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Dong Han

Publications and source records attributed to Dong Han.

At least 19 recordsLinked to original sources
PubMed2026

Remdesivir maintains antiviral potency against clinically relevant SARS-CoV-2 Nsp12 substitutions.

Remdesivir (RDV) is a nucleotide analog prodrug approved for COVID-19 treatment that inhibits the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp; nsp12). Although RDV maintains activity against circulating variants of concern, ongoing evaluation of resistance-associated substitutions is critical for clinical care, particularly in settings of prolonged viral replication such as immunocompromised individuals. We assessed the phenotypic impact of nsp12 substitutions identified from in vitro resistance selection, RDV clinical reports, and global sequence surveillance. Using a recombinant infectious SARS-CoV-2 reporter virus, we compared susceptibility of these nsp12 substitutions to RDV and its parent nucleoside, GS-441524. After confirming concordant resistance profiles between RDV and GS-441524, we assessed RDV susceptibility in a complementary non-infectious replicon system. In both systems, single nsp12 substitutions remained fully susceptible to RDV within their respective assay variability limits. Of the double substitutions tested, S759A/V792I conferred the largest reduction in antiviral susceptibility (∼15-fold) but was associated with impaired replication kinetics. Given the strong concordance between the two assays, the replicon system also enabled phenotypic characterization of substitutions E802A, E802D, and P323L/E802D that could not be rescued as infectious virus. Analysis of >17 million SARS-CoV-2 genomes in GISAID showed that all tested nsp12 substitutions had low prevalence (≤0.1%), except P323L (98.8%). Collectively, these data reinforce the high genetic barrier to RDV resistance, as reduced susceptibility is typically accompanied by substantial reductions in replication. Our findings support the continued clinical utility of RDV and highlight the complementary value of SARS-CoV-2 infectious virus and replicon systems for antiviral resistance surveillance and phenotyping.

COVID-19↗
PubMed2026

Adaptive Evolution of the PFK Gene Family in Chinese Longsnout Catfish, Leiocassis longirostris.

The Chinese longsnout catfish is a typical carnivorous fish with a relatively weak ability to utilize glucose. However, the genomic basis for its glucose metabolic adaptation remains unclear. In this study, we used comparative genomics methods to systematically analyze the evolutionary characteristics of glucose metabolism-related genes in the Chinese longsnout catfish, focusing on gene family evolution, patterns of expansion and contraction, and selective pressures. The results indicate that glucose metabolism-related genes have undergone significant reshaping during evolution. Genes involved in glucose digestion, absorption, and insulin signaling pathways demonstrate a tendency toward contraction, while those associated with protein and lipid metabolism exhibit expansion. This pattern is consistent with the species' long-term adaptation to a high-protein, high-fat diet. Comparative analysis further revealed that, compared to fish with different dietary habits, certain key genes involved in glycolysis in the Chinese longsnout catfish exhibit a reduction in copy number. Molecular evolutionary analysis showed that key genes involved in glycolysis and gluconeogenesis (including hexokinase 2 (hk2), phosphofructokinase, muscle/platelet (pfkm/p)) exhibit signs of accelerated evolution or positive selection. Notably, the PFK gene family exhibits complex evolutionary characteristics resulting from the combined effects of gene contraction, rapid evolution, and positive selection. In summary, this study reveals the genomic evolutionary basis for the glucose metabolic adaptation of the Chinese longsnout catfish and identifies the PFK gene family as a key candidate for elucidating its unique glucose metabolic characteristics.

Leiocassis longirostris↗
PubMed2026

Prenatal SNP-array chromosomal microarray analysis in 3,549 pregnancies: indication-specific yields and clinical implications.

