Molecular epidemiology and characterization of Streptococcus mutans strains in Korea / 대한구강보건학회지

OBJECTIVES: We investigated the characteristics of Streptococcus mutans in the national culture collection from Korea. Twenty-nine (dental plaque, n=27; endodontic infections, n=1; blood, n=1) isolates were included in this study.METHODS: Antimicrobial susceptibilities were tested using the disk diffusion test. Multilocus sequence typing (MLST), serotyping, and collagen-binding genes were used for polymerase chain reaction (PCR) and direct sequencing. A collagen-binding (to assess the adhesion properties) assay was performed. S. mutans demonstrated high susceptibility to antimicrobial agents. Differences in collagen-binding abilities of the cnm-positive and -negative groups were compared using the Mann-Whitney U test (P<0.05).RESULTS: MLST analyses revealed 25 sequence types (STs), 17 of which (ST213-ST229) contained new alleles. The strains were classified into four serotypes with the c type encompassing 79.3% of all strains, while the e, f, and k types representing 6.9% each. Analysis of the cnm and cbm genes, which encode the two surface adhesin components of S. mutans, revealed three cnm-positive strains, each displaying greater adhesion ability than those of the cnm-negative strains.CONCLUSIONS: This study highlights the presence of a wide variety of S. mutans genotypes in Korea. These findings may provide useful information regarding the pathogenesis of infectious diseases, such as dental caries.

Promotion of transplanted collagen scaffolds combined with brain-derived neurotrophic factor for axonal regeneration and motor function recovery in rats after transected spinal cord injury / 中国修复重建外科杂志

Objective: To evaluate the effect of the combination of collagen scaffold and brain-derived neurotrophic factor (BDNF) on the repair of transected spinal cord injury in rats. Methods: Thirty-two Sprague-Dawley rats were randomly divided into 4 groups: group A (sham operation group), T 9, T 10 segments of the spinal cord was only exposed; group B, 4-mm T 9, T 10 segments of the spinal cord were resected; group C, 4-mm T 9, T 10 segments of the spinal cord were resected and linear ordered collagen scaffolds (LOCS) with corresponding length was transplanted into lesion site; group D, 4-mm T 9, T 10 segments of the spinal cord were resected and LOCS with collagen binding domain (CBD)-BDNF was transplanted into lesion site. During 3 months after operation, Basso-Beattie-Bresnahan (BBB) locomotor score assessment was performed for each rat once a week. At 3 months after operation, electrophysiological test of motor evoked potential (MEP) was performed for rats in each group. Subsequently, retrograde tracing was performed for each rat by injection of fluorogold (FG) at the L 2 spinal cord below the injury level. One week later, brains and spinal cord tissues of rats were collected. Morphological observation was performed to spinal cord tissues after dehydration. The thoracic spinal cords including lesion area were collected and sliced horizontally. Thoracic spinal cords 1 cm above lesion area and lumbar spinal cords 1 cm below lesion area were collected and sliced coronally. Coronal spinal cord tissue sections were observed by the laser confocal scanning microscope and calculated the integral absorbance ( IA) value of FG-positive cells. Horizontal tissue sections of thoracic spinal cord underwent immunofluorescence staining to observe the building of transected spinal cord injury model, axonal regeneration in damaged area, and synapse formation of regenerated axons. Results: During 3 months after operation, the BBB scores of groups B, C, and D were significantly lower than those of group A ( P0.05). Immunofluorescence staining results of spinal cord tissue sections selected from dorsal to ventral spinal cord showed transected injured areas of spinal cords which were significantly different from normal tissues. The numbers of NF-positive axons in lesion center of group A were significantly larger than those of groups B, C, and D ( P<0.05), and in groups C and D than in group B ( P<0.05), and in group D than in group C ( P<0.05). Conclusion: The combined therapeutic approach containing LOCS and CBD-BDNF can promote axonal regeneration and recovery of hind limb motor function after transected spinal cord injury in rats.

