FCVB can effectively prevent many complications caused by direct SO endotamponade, such as secondary surgeries or SO dependence.

Three-Dimensional Bioprinting of Decellularized Extracellular Matrix-Based Bioinks for Tissue Engineering.

Three-dimensional (3D) bioprinting is one of the most promising additive manufacturing technologies for fabricating various biomimetic architectures of tissues and organs. In this context, the bioink, a critical element for biofabrication, is a mixture of biomaterials and living cells used in 3D printing to create cell-laden structures. Recently, decellularized extracellular matrix (dECM)-based bioinks derived from natural tissues have garnered enormous attention from researchers due to their unique and complex biochemical properties. This review initially presents the details of the natural ECM and its role in cell growth and metabolism. Further, we briefly emphasize the commonly used decellularization treatment procedures and subsequent evaluations for the quality control of the dECM. In addition, we summarize some of the common bioink preparation strategies, the 3D bioprinting approaches, and the applicability of 3D-printed dECM bioinks to tissue engineering. Finally, we present some of the challenges in this field and the prospects for future development.

Determination of PEGylation homogeneity of polyethylene glycol-modified canine uricase.

Polyethylene glycol-modified canine uricase (PEG-UHC) was prepared by modifying the ε-amino group of lysine residues on the canine uricase (UHC) protein to near-saturation with 5 kDa monomethoxyl-polyethylene glycol succinimide (mPEG-SPA-5k). In order to accurately determine the PEGylation uniformity of PEG-UHC, CZE, 3-8% gradient gel SDS-PAGE, and imaging CIEF (iCIEF) analyses were compared. CZE could not effectively separate PEG-UHC proteins with different degrees of modification, 3-8% gradient gel SDS-PAGE could separate PEG-UHC into seven gel bands; however, most of the gel bands were smeared or blurred, and the separation of PEG-UHC samples by iCIEF was significantly better than that by 3-8% gradient gel SDS-PAGE. Under denatured conditions, iCIEF separated 12 pI peaks, and could also accurately quantify the relative monomer PEG-UHC content. More than 85% of the total monomeric PEG-UHC was conjugated with 7-12 PEG molecules; of this 85%, approximately 40% was conjugated with 9-10 PEG molecules. These results demonstrated that iCIEF exhibits good potential for determining the PEGylation homogeneity of PEGylated protein drugs.

Reliability of 2 methods in maxillary transverse deficiency diagnosis.

INTRODUCTION: The objective of this research was to evaluate the reliability of 2 methods (Andrews' Element III analysis and Yonsei transverse analysis) in maxillary transverse deficiency diagnosis. METHODS: Plaster casts and cone-beam computed tomography images of 80 outpatients with skeletal Class I malocclusion (29 males and 51 females, mean age, 20.16 ± 8.22 years) were selected. Maxillary and mandibular width were measured, respectively, and independently by 2 examiners at an interval of 2 weeks, using Andrews' Element III analysis and Yonsei transverse analysis. Intraclass correlation coefficients and Bland-Altman plots of intraexaminer and interexaminer reliability were evaluated. After diagnosis, Cohen's kappa statistics were calculated to evaluate the diagnostic agreement. RESULTS: The intraclass correlation coefficients were all above 0.85, indicating good to excellent reliability. Compared with Andrews' Element III analysis, Yonsei transverse analysis had higher intraexaminer and interexaminer reliability in both maxillary and mandibular width measurements. Thirty-one to 42 of the patients were diagnosed with maxillary transverse deficiency by 2 examiners using 2 methods. The intraexaminer and interexaminer Cohen's kappa values of Yonsei transverse analysis were all higher than those of Andrews' Element III analysis. CONCLUSIONS: Both Andrews' Element III analysis and Yonsei transverse analysis had good to excellent reliability and substantial diagnostic agreement. Yonsei transverse analysis had higher reliability in maxillary and mandibular width measurements and higher diagnostic agreement, compared with Andrews' Element III analysis.