BACKGROUND: SNP-based chromosomal microarray analysis (CMA) is widely used in invasive prenatal diagnosis, yet real-world performance across contemporary referral pathways, especially in the NIPT era, remains incompletely characterized. METHODS: We retrospectively analyzed 3,549 prenatal invasive samples tested by SNP array, and evaluated diagnostic yield overall and by referral indication and ultrasound phenotype. RESULTS: In total, we identified 398 pathogenic or likely pathogenic (P/LP) variants across 386 fetuses, resulting in an overall diagnostic yield of 10.9% (386/3,549). These findings comprised 223 aneuploidies and 175 pathogenic CNVs. In contrast, variants of uncertain significance (VOUS) were detected in 12.0% (426/3,549) of cases. Diagnostic yields were heavily stratified by indication: yields peaked in NIPT high-risk referrals (38.9%) and were intermediate in ultrasound-based cases (~ 11%), but dropped significantly in the advanced maternal age (AMA; 4.2%) and serum screening (~ 5-6%) groups. Conversely, VOUS rates remained remarkably stable across all referral categories. Sub-analysis of ultrasound abnormalities revealed that multisystem anomalies conferred the highest risk (27.3%), driven predominantly by aneuploidies; among soft markers, increased nuchal translucency (NT) emerged as the strongest predictor of chromosomal pathology. CONCLUSIONS: In our cohort, SNP-array identified clinically actionable findings in 10.9% of cases. NIPT enriched diagnostic yields, particularly for aneuploidies, and NT thickness was strongly associated with pathogenic findings. These results support an indication-based approach to genomic testing, with NIPT as a triage tool for aneuploidy and CMA for high-risk populations, while improving VOUS counseling.

Humans↗
PubMed2024

No Remdesivir Resistance Observed in the Phase 3 Severe and Moderate COVID-19 SIMPLE Trials.

Remdesivir (RDV) is a broad-spectrum nucleotide analog prodrug approved for the treatment of COVID-19 in hospitalized and non-hospitalized patients with clinical benefit demonstrated in multiple Phase 3 trials. Here we present SARS-CoV-2 resistance analyses from the Phase 3 SIMPLE clinical studies evaluating RDV in hospitalized participants with severe or moderate COVID-19 disease. The severe and moderate studies enrolled participants with radiologic evidence of pneumonia and a room-air oxygen saturation of ≤94% or >94%, respectively. Virology sample collection was optional in the study protocols. Sequencing and related viral load data were obtained retrospectively from participants at a subset of study sites with local sequencing capabilities (10 of 183 sites) at timepoints with detectable viral load. Among participants with both baseline and post-baseline sequencing data treated with RDV, emergent Nsp12 substitutions were observed in 4 of 19 (21%) participants in the severe study and none of the 2 participants in the moderate study. The following 5 substitutions emerged: T76I, A526V, A554V, E665K, and C697F. The substitutions T76I, A526V, A554V, and C697F had an EC50 fold change of ≤1.5 relative to the wildtype reference using a SARS-CoV-2 subgenomic replicon system, indicating no significant change in the susceptibility to RDV. The phenotyping of E665K could not be determined due to a lack of replication. These data reveal no evidence of relevant resistance emergence and further confirm the established efficacy profile of RDV with a high resistance barrier in COVID-19 patients.

Humans↗
PubMed2006

Inhibition of MLK3-MKK4/7-JNK1/2 pathway by Akt1 in exogenous estrogen-induced neuroprotection against transient global cerebral ischemia by a non-genomic mechanism in male rats.