Expression,purification and renaturation of recombinant human collagen-binding bone morphogenetic protein-2 from Escherichia coli / 吉林大学学报(医学版)

Objective:To construct the Escherichia coli (E. coli)expression system for preparation of the bone morphogenetic protein-2 (BMP2)with collagen-binding domain (CBD),and to study the methods and conditions for expression, purification and renaturation of CBD-BMP2.Methods:CBD sequence was cloned into the N-terminal of BMP2 sequence, the recombinant vector pet21b/CBD-BMP2 was constructed and transformed into E.coli BL21.The expression of recombinant protein was induced using isopropylβ-D-thiogalactopyranoside (IPTG) at 37 ℃.Ni-NTA chelate chromatography was used to purify CBD-BMP-2.Denaturing CBD-BMP2 was refolded by dilution method using ultrapure water.The refolding CBD-BMP2 was filtered through a 0.22μm microfiltration membrane for degermation.The recovery rate was calculated by the ratio of the protein concentration before and after degermation. The expression, purification, and renaturation of recombinant protein were detected by SDS-PAGE method.The concentration of CBD-BMP2 was detected by BCA assay.Results:The recombinant vector pet21b/CBD-BMP2 was successfully transformed into E.coli BL21,and the recombinant protein was expressed as inclusion bodies in E.coli.The SDS-PAGE results showed denaturing protein was dissolved in supernatant of lysis buffer with 8 mol·L-1 urea and the purified recombinant protein existed in elution buffer B with relative molecular mass about 14 000.Two bands (14 000 and 28 000)were seen in the SDS-PAGE picture,which indicated that the monomer was successfully refolded into dimer by dilution method.The concentrations of recombinant protein before and after degermation were 110 and 80 mg · L-1 , respectively, and the recovery rate was about 73%. Conclusion:The recombinant vector pet21b/CBD-BMP2 is transformed into E.coli BL21 successfully,and the recombinant CBD-BMP2 is expressed and refolded efficiently. The methods of prokaryotic expression system for preparing recombinant CBD-BMP2 protein are established.

Experimental study on the periodontal regenerative capacity of moldable synthetic peptide domain gel in degree III furcation defect of beagles / 대한치주과학회지

PURPOSE: Osteopontin is one of the major non-collagenous protein of hard tissue. Use of peptide domain of biologically active protein has some advantages. The objective of this experimental study is evaluation of periodontal regenerative potency of synthetic peptide gel which containing collagen binding domain of osteopontin in the degree III periodontal defect of beagle dogs. MATERIAL AND METHODS: Experimental degree III furcation defect was made in the mandibular third and fourth premolar of beagles. Regenerative material was applied during flap operation. 8 weeks after regenerative surgery, all animals were sacrificed and histomorphometric measurement was performed to calculate the linear percentage of the new cementum formation and the volume percentage of new bone formation. RESULT: The linear percent of new cementum formation was 41.6% at control group and 67.1% at test group and there was statistically significant difference. The volume percent of new bone formation was 52.1% at control group and 58.9% at test group. CONCLUSION: As the results of present experiment, synthetic peptide gel containing collagen binding domain of osteopontin significantly increase new bone and cementum formation in the degree III furcation defect of canine mandible.

Establishment and clinical application of collagen binding assay for von Willebrand factor / 中华检验医学杂志

Objective To establish a new methA of detecting vWF function. Methas The capability of vWF to bind collagen was evaluated with ELISA. Results The assay′s sensitivity was 0.001 U/ml. Coefficient of variation for inner-batch and inter-batch were 3.34 and 6.70 respectively.The vWF:CBA value of plasma was(90.24?22.87)% in 20 normal subjects. The vWF:CBA value was (31.94?27.36)% in 54 vWD, (35.22?20.02)% in 10 type 1 vWD, (8.74?6.38)% in 10 type 2A vWD and (0.70?0.58)% in 6 type 3 vWD,the values of all four vWD groups were lower than that of normal group( P