Multicolor Tunable Polymeric Nanoparticle from the Tetraphenylethylene Cage for Temperature Sensing in Living Cells.

It is meaningful but challenging to develop a fluorescent probe for temperature sensing in living cells because it should possess the features of good cytocompatibility, easy read out, and high resolution. Herein, we successfully synthesized emissive star-like cage-based organic temperature-sensitive polymers that can assemble into nanoparticles in aqueous solution. The obtained nanoparticle can be easily tuned to full-color emission (including white light emission) with a temperature resolution of at least 0.5 °C by encapsulating different doses of guest dyes ((4-dimethylamino-2'-butoxychalcone (DMBC) and Nile Red (NR)) through a cascade Förster resonance energy transfer (FRET) effect. Moreover, the white light emission polymeric hybrid nanoparticles exhibit reversible stimuli response toward temperature and can be used as probes for temperature sensing in live cells through their fluorescent color variation between white and orange emission with good cytocompatibility.

Polyvinyl Alcohol-Supported AuAgNCs-CDs Film as a Selective Sensor for Gas Hydrogen Sulfide Detection in Air.

A novel strategy is reported for the synthesis of glutathione (GSH)-protected bimetallic nanoclusters, Au-AgNCs@GSH, and its fabrication with polyvinyl alcohol (PVA) into a film sensor for H2 S gas detection. Meanwhile, carbon dots (CDs) synthesized from polyethyleneimine (PEI) are introduced as an internal standard to correct the photobleaching of Au-AgNCs@GSH and uniformity of film. The joining of it greatly improves the chemical and structural stability of the composite film via multi cross-linking between PEI, PVA, and GSH. The PVA-AuAgNCs-CDs film exhibits an emission-quenching response to H2 S gas at atmosphere, which is highly repeatable, fast, sensitive, and can distinguish H2 S from other poisonous gases. Finally, the in-depth mechanism investigations reveal that the quenching response is attributed to decomposition of Au-AgNCs@GSH and the formation of Au2 S and Ag2 S in the composite film. As a sensor, the PVA-supported film combines the functions of fluorescent metal nanoclusters and polymer CDs, providing a portable device for the rapid detection of H2 S gas in air.

Ameloblastoma with mucous cells: A clinicopathological, BRAF mutation, and MAML2 rearrangement study.

OBJECTIVES: To investigate the clinicopathological features, BRAF V600E mutation, and MAML2 rearrangement of ameloblastoma with mucous cell differentiation. MATERIALS AND METHODS: Five cases of ameloblastoma with mucous cell differentiation were retrospectively studied. Clinicopathological features, BRAF V600E mutation, and MAML2 rearrangement were analyzed. Follow-up information was available for all cases. RESULTS: Of five cases, two cases were male and three were female, aged 18-55 years. Four cases were located in the mandible and one case in the maxilla. Histologically, four of the five cases (80%) presented with cystic features and three of the five cases (60%) with varying degrees of squamous metaplasia. The mucous cells were located in the epithelial islands or the luminal aspect of the cystic cavities. The BRAF V600E mutation was found in three of five cases (60%). All the cases showed no MAML2 rearrangement. Two cases were recurrent lesions, and one case had a local recurrence during the follow-up. CONCLUSIONS: Ameloblastoma with mucous cell differentiation is closely related to the cystic features, squamous metaplasia, and shows a high prevalence of BRAF V600E mutation. The absence of MAML2 rearrangement reveals that ameloblastoma with mucous cell differentiation and central mucoepidermoid carcinoma (MEC) are two distinct tumor entities.

Effects of Twin-block vs sagittal-guidance Twin-block appliance on alveolar bone around mandibular incisors in growing patients with Class II Division 1 malocclusion.