Numerous studies have demonstrated the neuroprotective effects of estrogen in experimental cerebral ischemia. To investigate molecular mechanisms of estrogen neuroprotection in global ischemia, immunoblotting, immunohistochemistry and Nissel-staining analysis were used. Our results showed that chronic pretreatment with beta-estradiol 3-benzoate (E2) enhanced Akt1 activation and reduced the activation of mixed-lineage kinase 3 (MLK3), mitogen-activated protein kinase kinase 4/7 (MKK4/7), and c-Jun N-terminal kinase 1/2 (JNK1/2) in the hippocampal CA1 subfield during reperfusion after 15 min of global ischemia. In addition, E2 reduced downstream JNK nuclear and non-nuclear components, c-Jun and Bcl-2 phosphorylation and Fas ligand protein expression induced by ischemia/reperfusion. Administration of phosphoinositide 3-kinase (PI3K) inhibitor LY 294,002 prevented both activation of Akt1 and inhibition of MLK3, MKK4/7 and JNK1/2. The interaction between ERalpha and the p85 subunit of PI3K was also examined. E2 and antiestrogen ICI 182,780 promoted and prevented this interaction, respectively. Furthermore, ICI 182,780 blocked both the activation of Akt1 and the inhibition of MLK3, MKK4/7 and JNK1/2. Photomicrographs of cresyl violet-stained brain sections showed that E2 reduced CA1 neuron loss after 5 days of reperfusion, which was abolished by ICI 182,780 and LY 294,002. Our data indicate that in response to estrogen, ERalpha interacts with PI3K to activate Akt1, which may inhibit the MLK3-MKK4/7-JNK1/2 pathway to protect hippocampal CA1 neurons against global cerebral ischemia in male rats.

Animals↗
PubMed2006

MAC mode atomic force microscopy studies of living samples, ranging from cells to fresh tissue.

Magnetic AC mode (MAC mode) atomic force microscopy (AFM), a novel type of tapping mode AFM in which the cantilever is driven directly by a magnetic field, is a powerful tool for imaging with high spatial resolution and better signal-to-noise in liquid environment. It may largely extend the application of AFM to living samples, especially those are sensitive to cantilever forces, even to multilayer tissue samples. However, there are few reports on the imaging of living cells by MAC mode AFM previously. In our present study, we explore the optimal imaging conditions of MAC mode AFM on living astrocytes and fresh arterial intima surface. We also used nude tips for PicoTREC panel (i.e., Aux in BNC, a new data collecting channel) to image living samples and discussed its difference with phase imaging. We show that living biological samples can be imaged by MAC mode AFM at details of comparable resolution as those by high resolution scanning electron microscopy. Furthermore, the combination of height, amplitude, phase and TREC panel signals provide abundant informations for the characteristics of living samples, such as topography, profile, stiffness and adhesion.

Animals↗
PubMed2006

Akt inhibits MLK3/JNK3 signaling by inactivating Rac1: a protective mechanism against ischemic brain injury.

The overall goal of this study was to determine the molecular basis by which mixed-lineage kinase 3 (MLK3) kinase and its signaling pathways are negatively regulated by the pro-survival Akt pathway in cerebral ischemia. We demonstrated that tyrosine phosphorylation of the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) underlies the increased Akt-Ser473 phosphorylation by orthovanadate. Co-immunoprecipitation analysis revealed that endogenous Akt physically interacts with Rac1 in the hippocampal CA1 region, and this interaction is promoted on tyrosine phosphatase inhibition. The elevated Akt activation can deactivate MLK3 by phosphorylation at the Ser71 residue of Rac1, a small Rho family of guanidine triphosphatases required for MLK3 autophosphorylation. Subsequently, inhibition of c-Jun N-terminal kinase 3 (JNK3) results in decreased serine phosphorylation of 14-3-3, a cytoplasmic anchor of Bax, and prevents ischemia-induced mitochondrial translocation of Bax, release of cytochrome c and activation of caspase 3. At the same time, the expression of Fas-ligand decreases in the CA1 region after inhibition of c-Jun activation. The neuroprotective effect of Akt activation is significant in the CA1 region after global cerebral ischemia. Our results suggest that the activation of the pro-apoptotic MLK3/JNK3 cascade induced by ischemic stress can be suppressed through activation of the anti-apoptotic phosphatidylinositol 3-kinase/Akt pathway, which provides a direct link between Akt and the family of stress-activated kinases.

14-3-3 Proteins↗
PubMed2006

Functional divergence of six isoforms of antifungal peptide Drosomycin in Drosophila melanogaster.

