Cultivation, screening, identification and transplantation of Muse cell from human umbilical cord-derived for spinal cord injury in rats / 中国骨伤

OBJECTIVE@#To investigate multilineage-differentiating stress-enduring (Muse) by immunomagnetic bead screening from Wharton's jelly mesenchymal stromal cells(WJ-MSCs), and explore transplantation of Muse cell for safety and effectivensess of sub acute cord injury in rats.@*METHODS@#Donated Wharton's Jelly-mesenchymal stromal cells (WJ-MSCs) were successfully derived from a human umbilical cord by a series of procedures namely physical isolation of Wharton's Jelly from cord membrane, collagenase and trypsin treatment and density gradient centrifugation. Magnetic activated cell sorting was performed to specifically select SSEA3+ Muse cells, and flow cytometry and immunocytochemistry were used to identify further. In vivo, spinal cord contusion injury model in rats was induced by NYU-III impactor, and were randomly divided and equally into four groups, namely group A (sham), group B (control), group C (Non-Muse cells transplantation) and group D (Muse cells transplantation). Laminectomy was conducted in group A but no spinal cord contusion injury. Laminectomy and cord injury were performed in group B, C and D, 10 g trip rod was freely falling down from 12.5 mm. Two weeks later, group B, C and D were received PBS injection, Non-Muse cells transplantation and Muse cells transplantation respectively, four-point injection were performed in each cord with totally 4×10⁵ cells. BBB scores were evaluated on 1 day, 1, 2, 3, 4, 5 and 6 week after injury. Four weeks after cell transplantation, the rats were sacrificed, and immunohistochemistry were carried out to observe survival, migration and differentiation of the injected cells.@*RESULTS@#The expression of CD105, CD90 and CD73 were over 99.5% in the derived WJ-MSCs population, but CD45 and CD14 were lower than 0.5%, positive rate of SSEA3+ was 1.46% under flow cytometer, However, after MACS sorting, the percentage of 92.0% Muse cells expressed SSEA3 and CD105, and immunohistochemistry results of SSEA3 showed typically membrane morphology with special processes. In vivo, BBB scores was 21 in group A at different time points. One-way ANOVA and LSD analysis showed that BBB scores in group C and D were significantly higher than that in group B (=0.004, 0.002), but there was no significantly difference between group C and D. Further intra-group paired t test showed that BBB score was significantly higher at 4 weeks than that 3 weeks in group C (=0.005). However, in group D, BBB scores were significantly higher at 4 and 6 week than those at 3 and 5 weeks, values were 0.005 and 0.016 respectively. Immunohistochemistry results showed that both Muse cells and Non-Muse cells could survive for 4 weeks in rats and they migrated from the four-point injection to injury site. But there showed more Muse cells survival than Non-Muse cells in the cord.@*CONCLUSIONS@#Immunomagnetic bead screening is efficient to select large number of purified SSEA3+ Muse cells. Muse cells could survive and target-migrate in injured cord to improve BBB scores continuously. Muse cells are a novel kind of seed cells in the spinal cord injury treatment.

Mannan-Binding Lectin Inhibits Candida Albicans-Induced DC Maturation and Cytokine Secretion / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the effects of mannan-binding lectin (MBL) on the maturation and cytokine secretion of human dendritic cells (DC) induced by Candida albicans (C. albicans).</p><p><b>METHODS</b>The plastic-adherent mononuclear cells were prepared from the blood of healthy adult volunteers. The human peripheral blood mononuclear cells-derived dendritic cells (MNC-DC) were induced by 5-day-culture in medium supplemented with rhGM-CSF and rhIL-4, and then cultured for 2 days in presence or absence of C. albicans at varying concentration of human MBL ranging from 1 to 20 mg/L. DC's shape and characters were observed under inverted microscopy, the expression of CD83 and CD86 on DC was analyzed by FACS. The levels of TNF-α and IL-6 were detected by ELISA. FACS also was used to investigate the interaction of MBL with immature DC(imDC) and C. albicans. Western blot was used to detect C. albicans-induced IκBα phosphorylation and p65/NF-κB translocation in DC.</p><p><b>RESULTS</b>MBL at higher concentrations (10-20 mg/L) down-regulated the expression of CD83 and CD86 on the monocyte-derived dentritic cells(MoDC) induced by C. albicans, and inhibited the production of TNF-α and IL-6 induced by C. albicans. FACS showed that MBL could not only bind to C. albicans but also bind to imDCs in a Ca2+-dependent manner. Western blot showed that MBL could decrease the phosphorylation of IκBα and the nuclear translocation of p65/ NF-κB.</p><p><b>CONCLUSION</b>MBL may inhibit TNF-α and IL-6 production induced by C. albicans in DC through NF-κB signaling pathways, suggesting that MBL can play some roles in the regulation of C. albicans-induced immune response.</p>

Distribution of Bifidobacterium in oral cavities of children and the relations with caries / 华西口腔医学杂志

<p><b>OBJECTIVE</b>To explore a selected-media of Bifidobacterium from oral cavity, to detect the distribution of Bifidobacterium in different sites of children and primarily investigate the relationship between oral Bifidobacterium and early childhood caries.</p><p><b>METHODS</b>70 children aged from 3 to 5-year-old were selected, 30 children were caries-free and 40 were severe early childhood caries (S-ECC). Saliva was collected and plaque samples from the 30 healthy subjects were pooled. For S-ECC group, plaques were collected separately from four different sites as follows: Saliva, surfaces of intact enamel, surfaces of white spot-lesions, and deep dentin-lesions. Samples would be grown in the selected-media, and the whole DNA of bacteria was extracted. Polymerase chain reaction was performed with specific primers and the results were analyzed by the electrophoresis.</p><p><b>RESULTS</b>Bifidobacterium were detected 0 in the caries-free children, while 47.5% in the S-ECC group. There was significant difference between two groups (P < 0.05) and there was no difference between different sites of teeth in S-ECC group (P > 0.05). 27.5% Bifidobacterium were detected in saliva, 27.5% on surfaces of intact enamel, 20.0% on surfaces of white spot-lesions and 22.5% in deep dentin-lesions. 10% Bifidobacterium dentium were detected in saliva, 7.5% on surfaces of intact enamel, 7.5% on surfaces of white spot-lesions and 10.0% in deep dentin-lesions.</p><p><b>CONCLUSION</b>One type of modified selected media of Bifidobacterium in oral cavity was explored. Bifidobacterium may be related to the occurrence of the S-ECC and has nothing to do with different sites of teeth in children.</p>