INTRODUCTION: The purpose of this study was to comparatively evaluate the effects of Twin-block (TB) appliance and sagittal-guidance Twin-block (SGTB) appliance on alveolar bone around mandibular incisors in growing patients with Class II Division 1 malocclusion, using cone-beam computed tomography. METHODS: The sample consisted of 25 growing patients with Class II Division 1 malocclusion (14 boys and 11 girls, mean age 11.92 ± 1.62 years) and was randomly distributed into the TB group (n = 13) and the SGTB group (n = 12). The treatment duration was 11.56 ± 1.73 months. Pretreatment (T1) and posttreatment (T2) cone-beam computed tomography scans were taken in both groups. Height, thickness at apex level, and volume of the alveolar bone around mandibular left central incisors were measured respectively on labial and lingual side, using Mimics software (version 19.0; Materialise, Leuven, Belgium). Based on the stable structures, 3-dimensional (3D) registrations of T1 and T2 models were taken to measure the sagittal displacement of incisors. Intragroup comparisons were evaluated by paired-samples t tests and Wilcoxon tests. Independent-samples t tests and Mann-Whitney U tests were used for intergroup comparisons. RESULTS: In both groups, alveolar bone height and volume on the labial side of the incisors significantly decreased after treatment (P <0.05). Lingual alveolar bone height, lingual and total alveolar bone volume, labial, lingual and total alveolar bone thickness showed no significant difference between T1 and T2 (P >0.05). In both groups the incisors tipped labially and drifted to the labial side. Compared with the TB group, less labial alveolar bone loss, less incisor proclination and crown edge drift were found in the SGTB group (P <0.05). CONCLUSIONS: Labial alveolar bone loss around mandibular incisors was observed after both types of appliances treatment in growing patients with Class II Division 1 malocclusion. Less labial alveolar bone loss, less incisor proclination, and crown edge drift were found in the SGTB group than in the TB group during treatment.

Homeobox protein MSX-1 inhibits expression of bone morphogenetic protein 2, bone morphogenetic protein 4, and lymphoid enhancer-binding factor 1 via Wnt/ß-catenin signaling to prevent differentiation of dental mesenchymal cells during the late bell stage.

Homeobox protein MSX-1 (hereafter referred to as MSX-1) is essential for early tooth-germ development. Tooth-germ development is arrested at bud stage in Msx1 knockout mice, which prompted us to study the functions of MSX-1 beyond this stage. Here, we investigated the roles of MSX-1 during late bell stage. Mesenchymal cells of the mandibular first molar were isolated from mice at embryonic day (E)17.5 and cultured in vitro. We determined the expression levels of ß-catenin, bone morphogenetic protein 2 (Bmp2), Bmp4, and lymphoid enhancer-binding factor 1 (Lef1) after knockdown or overexpression of Msx1. Our findings suggest that knockdown of Msx1 promoted expression of Bmp2, Bmp4, and Lef1, resulting in elevated differentiation of odontoblasts, which was rescued by blocking the expression of these genes. In contrast, overexpression of Msx1 decreased the expression of Bmp2, Bmp4, and Lef1, leading to a reduction in odontoblast differentiation. The regulation of Bmp2, Bmp4, and Lef1 by Msx1 was mediated by the Wnt/ß-catenin signaling pathway. Additionally, knockdown of Msx1 impaired cell proliferation and slowed S-phase progression, while overexpression of Msx1 also impaired cell proliferation and prolonged G1-phase progression. We therefore conclude that MSX-1 maintains cell proliferation by regulating transition of cells from G1-phase to S-phase and prevents odontoblast differentiation by inhibiting expression of Bmp2, Bmp4, and Lef1 at the late bell stage via the Wnt/ß-catenin signaling pathway.

Novel Bioconjugation Strategy Using Elevated Hydrostatic Pressure: A Case Study for the Site-Specific Attachment of Polyethylene Glycol (PEGylation) of Recombinant Human Ciliary Neurotrophic Factor.

