Subcutaneous Administration of PDGF-AA Improves the Functional Recovery After Spinal Cord Injury.

Previous studies by our group have demonstrated that the transplantation of exogenous platelet-derived growth factor (PDGF)-AA-overexpressing oligodendrocyte progenitor cells (OPCs) promotes tissue repair and recovery of neurological function in a rat model of spinal cord injury (SCI). However, it remains unclear whether treatment with PDGF-AA also affects endogenous oligodendrocytes (OLs) or even neurons, thus promoting further functional recovery after SCI. In the present study, we evaluated the therapeutic potential of PDGF-AA treatment by direct subcutaneous injection of PDGF-AA immediately after SCI. We demonstrated that PDGF-AA injection resulted in increased tissue sparing, myelination and functional recovery in rats following SCI. Further experimentation confirmed that PDGF-AA increased the survival of endogenous OPCs and OLs, and promoted the proliferation of OPCs and their differentiation into OLs. Moreover, PDGF-AA also protected motor neurons from death in the injured spinal cord. These results indicated that PDGF-AA administration may be an effective treatment for SCI.

[Analysis of Relationship between Serum Total Light Chain κ/λ Ratio and Proportion of Bone Marrow Plasma Cells in Patients with IgG type and IgA type Multiple Myeloma].

OBJECTIVE: To explore the relationship between serum total light chain κ/λ ratio (sTLC-κ/λ) and proportion of bone marrow plasma cells (BMPC) in patients with IgG type and IgA type multiple myeloma (MM) and its clinical significance. METHODS: The levels of serum IgG, IgA, κ type and λ type total light chain were detected in 79 newly diagnosed patients with IgG type (n=52) and IgA type (n=27) MM by immuno-nephelometric assay and the sTLC-κ/λ ratio was calculated. The proportion of BMPC was determined by bone marrow smears in the corresponding period, and the changes in sTLC-κ/λ ratio and the proportion of BMPC were observed in 19 patients with IgG type(n=16) and IgA type (n=3) MM undergoing treatment, 26 cases of non-phasmocytic proliferative diseases were enrolled in control group. RESULTS: In MM patients with IgGκ type and IgAκ type, the sTLC-κ/λ ratio was significantly higher than that in the control group (P<0.01), while in MM patients with IgGλ type and IgAλ type, the sTLC-κ/λ ratio was significantly lower than that in the control group (P<0.01). In MM patients with IgGκ, the sTLC-κ/λ ratio was significantly higher than that in MM patients with IgAκ(P<0.01), while the sTLC-κ/λ ratio in MM patients with IgGλ was significantly lower than that in MM patients with IgAλ. The sTLC-κ/λ ratios in MM patients with IgGκ and IgAκ were positively correlated with the concentrations of IgG (r=0.778,P=0.000) and IgA (r=0.601,P=0.039), while the sTLC-κ/λ ratios of patients with IgGλ and IgAλ were negativily correlated with the IgG(r=-0.586,P=0.01) and IgA level(r=-0.718,P=0.003). In addition, a correlation between each type MM was not found except the IgGκ type MM which had a positive correlation between the sTLC-κ/λ ratio and proportion of BMPC (r=0.579,P=0.002). Nonetheless, 18 of 19 patients with IgG type and IgA type MM undergoing treatment showed concordance between the sTLC-κ/λ ratio and proportion of BMPC change. CONCLUSION: There is a lower correlation between the sTLC-κ/λ ratio and the proportion of BMPC in MM patients with IgG type and IgA type, but there is a high concordance between the sTLC-κ/λ ratio and the proportion of BMPC change in the same patient and it suggests that the sTLC-κ/λ ratio plays an important role in the diagnosis and monitoring of IgG type and IgA type MM.

[Conditioned medium from rat RSC96 cells promotes proliferation of oligodendrocyte progenitor cells in vitro].