Drosomycin (Drs) gene encodes a 44-residue inducible antifungal peptide, Drosomycin, in Drosophila melanogaster. Six genes, Drs-lC, Drs-lD, Drs-lE, Drs-lF, Drs-lG and Drs-lI, show homology to the Drs form in a multigene family on the 3rd chromosome of D. melanogaster. It is the first experimental demonstration that the six members in the Drs family act as functional genes. To further delineate the functional divergence of these six members, their cDNA sequences were cloned respectively into the pET-3C vector and expressed in the E. coli. The antifungal activity of the expression products was assayed using the Cerletti's method. The results showed a difference among the six isoforms in antifungal activity against the tested fungal strains: in which Drs was most effective and showed antifungal activity to all seven fungal strains, whereas isoform Drs-lC was effective to six strains, Drs-lD was effective to five strains, Drs-lG was effective to four strains, and Drs-lE and Drs-lF were effective to only three strains. Drs-lI had no activity against any tested fungal strains. By comparing the variable residue sites of these six isoforms to that of Drosomycin in the three-dimensional structure, we suggested that the reduction in the antifungal activity was due to the variable residues that were not in the alpha-helix. In addition, two inserted residues (RV) in Drs-lI may affect the dimensional structure and resulted in a functional change. These results may explain the evolution of the Drosomycin multigene family and its functional divergence.

Amino Acid Sequence↗
PubMed2006

Biomechanopharmacology: a new borderline discipline.

Flowing blood is more than a drug transporter in pharmacology; its mechanical impact should also be considered. The in vitro pharmacological dose-response pattern of endothelial cellular functions can be significantly modified by in vivo shear stress. A new borderline discipline, biomechanopharmacology, is forming at the boundary between biomechanics and pharmacology. Biomechanopharmacology will probably consist of both the pharmacological intervention of signals induced by biomechanical factors and the biomechanical influence on pharmacokinetics and pharmacodynamics, in addition to the joint effect of biomechanical and pharmacological factors. Recent investigations show that exercise enhances the shear of pulsatile blood flow to stimulate angiogenesis. The benefits of exercise for gaining joint biomechanical and pharmacological effects should be emphasized.

Biomechanical Phenomena↗
PubMed2006

NMDA receptor/L-VGCC-dependent expression and AMPA/KA receptor-dependent activation of c-Jun induced by cerebral ischemia in rat hippocampus.

Over-activation of ionotropic glutamate receptors can cause an excessive influx of calcium ions into neurons, which subsequently triggers the degeneration and death of cells in a process known as excitotoxicity. Here, we examined the effects of modulating ionotropic glutamate receptors and L-type voltage-gated calcium channels (L-VGCC) on the expression and activation of c-Jun in hippocampus of SD rats after transient global ischemia. The total protein of c-Jun was altered by ischemia-reperfusion and reached its high levels at 3-6 h of reperfusion. However, the increased expression was prevented by pretreatment of ketamine (a non-competitive N-methyl-D-aspartate (NMDA) receptors antagonist) or nifedipine (a blocker of L-VGCC), but not by 6,7-dinitroquinoxaline-2,3(1H,4H)-dione (DNQX), an AMPA/KA receptor antagonist. On the other hand, c-Jun phosphorylation was significantly increased 3 h after reperfusion, which was inhibited by DNQX, but not ketamine or nifedipine. AP-1 binding activity reactions were also performed by electrophoretic mobility shift assay (EMSA), which detected similar results as those in Western blotting. Our results clearly showed that c-Jun expression is NMDA receptor/L-VGCC-dependent and c-Jun activation is AMPA/KA receptor-dependent, which expands our knowledge of the JNK-c-Jun signaling pathway in ischemic brain damage.

Animals↗
PubMed2006

Cytoskeletal response of microvessel endothelial cells to an applied stress force at the submicrometer scale studied by atomic force microscopy.