In this paper, we reported a novel strategy for the site-specific attachment of polyethylene glycol (PEGylation) of proteins using elevated hydrostatic pressure. The process was similar to the conventional one except the reactor was under elevated hydrostatic pressure. The model protein was recombinant human ciliary neurotrophic factor (rhCNTF), and the reagent was monomethoxy-polyethylene glycol-maleimide (mPEG-MAL). PEGylation with mPEG (40 kDa)-MAL at pH 7.0 under normal pressure for 5 h achieved a less than 5% yield. In comparison, when the pressure was elevated, the PEGylation yield was increased dramatically, reaching nearly 90% at 250 MPa. Furthermore, the following phenomena were observed: (1) high-hydrostatic-pressure PEGylation (HHPP) could operate at a low reactant ratio of 1:1.2 (rhCNTF to mPEG-MAL), while the conventional process needs a much-higher ratio. (2) Short and long chains of PEG gave a similar yield of 90% in HHPP, while the conventional yield for the short chain of the PEG was higher than that of the long chain. (3) The reaction pH in the range of 7.0 to 8.0 had almost no influence upon the yield of HHPP, while the PEGylation yield was significantly increased by a factor of three from pH 7.0 to 8.0 at normal pressure. Surface accessibility analysis was performed using GRASP2 software, and we found that Cys17 of rhCNTF was located at the concave patches, which may have steric hindrance for the PEG to approach. The speculated benefit of HHPP was the facilitation of target-site exposure, reducing the steric hindrance and making the reaction much easier. Structure and activity analysis demonstrated that the HHPP product was comparable to the PEGylated rhCNTF prepared through a conventional method. Overall, this work demonstrated that HHPP, as we proposed, may have application potentials in various conjugations of biomacromolecules.

The In vitro/vivo Evaluation of Prepared Gastric Floating Tablets of Berberine Hydrochloride.

Currently available antiulcer drugs suffered from serious side effects which limited their uses and prompted the need for a safe and efficient new antiulcer agent. The objective of this project work was to retain the drug in the stomach for better antiulcer activity and less side effects. Hence, the aim of our present work was to prepare a gastric floating tablet of Berberine hydrochloride (Ber) with suitable in vitro/vivo properties. In this study, different Ber gastric floating tablets were prepared by simple direct compression using various amounts of HPMCK15M and Carbopol 971PNF combined with other tablet excipients. The properties of the tablets including hardness, buoyancy, swelling ability, in vitro drug release, and in vivo pharmacokinetic study were evaluated. The obtained results disclosed that hardness, floating, swelling, and in vitro drug release of the Ber tablets depended mainly on the ratio of polymer combinations. Moreover, among six formulations, F3 exhibited desirable floating, swelling, and extended drug release. In addition, in vivo pharmacokinetic study suggested that prepared gastric floating tablets had significantly sustained-releasing effects compared with market tablets. Therefore, the developed gastric floating tablets of Ber could be an alternative dosage form for treatment of gastrointestinal disease.

Evaluation of the Transverse Displacement of the Proximal Segment After Bilateral Sagittal Split Ramus Osteotomy With Different Lingual Split Patterns and Advancement Amounts Using the Finite Element Method.

PURPOSE: To evaluate transverse displacement of the proximal segment after bilateral sagittal split ramus osteotomy (BSSO) advancement with different lingual split patterns and advancement amounts and to determine the influential factors related to mandibular width. MATERIALS AND METHODS: A 3-dimensional finite element model of the mandible including the temporomandibular joint was created for a presurgical simulation and for BSSO with lingual split patterns I (T1; Hunsuck split) and II (T2; Obwegeser split). The mandible was advanced 3 mm (A3) and 8 mm (A8) and fixated with a conventional titanium plate. Ansys software was used to measure the linear distances of the interproximal segments and to analyze the transverse displacement distribution of proximal segments after applying the load of masticatory muscle force groups. RESULTS: After surgical simulation, T1A3, T1A8, T2A3, and T2A8 showed increased transverse widths (mean, 2.99, 4.70, 2.36, and 4.42 mm, respectively). For transverse augmentation, there was a statistically significant difference between the 2 different mandibular advancement amounts in T1 and in T2 (P ≤ .000), but no significant differences was observed between T1 and T2 (P ≥ .058). The maximum transverse displacement distribution in the proximal segment was measured around the gonial area, and the early contact area was found near the border between the horizontal and sagittal osteotomy lines. CONCLUSION: Transverse displacements of proximal segments occur after BSSO advancement with T1 and T2 and transverse augmentation has statistically meaningful effects depending on the amount of advancement; however, no differences in transverse augmentation between T1 and T2 were identified. The fulcrum caused by the early contact between the proximal and distal segments could be an influential factor related to mandibular width.