OBJECTIVE: To investigate the effect of conditioned medium from rat RSC96 cells (RSC96-CM) on the proliferation of oligodendrocyte progenitor cells (OPCs) and explore the underlying mechanism. METHODS: OPCs isolated from the spinal cords of SD rats of embryonic day 15 using immunopanning were treated with RSC96-CM. The proliferation of OPCs was detected using 5-bromo-2'-deoxyuridine (BrdU) incorporation assay. The mRNA expressions of PDGF-AA and bFGF in RSC96 cells were detected using RT-PCR, and their protein concentrations in RSC96-CM were detected with enzyme-linked immunosorbent assay (ELISA). The effects of PDGF-AA and bFGF in RSC96-CM on OPC proliferation and the roles of ERK and JNK signaling pathways in RSC96-CM-induced OPC proliferation were determined by application of their specific inhibitors. RESULTS: The percentage of BrdU+ OPCs was significantly increased in response to treatment with RSC96-CM (P<0.05), reaching the peak level when 50% RSC96-CM was added in the cell culture. RSC96 cells expressed a substantial amount of PDGF-AA and bFGF mRNAs, and PDGF-AA and bFGF protein concentrations in RSC96-CM were higher than those in a conditioned medium (B104CM) we used previously by 0.87 and 0.92 folds, respectively. Both the specific inhibitor of PDGFR signal pathway (AG1295) and the specific inhibitor of bFGFR signal pathway (PD173074) significantly attenuated RSC96-CM-induced OPC proliferation. The specific inhibitors of ERK signal pathway (U0126) and JNK signal pathway (SP600125) significantly decreased the percentage of BrdU+ cells in RSC96-CM-induced OPCs (P<0.01). CONCLUSION: RSC96-CM can effectively promote OPC proliferation, possibly as a result of PDGF-AA and bFGF secretion by RSC96 cells to activate ERK1/2 and JNK signaling pathways. RSC96- CM can be used as a routine stimulator for promoting OPC proliferation.

Transplantation of PDGF-AA-Overexpressing Oligodendrocyte Precursor Cells Promotes Recovery in Rat Following Spinal Cord Injury.

Our previous study showed that Schwann cells (SCs) promote survival, proliferation and migration of co-transplanted oligodendrocyte progenitor cells (OPCs) and neurological recovery in rats with spinal cord injury (SCI). A subsequent in vitro study confirmed that SCs modulated OPC proliferation and migration by secreting platelet-derived growth factor (PDGF)-AA and fibroblast growth factor-2 (FGF)-2. We also found that PDGF-AA stimulated OPC proliferation and their differentiation into oligodendrocytes (OLs) at later stages. We therefore speculated that PDGF-AA administration can exert the same effect as SC co-transplantation in SCI repair. To test this hypothesis, in this study we investigated the effect of transplanting PDGF-AA-overexpressing OPCs in a rat model of SCI. We found that PDGF-AA overexpression in OPCs promoted their survival, proliferation, and migration and differentiation into OLs in vivo. OPCs overexpressing PDGF-AA were also associated with increased myelination and tissue repair after SCI, leading to the recovery of neurological function. These results indicate that PDGF-AA-overexpressing OPCs may be an effective treatment for SCI.

Morroniside protects SK-N-SH human neuroblastoma cells against H2O2-induced damage.

Oxidative stress-induced cell injury has been linked to the pathogenesis of neurodegenerative disorders such as spinal cord injury, Parkinson's disease, and multiple sclerosis. Morroniside is an antioxidant derived from the Chinese herb Shan-Zhu-Yu. The present study investigated the neuroprotective effect of morroniside against hydrogen peroxide (H2O2)-induced cell death in SK-N-SH human neuroblastoma cells. H2O2 increased cell apoptosis, as determined by flow cytometry and Hoechst 33342 staining. This effect was reversed by pretreatment with morroniside at concentrations of 1-100 µM. The increase in intracellular reactive oxygen species (ROS) generation and lipid peroxidation induced by H2O2 was also abrogated by morroniside. H2O2 induced a reduction in mitochondrial membrane potential, increased caspase-3 activity, and caused downregulation of B cell lymphoma-2 (Bcl-2) and upregulation of Bcl-2-associated X protein (Bax) expression. These effects were blocked by morroniside pretreatment. Thus, morroniside protects human neuroblastoma cells against oxidative damage by inhibiting ROS production while suppressing Bax and stimulating Bcl-2 expression, thereby blocking mitochondrial-mediated apoptosis. These results indicate that morroniside has therapeutic potential for the prevention and treatment of neurodegenerative diseases.

Differential effects of myelin basic protein-activated Th1 and Th2 cells on the local immune microenvironment of injured spinal cord.