Cytoskeleton fibers form an intricate three-dimensional network to provide structure and function to microvessel endothelial cells. During accommodation to blood flowing, stress fiber bundles become more prominent and align with the direction of blood flow. This network either mechanically resists the applied shear stress (lateral force) or, if deformed, is dynamically remodeled back to a preferred architecture. However, the detailed response of these stress fiber bundles to applied lateral force at submicrometer scales are as yet poorly understood. In our in vitro study, the tip, topography probe in lateral force microscopy of atomic force microscopy, acted as a tool for exerting quantitative vertical and lateral force on the filaments of the cytoskeleton. Moreover, the authors developed a formula to calculate the value of lateral force exerted on every point of the filaments. The results show that cytoskeleton fibers of healthy tight junctions in rat cerebral microvessel endothelial cells formed a cross-type network, and were reinforced and elongated in the direction of scanning under lateral force of 15-42 nN. Under peroxidation (H(2)O(2) of 300 micromol/L), the cytoskeleton remodeled at intercellular junctions, and changed over the meshwork structures into a dense bundle, that redistributed the stress. Once mechanical forces were exerted on an area, the cells shrank and lost morphologic tight junctions. It would be useful in our understanding of certain pathological processes, such as cerebral ischemia/reperfusion injury, which maybe caused by biomechanical forces and which are overlooked in current disease models.

Animals↗
PubMed2006

Factors affecting the efficiency and reversibility of roscovitine (ROS) block on the meiotic resumption of goat oocytes.

Goat oocytes from 2 to 4 and 0.8 to 1.2-mm follicles were freed (DOs) or not (COCs) of cumulus cells and cultured for different times in an inhibition medium supplemented with different concentrations of roscovitine (ROS). At the end of culture, oocytes were either cultured in a maturation medium for 24 hr and activated chemically for embryo development, or examined for GV chromatin configurations. Nuclear status was checked at different time points during maturation culture. Although both 200 and 250 microM ROS maintained 78-85% of oocytes at the GV stage for 24 hr, only oocytes blocked with 200 microM ROS developed to MII stage at a high rate after maturation culture. While few oocytes blocked with 200 microM ROS for 24 hr developed into morulae and none into blastocysts after activation, percentages of oocytes developing into morulae and blastocysts increased to the level of the control oocytes when the block time was reduced to 8 hr. While the GV and pMI stages were shortened with MI, and A/TI unaffected after oocytes were blocked for 8 hr, all the stages but A/TI were shortened after 24 hr of block. The sizes of nucleoli diminished with time and the GV chromatin configuration changed during ROS block. Significantly more DOs than COCs were blocked with 200 microM ROS, but none of the blocked DOs matured after drug withdrawal. However, maturation of the DOs improved significantly when ROS concentration was reduced to 150 microM or DOs were co-inhibited with COCs. The GV intact percentages of DOs did not differ after ROS inhibition with or without eCG, but those of COCs decreased significantly after ROS inhibited in the presence of eCG. When MII-incompetent oocytes from 0.8 to 1.2-mm follicles were inhibited with ROS for 8 and 24 hr prior to maturation culture, nuclear maturation improved significantly, activation rates were as high as that of the control oocytes, and some of the activated developed to 4- or 8-cell stages. It is concluded that (i) the efficiency and reversibility of ROS block was both drug concentration and exposure-time dependent; (ii) cumulus cells alleviated the toxicity of ROS on goat oocytes; (iii) eCG released goat oocytes from ROS block through the mediation of cumulus cells; (iv) ROS block quickened the nuclear maturation of goat oocytes and improved the developmental competence of meiosis-incompetent oocytes, possibly due to a sustained nuclear activity during inhibition culture; (v) oocyte nuclear maturation and activation did not depend upon cumulus expansion, but the embryo development occurred in association with cumulus expansion.

Animals↗
PubMed2006

Production of cloned goats by nuclear transfer of cumulus cells and long-term cultured fetal fibroblast cells into abattoir-derived oocytes.