Selection of Biopsy Site for Patients with Systematic Amyloidosis.

Objective To evaluate the sensitivities of various biopsy methods for the diagnosis of systematic amyloidosis (SA). Methods The clinical data and biopsy results of 194 SA patients who were treated in Peking Union Medical College Hospital from January 2009 to June 2015 were retrospectively analyzed. Results The highest sensitivity was achieved by biopsy of affected organs,with renal biopsy 97.4%,heart biopsy 95.0% and liver biopsy 87.5%. Among non-invasive biopsy methods,tongue biopsy was found to be 75% sensitive,followed by gingiva biopsy at 57%,abdominal fat pad aspiration at 57%,rectum biopsy at 16%,and bone marrow examination at 8%. Combination of tongue and abdominal fat pad biopsy yielded a detection rate of 93.1%. Conclusions Biopsy of the involved organ has the highest sensitivity. However,combination of multiple non-invasive biopsy methods may has sensitivity comparable to organ biopsy and is safer and more convenient.

[Study on the acid hydrolysis, fiber remodeling and bionics mineralization of rat tail tendon collagen type â ].

Objective: To produce bionic bone material that is consistent with human bone in chemical composition and molecular structure using rat tail tendon collagen type Ⅰ. Methods: The type Ⅰcollagen derived from rat tail was extracted by acetic acid to form collagen fibers. The reconstructed collagen fibers were placed in the mineralized solution to mimic bone mineralization for 2-6 days. Bone mineralization was observed by transmission electron microscopy and electron diffraction.Results: Collagen fibers with characteristic D-Band structure were reconstructed by using rat tail tendon collagen type Ⅰ extracted with acid hydrolysis method. Transmission electron microscopy and electron diffraction showed that calcium hydroxyapatite precursor infiltrated into the collagen fibers, and the collagen fibers were partially mineralized after 2 days of mineralization; the collagen fibers were completely mineralized and bionic bone material of typeⅠ collagen/calcium hydroxyapatite was formed after 6 days of mineralization.Conclusion: The collagen type Ⅰ can be extracted from rat tail tendon by acid hydrolysis method, and can be reformed and mineralized to form the bionic bone material which mimics human bone in chemical composition and the molecular structure.

Loss of PEG chain in routine SDS-PAGE analysis of PEG-maleimide modified protein.

SDS-PAGE represents a quick and simple method for qualitative and quantitative analysis of protein and protein-containing conjugates, mostly pegylated proteins. PEG-maleimide (MAL) is frequently used to site-specifically pegylate therapeutic proteins via free cysteine residue by forming a thiosuccinimide structure for pursuing homogeneous products. The C-S linkage between protein and PEG-MAL is generally thought to be relatively stable. However, loss of intact PEG chain in routine SDS-PAGE analysis of PEG-maleimide modified protein was observed. It is a thiol-independent thioether cleavage and the shedding of PEG chain exclusively happens to PEG-MAL modified conjugates although PEG-vinylsulfone conjugates to thiol-containing proteins also through a C-S linkage. Cleavage kinetics of PEG40k-MAL modified ciliary neurotrophic factor showed this kind of degradation could immediately happen even in 1 min incubation at high temperature and could be detected at physiological temperature and pH, although the rate was relatively slow. This may provide another degradation route for maleimide-thiol conjugate irrespective of reactive thiol, although the specific mechanism is still not very clear for us. It would also offer a basis for accurate characterization of PEG-MAL modified protein/peptide by SDS-PAGE analysis.