Myelin basic protein (MBP) activated T cells (MBP-T) play an important role in the damage and repair process of the central nervous system (CNS). However, whether these cells play a beneficial or detrimental role is still a matter of debate. Although some studies showed that MBP-T cells are mainly helper T (Th) cells, their subtypes are still not very clear. One possible explanation for MBP-T immunization leading to conflicting results may be the different subtypes of T cells are responsible for distinct effects. In this study, the Th1 and Th2 type MBP-T cells (MBP-Th1 and -Th2) were polarized in vitro, and their effects on the local immune microenvironment and tissue repair of spinal cord injury (SCI) after adoptive immunization were investigated. In MBP-Th1 cell transferred rats, the high levels of pro-inflammatory cells (Th1 cells and M1 macrophages) and cytokines (IFN-γ, TNF-α, -ß, IL-1ß) were detected in the injured spinal cord; however, the anti-inflammatory cells (Th2 cells, regulatory T cells, and M2 macrophages) and cytokines (IL-4, -10, and -13) were found in MBP-Th2 cell transferred animals. MBP-Th2 cell transfer resulted in decreased lesion volume, increased myelination of axons, and preservation of neurons. This was accompanied by significant locomotor improvement. These results indicate that MBP-Th2 adoptive transfer has beneficial effects on the injured spinal cord, in which the increased number of Th2 cells may alter the local microenvironment from one primarily populated by Th1 and M1 cells to another dominated by Th2, Treg, and M2 cells and is conducive for SCI repair.

Lower concentrations of methyl-ß-cyclodextrin combined with interleukin-2 can preferentially induce activation and proliferation of natural killer cells in human peripheral blood.

Previous studies have demonstrated that high concentrations of methyl-ß-cyclodextrin (MßCD, 10-15 mM) can interfere with the formation of lipid rafts and inhibit activation of lymphocytes. In this report, we determined that lower concentrations of MßCD (1-4 mM) could accelerate the proliferation of lymphocytes in human peripheral blood mononuclear cells (PBMCs). In the expanded cells, CD3(-)CD56(+) natural killer (NK) cells were the dominant subpopulation, and a significant dose-effect relationship existed between the proportion of NK cells and the concentration of MßCD. In the groups treated with 3-4 mM MßCD, the proportions of NK cells reached a level of more than 60%. When PBMCs were treated with MßCD, CD69 was more preferentially expressed on CD3(-)CD56(+) cells than on CD3(+) cells at 48 and 72 hours. The expression of CD25 had no distinct difference at 48 hours, but when recombinant human interleukin-2 (IL-2) was added for a further 24 hours, it was also preferentially expressed on NK cells. MßCD and IL-2 synergistically could also induce interferon-γ (IFN-γ) production in CD56(+) human PBMCs. Mechanistic studies revealed that IFN-γ production in response to MßCD plus IL-2 was IL-12 independent but depended on endogenous IL-18 and IL-1ß, and CD56(+)CD14(+) dendritic cell-like cells and B cells might mediate the ability of MßCD to activate NK cells. The MßCD-activated NK cells also had high cytotoxicity against the natural killer cell-sensitive K562 cells or lymphokine-activated killer cell-sensitive DAUDI cells in vitro. These studies indicated that lower concentrations of MßCD combined with IL-2 can preferentially induce activation and proliferation of NK cells in PBMCs.

Formation and aggregation of lipid rafts in γδ T cells following stimulation with Mycobacterium tuberculosis antigens.

Lipid rafts are plasma membrane microdomains that are implicated in diverse signaling pathways in immune cells. Based on the distinct types of T-cell receptors, two T-cell subpopulations have been identified: αß and γδ T cells. In humans, γδ T cells represent a relatively rare T lymphocyte population but play a critical role in the immune response to infection by Mycobacterium tuberculosis. It has been demonstrated that Mycobacterium tuberculosis antigens (Mtb-Ag) preferentially activate γδ T cells. Thus, we investigated whether lipid rafts are involved in the Mtb-Ag-mediated activation of γδ T cells. Human peripheral blood mononuclear cells (PBMCs) were stimulated with Mtb-Ag, and expression of a lipid raft marker ganglioside GM1 (GM1) was determined by flow cytometry. The aggregation of lipid rafts was evaluated by laser confocal microscopy. Non-stimulated fresh PBMCs minimally expressed GM1 (6.55 ± 2.01%) and had no aggregated rafts in γδ T cells. Mtb-Ag stimulation gradually increased the expression of GM1 in a time-dependent manner. At 72 h, the majority of γδ T cells expressed GM1 (88.69 ± 7.55%). Furthermore, accompanied with the increased expression of GM1, aggregation of lipid rafts became gradually visible in γδ T cells. The aggregated rafts, however, were not evenly distributed and only occurred over a small portion of GM1-positive cells. Pretreatment with methyl-ß-cyclodextrin, a cholesterol-depleting reagent, completely inhibited the Mtb-Ag-mediated aggregation of lipid rafts. These results demonstrate that lipid raft aggregation occurs in Mtb-Ag-activated γδ T cells, suggesting that lipid rafts are involved in activation of γδ T cells.