Dairy goats are ideal for the transgenic production of therapeutic recombinant proteins. The use of recombinant somatic cell lines for nuclear transfer (NT) allows the introduction of genes by transfection, increases the efficiency of transgenic animal production to 100%, and overcomes the problem of founder mosaicism. Although viable animals have been cloned via NT from somatic cells of 11 species, the efficiency has been extremely low. Both blastomere and somatic cell NT increased fetal loss and perinatal morbidity/mortality in cattle and sheep, but fetal loss and perinatal mortality appear to be relatively low in goats. In this study, we produced cloned goats by NT from cumulus cells and long-term cultured fetal fibroblast cells (FFCs) to abattoir-derived oocytes. NT embryos were constructed from electrofusion of cumulus cells (CCs), FFCs, or skin fibroblast cells (SFCs) with cytoplasts prepared from abattoir-derived ovaries. The NT embryos were activated with an optimized activating protocol (1 min exposure to 2.5 microM ionomycin followed by 2 hr incubation in 2mM 6-DMAP). Two viable cloned kids from CCs and one from long-term cultured FFCs (at passage 20-25) were born. Microsatellite analysis of 10 markers confirmed that all cloned offspring were derived from corresponding donor cells. To our knowledge, the production of cloned goat offspring using abattoir-derived oocytes receiving nuclei from CCs and long-term cultured FFCs has not been reported. The production of viable cloned animals after activation with reduced intensity of ionomycin and 6-DMAP treatment has also not been reported. Loss of cloned embryos was obvious after 45 and 90 days of pregnancy, and a lack of cotyledons, heart defects, and improperly closed abdominal wall were observed in the aborted fetuses and one cloned kid. The fusibility and in vitro developmental potential of embryos reconstructed from FFCs at passage 20-25 were significantly lower than those of embryos reconstructed from FFCs at passage 3-5, and the cloning efficiency of the long-term cultured cells was low (0.5%).

Abattoirs↗
PubMed2006

Interactive effects of granulosa cell apoptosis, follicle size, cumulus-oocyte complex morphology, and cumulus expansion on the developmental competence of goat oocytes: a study using the well-in-drop culture system.

Using a well-in-drop (WID) oocyte/embryo culture system that allows identification of follicular origin, we have investigated the effects of granulosa cells (GCs) apoptosis, follicle size, cumulus-oocyte complexes (COCs) morphology, and cumulus expansion on the developmental competence of goat oocytes matured and cultured individually following parthenogenetic activation. The WID system supported oocyte maturation and embryo development to a level similar to the conventional group system. The majority of goat oocytes acquired competence for development up to the 8-16 cell stage in follicles larger than 2 mm, but did not gain the ability to form morula/blastocyst (M/Bs) until follicles larger than 3 mm in diameter. The extent of atresia affected M/Bs formation. This effect varied according to the follicle size. Cumulus expansion increased with follicle size and decreased with increasing incidence of GCs apoptosis. Oocyte developmental potential was also correlated with cumulus expansion. Regardless of the degree of follicle atresia, 73-84% of the floating cells in the follicular fluid (FF) underwent apoptosis. Correlation between floating cell density in FF and oocyte developmental potency suggests the possibility to use the floating cell density as a simple and non-invasive marker for oocyte quality. It is concluded that the developmental potential of an oocyte is determined by multifactor interactions, and multiple factors must be considered together to accurately predict the quality of an oocyte.

Animals↗
PubMed2006

[ICCD-based fast fluorescence micro-imaging technique and preliminary application in living cells].

A fast fluorescence micro-imaging system using mainly intensified charge couple device (ICCD), argon-ion laser, and xenon lamp was set up, and its preliminary application in living cells was presented. Real-time observation and imaging of fast concentration and distribution changing of intracellular Ca2+ labeled by Fluo-3, a fluorescent Ca2+ indicator, in the proliferation process of rat cerebral micro-vessels endothelial cells (rCMECs) were carried out, and curves of artificial intensity versus imaging sequence of four typical points were obtained. It is shown that the ICCD-based fast fluorescence micro-imaging system is a powerful tool for recording the real-time fast processes in living cells.

Animals↗
PubMed2006

[Effects of BMP-2 gene therapy on vascularization in repairing bone defects].

OBJECTIVE: To study the effects of BMP-2 gene therapy on vascularization in repairing bone defects. METHODS: The isolated rabbit mesenchymal stem cells (rBMSC), after being transfected by adenovirus carrying BMP-2 gene (Ad-BMP-2) and seeded on xenogeneic bone scaffolds, were used to repair 1.5 cm-long radius bone defects. Five methods were in use in the experiments: Ad-BMP-2 infected rBMSC plus antigen-free bovine cancellous bone (BCB, Group A), rBMSC-BCB plus reconstructed hBMP-2 (Group B1), Ad-LacZ infected rBMSC-BCB (Group C), rBMSC-BCB (Group D) and only BCB scaffolds (Group E). After 4, 8, and 12 weeks of the operations, capillary vessel ink infusion, vascular endothelial growth factor ( VEGF) immunohistochemical staining and histological examination were conducted. RESULTS: After 4 weeks of the operations, usually in Group A one newly formed artery was found in every pore between the trabeculae of the BCB. The density of these intraosseous vessels was high in the periphery and decreasing towards the center of the grafts; by transmission electron microscopy, osteoblasts were always next to vascular endothelial cells and gradually developed into osteocytes with the increase of capillary vessel; VEGF expression were apparently enhanced in mesenchymocytes. CONCLUSIONS: BMP-2 gene therapy, by up-regulating VEGF expression, indirectly induces vascularization of grafts and is of great value to the treatment of bone in union and bone defects.

Animals↗
PubMed2006

[Vascularization in transplantation of gene modified tissue engineered bone for repairing bone defect].

OBJECTIVE: To study the vascularization of the composite of bone morphogenetic protein 2 (BMP-2) gene transfected marrow mesenchymal stem cells (MSCs) and biodegradable scaffolds in repairing bone defect. METHODS: Adenovirus vector carrying BMP-2 (Ad-BMP-2) gene transfected MSCs and gene modified tissue engineered bone was constructed. The 1.5 cm radial defect models were made on 60 rabbits, which were evenly divided into 4 groups randomly (n= 15, 30 sides). Different materials were used in 4 groups: Ad-BMP-2 transfected MSCs plus PLA/PCL (group A), Ad-Lacz transfected MSCs plus PLA/PCL (group B), MSCs plus PLA/PCL (group C) and only PLA/PCL scaffolds (group D). The X-ray, capillary vessel ink infusion, histology, TEM, VEGF expression and microvascular density counting (MVD) were made 4, 8, and 12 weeks after operation. RESULTS: In group A after 4 weeks, foliated formed bones image was observed in the transplanted bones, new vessels grew into the bones, the pores of scaffolds were filled with cartilage callus, osteoblasts with active function grew around the microvessels, and VEGF expression and the number of microvessels were significantly superior to those of other groups, showing statistically significant difference (P<0.01); after 8 weeks, increasingly more new bones grew in the transplanted bones, microvessels distended and connected with each other, cartilage callus changed into trabecular bones; after 12 weeks, lamellar bone became successive, marrow cavity recanalized, microvessels showed orderly longitudinal arrangement. In groups B and C, the capability of bone formation was weak, the regeneration of blood vessels was slow, after 12 weeks, defects were mostly repaired, microvessels grew among the new trabecular bones. In group D, few new vessels were observed at each time, after 12 weeks, broken ends became hardened, the defected area was filled with fibrous tissue. CONCLUSION: BMP-2 gene therapy, by up-regulating VEGF expression, indirectly induces vascularization of grafts, promotes the living of seed cells, and thus accelerates new bone formation.

Animals↗